AU2023398887A1

AU2023398887A1 - Bispecific antibody fusion molecules and methods of use thereof

Info

Publication number: AU2023398887A1
Application number: AU2023398887A
Authority: AU
Inventors: Xingyue AN; Guixian JIN; Stella Aviaty Martomo; Jeremy S. MYERS; Mohosin SARKAR; Eric M. Tam
Original assignee: Evolveimmune Therapeutics Inc
Current assignee: Evolveimmune Therapeutics Inc
Priority date: 2022-12-14
Filing date: 2023-12-14
Publication date: 2025-06-26
Also published as: IL321437A; CA3277013A1; WO2024130032A1; EP4634228A1

Abstract

The present disclosure provides bispecific antibodies or antigen binding fragments thereof that specifically bind to CD3 and CD20. The antibodies are optionally fused with a CD58 peptide or portions thereof. Described herein are methods of making and using the bispecific antibodies for treatment of cancers that express CD20.

Description

BISPECIFIC ANTIBODY FUSION MOLECULES AND METHODS OF USE THEREOF

RELATED APPLICATIONS

[0001] This application claims priority to, and the benefit of, U.S. Provisional Application No. 63/432,665, filed December 14, 2022, the contents of which are incorporated herein by reference in their entirety.

REFERENCE TO AN ELECTRONIC SEQUENCE LISTING [0002] The contents of the electronic sequence listing (EVIM- 003 001WO SeqList _ST26.xml; Size: 325,996 bytes; and Date of Creation: December 14, 2023) are herein incorporated by reference in its entirety.

BACKGROUND OF THE INVENTION

[0003] Bi-specific antibodies capable of targeting T cells to tumor cells have been identified and tested for their efficacy in the treatment of cancers. B-cell proliferative disorders describe a heterogeneous group of malignancies that includes both leukemias and lymphomas. Lymphomas develop from lymphatic cells and include two mam categories: Hodgkin lymphomas (HL) and the non-Hodgkin lymphomas (NHL). In the United States, lymphomas of B cell origin constitute approximately 80-85% of all non-Hodgkin lymphoma cases, and there is considerable heterogeneity within the B-cell subset, based upon genotypic and phenotypic expression paterns in the B-cell of origin. Despite the availability of various agents for the treatment of B-cell proliferative disorders, there is an ongoing need for development of safe and effective therapies to prolong remission and improve cure rates in patients.

[0004] A bispecific antibody that targets CD20 expressed on B cells and CD3 epsilon chain (CD3e) present on T cells leads to T-cell activation and T-cell mediated killing of B cells. In the presence of CD20 expressing B ceils, whether circulating or tissue resident, pharmacologically active doses of the bispecific antibody w ill trigger T-ceil activation and associated cytokine release. Parallel to B cell depletion in the peripheral blood, the bispecific antibody leads to a transient decrease of T cells in the peripheral blood within 24 hours after the first administration and to a peak in cytokine release, followed by rapid T- cell recovery and return of cytokine levels to baseline within 72 hours. Thus, in order to achieve complete elimination of tumor cells, there is a need for improved bispecific antibodies or multi-specific antibodies that conserve T-cell activation, improve T-cell persistence and delivers durable immune response to cancer cells.

SUMMARY OF THE INVENTION

[0005] This disclosure provides a bi specific antibody comprising a first antigen binding region that binds to CD3 and a second antigen binding region that binds to CD20, wherein the first antigen binding region that binds to CD3 comprises three heavy chain complementarity' determining regions (VHI CDRI, VH 1_CDR2, VH1 CDR3) and three light chain complementarity determining regions (VL1 CDRI, VL1 CDR2, VL1 CDR3), wherein a) VH 1_CDR1 comprises the amino acid sequence of SEQ ID NO: 29;

VH1__CDR2 comprises the amino acid sequence of SEQ ID NO: 34; VH1 CDR3 comprises the amino acid sequence of SEQ ID NO: 40; VL1 CDR1 comprises the ammo acid sequence of SEQ ID NO: 42; VL1__CDR2 comprises the amino acid sequence of SEQ ID NO: 43; and VL1_CDR3 comprises the amino acid sequence of SEQ ID NO: 47; b) VHI CDRI comprises the amino acid sequence of SEQ ID NO: 29; VH 1_CDR2 comprises the amino acid sequence of SEQ ID NO: 35; VH1 CDR3 comprises the amino acid sequence of SEQ ID NO: 38; VL1 CDR1 comprises tire ammo acid sequence of SEQ ID NO: 42; VL1 CDR2 comprises the amino acid sequence of SEQ ID NO: 43; and VL1 CDR3 comprises the amino acid sequence of SEQ ID NO: 47; c) VHI CDRI comprises the amino acid sequence of SEQ ID NO: 30; VH 1_CDR2 comprises the amino acid sequence of SEQ ID NO: 34; VH1_CDR3 comprises the amino acid sequence of SEQ ID NO: 37; VL1 CDR1 comprises the amino acid sequence of SEQ ID NO: 42; VL1 CDR2 comprises the amino acid sequence of SEQ ID NO: 43; and VL1 CDR3 comprises the amino acid sequence of SEQ ID NO: 45; d) VHI CDRI comprises the amino acid sequence of SEQ ID NO: 29; VH 1_CDR2 comprises the amino acid sequence of SEQ ID NO: 34; VH1_CDR3 comprises the amino acid sequence of SEQ ID NO: 39; VL1 CDRI comprises the amino acid sequence of SEQ ID NO: 42; VL1 CDR2 comprises the amino acid sequence of SEQ ID NO: 43; and VL1 CDR3 comprises the amino acid sequence of SEQ ID NO: 47; e) VHI CDRI comprises the ammo acid sequence of SEQ ID NO: 29; VH 1_CDR2 comprises the amino acid sequence of SEQ ID NO: 34; VHI CDR3 comprises the amino acid sequence of SEQ ID NO: 37; VL1 CDR1 comprises the amino acid sequence of SEQ ID NO: 42; VL1_CDR2 comprises the amino acid sequence of SEQ ID NO: 44; and VL1 CDR3 comprises the amino acid sequence of SEQ ID NO: 45; or f) VH1 CDR1 comprises the amino acid sequence of SEQ ID NO: 29; VH 1_CDR2 comprises the amino acid sequence of SEQ ID NO: 34; VH1 CDR3 comprises the amino acid sequence of SEQ ID NO: 37; VL1 _CDR 1 comprises the am ino acid sequence of SEQ ID NO: 42; VL1 CDR2 comprises the amino acid sequence of SEQ ID NO: 43; and VL1 CDR3 comprises the amino acid sequence of SEQ ID NO: 45; and wherein the second antigen binding region that binds to CD20 comprises three heavy chain complementarity determining regions (VH2 CDR1, VH2 CDR2, VH2 CDR3) and three light chain complementarity determining regions (VL2 CDR1, VL2 CDR2, VL2 CDR3), wherein i) VH2 CDR1 comprises the amino acid sequence of SEQ ID NO: 455; VH2 CDR2 comprises the amino acid sequence of SEQ ID NO: 457; VH2_CDR3 comprises the amino acid sequence of SEQ ID NO: 458; VL2 CDR1 comprises the amino acid sequence of SEQ ID NO: 463; VL2__CDR2 comprises the amino acid sequence of SEQ ID NO: 464; and VL2_CDR3 comprises the amino acid sequence of SEQ ID NO: 465; or ii) VH2_CDR1 comprises the amino acid sequence of SEQ ID NO: 459; VH2_CDR2 comprises the amino acid sequence of SEQ ID NO: 460; VH2 CDR3 comprises the amino acid sequence of SEQ ID NO: 462; VL2__CDR1 comprises tire ammo acid sequence of SEQ ID NO: 466; VL2_CDR2 comprises the amino acid sequence of SEQ ID NO: 467; and VL2_CDR3 comprises the amino acid sequence of SEQ ID NO: 468.

[0006] In some embodiments, the first antigen binding region that binds to CD3 comprises a variable heavy chain region ( VH 1 ) and a variable light chain region (VL1), wherein: a) VH1 comprises the amino acid sequence of SEQ ID NO: 19 and VL1 comprises the amino acid sequence of SEQ ID NO: 26; b) VH1 comprises the amino acid sequence of SEQ ID NO: 18 and VL1 comprises the amino acid sequence of SEQ ID NO: 26; c) VH1 comprises the amino acid sequence of SEQ ID NO: 17 and VL1 comprises the amino acid sequence of SEQ ID NO: 22; d) VH1 comprises the ammo acid sequence of SEQ ID NO: 16 and VL1 comprises the amino acid sequence of SEQ ID NO: 26; e) VH1 comprises the amino acid sequence of SEQ ID NO: 13 and VL1 comprises the amino acid sequence of SEQ ID NO: 27; or f) VH1 comprises the amino acid sequence of SEQ ID NO: 13 and VL1 comprises the amino acid sequence of SEQ ID NO: 22; and wherein the second antigen binding region that binds to CD20 comprises a variable heavy chain region (VH2) and a variable light chain region (VL2), wherein: i) VH2 comprises the amino acid sequence of SEQ ID NO: 451 and VL2 comprises the amino acid sequence of SEQ ID NO: 450; or ii) VH2 comprises the amino acid sequence of SEQ ID NO: 453 and VL2 comprises the amino acid sequence of SEQ ID NO: 452.

[0007] In some embodiments, the bispecific antibody has the following structure: a first heavy chain polypeptide (H1 ) comprising a variable region (VH1), and a constant region (CHI) having a constant region 1 domain (CH1___H1), a hinge region (H1H), a constant region 2 domain (CH1__H2) and a constant region 3 domain (CH1 H3); and a first light chain polypeptide (L1) comprising a variable region (VL1) and a constant region (CL1); and a second heavy chain polypeptide (H2) comprising a variable region (VH2), and a constant region (CH2) having a constant region 1 domain (CH2___H1), a hinge region (H2H), a constant region 2 domain (CH2 H2) and a constant region 3 domain (CH2__H3); and second light chain polypeptide (L2) comprising a variable region (VL2) and a constant region (CL2), and wherein i) the amino acid at position 39 (Kabat numbering) of the VH1 is a K and the amino acid at position 38 (Kabat numbering) of the VL1 is a D; ii) the amino acid at position 147 (EU numbering) of the CH1 H1 is a K and the amino acid at position 131 (EU numbering) of the CL1 is a D; iii) the amino acid at position 173 (EU numbering) of the CHI H1 is a C and the amino acid at position 162 (EU numbering) of the CL1 is a C; iv) the amino acid at position 220 (EU numbering) in the H1H is a S and the amino acid at position 214 (EU numbering) of the CL1 is a S; and i) the amino acid at position 39 (Kabat numbering) of the VH2 is a D and the amino acid at position 38 (Kabat numbering) of the VL2 is a K; and ii) the amino acid at position 147 (EU num bering) of the CH2_H1 is a D and the amino acid at position 180 (EU numbering) of the CL2 is a R.

[0008] In some embodiments, i) the amino acid at position 87 (Kabat numbering) of the VH1 and/or VH2 is a G; and ii) the amino acid at position 45 (Kabat numbering) of the VL1 and/or VL2 is a W.

[0009] In some embodiments, i) the CH1 H3 has a C at position 349, an S at position 366, an A at position 368 and a V at position 407 (EU numbering); and the CH2 _ H3 has a C at position 354 and a W at position 366 (EU numbering); ii) the CH2_H3 has a C at position 349, an S at position 366, an A at position 368 and a V at position 407 (EU numbering); and the CH1 H3 has a C at position 354 and a W at position 366 (EU numbering); iii) the CH1_H3 has a C at position 354, an S at position 366, an A at position 368 and a V at position 407 (EU numbering); and the CH2__H3 has a C at position 349 and a W at position 366 (EU numbering); or iv) the CH2 _ H3 has a C at position 354, an S at position 366, an A at position 368 and a V at position 407 (EU numbering); and the CH1 H3 has a C at position 349 and a W at position 366 (EU numbering).

[0010] In some embodiments, the amino acid at position 447 (EU numbering) of the CH1 H3 and/or of the CH2 _ H3 is deleted. In some embodiments, i) the H1H and/or the H2H has an A at positions 234 and 235 (EU numbering); ii) the H1H and/or the H2H has an A at positions 234, 235 and 237 (EU numbering); or iii) the HIH and/or the H2.H has an A at positions 234 and 235 and G at position 329 (EU numbering).

[0011] In some embodiments, the amino acid at position 446 and at position 447 (EU numbering) of the CH1 _ H3 and/or of the CH2 _ H3 is deleted. In some embodiments, i) tire HI H and/or the H2.H has an A at positions 234 and 235 (EU numbering); ii) the HIH and/or the H2H has an A at positions 234, 235 and 237 (EU numbering); or iii) the HIH and/or the H2H has an A at positions 234 and 235 and G at position 329 (EU numbering). [0012] In some embodiments, i) the CH1 _ H3 and/or the CH2 _ H3 has an A at position 297 (EU numbering); ii) the CH1 H3 and/or the CH2 _ H3 has a G at position 297 (EU numbering); or iii) the CH1 _ H3 and/or the CH2 _ H3 has a S at position 297 (EU numbering). In some embodiments, the CH1 _ H3 and/or the CH2 _ H3 has an S at position 331 (EU numbering).

[0013] In some embodiments, a polypeptide is fused to the N-tenninus or the C-terminus of the first heavy chain polypeptide or the second heavy chain polypeptide. In some embodiments, the polypeptide is fused to the C -terminus of the first heavy chain polypeptide.

[0014] In some embodiments, the polypeptide is fused via a linker peptide. In some embodiments, the linker peptide comprises the amino acid sequence of SEQ ID NO: 53. In some embodiments, the polypeptide comprises a CD58 or a fragment thereof. In some embodiments, the CD.58 comprises the amino acid sequence of SEQ ID NO: 49.

[0015] In some embodiments, the bispecific antibody comprises a VH1, a VL1, a VH2 and a VL2, wherein i) the VH1 comprises the amino acid sequence of SEQ ID NO: 19; the VL1 comprises the amino acid sequence of SEQ ID NO: 26; the VH2 comprises the amino acid sequence of SEQ ID NO: 453; and the VL2 comprises the amino acid sequence of SEQ ID NO: 452; ii) the VH1 comprises the amino acid sequence of SEQ ID NO: 18; the VL1 comprises the amino acid sequence of SEQ ID NO: 26; the VH2 comprises the amino acid sequence of SEQ ID NO: 453; and the VL2 comprises the amino acid sequence of SEQ ID NO: 452; iii) the VHI comprises the amino acid sequence of SEQ ID NO: 17; the VL1 comprises the amino acid sequence of SEQ ID NO: 22; the VH2 comprises the ammo acid sequence of SEQ ID NO: 453; and the VL2 comprises the amino acid sequence of SEQ ID NO: 452; iv) the VHI comprises the amino acid sequence of SEQ ID NO: 16; the VL1 comprises the amino acid sequence of SEQ ID NO: 26; the VH2 comprises the amino acid sequence of SEQ ID NO: 453; and the VL2 comprises the amino acid sequence of SEQ ID NO: 452; v) the VHI comprises the amino acid sequence of SEQ ID NO: 13; the VL1 comprises the amino acid sequence of SEQ ID NO: 27; the VH2 comprises the amino acid sequence of SEQ ID NO: 453; and the VL2 comprises the amino acid sequence of SEQ ID NO: 452; vi) the VHI comprises the amino acid sequence of SEQ ID NO: 13; the VLl comprises the amino acid sequence of SEQ ID NO: 22; the VH2 comprises the amino acid sequence of SEQ ID NO: 453; and the VL2 comprises the amino acid sequence of SEQ ID NO: 452; vii) the VHI comprises the amino acid sequence of SEQ ID NO: 19; the VLl comprises the amino acid sequence of SEQ ID NO: 26; the VH2 comprises the amino acid sequence of SEQ ID NO: 451 ; and the VL2 comprises the amino acid sequence of SEQ ID NO: 450; viii) the VHI comprises the ammo acid sequence of SEQ ID NO: 18; the VLl comprises the amino acid sequence of SEQ ID NO: 26; the VH2 comprises the amino acid sequence of SEQ ID NO: 451; and the VL2 comprises the amino acid sequence of SEQ ID NO: 450; ix) the VHI comprises the amino acid sequence of SEQ ID NO: 17; the VLl comprises the amino acid sequence of SEQ ID NO: 22; the VH2 comprises the amino acid sequence of SEQ ID NO: 451 ; and the VL2 comprises the amino acid sequence of SEQ ID NO: 450; x) the VHI comprises the amino acid sequence of SEQ ID NO: 16; the VLl comprises the amino acid sequence of SEQ ID NO: 26; the VH2 comprises the ammo acid sequence of SEQ ID NO: 451; and the VL2 comprises the amino acid sequence of SEQ ID NO: 450; xi) the VHI comprises the amino acid sequence of SEQ ID NO: 13; the VLl comprises the amino acid sequence of SEQ ID NO: 27; the VH2 comprises the amino acid sequence of SEQ ID NO: 451 ; and the VL2 comprises the amino acid sequence of SEQ ID NO: 450; or xii) the VHI comprises the amino acid sequence of SEQ ID NO: 13; the VLl comprises the amino acid sequence of SEQ ID NO: 22; the VH2 comprises the amino acid sequence of SEQ ID NO: 451 ; and the VL2 comprises the amino acid sequence of SEQ ID NO: 450.

[0016] In some embodiments, the bispecific antibody comprises a H1 , a Ll, a H2 and a L2, wherein i) the H1 comprises the amino acid sequence of SEQ ID NO: 491; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 490; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; ii) the H1 comprises the ammo acid sequence of SEQ ID NO: 491; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 566; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; iii) the H1 comprises the amino acid sequence of SEQ ID NO: 595; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 490; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; iv) the H1 comprises the amino acid sequence of SEQ ID NO: 595; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 566; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; v) the H1 comprises the amino acid sequence of SEQ ID NO: 494; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 493; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; vi) the H1 comprises the amino acid sequence of SEQ ID NO: 494; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 567; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; vii) the H1 comprises the amino acid sequence of SEQ ID NO: 596; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 493; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; viii) the H1 comprises the amino acid sequence of SEQ ID NO: 596; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 567; and the L2 comprises the amino acid sequence of SEQ ID NO: 471 ; ix) the H1 comprises the amino acid sequence of SEQ ID NO: 497; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 496; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; x) the H1 comprises the amino acid sequence of SEQ ID NO: 497; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 568; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xi) the H1 comprises the amino acid sequence of SEQ ID NO: 597; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 496; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xii) the H1 comprises the amino acid sequence of SEQ ID NO: 597; the L1 comprises the amino acid sequence of SEQ ID NO: 141 ; the H2 comprises the amino acid sequence of SEQ ID NO: 568; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xiii) the H1 comprises the amino acid sequence of SEQ ID NO: 500; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 499; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xiv) the H1 comprises the amino acid sequence of SEQ ID NO: 500; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 569; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xv) the H1 comprises the amino acid sequence of SEQ ID NO: 598; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 499; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xvi) the H1 comprises the amino acid sequence of SEQ ID NO: 598; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 569; and the L2 comprises the amino acid sequence of SEQ ID NO: 471 ; xvii) the H1 comprises the amino acid sequence of SEQ ID NO: 503; the L1 comprises the amino acid sequence of SEQ ID NO: 153; tire H2 comprises the amino acid sequence of SEQ ID NO: 502; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xviii) the H1 comprises the amino acid sequence of SEQ ID NO: 503; the L1 comprises the amino acid sequence of SEQ ID NO: 153; the H2 comprises the amino acid sequence of SEQ ID NO: 570; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xix) the H1 comprises the amino acid sequence of SEQ ID NO: 599; the L1 comprises the amino acid sequence of SEQ ID NO: 153; the H2 comprises the amino acid sequence of SEQ ID NO: 502; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xx) the H1 comprises the amino acid sequence of SEQ ID NO: 599; the L1 comprises the amino acid sequence of SEQ ID NO: 153; the H2 comprises the amino acid sequence of SEQ ID NO: 570; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xxi) the H1 comprises the amino acid sequence of SEQ ID NO: 506; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 505; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xxii) the H1 comprises the amino acid sequence of SEQ ID NO: 506; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 571; and the L2 comprises the amino acid sequence of SEQ ID NO: 471 ; xxiii) the H1 comprises the amino acid sequence of SEQ ID NO: 600; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 505; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xxiv) the H1 comprises the ammo acid sequence of SEQ ID NO: 600; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 571; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xxv) the H1 comprises the amino acid sequence of SEQ ID NO: 527; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 526; and the L2 comprises the amino acid sequence of SEQ ID NO: 525; xxvi) the H1 comprises the amino acid sequence of SEQ ID NO: 527; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 572; and the L2 comprises the amino acid sequence of SEQ ID NO: 525; xxvii) the H1 comprises tire ammo acid sequence of SEQ ID NO: 601; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 526; and the L2 comprises the amino acid sequence of SEQ ID NO: 525; xxviii) the H1 comprises the ammo acid sequence of SEQ ID NO: 601; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 572; and the L2 comprises the amino acid sequence of SEQ ID NO: 525; xxix) the H1 comprises the amino acid sequence of SEQ ID NO: 530; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 529; and the L2 comprises the amino acid sequence of SEQ ID NO: 528; xxx) the H1 comprises the amino acid sequence of SEQ ID NO: 530; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 573; and the L2 comprises the amino acid sequence of SEQ ID NO: 528; xxxi) the H1 comprises the ammo acid sequence of SEQ ID NO: 602; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 529; and the L2 comprises the amino acid sequence of SEQ ID NO: 528; xxxii) the H1 comprises the amino acid sequence of SEQ ID NO: 602; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 573; and the L2 comprises the amino acid sequence of SEQ ID NO: 528; xxxiii) the H1 comprises the amino acid sequence of SEQ ID NO: 533; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 532; and the L2 comprises the amino acid sequence of SEQ ID NO: 531; xxxiv) the H1 comprises the amino acid sequence of SEQ ID NO: 533; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 574; and the L2 comprises the amino acid sequence of SEQ ID NO: 531; xxxv) the HI comprises the amino acid sequence of SEQ ID NO: 603; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 532; and the L2 comprises the amino acid sequence of SEQ ID NO: 531; xxxvi) the H1 comprises the amino acid sequence of SEQ ID NO: 603; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 574; and the L2 comprises the amino acid sequence of SEQ ID NO: 531; xxxvii) the H1 comprises the amino acid sequence of SEQ ID NO: 536; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 535; and the L2 comprises the amino acid sequence of SEQ ID NO: 534; xxxviii) the H1 comprises the amino acid sequence of SEQ ID NO: 536; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 575; and the L2 comprises the amino acid sequence of SEQ ID NO: 534; xxxix) the H1 comprises the amino acid sequence of SEQ ID NO: 604; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 535; and the L2 comprises the amino acid sequence of SEQ ID NO: 534; xl) the HI comprises the amino acid sequence of SEQ ID NO: 604; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 575; and the L2 comprises the amino acid sequence of SEQ ID NO: 534; xli) the H1 comprises the amino acid sequence of SEQ ID NO: 539; the L1 comprises the amino acid sequence of SEQ ID NO: 153; the H2 comprises the amino acid sequence of SEQ ID NO: 538; and the L2 comprises the amino acid sequence of SEQ ID NO: 537; xlii) the H1 comprises the amino acid sequence of SEQ ID NO: 539; the L1 comprises the amino acid sequence of SEQ ID NO: 153; the H2 comprises the amino acid sequence of SEQ ID NO: 576; and the L2 comprises the amino acid sequence of SEQ ID NO: 537; xliii) the H1 comprises the amino acid sequence of SEQ ID NO: 605; the L1 comprises the amino acid sequence of SEQ ID NO: 153; the H2 comprises the amino acid sequence of SEQ ID NO: 538; and the L2 comprises the amino acid sequence of SEQ ID NO: 537; xliv) the HI comprises the amino acid sequence of SEQ ID NO: 605; the L1 comprises the amino acid sequence of SEQ ID NO: 153; the H2 comprises the amino acid sequence of SEQ ID NO: 576; and the L2 comprises the amino acid sequence of SEQ ID NO: 537; xlv) the HI comprises the amino acid sequence of SEQ ID NO: 542; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 541 ; and the L2 comprises the amino acid sequence of SEQ ID NO: 540; xlvi) the H1 comprises the ammo acid sequence of SEQ ID NO: 542; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 577; and the L2 comprises the amino acid sequence of SEQ ID NO: 540; xlvii) the H1 comprises the amino acid sequence of SEQ ID NO: 606; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 541 ; and the L2 comprises the amino acid sequence of SEQ ID NO: 540; or xlviii) the H1 comprises the amino acid sequence of SEQ ID NO: 606; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 577; and the L2 comprises the amino acid sequence of SEQ ID NO: 540.

[0017] In some embodiments, the bispecific antibody of the disclosure is an IgGl or an IgG4 antibody. In some embodiments, the bispecific antibody is a monoclonal antibody, a chimeric antibody or a humanized antibody.

[0018] The disclosure also provides a polynucleotide comprising a nucleic acid sequence encoding any one of the bispecific antibodies of the disclosure. The disclosure also provides a vector comprising the polynucleotide of the disclosure .

[0019] The disclosure also provides a pharmaceutical composition comprising any one of the bispecific antibodies of the disclosure, any one of the polynucleotides of the disclosure or any one of the vectors of the disclosure, and a pharmaceutically acceptable earner.

[0020] The disclosure also provides a method of treating a cancer expressing CD20 in a subject in need thereof comprising administering a therapeutically effective amount of anyone of the bispecific antibodies of the disclosure or any one of the pharmaceutical compositions of the disclosure. The disclosure also provides a method of T-cell re -targeting in a subject in need thereof comprising administering a therapeutically effective amount of any one of the bispecific antibodies of the disclosure or any one of the pharmaceutical compositions of the disclosure. The disclosure also provides a method of T-cell activation in a subject in need thereof compri sing administering a therapeutically effective amount of any one of the bispecific antibodies of the disclosure or any one of the pharmaceutical compositions of the disclosure.

[0021] In some embodiments, the subject has a B-cell cancer. In some embodiments, the B- cell cancer is follicular lymphoma, B-cell chronic lymphocytic leukemia, B-cell lymphoblastic lymphoma, Hodgkin lymphoma, non-Hodgkin’s lymphoma (NHL), chronic lymphoid leukemia (CLL), diffuse large B-cell lymphoma, marginal zone lymphoma, Mantle cell lymphoma, hairy cell leukemia, Burkitt lymphoma or small lymphocytic lymphoma (SLL), diffuse-large B cell lymphoma (DLBCL), a primary mediastinal (thymic) large B cell lymphoma (PMLBCL).

[0022] In some embodiments, the subject is simultaneously administered or previously administered with a therapeutically effective amount of an additional therapeutic agent. In some embodiments, the additional therapeutic agent is an anti-CD20 monospecific antibody. In some embodiments, the anti-CD20 monospecific therapy is rituximab. In some embodiments, the subject is previously administered with a therapeutically effective amount of rituximab.

[0023] In some embodiments, the additional therapeutic agent is a CAR-T cell therapy, an immune checkpoint inhibitor, a co-stimulatory ligand or a cytokine.

BRIEF DESCRIPTION OF THE DRAWINGS

[0024] FIGS. 1A-1F depicts schematic diagrams of antibodies with charged pair mutations, disulfide bond repositioning and knob into hole mutations. Grey shaded domains represent a first heavy chain polypeptide (H1) having a heavy chain variable region (VH1), having a constant region 1 domain (CHIHI or CH1 H1), a hinge region (H1H), a constant region 2 domain (CHI H2 or CH1_H2) and a constant region 3 domain (CH 1 H3 or CH1_H3); and a first light chain polypeptide (L1) comprising a variable region (VL1) and a constant region (CL I). White shaded domains represent a second heavy chain polypeptide (H2) comprising a variable region (VH2), and a constant region (CH2) having a constant region 1 domain (CH2H1 or CH2_H1), a hinge region (H2H), a constant region 2 domain (CH2«2or CH2 H2) and a constant region 3 domain (CH2H3 or CH2 _ H3 ); and second light chain polypeptide (L2) comprising a variable region (VL 2) and a constant region (CL2). The + and - symbols between the antigen binding domains represent the charged pair mutations. Lines between the ( CH1_H1 and CL1 domain and ( CH2_H1 and CL2 domain represent disulfide bonds, where a solid line represents an endogenous disulfide bond and a dashed line represents a repositioned disulfide bond. The protrusion and dent between the CH1_H3 and CH2 _ H3 domain represent knob into hole mutations. Charged pair mutations, disulfide bond repositioning and knob into hole mutations provide increased heavy chain and light chain heterodimerization, which is advantageous for production and purification of bispecific antibodies of the disclosure. FIG. IE depicts a schematic diagram of an exemplary bispecific antibody fusion molecule comprising an anti-CD3 arm, an anti-CD20 arm and a CD58 peptide fused to the CH1 H3 domain. FIG. 1F depicts a schematic diagram of an exemplary bispecific antibody fusion molecule comprising an anti-CD3 arm, an anti-CD20 arm and a CD58 peptide fused to the CH2 _ H3 domain.

[0025] FIG. 2 is a graph depicting preparative chromatograms obtained from protein A elution and size exclusion chromatography tandem purification of light chain pairing bispecific antibodies.

[0026] FIGS. 3A-3E are a series of graphs depi cting surface plasma resonance (SPR) binding sensorgrams of the captured recombinant CD3ε/5 interacting with light chain pairing bispecific antibodies. The sensograrns are recorded from single cycle kinetics with varying concentrations of bispecific antibodies. FIG. 3A show's EIP0866. FIG. 3B shows EIP0886. FIG. 3C shows EIP0892. FIG. 3D show's EIP0893. FIG. 3E shows EIP0929. [0027] FIG. 4 is a graph depicting differential scanning calorimetry analysis of light chain pairing bispecific antibodies.

[0028] FIGS. 5A-5B are two graphs depicting mass spectrometry analysis of intact and reduced masses of an exemplary light chain pairing bispecific antibody. FIG. 5A shows intact mass of EIP0866 determined after PNGAse F deglycosylation in non-reduced condition and chromatographic separation using reverse phase Cl 8 column. FIG. 5B shows reduced mass analysis of EIP0866 heavy chains and light chains determined after Rapid PNGAse F deglycosylation in reduced condition and chromatographic separation using reverse phase C18 column. [0029] FIG. 6 is a graph depicting functional evaluations of bispecific antibodies compared to controls in a tumor cell and PBMCs co-culture assay. JeKo-l-GFP-luc tumor cells were treated with PBMCs at 15: 1 E:T ratio with a serial dilution of bispecific antibody for 72 h. Tumor cells were counted by flow cytometry.

[0030] FIG. 7 is a graph depicting a cytotoxicity assay of bispecific antibodies compared to controls in a tumor cell and PBMCs co-culture assay. JeKo- 1 -GFP-luc tumor cells were treated with PBMCs at 15: 1 E:T ratio with a serial dilution of biologies for 72 h. Tumor cells were counted by flow cytometry.

[0031 ] FIG. 8 is a graph depicting a cytotoxicity assay of bispecific antibodies compared to control in tumor cell and PBMCs co-culture assay in an intermediate target density Raji- GFP-luc tumor ceil model. Tumor cells were treated with PBMCs at 15: 1 E:T ratio with a serial dilution of bispecific antibodies for 96 h. Tumor cells were counted by flow cytometry.

[0032] FIG. 9 is a graph depicting a cytotoxicity assay of bispecific antibodies compared to control in tumor cell and PBMCs co-culture assay in a low target density NALM6-GFP tumor cell model . Tumor cells were treated with PBMCs at 15 : 1 E:T ratio with a serial dilution of biologies for 96 h. Tumor ceils were counted by flow cytometry. The concentration of antibody is shown on the x-axis. The tumor cell counts are shown on the y- axis.

[0033] FIGS. 10A-10C are a series of graphs depicting a T cell repeat activation assay of bispecific antibodies on magnetic Dynabeads and of CD3/CD28 Dynabeads as a control. T cells were quantified after each round of activation. FIG. 10A shows the proliferation of T cells after five rounds of activation with bispecific antibodies and control. The round of T cell activation is shown on the x-axis. The T cell counts are shown on the y-axis. FIG. 10B shows a cytotoxicity assay of bispecific antibodies with T cells activated for five rounds and Raji-GFP-luc tumor cells co-culture. Tumor cells were treated with T cells activated for 5 rounds at 5: 1 E:T ratio for 72 h. Tumor cells were counted by flow cytometry . The concentration of antibody is shown on the x-axis. The live tumor cell counts are shown on the y-axis. FIG. 10C shows the expansion of CD8 T cells from a cytotoxicity assay performed with T cells activated for five rounds as described in FIG. 10B. The concentration of antibody is shown on the x-axis. The CD8 T cell counts are shown on the y-axis. [0034] FIG. 11 is a graph depicting tumor growth inhibition of a Burkitt’s lymphoma cell, Raji, in a human activated T cells and tumor cells co-engraftment study in a xenograft NSG mouse model. At day 0, 2x10⁶ activated T ceil and 10x10⁶ Raji-GFP-Iuc tumor cells were implanted subcutaneously with intravenous dosing of bispecific antibodies at day 0 and 7 days apart thereafter for 5 weeks. Tumor growth was monitored up to day 48.

[0035] FIG. 12 is a series of graphs showing thermal denaturation and melting temperature (Tm) of EIP0929, EIP0970, E1P0866, and a Glofitamab biosimilar benchmark determined by differential scanning calorimetry (DSC) using nanoDSC from TA Instrument, [0036] FIG. 13A is a series of graphs showing thermal forced denaturation of EIP0929, EIP0970, and EIP0866, and Glofitamab biosimilar benchmark was carried out by holding protein at different temperatures, and remaining intact protein was determined by analytical size exclusion chromatography.

[0037] FIG. 13B is a graph showing the thermal denaturing profiles of the fraction intact proteins remaining as a function of temperature.

[0038] FIGS. 14A and 14B are graphs showing a cytotoxicity assay of bispecific antibodies compared to controls and benchmarks in a tumor cell and PBMCs co-culture assay. Raji- GFP-luc tumor cells were treated with PBMCs at 15: 1 E:T ratio with a serial dilution of biologies for 72 h (FIG. 14A). Tumor cells and CD 19+ B cells were counted by flow cytometry (FIG. 14B).

[0039] FIG S. ISA, 15B, 15C and 15D are a series of graphs showing cytokine release and CD4+ T cell and CD8+ T cell proliferation from cytotoxicity assay of bispecific antibodies compared to controls and benchmarks in a tumor cell and PBMCs co-culture assay. Raji- GFP-luc tumor cells were treated w'ith PBMCs at 15: 1 E:T ratio with a serial dilution of biologies for 72 h. CD4+ T ceils and CD8+ T cells were counted by flow cytometry and the cytokine levels were measured by standard EL1SA. CD4+ T cell levels (FIG. ISA); CD8+ T cell levels (FIG. 15B), IL-2 levels (FIG. 15C); IFNy levels (FIG. 15D).

[0040] FIGS. 16A and 16B are a series of graphs showing a cytotoxicity assay of bispecific antibodies compared to controls and benchmarks in a DLBCL tumor cell line, Toledo, and PBMCs co-culture assay. Toledo tumor cells were treated with PBMCs at 15: 1 E:T ratio with a serial dilution of biologies for 72 h. Tumor cells (FIG. 16A) and CD 19+ B cells (FIG. 16B) were counted by flow cytometry. [0041] FIGS. 17A, 17B, 17C and 17D are a series of graphs showing cytokine induction and CD4+ T cell and CD8+ T cell proliferation from cytotoxicity' assay of bispecific antibodies compared to controls and benchmarks in a DLBCL tumor cell, Toledo, and PBMCs co-culture assay. Toledo tumor ceils were treated with PBMCs at 15: 1 E:T ratio with a serial dilution of biologies for 72 h. CD4+ T cells (FIG. 17A) and CD8+ T cells (FIG. 17B) were counted by flow cytometry and the IL-2 (FIG. 17C) and IFN_y (FIG. 17D) cytokine levels were measured by standard EL1SA.

[0042] FIG. 18A is a graph showing CD20 expression level on tumor cell lines NALM-6- GFP-luc (CD201ow) and JeKo-l -GFP (CD20high) determined by flow cytometry'.

[0043] FIG. 18B and FIG. 18C are a series of graphs showing a cytotoxicity assay of bispecific antibodies compared to controls and benchmarks in a ALL tumor cell line, NALM-6, and PBMCs co-culture assay. NALM-6 tumor cells were treated with PBMCs at 15: 1 E:T ratio with a serial dilution of biologies for 96 h. Tumor cells (FIG. 18B) and CD 19+ B cells (FIG. 18C) were counted by flow' cytometry.

[0044] FIG. 18D is a graph showing tumor growth inhibition of a low-antigen density' ALL tumor cell line, NALM-6, in an activated human T cells and tumor cells co-engraftment study in a xenograft NSG mouse model. At day 0, 2x10⁶ activated T cell and 10x10⁶ NALM-6-GFP-luc tumor cells were implanted subcutaneously with intravenous dosing of bispecific antibodies at day' 0 and 7 days apart for 3 weeks.

[0045] FIGS. 19A and 19B are a series of graphs showing single dose pharmacokinetics profile EIP0929 in NSG mice at 0.5 mg/kg dose (FIG. 19A) and 4 mg/kg dose (FIG. 19B). [0046] FIGS. 20A, 20B and 20C are a series of graphs showing EIP0929 induced complete B cell depletion in the periphery' of cynomolgus monkeys within 6 hours of 1st dose with expected T cell margination followed by T cell increases in subsequent doses. FIG. 20A shows Changes m B Cell Counts in PBMC. B cells were depleted in tire periphery within 6 hours of EIP0929 administration at all dose levels. Sustained B cell depletion for 168 hours post first dose was observed at 150 pg/kg. B cell levels remained below -15% of pre-dose levels at 150 pg/kg and below 35% at 10 and 50 pg/kg doses at the end of study. Sustained B cells depletion at 150 μg/kg is within 5-fold of the reported sustained B cell depletion for Glofltamab in NHP (30 μg/kg) 2 EIP0929 was significantly more potent than Epcoritamab doses reported for sustained B ceil depletion at 1 mg/kg.² FIGS. 20B and 20C shows T Cells Changes in PBMC. Expected CD8 (FIG. 20B) and CD4 (FIG. 20C) T cell margination was observed after the first dose in all dose cohorts and trended back towards pre-dose levels at 48 hours post dose. Increases in T cell numbers was observed at 50 pg/kg and 150 μg/kg after second and third dosing.

[0047] FIGS. 21A, 21B and 21C are a series of graphs showing that EIP0929 at 150 pg/kg induced near complete B cell depletion in lymph node and spleen in Cynomolgus monkeys. B Cell Depletion in the Mesenteric Lymph Node (FIG. 21 A) and Spleen (FIG. 21B) are shown. Dose-dependent B ceil depletion in the mesenteric lymph node and spleen was detected by flow cytometry at the end of study. IHC of Mesenteric Lymph Node and Spleen is shown (FIG. 21 C). IHC with an alternate B cell surface marker, CD 19 (instead of CD20 used in the flow cytometry), confirmed marked tissue B cell depletion in both the mesenteric lymph node and spleen.

[0048] FIGS. 22A and 22B are a series of graphs showing IL-6 and lower levels of IFNy and TNFa excursions were observed after first dose. IL-6 excursion within 6 hours of dosing was observed (FIG. 22A). Average IL-6 levels of -6000 pg/mL were observed within 6 hours of 50 and 150 pg/kg doses EIP0929 administrations that were trended towards baseline at 24 hours post dose. Reported levels after Glofitamab or Epcoritamab administration at doses which resulted in sustained peripheral B cell depletions were about 1.5- to 2-fold higher than EIP0929. IFNγ, TNFα and IL-2 excursions post first dose was observed (FIG. 22B). Lower levels of IFNy and TNFa excursion at 2 hours post first EIP0929 50 and 150 pg/kg doses were also observed. Reported levels after Epcoritamab administration at the dose which resulted in sustained peripheral B ceil depletions (intra venus (IV) and subcutaneous (SC) at I mg/kg were about 2.5- to 5-fold higher than EIP0929. The levels of IL-2 at 2 hours post EIP0929 administration are similar to the reported IL-2 levels after Epcoritamab at 1 mg/kg administration (SC) which could indicate that EIP0929 induced similar T cell activations with lower cytokine excursions compared to optimal SC administration of Epcoritamab.

[0049] FIG. 23 is a graph showing pharmacokinetics profiles of EIP0929 in Cynomolgus monkey determined from plasma concentration-time curves after single intravenous administration of EIP0929 at three different doses and monitoring plasma concentration up to 7 days.

DETAILED DESCRIPTION [0050] T cell retargeting (or T cell redirecting) bispecific antibodies is a class of therapeutics, capable of recruiting T cells to tumor cells and inducing tumor-specific (but MHC -independent) activation of T cell effector activities. The present disclosure is directed toward, T cell retargeting bispecific antibodies comprising an antigen binding domain that targets a CD3 portion of the T cell receptor for T cell recruitment, and an antigen bindi ng domain that targets a CD20 antigen. This targeting design promotes the recruitment of T cell and positions it in close contact with a CD20-expressing cell, resulting in the formation of an immunological synapse, local T cell activation and the subsequent destraction of the target cell, such as but not limited to a cancer cell, by perforin and granzyme released from T cell cytotoxic granules.

[0051] As the CD3 binding affinity of the T-cell retargeting bispecific antibodies is crucial for recruitment of T cells, the present invention also relates to the generation of a panel of antibodies that display different binding affinities. The affinity of the CD3 arm of a bispecific antibody can significantly modify the functional activity of the bispecific antibody. Thus, it is desirable and advantageous to have bispecific antibodies with varied affinities.

[0052] Additionally, bispecific antibodies disclosed herein may have a cytokine or costimulatory molecule fusion peptide that acts as antagonist to inhibit or block deleterious interactions or as an agonist to mimic or enhance physiological responses. Physiological responses include but are not limited to T-cell activation, T-cell proliferation, T-cell persistence and prevention of T-cell exhaustion. These properties are advantageous over conventional anti-CD20/anti-CD3-bispecific antibodies or tumor targeted co-stimulatory receptor agonists which do not optimally activate T-cells and induce (or promote) T-cell dysfunction. In accordance, cytokine and/or costimulatory fusion peptides are advantageous for enhancing the therapeutic potential of bispecific antibodies. In some embodiments, the costimulatory molecule of the anti-CD20/anti-CD3 bispecific antibodies of the present disclosure is CD58, or a fragment thereof.

[0053] The T cell retargeting bispecific antibodies of the disclosure have advantageous functional and biophysical properties relative to current anti-CD20/anti-CD3 clinical therapies (e.g. glofitamab and epcoritamab). Exemplary properties include: 1) increased thermal stability and half life which result in improved production capacity and yield, 2) sustained and complete tumor killing capacity and B cell depletion at lower doses, 3) increased T cell proliferation, activation and expansion at lower doses, 4) improved safety due to reduced inflammatory cytokine release, 5) improved pharmacokinetic profile and 6) efficacy in multiple modes of delivery (e.g., intravenous and subcutaneous). These properties of the bispecific antibodies of the disclosure, alone or in combination are advantageous over existing therapies for therapeutic application.

[0054] Bispecific or multispecific T cell retargeting agents of the disclosure share the druglike properties of human monoclonal antibodies. Further, T-cell retargeting bispecific and multispecific antibodies are advantageous over other existing therapies (e.g. CAR-T therapies) because it provides an off-the-shelf product with a high safety profile (e.g. mitigation of cytokine release syndrome and reduced levels of tonic signaling leading to T- cell dy sfunction) and the possibility of dose titration and escalation.

[0055] ANTIBODY COMPOSITIONS AND STRUCTURES

[0056] The present disclosure provides an antibody comprising the following domain structure: a) a first heavy chain polypeptide (H1) comprising a variable region (VH1), and a constant region (CH1) having a constant region 1 domain (CH1H1 or CH1__H1), a hinge region (H1H), a constant region 2 domain (CH1m or CH1 H2) and a constant region 3 domain (CHIH3 or CH1 _ H3 ); and a first light chain polypeptide (L1 ) comprising a variable region (VL1) and a constant region (CL1), and b) a second heavy chain polypeptide (H2) comprising a variable region (VH2), and a constant region (CH2) having a constant region 1 domain (CH2HI or CH2 H1), a hinge region (H2H), a constant region 2 domain (CH2H2 or CH2 _H2) and a constant region 3 domain (CH2H3 or CH2 _ H3 ); and second light chain polypeptide (L2) comprising a variable region (VL2) and a constant region (CL2). A schematic diagram of the antibody structure of the disclosure is shown in FIGS. 1A-1E.

[0057] As used herein, the term “antibody” refers to an immunoglobulin (Ig) molecule and immunologically active portions of an immunoglobulin molecule, i.e., molecules that contain an antigen binding site that specifically binds (immunoreacts with) an antigen. By “specifically bind” or “immunoreacts with” “or directed against” is meant that the antibody reacts with one or more antigenic determinants of the desired antigen and does not react with other polypeptides or binds at much lower affinity (K_d > 10^-6). Antibodies include, but are not limited to, polyclonal antibodies, monoclonal antibodies, chimeric antibodies. The antibody may be from recombinant sources and/or produced in transgenic animals.

[0058] The basic antibody structural unit is known to comprise a tetramer. Each tetramer is composed of two identical pairs of polypeptide chains, each pair having one “light” (about 25 kDa) and one “heavy” chain (about 50-70 kDa). The amino-terminal portion of each chain includes a variable region of about 100 to 110 or more amino acids primarily responsible for antigen recognition. The carboxy-terminal portion of each chain defines a constant region primarily responsible for effector function.

[0059] In general, antibody molecules obtained from humans relate to any of the classes IgG, IgM, IgA, IgE and IgD, which differ from one another by the nature of the heavy chain present in the molecule. Certain classes have subclasses as well, such as IgGl, IgG2, IgG4 and others. Furthermore, in humans, the light chain may be a kappa chain or a lambda chain. Accordingly, in one embodiment, the antibody disclosed herein is an IgG antibody. [0060] Antibodies may be purified by well-known techniques, such as affinity chromatography using protein A or protein G, which provide primarily the IgG fraction of immune serum. Subsequently, or alternatively, the specific antigen which is the target of the immunoglobulin sought, or an epitope thereof, may be immobilized on a column to purify the immune specific antibody by immunoaffinity chromatography. Purification of immunoglobulins is discussed, for example, by D. Wilkinson (The Scientist, published by The Scientist, Inc., Philadelphia PA, Vol. 14, No. 8 (April 17, 2000), pp. 25-28).

[0061] The term "antibody fragment" as used herein is intended to include without limitation, Fv, Fab, Fab', F(ab')2, scFv, dsFv, ds-scFv, dimers, minibodies, diabodies, and multimers thereof, multispecific antibody fragments and Domain Antibodies. Antibodies can be fragmented using conventional techniques. For example, F(ab')2 fragments can be generated by treating the antibody with pepsin. The resulting F(ab')2 fragment can be treated to reduce disulfide bridges to produce Fab' fragments. Papain digestion can lead to the formation of Fab fragments. Fab, Fab' and F(ab’)2, scFv, dsFv, ds-scFv, dimers, minibodies, diabodies, bispecific antibody fragments and other fragments can also be synthesized by recombinant techniques.

[0062] Techniques can be adapted for the production of single-chain antibodies specific to an antigenic protein of the disclosure (see e.g., U.S. Patent No. 4,946,778). In addition, methods can be adapted for the construction of Fab expression libraries (see e.g., Huse, et al., 1989 Science 246: 1275-1281) to allow rapid and effective identification of monoclonal Fab fragments with the desired specificity for a protein or derivatives, fragments, analogs or homologs thereof.

[0063] As used herein, the term “epitope” refers to the site on an antigen that is recognized by the antibodies and fragments disclosed herein. The term “epitope” includes any protein determinant capable of specific binding to an immunoglobulin. Epitopic determinants usually consist of chemically active surface groupings of molecules such as amino acids or sugar side chains and usually have specific three-dimensional structural characteristics, as well as specific charge characteristics. An antibody is said to specifically bind an antigen when the dissociation constant is < 1 micromolar; e.g., < 100 nM, preferably < 10 nM and more preferably < 1 nM.

[0064] Bispecific antibodies are antibodies that have binding specificities for at least two different antigens. The present disclosure provides a bispecific antibody having a first antigen binding region that binds to a first antigen (e.g. CD3) and a second antigen binding region that binds to a second antigen (e.g. disease associated antigen )

[0065] Antibodies with more than two valencies are also contemplated. For example, trispecific antibodies can be prepared. Tutt et al., J. Immunol. 147:60 (1991).

[0066] ANTIBODY VARIANTS

[0067] In some embodiments, amino acid sequence variants of the antibodies provided herein are contemplated. For example, it may be desirable to improve the heavy chain heterodimerization, light chain heterodimerization, binding affinity, and/or other biological properties of the antibody. Amino acid sequence variants of an antibody may be prepared by introducing appropriate modifications into the nucleotide sequence encoding the antibody, or by peptide synthesis. Such modifications include, for example, deletions from, and/or insertions into and/or substitutions of residues within the amino acid sequences of the antibody. Any combination of deletion, insertion, and substitution can be made to arrive at the final construct, provided that the final construct possesses the desired characteristics (e.g., light chain heterodimerization, heavy chain heterodimerization, antigen binding).

[0068 ] Amino acids may be grouped according to common side-chain properties:

(1) hydrophobic: Norleucine, Met, Ala, Vai, Leu, lle;

(2) neutral hydrophilic: Cys, Ser, Thr, Asn, Gin;

(3) acidic (negatively charged): Asp, Gin;

(4) basic (positively charged): His, Lys, Arg;

(5) residues that influence chain orientation: Gly, Pro; (6) aromatic: Trp, Tyr, Phe.

[0069] Functional variants of the antibody or antigen-binding fragments described herein are also encompassed by the present disclosure. The term "functional variant" as used herein includes modifications or chemical equivalents of the amino acid and nucleic acid sequences disclosed herein that perform substantially the same function as the polypeptides or nucleic acid molecules disclosed herein in substantially the same way. For example, functional variants of polypeptides disclosed herein include, without limitation, conservative amino acid substitutions.

[0070] A "conservative amino acid substitution" as used herein, is one in which one ammo acid residue is replaced with another amino acid residue that change an amino acid to a different amino acid with similar biochemical properties (e.g. charge, hydrophobicity and size). Variants of polypeptides also include additions and deletions to the polypeptide sequences disclosed herein. In addition, variant nucleotide sequences include analogs and derivatives thereof. A variant of the binding proteins disclosed herein include proteins that bind to the same antigen or epitope as the binding proteins.

[0071] In some embodiments, the charged amino acid residue is a naturally occurring amino acid or a non-naturally occurring amino acid. In some embodiments, the naturally occurring charged amino acid residue is an arginine, a lysine, a histidine, a glutamic acid or an aspartic acid.

[0072] Light Chain and Heavy Chain Substitution Variants

[0073] To generate a substantially homogeneous population of bispecific antibodies with the correct pairing of heavy chain and light chains (i.e. cognate pairing or heterodimerization of a light chain with the heavy chain necessary to form the variable domain or antigen binding domain of the original antibody ), the first heavy chain polypeptide (H1 ) has a strong preference for binding with the first light chain polypeptide (L1) relative to the second light chain polypeptide (L2); and the second heavy chain polypeptide (H2) has a strong preference for binding with the second light chain polypeptide (L2) relative to first light chain polypeptide (L1). In addition, the first heavy chain polypeptide (H1 ) and the second heavy chain polypeptide (H2) have a stronger preference for heterodimerization than homodimerization (i.e. heavy chain heterodimerization) . [0074] Antibody vanants having one or more amino acid substitutions are provided herein. Exemplar}/ substitutional mutagenesis sites include the charged substitution pairs shown in Tables 1-6.

[0075] Table 1. Kappa Light Chain and Heavy Chain - Constant Domain Mutations Pairs

All position information is reported using the EU numbering scheme

Wild type (WT) indicates the natural ammo acid at the indicated position

Charge pairs with negative and positive charge residues could be reversed between heavy and light chains, where D or E (negative charge) are replaced by K or R (positive charge) and cognate chain K or R (positive charge) are replaced by D or E (negative charge).

[0076] Table 2. Kappa Light Chain and Heavy Chain - Variable Domain Mutations Pairs

All position information is reported using the Kabat numbering scheme

Wild type (WT) indicates the natural ammo acid at the indicated position

Charged pairs with negative and positive charged residues could be reversed between heavy and light chains, where D or E (negative charged) are replaced by K or R (positive charged) and cognate chain K or R (positive charged) are replaced by D or E (negative charged).

[0077] Table 3. Lambda Light Chain and Heavy Chain - Constant Domain Mutations Pairs

All position information is reported using the EU numbering scheme

[0078] Table 4. Lambda Light Chain and Heavy Chain - Variable Domain Mutations Pairs

All position information is reported using the Kabat numbering scheme

[0079] Table 5. Kappa Constant Chain Cysteine Mutation Pairs

All position information is reported using the EU numbering scheme

[0081] In some embodiments, anti-CD3/anti-CD20 bispecific antibod ies of the present disclosure comprise antibody variants comprise substitutions in the variable heavy, variable light, constant heavy or constant light chain domains of the anti-CD3 or the anti-CD20 aims or both. Exemplary' variants include those described in PCT Application No.

PCT/US2023/064728 and PCT Publication No. WO2019/104075, which are incorporated by reference herein in their entirety.

[0082] In some embodiments, the antibody variant comprises the ‘'light chain pairing mutation set D” comprising the following substitutions: a) the heavy chain and light chain of the anti-CD3 arm comprise the following: i) the amino acid at position 39 (Kabat numbering) of the VH1 is a K and the amino acid at position 38 (Kabat numbering) of the VL1 is a D; ii) the amino acid at position 147 (EU numbering) of the CHI H1 is a K and the amino acid at position 131 (EU numbering) of the CL1 is a D; iii) the amino acid at position 173 (EU numbering) of the CH1__H1 is a C and the amino acid at position 162 (EU numbering) of the CL1 is a C; iv) the amino acid at position 220 (EU numbering) in the H1H is a S and the amino acid at position 214 (EU numbering) of the CL1 is a S; and b) the heavy chain and light chain of the anti-CD20 arm comprise the following: i)the amino acid at position 39 (Kabat numbering) of the VH2 is a D and the amino acid at position 38 (Kabat numbering) of the VL2 is a K; and ii) the amino acid at position 147 (EU numbering) of the CH2_H1 is a D and the amino acid at position 180 (EU numbering) of the CL2 is a R.

[0083] In some embodiments, the antibody variant comprises the “light chain pairing mutation set D” comprising the following substitutions: a) the heavy chain and light chain of the anti-CD20 arm comprise the following: i) the amino acid at position 39 (Kabat numbering) of the VH1 is a K and the amino acid at position 38 (Kabat numbering) of the VL1 is a D; ii) the amino acid at position 147 (EU numbering) of the CHI HI is a K and the amino acid at position 131 (EU numbering) of the CL1 is a D; iii) tire amino acid at position 173 (EU numbering) of the CH1_H1 is a C and the amino acid at position 162 (EU numbering) of the CL1 is a C; iv) the amino acid at position 220 (EU numbering) in the H1H is a S and the amino acid at position 214 (EU numbering) of tire CL1 is a S; and b) the heavy chain and light chain of the anti-CD3 arm comprise the following: i)the amino acid at position 39 (Kabat numbering) of the VH2 is a D and the amino acid at position 38 (Kabat numbering) of the VL2 is a K; and ii) the amino acid at position 147 (EU numbering) of the CH2_HI is a D and the amino acid at position 180 (EU numbering) of the CL2 is a R. [0084] It can be desirable to modify an antibody di sclosed herein with respect to effector function, so as to enhance, e.g., the effectiveness of the antibody in treating diseases and disorders. For example, cysteine residue(s) can be introduced into the Fc region, thereby allowing interchain disulfide bond formation in this region. The homodimeric antibody thus generated can have improved internalization capability and/or increased complement- mediated cell killing and antibody-dependent cellular cytotoxicity (ADCC). (See Caron et al., J Exp Med., 176: 1191-1195 (1992) and Shopes, J. Immunol., 148:2918-2922. (1992)). Alternatively, an antibody can be engineered that has dual Fc regions and can thereby have enhanced complement lysis and ADCC capabilities. (See Stevenson et al., Anti-Cancer Drug Design, 3:219-230 (1989)).

[0085] Certain antibody variants with improved or diminished binding to FcRs are described. (See, e.g., U.S. Patent No. 6.737,056; WO 2004/056312, and Shields et al., J. Biol. Chem. 9(2): 6591-6604 (2001).)

10086] In some embodiments, an antibody variant comprises an Fc region with one or more amino acid substitutions which improve ADCC, e.g., substitutions at positions 298, 333, and/or 334 of the Fc region (EU numbering of residues).

[0087] In some embodiments, alterations are made in the Fc region that result in altered (i.e., either improved or diminished) Clq binding and/or Complement Dependent Cytotoxicity (CDC), e.g., as described in US Patent No. 6,194,551 , WO 99/51642, and Idusogie et al. J. Immunol. 164: 4178-4184 (2000).

10088] Antibodies with increased half lives and improved binding to the neonatal Fc receptor (FcRn), which is responsible for the transfer of maternal IgGs to the fetus (Guyer et al., J. Immunol. 1 17:587 (1976) and Kim et al., J. Immunol. 24:249 (1994)), are described in US2005/0014934A1 (Hinton et al.). Those antibodies comprise an Fc region with one or more substitutions therein winch improve binding of the Fc region to FcRn. Such Fc variants include those with substitutions at one or more of Fc region residues: 238, 256, 265, 272, 286, 303, 305, 307, 311 , 312, 317, 340, 356, 360, 362, 376, 378, 380, 382, 413, 424 or 434, e.g., substitution of Fc region residue 434 (US Patent No. 7,371 ,826). See also Duncan & Winter, Nature 322:738-40 (1988); U.S. Patent No. 5,648,260; U.S. Patent No. 5,624,821 ; and WO 94/29351 concerning other examples of Fc region variants.

[0089] In some embodiments, antibodies may comprise a substitution mutation in the Fc region that reduces effector function. In some embodiments, tire substitution mutation is an aglycosylation site mutation. In some embodiments, the aglycosylation site mutation is at amino acid residue 297 and amino acid substitutions at residues 234, 235, 265 and 331 (EU numbering) to disrupt the Fc receptor binding interface. In some embodiments, the aglycosylation site mutation reduces effector function of the antibody.

[0090] In some embodiments, i) the H1H and/or the H2H has an A at positions 234 and 235 (EU numbering); or ii) the H1H and/or the H2H has an A at positions 234, 235 and 237 (EU numbering) iii) the H1H and/or the H2H has an A at positions 234 and 235 and G at position 329 (EU numbering). In some embodiments, i) the CH1_H3 and/or the CH2_H3 has an A at position 297 (EU numbering) ii) the CH1 H3 and/or the CH2 _ H3 has a G at position 297 (EU numbering); or iii) the CH1JH3 and/or the CH2 _ H3 has a S at position 297 (EU numbering). In some embodiments, the CH1 _ H3 and/or the CH2___H3 has an S at position 331 (EU numbering).

[0091] The use of knobs into holes as a method of producing multispecific antibodies is well known m the art. See U.S. Pat. No. 5,731,168 granted 24 Mar. 1998 assigned to Genentech, PCT Pub. No. W02009089004 published 16 Jul. 2009 and assigned to Amgen, and US Pat. Pub. No. 20090182127 published 16 Jul. 2009 and assigned to Novo Nordisk A/S. See also Marvin and Zhu, Acta Pharmacologica Sincia (2005) 26(6):649-658 and Kontermann (2005) Acta Pharacol. Sin., 26: 1-9.

[0092] A “'protuberance” refers to at least one amino acid side chain which projects from the interface of a first polypeptide and is therefore positionable in a compensatory cavity in the adjacent interface (i.e. the interface of a second polypeptide) so as to stabilize the heteromultimeric antibody, and thereby favor heteromultimeric antibody formation over homomultimeric antibody formation, for example. The protuberance may exist in the original interface or may be introduced synthetically (e.g. by altering nucleic acid encoding the interface). Normally, nucleic acid encoding tire interface of tire first polypeptide is altered to encode the protuberance. To achieve this, the nucleic acid encoding at least one “original” amino acid residue in the interface of the first polypeptide is replaced with nucleic acid encoding at least one “import” ammo acid residue which has a larger side chain volume than the original amino acid residue. It will be appreciated that there can be more than one original and corresponding import residue. Tire upper limit for the number of original residues which are replaced is the total number of residues in the interface of the first polypeptide.

[0093] The preferred import residues for the formation of a protuberance are generally naturally occurring amino acid residues and are preferably selected from arginine (R), phenylalanine (F), tyrosine (Y) and tryptophan (W). Most preferred are tn plophan and tyrosine. In one embodiment, the original residue for the formation of the protuberance has a small side chain volume, such as alanine, asparagine, aspartic acid, glycine, serine, threonine or valine. Exemplary'- amino acid substitutions in the CH1 _ H3 or CH2 _ H3 domain for forming the protuberance include without limitation the T366W substitution. [0094 ] A “cavity” refers to at least one amino acid side chain which is recessed from the interface of a second polypeptide and therefore accommodates a corresponding protuberance on the adjacent interface of a first polypeptide. The cavity may exist in the original interface or may be introduced synthetically (e.g. by altering nucleic acid encoding the interface). Normally, nucleic acid encoding the interface of the second polypeptide is altered to encode the cavity. To achieve this, the nucleic acid encoding at least one “original” ammo acid residue in the interface of the second polypeptide is replaced with DNA encoding at least one “import” amino acid residue which has a smaller side chain volume than the original amino acid residue. It will be appreciated that there can be more than one original and corresponding import residue. The upper limit for the number of original residues which are replaced is the total number of residues in tire interface of the second polypeptide. The side chain volumes of the various amino residues are shown in Table 1 above. The preferred import residues for the formation of a cavity are usually naturally occurring amino acid residues and are preferably selected from alanine (A), serine (S), threonine (T) and valine (V). Most preferred are serine, alanine or threonine. In one embodiment, the original residue for the formation of the cavity has a large side chain volume, such as tyrosine, arginine, phenylalanine or tryptophan. Exemplary amino acid substitutions in the CH1 _ H3 or CH2___H3 domain for generating the cavity include without limitation the T366S, L368A, Y407A, Y407T and Y407V substitutions. In some embodiments, the knob half-antibody comprises T366W substitution, and the hole half-antibody comprises the T366S/L368A/Y407V substitutions.

[0095] In some embodiments, the antibody variant comprises the following substitutions: the CH1 H3 has a C at position 349, an S at position 366, an A at position 368 and a V at position 407 (EU numbering); and the CH2 _H3 has a C at position 354 and a W at position 366 (EU numbering).

[0096] In some embodiments, the antibody variant comprises the following substitutions: the CH2 _ H3 has a C at position 349, an S at position 366, an A at position 368 and a V at position 407 (EU numbering); and the CHI 113 has a C at position 354 and a W at position 366 (EU numbering).

[0097] In some embodiments, the antibody variant comprises the following substitutions: the CH1_H3 has a C at position 354, an S at position 366, an A at position 368 and a V at position 407 (EU numbering); and the CH2__H3 has a C at position 349 and a W at position 366 (EU numbering);

[0098] In some embodiments, the antibody variant comprises the following substitutions: the CH2 _ H3 has a C at position 354, an S at position 366, an A at position 368 and a V at position 407 (EU numbering); and the CH 1 _H3 has a C at position 349 and a W at position 366 (EU numbering).

[0099] CLUSTER OF DIFFERENTIATION 3 (CD3)

[00100] The present disclosure provides a bispecific antibody comprising a first antigen binding domain that binds to a CDS expressed on a T-cell, and a second antigen binding domain that binds to a CD20 antigen on the surface of a cancer cell. Antibodies of the invention are useful, for example, for treating or delaying the progression of a cell proliferative disorder (e.g., cancer expressing CD20).

[00101] The term "‘duster of differentiation 3” or “CD3,” as used herein, refers to any native CD3 from any vertebrate source, including mammals such as primates (e.g. humans, cynomolgus monkey) and rodents (e.g., mice and rats), unless otherwise indicated, including, for example, CD3ε, CD3y, CD3a, and CD3p chains. CD3 is a cell surface complex expressed on T cells in association with the T cell receptor. The CD3 complex is required for the activation of CD8+ and CD4+ T lymphocytes. It is formed of three different but highly related chains: one CD3 gamma chain, one CD3 delta chain, and two CD3 epsilon chains, which associate with each other to form a CD3 epsilon/gamma heterodimer, and a CD3 epsilon/delta heterodimer. The two C'D3 heterodimers, together with the T ceil receptor (TCR) and the signal-transducing zeta chain homodimer form the T cell receptor complex.

[00102] The term encompasses ‘‘full-length” unprocessed CD3 (e.g., unprocessed or unmodified CD3e or CD3γ), as well as any form of CD3 that results from processing in the cell. The term also encompasses naturally occurring variants of CD3, including, for example, splice variants or allelic variants. CD3 includes, for example, human CD3ε protein (NCBI RefSeq No. NP 000724), which is 207 amino acids in length.

[00103] In some embodiments, the invention provides isolated antibodies that bind to CD3. In some embodiments, the invention provides antibodies that bind to CD3ε. In some instances the anti-CD3ε antibody binds to a human CD3ε polypeptide or a cynomolgus monkey (cyno) CD3ε polypeptide. In some instances, the human CD3 polypeptide or tire cyno CD3 polypeptide is a human CD3ε polypeptide (SEQ ID NO: 419) or a cyno CD3e. polypeptide (SEQ ID NO: 420), respectively. In some instances, the anti-

CD3 antibody binds to an epitope within a fragment of CD3ε (e.g., human CD3ε) consisting of amino acid residues 1-26 or amino acid residues 1-27 of human CD3e (SEQ ID NO: 419).

[00104] A useful method for identification of residues or regions of an antibody that may be targeted for mutagenesis is called “alanine scanning mutagenesis” as described by Cunningham and Wells (1989) Science, 2,44: 1081-1085. In this method, a residue or group of target residues (e.g., charged residues such as Arg, Asp, His, Lys, and Glu) are identified and replaced by a neutral or negatively charged amino acid (e.g., alanine or polyalanine) to determine whether the interaction of the antibody with antigen is affected. Further substitutions may be introduced at the amino acid locations demonstrating functional sensitivity to the initial substitutions. Alternatively, or additionally, a crystal structure of an antigen-antibody complex is used to identify contact points between the antibody and antigen. Such contact residues and neighboring residues may be targeted or eliminated as candidates for substitution. Variants may be screened to determine whether they contain the desired properties.

[00105] In some embodiments, alanine scanning mutagenesis was performed on the “SP34” anti- CD3ε antibody to produce affinity modulated anti-CD3ε antibodies of the invention. [00106] In some embodiments, the first antigen binding region that binds to CD3 comprises any one of the VH and VL sequences listed in Table 7. In Table 7, the underlined sequences are CDR sequence according to Kabat and the bolded sequences are CDR sequences according to Chothia.

[00107] In some embodiments, a anti-CD3 antibody of the disclosure comprises: a) a heavy chain variable region (VH) comprising a VH complementarity determining region 1 (VH_CDR1), a VH complementarity determining region 2 (VH_CDR2) and a VH complementarity determining region 3 (VH_CDR3); and b) a light chain variable region (VL) comprising a VL complementarity determining region 1 (VL_CDR1), a VL complementarity determining region 2 (VL_CDR2) and a VL complementarity determining region 3 (VL_CDR3). Tables 8 and 9 provide exemplary of CDR sequences of the anti-CD3 antibodies provided herein. [00108] Table 7. Anti-CD3 Variable Heavy Chain and Variable Light Chain Domains

[00109] Table 8. Anti-CD3 Heavy Chain CDRs [00110] Table 9. Anti-CD3 Light Chain CDRs

[00111] Provided herein are bispecific antibodies comprising a first antigen binding domain that binds a first antigen (e.g. CD3) and a second antigen binding domain that binds to a second antigen (e.g. CD20). In some embodiments the bispecific antibody has the following structure: a first heavy chain polypeptide (H1) comprising a variable region (VH1), and a constant region (CH1) having a constant region 1 domain (CH1_H1), a hinge region (H1H), a constant region 2 domain (CH1 H2) and a constant region 3 domain (CH1 H3); and a first light chain polypeptide (L1) comprising a variable region (VL1) and a constant region (CL1), and a second heavy chain polypeptide (H2) comprising a variable region (VH2), and a constant region (CH2) having a constant region 1 domain (CH2 H1), a hinge region (H2H), a constant region 2 domain (CH2 H2) and a constant region 3 domain (CH2 _ H3 ); and second light chain polypeptide (L2) comprising a variable region (VL2) and a constant region (CL2). [00112] In some embodiments, the bispecific antibody of the disclosure comprises a first antigen binding domain (e.g. binding to CD3) comprising any one of the VH1 and VL1 sequences listed in Table 7. In Table 7, the underlined sequences are CDR sequence according to Kabat and the bolded sequences are CDR sequences according to Chothia. [00113] In some embodiments, the bispecific antibody of the disclosure comprises a first antigen binding domain (e.g. binding to CD3ε) comprising: a) a heavy chain variable region (VH1) comprising a VH complementarity determining region 1 (VH1 CDR1), a VH complementarity determining region 2 (VH1_ CDR2) and a VH complementarity determining region 3 (VH1 CDR3); and b) a light chain variable region (VL) comprising a VL complementarity determining region 1 (VL1 CDR1), a VL complementarity determining region 2 (VL1 CDR2) and a VL complementarity determining region 3 (VL1 CDR3). Tables 8 and 9 provide exemplary of CDR sequences of the anti-CD3 antibodies provided herein.

[00114] In some embodiments, the bispecific antibody comprises any one of the anti-CD3 antibodies of the disclosure. Exemplary' anti-CD3 antibodies of the invention include CD3- Al, CD3-A2, CD3-A3, CD3-A4, CD3-A5, CD3-A6, CD3-A7, CD3-A8, CD3-A9, CD3- A10, CD3-A11, CD3-A12 and CD3-A13.

[00115] In some embodiments, the binding affinity (KD) of the first antigen binding region of the bispecific antibody that binds to CD3 (e.g., human CD3ε (e.g., (SEQ ID NO: 419) or cynomolgus CD3g (e.g., (SEQ ID NO: 422)) is about 0.001 nM to about 5000 nM. In some embodiments, the binding affinity to CD3 is about 0.001 nM to about 0.01 nM, about 0.01 to about 0.1 nM or about 0.1 to about InM. In some embodiments, the binding affinity' is about 1 nM to about 1000 nM, about 10 nM to about 1000 nM or about 100nM to about 1000 nM.

[00116] In some embodiments, the binding affinity is about 50 nM to about 5000 nM, about 50 nM to about 4000 nM, about 50 nM to about 3000 nM, about 50 nM to about 2000 nM, about 50 nM to about 1000 nM, about 50 nM to about 900 nM, about 50 nM to about 800 nM, about 50 nM to about 700 nM, about 50 nM to about 600 nM, about 50 nM to about 500 nM, about 50 nM to about 400 nM, about 50 nM to about 300 nM, about 50 nM to about 200 nM, about 50 nM to about 100 nM or about 50 nM to about 500 nM. In some embodiments, the binding affinity is about 50 nM to about 200 nM. [00117] In some embodiments, the binding affinity is about 10 nM to about 2,0 nM, about 20 nM to about 30 nM, about 30 nM to about 40 nM, about 50 nM to about 60 nM, about 60 nM^ to about 70 nM, about 70 nM to about 80 nM, about 80 nM to about 90 nM, about 90 nM to about 100 nM, about 100 nM to about 110 nM, about 110 nM to about 120 nM, about 120 nM to about 130 nM, about 130 nM to about 140 nM, about 150 nM to about 160 nM, about 160 nM to about 170 nM, about 170 nM to about 180 nM, about 180 nM to about 190 nM, or about 190 nM to about 200 nM.

[00118] In some embodiments, the binding affinity is less than about 5000 nM, 4000 nM, 3000 nM, 2000 nM, 1000 nM, 900 nM, 800 nM, 700 nM, 600 nM, 500 nM, 400 nM, 300 nM, 275 nM, 250 nM, 225 nM, 200 nM, 175 nM, 150 nM, 125 nM, 100 nM, 90 nM, 80 nM, 70 nM, 60 nM, 50 nM, 40 nM, 30 nM, 20 nM, 10 nM, 9.5 nM, 9nM, 8.5 nM, 8 nM, 7.5 nM, 7 nM, 6.5 nM, 6 nM, 5 nM, 4.5 nM, 4 nM, 3.5 nM, 3 nM, 2.5 nM, 2 nM, 1.5 nM, 1 nM, or 0.5 nM.

[00119] CD20 ANTIGEN

[00120] The present disclosure provides a bispecific antibody comprising a first antigen binding domain that binds to a CD3 expressed on a T-cell, and a second antigen binding domain that binds to a CD20 antigen on the surface of a cancer cell. Antibodies of the invention are useful, for example, for treating or delaying the progression of a cell proliferative disorder (e.g., cancer expressing CD20).

[00121] “CD20” as used herein refers to the human B-lymphocyte antigen CD20 (also known as CD20, human B-lymphocyte -restricted differentiation antigen, B-lymphocyte surface antigen Bl, Leu-16, Bp35, BMS, and LFS; the sequence is characterized by the SwissProt database entry P 11836), which is a hydrophobic transmembrane protein with a molecular weight of approximately 35 kD located on pre-B and mature B lymphocytes. (Valentine, M. A., et al., J. Biol. Chem. 264(19) (1989 11282-11287: Tedder, T. F., et al, Proc. Natl. Acad. Sci. U.S.A. 85 (1988) 2,08-12; Stamenkovic, I., et al., J. Exp. Med. 167 (1988) 1975-80; Emfeld, D. A., et al, EMBO J. 7 (1988) 711-7; Tedder, T. F, et al, J. Immunol. 142 (1989) 2560-8). CD20 is found on the surface of greater than 90% of B cells from peripheral blood or lymphoid organs and is expressed during early pre-B cell development and remains until plasma cell differentiation. CD20 is present on both normal B cells as well as malignant B cells. In particular, CD20 is expressed on greater than 90% of B cell non-Hodgkin's lymphomas (NHL) (Anderson, K. C, et al., Blood 63(6) (1984) 1424- 1433) but is not found on hematopoietic stem cells, pro-B cells, normal plasma cells, or other normal tissues (Tedder, T. F, et al, J. Immunol. 135(2)( 1985) 973-979).

100122] The 85 amino acid carboxyl-terminal region of the CD20 protein is located within the cytoplasm. The length of this region contrasts with that of other B cell-specific surface structures such as IgM, IgD, and IgG heavy chains or histocompatibility antigens class II a or B chains, which have relatively short intracytoplasmic regions of 3, 3, 28, 15, and 16 amino acids, respectively' (Komaromy, M, et al., NAR 11 (1983) 6775-6785). Of the last 61 carboxy] -termin al amino acids, 21 are acidic residues, whereas only 2 are basic, indicating that this region has a strong net negative charge. The GenBank Accession No. is NP- 690605. It is thought that CD20 might be involved in regulating an early step(s) in the activation and differentiation process of B cells (Tedder, T. F, et al., Eur. J. Immunol. 16 (1986) 881-887) and could function as a calcium ion channel (Tedder. T. F, et al., J. Cell. Biochem. 14D (1990) 195).

[00123] The terms “CD20” and “'CD20 antigen” are used interchangeably herein, and include any variants, isoforms and species homologs of human CD20 which are naturally expressed by cells or are expressed on cells transfected with the CD20 gene. Binding of an antibody of the invention to the CD20 antigen mediate tire killing of cells expressing CD20 (e.g, a tumor cell) by inactivating CD20. The killing of the cells expressing CD20 may occur by one or more of the following mechanisms: Cell death/apoptosis induction, ADCC and CDC.

[00124] Synonyms of CD20, as recognized in the art, includ B-lymphocyte antigen CD20, B-lymphocyte surface antigen Bl, Leu-16, Bp35, BMS, and LF5.

[00125] The term “anti-CD20 antibody” according to the invention is an antibody that binds specifically to CD20 antigen. Depending on binding properties and biological activities of anti-CD20 antibodies to the CD20 antigen, two types of anti-CD20 antibodies (type I and type II anti-CD20 antibodies) can be distinguished according to Cragg, M. S, et al., Blood 103 (2004) 2738-2743; and Cragg, M. S, et al. Blood 101 (2003) 1045-1052.

[00126] The two different types of anti-CD20 antibodies differ significantly in their mode of CD20 binding and biological activities (Cragg, M. S, et al. Blood 103 (2004) 2738- 2743; and Cragg, M. S, et al. Blood 101 (2003) 1045-1052). Type I antibodies, as e.g. rituximab, are potent in complement mediated cytotoxicity, whereas type II antibodies, as e.g. Tositumomab (Bl), 11B8, AT80 or humanized B-Lyl antibodies, effectively initiate target cell death via caspase-independent apoptosis with concomitant phosphatidylserine exposure.

[00127] In some instances, CD20 may be expressed in low' copy number on the target cell (e.g. tumor cell). For example, in some instances, CD20 is expressed or present at less than 35,000 copies per target cell . In some embodiments, the low copy number cell surface CD20 is present between 100 and 35,000 copies per target cell; between 100 and 30,000 copies per target cell; between 100 and 25,000 copies per target cell; between 100 and 20,000 copies per target cell; between 100 and 15,000 copies per target cell; between 100 and 10,000 copies per target cell; between 100 and 5,000 copies per target cell; between 100 and 2,000 copies per target cell; between 100 and 1,000 copies per target cell; or between 100 and 500 copies per target cell. Copy number of the cell surface CD20 can be determined, for example, using a standard Scratchcard plot.

[00128] This disclosure provides an bispecific antibody comprising a first antigen binding region that binds CD3 and a second binding region that binds CD20.

[00129] In some embodiments the bispecific antibody has the following structure: a first heavy chain polypeptide (H1 ) comprising a variable region (VH1), and a constant region (CHI) having a constant region 1 domain (CHI H1 ), a hinge region (H1H), a constant region 2 domain (Cl 11 1 12) and a constant region 3 domain (CH1__H3); and a first light chain polypeptide (L1) comprising a. variable region (VL1) and a constant region (CL1), and a second heavy chain polypeptide (H2) comprising a variable region (VH2), and a constant region (Ci 12) having a constant region 1 domain (CH2 H1), a hinge region (H2H), a constant region 2 domain (CH2 H2) and a constant region 3 domain (CH2 _ H3 ); and second light chain polypeptide (L2) comprising a variable region (VL2) and a constant region (CL2).

[00130] For example, the second binding region that binds to CD20 can be derived from the binding regions of an anti-CD20 antibody. Exemplary anti-CD20 antibodies include but are not limited to obinutuzumab (GA101; GAZYVA® or GAZYVARO®), tositumomab, rituximab, ofatumumab, veltuzumab, ocaratuzumab, ocrelizumab, PRO 131921, ublituximab, HI47 IgG3 (ECACC, hybridoma), 2C6 IgGl (as disclosed in WO 2005/103081 ), 2F2 IgGl (as disclosed in WO 2004/035607 and WO 2005/103081) and

2H7 IgGl (as disclosed in WO 2004/056312). [00131] In some embodiments, a second antigen binding region that binds CD20 comprises any one of the VH and VL sequences listed in Table 10. In Table 10, the underlined sequences are CDR sequence according to Kabat and the bolded sequences are CDR sequences according to Chothia.

[00132] In some embodiments, the second binding region that binds to CD20 comprises: a) a heavy chain variable region (VH) comprising a VH complementarity determining region 1 (VH_CDR1), a VH complementarity determining region 2 (VH_CDR2) and a VH complementarity determining region 3 (VH_CDR3); and b) a light chain variable region (VL) comprising a VL complementarity determining region 1 (VL_CDR1), a VL complementarity determining region 2 (VL_CDR2) and a VL complementarity determining region 3 (VL_CDR3). Tables 11 and 12 provide exemplary of CDR sequences of the anti-CD20 antibodies.

[00133] Table 10. Anti-CD20 Variable Heavy Chain and Variable Light Chain Domains

[00134] Table 11. Anti-CD20 Heavy Chain CDRs

[00135] Table 12. Anti-CD20 Light Chain CDRs [00136] In some embodiments, the second antigen binding region comprises a) a heavy chain variable region (VH) comprising a i) a VH complementarity determining region 1 (VH_CDR1) comprising the amino acid sequence of SEQ ID NO: 459 or 33, ii) a VH complementarity determining region 2 (VH_CDR2) comprising the amino acid sequence of SEQ ID NO: 460 or 461 , iii) a VH complementarity determining region 3 (VH_CDR3) comprising the amino acid sequence of SEQ ID NO: 462; and b) a light chain variable region (VL) comprising a i) a VL complementarity determining region 1 (VL_CDR1) comprising the amino acid sequence of SEQ ID NO: 466, ii) a VL complementarity determining region 2 (VL_CDR2) comprising the amino acid sequence of SEQ ID NO: 467, iii) a VL complementarity determining region 3 (VL_CDR3) comprising the ammo acid sequence of SEQ ID NO: 468.

[00137] In some embodiments, the second antigen binding region comprises a VH region comprising a VH_CDR1 comprising the amino acid sequence of SEQ ID NO: 455, a VH_CDR2. comprising the amino acid sequence of SEQ ID NO: 457, and a VH_CDR3 comprising the amino acid sequence of SEQ ID NO: 458; and a VL region comprising a VL_CDR1 comprising the amino acid sequence of SEQ ID NO: 463, aVL_CDR2 comprising the amino acid sequence of SEQ ID NO: 464, and a VL_CDR3 comprising the amino acid sequence of SEQ ID NO: 465.

[00138] In some embodiments, the second antigen binding region comprises a VH region comprising a VH_CDR1 comprising the amino acid sequence of SEQ ID NO: 454, a VH_CDR2. comprising the amino acid sequence of SEQ ID NO: 456, and a VH_CDR3 comprising the amino acid sequence of SEQ ID NO: 458; and a VL region comprising a VL_CDR1 comprising the amino acid sequence of SEQ ID NO: 463, a VL_CDR2 comprising the amino acid sequence of SEQ ID NO: 464, and a VL_CDR3 comprising the amino acid sequence of SEQ ID NO: 465.

[00139] In some embodiments, the second antigen binding region comprises a VH region comprising the amino acid sequence shown in SEQ ID NO: 451 and a VL region comprising the amino acid sequence shown in SEQ ID NO: 450.

[00140] In some embodiments, the second antigen binding region comprises a VH region comprising a VH_CDR1 comprising the amino acid sequence of SEQ ID NO: 459, a VH_CDR2 comprising the amino acid sequence of SEQ ID NO: 460, and a VH_CDR3 comprising the amino acid sequence of SEQ ID NO: 462; and a VL region comprising a VL_CDR1 comprising the amino acid sequence of SEQ ID NO: 466, a VL_CDR2 comprising the amino acid sequence of SEQ ID NO: 467, and a VL_CDR3 comprising the amino acid sequence of SEQ ID NO: 468.

[00141] In some embodiments, the second antigen binding region comprises a VH region comprising a VH_CDR1 comprising the amino acid sequence of SEQ ID NO: 33, a VH_CDR2 comprising the amino acid sequence of SEQ ID NO: 461, and a VH_CDR3 comprising tire amino acid sequence of SEQ ID NO: 462; and a VL region comprising a VL_CDR1 comprising the amino acid sequence of SEQ ID NO: 466, a VL_CDR2 comprising the amino acid sequence of SEQ ID NO: 467, and a VL_CDR3 comprising the amino acid sequence of SEQ ID NO: 468.

[00142] In some embodiments, the second antigen binding region comprises a VH region comprising the amino acid sequence shown in SEQ ID NO: 453 and a VL region comprising the amino acid sequence shown in SEQ ID NO: 452.

[00143] EXEMPLARY BISPECIFIC ANTIBODIES THAT BIND TO CD3 AND CD20

[00144] Provided herein are bispecific antibodies comprising a first antigen binding domain that binds a first antigen (e.g. CD3ε) and a second antigen binding domain that binds to a CD20. In some embodiments the bispecific antibody has the following structure: a first heavy chain polypeptide (H1) comprising a variable region ( VH 1), and a constant region (CHI) having a constant region 1 domain ( CH1_H1), a hinge region (H1H), a constant region 2 domain (CHI EI2) and a constant region 3 domain ( CH1_H3); and a first light chain polypeptide (L1 ) comprising a variable region (VL1) and a constant region (CL1), and a second heavy chain polypeptide (H2) comprising a variable region (VH2), and a constant region (CH2) having a constant region 1 domain (CH2 H1), a hinge region (H2H), a constant region 2 domain (CH2 H2) and a constant region 3 domain (CH2_H3); and second light chain polypeptide (L2) comprising a variable region ( VL2) and a constant region (CL2).

[00145] In some embodiments, the bispecific antibody of the disclosure comprises a first antigen binding domain (e.g. binding to CD3ε) comprising: a) a heavy chain variable region (VH1) comprising a VH complementarity determining region 1 (VH 1_CDR1), a VH complementarity determining region 2 (VH 1_CDR2) and a VH complementarity determining region 3 (VH1 CDR3); and b) a light chain variable region (VL) comprising a VL complementarity determining region 1 (VL1 CDR1), a VL complementarity determining region 2 (VL1 CDR2) and a VL complementarity determining region 3 (VL1 CDR3); and a second antigen binding domain (e.g. binding to CD20) comprising: a) a heavy chain variable region (VH2) comprising a VH complementarity determining region 1 (VH2_CDR1), a VH complementarity determining region 2 (VH2_CDR2) and a VH2 complementarity determining region 3 (VH2 CDR3); and b) a light chain variable region (VL2) comprising a VL complementarity determining region 1 (VL2 CDR1), a VL complementarity determining region 2 (VL2 CDR2) and a VL complementarity determining region 3 (VL2 CDR3). Tables 8 and 9 provide exemplary of CDR sequences of the anti-CD3e antibodies provided herein. Tables 11 and 12 provide exemplary of CDR sequences of the anti-CD20 antibodies provided herein.

[00146] In some embodiments, the bispecific antibody of the disclosure comprises a first antigen binding domain (e.g. binding to CD3ε) comprising any one of the VH1 and VL1 sequences listed in Table 7 and a second antigen binding domain (e.g. binding to CD20) comprising any one of the VH2 and VL2 sequences listed in Table 10,

[00147] In some embodiments, the bispecific antibody of the disclosure comprises a. first heavy chain polypeptide (H1 ) and a first light chain polypeptide (L1 ); and a second heavy chain polypeptide (H2) and a second light chain polypeptide (L2) comprising any one of the sequence listed in Table 13 and Table 16. The italicized sequences are the heavy chain variable regions and the light chain variable regions. The underlined sequences are CDRs according to Kabat and the bolded sequences are CDRs according to Chothia.

[00148] In some embodiments, the bi specific antibody of the disclosure provided herein comprises a H1 that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94 %, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or at least 100% identical to the amino acid sequence of the sequences listed in Table 13 and Table 16.

[00149] In some embodiments, the bispecific antibody of the disclosure provided herein comprises a L1 that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or at least 100% identical to the amino acid sequence of the sequences listed in Table 13 and Table 16.

[00150] In some embodiments, the bispecific antibody of the disclosure provided herein comprises a H2 that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or at least 100% identical to the amino acid sequence of the sequences listed in Table 13 and Table 16.

[00151 ] In some embodiments, the bi specific antibody of the disclosure provided herein comprises a L2 that is at least 90%, at least 91 %. at least 92%, at least 93%, at least 94%, at least 95%. at least 96%, at least 97%, at least 98%, at least 99% or at least 100% identical to the amino acid sequence of the sequences listed in Table 13 and Table 16.

[00152] In some embodiments, the H1 amino acid sequence is numbered in accordance with SEQ ID NO: 473. In some embodiments the L1 amino acid sequence is numbered in accordance with SEQ ID NO: 149. In some embodiments, the H2 amino acid sequence is numbered in accordance with SEQ ID NO: 472. In some embodiments, the L2 amino acid sequence is numbered in accordance with SEQ ID NO: 471.

[00153] Table 13. Exemplary Bispecific Antibodies that bind CD3e and CD20

[00154] Exemplary, anti-CD3e x anti-CD20 bispecific antibodies of the invention include

EIP0960, EIP0960-2, EIP0960-3, EIP0960-4, EIP0963, EIP0963-2, EIP0963-3, EIP0963-4,

EIP0961, EIP0961-2, EIP0961-3, EIP0961-4, EIP0964, EIP0964-2, EIP0964-3, EIP0964-4, EIP0962, EIP0962-2, EIP0962-3, EIP0962-4, EIP0865, EIP0865-2, EIP0865-3, EIP0865-4,

EIP0826, EIP0826-2, EIP0826-3, EIP0826-4, EIP0825, EIP0825-2, EIP0825-3, EIP0825-4,

EIP0824, EIP0824-2, EIP0824-3, EIP0824-4, EIP0846, EIP0846-2, EIP0846-3, EIP0846-4, EIP0965, EIP0965-2, E1P0965-3, EIP0965-4, EIP0867, E1P0867-2, EIP0867-3, and EIP0867-4.

[00155] In some embodiments, the bispecific antibody EIP0960-1, ETP0960-2, EIP0960-3, EIP0960-4, EIP0826-1, EIP0826-2, EIP0826-3, and EIP0826-4, comprises a first antigen binding domain that binds CD3ε, comprising a VH 1_CDR1 comprising the amino acid sequence of SEQ ID NO: 29; a VH 1_CDR2 comprising the amino acid sequence of SEQ ID NO: 34; a VH1 CDR3 comprising the amino acid sequence of SEQ ID NO: 40; a VL1 CDRI comprising the amino acid sequence of SEQ ID NO: 42; a VL1 CDR2 comprising the amino acid sequence of SEQ ID NO: 43; and a VL1 CDR3 comprising the amino acid sequence of SEQ ID NO: 47.

[00156] In some embodiments, the bispecific antibody EIP0963-I, E1P0963-2, EIP0963-3, EIP0963-4, EIP0825-1, EIP0825-2, EIP0825-3, and EIP0825-4, comprises a first antigen binding domain that binds CD3e, comprising a VH 1_CDR1 comprising the amino acid sequence of SEQ ID NO: 29; a VH 1_CDR2 comprising the amino acid sequence of SEQ ID NO: 35; a VH 1_CDR3 comprising the amino acid sequence of SEQ ID NO: 38; a VL1 CDR1 comprising the amino acid sequence of SEQ ID NO: 42; a VL1 CDR2 comprising the amino acid sequence of SEQ ID NO: 43; and a VL1 CDR3 comprising the amino acid sequence of SEQ ID NO: 47.

[00157] In some embodiments, the bispecific antibody EIP0961-1, EIP0961-2, EIP0961-3, EIP096I-4, EIP0824-1, EIP0824-2, EIP0824-3, and EIP0824-4, comprises a first antigen binding domain that binds CD3ε, comprising a VH 1_CDR1 comprising the amino acid sequence of SEQ ID NO: 30; a VH 1_CDR2 comprising the ammo acid sequence of SEQ ID NO: 34; a VH1_CDR3 comprising the amino acid sequence of SEQ ID NO: 37; a VL1 CDR1 comprising the amino acid sequence of SEQ ID NO: 42; a VL1 CDR2 comprising the amino acid sequence of SEQ ID NO: 43; and a VL1 CDR3 comprising the amino acid sequence of SEQ ID NO: 45.

[00158] In some embodiments, the bispecific antibody EIP0964-1, EIP0964-2, EIP0964-3, EIP0964-4, EIP0846-1, EIP0846-2, EIP0846-3, and EIP0846-4, comprises a first antigen binding domain that binds CD3ε, comprising a VH 1_CDR1 comprising the amino acid sequence of SEQ ID NO: 29; a VH1_CDR2 comprising the amino acid sequence of SEQ ID NO: 34; a VH1 CDR3 comprising the amino acid sequence of SEQ ID NO: 39; a VL1 CDR1 comprising the amino acid sequence of SEQ ID NO: 42; a VL1 CDR2 comprising the amino acid sequence of SEQ ID NO: 43; and a VL1 CDR3 comprising the amino acid sequence of SEQ ID NO: 47.

[00159] In some embodiments, the bispecific antibody EIP0962-1, ETP0962-2, EIP0962-3, EIP0962-4, EIP0965-1, EIP0965-2, EIP0965-3, and EIP0965-4, comprises a first antigen binding domain that binds CD3ε, comprising a VH1 CDR1 comprising the amino acid sequence of SEQ ID NO: 29; a VH 1_CDR2 comprising the amino acid sequence of SEQ ID NO: 34; a VH1 CDR3 comprising the amino acid sequence of SEQ ID NO: 37; a VL1 CDR1 comprising the amino acid sequence of SEQ ID NO: 42; a VL1 CDR2 comprising the amino acid sequence of SEQ ID NO: 44; and a VL1 CDR3 comprising the amino acid sequence of SEQ ID NO: 45.

[00160] In some embodiments, the bispecific antibody EIP0865-1, E1P0865-2, EIP0865-3, and EIP0865-4, EIP0867-1, EIP0867-2, EIP0867-3, and EIP0867-4, comprises a first antigen binding domain that binds CD3ε, comprising a VH1 CDR1 comprising the amino acid sequence of SEQ ID NO: 29; a VH1 _CDR2 comprising the amino acid sequence of SEQ ID NO: 34; a VHI CDR3 comprising the amino acid sequence of SEQ ID NO: 37; a VL1 CDR1 comprising the amino acid sequence of SEQ ID NO: 42; a VL1 CDR2 comprising the amino acid sequence of SEQ ID NO: 43; and a VL1 CDR3 comprising the amino acid sequence of SEQ ID NO: 45.

[00161] In some embodiments, the bispecific antibody EIP0960, EIP0960-2, EIP0960-3, EIP0960-4, EIP0963, EIP0963-2, EIP0963-3, EIP0963-4, EIP0961, EIP0961-2, EIP0961-3, EIP0961-4, EIP0964, EIP0964-2, EIP0964-3, EIP0964-4, EIP0962, EIP0962-2, EIP0962-3, EIP0962-4, E1P0865, E1P0865-2, EIP0865-3, and EIP0865-4, comprises a second antigen binding domain that binds CD20, comprising a VH2 CDR1 comprising the amino acid sequence of SEQ ID NO: 459; a VH2 CDR2 comprising the amino acid sequence of SEQ ID NO: 460; a VH2 CDR3 comprising the amino acid sequence of SEQ ID NO: 462; a VL2_CDR1 comprising the amino acid sequence of SEQ ID NO: 466; a VL2_CDR2 comprising the amino acid sequence of SEQ ID NO: 467; and a VL2 CDR3 comprising the amino acid sequence of SEQ ID NO: 468. [00162] In some embodiments, the bispecific antibody EIP0826, EIP0826-2, EIP0826-3, EIP0826-4, EIP0825, ETP0825-2, EIP0825-3, EIP0825-4, EIP0824, EIP0824-2, EIP0824-3, EIP0824-4, EIP0846, EIP0846-2, EIP0846-3, EIP0846-4, EIP0965, EIP0965-2, EIP0965-3, EIP0965-4, EIP0867, E1P0867-2, EIP0867-3, and EIP0867-4, comprises a second antigen binding domain that binds CD20, comprising a VH2 CDR1 comprising the amino acid sequence of SEQ ID NO: 455; a VH2 CDR2 comprising the amino acid sequence of SEQ ID NO: 457; a VH2 CDR3 comprising the amino acid sequence of SEQ ID NO: 458; a VL2_CDR1 comprising the amino acid sequence of SEQ ID NO: 463; a VL2_CDR2 comprising the amino acid sequence of SEQ ID NO: 464; and a VL2 CDR3 comprising the amino acid sequence of SEQ ID NO: 465. In some embodiments, the bispecific antibody EIP0960 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 19, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00163] In some embodiments, the bispecific antibody EIP0960-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 19, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00164] In some embodiments, the bispecific antibody EIP0960-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 19, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising tire ammo acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00165] In some embodiments, the bispecific antibody EIP0960-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 19, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00166] In some embodiments, the bispecific antibody EIP0963 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00167] In some embodiments, the bispecific antibody EIP0963-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00168] In some embodiments, die bispecific antibody EIP0963-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00169] In some embodiments, the bispecific antibody EIP0963-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising die amino acid sequence of SEQ ID NO: 452.

[00170] In some embodiments, the bispecific antibody E1P0961 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 17, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00171] In some embodiments, the bispecific antibody EIP0961-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 17, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00172] In some embodiments, the bispecific antibody EIP0961-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 17, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00173] In some embodiments, the bispecific antibody EIP0961 -4 comprises a VH1 comprising die amino acid sequence of SEQ ID NO: 17, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00174] In some embodiments, the bispecific antibody EIP0964 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00175] In some embodiments, die bispecific antibody EIP0964-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00176] In some embodiments, die bispecific antibody EIP0964-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00177] In some embodiments, the bispecific antibody EIP0964-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising die amino acid sequence of SEQ ID NO: 452.

[00178] In some embodiments, the bispecific antibody E1P0962 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 27, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00179] In some embodiments, the bispecific antibody EIP0962-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 27, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00180] In some embodiments, the bispecific antibody EIP0962-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the ammo acid sequence of SEQ ID NO: 27, a VH2 comprising the ammo acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00181] In some embodiments, the bispecific antibody EIP0962-4 comprises a VH1 comprising die amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 27, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00182] In some embodiments, the bispecific antibody EIP0865 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00183] In some embodiments, die bispecific antibody EIP0865-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the ammo acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00184] In some embodiments, die bispecific antibody EIP0865-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the ammo acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00185] In some embodiments, the bispecific antibody EIP0865-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising die amino acid sequence of SEQ ID NO: 452.

[00186] In some embodiments, the bispecific antibody E1P0826 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 19, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00187] In some embodiments, the bispecific antibody EIP0826-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 19, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00188] In some embodiments, the bispecific antibody EIP0826-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 19, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00189] In some embodiments, the bispecific antibody EIP0826-4 comprises a VH1 comprising die amino acid sequence of SEQ ID NO: 19, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00190] In some embodiments, the bispecific antibody EIP0825 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451 , and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00191] In some embodiments, die bispecific antibody EIP0825-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451 , and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00192] In some embodiments, die bispecific antibody EIP0825-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00193] In some embodiments, the bispecific antibody EIP0825-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising die amino acid sequence of SEQ ID NO: 450.

[00194] In some embodiments, the bispecific antibody E1P0824 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 17, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00195] In some embodiments, the bispecific antibody EIP0824-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 17, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00196] In some embodiments, the bispecific antibody EIP0824-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 17, a VL1 comprising the ammo acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00197] In some embodiments, the bispecific antibody EIP0824-4 comprises a VH1 comprising die amino acid sequence of SEQ ID NO: 17, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00198] In some embodiments, the bispecific antibody EIP0846 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451 , and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00199] In some embodiments, die bispecific antibody EIP0846-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00200] In some embodiments, the bispecific antibody EIP0846-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00201] In some embodiments, the bispecific antibody EIP0846-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00202] In some embodiments, the bispecific antibody E1P0965 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 27, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00203] In some embodiments, the bispecific antibody EIP0965-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 27, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00204] In some embodiments, the bispecific antibody EIP0965-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the ammo acid sequence of SEQ ID NO: 27, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00205] In some embodiments, the bispecific antibody EIP0965-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 27, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00206] In some embodiments, the bispecific antibody EIP0867 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00207] In some embodiments, the bispecific antibody EIP0867-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the ammo acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 45 I, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00208] In some embodiments, the bispecific antibody EIP0867-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the ammo acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00209] In some embodiments, the bispecific antibody EIP0867-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00210] In some embodiments, the bispecific antibody E1P0960-1 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 473, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 472, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00211] In some embodiments, the bispecific antibody EIP0960-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 473, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 554, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00212] In some embodiments, the bispecific antibody EIP0960-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 583, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 472, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00213] In some embodiments, the bispecific antibody EIP0960-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 583, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 554, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00214] In some embodiments, the bispecific antibody EIP0963-1 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 476, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 475, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00215] In some embodiments, the bispecific antibody EIP0963-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 476, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 555, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00216] In some embodiments, the bispecific antibody EIP0963-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 584, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 475, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00217] In some embodiments, the bispecific antibody EIP0963-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 584, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

555, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00218] In some embodiments, the bispecific antibody EIP0961-1 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 479, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 478, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00219] In some embodiments, the bispecific antibody EIP0961-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 479, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO:

556, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00220] In some embodiments, the bispecific antibody EIP0961-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 585, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 478, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00221] In some embodiments, the bispecific antibody EIP0961 -4 comprises a. H1 comprising the amino acid sequence of SEQ ID NO: 585, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 556, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00222] In some embodiments, the bispecific antibody EIP0964-1 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 482, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 481, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00223] In some embodiments, the bispecific antibody EIP0964-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 482, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 557, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00224] In some embodiments, the bispecific antibody EIP0964-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 586, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 481, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00225] In some embodiments, the bispecific antibody EIP0964-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 586, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

557, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00226] In some embodiments, the bispecific antibody EIP0962-1 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 485, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 484, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00227] In some embodiments, the bispecific antibody EIP0962-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 485, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO:

558, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00228] In some embodiments, the bispecific antibody EIP0962-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 587, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 484, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00229] In some embodiments, the bispecific antibody EIP0962-4 comprises a. H1 comprising the amino acid sequence of SEQ ID NO: 587, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 558, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00230] In some embodiments, the bispecific antibody EIP0865-1 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 488, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 487, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00231] In some embodiments, the bispecific antibody EIP0865-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 488, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 559, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00232] In some embodiments, the bispecific antibody EIP0865-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 588, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 487, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00233] In some embodiments, the bispecific antibody EIP0865-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 588, a L1 comprising the amino acid sequence of SEQ ID NO: 141 , a H2 comprising the amino acid sequence of SEQ ID NO:

559, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00234] In some embodiments, the bispecific antibody EIP0826-1 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 509, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 508, and a L2 comprising the amino acid sequence of SEQ ID NO: 507.

[00235] In some embodiments, the bispecific antibody EIP0826-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 509, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

560, and a L2 comprising the amino acid sequence of SEQ ID NO: 507.

[00236] In some embodiments, the bispecific antibody EIP0826-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 589, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 508, and a L2 comprising the amino acid sequence of SEQ ID NO: 507.

[00237] In some embodiments, the bispecific antibody EIP0826-4 comprises a. H1 comprising the amino acid sequence of SEQ ID NO: 589, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 560, and a L2 comprising the amino acid sequence of SEQ ID NO: 507.

[00238] In some embodiments, the bispecific antibody EIP0825-1 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 512, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 511, and a L2 comprising the amino acid sequence of SEQ ID NO: 510.

[00239] In some embodiments, the bispecific antibody EIP0825-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 512, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 561, and a L2 comprising the amino acid sequence of SEQ ID NO: 510.

[00240 ] In some embodiments, the bispecific antibody EIP0825-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 590, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 51 1, and a L2 comprising the amino acid sequence of SEQ ID NO: 510.

[00241] In some embodiments, the bispecific antibody EIP0825-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 590, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

561, and a L2 comprising the amino acid sequence of SEQ ID NO: 510.

[00242] In some embodiments, the bispecific antibody E1P0824-1 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 515, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 514, and a L2 comprising the amino acid sequence of SEQ ID NO: 513.

[00243] In some embodiments, the bispecific antibody EIP0824-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 515, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO:

562, and a L2 comprising the amino acid sequence of SEQ ID NO: 513.

[00244] In some embodiments, the bispecific antibody EIP0824-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 591, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 514, and a L2 comprising the amino acid sequence of SEQ ID NO: 513.

[00245] In some embodiments, the bispecific antibody EIP0824-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 591, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 562, and a L2 comprising the amino acid sequence of SEQ ID NO: 513.

[00246] In some embodiments, the bispecific antibody EIP0846-1 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 518, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 517, and a L2 comprising the amino acid sequence of SEQ ID NO: 516.

[00247] In some embodiments, the bispecific antibody EIP0846-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 518, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 563, and a L2 comprising the amino acid sequence of SEQ ID NO: 516.

[00248] In some embodiments, the bispecific antibody EIP0846-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 592, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 517, and a L2 comprising the amino acid sequence of SEQ ID NO: 516.

[00249] In some embodiments, the bispecific antibody EIP0846-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 592, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

563, and a L2 comprising the amino acid sequence of SEQ ID NO: 516.

[00250] In some embodiments, the bispecific antibody EIP0965-1 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 521, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 520, and a L2 comprising the amino acid sequence of SEQ ID NO: 519.

[00251] In some embodiments, the bispecific antibody EIP0965-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 521 , a L 1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO:

564, and a L2 comprising the amino acid sequence of SEQ ID NO: 519.

[00252] In some embodiments, the bispecific antibody EIP0965-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 593, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 520, and a L2 comprising the amino acid sequence of SEQ ID NO: 519.

[00253] In some embodiments, the bispecific antibody EIP0965-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 593, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 564, and a L2 comprising the amino acid sequence of SEQ ID NO: 519.

[00254] In some embodiments, the bispecific antibody EIP0867-1 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 524, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 523, and a L2 comprising the amino acid sequence of SEQ ID NO: 522.

[00255] In some embodiments, the bispecific antibody EIP0867-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 524, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 565, and a L2 comprising the amino acid sequence of SEQ ID NO: 522.

[00256] In some embodiments, die bispecific antibody EIP0867-3 comprises a H1 comprising die amino acid sequence of SEQ ID NO: 594, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 523, and a L2 comprising the amino acid sequence of SEQ ID NO: 522.

[00257] In some embodiments, the bispecific antibody EIP0867-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 594, a L1 comprising the amino acid sequence of SEQ ID NO: 141 , a H2 comprising the amino acid sequence of SEQ ID NO: 565, and a L2 comprising die amino acid sequence of SEQ ID NO: 522.

[00258] Any one of the bispecific antibodies shown above in Table 13 can be further modified by substituting any one of the anti-CD3e antigen binding regions with any one of the anti-CD3ε binding regions shown in Tables 7-9. For example, the anti-CD3ε antigen binding regions of bispecific antibody “E1P0960” can be substituted with any one of the anti-CD3ε binding regions shown in Tables 7-9 to produce the bispecific antibodies of the invention. Exemplar} antibodies are shown in Table 13.

[00259] FUSION PEPTIDES

[00260] Provided herein is an antibody (e.g. monospecific antibody or bispecific antibody) that has a fusion peptide fused to the N-tenn inus or the C -terminus of the first heavy chain polypeptide or the second heavy chain polypeptide.

[00261] Critical to the initial T cell response is the capacity for T cells to detect foreign and mutated proteins through their T cell receptor. This response, often referred to as signal 1 of T cell activation, occurs when the T cell receptor engages a cell that displays a foreign or mutated protein fragment or antigen in a specific protein complex called the Major Histocompatibility Complex I (MHCI). The activation of the T cell receptor is by itself both activating and auto-regulatory' to T cells. Strong binding of the TCR to an MHCI complex creates chronic activation of the TCR. This form of signal is associated with T cells that are reactive to self-antigens. T cells are programed to inactivate when they experience this form activation. T cells with TCR that bind weaker, but sufficient for activation, experience acute signaling with the potential to remain active and differentiate into memory T cells. Ulis is emerging as important consideration in the design the T cell therapeutics. [00262] T cell cytokine activation, often referred to signal 3, is important in T cell transitions, either from non-dividing to a state of rapid cell division or from one phenotypic state to another. T cell cytokine receptors bind to cytokines that are produced by immune and non-immune cells and depending on the cytokine and the state of the T cell at the time of receiving the cytokine signal can induce cell proliferation, can sustain vitality', or can induce differentiation of T cells into a specialized cell state appropriate for sustained activation or inactivation following infection.

[00263] One example is the transition that naive cells experience through cytokines which can induce naive T cells to proliferate and promote T cell differentiation into memory' T cells. Exemplary' cytokines include but are not limited to IL-2, IL-7, IL-10, IL-12, IL-15, IL- 18 and IL-21.

[00264] Costimulatory receptor activation, referred to as signal 2 provides a context specific cell-to-cell reinforcement of T activation. The most recognized form of costimulation occurs when T cells interact with activated antigen presenting cells through the T cell costimulatory receptor CD28 with CD80 and CD86 ligands found on APCs. These interactions can " prime” specific T cells armed with T cell receptors responsive to pathogen or cancer proteins.

[00265] Less appreciated is costimulation induced at the site of infection and malignancies. This includes costimulation that acts through CD2 and NKG2D receptors responsive to ligands like CD58 and UL16 binding proteins (e.g. ULBP2/5/6) that are induced in immune cells and epithelial cells upon viral infection. These signals provide not only reinforcement of T activation, but confirmation that the T cell’s lethal effector activities are targeted with single cell accuracy. While many costimulatory' receptors have been discovered, the importance of each receptor’s specific context and the impact of concurrent signaling of multiple costimulatory' receptors remains largely unknown and an area to greatly advance our understanding of T cell biology and creating possibilities for novel tumor-targeted T cell therapeutic development.

[00266] Costimulatory ligands include but are not limited to CD48, CD58, CD86, TNFSF9, OX40L, 4-1BBL, GITL, ( 1)70, CD80, MR 1 , TNFSF4, ICOSL or ICOSLG.

[00267] CD58 is advantageous over other costimulatory ligands in that it is the primary' costimulatory' pathway available at the tumor site as tumor infiltrating T lymphocytes often lose expression of other costimulatory receptors like CD28, or due to the low immunogenicity of tumor cells, tumor cells do not sufficiently activate T cell, thus limiting the potential of inducible costimulatory receptors like 41BB.

[00268] As described previously, the anti-CD3ε antibodies of the disclosure induce varying levels of T cell receptor activation that confer alteration m T cell vitality and cytokine production. Accordingly, a fusion of the costimulatory ligand CD58 to the anti-CD3e bispecific antibody provides integrated costimulatory T cell activation for optimal T cell activation.

[00269] In some embodiments, the bispecific antibody has a peptide fused to the N- terminus of the first heavy chain polypeptide (H1 ). In some embodiments, the bispecific antibody has a peptide fused to the C -terminus of the first heavy chain polypeptide (H1). In some embodiments, the bispecific antibody has a polypeptide fused to the N-terminus of the second heavy chain polypeptide (1 12). In some embodiments, the bi specific antibody has a peptide fused to the C-terminus of the second heavy chain polypeptide (H2). Exemplary' peptides include but are not limited to IL-2, IL-7, IL-10, IL-12, IL-15, IL-18, IL-21 or portions thereof. Exemplary' peptides include but are not limited to CD48, CD58, CD86, TNFSF9, OX40L, 4-1BBL, GITL, CD70, CD80, MR1, TNFSF4, ICOSL, ICOSLG or portions thereof Exemplary peptide sequences that are fused to the bispecific antibodies include but are not limited to those listed in Table 14.

[00270] Table 14. Exemplary Fusion Peptide Sequences

[00271] In some embodiments the polypeptide is fused directly to the bispecific antibody. In some embodiments, the polypeptide is fused indirectly through a linker. In some embodiments, the bispecific antibody fused with a peptide comprises a linker sequence.

Exemplary linker sequences include but are not limited to those listed in Table 15. [00272] Table 15. Exemplary Linker Sequences

[00273] In some embodiments, die CD3 x CD20 bispecific antibodies of the invention have a CD58 fusion peptide (SEQ ID NO: 49) fused indirectly at the C-terminus of the first heavy chain polypeptide (H1 ) using linker-1 (SEQ ID NO: 52), In some embodiments, the CD3 x CD20 bispecific antibodies of the invention have a CD58v* fusion peptide (SEQ ID NO: 50) fused indirectly at the C-terminus of the first heavy chain polypeptide (H1 ) using linker- 1 (SEQ ID NO: 52).

[00274] In some embodiments, the CD3 x CD20 bispecific antibodies of the invention have a CD58 fusion peptide (SEQ ID NO: 49) fused indirectly at the C-terminus of the first heavy chain polypeptide (H1 ) using linker-2 (SEQ ID NO: 53). In some embodiments, the CD3 x CD20 bispecific antibodies of the inventi on have a CD58v* fusion peptide (SEQ ID NO: 50) fused indirectly at the C-terminus of the first heavy chain polypeptide (H1 ) using linker-2 (SEQ ID NO: 53).

[00275] In some embodiments, the CD3 x CD20 bispecific antibodies of the invention have a CD58 fusion peptide (SEQ ID NO: 49) fused indirectly at the C-terminus of the first heavy chain polypeptide (H1 ) using linker-3 (SEQ ID NO: 54). In some embodiments, the CD3 x CD20 bispecific antibodies of the invention have a CD58v* fusion peptide (SEQ ID NO: 50) fused indirectly at the C-terminus of the first heavy chain polypeptide (H1) using linker-3 (SEQ ID NO: 54).

[00276] Exemplary CD3 x CD20 bispecific antibodies having a CD58 fusion peptide are shown m Table 16.

[00277] Table 16. Exemplary Bispecific Antibodies that bind to CD3ε and CD20 with a C-terminus CD58 fusion peptide

[00278] Exemplary', anti-CD3ε x anti-CD20 bispecific antibodies with a C-temiinus fusion peptide of the invention include ETP0969, EIP0969-2, EIP0969-3, EIP0969-4, EIP0892, EIP0892-2, EIP0892-3, EIP0892-4, EIP0929, EIP0929-2, EIP0929-3, EIP0929-4, EIP0893, EIP0893-2, EIP0893-3, EIP0893-4, EIP0886, EIP0886-2, EIP0886-3, E1P0886-4, EIP0866, EIP0866-2, EIP0866-3, EIP0866-4, EIP0966, EIP0966-2, EIP0966-3, EIP0966-4, EIP0967, EIP0967-2, EIP0967-3, EIP0967-4, EIP0930, EIP0930-2, EIP0930-3, EIP0930-4, EIP0968, EIP0968-2, EIP0968-3, EIP0968-4, EIP0891, EIP0891-2, EIP0891-3, EIP0891-4, EIP0868, EIP0868-2, EIP0868-3, and EIP0868-4.

[00279] In some embodiments, the bispecific antibody EIP0969, EIP0969-2, EIP0969-3, EIP0969-4, EIP0966, EIP0966-2, EIP0966-3, and EIP0966-4, comprises a first antigen binding domain that binds CD3e, comprising a VH1 CDR1 comprising the amino acid sequence of SEQ ID NO: 29; a VH1_CDR2 comprising the amino acid sequence of SEQ ID NO: 34; a VH1 CDR3 comprising the amino acid sequence of SEQ ID NO: 40; a VL1 CDR1 comprising the amino acid sequence of SEQ ID NO: 42; a VL1 CDR2 comprising the amino acid sequence of SEQ ID NO: 43; and a VL1 CDR3 comprising the amino acid sequence of SEQ ID NO: 47. [00280] In some embodiments, the bispecific antibody EIP0892, EIP0892-2, EIP0892-3, EIP0892-4, EIP0967, ETP0967-2, EIP0967-3, and EIP0967-4, comprises a first antigen binding domain that binds CD3ε, comprising a VH1 CDR1 comprising the amino acid sequence of SEQ ID NO: 29; a VH 1_CDR2 comprising the ammo acid sequence of SEQ ID NO: 35; a VH1_CDR3 comprising the amino acid sequence of SEQ ID NO: 38; a VL1 CDR1 comprising the amino acid sequence of SEQ ID NO: 42; a VL1 CDR2 comprising the amino acid sequence of SEQ ID NO: 43; and a VL1 CDR3 comprising the amino acid sequence of SEQ ID NO: 47.

[00281] In some embodiments, the bispecific antibody EIP0929, EIP0929-2, EIP0929-3, EIP0929-4, EIP0930, EIP0930-2, EIP0930-3, and EIP0930-4, comprises a first antigen binding domain that binds CD3ε, comprising a VH1 CDR1 comprising the amino acid sequence of SEQ ID NO: 30; a VH1_CDR2 comprising the amino acid sequence of SEQ ID NO: 34; a VH1 CDR3 comprising the amino acid sequence of SEQ ID NO: 37; a VL1 CDR1 comprising the amino acid sequence of SEQ ID NO: 42; a VL1 CDR2 comprising the amino acid sequence of SEQ ID NO: 43; and a VL1 CDR3 comprising the amino acid sequence of SEQ ID NO: 45.

[00282] In some embodiments, die bispecific antibody EIP0893, EIP0893-2, EIP0893-3, EIP0893-4, EIP0968, E1P0968-2, EIP0968-3, and EIP0968-4, comprises a first antigen binding domain that binds CD3ε, comprising a VH1 CDR1 comprising the amino acid sequence of SEQ ID NO: 29; a VH1_CDR2 comprising the amino acid sequence of SEQ ID NO: 34; a VH1 CDR3 comprising the amino acid sequence of SEQ ID NO: 39; a VL1_CDRI comprising the amino acid sequence of SEQ ID NO: 42; a VL1_CDR2 comprising the amino acid sequence of SEQ ID NO: 43; and a VL1 CDR3 comprising the amino acid sequence of SEQ ID NO: 47.

[00283] In some embodiments, the bispecific antibody EIP0886, EIP0886-2, EIP0886-3, EIP0886-4, EIP0891, EIP089I-2, EIP0891-3, and EIP0891-4, comprises a first antigen binding domain that binds CD3ε, comprising a VH1 CDR1 comprising the amino acid sequence of SEQ ID NO: 29; a VH 1_CDR2 comprising the amino acid sequence of SEQ ID NO: 34; a VH1_CDR3 comprising the amino acid sequence of SEQ ID NO: 37; a VL1 CDRI comprising the amino acid sequence of SEQ ID NO: 42; a VL1 CDR2 comprising the amino acid sequence of SEQ ID NO: 44; and a VL1 CDR3 comprising the amino acid sequence of SEQ ID NO: 45. [00284] In some embodiments, the bispecific antibody EIP0866, EIP0866-2, EIP0866-3, EIP0866-4, EIP0868, ETP0868-2, EIP0868-3, and EIP0868-4, comprises a first antigen binding domain that binds CD3ε, comprising a VH1 CDR1 comprising the amino acid sequence of SEQ ID NO: 29; a VH 1_CDR2 comprising tire amino acid sequence of SEQ ID NO: 34; a VH1_CDR3 comprising the amino acid sequence of SEQ ID NO: 37; a VL1 CDR1 comprising the amino acid sequence of SEQ ID NO: 42; a VL1 CDR2 comprising the amino acid sequence of SEQ ID NO: 43; and a VL1 CDR3 comprising the amino acid sequence of SEQ ID NO: 45.

[00285] In some embodiments, the bispecific antibody EIP0969, EIP0969-2, EIP0969-3, EIP0969-4, EIP0892, EIP0892-2, EIP0892-3, EIP0892-4, EIP0929, EIP0929-2, EIP0929-3, EIP0929-4, EIP0893, E1P0893-2, EIP0893-3, EIP0893-4, EIP0886, EIP0886-2, EIP0886-3, EIP0886-4, EIP0866, EIP0866-2, EIP0866-3 and EIP0866-4, comprises a second antigen binding domain that binds CD20, comprising a VH2 CDR1 comprising the amino acid sequence of SEQ ID NO: 459; a VH2 CDR2 comprising the amino acid sequence of SEQ ID NO: 460; a VH2_CDR3 comprising the amino acid sequence of SEQ ID NO: 462; a VL2 CDR1 comprising the amino acid sequence of SEQ ID NO: 466; a VL2 CDR2 comprising the amino acid sequence of SEQ ID NO: 467; and a VL2 CDR3 comprising the amino acid sequence of SEQ ID NO: 468.

[00286] In some embodiments, the bispecific antibody EIP0966, EIP0966-2, EIP0966-3, EIP0966-4, EIP0967, EIP0967-2, ETP0967-3, EIP0967-4, EIP0930, EIP0930-2, EIP0930-3, EIP0930-4, EIP0968, EIP0968-2, EIP0968-3, EIP0968-4, EIP0891, EIP0891-2, EIP0891-3, EIP089I-4, EIP0868, EIP0868-2, EIP0868-3, and EIP0868-4, comprises a second antigen binding domain that binds CD20, comprising a VH2 CDR1 comprising the amino acid sequence of SEQ ID NO: 455; a VH2 CDR2 comprising the amino acid sequence of SEQ ID NO: 457; a VH2 CDR3 comprising the ammo acid sequence of SEQ ID NO: 458; a VL2 CDR1 comprising the amino acid sequence of SEQ ID NO: 463; a VL2 CDR2 comprising the amino acid sequence of SEQ ID NO: 464; and a VL2 CDR3 comprising the amino acid sequence of SEQ ID NO: 465.

[00287] In some embodiments, the bispecific antibody EIP0969 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 19, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452. [00288] In some embodiments, the bispecific antibody EIP0969-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 19, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00289] In some embodiments, the bispecific antibody EIP0969-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 19, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising the ammo acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00290] In some embodiments, the bispecific antibody EIP0969-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 19, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00291] In some embodiments, the bispecific antibody EIP0892 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00292] In some embodiments, the bispecific antibody EIP0892-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00293] In some embodiments, the bispecific antibody EIP0892-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00294] In some embodiments, the bispecific antibody EIP0892-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00295] In some embodiments, the bispecific antibody EIP0929 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 17, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452. [00296] In some embodiments, the bispecific antibody EIP0929-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 17, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00297] In some embodiments, the bispecific antibody EIP092.9-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 17, a VL1 comprising the ammo acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00298] In some embodiments, the bispecific antibody EIP0929-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 17, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00299] In some embodiments, the bispecific antibody EIP0893 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00300] In some embodiments, the bispecific antibody EIP0893-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00301] In some embodiments, the bispecific antibody EIP0893-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00302] In some embodiments, the bispecific antibody EIP0893-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00303] In some embodiments, the bispecific antibody EIP0886 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 27, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452. [00304] In some embodiments, the bispecific antibody EIP0886-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 27, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00305] In some embodiments, the bispecific antibody EIP0886-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the ammo acid sequence of SEQ ID NO: 27, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00306] In some embodiments, the bispecific antibody EIP0886-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 27, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00307] In some embodiments, the bispecific antibody EIP0866 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00308] In some embodiments, the bispecific antibody EIP0866-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the ammo acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00309] In some embodiments, the bispecific antibody EIP0866-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the ammo acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00310] In some embodiments, the bispecific antibody EIP0866-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 453, and a VL2 comprising the amino acid sequence of SEQ ID NO: 452.

[00311] In some embodiments, the bispecific antibody EIP0966 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 19, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450. [00312] In some embodiments, the bispecific antibody EIP0966-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 19, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00313] In some embodiments, the bispecific antibody EIP0966-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 19, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00314] In some embodiments, the bispecific antibody EIP0966-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 19, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 45 I, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00315] In some embodiments, the bispecific antibody EIP0967 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2. comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00316] In some embodiments, the bispecific antibody EIP0967-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00317] In some embodiments, the bispecific antibody EIP0967-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00318] In some embodiments, the bispecific antibody EIP0967-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 18, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00319] In some embodiments, the bispecific antibody EIP0930 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 17, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450. [00320] In some embodiments, the bispecific antibody EIP0930-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 17, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00321] In some embodiments, the bispecific antibody EIP0930-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 17, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00322] In some embodiments, the bispecific antibody EIP0930-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 17, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 45 I, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00323] In some embodiments, the bispecific antibody EIP0968 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2. comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00324] In some embodiments, the bispecific antibody EIP0968-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the ammo acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00325] In some embodiments, the bispecific antibody EIP0968-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00326] In some embodiments, the bispecific antibody EIP0968-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 16, a VL1 comprising the amino acid sequence of SEQ ID NO: 26, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00327] In some embodiments, the bispecific antibody EIP0891 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 27, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450. [00328] In some embodiments, the bispecific antibody EIP0891-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 27, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00329] In some embodiments, the bispecific antibody EIP0891-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the ammo acid sequence of SEQ ID NO: 27, a VH2 comprising the ammo acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00330] In some embodiments, the bi specific antibody EIP0891 -4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 27, a VH2 comprising the amino acid sequence of SEQ ID NO: 45 I, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00331] In some embodiments, the bispecific antibody EIP0868 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2. comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00332] In some embodiments, the bispecific antibody EIP0868-2 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the ammo acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00333] In some embodiments, the bispecific antibody EIP0868-3 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00334] In some embodiments, the bispecific antibody EIP0868-4 comprises a VH1 comprising the amino acid sequence of SEQ ID NO: 13, a VL1 comprising the amino acid sequence of SEQ ID NO: 22, a VH2 comprising the amino acid sequence of SEQ ID NO: 451, and a VL2 comprising the amino acid sequence of SEQ ID NO: 450.

[00335] In some embodiments, the bispecific antibody EIP0960 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 473, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 472, and a L2 comprising the amino acid sequence of SEQ ID NO: 471. [00336] In some embodiments, the bispecific antibody EIP0960-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 473, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 554, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00337] In some embodiments, the bispecific antibody EIP0960-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 583, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 472, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00338] In some embodiments, the bispecific antibody EIP0960-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 583, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

554, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00339] In some embodiments, the bispecific antibody EIP0963 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 476, a L1 comprising the ammo acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 475, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00340] In some embodiments, the bispecific antibody EIP0963-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 476, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

555, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00341] In some embodiments, the bispecific antibody EIP0963-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 584, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 475, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00342] In some embodiments, the bispecific antibody EIP0963-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 584, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 555, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00343] In some embodiments, the bispecific antibody EIP0961 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 479, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 478, and a L2 comprising the amino acid sequence of SEQ ID NO: 471. [00344] In some embodiments, the bispecific antibody EIP0961-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 479, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 556, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00345] In some embodiments, the bispecific antibody EIP0961-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 585, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 478, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00346] In some embodiments, the bispecific antibody EIP0961 -4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 585, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO:

556, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00347] In some embodiments, the bispecific antibody EIP0964 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 482, a L1 comprising the ammo acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 481, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00348] In some embodiments, the bispecific antibody E1P0964-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 482, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

557, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00349] In some embodiments, the bispecific antibody EIP0964-3 comprises a HI comprising the amino acid sequence of SEQ ID NO: 586, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 481, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00350] In some embodiments, the bispecific antibody E1P0964-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 586, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 557, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00351] In some embodiments, the bispecific antibody EIP0962 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 485, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 484, and a L2 comprising the amino acid sequence of SEQ ID NO: 471. [00352] In some embodiments, the bispecific antibody EIP0962-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 485, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 558, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00353] In some embodiments, the bispecific antibody EIP0962-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 587, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 484, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00354] In some embodiments, the bispecific antibody EIP0962-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 587, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO:

558, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00355] In some embodiments, the bispecific antibody EIP0865 comprises a HI comprising the amino acid sequence of SEQ ID NO: 488, a L1 comprising the ammo acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 487, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00356] In some embodiments, the bispecific antibody EIP0865-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 488, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO:

559, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00357] In some embodiments, the bispecific antibody EIP0865-3 comprises a HI comprising the amino acid sequence of SEQ ID NO: 588, a L1 comprising the amino acid sequence of SEQ ID NO: 141 , a H2 comprising the amino acid sequence of SEQ ID NO: 487, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00358] In some embodiments, the bispecific antibody E1P0865-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 588, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 559, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00359] In some embodiments, the bispecific antibody EIP0826 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 509, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 508, and a L2 comprising the amino acid sequence of SEQ ID NO: 507. [00360] In some embodiments, the bispecific antibody EIP0826-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 509, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 560, and a L2 comprising the amino acid sequence of SEQ ID NO: 507.

[00361] In some embodiments, the bispecific antibody EIP0826-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 589, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 508, and a L2 comprising the amino acid sequence of SEQ ID NO: 507.

[00362] In some embodiments, the bispecific antibody EIP0826-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 589, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

560, and a L2 comprising the amino acid sequence of SEQ ID NO: 507.

[00363] In some embodiments, the bispecific antibody EIP0825 comprises a HI comprising the amino acid sequence of SEQ ID NO: 512, a L1 comprising the ammo acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 511, and a L2 comprising the amino acid sequence of SEQ ID NO: 510.

[00364] In some embodiments, the bispecific antibody EIP0825-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 512, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

561, and a L2 comprising the amino acid sequence of SEQ ID NO: 510.

[00365] In some embodiments, the bispecific antibody EIP0825-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 590, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 511, and a L2 comprising the amino acid sequence of SEQ ID NO: 510.

[00366] In some embodiments, the bispecific antibody EIP0825-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 590, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 561, and a L2 comprising the amino acid sequence of SEQ ID NO: 510.

[00367] In some embodiments, the bispecific antibody EIP0824 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 515, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 514, and a L2 comprising the amino acid sequence of SEQ ID NO: 513. [00368] In some embodiments, the bispecific antibody EIP0824-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 515, a L I comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 562, and a L2 comprising the amino acid sequence of SEQ ID NO: 513.

[00369] In some embodiments, the bispecific antibody EIP0824-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 591, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 514, and a L2 comprising the amino acid sequence of SEQ ID NO: 513.

[00370] In some embodiments, the bispecific antibody EIP0824-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 591, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO:

562, and a L2 comprising the amino acid sequence of SEQ ID NO: 513.

[00371] In some embodiments, the bispecific antibody EIP0846 comprises a HI comprising the amino acid sequence of SEQ ID NO: 518, a L1 comprising the ammo acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 517, and a L2 comprising the amino acid sequence of SEQ ID NO: 516.

[00372] In some embodiments, the bispecific antibody EIP0846-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 518, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

563, and a L2 comprising the amino acid sequence of SEQ ID NO: 516.

[00373] In some embodiments, the bispecific antibody EIP0846-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 592, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 517, and a L2 comprising the amino acid sequence of SEQ ID NO: 516.

[00374] In some embodiments, the bispecific antibody E1P0846-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 592, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 563, and a L2 comprising the amino acid sequence of SEQ ID NO: 516.

[00375] In some embodiments, the bispecific antibody EIP0965 comprises a HI comprising the amino acid sequence of SEQ ID NO: 521 , a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 520, and a L2 comprising the amino acid sequence of SEQ ID NO: 519. [00376] In some embodiments, the bispecific antibody EIP0965-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 521 , a L I comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 564, and a L2 comprising the amino acid sequence of SEQ ID NO: 519.

[00377] In some embodiments, the bispecific antibody EIP0965-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 593, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 520, and a L2 comprising the amino acid sequence of SEQ ID NO: 519.

[00378] In some embodiments, the bispecific antibody EIP0965-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 593, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO:

564, and a L2 comprising the amino acid sequence of SEQ ID NO: 519.

[00379] In some embodiments, the bispecific antibody EIP0867 comprises a HI comprising the amino acid sequence of SEQ ID NO: 524, a L1 comprising the ammo acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 523, and a L2 comprising the amino acid sequence of SEQ ID NO: 522.

[00380] In some embodiments, the bispecific antibody EIP0867-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 524, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO:

565, and a L2 comprising the amino acid sequence of SEQ ID NO: 522.

[00381] In some embodiments, the bispecific antibody EIP0867-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 594, a L1 comprising the amino acid sequence of SEQ ID NO: 141 , a H2 comprising the amino acid sequence of SEQ ID NO: 523, and a L2 comprising the amino acid sequence of SEQ ID NO: 522.

[00382] In some embodiments, the bispecific antibody EIP0867-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 594, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 565, and a L2 comprising the amino acid sequence of SEQ ID NO: 522.

[00383] In some embodiments, the bispecific antibody EIP0969 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 491 , a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 490, and a L2 comprising the amino acid sequence of SEQ ID NO: 471. [00384] In some embodiments, the bispecific antibody EIP0969-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 491 , a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 566, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00385] In some embodiments, the bispecific antibody EIP0969-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 595, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 490, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00386] In some embodiments, the bispecific antibody EIP0969-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 595, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

566, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00387] In some embodiments, the bispecific antibody EIP0892 comprises a HI comprising the amino acid sequence of SEQ ID NO: 494, a L1 comprising the ammo acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 493, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00388] In some embodiments, the bispecific antibody EIP0892-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 494, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

567, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00389] In some embodiments, the bispecific antibody EIP0892-3 comprises a HI comprising the amino acid sequence of SEQ ID NO: 596, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 493, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00390] In some embodiments, the bispecific antibody EIP0892-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 596, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 567, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00391] In some embodiments, the bispecific antibody EIP0929 comprises a HI comprising the amino acid sequence of SEQ ID NO: 497, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 496, and a L2 comprising the amino acid sequence of SEQ ID NO: 471. [00392] In some embodiments, the bispecific antibody EIP0929-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 497, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 568, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00393] In some embodiments, the bispecific antibody EIP0929-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 597, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 496, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00394] In some embodiments, the bispecific antibody EIP0929-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 597, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO:

568, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00395] In some embodiments, the bispecific antibody EIP0893 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 500, a L1 comprising the ammo acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 499, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00396] In some embodiments, the bispecific antibody E1P0893-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 500, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

569, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00397] In some embodiments, the bispecific antibody EIP0893-3 comprises a HI comprising the amino acid sequence of SEQ ID NO: 598, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 499, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00398] In some embodiments, the bispecific antibody E1P0893-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 598, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 569, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00399] In some embodiments, the bispecific antibody EIP0886 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 503, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 502, and a L2 comprising the amino acid sequence of SEQ ID NO: 471. [00400] In some embodiments, the bispecific antibody EIP0886-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 503, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 570, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00401] In some embodiments, the bispecific antibody EIP0886-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 599, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 502, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00402] In some embodiments, the bispecific antibody EIP0886-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 599, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO:

570, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00403] In some embodiments, the bispecific antibody EIP0866 comprises a HI comprising the amino acid sequence of SEQ ID NO: 506, a L1 comprising the ammo acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 505, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00404] In some embodiments, the bispecific antibody EIP0866-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 506, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO:

571, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00405] In some embodiments, the bispecific antibody EIP0866-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 600, a L1 comprising the amino acid sequence of SEQ ID NO: 141 , a H2 comprising the amino acid sequence of SEQ ID NO: 505, and a L2 comprising the amino acid sequence of SEQ ID NO: 471 .

[00406] In some embodiments, the bispecific antibody E1P0866-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 600, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 571, and a L2 comprising the amino acid sequence of SEQ ID NO: 471.

[00407] In some embodiments, the bispecific antibody EIP0966 comprises a HI comprising the amino acid sequence of SEQ ID NO: 527, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 526, and a L2 comprising the amino acid sequence of SEQ ID NO: 525. [00408] In some embodiments, the bispecific antibody EIP0966-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 527, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 572, and a L2 comprising the amino acid sequence of SEQ ID NO: 525.

[00409] In some embodiments, the bispecific antibody EIP0966-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 601, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 526, and a L2 comprising the amino acid sequence of SEQ ID NO: 525.

[00410] In some embodiments, the bispecific antibody EIP0966-4 comprises a HI comprising the amino acid sequence of SEQ ID NO: 601, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

572, and a L2 comprising the amino acid sequence of SEQ ID NO: 525.

[00411] In some embodiments, the bispecific antibody EIP0967 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 530, a L1 comprising the ammo acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 529, and a L2 comprising the amino acid sequence of SEQ ID NO: 528.

[00412] In some embodiments, the bispecific antibody EIP0967-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 530, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

573, and a L2 comprising the amino acid sequence of SEQ ID NO: 528.

[00413] In some embodiments, the bispecific antibody EIP0967-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 602, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 529, and a L2 comprising the amino acid sequence of SEQ ID NO: 528.

[00414] In some embodiments, the bispecific antibody E1P0967-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 602, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 573, and a L2 comprising the amino acid sequence of SEQ ID NO: 528.

[00415] In some embodiments, the bispecific antibody EIP0930 comprises a HI comprising the amino acid sequence of SEQ ID NO: 533, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 532, and a L2 comprising the amino acid sequence of SEQ ID NO: 531. [00416] In some embodiments, the bispecific antibody EIP0930-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 533, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 574, and a L2 comprising the amino acid sequence of SEQ ID NO: 531.

[00417] In some embodiments, the bispecific antibody E1P0930-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 603, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 532, and a L2 comprising the amino acid sequence of SEQ ID NO: 531 .

[00418] In some embodiments, the bispecific antibody EIP0930-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 603, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO:

574, and a L2 comprising the amino acid sequence of SEQ ID NO: 531.

[00419] In some embodiments, the bispecific antibody EIP0968 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 536, a L1 comprising the ammo acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 535, and a L2 comprising the amino acid sequence of SEQ ID NO: 534.

[00420] In some embodiments, the bispecific antibody E1P0968-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 536, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO:

575, and a L2 comprising the amino acid sequence of SEQ ID NO: 534.

[00421] In some embodiments, the bispecific antibody EIP0968-3 comprises a HI comprising the amino acid sequence of SEQ ID NO: 604, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 535, and a L2 comprising the amino acid sequence of SEQ ID NO: 534.

[00422] In some embodiments, the bispecific antibody E1P0968-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 604, a L1 comprising the amino acid sequence of SEQ ID NO: 149, a H2 comprising the amino acid sequence of SEQ ID NO: 575, and a L2 comprising the amino acid sequence of SEQ ID NO: 534.

[00423] In some embodiments, the bispecific antibody EIP0891 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 539, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 538, and a L2 comprising the amino acid sequence of SEQ ID NO: 537. [00424] In some embodiments, the bispecific antibody EIP0891-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 539, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 576, and a L2 comprising the amino acid sequence of SEQ ID NO: 537.

[00425] In some embodiments, the bispecific antibody EIP0891-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 605, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO: 538, and a L2 comprising the amino acid sequence of SEQ ID NO: 537.

[00426] In some embodiments, the bispecific antibody EIP0891 -4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 605, a L1 comprising the amino acid sequence of SEQ ID NO: 153, a H2 comprising the amino acid sequence of SEQ ID NO:

576, and a L2 comprising the amino acid sequence of SEQ ID NO: 537.

[00427] In some embodiments, the bispecific antibody EIP0868 comprises a HI comprising the amino acid sequence of SEQ ID NO: 542, a L1 comprising the ammo acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 541, and a L2 comprising the amino acid sequence of SEQ ID NO: 540.

[00428] In some embodiments, the bispecific antibody EIP0868-2 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 542, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO:

577, and a L2 comprising the amino acid sequence of SEQ ID NO: 540.

[00429] In some embodiments, the bispecific antibody EIP0868-3 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 606, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 541, and a L2 comprising the amino acid sequence of SEQ ID NO: 540.

[00430] In some embodiments, the bispecific antibody EIP0868-4 comprises a H1 comprising the amino acid sequence of SEQ ID NO: 606, a L1 comprising the amino acid sequence of SEQ ID NO: 141, a H2 comprising the amino acid sequence of SEQ ID NO: 577, and a L2 comprising the amino acid sequence of SEQ ID NO: 540.

[00431] Any one of the anti-CD3/anti-20 bispecific antibodies shown above in Table 16 can be further modified by substituting any one of the anti-CD3g antigen binding regions with any one of the anti-CD3e binding regions shown in Tables 7-9. For example, the anti- CD3e antigen binding regions of bispecific antibody “EIP0969” can be substituted with any one of the anti-CD3e binding regions shown in Tables 7-9 to produce the bispecific antibodies of the invention. Exemplary antibodies are shown in Table 13 and Table 16. [00432] METHODS OF PRODUCTION

[00433] Various procedures known within the art may be used for the production of polyclonal or monoclonal antibodies directed against a given target, such as, for example, CD20, a disease associated antigen or other target, or against derivatives, fragments, analogs homologs or orthologs thereof. (See, for example, Antibodies: A Laboratory Manual, Harlow E, and Lane D, 1988, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, incorporated herein by reference).

[00434] Antibodies are purified by well-known techniques, such as affinity chromatography using protein A or protein G, which provide primarily the IgG fraction of immune serum. Subsequently, or alternatively, the specific antigen which is the target of the immunoglobulin sought, or an epitope thereof, may be immobilized on a column to purify the immune specific antibody by immunoaffinity chromatography. Purification of immunoglobulins is discussed, for example, by D. Wilkinson (The Scientist, published by Tire Scientist, Inc., Philadelphia PA, Vol. 14, No. 8 (April 17, 2000), pp. 25-28).

[00435] In some embodiments, the antibodies of the invention are monoclonal antibodies. Monoclonal antibodies are generated, for example, by using the procedures set forth m the Examples provided herein. Antibodies are also generated, e.g., by immunizing BALB/c mice with combinations of cell transfectants expressing high levels of a given target on their surface. Hybridomas resulting from myeloma/B cell fusions are then screened for reactivity' to the selected target.

[00436] Monoclonal antibodies are prepared, for example, using hybridoma methods, such as those described by Kohler and Milstein, Nature, 256:495 (1975). In a hybridoma method, a mouse, hamster, or other appropriate host animal, is typically immunized with an immunizing agent to elicit lymphocytes that produce or are capable of producing antibodies that will specifically bind to the immunizing agent. Alternatively, the lymphocytes can be immunized in vitro.

[00437] The immunizing agent will typically include the protein antigen, a fragment thereof or a fusion protein thereof Generally, either peripheral blood lymphocytes are used if cells of human origin are desired, or spleen cells or lymph node cells are used if non-human mammalian sources are desired. The lymphocytes are then fused with an immortalized cell line using a suitable fusing agent, such as polyethylene glycol, to form a hybridoma cell (Goding, Monoclonal Antibodies: Principles and Practice, Academic Press, (1986) pp. 59- 103). Immortalized cell lines are usually transformed mammalian cells, particularly myeloma cells of rodent, bovine and human origin. Usually, rat or mouse myeloma cell lines are employed. The hybridoma cells can be cultured in a suitable culture medium that preferably contains one or more substances that inhibit the growth or survival of the unfused, immortalized cells. For example, if the parental cells lack the enzyme hypoxanthine guanine phosphoribosyl transferase (HGPRT or HPRT), the culture medium for the hybridomas typically will include hypoxanthine, aminopterin, and thymidine (“HAT medium”), which substances prevent the growth of HGPRT-deficient cells.

[00438] Preferred immortalized cell lines are those that fuse efficiently, support stable high level expression of antibody by the selected antibody-producing cells, and are sensitive to a medium such as HAT medium. More preferred immortalized cell lines are murine myeloma lines, which can be obtained, for instance, from the Salk Institute Cell Distribution Center, San Diego, California and the American Type Culture Collection, Manassas, Virginia. Human myeloma and mouse-human heteromyeloma cell lines also have been described for the production of monoclonal antibodies. (See Kozbor, J. Immunol., 133:3001 (1984); Brodeur et al., Monoclonal Antibody Production Techniques and Applications, Marcel Dekker, Inc., New York, (1987) pp. 51-63)).

[00439] Tire culture medium in which the hybridoma cells are cultured can then be assayed for the presence of monoclonal antibodies directed against tire antigen. Preferably, the binding specificity of monoclonal antibodies produced by the hybridoma cells is determined by immunoprecipitation or by an in vitro binding assay, such as radioimmunoassay (RIA) or enzyme-linked immunoabsorbent assay (EL1SA). Such techniques and assays are known in the art. Tire binding affinity of the monoclonal antibody can, for example, be determined by the Scatchard analysis of Munson and Pollard, Anal. Biochem., 107:220 (1980). Moreover, in therapeutic applications of monoclonal antibodies, it is important to identify antibodies having a high degree of specificity and a high binding affinity for the target antigen.

[00440] After the desired hybridoma cells are identified, the clones can be subcioned by limiting dilution procedures and grown by standard methods. (See Goding, Monoclonal Antibodies: Principles and Practice, Academic Press, (1986) pp. 59-103). Suitable culture media for this purpose include, for example, Dulbecco's Modified Eagle’s Medium and RPMI-1640 medium. Alternatively, the hybridoma cells can be grown in vivo as ascites in a mammal.

[0044 H The monoclonal antibodies secreted by the subclones can be isolated or purified from the culture medium or ascites fluid by conventional immunoglobulin purification procedures such as, for example, protein A-Sepharose, hydroxylapatite chromatography, gel electrophoresis, dialysis, or affinity chromatography.

[00442] Monoclonal antibodies can also be made by recombinant DNA methods, such as those described in U.S. Patent No. 4,816,567. DNA encoding the monoclonal antibodies of the invention can be readily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes that are capable of binding specifically to genes encoding the heavy and light chains of murine antibodies). The hybridoma cells of the invention serve as a preferred source of such DNA. Once isolated, the DNA can be placed into expression vectors, which are then transfected into host ceils such as simian COS cells, Chinese hamster ovary (CHO) cells, or myeloma cells that do not otherwise produce immunoglobulin protein, to obtain the synthesis of monoclonal antibodies in the recombinant host cells. The DNA also can be modified, for example, by substituting the coding sequence for human heavy and light chain constant domains in place of the homologous murine sequences (see U.S. Patent No. 4,816,567; Morrison, Nature 368, 812- 13 (1994)) or by covalently joining to the immunoglobulin coding sequence all or part of the coding sequence for a non -immunoglobulin polypeptide. Such a non-immunoglobulm polypeptide can be su bstituted for tire constant domains of an antibody of the invention, or can be substituted for the variable domains of one antigen-combining site of an antibody of the invention to create a chimeric bivalent antibody.

[00443] Monoclonal antibodies of the invention include humanized antibodies or human antibodies. These antibodies are suitable for administration to humans without engendering an immune response by the human against the administered immunoglobulin. Humanized forms of antibodies are chimeric immunoglobulins, immunoglobulin chains or fragments thereof (such as Fv, Fab, Fab', F(ab')?. or other antigen -binding subsequences of antibodies) that are principally comprised of the sequence of a human immunoglobulin, and contain minimal sequence derived from a non-human immunoglobulin. Humanization is performed, e.g., by following the method of Winter and co-workers (Jones et al., Nature, 321 :522-525 (1986); Riechmann et al., Nature, 332:323-327 (1988); Verhoeyen et al., Science, 239:1534- 1536 (1988)), by substituting rodent CDRs or CDR sequences for the corresponding sequences of a human antibody. (See also U.S. Patent No. 5,225,539). In some instances, Fv framework residues of the human immunoglobulin are replaced by corresponding nonhuman residues. Humanized antibodies also comprise, e.g., residues which are found neither in the recipient antibody nor in the imported CDR or framework sequences. In general, the humanized antibody includes substantially all of at least one, and typically two, variable domains, in which all or substantially all of the CDR regions correspond to those of a nonhuman immunoglobulin and all or substantially all of the framework regions are those of a human immunoglobulin consensus sequence. Tire humanized antibody optimally also includes at least a portion of an immunoglobulin constant region (Fc), typically that of a human immunoglobulin (Jones et al., 1986; Riechmann et al., 1988; and Presta, Curr. Op. Struct. Biol., 2:593-596 (1992)).

[00444] Fully human antibodies are antibody molecules in which the entire sequence of both the light chain and the heavy chain, including the CDRs, arise from human genes. Such antibodies are termed "‘human antibodies”, or “folly human antibodies” herein. Monoclonal antibodies can be prepared by using trioma technique; the human B-cell hybridoma technique (see Kozbor, et al., 1983 Immunol Today 4: 72); and the EBV hybridoma technique to produce monoclonal antibodies (see Cole, et al., 1985 In: MONOCLONAL ANTIBODIES AND CANCER THERAPY, Alan R. Liss, Inc., pp. 77-96). Monoclonal antibodies may be utilized and may be produced by using human hybridomas (see Cote, et al., 1983. Proc Natl Acad Sei USA 80: 2026-2030) or by transforming human B-cells with Epstein Barr Virus in vitro (see Cole, et al., 1985 In: MONOCLONAL ANTIBODIES AND CANCER THERAPY, Alan R. Liss, Inc., pp. 77-96).

[00445] In addition, human antibodies can also be produced using additional techniques, including phage display libraries. (See Hoogenboom and Winter, J. Mol. Biol., 227:381 (1991); Marks et al., J. Mol. Biol., 222:581 (1991)). Similarly, human antibodies can be made by introducing human immunoglobulin loci into transgenic animals, e.g., mice in which the endogenous immunoglobulin genes have been partially or completely inactivated. Upon challenge, human antibody production is observed, which closely resembles that seen in humans in all respects, including gene rearrangement, assembly, and antibody repertoire. Ulis approach is described, for example, in U.S. Patent Nos. 5,545,807; 5,545,806;

5,569,825; 5,625,126; 5,633,425; 5,661,016, and in Marks et al., Bio/Technology 10, 779 783 (1992); Lonberg et al., Nature 368 856-859 (1994); Morrison, Nature 368, 812-13 (1994); Fishwild et al, Nature Biotechnology 14, 845-51 (1996); Neuberger, Nature Biotechnology 14, 826 (1996); and Lonberg and Huszar, Intern. Rev. Immunol. 13 65-93 (1995).

[00446] Human antibodies may additionally be produced using transgenic nouhuman animals which are modified so as to produce fully human antibodies rather than the animal’s endogenous antibodies in response to challenge by an antigen. (See PCT publication W094/02602). The endogenous genes encoding the heavy and light immunoglobulin chains in the nonhuman host have been incapacitated, and active loci encoding human heavy and light chain immunoglobulins are inserted into the host’s genome. The human genes are incorporated, for example, using yeast artificial chromosomes containing the requisite human DNA segments. An animal which provides all the desired modifications is then obtained as progeny by crossbreeding intermediate transgenic animals containing fewer than the full complement of the modifications. An example of such a nonhuman animal is a mouse termed the Xenomouse™ as disclosed in PCT publications WO 96/33735 and WO 96/34096. This animal produces B cells which secrete fully human immunoglobulins. The antibodies can be obtained directly from the animal after immunization with an immunogen of interest, as, for example, a preparation of a polyclonal antibody, or alternatively from immortalized B cells derived from the animal, such as hybridomas producing monoclonal antibodies. Additionally, the genes encoding the immunoglobulins with human variable regions can be recovered and expressed to obtain the antibodies directly, or can be further modified to obtain analogs of antibodies such as, for example, single chain Fv (scFv) molecules.

[00447] An example of a method of producing a nonhuman host, exemplified as a mouse, lacking expression of an endogenous immunoglobulin heavy chain is disclosed in U.S. Patent No. 5,939,598. It can be obtained by a method, which includes deleting the J segment genes from at least one endogenous heavy chain locus in an embryonic stem cell to prevent rearrangement of the locus and to prevent formation of a transcript of a rearranged immunoglobulin heavy chain locus, the deletion being effected by a targeting vector containing a gene encoding a selectable marker; and producing from the embryonic stem cell a transgenic mouse whose somatic and germ cells contain die gene encoding the selectable marker. [00448] One method for producing an antibody of interest, such as a human antibody, is disclosed in U.S. Patent No. 5,916,771. This method includes introducing an expression vector that contains a nucleotide sequence encoding a heavy chain into one mammalian host cell in culture introducing an expression vector containing a nucleotide sequence encoding a light chain into another mammalian host cell, and fusing the two cells to form a hybrid cell. The hybrid cell expresses an antibody containing the heavy chain and the light chain. [00449] In a further improvement on this procedure, a method for identifying a clinically relevant epitope on an immunogen and a correlative method for selecting an antibody that binds specifically to the relevant epitope with high affinity are disclosed in PCT publication WO 99/53049.

[00450] The antibody can be expressed by a vector containing a DNA segment encoding the single chain antibody described above.

[00451] These can include vectors, liposomes, naked DNA, adjuvant-assisted DNA. gene gun, catheters, etc. Vectors include chemical conjugates such as described in WO 93/64701, which has targeting moiety (e.g.. a ligand to a cellular surface receptor), and a nucleic acid binding moiety (e.g., polylysine), viral vector (e.g.. a. DNA or RNA viral vector), fusion proteins such as described in PCT/US 95/02140 (WO 95/22618) which is a fusion protein containing a target moiety (e.g., an antibody specific for a target cell) and a nucleic acid binding moiety (e.g., a protamine), plasmids, phage, etc. The vectors can be chromosomal, non-chromosomal or synthetic.

[00452] Preferred vectors include viral vectors, fusion proteins and chemical conjugates. Retroviral vectors include moloney murine leukemia viruses. DNA viral vectors are preferred. These vectors include pox vectors such as orthopox or avipox vectors, herpesvirus vectors such as a herpes simplex I virus (HSV) vector (see Geller, A. I. et al., J. Neurochem, 64:487 (1995); Lim, F., et al., in DNA Cloning: Mammalian Systems, D. Glover, Ed. (Oxford Univ. Press, Oxford England) (1995); Geller, A. I. et al.. Proc Natl. Acad. Sci.: U.S.A. 90:7603 (1993); Geller, A. I„ et al„ Proc Natl. Acad. Set USA 87: 1 149 (1990), Adenovirus Vectors (see LeGal LaSalle et al., Science, 259:988 (1993); Davidson, et al., Nat. Genet 3:219 (1993); Yang, et al., J. Virol. 69:2004 (1995) and Adeno-associated Virus Vectors (see Kaplitt, M. G. et al., Nat. Genet. 8: 148 (1994).

[00453] Pox viral vectors introduce the gene into the cell’s cytoplasm. Avipox virus vectors result in only a short term expression of the nucleic acid. Adenovirus vectors, adeno- associated virus vectors and herpes simplex virus (HSV) vectors are preferred for introducing the nucleic acid into neural cells. The adenovirus vector results in a shorter term expression (about 2 months) than adeno-associated virus (about 4 months), which in turn is shorter than HSV vectors. The particular vector chosen will depend upon the target cell and the condition being treated. The introduction can be by standard techniques, e.g., infection, transfection, transduction or transformation. Examples of modes of gene transfer include e.g., naked DMA, (Ca)₂(PO₄)₃ precipitation, DEAE dextran, electroporation, protoplast fusion, lipofection, cell microinjection, and viral vectors.

[00454] The vector can be employed to target essentially any desired target cell. For example, stereotaxic injection can be used to direct the vectors (e.g., adenovirus, HSV) to a desired location. Additionally, the particles can be delivered by intracerebroventricular (rev) infusion using a minipump infusion system, such as a Synchro Med Infusion System. A method based on bulk flow, termed convection, has also proven effective at delivering large molecules to extended areas of the brain and may be useful in delivering the vector to the target cell. (See Bobo et al.. Proc. Natl. Acad. Sei. USA 91 :2076-2080 (1994); Morrison et al., Am. J. Physiol. 266:292-305 (1994)). Other methods that can be used include catheters, intravenous, parenteral, intraperitoneal and subcutaneous injection, and oral or other known routes of administration.

[00455] Bispecific antibodies are antibodies that have binding specificities for at least two different antigens. In the present case, one of the binding specificities is for a first target such as CD3ε or any fragment thereof. The second binding target is a disease associated antigen such as CD20 or any fragment thereof.

[00456] Methods for making bispecific antibodies are known in the art. Traditionally, the recombinant production of bispecific antibodies is based on the co-expression of two immunoglobulin heavy -chain/light-chain pairs, where the two heavy chains have different specificities (Milstein and Cuello, Nature, 305:537-539 (1983)). Because of the random assortment of immunoglobulin heavy and light chains, these hybridomas (quadromas) produce a potential mixture often different antibody molecules, of which only one has the correct bispecific structure. The purification of the correct molecule is usually accomplished by affinity chromatography steps. Similar procedures are disclosed in WO 93/08829, published 13 May 1993, and in Traunecker et al., EMBO J., 10:3655-3659 (1991). [00457] Bispecific and/or monovalent antibodies of the invention can be made using any of a variety of art-recognized techniques, including those disclosed in co-pending application WO 2012/023053, filed August 16, 2011, the contents of which are hereby incorporated by reference tn their entirety. The methods described in WO 2012/023053 generate bispecific antibodies that are identical in structure to a human immunoglobulin. This type of molecule is composed of two copies of a unique heavy chain polypeptide, a first light chain variable region fused to a constant Kappa domain and second light chain variable region fused to a constant Lambda domain. Each combining site displays a different antigen specificity to which both the heavy and light chain contribute. Tire light chain variable regions can be of die Lambda or Kappa family and are preferably fused to a Lambda and Kappa constant domains, respectively. This is preferred in order to avoid the generation of non-natural polypepti de junctions. However, it is also possible to obtain bispecific antibodies of the invention by fusing a Kappa light chain variable domain to a constant Lambda domain for a first specificity and fusing a Lambda light chain variable domain to a constant Kappa domain for the second specificity. The bispecific antibodies described in WO 2012/023053 are referred to as IgCsKA antibodies or “ κλ bodies,” a new fully human bispecific IgG format. This κλ-body format allows the affinity purification of a bispecific antibody that is undistinguishable from a standard IgG molecule with characteristics that are undistinguishable from a standard monoclonal antibody and, therefore, favorable as compared to previous formats.

[00458] An essential step of the method is the identification of two antibody Fv regions (each composed by a variable light chain and variable heavy chain domain) having different antigen specificities that share the same heavy chain variable domain. Numerous methods have been described for the generation of monoclonal antibodies and fragments thereof. (See, e.g., Antibodies: A Laboratory Manual, Harlow E, and Lane D, 1988, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, incorporated herein by reference). Fully human antibodies are antibody molecules in which the sequence of both the light chain and the heavy chain, including the CDRs 1 and 2, arise from human genes. The CDR3 region can be of human origin or designed by- synthetic means. Such antibodies are termed "‘human antibodies”, or “fully human antibodies” herein. Human monoclonal antibodies can be prepared by using the trioma technique; the human B-cell hybridoma technique (see

Kozbor, et al., 1983 Immunol Today 4: 72); and the EBV hybridoma technique to produce human monoclonal antibodies (see Cole, et al., 1985 In: MONOCLONAL ANTIBODIES AND CANCER THERAPY, Alan R. Liss, Inc., pp. 77-96). Human monoclonal antibodies may be utilized and may be produced by using human hybridomas (see Cote, et al., 1983. Proc Natl Acad Sci USA 80: 2026-2030) or by transforming human B-cells with Epstein Barr Virus in vitro (see Cole, et al., 1985 In: MONOCLONAL ANTIBODIES AND CANCER THERAPY, Alan R. Liss, Inc., pp. 77-96).

[00459] Monoclonal antibodies are generated, e.g. , by immunizing an animal with a target antigen or an immunogenic fragment, derivative or variant thereof. Alternatively, the animal is immunized with cells transfected with a vector containing a nucleic acid molecule encoding the target antigen, such that the target antigen is expressed and associated with the surface of the transfected cells. A variety of techniques are well-known in the art for producing xenogenic non-human animals. For example, see U.S. Pat. No. 6,075,181 and No. 6,150,584, which is hereby incorporated by reference in its entirety.

[00460] Alternatively, the antibodies are obtained by screening a library that contains antibody or antigen binding domain sequences for binding to the target antigen. This library is prepared, e.g., in bacteriophage as protein or peptide fusions to a bacteriophage coat protein that is expressed on the surface of assembled phage particles and the encoding DNA sequences contained within tire phage particles (i.e., ‘"phage displayed library”).

Alternatively, a library’ can be prepared in yeast as protein or peptide fissions to a cell wall protein on the surface of yeast cells and encoding DNA sequences contained within the yeast cells (i.e. “yeast display library”).

[00461] Hybridomas resulting from myeloma/B cell fusions are then screened for reactivity to the target antigen. Monoclonal antibodies are prepared, for example, using hybridoma methods, such as those described by Kohler and Milstein, Nature, 256:495 (1975). In a hybridoma method, a mouse, hamster, or other appropriate host animal, is typically immunized with an immunizing agent to elicit lymphocytes that produce or are capable of producing antibodies that will specifically bind to the immunizing agent. Alternatively, the lymphocytes can be immunized in vitro.

[00462] Although not strictly impossible, the serendipitous identification of different antibodies having the same heavy chain variable domain but directed against different antigens is highly unlikely. Indeed, in most cases the heavy chain contributes largely to the antigen binding surface and is also the most variable in sequence. In particular the CDR3 on the heavy chain is the most diverse CDR in sequence, length and structure. Thus, two antibodies specific for different antigens will almost invariably carry different heavy chain variable domains.

[00463] The methods disclosed in co-pending application WO 2012/023053 overcomes this lim itation and greatly facilitates the isolation of antibodies having the same heavy chain variable domain by the use of antibody libraries in which the heavy chain variable domain is the same for all the library members and thus the diversity is confined to the light chain variable domain. Such libraries are described, for example, in co-pending applications WO 2010/135558 and WO 2011/084255, each of which is hereby incorporated by reference in its entirety. However, as the light chain variable domain is expressed in conjunction with the heavy variable domain, both domains can contribute to antigen binding. To further facilitate the process, antibody libraries containing the same heavy chain variable domain and either a diversity' of Lambda variable light chains or Kappa variable light chains can be used in parallel for in vitro selection of antibodies against different antigens. This approach enables the identi fication of two antibodies having a common heavy chain but one earning a Lambda light chain variable domain and the other a Kappa light chain variable domain that can be used as building blocks for the generation of a bispecific antibody in the full immunoglobulin format of the invention. The bispecific antibodies of the invention can be of different Isotypes and their Fc portion can be modified in order to alter the bind properties to different Fc receptors and in this ivay modify the effectors functions of the antibody as well as it pharmacokinetic properties. Numerous methods for the modification of the Fc portion have been described and are applicable to antibodi es of the invention, (see for example Strohl, WR Curr Opin Biotechnol 2009 (6):685-91 ; U.S. Pat. No. 6,528,624; PCT/US2009/0191199 filed Jan 9, 2009). The methods of the invention can also be used to generate bispecific antibodies and antibody mixtures in a F(ab’)2 format that lacks the Fc portion.

[00464] The common heavy chain and two different light chains are co-expressed into a single cell to allow for the assembly of a bispecific antibody of the invention. If all the polypeptides get expressed at the same level and get assembled equally well to form an immunoglobulin molecule then the ratio of monospecific (same light chains) and bispecific (two different light chains) should be 50%. However, it is likely that different light chains are expressed at different levels and/or do not assemble with the same efficiency. Therefore, a means to modulate the relative expression of the different polypeptides is used to compensate for their intrinsic expression characteristics or different propensities to assemble with the common heavy chain. This modulation can be achieved via promoter strength, tire use of internal ribosome entry sites (IRES) featuring different efficiencies or other types of regulatory elements that can act at transcriptional or translational levels as well as acting on mRNA stability. Different promoters of different strength could include CMV (Immediate-early Cytomegalovirus virus promoter); EFl-la (Human elongation factor la-subunit promoter); Ubc (Human ubiquitin C promoter); SV40 (Simian virus 40 promoter). Different IRES have also been described from mammalian and viral origin. (See e.g., Hellen CU and Sarnow P. Genes Dev 2001 15: 1593-612). These IRES can greatly differ in their length and ribosome recruiting efficiency. Furthermore, it is possible to further tune the activity by introducing multiple copies of an IRES (Stephen et al. 2000 Proc Natl Acad Sci USA 97: 1536-1541). The modulation of the expression can also be achieved by multiple seq uential transfections of cells to increase the copy number of individual genes expressing one or the other light chain and thus modify their relative expressions. The Examples provided herein demonstrate that controlling the relative expression of the different chains is critical for maximizing the assembly and overall yield of the bispecific antibody.

[00465] The co-expression of the heavy chain and two light chains generates a mixture of three different antibodies into the cell culture supernatant: two monospecific bivalent antibodies and one bispecific bivalent antibody. The latter has to be purified from tire mixture to obtain the molecule of interest. The method described herein greatly facilitates this purification procedure by the use of affinity' chromatography media that specifically interact with the Kappa or Lambda light chain constant domains such as the CaptureSelect Fab Kappa and CaptureSelect Fab Lambda affinity matrices (BAC BV, Holland). This multi-step affinity chromatography purification approach is efficient and generally applicable to antibodies of the invention. This is in sharp contrast to specific purification methods that have to be developed and optimized for each bispecific antibodies derived from quadromas or other cell lines expressing antibody mixtures. Indeed, if the biochemical characteristics of the different antibodies in the mixtures are similar, their separation using standard chromatography technique such as ion exchange chromatography can be challenging or not possible at all. [00466] Other suitable purification methods include those disclosed in co-pending application PCT/IB2012/003028, filed on October 19, 2012, published as WO2013/088259, die contents of which are hereby incorporated by reference in their entirety.

[00467] In some embodiments of producing bispecific antibodies, antibody variable domains with the desired binding specificities (antibody-antigen combining sites) can be fused to immunoglobulin constant domain sequences. The fusion preferably is with an immunoglobulin heavy-chain constant domain, comprising at least part of the hinge, CH2, and CH3 regions. It is preferred to have the first heavy-chain constant region (CHI) containing the site necessary for light-chain binding present in at least one of the fusions. DNAs encoding the immunoglobulin heavy-chain fusions and, if desired, the immunoglobulin light chain, are inserted into separate expression vectors, and are cotransfected into a suitable host organism. For further details of generating bispecific antibodies see, for example, Suresh et al., Methods in Enzymology, 121:210 (1986).

[00468] According to another approach described in WO 96/27011, the interface between a pair of anti body molecules can be engineered to maximize the percentage of heterodimers which are recovered from recombinant cell culture. The preferred interface includes at least a part of the CH3 region of an antibody constant domain. In this method, one or more small amino acid side chains from the interface of the first antibody molecule are replaced with larger side chains (e.g., tyrosine or tryptophan). Compensatory “cavities” of identical or similar size to the large side chain(s) are created on the interface of the second antibody molecule by replacing large amino acid side chains with smaller ones (e.g., alanine or threonine). This provides a mechanism for increasing the yield of the heterodimer over other unwanted end-products such as homodimers.

[00469] Techniques for generating bispecific antibodies from antibody fragments have been described in the literature. For example, bispecific antibodies can be prepared using chemical linkage. The bispecific antibodies produced can be used as agents for the selective immobilization of enzymes.

[00470] Various techniques for making and isolating bispecific antibody fragments directly from recombinant cell culture have also been described. For example, bispecific antibodies have been produced using leucine zippers. Kostelny et al., J. Immunol. 148(5): 1547-1553 (1992). The leucine zipper peptides from the Fos and Jun proteins were linked to the Fab’ portions of two different antibodies by gene fusion. The antibody homodimers were reduced at the hinge region to form monomers and then re-oxidized to form the antibody heterodimers. This method can also be utilized for the production of antibody homodimers. The “diabody” technology described by Hollinger et al., Proc. Natl. Acad. Sci. USA 90:6444-6448 (1993) has provided an alternative mechanism for making bispecific antibody fragments. The fragments comprise a heavy-chain variable domain (VH) connected to a light-chain variable domain (VL) by a linker which is too short to allow pairing between the two domains on the same chain. Accordingly, the VH and VL domains of one fragment are forced to pair with the complementary VL and VH domains of another fragment, thereby forming two antigen-binding sites. Another strategy for making bispecific antibody fragments by the use of single-chain Fv (sFv) dimers has also been reported. See, Gruber et al., J. Immunol. 152:5368 (1994).

[00471] Antibodies with more than two valencies are contemplated. For example, trispecific antibodies can be prepared. Tutt et al., J. Immunol. 147:60 (1991).

[00472] Exemplary bispecific antibodies can bind to two different epitopes, at least one of which originates in the protein antigen of the invention. Alternatively, an anti -antigenic arm of an immunoglobulin molecule can be combined with an arm which binds to a triggering molecule on a leukocyte such as a T-cell receptor molecule (e.g., CD2, CD3, CD28, or B7), or Fc receptors for IgG (FcyR), such as FcyRl (CD64), FcvRII (CD32) and FcyRIII (CD 16) so as to focus cellular defense mechanisms to the cell expressing the particular antigen. Bispecific antibodies can also be used to direct cytotoxic agents to cells which express a particular antigen. These antibodies possess an antigen-binding arm and an arm which binds a cytotoxic agent or a radionuclide chelator, such as EOTUBE, DPTA, DOTA, or TETA. Another bispecific antibody of interest binds the protein antigen described herein and further binds tissue factor (TF).

[00473] Heteroconjugate antibodies are also within the scope of the present invention. Heteroconjugate antibodies are composed of two covalently joined antibodies. Such antibodies have, for example, been proposed to target immune system cells to unwanted cells (see U.S. Patent No. 4,676,980), and for treatment of HIV infection (see WO 91/00360; WO 92/200373; EP 03089). It is contemplated that the antibodies can be prepared in vitro using known methods in synthetic protein chemistry, including those involving crosslinking agents. For example, immunotoxins can be constructed using a disulfide exchange reaction or by forming a thioether bond. Examples of suitable reagents for this purpose include iminothiolate and methyl -4-mercaptobutyrimidate and those disclosed, for example, in U.S. Patent No. 4,676,980.

[00474] It can be desirable to modify the antibody of the invention with respect to effector function, so as to enhance, e.g., the effectiveness of the antibody m treating cancer and/or other diseases and disorders associated with aberrant CD20 expression and/or activity. For example, cysteine residue(s) can be introduced into the Fc region, thereby allowing interchain disulfide bond formation in this region. The homodimeric antibody thus generated can have improved internalization capability and/or increased complement- mediated cell killing and antibody -dependent cellular cytotoxicity (ADCC). (See Caron et al., J. Exp Med., 176: 1 191-1195 (1992) and Shopes, J. Immunol., 148: 2918-2922 (1992)). Alternatively, an antibody can be engineered that has dual Fc regions and can thereby have enhanced complement lysis and ADCC capabilities. (See Stevenson et al., Anti-Cancer Drug Design, 3: 219-230 (1989)).

[00475] Tire invention also pertains to immunoconjugates comprising an antibody conjugated to a cytotoxic agent such as a toxin (e.g. , an enzymatically' active toxin of bacterial, fungal, plant, or animal origin, or fragments thereof), or a radioactive isotope (i.e. , a radioconjugate).

[00476] Enzymatically active toxins and fragments thereof that can be used include diphtheria A chain, nonbinding active fragments of diphtheria toxin, exotoxin A chain (from Pseudomonas aeruginosa), ricin A chain, abrin A chain, modeccin A chain, alpha-sarcin, Aleurites fordii proteins, dianthin proteins, Phytolaca americana proteins (PAPI, PAPII, and PAP-S), momordica charantia inhibitor, curcin, crotin, sapaonaria officinalis inhibitor, gelonin, mitogellin, restrictocin, phenomycin, enomycin, and the tricothecenes. A variety of radionuclides are available for the production of radioconjugated antibodies. Examples include ²¹²Bi, ¹³¹I, ¹³¹n, ⁹⁰Y, and ¹⁸⁶Re.

[00477] Conjugates of the antibody and cytotoxic agent are made using a variety of bifunctional protein-coupling agents such as N-succinimidyl-3-(2-pyridyldithiol) propionate (SPDP), iminothiolane (IT), bifunctional derivatives of imidoesters (such as dimethyl adipimidate HCL), active esters (such as disuccinimidyl suberate), aldehydes (such as glutareldehyde), bis-azido compounds (such as bis (p-azidobenzoyl) hexanedi amine), bis- diazonium derivatives (such as bis-(p-diazoniumbenzoyl)-ethylenediamine), diisocyanates (such as tolyene 2,6-diisocyanate), and bis-active fluorine compounds (such as 1,5-difluoro- 2,4-dinitrobenzene). For example, a ricin immunotoxin can be prepared as described in Vitetta et al., Science 238: 1098 (1987), Carbon- 14-labeled l-isothiocyanatobenzyl-3- methyldiethylene triaminepentaacetic acid (MX-DTPA) is an exemplary chelating agent for conjugation of radionucleotide to the antibody. (See WO94/11026).

[00478] Those of ordinary skill in the art will recognize that a large variety of possible moieties can be coupled to the resultant antibodies of the invention. (See, for example, ‘"Conjugate Vaccines’’, Contributions to Microbiology and Immunology, J. M. Cruse and R. E. Lewis, Jr (eds), Carger Press, New Y ork, (1989), the entire contents of which are incorporated herein by reference).

[00479] Coupling may be accomplished by any chemical reaction that will bind the two molecules so long as the antibody and tire other moiety retain their respective activities. This linkage can include many chemical mechanisms, for instance covalent binding, affinity binding, intercalation, coordinate binding and complexation. The preferred binding is, however, covalent binding. Covalent binding can be achieved either by direct condensation of existing side chains or by the incorporation of external bridging molecules. Many bivalent or polyvalent linking agents are usefill in coupling protein molecules, such as the antibodies of the present invention, to other molecules. For example, representative coupling agents can include organic compounds such as thioesters, carbodiimides, succinimide esters, diisocyanates, glutaraldehyde, diazobenzenes and hexamethylene diamines. This listing is not intended to be exhaustive of the various classes of coupling agents known in tire art but, rather, is exemplary' of the more common coupling agents. (See Killen and Lindstrom, Jour. Immun. 133: 1335-2549 (1984); Jansen et al., Immunological Review's 62: 185-216 (1982); and Vitetta et al.. Science 238: 1098 (1987).

[00480] Preferred linkers are described in the literature. (See, for example, Ramakrishnan, S. et al., Cancer Res. 44:201-208 (1984) describing use of MBS (M-maleimidobenzoyl-N- hydroxysuccinimide ester). See also, U.S. Patent No. 5,030,719, describing use of halogenated acetyl hydrazide derivative coupled to an antibody by way of an oligopeptide linker. Particularly preferred linkers include: (i) EDC (l-ethyl-3-(3-dimethylamino-propyl) carbodiimide hydrochloride; (ii) SMPT (4-succinimidyloxycarbonyl-alpha-methyl-alpha-(2- pridyl-dithio)-toluene (Pierce Chem. Co., Cat. (21558G); (iii) SPDP (succmimidyl-6 [3-(2- pyridyldithio) propionamido]hexanoate (Pierce Chem. Co., Cat #21651G); (iv) Sulfo-LC- SPDP (sulfosuccinimidyl 6 [3-(2-pyridyldithio)-propianamide] hexanoate (Pierce Chem. Co. Cat. &2165-G); and (v) sulfo-NHS (N-hydroxysulfo-succinimide: Pierce Chem. Co., Cat. #24510) conjugated to EDC.

[00481] The linkers described above contain components that have different attributes, thus leading to conjugates with differing physio-chemical properties. For example, sulfo-NHS esters of alkyl carboxylates are more stable than sulfo-NHS esters of aromatic carboxylates. NHS-ester containing linkers are less soluble than sulfo-NHS esters. Further, the linker SMPT contains a sterically hindered disulfide bond, and can form conjugates with increased stability. Disulfide linkages, are in general, less stable than other linkages because the disulfide linkage is cleaved in vitro, resulting in less conjugate available, Sulfo-NHS, in particular, can enhance the stability of carbodimide couplings. Carbodimide couplings (such as EDC) when used in conjunction with sulfo-NHS, forms esters that are more resistant to hydrolysis than the carbodimide coupling reaction alone.

[00482] Tire antibodies disclosed herein can also be formulated as immunoliposomes. Liposomes containing the antibody are prepared by methods known in the art, such as described in Epstein et al.. Proc. Natl. Acad. Sei. USA, 82: 3688 (1985); Hwang et al.. Proc. Natl Acad. Sci. USA, 77: 4030 (1980); and U.S. Pat. Nos. 4,485,045 and 4,544,545. Liposomes with enhanced circulation time are disclosed in U.S. Patent No. 5,013,556.

[00483] Particularly useful liposomes can be generated by the reverse-phase evaporation method with a lipid composition comprising phosphatidylcholine, cholesterol, and PEG- derivatized phosphatidylethanolamine (PEG-PE). Liposomes are extruded through filters of defined pore size to yield liposomes with the desired diameter. Fab' fragments of the antibody of the present invention can be conjugated to the liposomes as described in Martin et al., J. Biol. Chem., 257: 286-288 ( 1982) via a disulfide-interchange reaction.

[00484] METHODS OF USE

[00485] Any of the bispecific antibodies of the disclosure (e.g. anti-CD3ε and anti-CD20 antibody; anti-CD3ε and anti-CD20 antibody with CD58 fusion peptide) an may be used in therapeutic methods. In some embodiments, a bispecific antibody may be for use as a medicament is provided. In some embodiments, a bispecific antibody for use in treating or delaying progression of a cell proliferative disorder (e.g., cancer) e.g., esophageal cancer or an adenocarcinoma) or an autoimmune disorder (e.g., arthritis) is provided. In some embodiments, a bispecific antibody for use in a method of treatment is provided. In some embodiments, the invention provides a bispecific antibody for use in a method of treating an individual having a cell proliferative disorder comprising administering to the individual an effective amount of the bispecific antibody . In some embodiments, the method further comprises administering to the individual an effective amount of at least one additional therapeutic agent, for example, as described below. In further embodiments, the invention provides a bispecific antibody for use in enhancing immune function in an individual having a cell proliferative disorder. In some embodiments, the invention provides a bispecific antibody for use in a method of enhancing immune function in an individual having a cell proliferative disorder comprising administering to the individual an effective of the bispecific antibody to activate effector cells (e.g., T cells, e.g., CD8+ and/or CD4+ T cells), expand (increase) an effector cell population, reduce a target cell population, and/or kill a target cell (e.g., target tumor cell). An "individual" according to any of the abo ve embodiments may be a human.

[00486 j In some aspects, the invention provides for the use of bispecific antibodies of the disclosure (e.g. anti-CD3e and anti-CD20 antibody; anti-CD3ε and anti-CD20 antibody with CD58 fusion peptide) in the manufacture or preparation of a medicament. In one embodiment, the medicament is for treatment of a cell proliferative disorder (e.g., cancer, e.g., esophageal cancer or an adenocarcinoma) or an autoimmune disorder (e.g., arthritis). In some embodiments, the medicament is for use in a method of treating a cell proliferative disorder or an autoimmune disorder comprising administering to an individual having a cell proliferative disorder or an autoimmune disorder an effective amount of the medicament. In some embodiments, the method further comprises administering to the individual an effective amount of at least one additional therapeutic agent, for example, as described below. In some embodiments, the medicament is for activating effector cells (e.g., T cells, e.g., CD8+ and/or CD4+ T cells), expanding (increasing) an effector cell population, reducing a target cell (e.g., a cell expressing CD20) population, and/or killing target cells (e.g., target tumor cells) in the individual. In some embodiments, the medicament is for use in a method of enhancing immune function in an individual having a cell proliferative disorder or an autoimmune disorder comprising administering to the individual an amount effective of the medicament to activate effector cells (e.g., T cells, e.g., CD8+ and/or CD4+ T cells), expand (increase) an effector cell population, reduce a target cell (e.g., a cell expressing CD20) population, and/or kill a target ceil (e.g., target tumor cell). An "individual" according to any of the above embodiments may be a human. [00487] In some embodiments, the invention provides a method for treating a cell proliferative disorder (e.g., cancer). In one embodiment, the method comprises administering to an individual having such a cell proliferative disorder an effective amount of an bispecific antibody. In some embodiments, the method further comprises administering to the individual an effective amount of at least one additional therapeutic agent, for example, as described below. An "individual" according to any of the above embodiments may be a human. In a further aspect, the invention provides a method for enhancing immune function in an individual having a cell proliferative disorder or an autoimmune disorder in an individual having a cell proliferative disorder or an autoimmune disorder. In one embodiment, the method comprises administering to the individual an effective amount of a bispecific antibody to activate effector cells (e.g., T cells, e.g., CD8+ and/or CD4+ T cells), expand (increase) an effector cell population, reduce a target cell (e.g., a cell expressing CD20) population, and/or kill a target cell (e.g., target tumor cell). [00488] In some embodiments, the invention provides a method for treating a cancer that expresses CD2.0. In some embodiments, the cancer that expressed CD20 is a hematological cancer, such as a B cell cancer (for example, mature B-cell lymphoma). In some embodiments, the method of treating a cancer comprises administering an effective amount of the bispecific antibody of the disclosure (e.g. anti-CD3ε and anti-CD20 antibody; anti- CD3ε and anti-CD20 antibody with CD58 fusion peptide). In a further aspect of the embodiment, the mature B-cell lymphoma is a Nou-Hodgkin's Lymphoma (NHL). In a further aspect of the embodiment, the NHL is selected from tire group comprising: germinal-center B-cell-like (GCB) DLBCL, activated B-cell like (ABC) DLBCL, follicular lymphoma (FL), mantle cell lymphoma (MCL), acute myeloid leukemia (AML), chronic lymphoid leukemia (CLL), marginal zone lymphoma (MZL), small lymphocytic leukemia (SLL), lymphoplasmacytic lymphoma (LL), Waldenstrom macroglobulinemia (W), central nervous system lymphoma (CNSL), Burkitt's lymphoma (BL), B-cell prolymphocytic leukemia, Splenic marginal zone lymphoma, Hairy cell leukemia, Splenic lymphoma/Ieukemia, unclassifiable, Splenic diffuse red pulp small B-cell lymphoma, Hairy cell leukemia variant, Waldenstrom macroglobulinemia, Heavy chain diseases, a Heavy' chain disease, y Heavy chain disease, p Heavy chain disease, Plasma cell myeloma, Solitary' plasmacytoma of bone, [Extraosseous plasmacytoma, Extranodal marginal zone lymphoma of mucosa-associated lymphoid tissue (MALT lymphoma), N odal marginal zone lymphoma, Pediatric nodal marginal zone lymphoma, Pediatric follicular lymphoma. Primary' cutaneous follicle centre lymphoma, T-cell/histiocyte rich large B-cell lymphoma, Primary DLBCL of the CNS, Primary' cutaneous DLBCL, leg type, EBV-positive DLBCL of the elderly, DLBCL associated with chronic inflammation, Lymphomatoid granulomatosis, Primary’ mediastinal (thymic) large B-cell lymphoma, Intravascular large B-cell lymphoma, ALK-positive large B-cell lymphoma, Plasmablastic lymphoma, Large B-cell lymphoma arising in HHV8-associated multicentric Castleman disease, Primary effusion lymphoma: B-cell lymphoma, unclassifiable, with features intermediate between diffuse large B-cell lymphoma and Burkit lymphoma, and B-cell lymphoma, unclassifiable, with features intermediate between diffuse large B-cell lymphoma and classical Hodgkin lymphoma. In some embodiments of tire invention, the method comprises treating a cancer comprising germinal -center B-cell like (GCB) DLBCL, activated B-cell-like (ABC) DLBCL, follicular lymphoma (FL), mantle cell lymphoma (MCL), acute myeloid leukemia (AML), chronic lymphoid leukemia (CLL), marginal zone lymphoma (MZL), small lymphocytic leukemia (SLL), lymphoplasmacytic lymphoma (LL), Waldenstrom macroglobulinemia (WM), central nervous system lymphoma (CNSL), or Burkit's lymphoma (BL).

^10489^ In some embodiments, the invention provides a method for treating an inflammatory' disease or a immunological disease. Exemplary inflammatory or immunological diseases include but are not limited to neurological diseases, pulmonological diseases, rheumatological diseases, hematological diseases, dermatological diseases, nephrological diseases and endocrinological diseases. Exemplary neurological disease include but are not limited to myasthenia gravis, relapsing-remiting multiple sclerosis (RMS), primary progressive multiple sclerosis (PPMS) and neuromyelitis optica.

Exemplary' pulmonological diseases include but are not limited to progressi ve pulmonary fibrosis including either usual interstitial pneumonia ( U IP) and non-specific interstitial pneumonia (NSIP), and idiopathic pulmonary fibrosis. Exemplary' rheumatological diseases include but are not limited to ANCA-associated vasculitis, Behcet’s syndrome, Castelman’s disease, cryoglobulinemia, granulomatosis with polyangiitis (GPA), IgG4-mediated diseases, juvenile idiopathic arthritis, microscopic polyangiitis (MPA), myositis, rheumatoid arthritis, SLE, Sjogren’s syndrome, and Systemic sclerosis/ scleroderma. Exemplary' hematological diseases include but are not limited to antiphospholipid syndrome, autoimmune hemoh tic anemia, immune thrombocytopenic purpura. Exemplary dermatological diseases include but are not limited to acquired angioedema with Cl deficiency, bullous pemphigoid and pemphigus vulgaris. Exemplary nephrological diseases include but are not limited to IgA nephropathy, membranous nephropathy and nephrotic syndrome. Exemplary endocrinological diseases include but are not limited to autoimmune hepatitis. Graves’ disease and Type 1 Diabetes.

[00490] In some embodiments, the invention provides pharmaceutical formulations comprising any of the bispecific antibodies provided herein, e.g, for use in any of the above therapeutic methods. In one embodiment, a pharmaceutical formulation comprises any of die bispecific antibodies provided herein and a pharmaceutically acceptable carrier. In another embodiment, a pharmaceutical formulation comprises any of the bispecific antibodies provided herein and at least one additional therapeutic agent, for example, as described herein.

[00491] Antibodies of the invention can be used either alone or m combination with other agents in a therapy. For instance, an antibody of the invention may be co-adm mistered with at least one additional therapeutic agent. In some embodiments, an additional therapeutic agent is a chemotherapeutic agent, growth inhibitory agent, cytotoxic agent, agent used in radiation therapy, anti-angiogenesis agent, apoptotic agent, anti-tubulin agent, or other agent, such as a epidermal growth factor receptor (EGFR) antagonist (e.g, a tyrosine kinase inhibitor), HER1/EGFR inhibitor (e.g, erlotinib (Tarceva™), platelet derived growth factor inhibitor (e.g, Gleevec™ (Imatinib Mesylate)), a COX-2 inhibitor (e.g, celecoxib), interferon, cytokine, antibody' other than the anti-CD3 antibody' of the invention, such as an antibody that bind to one or more of the following targets ErbB2, ErbB3, ErbB4, PDGFR- beta, BlyS, APRIL, BCMA VEGF, or VEGF receptor(s), TRAIL/Apo2, PD-1 (e.g.

Nivolumab, Pembrolizumab, Cemiplimab), PD-L1 (Atezolizumab, Avelumab, Durvalumab), PD-L2, or another bioactive or organic chemical agent. In some embodiments, the invention provides a method wherein the additional therapeutic agent is a glucocorticoid. In one embodiment, the glucocorticoid is dexamethasone.

[00492] In some embodiments, the additional therapeutic agent is a monoclonal anti-CD20 antibody. Exemplary- anti-CD20 antibodies include but are not limited to rituximab (RITUXAN®), obinutuzumab (GAZYVA®), tocilizumab (ACTEMRA® / RoACTEMRA®), ocrelizumab (OCREVUS®), ofatumumab (KESIMPTA®), ibritumomab (ZEA AL1N®). In some embodiments, the additional therapeutic agent is rituximab.

[00493] In some embodiments, the additional therapeutic agent is a checkpoint inhibitor. The term “inhibition” or “inhibitor” includes a reduction m a certain parameter, e.g., an activity, of a given molecule, e.g., an immune checkpoint inhibitor. For example, inhibition of an activity, e.g., an activity of, e.g., PD-1, PD-L1, CTLA-4, TIM-3, CEACAM (e.g., CEACAM-1, CEACAM-3 and/or CEACAM-5), LAG-3, VISTA, BTLA, TIGIT, LAIR1, CD160, 2B4 or TGFR beta, of at least 5%, 10%, 20%, 30%, 40% or more is included by this term. Tire level of inhibition need not be 100%.

[00494] In some embodiments, the checkpoint inhibitor is a PD-1 inhibitor. In some embodiments, the PD-1 inhibitor is an anti-PDl antibody. In some embodiments, the PD-1 inhibitor is an anti PD-1 monoclonal antibody. Exemplary anti -PD-1 monoclonal antibodies include, but are not limited to cemiplimab (Libtayo), nivolumab (Opdivo), pembrolizumab (Keytruda). In some embodiments, the checkpoint inhibitor is a PD-L1 inhibitor. Exemplary PD-L1 inhibitors include but are not lim ited to avelumab (Bavencio), durvalumab (Imfinzi) and atezolizumab (Tecentriq).

[00495] Such combination therapies noted above encompass combined administration (where two or more therapeutic agents are included in the same or separate formulations), and separate administration, in which case, administration of the antibody of the invention can occur prior to, simultaneously, and/or following, administration of the additional therapeutic agent or agents. In one embodiment, administration of the bispecific antibody and administration of an additional therapeutic agent occur within about one month, or within about one, two or three weeks, or -within about one, two, three, four, five, or six days, of each other. Bispecific antibodies of the disclosure can also be used in combination with radiation therapy.

[00496] In some embodiments, the additional therapeutic agent is a chimeric antigen receptor (CAR) T cell therapy. In some embodiments, the CAR-T cell therapy specifically binds CD 19. Exemplary' CAR-T cell therapies that specifically bind CD 19 include but are not limited to BREYANZI® (lisocabtagene maraleucel), TECARTUS™ (brexucabtagene autoleucel), KYMRIAH™ (tisagenlecleucel), YESCARTA™ (axicabtagene ciloleucel), ABECMA® (idecabtagene vicleucel), or CARVYKTI™ (ciltacabtagene autoleucel). [00497] An antibody of the invention (and/or any add itional therapeutic agent) can be administered by any suitable means, including parenteral, intrapulmonary, and intranasal, and, if desired for local treatment, intralesional administration. Parenteral infusions include intramuscular, intravenous, intraarterial, intraperitoneal, or subcutaneous administration. In some embodiments, the antibody is administered by subcutaneous administration. In some embodiments, an anti-CD3ε antibody administered by subcutaneous injection exhibits a less toxic response m a patient than the same anti-CD3ε antibody administered by intravenous injection. Dosing can be by any suitable route, for example, by injections, such as intravenous or subcutaneous injections, depending in part on whether the administration is brief or chronic. Various dosing schedules including but not limited to single or multiple administrations over various time-points, bolus administration, and pulse infusion are contemplated herein.

[00498] Antibodies of the invention would be formulated, dosed, and administered in a fashion consistent with good medical practice. Factors for consideration in this context include the particular disorder being treated, the particular mammal being treated, the clinical condition of the individual patient, the cause of the disorder, the site of delivery of die agent, the method of administration, the scheduling of administration, and other factors known to medical practitioners. The antibody need not be, but is optionally formulated with one or more agents currently used to prevent or treat the disorder in question. The effective amount of such other agents depends on the amount of antibody present in the formulation, the type of disorder or treatment, and other factors discussed above. These are generally used in the same dosages and with administration routes as described herein, or about from 1 to 99% of the dosages described herein, or in any dosage and by any route that is empirically/clinically determined to be appropriate.

[00499] For the prevention or treatment of disease, the appropriate dosage of an antibody of the invention (when used alone or in combination with one or more other additional therapeutic agents) will depend on the type of disease to be treated, the type of antibody, the severity and course of the disease, whether the antibody is administered for preventive or therapeutic purposes, previous therapy, the patient's clinical history’ and response to the antibody, and the discretion of the atending physician. Tire antibody is suitably administered to the patient at one time or over a series of treatments. [00500] As a general proposition, the therapeutically effecti ve amount of the bispecific anti- CD3ε and anti-CD20 antibody administered to human will be in the range of about 0,01 to about 100 mg/kg of patient body weight whether by one or more administrations. In some embodiments, the antibody used is about 0.01 to about 45 mg/kg, about 0.01 to about 40 mg kg, about 0.01 to about 35 mg/kg. about 0.01 to about 30 mg/kg, about 0.01 to about 25 mg/kg, about 0.01 to about 20 mg/kg, about 0.01 to about 15 mg kg, about 0.01 to about 10 mg/kg, about 0.01 to about 5 mg/kg, or about 0.01 to about 1 mg/kg administered daily, for example. In one embodiment, an bispecific antibody described herein is administered to a human at a dose of about 100 mg, about 200 mg, about 300 mg, about 400 mg, about 500 mg, about 600 mg, about 700 mg, about 800 mg, about 900 mg, about 1000 mg, about 1100 mg, about 1200 mg, about 1300 mg or about 1400 mg on day 1 of 21 -day cycles. The dose may be administered as a single dose or as multiple doses (e.g., 2 or 3 doses), such as infusions. For repeated administrations over several days or longer, depending on the condition, the treatment would generally be sustained until a desired suppression of disease symptoms occurs. One exemplary’ dosage of the antibody' would be in the range from about 0.05 mg/kg to about 10 mg/kg. Tirus, one or more doses of about 0.5 mg/kg, 2.0 mg/kg, 4.0 mg kg, or 10 mg/kg (or any combination thereof) may be administered to the patient. Such doses may be administered intermittently, for example, every week or every’ three weeks (e.g., such that the patient receives from about two to about twenty, or, for example, about six doses of the bispecific antibody). An initial higher loading dose, followed by' one or more lower doses may be administered. The progress of this therapy is easily monitored by conventional techniques and assays.

[00501 ] In some embodiments, the methods may further comprise an additional therapy. The additional therapy may be radiation therapy, surgery', chemotherapy, gene therapy, DMA therapy, viral therapy, RNA therapy, immunotherapy, bone marrow transplantation, nanotherapy, monoclonal antibody' therapy, or a combination of the foregoing. The additional therapy may be in the form of adjuvant or neoadjuvant therapy. In some embodiments, the additional therapy is the administration of small molecule enzymatic inhibitor or anti -metastatic agent. In some embodiments, the additional therapy is the administration of side-effect limiting agents (e.g., agents intended to lessen the occurrence and/or severity of side effects of treatment, such as anti-nausea agents, etc.). In some embodiments, the additional therapy' is radiation therapy. In some embodiments, the additional therapy is surgery. In some embodiments, the additional therapy is a combination of radiation therapy and surgery. In some embodiments, the additional therapy is gamma irradiation. In some embodiments, the additional therapy may be a separate administration of one or more of the therapeutic agents described above.

[00502] It will be appreci ated that administration of therapeutic entities in accordance with the invention will be administered with suitable carriers, excipients, and other agents that are incorporated into formulations to provide improved transfer, delivery, tolerance, and the like. A multitude of appropriate formulations can be found in the formulary’ known to all pharmaceutical chemists: Remington's Pharmaceutical Sciences (15th ed, Mack Publishing Company, Easton, PA (1975)), particularly Chapter 87 by Blaug, Seymour, therein. These formulations include, for example, powders, pastes, ointments, jellies, waxes, oils, lipids, lipid (cationic or anionic) containing vesicles (such as Lipofectin™), DNA conjugates, anhydrous absorption pastes, oil-in-water and water-in-oil emulsions, emulsions carbowax (polyethylene glycols of various molecular weights), semi-solid gels, and semi-solid mixtures containing carbow ax . Any of the foregoing mixtures may be appropriate in treatments and therapies in accordance with the present invention, provided that the active ingredient in the formulation is not inactivated by the formulation and the formulation is physiologically compatible and tolerable with the route of administration. See also Baldrick P. “Pharmaceutical excipient development: the need for preclinical guidance.” Regul.

Toxicol Pharmacol. 32(2):210-8 (2000), Wang W. “Lyophilization and development of solid protein pharmaceuticals.” Int. J. Pharm. 203(1-2): 1-60 (2000), Charman WN “Lipids, lipophilic drags, and oral drug delivery-some emerging concepts.” J Pharm Sci. 89(8):967- 78 (2000), Powell et al. “Compendium of excipients for parenteral formulations” PDA J Pharm Sci Technol. 52:238-31 1 (1998) and the citations therein for additional information related to formulations, excipients and carriers well known to pharmaceutical chemists. [00503] Therapeutic formulations of the invention, which include an antibody of the invention, are used to treat or alleviate a symptom associated with a cancer, such as, by way of non-limiting example, leukemias, lymphomas, breast cancer, colon cancer, ovarian cancer, bladder cancer, prostate cancer, glioma, lung & bronchial cancer, colorectal cancer, pancreatic cancer, esophageal cancer, liver cancer, urinary bladder cancer, kidney and renal pelvis cancer, oral cavity & pharynx cancer, uterine corpus cancer, and/or melanoma The present invention also provides methods of treating or alleviating a symptom associated with a cancer. A therapeutic regimen is carried out by identifying a subject, e.g., a human patient suffering from (or at risk of developing) a cancer, using standard methods.

[00504 [ Efficaciousness of treatment is determined in association with any known method for diagnosing or treating the particular immune-related disorder. Alleviation of one or more symptoms of the immune -related disorder indicates that the antibody confers a clinical benefit.

[00505] Methods for the screening of antibodies that possess the desired specificity include, but are not limited to, enzyme linked immunosorbent assay (EL1SA) and other immunologically mediated techniques known within the art.

[00506j Antibodies directed against a target such as CD3ε, CD20, or a combination thereof (or a fragment thereof), may be used in methods known within the art relating to tire localization and/or quantitation of these targets, e.g., for use in measuring levels of these targets within appropriate physiological samples, for use in diagnostic methods, for use in imaging the protein, and the like). In a given embodiment, antibodies specific any of these targets, or derivative, fragment, analog or homolog thereof, that contain the antibody derived antigen binding domain, are utilized as pharmacologically active compounds (referred to hereinafter as “Therapeutics”).

[00507J An antibody of the invention can be used to isolate a particular target using standard techniques, such as immunoaffinity, chromatography or immunoprecipitation. Antibodies of the invention (or a fragment thereof) can be used diagnostically to monitor protein levels m tissue as part of a clinical testing procedure, e.g., to determine the efficacy of a given treatment regimen. Detection can be facilitated by coupling (i.e., physically linking) the antibody to a detectable substance. Examples of detectable substances include various enzymes, prosthetic groups, fluorescent materials, luminescent materials, bioluminescent materials, and radioactive materials. Examples of suitable enzymes include horseradish peroxidase, alkaline phosphatase, p-galactosidase, or acetylcholinesterase; examples of suitable prosthetic group complexes include streptavidin/biotin and avidin/biotin; examples of suitable fluorescent materials include urnbelliferone, fluorescein, fluorescein isothiocyanate, rhodamine, dichlorotriazinylamine fluorescein, dansyl chloride or phycoerythrin; an example of a luminescent material includes luminol; examples of bioluminescent materials include luciferase, luciferin, and aequorin, and examples of suitable radioactive material include ¹³¹1, ^3sS or ³H. [00508] Antibodies of the invention, including polyclonal, monoclonal, humanized and fully human antibodies, may be used as therapeutic agents. Such agents will generally be employed to treat or prevent a disease or pathology associated with aberrant expression or activation of a given target in a subject. An antibody preparation, preferably one having high specificity and high affinity for its target antigen, is administered to the subject and will generally have an effect due to its binding with the target. Administration of the antibody may abrogate or inhibit or interfere with the signaling function of the target. Administration of the antibody may abrogate or inhibit or interfere with the binding of the target with an endogenous ligand to which it naturally binds.

[00509] A therapeutically effective amount of an antibody of the invention relates generally to the amount needed to achieve a therapeutic objective. As noted above, this may be a binding interaction between the antibody and its target antigen that, in certain cases, interferes with the functioning of the target. The amount required to be administered will furthermore depend on the binding affinity of the antibody for its specific antigen, and will also depend on the rate at which an administered antibody is depleted from the free volume other subject to which it is administered. Common ranges for therapeutically effective dosing of an antibody or antibody fragment of the invention may be, by way of nonlimiting example, from about 0.1 mg/kg body weight to about 50 mg/kg body weight. Common dosing frequencies may range, for example, from twice daily to once a week.

[00510] Antibodies or a fragment thereof of the invention can be administered for the treatment of a variety of diseases and disorders in the form of pharmaceutical compositions. Principles and considerations involved in preparing such compositions, as well as guidance in the choice of components are provided, for example, in Remington: Tire Science And Practice Of Pharmacy 19th ed. (Alfonso R. Gennaro, et al., editors) Mack Pub. Co., Easton, Pa.: 1995; Drug Absorption Enhancement: Concepts, Possibilities, Limitations, And Trends, Harwood Academic Publishers, Langhome, Pa., 1994; and Peptide And Protein Drag Deliver}' (Advances In Parenteral Sciences, Vol. 4), 1991, M. Dekker, New York.

[00511] Where antibody fragments are used, the smallest inhibitory fragment that specifically binds to the binding domain of the target protein is preferred. For example, based upon the variable-region sequences of an antibody, peptide molecules can be designed that retain the ability to bind the target protein sequence. Such peptides can be synthesized chemically and/or produced by recombinant DNA technology. (See, e.g., Marasco et al., Proc. Natl. Acad. Sci. USA, 90: 7889-7893 (1993)). The formulation can also contain more than one active compound as necessary for the particular indication being treated, preferably those with complementary activities that do not adversely affect each other. Alternatively, or m addition, the composition can comprise an agent that enhances its function, such as, for example, a cytotoxic agent, cytokine, chemotherapeutic agent, or growth-inhibitory agent. Such molecules are suitably present in combination in amounts that are effective for tire purpose intended.

[00512] The active ingredients can also be entrapped in microcapsules prepared, for example, by coacervation techniques or by interfacial polymerization, for example, hydroxymethylcellulose or gelatin-microcapsules and poly-(methylmethacrylate) microcapsules, respectively, in colloidal drug delivery systems (for example, liposomes, albumin microspheres, microemulsions, nano-particles, and nanocapsules) or in macroem ulsions .

[00513] The formulations to be used for m vivo administration must be sterile. This is readily' accomplished by' filtration through sterile filtration membranes.

[00514] Sustained-release preparations can be prepared. Suitable examples of sustained- release preparations include semipermeable matrices of solid hydrophobic polymers containing the antibody, which matrices are in the form of shaped articles, e.g., films, or microcapsules. Examples of sustained-release matrices include polyesters, hydrogel s (for example, poly(2-hydroxyethyl-methacrylate), or poly(vinylalcohol)), polylactides (U.S. Pat. No. 3,773,919), copolymers of L-glutamic acid and γ ethyl-L-glutamate, non-degradable ethylene-vinyl acetate, degradable lactic acid-glycolic acid copolymers such as the LUPRON DEPOT (injectable microspheres composed of lactic acid -glycolic acid copolymer and leuprolide acetate), and poly-D-(-)-3 -hydroxybutyric acid. While polymers such as ethylene-vinyl acetate and lactic acid-glycolic acid enable release of molecules for over 100 days, certain hydrogels release proteins for shorter time periods.

[00515] An antibody according to the invention can be used as an agent for detecting the presence of a given target (or a protein fragment thereof) in a sample. In some embodiments, the antibody’ contains a detectable label. Antibodies are polyclonal, or more preferably, monoclonal. An intact antibody, or a fragment thereof (e.g. , Fab, scFv, or F(ab)2) is used. The term “labeled”, with regard to the probe or antibody, is intended to encompass direct labeling of the probe or antibody by coupling (i.e., physically linking) a detectable substance to the probe or antibody, as well as indirect label ing of the probe or antibody by reactivity with another reagent that is directly labeled. Examples of indirect labeling include detection of a primary antibody using a fluorescently-labeled secondary antibody and endlabeling of a DNA probe with biotin such that it can be detected with fluorescently -labeled streptavidin. The term "‘biological sample” is intended to include tissues, cells and biological fluids isolated from a subject, as well as tissues, cells and fluids present within a subject. Included within the usage of the term “biological sample”, therefore, is blood and a fracti on or component of blood including blood serum, blood plasm a, or lymph. That is, the detection method of the invention can be used to detect an analyte mRNA, protein, or genomic DNA in a biological sample in vitro as well as in vivo. For example, in vitro techniques for detection of an analyte mRNA include Northern hybridizations and in situ hybridizations. In vitro techniques for detection of an analyte protein include enzyme linked immunosorbent assays (EL1SAs), Western blots, immunoprecipitations, and immunofluorescence. In vitro techniques for detection of an analyte genomic DNA include Southern hybridizations. Procedures for conducting immunoassays are described, for example in “EL1SA: Theory and Practice: Methods in Molecular Biology”, Vol. 42, J. R. Crowther (Ed.) Human Press, Totowa, NJ, 1995; “Immunoassay”, E. Diamandis and T. Christopoulus, Academic Press, Inc., San Diego, CA, 1996; and “Practice and Theory of Enzyme Immunoassays”, P. Tijssen, Elsevier Science Publishers, Amsterdam, 1985.

Furthermore, in vivo techniques for detection of an analyte protein include introducing into a subject a labeled anti-analyte protein antibody. For example, the antibody can be labeled with a radioactive marker whose presence and location in a subject can be detected by standard imaging techniques.

[00516] PHARMACEUTICAL COMPOSITIONS

[00517] The antibodies of the invention (also referred to herein as “active compounds”), and deri vatives, fragments, analogs and homologs thereof, can be incorporated into pharmaceutical compositions suitable for administration. Such compositions typically comprise the antibody and a pharmaceutically acceptable carrier. As used herein, the term “pharmaceutically' acceptable carrier” is intended to include any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like, compatible with pharmaceutical administration. Suitable carriers are described in the most recent edition of Remington’s Pharmaceutical Sciences, a standard reference text in the field, which is incorporated herein by reference. Preferred examples of such carriers or diluents include, but are not limited to, water, saline, ringer’s solutions, dextrose solution, and 5% human serum albumin. Liposomes and non-aqueous vehicles such as fixed oils may also be used. The use of such media and agents for pharmaceutically active substances is well known in the art. Except insofar as any conventional media or agent is incompatible with the active compound, use thereof in the compositions is contemplated. Supplementary active compounds can also be incorporated into the compositions.

[00518] A pharmaceutical composition of the invention is formulated to be compatible with its intended route of administration. Examples of routes of administration include parenteral, e.g., intravenous, intradermal, subcutaneous, oral (e.g., inhalation), transdermal (i.e., topical), transmucosal, and rectal administration. Solutions or suspensions used for parenteral, intradermal, or subcutaneous application can include the following components: a sterile diluent such as water for injection, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents; antibacterial agents such as benzyl alcohol or methyl parabens; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as ethylenediarninetetraacetic acid (EDTA); buffers such as acetates, citrates or phosphates, and agents for the adjustment of tonicity such as sodium chloride or dextrose. The pH can be adjusted with acids or bases, such as hydrochloric acid or sodium hydroxide. The parenteral preparation can be enclosed in ampoules, disposable syringes or multiple dose vials made of glass or plastic.

[00519] Pharmaceutical compositions suitable for injectable use include sterile aqueous solutions (where water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersion. For intravenous administration, suitable carriers include physiological saline, bacteriostatic water, Cremophor EL™ (BASF, Parsippany, N . J.) or phosphate buffered saline (PBS). In all cases, the composition must be sterile and should be fluid to the extent that easy syringeability exists. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms such as bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol, and the like), and suitable mixtures thereof. The proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersion and by the use of surfactants. Prevention of the action of microorganisms can be achieved by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like. In many cases, it will be preferable to include isotonic agents, for example, sugars, polyalcohols such as manitol, sorbitol, sodium chloride in the composition. Prolonged absorption of the injectable compositions can be brought about by including in the composition an agent which delays absorption, for example, aluminum monostearate and gelatin.

[00520] Sterile injectable solutions can be prepared by incorporating the active compound in the required amount in an appropriate solvent with one or a combination of ingredients enumerated above, as required, followed by filtered sterilization. Generally, dispersions are prepared by incorporating the active compound into a sterile vehicle that contains a basic dispersion medium and the required other ingredients from those enumerated above. In the case of sterile powders for the preparation of sterile injectable solutions, methods of preparation are vacuum dry ing and freeze-drying that yi elds a powder of the active ingredient plus any additional desired ingredient from a previously sterile-filtered solution thereof.

[00521] Oral compositions generally include an inert diluent or an edible carrier. They can be enclosed in gelatin capsules or compressed into tablets. For the purpose of oral therapeutic administration, the active compound can be incorporated with excipients and used in the form of tablets, troches, or capsules. Oral compositions can also be prepared using a fluid carrier for use as a mouthwash, wherein the compound in the fluid carrier is applied orally and swished and expectorated or swallowed. Pharmaceutically compatible binding agents, and/or adjuvant materials can be included as part of the composition. The tablets, pills, capsules, troches and the like can contain any of the following ingredients, or compounds of a similar nature: a binder such as microcrystalline cellulose, gum tragacanth or gelatin; an excipient such as starch or lactose, a disintegrating agent such as alginic acid, Primogel, or corn starch; a lubricant such as magnesium stearate or Sterotes; aglidant such as colloidal silicon dioxide; a sweetening agent such as sucrose or saccharin; or a flavoring agent such as peppermint, methyl salicylate, or orange flavoring. [00522] For administration by inhalation, the compounds are delivered in the form of an aerosol spray from pressured container or dispenser which contains a suitable propellant, e.g., a gas such as carbon dioxide, or a nebulizer.

[00523] Systemic administration can also be by transmucosal or transdermal means. For transmucosal or transdermal administration, penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art, and include, for example, for transmucosal administration, detergents, bile salts, and fusidic acid derivatives. Transmucosal administration can be accomplished through the use of nasal sprays or suppositories. For transdermal administration, the active compounds are formulated into ointments, salves, gels, or creams as generally known in the art.

[00524] The compounds can also be prepared in the form of suppositories (e.g., with conventional suppository bases such as cocoa butter and other glycerides) or retention enemas for rectal delivery.

[00525] In one embodiment, the active compounds are prepared with carriers that will protect the compound against rapid elim ination from the body, such as a controlled release formulation, including implants and microencapsulated delivery' systems. Biodegradable, biocompatible polymers can be used, such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid. Methods for preparation of such formulations will be apparent to those skilled in the art. The materials can also be obtained commercially from Alza Corporation and Nova Pharmaceuticals, Inc. Liposomal suspensions (including liposomes targeted to infected cells with monoclonal antibodies to viral antigens) can also be used as pharmaceutically acceptable carriers. These can be prepared according to methods known to those skilled in the art, for example, as described in U.S. Patent No. 4,522,811.

[00526] It is especially advantageous to formulate oral or parenteral compositions m dosage unit form for ease of administration and uniformity' of dosage. Dosage unit form as used herein refers to physically discrete units suited as unitary' dosages for the subject to be treated; each unit containing a predetermined quantity of active compound calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier. The specification for the dosage unit forms of the invention are dictated by and directly dependent on the unique characteristics of the active compound and the particular therapeutic effect to be achieved, and the limitations inherent in the art of compounding such an active compound for the treatment of individuals.

[00527] The pharmaceutical compositions can be included in a container, pack, or dispenser together with instructions for administration.

[00528] The invention will be further described in the following examples, which do not limit the scope of the invention described in the claims,

[00529] DEFINITIONS

[00530] Unless otherwise defined, scientific and technical terms used in connection with the present invention shall have the meanings that are commonly understood by those of ordinary' skill in the art. Further, unless otherwise required by context, singular terms shall include pluralities and plural tenns shall include the singular. Generally, nomenclatures utilized in connection with, and techniques of, cell and tissue culture, molecular biology, and protein and oligo- or polynucleotide chemistry and hybridization described herein are those -well-known and commonly used in the art. Standard techniques are used for recombinant DNA, oligonucleotide synthesis, and tissue culture and transformation (e.g., electroporation, lipofection). Enzymatic reactions and purification techniques are performed according to manufacturer's specifications or as commonly accomplished in the art or as described herein. Tire foregoing techniques and procedures are generally performed according to conventional methods well known in the art and as described in various general and more specific references that are cited and discussed throughout the present specification. See e.g., Sambrook et al. Molecular Cloning: A Laboratory' Manual (2d ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (1989)). The nomenclatures utilized in connection with, and the laboratory procedures and techniques of, analytical chemistry, synthetic organic chemistry, and medicinal and pharmaceutical chemistry described herein are those well-known and commonly used in the art. Standard techniques are used for chemical syntheses, chemical analyses, pharmaceutical preparation, formulation, and delivery', and treatment of patients.

[00531] As utilized in accordance with the present disclosure, the following terms, unless otherwise indicated, shall be understood to have the following meanings: [00532] As used herein, the term “antibody” refers to immunoglobulin molecules and immunologically active portions of immunoglobulin (Ig) molecules, i.e., molecules that contain an antigen binding site that specifically binds (immunoreacts with) an antigen. By “specifically bind” or “immunoreacts with” or “immunospecifically bind” is meant that the antibody reacts with one or more antigenic determinants of the desired antigen and does not react with other polypeptides or binds at much lower affinity (Kd > 10'⁶). Antibodies include, but are not limited to, polyclonal, monoclonal, chimeric, dAb (domain antibody), single chain, Fab, Fab’ and F(ab’)2 fragments, scFvs, and an Fab expression library.

[00533] The basic antibody structural unit is known to comprise a tetramer. Each tetramer is composed of two identical pairs of polypeptide chains, each pair having one “light” (about 25 kDa) and one “heavy” chain (about 50-70 kDa). The ammo-terminal portion of each chain includes a variable region of about 100 to 110 or more amino acids primarily responsible for antigen recognition. The carboxy-temiinal portion of each chain defines a constant region primarily responsible for effector function. In general, antibody molecules obtained from humans relate to any of the classes IgG, IgM, IgA, IgE and IgD, which d iffer from one another by the nature of the heavy chain present in the molecule. Certain classes have subclasses as well, such as IgGi, IgGr, IgG4 and others. Furthermore, in humans, the light chain may be a kappa chain or a lambda chain.

[00534] The term “monoclonal antibody” (MAb) or “monoclonal antibody composition”, as used herein, refers to a population of antibody molecules that contain only one molecular species of antibody molecule consisting of a unique light chain gene product and a unique heavy chain gene product. In particular, the complementarity determining regions (CDRs) of the monoclonal antibody are identical in all the molecules of the population. MAbs contain an antigen binding site capable of immunoreacting with a particular epitope of the antigen characterized by a unique binding affinity for it.

[00535] The term “antigen binding region” or “antigen-binding site” or “binding portion” refers to the part of the immunoglobulin molecule that participates in antigen binding. Tire antigen binding site is formed by amino acid residues of the N-terminal variable (“V”) regions of the heavy (“H”) and light (“L”) chains. Three highly divergent stretches within the V regions of the heavy and light chains, referred to as “hypervariable regions,” are interposed between more conserved flanking stretches known as “framework regions,” or “FRs”. Tirus, the term “FR” refers to amino acid sequences which are naturally found between, and adjacent to, hypervariable regions in immunoglobulins. In an antibody molecule, the three hypervariable regions of a light chain and the three hypervariable regions of a heavy chain are disposed relative to each other in three dimensional space to form an antigen-binding surface. The antigen-binding surface is complementary' to the three-dimensional surface of a bound antigen, and the three hypervariable regions of each of the heavy and light chains are referred to as “complementarity-determining regions,” or “CDRs.” Various methods are known in the art for numbering tire amino acids sequences of antibodies and identification of the complementary determining regions. For example, the Kabat numbering system (See Rabat, E.A., et al.. Sequences of Protein of immunological interest, Fifth Edition, US Department of Health and Human Services, US Government Printing Office (1991)) or the IMGT numbering system (See IMGTy the international ImMunoGeneTics information system^® . Available online: http://www.imgt.org/). The IMGT numbering system is routinely used and accepted as a reliable and accurate system in the art to determine amino acid positions in coding seq uences, alignment of alleles, and to easily compare sequences in immunoglobulin (IG) and T-cell receptor (TR) from all vertebrate species. The accuracy and the consistency of the IMGT data are based on IMGT- 0NT0L0GY, the first, and so far unique, ontology for immunogenetics and immunoinformatics (See Lefranc. M.P. et al., Biomolecules, 2014 Dec; 4(4), 1102-1139). IMGT tools and databases ran against IMGT reference directories built from a large repository' of sequences. In the IMGT system the IG V-DOMAIN and IG C -DOMAIN are delimited taking into account the exon delimitation, whenever appropriate. The re lore, the availability’ of more sequences to the IMGT database, the IMGT exon numbering system can be and “is used” by those skilled in the art reliably to determine ammo acid positions in coding sequences and for alignment of alleles. Additionally, correspondences between the IMGT unique numbering with other numberings (i.e., Rabat) are available in the IMGT Scientific chart (See Lefranc. M.P. et al., B iomolecules, 2014 Dec; 4(4), 1102-1139). [00536] The term "hvpcrvanablc region" or “variable region” refers to the ammo acid residues of an antibody that are typically responsible for antigen-binding. The hypervariable region generally comprises amino acid residues from a "complementarity determining region" or "CDR" (e.g., around about residues 24-34 (L1), 50-56 (L2) and 89-97 (L3) in the VL, and around about 31-35 (HI), 50-65 (H2) and 95-102 (H3) in the VH when numbered in accordance with the Kabat numbering system; Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Send ee, National Institutes of Health , Bethesda, Md. (1991)); and/or those residues from a "hypervariable loop" (e.g., residues 24- 34 (L1), 50-56 (L2) and 89-97 (L3) in the VL, and 26-32 (HI), 52-56 (H2) and 95-101 (H3) in the VH when numbered in accordance with the Chothia numbering system; Chothia and Lesk, J. Mol . Biol. 196:901-917 (1987)); and/or those residues from a "hypervariable loop" VCDR (e.g., residues 27-38 (L1), 56-65 (L2) and 105-120 (L3) in the VL, and 27-38 (HI), 56-65 (H2) and 105-120 (H3) in the V_H when numbered in accordance with the IMGT numbering system; Lefranc, M.P. et al. Nucl. Acids Res. 27:209-212 (1999), Ruiz, M. e al. Nucl. Acids Res. 28:219-221 (2000)). Optionally, the antibody has symmetrical insertions at one or more of the following points 28, 36 (L1), 63, 74-75 (L2) and 123 (L3) in the VL, and 28, 36 (HI), 63, 74-75 (H2) and 123 (H3) in the V_H when numbered in accordance with AHo; Honneger, A. and Plunkthun, A. J. Mol. Biol. 309:657- 670 (2001)).

[00537 ] As used herein, the term ‘'epitope” includes any protein determinant capable of specific binding to an immunoglobulin, an scFv, or a T-cell receptor. The term “epitope” includes any protein determinant capable of specific binding to an immunoglobulin or T- cell receptor. Epitopic determinants usually consist of chemically active surface groupings of molecules such as amino acids or sugar side chains and usually have specific three dimensional structural characteristics, as well as specific charge characteristics. For example, antibodies may be raised against N-terminal or C-terminal peptides of a polypeptide. An antibody is the to specifically bind an antigen when the dissociation constant is < 1 pM; e.g., < 100 nM, preferably < 10 nM and more preferably < 1 nM. [00538 ] As used herein, the terms “immunological binding,” and “immunological binding properties” refer to the non-covalent interactions of the type which occur between an immunoglobulin molecule and an antigen for which the immunoglobulin is specific. The strength, or affinity of immunological binding interactions can be expressed in terms of the dissociation constant (Kd) of the interaction, wherein a smaller Kd represents a greater affinity. Immunological binding properties of selected polypeptides can be quantified using methods well known in the art. One such method entails measuring the rates of antigenbinding site/antigen complex formation and dissociation, wherein those rates depend on the concentrations of the complex partners, the affinity of the interaction, and geometric parameters that equally influence the rate in both directions. Thus, both the “on rate constant” ( K_on) and the “off rate constant” (K_off) can be determined by calculation of the concentrations and the actual rates of association and dissociation. (See Nature 361: 186-87 (1993)). The ratio of K_off /K_on enables the cancellation of all parameters not related to affinity, and is equal to the dissociation constant K_d. (See, generally, Davies et al. (1990) Annual Rev Biochem 59:439-473). An antibody of the present invention is the to specifically bind to its target, when the equilibrium binding constant (Kd) is <1 μM, e.g., < 100 nM, preferably < 10 nM, and more preferably < 1 nM, as measured by assays such as radioligand binding assays or similar assays known to those skilled in the art.

[00539 j The term “isolated polynucleotide” as used herein shall mean a polynucleotide of genomic, cDNA, or synthetic origin or some combination thereof, which by virtue of its origin the “isolated polynucleotide” (1) is not associated with all or a portion of a polynucleotide in wfiich the “isolated polynucleotide” is found in nature, (2) is operably linked to a polynucleotide which it is not linked to in nature, or (3) does not occur in nature as part of a larger sequence. Polynucleotides in accordance with the invention include the nucleic acid molecules encoding the heavy chain immunoglobulin molecules, and nucleic acid molecules encoding tire light chain immunoglobulin molecules described herein. [00540] The term “isolated protein” referred to herein means a protein of cDNA, recombinant RNA, or synthetic origin or some combination thereof, which by virtue of its origin, or source of derivation, the “isolated protein” (1) is not associated with proteins found nature, (2) is free of other proteins from the same source, e.g., free of marine proteins, (3) is expressed by a cell from a different species, or (4) does not occur in nature. [00541] The term “polypeptide” is used herein as a generic term to refer to native protein, fragments, or analogs of a polypeptide sequence. Hence, native protein fragments, and analogs are species of the polypeptide genus. Polypeptides in accordance with the invention comprise the heavy chain immunoglobulin molecules, and the light chain immunoglobulin molecules described herein, as well as antibody molecules formed by combinations comprising the heavy chain immunoglobulin molecules with light chain immunoglobulin molecules, such as kappa light chain immunoglobulin molecules, and vice versa, as well as fragments and analogs thereof.

[00542] The term “naturally-occurring” as used herein as applied to an object refers to the fact that an object can be found in nature. For example, a polypeptide or polynucleotide sequence that is present in an organism (including viruses) that can be isolated from a source in nature and which has not been intentionally modified by man in the laboratory or otherwise is naturally -occurring.

[00543] The term “operably linked” as used herein refers to positions of components so described are in a relationship permitting them to function in their intended manner. A control sequence “operably linked” to a coding sequence is ligated in such a way that expression of the coding sequence is achie ved under conditions compatible with the control sequences.

[00544] The term “control sequence” as used herein refers to polynucleotide sequences which are necessary to affect the expression and processing of coding sequences to which they are ligated. The nature of such control sequences differs depending upon the host organism in prokaryotes, such control sequences generally include promoter, ribosomal binding site, and transcription termination sequence in eukaryotes, generally, such control sequences include promoters and transcription termination sequence. The term “control sequences” is intended to include, at a minimum, all components whose presence is essential for expression and processing, and can also include additional components whose presence is advantageous, for example, leader sequences and fusion partner sequences. The term “polynucleotide” as referred to herein means a polymeric boron of nucleotides of at least 10 bases in length, either ribonucleotides or deoxynucleotides or a modified form of either type of nucleotide. Tire term includes single and double stranded forms of DNA.

[00545] As used herein, the twenty conventional amino acids and their abbreviations follow conventional usage. See Immunology - A Synthesis (2nd Edition, E.S. Golub and D R. Gren, Eds,, Sinauer Associates, Sunderland Mass. (1991)). Stereoisomers (e.g, D- amino acids) of the twenty conventional amino acids, unnatural amino acids such as a-, a- disubstituted amino acids, N-alkyl ammo acids, lactic acid, and other unconventional ammo acids may also be suitable components for polypeptides of the present invention. Examples of unconventional amino acids include: 4 hydroxyproline, γ-carboxyglutamate, ε-N,N,N- trimethyllysine, ε -N -acetyllysine, O-phosphoserine, N- acetylserine, N-formylmethionine, 3-methylhistidine, 5 -hydroxy lysine, σ-N-methylarginine, and other similar amino acids and imino acids (e.g.. 4- hydroxyproline). In tire polypeptide notation used herein, the left-hand direction is the amino terminal direction and the right-hand direction is the carboxy-terminal direction, in accordance with standard usage and convention. [00546] As applied to polypeptides, the term “'substantial identity” means that two peptide sequences, when optimally aligned, such as by the programs GAP or BESTFIT using default gap weights, share at least 80 percent sequence identity, preferably at least 90 percent sequence identity, more preferably at least 95 percent sequence identity, and most preferably at least 99 percent sequence identity.

[00547] Preferably, residue positions which are not identical differ by conservative amino acid substitutions.

[00548] Conservative amino acid substitutions refer to the interchangeability of residues having similar side chains. For example, a group of amino acids having aliphatic side chains is glycine, alanine, valine, leucine, and isoleucine: a group of amino acids having aliphatic- hydroxyl side chains is serine and threonine: a group of amino acids having amide- containing side chains is asparagine and glutamine; a group of amino acids having aromatic side chains is phenylalanine, tyrosine, and tryptophan; a group of amino acids having basic side chains is lysine, arginine, and histidine; and a group of amino acids having sulfur- containing side chains is cysteine and methionine. Preferred conservative amino acids substitution groups are: valine-leucine-isoleucine, phenylalanine-tyrosine, lysme-arginine, alanine valine, glutamic- aspartic, and asparagine-glutamine.

[00549] As discussed herein, minor variations in the amino acid sequences of antibodies or immunoglobulin molecules are contemplated as being encompassed by the present invention, providing that the variations in the ammo acid sequence maintain at least 75%, more preferably at least 80%, 90%, 95%, and most preferably 99%. In particular, conservative amino acid replacements are contemplated. Conservative replacements are those that take place within a family of amino acids that are related in their side chains. Genetically encoded amino acids are generally divided into families: (1) acidic amino acids are aspartate, glutamate; (2) basic amino acids are lysine, arginine, histidine; (3) non-polar amino acids are alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan, and (4) uncharged polar amino acids are glycine, asparagine, glutamine, cysteine, serine, threonine, tyrosine. The hydrophilic amino acids include argmine, asparagine, aspartate, glutamine, glutamate, histidine, lysine, serine, and threonine. The hydrophobic amino acids include alanine, cysteine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, tyrosine and valine. Other families of amino acids include (i) serine and threonine, which are the aliphatic-hydroxy family; (ii) asparagine and glutamine, which are the amide containing family; (hi) alanine, valine, leucine and isoleucine, which are the aliphatic family; and (iv) phenylalanine, tryptophan, and tyrosine, which are the aromatic family. For example, it is reasonable to expect that an isolated replacement of a leucine with an isoleucine or valine, an aspartate with a glutamate, a threonine with a serine, or a similar replacement of an amino acid wuth a structurally related amino acid will not have a major effect on the binding or properties of the resulting molecule, especially if the replacement does not involve an amino acid within a framework site. Whether an amino acid change results in a functional peptide can readily be determined by assaying the specific activity of the polypeptide derivative. Assays are described in detail herein. Fragments or analogs of antibodies or immunoglobulin molecules can be readily prepared by those of ordinary skill in the art. Preferred amino- and carboxy-termini of fragments or analogs occur near boundaries of functional domains. Structural and functional domains can be identified by comparison of the nucleotide and/or amino acid sequence data to public or proprietary sequence databases. Preferably, computerized comparison methods are used to identify sequence motifs or predicted protein conformation domains that occur in other proteins of known structure and/or function. Methods to identify protein sequences that fold into a known three-dimensional structure are known. Bowie et al. Science 253: 164 (1991). Thus, the foregoing examples demonstrate that those of skill m the art can recognize sequence motifs and structural conformations that may be used to define structural and functional domains in accordance with the invention.

[ 00550 ] Preferred ammo acid substitutions are those which: (1) reduce susceptibility to proteolysis, (2) reduce susceptibility to oxidation, (3) alter binding affinity for forming protein complexes, (4) alter binding affinities, and (4) confer or modify other physicochemical or functional properties of such analogs. Analogs can include various muteins of a sequence other than the naturally -occurring peptide sequence. For example, single or multiple amino acid substitutions (preferably conservative amino acid substitutions) may be made in the naturally- occurring sequence (preferably in the portion of the polypeptide outside the domain(s) forming intermolecular contacts. A conservative amino acid substi tution should not substantially change the structural characteristics of the parent sequence (e.g., a replacement amino acid should not tend to break a helix that occurs in the parent sequence, or disrupt other types of secondary structure that characterizes the parent sequence). Examples of art-recognized polypeptide secondary and tertiary structures are described in Proteins, Structures and Molecular Principles (Creighton, Ed., W. H. Freeman and Company, New York (1984)); Introduction to Protein Structure (C. Branden and J. Tooze, eds., Garland Publishing, New York, NY . (1991)); and Thornton et at. Nature 354: 105 (1991).

[00551] As used herein, the terms " label” or “labeled” refers to incorporation of a detectable marker, e.g., by incorporation of a radiolabeled amino acid or atachment to a polypeptide of biotinyl moieties that can be detected by marked avidin (e.g., streptavidin containing a fluorescent marker or enzymatic activity that can be detected by optical or calorimetric methods). In certain situations, the label or marker can also be therapeutic. Various methods of labeling polypeptides and glycoproteins are known in the art and may be used. Examples of labels for polypeptides include, but are not limited to, the following: radioisotopes or radionuclides (e.g., ³H, ¹⁴C, ¹⁵N, ³⁵S, ⁹⁰Y, ⁹ ₉Tc, ^{1 11}In, ¹²⁵I, ¹³¹I), fluorescent labels (e.g., FITC, rhodamine, lanthanide phosphors), enzymatic labels (e.g., horseradish peroxidase, p-galactosidase, luciferase, alkaline phosphatase), chemiluminescent, biotinyl groups, predetermined polypeptide epitopes recognized by a secondary reporter (e.g., leucine zipper pair sequences, binding sites for secondary antibodies, metal binding domains, epitope tags). In some embodiments, labels are attached by spacer arms of various lengths to reduce potential steric hindrance. The term “pharmaceutical agent or drug” as used herein refers to a chemical compound or composition capable of inducing a desired therapeutic effect when properly administered to a patient.

[00552] Other chemistry terms herein are used according to conventional usage in the art, as exemplified by The McGraw-Hill Dictionary of Chemical Terms (Parker, S., Ed., McGraw- Hill, San Francisco (1985)).

[00553] As used herein, “substantially pure” means an object species is the predominant species present (i.e., on a molar basis it is more abundant than any other individual species in the composition), and preferably a substantially purified fraction is a composition wherein the object species comprises at least about 50 percent (on a molar basis) of all macromolecular species present.

[00554] Generally, a substantially pure composition will comprise more than about 80 percent of all macromolecular species present in the composition, more preferably more than about 85%, 90%, 95%, and 99%. Most preferably, the object species is purified to essential homogeneity (contaminant species cannot be detected in the composition by conventional detection methods) wherein the composition consists essentially of a single macromolecular species.

[00555] The term patient includes human and veterinary subjects.

[00556] EXAMPLES

[00557] EXAMPLE 1: Design and Expression of Bispecific Antibodies with Heavy Chain and Light Chain Heterodimerization

[00558] A schematic of the anti-CD3/anti-CD20 targeted bispecific antibodies of the present disclosure is represented in FIG. IE and is composed of the following: (1) a first antigen binding region that binds CD 3 comprising SP34 antibody variants (Table 7) (“anti- CD3” or “anti-CD3 arm”), (2) a second antigen binding region that binds CD20 derived from ofatumumab or obinutuzumab (“anti-CD20” or “anti-CD20 arm”), (3) knob-in-hole mutations in the heavy chain as described in US 63/368,852, (4) mutations corresponding to set D that enforce light chain heterodimerization, (5) a protein linker, and (6) full length CD58 peptide fused to the C -terminal of the heavy chain of the CD3 binding arm.

[00559] Challenges for bispecific IgG-like antibody format designs are (i) efficient heavy chain heterodimerization and, (ii) efficient light chain heterodimerization for correct cognate pairing with good specificity. Thus, bispecific or multispecific antibodies were designed to include (i) knob-into-hole mutations to promote efficient heavy chain heterodimerization and (ii) charged pairing (between VH1 and VL1 interface and/or VH2 and VL2 interface, and CH1 H1 and CL1 interface and/or CH2 H1 and CL2 interface) and disulfide stabilization mutations (between CHI H1 and CL1 interface and/or CH2 H1 and CL2 interface), which together promote correct cognate pairing of heavy chain and light chains. Correct pairing of heavy and light chains is advantageous for efficient large scale antibody production and purification.

[00560] Bioluminate modeling suite and Pymol molecular graphics system from Schrodinger were used to perform all molecular modeling works. A high resolution crystal structure of Trastuzumab (PDB ID: 1N8Z) was used as a backbone template for homology modeling of VH2-CH2 JTl/VL2-CL2kappa antigen binding fragment (Fab) of the disease- associated antigen (DAA) binding portion of the antibody, and a high resolution crystal structure of Pertuzumab (PDB ID: 4LLU) was used as a backbone template for homology modeling of VH1 -CH 1 III /VL1 -CL 1 lambda (Fab) of the CD3 binding portion of the antibody. [00561] The evaluation of impact of individual mutation sets for charged pairs or knob into hole mutations on the Fab stability and on the respective affinity of each paratope was carried out with the BioLuminate Residue Scanning Tool. Residue pairs for disulfide repositioning were identified using PyMol molecular graphics system through structure guided design and validated by homology modeling or Bioluminate Cys-Cys engineering tool. Exemplary' charged pairs and disulfide repositioning mutations are shown in Tables 1- 6.

[00562] Exemplary bispecific antibodies of the disclosure are shown in FIGS. 1A-1E. Specifically, FIG. IE shows a bispecific antibody with a first antigen binding region that binds CD3 and a second antigen binding region that binds CD20 and a CD58 peptide fused the C-terminal of the first heavy chain. Exemplary antibodies, EIP0866, EIP0886, EIP0893, EIP0929 and EIP0892 comprise the mutations shown in Table 17. Further, each antibody comprises light chain pairing mutation set D shown in Table 18. This structural design of the antibody provides improved specificity and complete cognate pairing, w'hich is advantageous for functional antibody expression and purification

[00563] Table 17. Nomenclature and mutations associated with CD3 variants

[00564] Table 18. Light chain pairing mutation set D [00565] Antibody Expression and Purification

[00566] DNA sequences corresponding to antibody heavy and light chains were synthesized in pDT5 vector (ATUM, Newark CA). Plasmid DNA (1 μg/ml) were transfected into Expi293 cells (ThermoFisher) according to manufacturer’s protocols. Cells were grown in flasks with rotation (125 rpm) at 37°C with 8% CO2. Five days post-transfection, the conditioned media was harvested from the cells by centrifugation (3000 x g) for 30 minutes and filtered using 0.45 pm filter. Antibodies were then purified using an AKTA Avant chromatography system (Cytiva) and a tandem purification method using HiTrap Mabselect Protein A and HiLoad Superdex 16/600 200 pg columns (Cytiva). Antibodies were stored in PBS, pH 7.2 at 4°C following purification and prior to analysis.

[00567] Biophysical characterization of the anti-CD3 arm of the anti-CD3/anti-CD20 bispecific antibodies is as described in PCT application No. PCT/US2023/064728 and is incorporated by reference herein.

[00568] Anti-CD3/anti-CD20 targeted bispecifics and bispecific fusions were produced in transient Expi293 system. The preparative size exclusion chromatogram of exemplary bispecific antibodies (EIP0866, EIP0886, EIP0892, EIP0893, EIP0929) of the present dislcosure following protein A purification is shown in FIG. 2.

[00569] Surface Plasmon Resonance (SPR) Biacore 8K kinetic binding analysis [00570] Exemplary bispecific antibodies were advanced for further analysis via surface plasma resonance (SPR) for binding to recombinant CD3e/d heterodimer. The kinetic constants (association, dissociation and equilibrium) are shown in Table 19.

[00571] Analysis was performed on Biacore 8K+ system (Cytiva). Biotinylated human

CD3 -epsilon and CDS -delta heterodimer (Aero Biosystems) was captured on the streptavidin sensor chip (Cytiva) with the RU values ranging from 50-70. Protein samples at different concentrations diluted in the running buffer (10 mM HEPES, 150 mM NaCl, 0.05% v/v Surfactant P20, pH 7.4 ) were then injected to each channel at a flow' rate of 30 pL/min for the multi-cycle kinetics/affinity analysis. The binding kinetics were carried out at 25 °C and the association contact and dissociation times used were 60-120 seconds and 120-420 seconds. The chip surface was regenerated by injecting 10 mM glycine, pH 1.5, at a flow rate of 30 μL/min for 60 sec. The kinetic and affinity constants are determined by fiting the data to a 1 : 1 binding model using the Biacore Insight Evaluation Software (Cytiva).

100572] Table 19: Association, dissociation, and equilibrium constants for CD3 binding of CD20-targeted bispecific fusions

[ 00573] FIGS. 3A- 3E are a series of graphs depicting surface plasma resonance (SPR) binding sensorgrams of the captured recombinant CD3e/d interacting with light chain pairing bispecific antibodies (EIP0866, EIP0886, EIP0892, EIP0893, EIP0929) on Biacore 8K. The sensograms are recorded from single cycle kinetics with varying concentrations of bispecific antibodies and the data are processed on Biacore evaluation program using 1 : 1 kinetic model fit. FIG. 3A shows EIP0866. FIG. 3B shows EIP0886. FIG. 3C shows EIP0892. FIG. 3D shows EIP0893. FIG. 3E shows EIP0929.

[00574] Differential Scanning Calorimetry

[00575] The thermal stability of CD20 targeted bispecific fissions were assessed using differential scanning calorimetry (DSC).

[00576] Protein samples were prepared m PBS at 1 mg/mL concentration, loaded into sample cell and scanned at l°C/min increment from 25°C to 95°C. Data was analyzed using NanoAnalyzer program subtracting PBS buffer background from each individual scan. Differential scanning calorimetry analysis of exemplary bispecific antibodies is shown in FIG. 4. As shown m FIG. 4, all proteins show first major melting transition between 67- 70 degrees Celsius. A second melting transition corresponding to the Fab region can be observed as either shoulder or small peak following first transition.

[00577] Mass Spectrometry

[00578] Mass spectrometry was used to assess the degree of mispairing of peptide chain s for exemplary bispecific antibodies, as shown in FIGS. 5A-5B. [00579] The intact mass of the purified fusion proteins and antibody chains was confirmed by Xevo G2-XS QTof Quadrupole Time-of-Flight Mass Spectrometry coupled to a Acquity UHPLC system (Waters) equipped with a Protein BEH C4 (300 A 1.7 pm) column, as depicted FIGS. 5A-5B. Antibody samples were deglycosylated first using rapid PNGase F enzyme in reducing and non-reducing conditions following supplier’s protocols. Reaction mixture was diluted to 1: 10 in 50% Acetonitrile containing 0.1% Formic acid and 2 pL of which was injected into LC-MS. Tie total ion chromatogram and m/z data of the proteins were acquired with gradient ran of 10 to 70 mL HPLC grade acetonitrile over 12 min. Mass of the protein samples was deconvoluted from to tal ion chromatogram using BYOS software from Protein Metrics. FIG. 5A shows intact mass of EIP0866 determined after PNGAse F deglycosylation in non-reduced condition and chromatographic separation using reverse phase C18 column. FIG. 5B shows reduced mass analysis of EIP0866 heavy' chains and light chains determined after Rapid PNGAse F deglycosylation in reduced condition and chromatographic separation using reverse phase C18 column.

[00580] The expected mass for a correctly- paired bispecific fusion was observed for EIP0866 by non-reduced intact antibody mass analysis (FIG. 5A). Four distinct polypeptide chains corresponding to two light chains and two heavy chains of the expected mass were also observed upon reduction (FIG. SB).

[00581] EXAMPLE 2: Characterization of aCD20-aCD3 Bispecific Antibodies With Mutations That Promote Heavy Chain And Light Chain Heterodimerization

[00582] Using tire methods described above, aCD20-aCD3 bispecific antibodies (e.g. EIP0866 and EIP0865) were constructed and tested for efficacy in T-cell mediated cytotoxicity.

[00583] Isolation and Activation of T-cells for Cytotoxicity Assays

[00584] T cells were isolated from healthy human PBMCs donors using StemCell T-cell isolation kit and resuspended in RPM1 was supplemented with 10% FBS, IX penicillin/streptomycin. To activate T cells, Dynabeads coated with aCD3 and aCD28 antibodies were added to the T cells at a ratio of 25 uL beads per million cells and incubated at 37°C in 5% CO2 for 48 hours. Following removal of Dynabeads, T cells were incubated with IL-7 (10 ng/ml) and IL- 15 (10 ng/ml) in supplemented in media for an additional 7 days. The supplemented media was replaced with freshly thawed cytokines (IL7 and IL15) every' 48 hours. Human tumor cells were seeded at 10,000 cells per well in 96- well tissue culture plates (Perkin Elmer) and incubated for 24 hours at 37 °C with 5 % CO2. The following day, naive or activated human T cells (50-100,000) or PBMCs (250-300,000) were added to tumor cells in the presence or absence of antibodies and incubated for up to 3-7 days. Cytolysis of tumor cells were measured by using the LDH-Glo cytotoxicity assay (Promega) or cytolysis of Green Fluorescent Protein (GFP) engineered tumor cells were visualized using fluorescent plate reader (Ensight, Perkin Elmer).

[00585] Human tumor cells engineered to express Green Fluorescent Protein (GFP) (10,000 cells) were incubated activated human T cells (50,000) or human PBMCs (150,000) in the presence of antibodies and incubated for 72- 96 hours at 37C with 5 % CO2. Cytolysis of GFP positive tumor cells was determined by flow cytometric analysis (FACSymphony, BD Biosciences). Tumor cytotoxicity curves were generated by plotting tumor cell counts versus concentration using Prism software (GraphPad).

[00586] T-cell repeat activation assay

[00587] Bispecific and bispecific fusion proteins were immobilized on magnetic Dynabeads including CD3 and CD28 antibodies as a control. In each round, T cells were activated with protein/Dynabeads for 48 h. Subsequently beads were removed, and the T cells are counted by flow cytometry and subjected to the next round of activation.

[00588] The bispecific variants with complete cognate pairing were then tested for their functional cytolytic activity using co-culture cytotoxicity assays.

[00589] A surrogate of Mosunetuzumab, a clinically approved CD20 bispecific antibody, was generated by following cross-mab technology described by Schaefer W et. al. (Schaefer W . PNAS, 2011, 108(27), 1 1 187-11 192), which allowed for the purification of the bi specific antibody using a single cell line while avoiding the oxidative/reduction step. The cytolytic potency of exemplary anti-CD3/anti-CD20 bispecific and bispecific fusions were compared to the Mosunetuzumab surrogate in a human PMBC co-culture assay w'ith JeKo- 1-GFP tumor cells. The use of Ofatumumab as a the CD20 targeting antibody in the bispecific (EIP0865) resulted in - 1000-fold improvement in activity compared to the Mosunetuzumab surrogate (FIG. 6). The fusion of CD58 to both EIP0865 and Mosunetuzumab surrogate improved activity 2-3 fold as seen with EIP0866 (CD20 x CD3- 01 x CD58) and Mosunetuzumab surrogate x CD58. EIP0607 (Control x CD3-A6 x CD58) which targets an irrelevant antigen did not show any activity in this assay. As shown in FIG. 7, bispecific fusions with attenuated CD3 affinities: EIP0866 (CD20 x CD3-A2 x CD58), EIP0893 (CD20 x CD3-A5 x CD58), EIP0929 (CD20 x CD3-A6 x CD58) and EIP0892 (CD20 x CD3-A8 x CD58) were all observed to have similar potency as EIP0866 (CD2.0 x CD3-01 x CD58) and were all vastly superior to Mosunetuzumab.

[00590] Tire cytolytic potency of anti-CD3/anti-CD20 targeted bispecific fusions compared to Glofitamab, a CD20 bispecific under clinical evaluation in a human PMBC co-culture assays, was evaluated with either Raji-GFP and NALM6-GFP tumor cells, as depicted in FIG. 8 and FIG. 9, respectively. ETP0866 (CD20 x CD3-01 x CD58) showed equivalent cell killing to the Glofitamab in Raji-GFP cells while the activity of EIP0929 (CD20 x CD3-A6 x CD58) was slightly diminished (FIG. 8). NALM6 tumor cells are known to express lower CD20 levels than Raji cells. Here, while Glofitamab was potent at single digit picomolar concentrations, the maximal tumor cell killing achieved was -50% at several of the highest concentrations tested (FIG. 9). Both EIP0866 and EIP0929 were active at slightly higher concentrations than the Glofitamab but importantly were able to achieve maximal tumor cell killing of 80% at the highest concentrations tested.

[00591] The advantage of the providing costimulation and CD3 binding simultaneously versus CD3 alone for CD20 targeted antibodies was clearly demonstrated using T cell repeat activation assay. The CD20 bispecific fusions EIP0866 (CD20 x CD3-01 x CD58) and EIP0929 (CD20 x CD3-A6 x CD58), and Glofitamab were immobilized on Dynabeads and incubated with naive human T cells for successive rounds of 48 hour activation. As shown in FIG. 10A, both EIP0866 and Glofitamab failed to stimulate T cells to proliferate beyond the initial cell count while the EIP0929 stimulated T cells to proliferate to a much greater extent exceeding that of the CD3/CD28 antibody Dynabead control. This suggests that in addition to the presence of co-stimulation, the strength of tire CD3 binding is an important consideration. Functionally, T cells activated EIP0866 and EIP0929 were equally potent in cytotoxicity assay with Raji tumor cells in contrast to T cells activated with Glofitamab, as shown in FIG. 10B. In addition, as shown in FIG. 10C, the bispecific fusions continue to induce CD8 T cell expansion during the cytotoxicity assay whereas Glofitamab does not. [00592] EXAMPLE 3 - Production and biophysical characterization of aCD20-aCD3 Bi specific Antibodies with Fusion Peptides

[00593] Bispecific fusion was generated by fusing the full-length extracellular domain of CD58 (UNIPROTKB: Pl 9256, ammo acids: 29-216), CD58v (ammo acids: 29-122), IL7 (UNIPROTKB: P13232, amino acids: 26-177) using glycine serine linker to the carboxylterminus of the hole heavy chain of the CD3 specific heavy chain, as shown in FIG. 1. Bispecific antibodies were fused with CD58 at the carboxyl-terminus of the hole heavy chain to create a series of bispecific CD58 fusions. Listed in Table 17 are anti-CD20/anti- CD3 bispecific CD58 fusions and their cognate bispecific antibodies.

[00594] EXAMPLE 4: In Vivo Characterization of Bispecific Antibodies with fusion peptides

[00595] Activation and Expansion of T cells for in vivo studies

[00596] T-cells were isolated from healthy PBMCs donors using StemCell T-cell isolation kit. OpTmizer media was supplemented with 2% human AB serum, IX penicillin/ streptomycin, 4 mM glutaMAX, 2 mM glutamine. Cells were incubated at 37°C in 5% CO2 in the supplemented OpTmizer media at 5X10⁵ cells/mL and T cells were treated with anti-CD3 and anti-CD28 antibody coated Dynabeads at a ratio of 25 uL Dynabeads per million cells for 48 hours. Following removal of Dynabeads, T cells were incubated with IL-7 (10 ng/ml) and IL-15 (10 ng/ml) in supplemented OpTmizer media for an additional 6 days. The supplemented media was replaced with freshly thawed cytokines (IL7 and IL 15) every 48 hours. On the day of adoptive transfer or Co-engraftment, the T- cells are harvested and a hemocytometer was used to determine cell number and viability. T cells were suspended in cold PBS and stored on ice for no longer than 30 minutes before injecting into animals.

[00597] Tumor studies with co-engraftment of human T cells

[00598] Tumor growth inhibition of exemplary anti-CD3/anti-CD20 bispecific antibodies was examined using a Burkit’s lymphoma cell line, Raji, co-engrafted with activated human T cells and a xenograft NSG mouse model.

[00599] Ten million Raji-GFP-luc cells and 2 million activated T cells were implanted subcutaneously into flank of NSG mice (NOD.Cg-Prkdcscid 112rgtmlWjl/SzJ, Jackson Laboratory). Tire CD20 benchmark bispecific and CD20 bispecific fusions were dosed weekly starting at day 0 for 3 weeks. Tumor growth was monitored using electronic calipers and volume were calculated according to the formula: π/6 X (length x width²).

[ 00600 ] At day 0, 1x10⁶ activated T cell and 10x10⁶ Raji-GFP-luc tumor cells were implanted subcutaneously. Intravenous dosing of EIP0614 (0.5 mg/kg), Glofitamab- biosimilar (0.15 mg/kg and 0.5 mg/kg), EIP0866 (0.15 mg/kg and 0.5 mg/kg), and EIP0929 (or 0.15 mg/kg and 0.5 mg/kg) was conducted as indicated above, out to day 20. Tumor volume following each treatment was analyzed as shown in FIG. 11 . As shown in FIG. 11, treatment with EIP0929 at both doses, and EIP0866 at 0.5 mg/kg resulted in reduction of tumor volume at day 7 and maintained out to day 20 compared to treatment with EIP0614. EIP0929 (CD20 x CD3-A6 x CD58) showed equivalent potency as Glofitamab-biosimilar at both dose levels tested. While EIP0866 (CD20 x CD3-01 x CD58) also displayed similar potency to Glofitamab-biosimilar at 0.5 mg/kg dose level, a marked reduction in potency at the 0.15 mg/kg dose level was observed. EIP0614 (Control x CD3-A5 x CD58) which targets an irrelevant antigen was used as a control in this study.

[00601 ] These data shown that treatment with EIP0887 and EIP0929 at the indicated doses are more effective at reducing tumor volume in vivo compared to EIP0614 and EIP0866 (0.15mg/kg).

[00602] EXAMPLE 5: Characterization of exemplary anti-CD3 and anti-CD20 Bispecific Antibodies with CD58 Fusion Peptides

[00603] Thermal stability assessment

[00604] Thermal stability is an important biophysical attribute for biological therapeutics. Poor stability may affect the solubility of the biologic and can lead to the aggregation. The thermal stability of exemplary antibodies with respect to Tonset and TmS were determined by differential scanning calorimetry (DSC) using nanoDSC from TA Instrument. Samples were prepared in IxPBS at a concentration of 1 mg/rnL and each sample was run in duplicate and scanned at 1 °C/min from 25 to 95 °C. A buffer background was subtracted from each individual scan leaving the partial molar heat capacity, which was normalized using the moles of the protein in the active cell volume. Data was analyzed from each individual scan using NanoAnalyzer program provided by the supplier.

[00605] Thermal denaturation and exact melting temperatures (Tm and Tonset) of the biologies were determined by differential scanning calorimetry (DSC) as shown in FIG. 12. EIP0929 shows a major unfolding at Tm 69.9x0.3 °C, a second unfolding at 77.5x0.3 °C with a T_onset at 57±0.5 °C, whereas Glofitamab biosimilar shows first unfolding at 62.9±0.3 °C and a second unfolding at 78.9±0.3 °C. Thus, ETP0929 shows a superior biophysical stability profile compared to clinically approved benchmark Glofitamab.

[00606] A thermal forced aggregation assay was also used to determine the stability of the antibodies. The fusion protein and antibody samples in 2, mg/mL concentration were subjected to thermal denaturation for 2 h at varying degrees of temperatures, from 35 °C to 90 °C at 5 °C interval. Following cooling down the samples to 4 °C, the denatured aggregated protein fractions were removed from the samples by centrifugation at 10,000 xg for 5 min. The cleared samples were analyzed by analytical size exclusion chromatography (aSEC) to determine the remaining intact protein fractions from integrated peak area of the chromatograms. Results show' that EIP0929 and EIP0970 have a similar stability to a clinical grade antibodies (e.g. trastuzumab) and about 7 to 10 °C beter stability profile compared to Glofitamab biosimilar (FIGS. 13A and 13B).

[00607] In vitro cytotoxicity assays and in vivo efficacy study

[00608] Tire cytolytic potency of EIP0929, controls and clinical benchmark surrogates were evaluated m a tumor cells and PBMCs co-culture assay settings using high CD20 expressing Burkitt lymphoma cell line (Raji), diffuse large B cell lymphoma (DLBCL) cell line (Toledo), and a low CD20 expressing acute lymphocytic leukemia (ALL) cell line (NALM-6). JeKo-1, Raji, and NALM-6 cells w'ere engineered to overexpress GFP in-house. [00609] The efficacy of the bispecific antibodies to mediate tumor cell lysis was evaluated in PBMC -tumor cell co-culture assays. PBMC from healthy human donors were added at 120,000 per well in 96-well cell -repellent plates (Greiner Bio-One), and CellTrace Violet (Invitrogen) - labeled or GFP -expressing human tumor cells were seeded at 8,000 cells per well, in the presence or absence of serial dilution of antibodies and incubated at 37 °C. Research grade Epcoritamab surrogate was purchased from ProSci Incorporated. Research grade Mosunetuzumab surrogate was purchased from Ichorbio or generated in-house. Research grade Odronextamab was purchased from Ichorbio. After 3 days, the plate was centrifugated and the supernatant was collected and frozen for further cytokine analysis. The cell pellets were stained with Zombie NIR™ Fixable Viability Kit (BioLegend), followed by Fc receptor blocking and surface marker staining. Flow antibodies again surface cell markers (CD3, CD4, CD8, CD 19) were purchased from BioLegend and BI) Bioscience, and stained cells were analyzed by flow cytometry. Numbers of tumor cells, CD4 T cells, CD8 T cells, and B cells were analyzed in FlowJo software. Cytolysis of tumor cells, depletion of B cells, CD4 T cells and CD8 T cells proliferation curves were generated by either plotting the cell counts or normalized cell counts versus bispecific antibodies concentration using Prism software (GraphPad). Four-parameter non-linear regression analysis was used to obtain EC50s using Prism software (GraphPad).

[00610] Raji-GFP cells were cocultured with PBMCs in a 15: 1 E:T ratio and treated with a serial dilution of biologies and evaluated for tumor cell killing, B cell depletion, T cell counts and induction of cytokine levels in the assay.

[00611 ] Table 5,1 shows a comparison of ECso values of tumor cell killing, B cell depletion, CD4+ and CD8 T cell proliferation, and cytokine induction from Raji cells and PBMCs coculture assay. Table 5.2 shows a comparison of ECso values of tumor cell killing, B cell depletion, CD4+ and CD8 T cell proliferation, and cytokine induction from Toledo cells and PBMCs coculture assay.

[00612] Table 5.1

*estimated from the curve

[00613] Table 5.2

* estimated from the curve [00614] As shown in Figures 14A-14B, 15A-15D and Table 5.1, EIP0929 demonstrated -10-30 fold higher potency than Mosunetuzumab and Epcoritamab (clinically approved benchmarks against B cell lymphoma).

[00615] EIP0866, which has a strong CD 3 affinity binding arm, showed similar potency of Glofitamab, while EIP0929, which has a moderate CD3 affinity binding arm, showed a potency of within about 5-10 fold of Glofitamab, in tumor killing and B cell depletion. E1P0929 induced about 5-10 fold more CD8+/CD4+ T cell expansion potency over Glofitamab and other benchmarks. Further, treatment with EIP0929 resulted in IFNy and IL-2 induction levels that were much lower than those from Glofitamab. A similar potency profile, cytokine induction profile and CD8+/CD4+ proliferation of EIP0929 were observed from a coculture assay with an intermediate CD20 density cell line Toledo (FIGS. I6A and 16B, 17A-D, and Table 5.2),

[00616] But, in a low CD20 expressing cell line (NALM-6) tumor killing assay, EIP0929 resulted in a higher level of killing of about 80% than Glofitamab, which only showed a level of killing of about 60% (FIGS. 18B and 18C, and FIG. 9). This is also reflected in in vivo tumor regression study with NALM-6 tumor cells and activated human T cells co- engraftment study in NSG mice (FIG. 18D). Treatment with EIP0929 at 2 mg/kg resulted in reduction of tumor volume at day 7, which was maintained out to day 22 compared to treatment with EIP0614. EIP0929 also showed a stronger tumor regression response compared to Glofitamab-biosimilar at the same dose level. Together, this demonstrates that anti-CD3 and anti-CD20 bispecific antibodies with CD58 fusion peptides as potent therapeutics which have advantageous tumor killing properties relative to clinically approved biosimilars.

[00617] Mouse pharmacokinetics

[00618] A single dose pharmacokinetics study was earned out using a single intravenous dose of 0.5 mg/kg or 4 mg/kg in NSG mice (n=3). A series of blood samples were collected from die retro-orbital sinus (RO) of each mouse at 10 min, 2 h, 6 h, 24 h, 48 h, 72 h, 7 day, 10 day, and 14 day. The blood samples were centrifuged at 2500 rpm for 10 min at 4 °C and the resulting plasma were stored at -80 °C until analysis. The plasma concentrations of EIP0929 were determined by a custom sandwich EL1SA using CD20 for capture and antihuman IgG for detection. The data analysis was carried out tn GraphPad Prism using two- compartmental m odel system and the half -life of the biologic was determined from the linear log of elimination phase. From the serum concentration-time profile in both dose groups exhibited a favorable PK profile of half-life (T1/2) of about 10 days (FIGS. 19A- 19B), which is within the expected range of a therapeutic antibody. Together this demonstrates the safety and potency of anti-CD3 and anti-CD20 bispecific antibodies with CD58 fusion peptides and their application as a therapeutic antibody.

[00619] Tumor studies with co-engraftment of human T cells

[00620] Tumor growth inhibition of exemplary' anti-CD3/anti-CD20 bispecific antibodies was examined using a Burkit’s lymphoma cell line (Raji), and a Acute lymphoblastic leukemia (ALL) cell line (NALM-6), co-engrafted with activated human T cells in a xenograft NSG mouse model.

[00621] Ten million Raji-GFP-luc or NALM-6-GFP-luc cells and 2 million activated T cells were implanted subcutaneously into flank of NSG mice. The CD20 benchmark bispecific and CD20 bispecific fusions were dosed weekly starting at day 0 for 3 weeks. Tumor growth was monitored using electronic calipers and volume were calculated according to the formula: rc/6 x (length x width²).

[00622] At day 0, 2x 10⁶ activated T cell and 10x10⁶ Raji-GFP-luc tumor cells were implanted subcutaneously. Intravenous dosing of EIP0614 (0.5 mg/kg), Glofitamab- biosimilar (0.15 mg/kg and 0.5 mg/kg), EIP0866 (0.15 mg/kg and 0.5 mg/kg), and EIP0929 (or 0.15 mg/kg and 0.5 mg/kg) was conducted as indicated above, out to day 48. Tumor volume following each treatment w^ras analyzed as shown in FIG. 11 . As shown in FIG. 11, treatment with EIP0929 at both doses, and EIP0866 at 0.5 mg/kg resulted in reduction of tumor volume at day 7 and maintained out to day 48 compared to treatment with E1P0614. EIP0929 (CD20 x CD3-A6 x CD58) showed equivalent potency as Glofitamab-biosimilar at both dose levels tested. While EIP0866 (CD20 x CD3-01 x CD58) also displayed similar potency to Glofitamab at 0.5 mg/kg dose level, a marked reduction in potency at the 0.15 mg/kg dose level was observed. EIP0614 (Control x CD3-A5 x CD58) which targets an irrelevant antigen was used a control in this study.

[00623] At day 0, 2x10⁶ activated T cell and 10x10⁶ NALM-6-GFP-luc tumor cells were implanted subcutaneously. Intravenous dosing of E1P0614 (10 mg/kg), Glofitamab- biosimilar (2 mg/kg), and EIP0929 (2 mg/kg) was conducted as indicated above, out to day 21 . Tumor volume following each treatment w^ras analyzed by caliper measurement as shown in FIGS. 19A-19B. The treatment with EIP0929 at 2 mg/kg resulted in reduction of tumor volume at day 7 and maintained up to day 22 compared to treatment with EIP0614. In this low antigen density tumor model EIP0929 showed stronger tumor regression response compared to Glofitamab-biosimilar at the dose level tested. These data show that treatment with EIP0887 and EIP0929 at the indicated doses are more effective at reducing tumor volume in vivo compared to EIP0614 and/or EIP0866.

100624 ] Multidose Pharmacodynamic Non-GLP toxicology study of EIP0929 in Cynomolgus Monkey

[00625] Table 5.3

[00626] Pharmacokinetics sample collection and bioanalysis

[00627] Blood samples (approximately 0.5 mL) were collected via a femoral vein on Days 1 and 8, 15, and 22 of the dosing phase. Samples were collected predose (Day I only), 58 minutes after the start of infusion, and approximately 0.5, 2, 6, 24, 48, 72 and 168 hours postdose. Blood samples were collected into K2 EDTA-tubes and centrifuged (2500 rpm for 10 minutes at 4°C). Resulting plasma was recovered, divided into two aliquots (sets 1 and 2), and stored frozen (< -60°C) before analysis.

[00628] Serial dilutions of plasma samples from each time point (from treatment and vehicle group) were prepared in blocking buffer (IxPBS, 0.02% polysorbate 80) and analyzed alongside purified EIP0929 protein standards. Four-fold serial dilutions of the standard were made in blocking buffer plus 2-5% normal cynomolgus monkey serum in duplicate to generate a standard curve. All samples were evaluated with a custom sandwich EL1SA using recombinant CD20 capture and anti-human IgG-HRP detection and chemiluminescent peroxidase substrate. Luminescence signals were measured on an EnSight (PerkinElmer) plate reader using default luminescence detection settings. [00629] Plasma concentrations of the antibody (for each time point and dose groups) were determined against a standard curve (4PL) generated from purified EIP0929 using GraphPad Prism. The pharmacokinetics profile for each dose group up to 168 h were determined in GraphPad Prism using two-compartmental model analysis and the half -life of the antibody was determined from the linear log of elimination phase.

[00630] Peripheral Blood Immunophenotyping

[00631] Blood (approximately 1 mL) was collected for immunophenotyping (1PT) with K2 EDTA as anticoagulant from non-fasted animals, unless fasted for other procedures, via a femoral vein on Day 1 (Predose and 6 hours post the end of infusion) and once prior to dosing on Days 2, 3, 8, 9, 15, 16, 22, 23, and 29. A heterogeneous lymphocyte gating strategy of CD45 fluorescent staining and side scatter (SSC) demarcation (CD45^bright

SSC^dim) was used to delineate lymphocyte populations by flow cytometry . [00632 ] Table 5.4 Peripheral Blood Immunophenotyping Tests

[00633 ] Peripheral Blood Immunophenotyping

[00634] Cytokine Analysis

[00635] Blood (approximately 0.5 mL) was collected for cytokine analysis from non-fasted animals, unless fasted for other procedures, via the femoral vein once during the predose phase and on Days 1, 2, 8, 9, 15, 16, 22, 23, and 29 of the dosing phase. Blood was collected predose and 2 and 6 hours post the end of infusion on Days 1, 8, 15, and 22 and once on Days 2, 9, 16, 23, and 29. Blood was collected into serum separator tabes (without anticoagulant), allowed to clot at room temperature for at least 30 minutes prior to centrifugation, and centrifuged within 1 hour of collection. Serum was harvested and split into two, approximately equal aliquots and stored on dry ice until placed in a freezer, set to maintain -60 to -80°C, until analyzed. Cytokine determination (IFN-y, TNF-a, IL-2, and IL- 6) was performed using Luminex. [00636] Complete or near complete depletion of B cells in the periphery' was observed

6 hours post first dose of EIP0929 in all treated animals compared to controls (FIG. 20A). B cells in die periphery remained depleted throughout the remainder of the study for most animals. Starting on Day 15, B cells remained below' 35% for all treated animals. Sustained B cells depletion at 150 pg/kg is wdthin 5 -fold of the reported sustained B cell depletion for Glofitamab in NHP (30 pg/kg) (Frances N et al,, Journal of Pharmaceutical Sciences 1 1 1 (2022) 1208-1218).

[00637] EIP0929, was significantly more potent than Epcoritamab doses reported for sustained B cell depletion at 1 mg/kg (Engelberts PJ et al., EBioMedicine 52 (2020) 102625).

[00638] Expected T cell margination in the periphery observed beginning 6 hours post first dose of EIP0929 in all treated animals compared to controls (FIGS. 20B and 20C). T cell levels trended back to baseline or slightly above prior to 2^nd dose and were above baseline after 2^nd and 3^rd dosing for animals treated >50 pg/kg/dose.

[00639] Immunophenotypmg

[00640] Tissue Collection for Immunophenotypmg

[00641] On Day 29 (Groups 2, 3, and 4) or Day 30 (Group 1) of the dosing phase, all animals/sex/group had spleen and mesenteric lymph nodes collected. Following macroscopic examination and organ weight collection, sections of tissue for immunophenotypmg were weighed and placed into a media-containing tube for flow' cytometry analysis. Remaining spleen samples were preserved in 10% neutral-buffered formalin for histology and histopathology.

[00642] Table 5.5 Spleen Tissue Immunophenotypmg

[00644] Histology

[00645] Tissues from each animal at term inal necropsy were embedded in paraffin.

Embedded spleen and mesenteric lymph node tissues were sectioned at a nominal 5 pm and stained with hematoxylin and eosin. The tissue blocks of the mesenteric lymph nodes and spleens were serial sectioned for histopathology and immunohistochemistry for the detection of CD 19.

[00646] Tissue phenotyping revealed that in the spleen and lymph node, decreases in the relative percentages of B cells were noted in most treated animals compared to controls (FIG, 21A-21B). B cell depletions were lowest in animals administered 10 pg/kg/dose with more depletions in animals administered 50 or 150 pg/kg/dose (Table 5.7). Near complete B ceil depletion was observed at 150 pg/kg dose.

[00647] Table 5.7

[00648] Immunohistochemistry of spleens and mesenteric lymph nodes at terminal necropsy showed minimal to marked decreased splenic CD 19 stain and minimal to marked decreased mesenteric lymph node CD19 stain for animals administered >10 pg/kg/dose (FIG. 21C).

[00649] IL-6 excursions were noted at 2 and 6 hours post first dose in all treated animals compared to controls. IL-6 levels trended back to baseline at 24 hours post first dose (FIG.

22A). Lower levels excursions were also noted at 2 and 6 hours post second dose that trended back to baseline at 24 hours post 2nd dose. Reported levels after Glofitamab or Epcoritamab administration at doses which resulted in sustained peripheral B cell depletions resulted in IL6 levels that were about 1 .5- to 2-fold higher than ETP0929 (Frances N et al., Journal of Pharmaceutical Sciences 1 11 (2022) 1208--- 1218; Engelberts PJ et al., EBioMedicine 52 (2020) 102625).

[00650] IL-2 excursions were noted at 2 and 6 hours post first dose in animals administered >50 pg/kg/dose (FIG. 22B). TNF-a excursions were noted at 2 hours post first dose in a subset of animals administered 50 or 150 pg/kg/dose, compared to controls (FIG. 22B).

IFNy excursions were noted at 2 hours in only three animals administered 50 or 150 pg/kg/dose compared to controls (FIG. 22B). Reported levels of IFNy after Epcoritamab administration at the dose which resulted in sustained peripheral B cell depletions (intravenous (IV) and subcutaneous (SC) at 1 mg/kg were about 2.5- to 5-fold higher than EIP0929 (Engelberts PJ et al., EBioMedicine 52 (2020) 102625). The levels of IL-2 at 2 hours post EIPO0929 administration are similar to the reported IL-2 levels after Epcoritamab at 1 mg/kg administration (SC) (Engelberts PJ et al., EBioMedicine 52 (2020) 102625), which could indicate that EIP0929 induced similar T cell activations with lower cytokine excursions compared to optimal SC administration of Epcoritamab.

[00651] A two compartmental model was used to determine the pharmacokinetics profiles in Cynomolgtis monkeys. Data analysis from plasma concentration-time profiles of doses ranging 10 pg/kg to 150 pg/kg demonstrated a very rapid distribution of EIP0929 with a mean half-life of about 3.2 days (FIG. 23). This shows the exposure of E1P0929 is about ~10 fold higher than the reported value for clinically approved biologic Glofitamab in a similar dose range (Frances N et al.. Journal of Pharmaceutical Sciences 1 1 1 (2022) 1208-1218).

OTHER EMBODIMENTS

[00652] While the invention has been described in conjunction with the detailed description thereof, the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims.

[00653] Additional embodiments of the disclosure include the following:

Embodiment 1. A bispecific antibody comprising a first antigen binding region that binds to CD3 and a second antigen binding region that binds to CD20, wherein the first antigen binding region that binds to CD3 comprises three heavy chain complementarity determining regions (CDRH1, CDRH2, CDRH3) and three light chain complementarity determining regions (CDRL1, CDRL2, CDRL3), wherein a) CDRH1 comprises the amino acid sequence of SEQ ID NO: 29;

CDRII2 comprises the amino acid sequence of SEQ ID NO: 34; CDRH3 comprises the amino acid sequence of SEQ ID NO: 40; CDRL1 comprises the amino acid sequence of SEQ ID NO: 42;

CDRL2 comprises the amino acid sequence of SEQ ID NO: 43; and

CDRL3 comprises the amino acid sequence of SEQ ID NO: 47; b) CDRH1 comprises the amino acid sequence of SEQ ID NO: 29;

CDRH2 comprises the amino acid sequence of SEQ ID NO: 35;

CDRH3 comprises the amino acid sequence of SEQ ID NO: 38;

CDRL1 comprises the amino acid sequence of SEQ ID NO: 42;

CDRL2 comprises the amino acid sequence of SEQ ID NO: 43; and

CDRL3 comprises the amino acid sequence of SEQ ID NO: 47; c) CDRH1 comprises the amino acid sequence of SEQ ID NO: 30;

CDRH2 comprises the amino acid sequence of SEQ ID NO: 34;

CDRH3 comprises the amino acid sequence of SEQ ID NO: 37;

CDRL1 comprises the amino acid sequence of SEQ ID NO: 42;

CDRL2 comprises the amino acid sequence of SEQ ID NO: 43; and

CDRL3 comprises the amino acid sequence of SEQ ID NO: 45; d) CDRIII comprises the amino acid sequence of SEQ ID NO: 29;

CDRH2 comprises the amino acid sequence of SEQ ID NO: 34;

CDRH3 comprises the amino acid sequence of SEQ ID NO: 39;

CDRL1 comprises the amino acid sequence of SEQ ID NO: 42;

CDRL2 comprises the amino acid sequence of SEQ ID NO: 43; and

CDRL3 comprises the amino acid sequence of SEQ ID NO: 47; e) CDRH1 comprises the amino acid sequence of SEQ ID NO: 29;

CDRII2 comprises the amino acid sequence of SEQ ID NO: 34;

CDRH3 comprises the amino acid sequence of SEQ ID NO: 37;

CDRL1 comprises tire amino acid sequence of SEQ ID NO: 42;

CDRL2 comprises the amino acid sequence of SEQ ID NO: 44; and

CDRL3 comprises the amino acid sequence of SEQ ID NO: 45; or f) CDRH1 comprises the amino acid sequence of SEQ ID NO: 29;

CDRH2 comprises the amino acid sequence of SEQ ID NO: 34;

CDRH3 comprises the amino acid sequence of SEQ ID NO: 37;

CDRL1 comprises the amino acid sequence of SEQ ID NO: 42;

CDRL2 comprises tire amino acid sequence of SEQ ID NO: 43; and CDRL3 comprises the amino acid sequence of SEQ ID NO: 45; and wherein the second antigen binding region that binds to CD20 comprises three heavy chain complementarity determining regions (CDRH1, CDRH2, CDRH3) and three light chain complementarity determining regions (CDRL1, CDRL2, CDRL3): i) CDRH1 comprises the amino acid sequence of SEQ ID NO: 454;

CDRH2 comprises the amino acid sequence of SEQ ID NO: 456;

CDRH3 comprises the ammo acid sequence of SEQ ID NO: 458;

CDRL1 comprises the amino acid sequence of SEQ ID NO: 463;

CDRL2 comprises the amino acid sequence of SEQ ID NO: 464; and

CDRL3 comprises the amino acid sequence of SEQ ID NO: 465; or ii) CDRH1 comprises the amino acid sequence of SEQ ID NO: 459;

CDRH2 comprises the amino acid sequence of SEQ ID NO: 460;

CDRH3 comprises the amino acid sequence of SEQ ID NO: 462;

CDRL1 comprises the amino acid sequence of SEQ ID NO: 466;

CDRL2 comprises the amino acid sequence of SEQ ID NO: 467; and

CDRI.3 comprises the amino acid sequence of SEQ ID NO: 468.

Embodiment 2. The bispecific antibody of embodiment 1, wherein the first antigen binding region that binds to CD3 comprises a variable heavy chain region (VH) and a variable light chain region (VL), wherein: a) VH comprises the amino acid sequence of SEQ ID NO: 19 and

VL comprises the ammo acid sequence of SEQ ID NO: 26; b) VH comprises the amino acid sequence of SEQ ID NO: 18 and

VL comprises the ammo acid sequence of SEQ ID NO: 26; c) VH comprises the amino acid sequence of SEQ ID NO: 17 and

VL comprises the amino acid sequence of SEQ ID NO: 22; d) VH comprises the amino acid sequence of SEQ ID NO: 16 and

VL comprises the amino acid sequence of SEQ ID NO: 26; e) VH comprises the amino acid sequence of SEQ ID NO: 13 and

VL comprises the amino acid sequence of SEQ ID NO: 27; or f) VH comprises the amino acid sequence of SEQ ID NO: 13 and

VL comprises the amino acid sequence of SEQ ID NO: 22; and wherein the second antigen binding region that binds to CD20 comprises a variable heavy chain region (VH) and a variable light chain region (VL), wherein: i) VH comprises the amino acid sequence of SEQ ID NO: 451 and VL comprises the amino acid sequence of SEQ ID NO: 450; or ii) VH comprises the amino acid sequence of SEQ ID NO: 453 and VL comprises the amino acid sequence of SEQ ID NO: 452.

Embodiment 3. The bispecific antibody of any one of embodiments 1-2, wherein the bispecific antibody has the following structure: a first heavy chain polypeptide (H1 ) comprising a variable region (VH1), and a constant region (CH1) having a constant region 1 domain (CHI HI), a hinge region (H1H), a constant region 2 domain (CH1 H2) and a constant region 3 domain (CH1 _ H3 ); and a first light chain polypeptide (L1) comprising a variable region (VL1) and a constant region (CL1); and a second heavy chain polypeptide (H2) comprising a variable region (VH2), and a constant region (CH2) having a constant region 1 domain (CH2 H1 ), a hinge region (H2H), a constant region 2 domain (CH2 H2) and a constant region 3 domain (CH2__H3); and second light chain polypeptide (L2) comprising a variable region (VL2) and a constant region (CL2), and wherein i) the amino acid at position 39 (Kabat numbering) of the VH1 is a K and the amino acid at position 38 (Kabat numbering) of the VL1 is a D; ii) the ammo acid at position 147 (EU numbering) of the CH1_H1 is a K and the amino acid at positi on 131 (EU numbering) of the CL1 is a D; Hi) the amino acid at position 173 (EU numbering) of the CH1 H1 is a C and the amino acid at position 162 (EU numbering) of the CL1 is a C; iv) the ammo acid at position 220 (EU numbering) in the H1H is a S and the amino acid at position 214 (EU numbering) of the CL1 is a S; and i) the amino acid at position 39 (Kabat numbering) of the VH2 is a D and the amino acid at position 38 (Kabat numbering) of the VL2 is a K; and ii) the amino acid at position 147 (EU numbering) of the CH2_H1 is a D and the amino acid at positi on 180 (EU numbering) of the CL2 is a R.

Embodiment 4. The bispecific antibody of embodiment 3, wherein i) the amino acid at position 87 (Kabat numbering) of the VH1 and/or VH2 is a G; and ii) the amino acid at position 45 (Kabat numbering) of the VL1 and/or VL2 is aW. Embodiment 5. The antibody of any one of embodiments 3-4, wherein: i) the CH1__H3 has a C at position 349, an S at position 366, an A at position 368 and a V at position 407 (EU numbering); and the CH2 _ H3 has a C at position 354 and a W at position 366 (EU numbering); ii) the CH2 _ H3 has a C at position 349, an S at position 366, an A at position 368 and a V at position 407 (EU numbering); and the CH1_H3 has a C at position 354 and a W at position 366 (EU numbering); iii) the CH1_H3 has a C at position 354, an S at position 366, an A at position 368 and a V at position 407 (EU numbering); and the CH2_H3 has a C at position 349 and a W at position 366 (EU numbering); or iv) the CH2 _ H3 has a C at position 354, an S at position 366, an A at position 368 and a V at position 407 (EU numbering); and the CH1_H3 has a C at position 349 and a W at position 366 (EU numbering).

Embodiment 6. The antibody of any one of embodiments 3-5, wherein the amino acid at position 447 (EU numbering) of the CH1 _ H3 and/or of the CH2_H3 is deleted.

Embodiment 7. The antibody of any one of embodiments 3-6, wherein: i) the H1H and/or the H2H has an A at positions 234 and 235 (EU numbering); ii) the H1H and/or the H2H has an A at positions 234, 235 and 237 (EU numbering); or iii) the H1H and/or the H2H has an A at positions 234 and 235 and G at position 329 (EU numbering).

Embodiment 8. The antibody of any one of embodiments 3-7, wherein i) the CH1 H3 and/or the CH2 _ H3 has an A at position 297 (EU numbering); ii) the CH1__H3 and/or the CH2__H3 has a G at position 297 (EU numbering); or iii) the CH1 H3 and/or the CH2 _ H3 has a S at position 297 (EU numbering).

Embodiment 9. The antibody of any one of embodiments 3-8, wherein the CH1 _ H3 and/or the CH2 _ H3 has an S at position 331 (EU numbering).

Embodiment 10. The bispecific antibody of any one of embodiments 1-9, wherein a polypeptide is fused to the N-terminus or the C-terminus of the first heavy chain polypeptide or the second heavy chain polypeptide.

Embodim ent 11. The bispecific antibody of embodiment 10, wherein the polypeptide is fused to the C-terminus of the first heavy chain polypeptide.

Embodiment 12. The bispecific antibody of any one of embodiments 10-11, wherein the polypeptide is fused via a linker peptide. Embodim ent 13. The bispecific antibody of embodiment 12, wherein the linker peptide comprises the amino acid sequence of SEQ ID NO: 53,

Embodiment 14. The bispecific antibody of any one of embodiments 10-13, wherein the polypeptide comprises a CD58 or a fragment thereof.

Embodiment 15. The bispecific antibody of embodiment 14, wherein the CD58 comprises the amino acid sequence of SEQ ID NO: 49.

Embodiment 16. The bispecific antibody of any one of embodiments 1-15, wherein the bispecific antibody is an IgGl or an IgG4 antibody.

Embodiment 17. The bispecific antibody of any one of embodiments 1-16, wherein the bispecific antibody is a monoclonal antibody, a chimeric antibody or a humanized antibody. Embodiment 18. A polynucleotide comprising a nucleic acid sequence encoding the bi specific antibody according to any one of embodiments 1-17.

Embodiment 19. A vector comprising the polynucleotide of embodiment 18.

Embodiment 20. A pharmaceutical composition comprising the bispecific antibody of any one of embodiments 1-17, the polynucleotide of embodiment 18 or the vector of embodiment 19 and a pharmaceutically acceptable carrier.

Embodiment 21. A method of treating a cancer expressing CD20 in a subject in need thereof comprising administering a therapeutically effective amount of the pharmaceutical composition of embodiment 2,0.

Embodiment 22. A method of T-cell re-targeting in a subject in need thereof comprising administering a therapeutically effective amount of the pharmaceutical composition of embodiment 20.

Embodiment 23. A method of T-cell activation in a subject in need thereof comprising administering a therapeutically effective amount of the pharmaceutical composition of embodiment 20.

Embodiment 24. The method of any one of embodiments 21-23, wherein the subject has a B-cell cancer.

Embodiment 25. The method of embodiment 24, wherein the B-cell cancer is follicular lymphoma, B-cell chronic lymphocytic leukemia, B-cell lymphoblastic lymphoma, Hodgkin lymphoma, non-Hodgkin’s lymphoma (NHL), chronic lymphoid leukemia (CLL), diffuse large B-cell lymphoma, marginal zone lymphoma, Mantle cell lymphoma, hairy cell leukemia, Burkitt lymphoma or small lymphocytic lymphoma (SLL), diffuse-large B cell lymphoma (DLBCL), a primary mediastinal (thymic) large B cell lymphoma (PMLBCL). Embodiment 26. The method of any one of embodiments 21-25, wherein the subject is simultaneously administered or previously administered with a therapeutically effective amount of an additional therapeutic agent.

Embodiment 27. Tire method of embodiment 26, wherein the additional therapeutic agent is an anti-CD20 monospecific antibody.

Embodiment 28. The method of embodiment 27, wherein the anti-CD20 monospecific therapy is rituximab.

Embodiment 29. The method of embodiment 26, wherein the additional therapeutic agent is a CAR-T ceil therapy, an immune checkpoint inhibitor, a co-stimulatory ligand or a cytokine.

Claims

CLAIMS What is claimed is:

1. A bispecific antibody comprising a first antigen binding region that binds to CD3 and a second antigen binding region that binds to CD20, wherein the first antigen binding region that binds to CD 3 comprises three heavy chain complementarity determining regions (VH1 CDR1, VH 1_CDR2, VH1 CDR3) and three light chain complementarity determining regions (VL1 CDR1, VL1 CDR2, VL1 CDR3), wherein a) VH1 CDR1 comprises the amino acid sequence of SEQ ID NO: 29; VH 1_CDR2 comprises the amino acid sequence of SEQ ID NO: 34;

VH1 CDR3 comprises the amino acid sequence of SEQ ID NO: 40; VL1 CDRI comprises the amino acid sequence of SEQ ID NO: 42;

VL1 CDR2 comprises the amino acid sequence of SEQ ID NO: 43; and

VL1 CDR3 comprises the amino acid sequence of SEQ ID NO: 47; b) VH1__CDR1 comprises the amino acid sequence of SEQ ID NO: 29; VH 1_CDR2 comprises the amino acid sequence of SEQ ID NO: 35;

VH1 CDR3 comprises the amino acid sequence of SEQ ID NO: 38; VL1 CDRI comprises the amino acid sequence of SEQ ID NO: 42;

VL1 CDR2 comprises the amino acid sequence of SEQ ID NO: 43; and

VL1 CDR3 comprises the amino acid sequence of SEQ ID NO: 47; c) VH1 CDR1 comprises the amino acid sequence of SEQ ID NO: 30; VH 1_CDR2 comprises the amino acid sequence of SEQ ID NO: 34;

VH1 CDR3 comprises the amino acid sequence of SEQ ID NO: 37; VL1 CDRI comprises the amino acid sequence of SEQ ID NO: 42;

VL1 CDR2 comprises the amino acid sequence of SEQ ID NO: 43; and

VL1 CDR3 comprises the amino acid sequence of SEQ ID NO: 45; d) VH1 CDR1 comprises the amino acid sequence of SEQ ID NO: 29; VH 1_CDR2 comprises the amino acid sequence of SEQ ID NO: 34;

VH1 CDR3 comprises the amino acid sequence of SEQ ID NO: 39; VL1 CDRI comprises the amino acid sequence of SEQ ID NO: 42;

VL1 CDR2 comprises the amino acid sequence of SEQ ID NO: 43; and

VL1 CDR3 comprises the amino acid sequence of SEQ ID NO: 47; e) VH1__CDR1 comprises the amino acid sequence of SEQ ID NO: 29; VH 1_CDR2 comprises the amino acid sequence of SEQ ID NO: 34;

VH1 CDR3 comprises the amino acid sequence of SEQ ID NO: 37;

VL1 CDR1 comprises the amino acid sequence of SEQ ID NO: 42;

VL1 CDR2 comprises the amino acid sequence of SEQ ID NO: 44; and

VL1 CDR3 comprises the amino acid sequence of SEQ ID NO: 45; or f) VH1 CDR1 comprises the amino acid sequence of SEQ ID NO: 29; VH 1_CDR2 comprises the amino acid sequence of SEQ ID NO: 34;

VH1 CDR3 comprises the amino acid sequence of SEQ ID NO: 37;

VL1 CDR1 comprises the amino acid sequence of SEQ ID NO: 42;

VL1 CDR2 comprises the amino acid sequence of SEQ ID NO: 43; and

VL1 CDR3 comprises the amino acid sequence of SEQ ID NO: 45; and wherein the second antigen binding region that binds to CD20 comprises three heavy chain complementarity determining regions (VH2_CDR1, VH2_CDR2, VH2 CDR3) and three light chain complementarity determining regions (VL2 CDR1 , VI.2 CDR2. VI.2 CDR3), wherein i) VH2 CDR1 comprises the amino acid sequence of SEQ ID NO: 455;

VH2_CDR2 comprises the amino acid sequence of SEQ ID NO: 457;

VH2 CDR3 comprises the amino acid sequence of SEQ ID NO: 458;

VL2 CDR1 comprises the amino acid sequence of SEQ ID NO: 463;

VL2 CDR2 comprises the amino acid sequence of SEQ ID NO: 464; and

VL2 CDR3 comprises the amino acid sequence of SEQ ID NO: 465; or ii) VH2 CDR1 comprises the amino acid sequence of SEQ ID NO: 459;

VH2 CDR2 comprises the amino acid sequence of SEQ ID NO: 460;

VH2 CDR3 comprises the amino acid sequence of SEQ ID NO: 462;

VL2 CDR 1 comprises the amino acid sequence of SEQ ID NO: 466;

VL2 CDR2 comprises the amino acid sequence of SEQ ID NO: 467; and

VL2 CDR3 comprises the amino acid sequence of SEQ ID NO: 468.

2, The bi specific antibody of claim 1, wherein the first antigen binding region that binds to CD3 comprises a variable heavy chain region (VH1) and a variable light chain region (VL1), wherein: a) VH1 comprises the amino acid sequence of SEQ ID NO: 19 and

VL I comprises the amino acid sequence of SEQ ID NO: 26; b) VH1 comprises the amino acid sequence of SEQ ID NO: 18 and VL1 comprises the amino acid sequence of SEQ ID NO: 26; c) VH1 comprises the amino acid sequence of SEQ ID NO: 17 and VL1 comprises the amino acid sequence of SEQ ID NO: 22; d) VH1 comprises the ammo acid sequence of SEQ ID NO: 16 and VL1 comprises the amino acid sequence of SEQ ID NO: 26; e) VH1 comprises the amino acid sequence of SEQ ID NO: 13 and VL1 comprises the amino acid sequence of SEQ ID NO: 27; or f) VH1 comprises the amino acid sequence of SEQ ID NO: 13 and VL1 comprises the amino acid sequence of SEQ ID NO: 22; and wherein the second antigen binding region that binds to CD20 comprises a variable heavy chain region (VH2) and a variable light chain region (VL2), wherein: i) VH2 comprises the amino acid sequence of SEQ ID NO: 451 and VL2 comprises the amino acid sequence of SEQ ID NO: 450; or ii) VH2 comprises the amino acid sequence of SEQ ID NO: 453 and VL2 comprises the amino acid sequence of SEQ ID NO: 452.

3. The bispecific antibody of any one of claims 1-2, wherein the bispecific antibody has the following structure: a first heavy chain polypeptide (H1 ) comprising a variable region (VH1), and a constant region (CHI) having a constant region 1 domain (CHI JH1), a hinge region (H1H), a constant region 2 domain (CH1 H2) and a constant region 3 domain (CHI __H3); and a first light chain polypeptide (L1) comprising a variable region (VL1) and a constant region (CL1); and a second heavy chain polypeptide (H2) comprising a variable region (VH2), and a constant region (CH2) having a constant region 1 domain (CH2 H1), a hinge region (H2H), a constant region 2 domain (CH2JH2) and a constant region 3 domain (CH2__H3); and second light chain polypeptide (L2) comprising a variable region (VL2) and a constant region (CL2), and wherein i) the amino acid at position 39 (Kabat numbering) of the VH1 is a K and the amino acid at position 38 (Kabat numbering) of the VL1 is a D; ii) the amino acid at position 147 (EU numbering) of the CH1_H1 is a K and the amino acid at positi on 131 (EU numbering) of the CL1 is a D; iii) the amino acid at position 173 (EU numbering) of the CH1 H1 is a C and the amino acid at position 162 (EU numbering) of the CL1 is a C; iv) the ammo acid at position 220 (EU numbering) in the H1H is a S and the amino acid at position 214 (EU numbering) of the CL1 is a S; and i) the amino acid at position 39 (Kabat numbering) of the VH2 is a D and the amino acid at position 38 (Kabat numbering) of the VL2 is a K; and ii) the amino acid at position 147 (EU numbering) of the CH2_H1 is a D and the amino acid at positi on 180 (EU numbering) of the CL2 is a R.

4. The bispecific antibody of claim 3, wherein i) the amino acid at position 87 (Kabat numbering) of the VH1 and/or VH2 is a G; and ii) the amino acid at position 45 (Kabat numbering) of the VL1 and/or VL2 is a W.

5. The antibody of any one of ciaims 3-4, wherein: i) the CH1 H3 has a C at position 349, an S at position 366, an A at position 368 and a V at position 407 (EU numbering); and the CH2JH3 has a C at position 354 and a W at position 366 (EU numbering); ii) the CH2_H3 has a C at position 349, an S at position 366, an A at position 368 and a V at position 407 (EU numbering); and the CH1 H3 has a C at position 354 and a W at position 366 (EU numbering); iii) the CH1_H3 has a C at position 354, an S at position 366, an A at position 368 and a V at position 407 (EU numbering); and the CH2 _ H3 has a C at position 349 and a W at position 366 (EU numbering); or iv) the CH2_H3 has a C at position 354, an S at position 366, an A at position 368 and a V at position 407 (EL⁷ numbering); and the CH1 H3 has a C at position 349 and a W at position 366 (EU numbering).

6. The antibody of any one of claims 3-5, wherein the amino acid at position 447 (EU numbering) of the CH1 _ H3 and/or of the CH2 _ H3 is deleted.

7. The antibody of any one of claims 3-6, wherein: i) the H1H and/or the H2H has an A at positions 2.34 and 235 (EU numbering); ii) the H1H and/or the H2H has an A at positions 234, 235 and 237 (EU numbering); or iii) the H1H and/or the H2H has an A at positions 234 and 235 and G at position 329 (EU numbering).

8. The antibody of any one of claims 3-7, wherein i) the CH1 _ H3 and/or tire CH2 _ H3 has an A at position 297 (EU numbering); ii) the CH1__H3 and/or the CH2 _ H3 has a G at position 297 (EU numbering); or iii) the CH1 H3 and/or the CH2 _ H3 has a S at position 297 (EU numbering).

9. The anti body of any one of claims 3-8, wherein the CH1_H3 and/or the CH2_H3 has an S at position 331 (EU numbering).

10. The bispecific antibody of any one of claims 1-9, wherein a polypeptide is fused to the N-temiinus or the C -terminus of the first heavy chain polypeptide or the second heavy chain polypeptide.

11. The bi specific antibody of claim 10, wherein the polypeptide is fused to the C- terminus of the first heavy chain polypeptide.

12. The bispecific antibody of any one of claims 10-11, wherein the polypeptide is fused via a linker peptide.

13. The bispecific antibody of claim 12, wherein the linker peptide comprises the amino acid sequence of SEQ ID NO: 53.

14. The bispecific antibody of any one of claims 10-13, wherein the polypeptide comprises a CD58 or a fragment thereof.

15. The bispecific antibody of claim 14, wherein the CD58 comprises the amino acid sequence of SEQ ID NO: 49.

16. The bispecific antibody of any one of claims 1-15, wherein i) the VHI comprises the amino acid sequence of SEQ ID NO: 19; the VL1 comprises the ammo acid sequence of SEQ ID NO: 26; the VH2 comprises the amino acid sequence of SEQ ID NO: 453; and the VL2 comprises the amino acid sequence of SEQ ID NO: 452; ii) the VHI comprises the amino acid sequence of SEQ ID NO: 18; the VL1 comprises the amino acid sequence of SEQ ID NO: 26; the VH2 comprises the amino acid sequence of SEQ ID NO: 453; and the VL2 comprises the amino acid sequence of SEQ ID NO: 452; iii) the VHI comprises the amino acid sequence of SEQ ID NO: 17; the VL1 comprises the amino acid sequence of SEQ ID NO: 22; the VH2 comprises the amino acid sequence of SEQ ID NO: 453; and die VL2 comprises the amino acid sequence of SEQ ID NO: 452; iv) the VHI comprises the ammo acid sequence of SEQ ID NO: 16; the VL1 comprises the amino acid sequence of SEQ ID NO: 26; the VH2 comprises the amino acid sequence of SEQ ID NO: 453; and the VL2 comprises the ammo acid sequence of SEQ ID NO: 452; v) the VHI comprises the amino acid sequence of SEQ ID NO: 13; the VL1 comprises the amino acid sequence of SEQ ID NO: 27; die VH2 comprises the amino acid sequence of SEQ ID NO: 453; and the VL2 comprises the amino acid sequence of SEQ ID NO: 452; vi) the VHI comprises the amino acid sequence of SEQ ID NO: 13; the VL1 comprises the amino acid sequence of SEQ ID NO: 22; the VH2 comprises die amino acid sequence of SEQ ID NO: 453; and the VL2 comprises the amino acid sequence of SEQ ID NO: 452; vii) the VHI comprises the amino acid sequence of SEQ ID NO: 19; die VL1 comprises the amino acid sequence of SEQ ID NO: 26; the VH2 comprises the amino acid sequence of SEQ ID NO: 451; and the VL2 comprises the amino acid sequence of SEQ ID NO: 450; viii) the VH1 comprises the amino acid sequence of SEQ ID NO: 18; tlie VL1 comprises the amino acid sequence of SEQ ID NO: 26; die VH2 comprises the amino acid sequence of SEQ ID NO: 451; and the VL2 comprises the amino acid sequence of SEQ ID NO: 450; ix) the VHI comprises the amino acid sequence of SEQ ID NO: 17; the VL1 comprises the ammo acid sequence of SEQ ID NO: 22; the VH2 comprises the amino acid sequence of SEQ ID NO: 451; and the VL2 comprises the amino acid sequence of SEQ ID NO: 450; x) the VHI comprises the amino acid sequence of SEQ ID NO: 16; die VL1 comprises the amino acid sequence of SEQ ID NO: 26; the VH2 comprises the amino acid sequence of SEQ ID NO: 451 ; and the VL2 comprises the amino acid sequence of SEQ ID NO: 450; xi) the VHI comprises the amino acid sequence of SEQ ID NO: 13; the VL1 comprises the amino acid sequence of SEQ ID NO: 27; the VH2 comprises the amino acid sequence of SEQ ID NO: 451; and die VL2 comprises the amino acid sequence of SEQ ID NO: 450; or xii) tire VHI comprises the amino acid sequence of SEQ ID NO: 13; the VL1 comprises the amino acid sequence of SEQ ID NO: 22; the VH2 comprises the amino acid sequence of SEQ ID NO: 451 ; and the VL2 comprises the ammo acid sequence of SEQ ID NO: 450.

17. The bispecific antibody of claim 16, wherein i) the H1 comprises the amino acid sequence of SEQ ID NO: 491; die L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 490; and the L2 comprises the amino acid sequence of SEQ ID NO: 471 ; ii) die H1 comprises the amino acid sequence of SEQ ID NO: 491; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 566; and die L2 comprises the amino acid sequence of SEQ ID NO: 471; iii) the H1 comprises the amino acid sequence of SEQ ID NO: 595; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 490; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; iv) the H1 comprises the amino acid sequence of SEQ ID NO: 595; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 566; and the L2 comprises the ammo acid sequence of SEQ ID NO: 471; v) the H1 comprises the amino acid sequence of SEQ ID NO: 494; the L1 comprises the ammo acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 493; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; vi) the H1 comprises the amino acid sequence of SEQ ID NO: 494; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 567; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; vii) the H1 comprises the ammo acid sequence of SEQ ID NO: 596; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the ammo acid sequence of SEQ ID NO: 493; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; viii) the H1 comprises the ammo acid sequence of SEQ ID NO: 596; the L1 comprises the ammo acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 567; and the L2 comprises the ammo acid sequence of SEQ ID NO: 471; ix) the H1 comprises the amino acid sequence of SEQ ID NO: 497; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 496; and the L2 comprises the amino acid sequence of SEQ ID NO: 471 ; x) the H1 comprises the amino acid sequence of SEQ ID NO: 497; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 568; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xi) the H1 comprises the amino acid sequence of SEQ ID NO: 597; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 496; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xii) the H1 comprises the amino acid sequence of SEQ ID NO: 597; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 568; and the L2 comprises the ammo acid sequence of SEQ ID NO: 471; xiii) the H1 comprises the amino acid sequence of SEQ ID NO: 500; the L1 comprises the ammo acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 499; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xiv) the H1 comprises the amino acid sequence of SEQ ID NO: 500; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 569; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xv) the H1 comprises the amino acid sequence of SEQ ID NO: 598; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises tire ammo acid sequence of SEQ ID NO: 499; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xvi) the H1 comprises the amino acid sequence of SEQ ID NO: 598; the L1 comprises the ammo acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 569; and the L2 comprises the ammo acid sequence of SEQ ID NO: 471; xvii) the H1 comprises the amino acid sequence of SEQ ID NO: 503; the L1 comprises the amino acid sequence of SEQ ID NO: 153; the H2 comprises the amino acid sequence of SEQ ID NO: 502; and the L2 comprises the amino acid sequence of SEQ ID NO: 471 ; xviii) the H1 comprises the amino acid sequence of SEQ ID NO: 503; the L1 comprises the amino acid sequence of SEQ ID NO: 153; the H2 comprises the amino acid sequence of SEQ ID NO: 570; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xix) the H1 comprises the amino acid sequence of SEQ ID NO: 599; the L1 comprises the amino acid sequence of SEQ ID NO: 153; the H2 comprises the amino acid sequence of SEQ ID NO: 502; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xx) the H1 comprises the ammo acid sequence of SEQ ID NO: 599; the L1 comprises the amino acid sequence of SEQ ID NO: 153; the H2 comprises the amino acid sequence of SEQ ID NO: 570; and the L2 comprises the ammo acid sequence of SEQ ID NO: 471; xxi) the H1 comprises the amino acid sequence of SEQ ID NO: 506; the L1 comprises the ammo acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 505; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xxii) the H1 comprises the amino acid sequence of SEQ ID NO: 506; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 571; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xxiii) the H1 comprises the amino acid sequence of SEQ ID NO: 600; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the ammo acid sequence of SEQ ID NO: 505; and the L2 comprises the amino acid sequence of SEQ ID NO: 471; xxiv) the H1 comprises the amino acid sequence of SEQ ID NO: 600; the L1 comprises the ammo acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 571; and the L2 comprises the ammo acid sequence of SEQ ID NO: 471; xxv) the H1 comprises the amino acid sequence of SEQ ID NO: 527; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 526; and the L2 comprises the amino acid sequence of SEQ ID NO: 525; xxvi) the H1 comprises the ammo acid sequence of SEQ ID NO: 527; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 572; and the L2 comprises the amino acid sequence of SEQ ID NO: 525; xxvii) the H1 comprises the amino acid sequence of SEQ ID NO: 601; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 526; and the L2 comprises the amino acid sequence of SEQ ID NO: 525; xxviii) the H1 comprises the amino acid sequence of SEQ ID NO: 601; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 572; and the L2 comprises the ammo acid sequence of SEQ ID NO: 525; xxix) the H1 comprises the amino acid sequence of SEQ ID NO: 530; the L1 comprises the ammo acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 529; and the L2 comprises the amino acid sequence of SEQ ID NO: 528; xxx) the H1 comprises the amino acid sequence of SEQ ID NO: 530; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 573; and the L2 comprises the amino acid sequence of SEQ ID NO: 528; xxxi) the H1 comprises the amino acid sequence of SEQ ID NO: 602; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the ammo acid sequence of SEQ ID NO: 529; and the L2 comprises the amino acid sequence of SEQ ID NO: 528; xxxii) the H1 comprises the amino acid sequence of SEQ ID NO: 602; the L1 comprises the ammo acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 573; and the L2 comprises the ammo acid sequence of SEQ ID NO: 528; xxxiii) the H1 comprises the amino acid sequence of SEQ ID NO: 533; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 532; and the L2 comprises the amino acid sequence of SEQ ID NO: 531 ; xxxiv) the H1 comprises the amino acid sequence of SEQ ID NO: 533; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 574; and the L2 comprises the amino acid sequence of SEQ ID NO: 531; xxxv) the H1 comprises the amino acid sequence of SEQ ID NO: 603; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 532; and the L2 comprises the amino acid sequence of SEQ ID NO: 531; xxxvi) the H1 comprises the amino acid sequence of SEQ ID NO: 603; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 574; and the L2 comprises the ammo acid sequence of SEQ ID NO: 531; xxxvii) the H1 comprises the amino acid sequence of SEQ ID NO: 536; the L1 comprises the ammo acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 535; and the L2 comprises the amino acid sequence of SEQ ID NO: 534; xxxviii) the H1 comprises the amino acid sequence of SEQ ID NO: 536; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 575; and the L2 comprises the amino acid sequence of SEQ ID NO: 534; xxxix) the H1 comprises the amino acid sequence of SEQ ID NO: 604; the L1 comprises the amino acid sequence of SEQ ID NO: 149; the H2 comprises the ammo acid sequence of SEQ ID NO: 535; and the L2 comprises the amino acid sequence of SEQ ID NO: 534; xl) the HI comprises the amino acid sequence of SEQ ID NO: 604; the L1 comprises the ammo acid sequence of SEQ ID NO: 149; the H2 comprises the amino acid sequence of SEQ ID NO: 575; and the L2 comprises the ammo acid sequence of SEQ ID NO: 534; xli) the H1 comprises the amino acid sequence of SEQ ID NO: 539; the L1 comprises the amino acid sequence of SEQ ID NO: 153; the H2 comprises the amino acid sequence of SEQ ID NO: 538; and the L2 comprises the amino acid sequence of SEQ ID NO: 537; xlii) the H1 comprises the amino acid sequence of SEQ ID NO: 539; the L1 comprises the amino acid sequence of SEQ ID NO: 153; the H2 comprises the amino acid sequence of SEQ ID NO: 576; and the L2 comprises the amino acid sequence of SEQ ID NO: 537; xliii) the H1 comprises the amino acid sequence of SEQ ID NO: 605; the L1 comprises the amino acid sequence of SEQ ID NO: 153; the H2 comprises the amino acid sequence of SEQ ID NO: 538; and the L2 comprises the amino acid sequence of SEQ ID NO: 537; xliv) the H1 comprises the amino acid sequence of SEQ ID NO: 605; the L1 comprises the amino acid sequence of SEQ ID NO: 153; the H2 comprises the amino acid sequence of SEQ ID NO: 576; and the L2 comprises the ammo acid sequence of SEQ ID NO: 537; xlv) the H1 comprises the amino acid sequence of SEQ ID NO: 542; the L1 comprises the ammo acid sequence of SEQ ID NO: 141; die H2 comprises the amino acid sequence of SEQ ID NO: 541; and the L2 comprises the amino acid sequence of SEQ ID NO: 540; xlvi) the H1 comprises the amino acid sequence of SEQ ID NO: 542; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 577; and the L2 comprises the amino acid sequence of SEQ ID NO: 540; xlvii) the H1 comprises the amino acid sequence of SEQ ID NO: 606; the L1 comprises the amino acid sequence of SEQ ID NO: 141; the H2 comprises tire ammo acid sequence of SEQ ID NO: 541; and the L2 comprises the amino acid sequence of SEQ ID NO: 540; or xlviii) the H1 comprises the amino acid sequence of SEQ ID NO: 606; the L1 comprises the ammo acid sequence of SEQ ID NO: 141; the H2 comprises the amino acid sequence of SEQ ID NO: 577; and the L2 comprises the ammo acid sequence of SEQ ID NO: 540.

18. The bispecific antibody of any one of claims 1-17, wherein the bispecific antibody is an IgGl or an lgG4 antibody.

19. The bispecific antibody of any one of claims 1-18, wherein the bispecific antibody is a monoclonal antibody, a chimeric antibody or a humanized antibody.

20. A polynucleotide comprising a nucleic acid sequence encoding the bispecific antibody according to any one of claims 1 - 19.

21. A vector comprising the polynucleotide of claim 20.

22. A pharmaceutical composition comprising the bispecific antibody of any one of claims 1-19, the polynucleotide of claim 20 or the vector of claim 21 and a pharmaceutically acceptable carrier.

23. A method of treating a cancer expressing CD20 in a subject in need thereof comprising administering a therapeutically effective amount of the pharmaceutical composition of claim 22.

24. A method of T-cell re -targeting in a subject in need thereof comprising administering a therapeutically effective amount of the pharmaceutical composition of claim 22.

25. A method of T-cell activation in a subject in need thereof comprising administering a therapeutically effective amount of the pharmaceutical composition of claim 22.

26. The method of any one of claim s 23-25, wherein the subject has a B-cell cancer.

27. The method of claim 26, wherein the B-cell cancer is follicular lymphoma, B-cell chronic lymphocytic leukemia, B-cell lymphoblastic lymphoma, Hodgkin lymphoma, nonHodgkin’s lymphoma (NHL), chronic lymphoid leukemia (CLL), diffuse large B-cell lymphoma, marginal zone lymphoma, Mantle cell lymphoma, hairy cell leukemia, Burkitt lymphoma or small lymphocytic lymphoma (SLL), diffuse-large B cell lymphoma (DLBCL), a primary mediastinal (thymic) large B cell lymphoma (PMLBCL).

28. The method of any one of claims 23-27, wherein the subject is simultaneously administered or previously administered with a therapeutically' effective amount of an additional therapeutic agent.

29. The method of claim 2,8, wherein the additional therapeutic agent is an anti-CD20 monospecific antibody.

30. The method of claim 29, wherein the anti-CD20 monospecific therapy is rituximab.

31. The method of claim 28, wherein the additional therapeutic agent is a CAR-T cell therapy, an immune checkpoint inhibitor, a co-stimulatory ligand or a cytokine.