WO2022133492A1

WO2022133492A1 - Compositions and methods for generating anti-tumor immune response

Info

Publication number: WO2022133492A1
Application number: PCT/US2021/073007
Authority: WO
Inventors: Lan Huang; James R. Tonra
Original assignee: BeyondSpring Pharmaceuticals Inc
Current assignee: BeyondSpring Pharmaceuticals Inc
Priority date: 2020-12-18
Filing date: 2021-12-17
Publication date: 2022-06-23
Anticipated expiration: 2023-06-18
Also published as: TW202302107A

Abstract

Disclosed herein are compositions comprising dehydrophenylahistin analogs such as plinabulin and an anti-CD47 agent for treating cancer. Also disclosed here are methods of treating cancer by co-administering the dehydrophenylahistin analog and one or more anti-CD47 agent to a subject in need thereof. Some embodiments, the one or more anti-CD47 agent is an anti-CD47 antibody molecule capable of binding CD47, as well as fragments of these monospecific antibodies that are immunologically active and still bind to CD47.

Description

COMPOSITIONS AND METHODS FOR GENERATING ANTI TUMOR IMMUNE

RESPONSE

CROSS REFERENCE TO RELATED APPLIATIONS

[0001] This patent application claims the benefit of priority to U.S. Provisional Application No. 63/144348, filed February 1, 2021, and U.S. Provisional Application No. 63/127405, filed December 18, 2020. All of the foregoing applications are fully incorporated herein by reference in their entireties for all purposes.

REFERENCE TO SEQUENCE LISTING

[0002] The present application is filed with a Sequence Listing in electronic format. The Sequence Listing is provided as a file entitled DALWC.199WO_SEQLIST.txt, created December 17, 2021, which is approximately 288 KB in size. The information in the electronic format of the sequence listing is incorporated herein by reference in its entirety.

FIELD OF THE DISCLOSURE

[0003] The present disclosure relates to the field of chemistry and medicine. More particularly, the present disclosure relates to dehydrophenylahistin analogs, compositions including dehydrophenylahistin analogs, and their use in treatment.

BACKGROUND

[0004] The tumor growth promoting M2 phenotype is dominant in tumor associated macrophages (TAM) and correlates to poor prognosis. As Ml polarized macrophages possess anti-tumor functions, in-vivo reprogramming of macrophages is a promising strategy for cancer treatment. The alternate polarization of TAMs from M2 to Ml may elicit tumor- specific immune responses.

[0005] CD47 is a cell surface receptor comprised of an extracellular IgV set domain, a 5 transmembrane domain, and a cytoplasmic tail that is alternatively spliced. Two ligands bind CD47: signal inhibitory receptor protein a (SIRPa) and thrombospondin- 1 (TSP1). CD47 expression and/or activity have been implicated in a number of diseases and disorders. [0006] Tumor cells hijack the immunosuppressive mechanism by overexpressing CD47, which efficiently helps them to escape immune surveillance and killing by innate immune cells. (Majeti R, Ch et al., CD47 is an adverse prognostic factor and therapeutic antibody target on human acute myeloid leukemia stem cells, Cell. 2009 Jul. 23; 138(2):286-99; S. Jaiswal et al., CD47 is upregulated on circulating hematopoietic stem cells and leukemia cells to avoid phagocytosis, Cell. 2009 Jul. 23; 138(2):271- 85). CD47 expression is upregulated in most human cancers (e.g., NHL, AML, breast, colon, glioblastoma, glioma, ovarian, bladder and prostate cancers) and increased levels of CD47 expression clearly correlate with aggressive disease and poor survival. (Majeti R, et al., Cell. 2009 Jul. 23; 138(2):286-99; S. Jaiswal et al., Cell. 2009 Jul. 23; 138(2):271-85; Willingham S B, et al., The CD47-signal regulatory protein alpha (SIRPa) interaction is a therapeutic target for human solid tumors, Proc Natl Acad Sci USA. 2012 Apr. 24; 109(17):6662-7; Chao M P, et al., Therapeutic antibody targeting of CD47 eliminates human acute lymphoblastic leukemia, Cancer Res. 2011 Feb. 15; 71(4): 1374-84).

SUMMARY OF THE INVENTION

[0007] Some aspects of the disclosure relate to a method of treating a subject with cancer. In some embodiments, the method comprises administering a therapeutic dose of a compound of Formula (I):

wherein

Ri, R4, and Re, are each separately selected from the group consisting of a hydrogen atom, a deuterium atom, a halogen atom, and saturated Ci,-C24 alkyl, unsaturated Ci- C24 alkenyl, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, azido, substituted nitro, phenyl, and substituted phenyl groups, hydroxy, carboxy, — CO — O — R7, cyano, alkylthio, halogenated alkyl including polyhalogenated alkyl, halogenated carbonyl, and carbonyl — CH2CO — R7, wherein R7 is selected from a hydrogen atom, a halogen atom, and saturated C1-C24 alkyl, unsaturated C1-C24 alkenyl, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, azido, substituted nitro, phenyl, and substituted phenyl groups;

Ri' and Ri" are each independently selected from the group consisting of a hydrogen atom, a deuterium atom, a halogen atom, and saturated C1-C24 alkyl, unsaturated Ci- C24 alkenyl, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, azido, substituted nitro, phenyl, and substituted phenyl groups, hydroxy, carboxy, — CO — O — R7, cyano, alkylthio, halogenated alkyl including polyhalogenated alkyl, halogenated carbonyl, and carbonyl — CH2CO — R7, wherein R7 is selected from a hydrogen atom, a halogen atom, and saturated C1-C24 alkyl, unsaturated C1-C24 alkenyl, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, azido, substituted nitro, phenyl, and substituted phenyl groups;

R, Ri' and Ri" are either covalently bound to one another or are not covalently bound to one another;

R2, R3, and R5 are each separately selected from the group consisting of a hydrogen atom, a deuterium atom, a halogen atom, and saturated C1-C12 alkyl, unsaturated Ci- C12 alkenyl, acyl, cycloalkyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, and substituted nitro groups, sulfonyl and substituted sulfonyl groups; m is an integer equal to zero, one or two;

Xi and X2 are separately selected from the group consisting of an oxygen atom, a nitrogen atom and a sulfur atom, and

Y is selected from the group consisting of a nitrogen atom, a substituted nitrogen atom with a R5 group from above, an oxygen atom, a sulfur atom, a oxidized sulfur atom, a methylene group and a substituted methylene group; Z, for each separate n, if non-zero, and Zi, Z2, Za and Z4 are each separately selected from a carbon atom, a sulfur atom, a nitrogen atom or an oxygen atom; and the dashed bonds may be either single or double bonds an anti-CD47 agent.

[0008] In some embodiments, the anti-CD47 agent is an antibody or antigenbinding fragment that specifically binds to CD47. In some embodiments, the antibody that specifically binds to CD47 is an isolated monoclonal antibody. In some embodiments, the anti-CD47 agent is an antibody or antigen-binding fragment that specifically binds to SIRPa. In some embodiments, the antibody that specifically binds to SIRPa is an isolated monoclonal antibody. In some embodiments, the antibody promotes macrophage-mediated phagocytosis of a CD47-expressing cell. In some embodiments, the anti-CD47 agent is a soluble CD47 binding SIRPa fragment. In some embodiments, the anti-CD47 agent prevents interaction between CD47 and SIRPa. In some embodiments, the anti-CD47 agent is an IgG isotype selected from the group consisting of IgG 1 isotype, IgG2 isotype, IgG3 isotype, IgG4 isotype, IG-G1-N297Q, IG4-S228P, and IG64 PE. In some embodiments, the cancer is selected from non-Hodgkin lymphoma, acute lymphoblastic leukemia (ALL), T-ALL, B- ALL, acute myelogenous leukemia (AML), B -lymphoblastic leukemia/lymphoma; diffuse large B cell lymphoma (DLBCL); B-cell chronic lymphocytic leukemia/small lymphocytic lymphoma, chronic lymphocytic leukemia (CLL), chronic myelocytic leukemia (CML), Burkitt's lymphoma, follicular lymphoma, SLL, marginal zone lymphoma, CNS lymphoma, Richter's Syndrome, multiple myeloma, myelofibrosis, polycythemia vera, cutaneous T-cell lymphoma, MGUS, myelodysplastic syndrome (MDS), immunoblastic large cell lymphoma, precursor B -lymphoblastic lymphoma and anaplastic large cell lymphoma. In some embodiments, the compound of formula (I) is administered at a dose from about 5 mg/m² to 150 mg/m². In some embodiments, the compound of formula (I) is administered orally, sublingually, buccally, subcutaneously, intravenously, intranasally, intratumorally, topically, transdermally, intradermally, intraperitoneally, intramuscularly, intrapulmonarilly, vaginally, rectally, or intraocularly. In some embodiments, the compound of formula (I) is administered in combination with radiation. In some embodiments, the compound of formula (I) is administered once a week. In some embodiments, the compound of formula (I) is administered once on each of day 1 and day 8 of a three- week (21 day) treatment cycle. In some embodiments, the cancer is non-Hodgkin’s lymphoma.

[0009] In some embodiments, the compound of Formula (I) includes plinabulin, (3Z,6Z)-3-(phenyl-2,3,4,5,6-d5)-methylene-6-((5-(tert-butyl)-lH-imidazol-4- yl)methylene)piperazine-2, 5-dione; (3Z, 6Z)-3-(phenyl-2, 3,4,5, 6-d5)-methylene-d-6-((5-(tert- butyl)-lH-imidazol-4-yl)methylene)piperazine-2, 5-dione; (3Z,6Z)-3-(phenylmethylene-d)-6- ((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(phenyl- 2,3,4,5,6-d5)-methylene-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2,5- dione; (3Z,6Z)-3-(phenylmethylene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene- d)piperazine-2, 5-dione; (3Z,6Z)-3-(phenyl-2,3,4,5,6-d5)-methylene-d-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(4-Fluoro-(phenyl-2,3,5,6-d4))- methylene-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene)piperazine-2, 5-dione; (3Z,6Z)-3-

(4-Fluoro-(phenyl-2,3,5,6-d4))-methylene-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene- d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-fluorobenzylidene)-6-((5-(tert-butyl)-lH-imidazol-4- yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-benzoylbenzylidene)-6-((5-(tert-butyl)- lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-(4- fluorobenzoyl)benzylidene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2,5- dione; (3Z,6Z)-3-(3-(4-methoxybenzoyl)benzylidene)-6-((5-(tert-butyl)-lH-imidazol-4- yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-methoxybenzylidene)-6-((5-(tert-butyl)- lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; or (3Z,6Z)-3-(3-

(trifluoromethyenzydene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2,5- dione.

[0010] Some aspects of the present disclosure provide for a method of treating a subject with cancer. In some embodiments, the method includes administering a therapeutic dose of plinabulin and an anti-CD47 agent. In some embodiments, the anti-CD47 agent is an antibody or antigen-binding fragment that specifically binds to CD47. In some embodiments, the antibody that specifically binds to CD47 is an isolated monoclonal antibody. In some embodiments, the anti-CD47 agent is an antibody or antigen-binding fragment that specifically binds to SIRPa. In some embodiments, the antibody that specifically binds to SIRPa is an isolated monoclonal antibody. In some embodiments, the antibody promotes macrophage-mediated phagocytosis of a CD47-expressing cell. In some embodiments, the anti-CD47 agent is a soluble CD47 binding SIRPa fragment. In some embodiments, the anti-CD47 agent prevents interaction between CD47 and SIRPa. In some embodiments, the anti-CD47 agent is an IgG isotype selected from the group consisting of IgGl isotype, IgG2 isotype, IgG3 isotype, IgG4 isotype, IG-G1-N297Q, IG4-S228P, and IG64 PE. In some embodiments, the cancer is selected from non-Hodgkin lymphoma, acute lymphoblastic leukemia (ALL), T-ALL, B-ALL, acute myelogenous leukemia (AML), B- lymphoblastic leukemia/lymphoma; diffuse large B cell lymphoma (DLBCL); B-cell chronic lymphocytic leukemia/small lymphocytic lymphoma, chronic lymphocytic leukemia (CLL), chronic myelocytic leukemia (CML), Burkitt's lymphoma, follicular lymphoma, SLL, marginal zone lymphoma, CNS lymphoma, Richter's Syndrome, multiple myeloma, myelofibrosis, polycythemia vera, cutaneous T-cell lymphoma, MGUS, myelodysplastic syndrome (MDS), immunoblastic large cell lymphoma, precursor B -lymphoblastic lymphoma and anaplastic large cell lymphoma. In some embodiments, the plinabulin is administered at a dose from about 5 mg/m² to 150 mg/m². In some embodiments, the plinabulin is administered orally, sublingually, buccally, subcutaneously, intravenously, intranasally, intratumorally, topically, transdermally, intradermally, intraperitoneally, intramuscularly, intrapulmonarilly, vaginally, rectally, or intraocularly. In some embodiments, the plinabulin is administered in combination with radiation. In some embodiments, the plinabulin is administered once a week. In some embodiments, the plinabulin is administered once on each of day 1 and day 8 of a three- week (21 day) treatment cycle. In some embodiments, the cancer is non-Hodgkin’ s lymphoma.

[0011] Some aspects of the disclosure provide for a method of treating cancer in a subject in need thereof, wherein the cancer comprises cells that express CD-47. In some embodiments, the method includes administering to the subject an effective amount of one or more anti-CD47 agent and plinabulin. In some embodiments, the one or more anti-CD47 agent is a monoclonal antibody that inhibits the interaction between human CD47 and SIRPa. In some embodiments, the monoclonal antibody is a human antibody. In some embodiments, the monoclonal antibody comprises a human kappa constant region. In some embodiments, the monoclonal antibody is administered prior to the administration of plinabulin. In some embodiments, the monoclonal antibody is administered after the administration of plinabulin. In some embodiments, the monoclonal antibody is an optimized antibody molecule. In some embodiments, the one or more anti-CD47 agent is selected from rituximab or daratumumab. In some embodiments, the cancer is selected from acute lymphoblastic leukemia (ALL), T-ALL, B-ALL, acute myelogenous leukemia (AML), Non-Hodgkin lymphoma, B -lymphoblastic leukemia/lymphoma; diffuse large B cell lymphoma (DLBCL); B-cell chronic lymphocytic leukemia/small lymphocytic lymphoma, chronic lymphocytic leukemia (CLL), chronic myelocytic leukemia (CML), Burkitt's lymphoma, follicular lymphoma, SLL, marginal zone lymphoma, CNS lymphoma, Richter's Syndrome, multiple myeloma, myelofibrosis, polycythemia vera, cutaneous T-cell lymphoma, MGUS, myelodysplastic syndrome (MDS), immunoblastic large cell lymphoma, precursor B -lymphoblastic lymphoma and anaplastic large cell lymphoma. In some embodiments, the cancer is a cancer of a tissue selected from the group consisting of: lung, pancreas, breast, liver, ovary, testicle, kidney, bladder, spine, brain, cervix, endometrium, colon/rectum, anus, esophagus, gallbladder, gastrointestinal tract, skin, prostate, pituitary, stomach, uterus, vagina, and thyroid. In some embodiments, the one or more anti-CD47 agent is administered in combination with a pharmaceutically acceptable carrier or diluent. In some embodiments, the one or more anti-CD47 agent is administered subcutaneously. In some embodiments, the one or more anti-CD47 agent is administered intravenously.

[0012] Some aspects of the disclosure provide for a pharmaceutical composition comprising plinabulin and one or more anti-CD47 agent. In some embodiments, the anti- CD47 agent is an antibody or antigen -binding fragment that specifically binds to CD47. In some embodiments, the antibody that specifically binds to CD47 is an isolated monoclonal antibody. In some embodiments, the anti-CD47 agent is an antibody or antigen-binding fragment that specifically binds to SIRPa. In some embodiments, the antibody that specifically binds to SIRPa is an isolated monoclonal antibody. In some embodiments, the antibody promotes macrophage-mediated phagocytosis of a CD47-expressing cell. In some embodiments, the anti-CD47 agent is a soluble CD-47 binding SIRPa fragment. In some embodiments, the anti-CD47 agent prevents interaction between CD47 and SIRPa. In some embodiments, the anti-CD47 agent is an IgG isotype selected from the group consisting of IgGl isotype, IgG2 isotype, IgG3 isotype, IgG4 isotype, IG-G1-N297Q, IG4-S228P, and IG64 PE. In some embodiments, the plinabulin is in an amount from about 5 mg to 150 mg. [0013] Some aspects of the disclosure provide for a method of treating a subject with a cancer cell expressing CD-47. In some embodiments, the method comprises detecting tumor cell expression of CD-47 in a subject, and providing the subject a therapeutic amount of plinabulin and an anti-CD47 agent. In some embodiments, detecting a tumor cell expression of CD-47 comprises at least one of immunocytochemistry, proteomics, mRNA quantification, or a combination thereof, used to detect tumor cell expression of CD-47 in a subject.

BRIEF DESCRIPTION OF THE DRAWINGS

[0014] The disclosure is best understood from the following detailed description when read in conjunction with the accompanying drawings. It is emphasized that, according to common practice, the various features of the drawings may not be to-scale. On the contrary, the dimensions of the various features may be arbitrarily expanded or reduced for clarity. Included in the drawings are the following figures.

[0015] FIG. 1A illustrates two scatterplot graphs depicting Ml polarization of intratumoral TAM cells from C57BV6 mice treated with plinabulin and M1/M2 ratio (macrophages).

[0016] FIG. IB illustrates a line graph depicting survival of C57BI/6 and Rag -/- mice treated with plinabulin.

[0017] FIG. 2A illustrates bar graphs (left) and histrograms (right) depicting M2 to Ml polarization of macrophages derived from murine tumors.

[0018] FIG. 2B illustrates bar graphs (top) and histograms (bottom) depicting murine bone marrow and human monocytes following ex vivo treatment with plinabulin.

[0019] FIG. 3 illustrates density plots depicting increased phagocytosis following treatment with plinabulin.

[0020] FIG. 4 illustrates line graphs depicting plinabulin induces an increase of Ml markers.

[0021] FIG. 5A illustrates a graph depicting mice bearing MC38 tumors (50-100 mm^) _were i_eft untreated or treated with plinabulin (7 mg/kg) administered peri-tumoral on days 0, 1, 2, 4, 7, 9, 11 post treatment start (first dose administered 18 days post MC38 cell innoculation). [0022] FIG. 5B illustrates a bar graph depicting tumor volume (left) and percentage of live CD45" cells out of total cells (i.e. tumor cells, right) 7 days post plinabulin treatment start in plinabulin-treated or control mice.

[0023] FIG. 5C illustrates a line graph depicting Kaplan-Meier survival to humane endpoint curve of plinabulin-treated vs control mice (log-rank test p-value is shown).

[0024] FIG. 5D illustrates a graph depicting mice bearing MC38 tumors (50-100 mm3) were left untreated or treated with plinabulin (7.5 mg/kg) administered intra-peritoneal twice a day on days 0, 2, 4, post treatment start (first dose administered 18 days post MC38 cell innoculation). Tumors were collected for flow cytometry analyses on day 7 post treatment start.

[0025] FIG. 5E illustrates a bar graph depicting the percentage of intratumoral CD4⁺ (left) and CD8⁺ (right) T cells out of total live CD45⁺ CD3⁺ cells in plinabulin-treated and untreated MC38 tumors.

[0026] FIG. 5F illustrates a bar graph depicting the percentage of intratumoral Tregs (left) and CD8+ T cells to Treg ratio (right) in plinabulin treated and untreated MC38 tumors.

[0027] FIG. 5G illustrates a bar graph depicting the percentage of IFNy⁺CD4⁺ and IFNy " CD8⁺ cells after ex vivo anti-CD3 and anti-CD28 mAb re-stimulation of tumor infiltrating immune cells from plinabulin-treated or untreated MC38 tumors.

[0028] FIG. 5H illustrates a line graph depicting Kaplan-Meier survival curve of plinabulin-treated vs untreated mice at the experiment endpoint in wild type C57BL/6 (WT) or T- and B-cell deficient C57BL/6 RAG ^/_ mice.

[0029] FIG. 51 illustrates a bar graph depicting frequency of CDl lb⁺ CDl lc⁺ positive DCs (of all CD45+ cells, left) and F4/80⁺ Ly6C“ Ly6G“ TAMs (of CDl lb⁺ population, right) in MC38-tumor bearing mice treated or untreated with plinabulin.

[0030] FIG. 5 J illustrates a bar graph depicting M1/M2 ratio (defined as the ratio between CD80+ and CD206+ TAMs) in plinabulin-treated MC38-tumor bearing mice and in untreated animals.

[0031] FIG. 6A illustrates a graph depicting MC38 tumors (400-600 mm3) from C57BL/6 mice were isolated (day -1). Tumor infiltrating TAMs were sorted by FACS and treated with plinabulin or relevant controls (day 0). Macrophage polarization was assessed by flow cytometry (day 2).

[0032] FIG. 6B illustrates Left: Histograms depicting expression of CD80 in murine TAMs treated ex vivo for 48 hours with plinabulin (1000 nM or 200 nM), IL-4 (25 ng/mL) or LPS (20 ng/mL) and IFN-y (50 ng/mL) combination and in untreated control. Right: gMFI of CD80 expression of murine TAMs in different treatment conditions (as indicated) after 48 hours.

[0033] FIG. 6C illustrates density plots depicting CD80 and CD206 expression in murine TAMs treated with plinabulin or in control conditions for 48 hours.

[0034] FIG. 6D illustrates a bar depicting quantification of M1/M2 ratio in murine TAMs treated with plinabulin or in control conditions for 48 hours.

[0035] FIG. 6E illustrates a graph depicting murine BMDMs were generated by culturing murine bine marrow cells with M-CSF (20 ng/mL) and IL-4 (25 ng/mL) for 7 days. BMDMs were treated with plinabulin or control conditions (concentrations as if Fig. 5B) for 2 days prior to assessment with flow cytometry.

[0036] FIG. 6F illustrates bar graphs depicting frequency of CD80+ (left) or CD206+ (right) cells out of F4/80+ BMDMs, treated with plinabulin or control conditions.

[0037] FIG. 6G illustrates a bar graph depicting quantification of M1/M2 ratio in murine BMDMs treated with plinabulin or control conditions for 48 hours.

[0038] FIG. 6H illustrates a graph depicting experimental outline of macrophage generation with M-CSF (50 ng/mL) and IL-4 (25 ng/mL) from healthy donor PBMCs and treatment with plinabulin at indicated doses or controls (LPS at 25 ng/mL and IFN-y at 50 ng/mL).

[0039] FIG. 61 illustrates bar graphs depicting frequency of CD80⁺ (left) or CD206+ (right) cells out of CDl lb⁺ CD 14+ human macrophages, treated with plinabulin or control conditions.

[0040] FIG. 6J illustrates a bar graph depicting quantification of M1/M2 ratio in human CD 14+ derived macrophages treated with plinabulin or control conditions for 48 hours. [0041] FIG. 7A illustrates a graph depicting experimental outline of macrophage generation from healthy donor PBMCs, CTV labelling and treatment with plinabulin or controls prior to analysis by flow cytometry and multiplex cytokine analysis.

[0042] FIG. 7B illustrates histograms depicting CTV expression i.e. proliferation of CD86⁺ (left) or CD206+ (right) human macrophages treated for 48 hours with plinabulin (1000 nM or 200 nM), IL-4 (25 ng/mL), LPS (25 ng/mL), and IFN-y (50 ng/mL), combination or untreated.

[0043] FIG. 7C illustrates bar graphs depicting quantification of CTV signal as gMFI in CD86+ (left) or CD206+ (right) human macrophages treated for 48 hours with plinabulin or control treatments.

[0044] FIG. 7D illustrates bar graphs depicting percentage of Annexin V⁺ cells (left) and gMFI of AnnexinV (right) in human macrophages treated for 48 hours with plinabulin or control conditions.

[0045] FIG. 7E illustrates line graphs depicting quantification of pro- inflammatory cytokinesIL-ip IL-6 and IL12p40 in the supernatant of human macrophages from four healthy donors treated for 0, 24 or 48 hours with plinabulin (top) or LPS (25 ng/mL), and IFN-y (50 ng/mL), combination treatment (bottom).

[0046] FIG. 7F illustrates line graphs depicting quantification of iNOS mRNA expression by qPCR in human macrophages after 4 or 8 hours of treatment with plinabulin or LPS (25 ng/mL), and IFN-y (50 ng/mL), combination.

[0047] FIG. 8A illustrates a graph depicting experimental outline of macrophage generation from healthy donor PBMCs and treatment with plinabulin at indicated doses or controls in the presence of a JNK inhibitor SP600125 (iJNK, 20 |lM).

[0048] FIG. 8B illustrates bar graphs depicting percentage of CD86+ (left) or CD163+ (right) cells out of CDl lb⁺ CD 14+ human macrophages, treated with plinabulin or control conditions in the presence or absence of a JNK inhibitor.

[0049] FIG. 8C illustrates bar graphs depicting CD86+ (left) or CD163+ (right) events out of CDl lb+ CD 14+ human macrophages, treated with plinabulin or control conditions in the presence or absence of a JNK inhibitor, calculated using counting beads on flow cytometry. [0050] FIG. 8D illustrates a bar graph depicting the percentage of live cells (cells negative for the live cell exclusion dye) out of total human macrophages, treated with plinabulin or control conditions, measured by flow cytometry.

[0051] FIG. 9A illustrates a graph depicting experimental outline of macrophage generation from healthy donor PBMCs, treatment with plinabulin or controls and co-culture with CTV-labelled HuT 78 tumor cells for 48 hours.

[0052] FIG. 9B illustrates bar graphs depicting frequency of live HuT 78 tumor cells alone in culture or after co-culture with human macrophages that were pre-treated with plinabulin or control treatments at a 5:1 (left) or 10:1 (right) E:T ratio (E i.e. effector= macrophages; T i.e. tumor cells= HuT 78 tumor cells).

[0053] FIG. 9C illustrates histograms depicting CTV signal i.e. proliferation of HuT 78 tumor cells after co-culture with human macrophages pre-treated with plinabulin or control treatments at a 5:1 (left) or 10:1 (right) E:T ratio. Left: Frequency of Fas⁺ HuT 78 tumor cells after co-culture with human macrophages that were pre-treated with plinabulin or control conditions. Right: Frequency of Fas-L⁺ Ml macrophages (CD86+) treated with plinabulin or control conditions.

[0054] FIG. 9D illustrates bar graphs depicting quantification of CTV signal as gMFI in HuT 78 tumor cells after co-culture with human macrophages pre-treated with plinabulin or control treatments at a 5:1 (left) or 10:1 (right) E:T ratio.

[0055] FIG. 9E illustrates a bar graph depicting an increase in Fas+ tumor cells in co-culture with plinabulin-treated macrophages, compared to untreated cells and positive control.

[0056] FIG. 9F illustrates a bar graph depicting a dose-dependent increase in Fas- L expression on human macrophages treated with plinabulin.

[0057] FIG. 10A illustrates a graph depiciting experimental outline of TAM isolation from ovarian patient tumor digests, CTV labelling and treatment with plinabulin or controls.

[0058] FIG. 10B illustrates bar graphs depicting frequency of CD86+ (left) or CD206+ (right) cells out of CDl lb⁺ CD 14+ human tumor infiltrating macrophages, treated with plinabulin or control conditions for 48 hours. [0059] FIG. 10C illustrates histograms depicting CTV signal i.e. proliferation of human ovarian TAMs treated ex vivo for 48 hours with plinabulin (1000 nM or 200 nM) or control treatments.

[0060] FIG. 10D illustrates bar graphs depicting quantification of CTV signal as gMFI in human ovarian TAMs treated ex vivo for 48 hours with plinabulin (1000 nM or 200 nM) or control treatments.

[0061] FIG. 11A illustrates line graphs depicting EMT-6 tumor growth in untreated (black lines) or plinabulin-treated animals (blue lines; used at the dose of 15 mg/kg) over time.

[0062] FIG. 11B illustrates line graphs depicting Kaplan-Meier survival to humane end-point curve of EMT-6 tumour bearing, plinabulin-treated vs untreated mice. Statistical significance was determined by log-rank Mantel-Cox test with p value indicated on the graph.

[0063] FIG. 11C illustrates bar graphs depicting the percentage of TNFoc⁺CD4⁺ and TNFoc⁺CD8⁺ cells after ex vivo anti-CD3 and anti-CD28 mAb re-stimulation of intratumoral CD4+ and CD8+ T cells from plinabulin-treated or untreated MC38 tumors.

[0064] FIG. 12A illustrates a bar graph depicting the percentage live murine BMDMs following treatment with Plinabulin or controls as per schematic in Fig. 6E.

[0065] FIG. 12B illustrates a bar graph depicting frequency of CD86+ cells out of F4/80+ BMDMs, treated with plinabulin or control treatments.

[0066] FIG. 12C illustrates a bar graph depicting quantification of M1/M2 ratio in BMDMs treated with plinabulin or control treatments.

[0067] FIG. 12D illustrates a bar graph depicting frequency of CD163+ cells out of CD1 lb⁺ CD14+ human macrophages, treated with plinabulin or control treatments.

[0068] FIG. 12E illustrates a line graph depicting frequency of CD86⁺ cells out of CDl lb+ CD14+ human macrophages after 48, 72 or 86 hours of treatment with plinabulin or control treatments.

[0069] FIG. 12F illustrates a line graph depicting frequency of CD206+ cells out of CDl lb⁺ CD14+ human macrophages after 48, 72 or 86 hours of treatment with plinabulin or control treatments. [0070] FIG. 13 A illustrates a line graph depicting quantification of IL lb mRNA expression by qPCR in human macrophages after 6 or 18 hours of treatment with plinabulin or LPS and IFN-y combination.

[0071] FIG. 13B illustrates a line graph depicting Quantification of Tgfb, Egr2, 114 and Ccll7 mRNA expression by qPCR in human macrophages after 4, 8 or 20 hours of treatment with plinabulin or LPS and IFN-y combination.

[0072] FIG. 14 illustrates bar graphs depicting the frequency of plinabulin or control-treated live human macrophages after co-culture with Fas⁺ HuT 78 tumor cells at 5:1 (left) or 10:1 (right) E:T ratio.

[0073] FIG. 15 illustrates bar graphs depicting frequency of CD80+ and CD163+ cells in TAMs isolated from untreated human ovarian tumor digest.

[0074] FIG. 16 illustrate a bar graph depicting the percentage of phagocytosis index of Plinabulin.

[0075] FIG. 17 illustrate a bar graph depicting the percentage of phagocytosis index of anti-CD47.

DETAILED DESCRIPTION

[0076] Methods and compositions provided herein are useful in treating, delaying the progression of, preventing relapse of or alleviating a symptom of a cancer or other neoplastic condition using a synergistic combination of agents targeting CD47 and a compound of Formula (I). In some embodiments, the compound of Formula (I) is plinabulin. Plinabulin, (3Z,6Z)-3-Benzylidene-6-{ [5-(2-methyl-2-propanyl)-l/Z-imidazol-4- yl]methylene]-2,5-piperazinedione, is a synthetic analog of the natural compound phenylahistin. Plinabulin can be readily prepared according to methods and procedures detailed in U.S. Patent Nos. 7,064,201 and 7,919,497, which are incorporated herein by reference in their entireties. While not being bound by any particular theory, it was surprisingly discovered that plinabulin enhances polarization of TAMs from M2 to Ml phenotype. This effect permits plinabulin to work synergistically with agents targeting CD47. In some aspects, the agents targeting CD47 include one or more monoclonal antibody or an antigen-binding fragment. In some embodiments, the one or more monoclonal antibody or the antigen-binding fragment are capable of modulating, e.g., blocking, inhibiting, reducing, antagonizing, neutralizing or otherwise interfering with CD47 expression, activity and/or signaling. In some embodiments, the one or more anti-CD47 agent is an antibody or antigen-binding fragment that specifically binds to CD47.

[0077] Before the present disclosure is further described, it is to be understood that this invention is not limited to particular embodiments described, as such may, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting, since the scope of the present invention will be limited only by the appended claims.

[0078] Where a range of values is provided, it is understood that each intervening value, to the tenth of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limit of that range and any other stated or intervening value in that stated range, is encompassed within the invention. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges and are also encompassed within the invention, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the invention.

[0079] Methods recited herein may be carried out in any order of the recited events which is logically possible, as well as the recited order of events.

[0080] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present invention, the preferred methods and materials are now described.

[0081] All publications mentioned herein are incorporated herein by reference to disclose and describe the methods and/or materials in connection with which the publications are cited.

[0082] It must be noted that as used herein and in the appended claims, the singular forms “a”, “an”, and “the” include plural referents unless the context clearly dictates otherwise. It is further noted that the claims may be drafted to exclude any optional element. As such, this statement is intended to serve as antecedent basis for use of such exclusive terminology as “solely,” “only” and the like in connection with the recitation of claim elements, or use of a “negative” limitation.

[0083] The publications discussed herein are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the present invention is not entitled to antedate such publication by virtue of prior invention. Further, the dates of publication provided may be different from the actual publication dates which may need to be independently confirmed.

Definitions

[0084] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of ordinary skill in the art to which this disclosure belongs. All patents, applications, published applications, and other publications are incorporated by reference in their entirety. In the event that there is a plurality of definitions for a term herein, those in this section prevail unless stated otherwise.

[0085] The term “agent” is used herein to denote a chemical compound, a mixture of chemical compounds, a biological macromolecule, or an extract made from biological materials.

[0086] The term “antibody” or “antibody moiety” is intended to include any polypeptide chain-containing molecular structure with a specific shape that fits to and recognizes an epitope, where one or more non-covalent binding interactions stabilize the complex between the molecular structure and the epitope. Antibodies utilized in the present disclosure may be polyclonal antibodies or monoclonal antibodies. Antibodies also include free antibodies and antigen binding fragments derived therefrom, and conjugates, e.g. pegylated antibodies, drug, radioisotope, or toxin conjugates, and the like. Monoclonal antibodies directed against a specific epitope, or combination of epitopes, will allow for the targeting and/or depletion of cellular populations expressing the marker. Various techniques can be utilized using monoclonal antibodies to screen for cellular populations expressing the marker(s), and include magnetic separation using antibody-coated magnetic beads, “panning” with antibody attached to a solid matrix (i.e., plate), and flow cytometry (See, e.g., U.S. Pat. No. 5,985,660; and Morrison et al. Cell, 96:737-49 (1999)). These techniques allow for the screening of particular populations of cells; in immunohistochemistry of biopsy samples; in detecting the presence of markers shed by cancer cells into the blood and other biologic fluids, and the like. Humanized versions of such antibodies are also within the scope of this disclosure. Humanized antibodies are especially useful for in vivo applications in humans due to their low antigenicity.

[0087] The terms “cancer”, “neoplasm”, “tumor”, and “carcinoma”, are used interchangeably herein to refer to cells which exhibit relatively autonomous growth, so that they exhibit an aberrant growth phenotype characterized by a significant loss of control of cell proliferation. In general, cells of interest for detection or treatment in the present application include precancerous (e.g., benign), malignant, pre-metastatic, metastatic, and non-metastatic cells. Detection of cancerous cells is of particular interest.

[0088] The term “CD47” as used herein refers a protein encoded by the CD47 gene. CD47 is overexpressed on the surface of many types of cancer cells. CD47 forms a signaling complex with signal-regulatory protein a (SIRPa), enabling the escape of these cancer cells from macrophage-mediated phagocytosis.

[0089] As used herein, “CD47 polypeptides” denotes the three transcript variants of human CD47 (variant 1, NM 001777; variant 2, NM 198793; and variant 3, NM 001025079) encode three isoforms of CD47 polypeptide. CD47 isoform 1 (NP 001768), the longest of the three isoforms, is 323 amino acids long. CD47 isoform 2 (NP 942088) is 305 amino acid long. CD47 isoform 3 is 312 amino acids long. The three isoforms are identical in sequence in the first 303 amino acids. Amino acids 1-8 comprise the signal sequence, amino acids 9-142 comprise the CD47 immunoglobulin like domain, which is the soluble fragment, and amino acids 143-300 is the transmembrane domain.

[0090] The first 142 amino acids of CD47 polypeptide comprise the extracellular region of CD47 (SEQ ID NO: 1). The three isoforms have identical amino acid sequence in the extracellular region, and thus any of the isoforms are can be used to generate soluble CD47. “Soluble CD47” is a CD47 protein that lacks the transmembrane domain. Soluble CD47 is secreted out of the cell expressing it, instead of being localized at the cell surface.

[0091] As used herein, a “fusion” polypeptide denotes a polypeptide comprising a polypeptide or portion (e.g., one or more domains) thereof fused or bonded to heterologous polypeptide. A fusion soluble CD47 protein, for example, will share at least one biological property in common with a native sequence soluble CD47 polypeptide. Examples of fusion polypeptides include immunoadhesins, as described above, which combine a portion of the CD47 polypeptide with an immunoglobulin sequence, and epitope tagged polypeptides, which comprise a soluble CD47 polypeptide or portion thereof fused to a “tag polypeptide”. The tag polypeptide has enough residues to provide an epitope against which an antibody can be made, yet is short enough such that it does not interfere with biological activity of the CD47 polypeptide. Suitable tag polypeptides generally have at least six amino acid residues and usually between about 6-60 amino acid residues.

[0092] 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 Kd. (See, generally, Davies et al. (1990) Annual Rev Biochem 59:439-473). An antibody of the present invention is said to specifically bind to CD47, when the equilibrium binding constant (Kd) is ^1 pM, preferably ^100 nM, more preferably ^10 nM, and most preferably ^100 pM to about 1 pM, as measured by assays such as radioligand binding assays, surface plasmon resonance (SPR), flow cytometry binding assay, or similar assays known to those skilled in the art.

[0093] 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 which 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.

[0094] 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 in 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.

[0095] 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.

[0096] The terms “immunospecifically binds,” “immunospecifically recognizes,” “specifically binds,” and “specifically recognizes” are analogous terms in the context of antibodies and refer to molecules that bind to an antigen/epitope as such binding is understood by one skilled in the art. For example, a molecule (e.g., an antibody) that specifically binds to an antigen may bind to other peptides or polypeptides, generally with lower affinity as determined by, e.g., immunoassays, surface plasmon resonance assays, for example, Biacore™ KinExA platform (Sapidyne Instruments, Boise, Id.), or other assays known in the art. In a specific embodiment, molecules that specifically bind to an antigen bind to the antigen with a K_athat is at least 2 logs, 2.5 logs, 3 logs, 4 logs or greater than the Kawhen the molecules bind to another antigen. In another specific embodiment, molecules that specifically bind to an antigen do not cross react with other proteins. In another specific embodiment, molecules that specifically bind to an antigen do not cross react with other non- CD47 proteins.

[0097] The term “monoclonal antibody” as used herein is a well-known term of art that refers to an antibody obtained from a population of homogenous or substantially homogeneous antibodies. The term “monoclonal” is not limited to any particular method for making the antibody. Generally, a population of monoclonal antibodies can be generated by cells, a population of cells, or a cell line. In specific embodiments, a “monoclonal antibody,” as used herein, is an antibody produced by a single cell or cell line wherein the antibody immunospecifically binds to a CD47 epitope as determined, e.g., by ELISA or other antigenbinding or competitive binding assay known in the art or in the Examples provided herein. In particular embodiments, a monoclonal antibody can be a chimeric antibody or a humanized antibody. In certain embodiments, a monoclonal antibody is a monovalent antibody or multivalent (e.g., bivalent) antibody.

[0098] The term “non-natural amino acid” as used herein refers to an amino acid that is not a proteinogenic amino acid, or a post-translationally modified variant thereof. In particular, the term refers to an amino acid that is not one of the 20 common amino acids or pyrrolysine or selenocysteine, or post-translationally modified variants thereof.

[0099] The term “polyclonal antibodies” as used herein refers to an antibody population that includes a variety of different antibodies that immunospecifically bind to the same and/or to different epitopes within an antigen or antigens.

[0100] The terms “variable region” or “variable domain” as used herein refer to a portion of an antibody, generally, a portion of an antibody light or heavy chain, typically about the amino-terminal 110 to 120 amino acids in a mature heavy chain and about the amino-terminal 90 to 100 amino acids in a mature light chain. Variable regions comprise complementarity determining regions (CDRs) flanked by framework regions (FRs). Generally, the spacial orientation of CDRs and FRs are as follows, in an N-terminal to C- terminal direction: FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4. Without wishing to be bound by any particular mechanism or theory, it is believed that the CDRs of the light and heavy chains are primarily responsible for the interaction of the antibody with antigen and for the specificity of the antibody for an epitope. In a specific embodiment, numbering of amino acid positions of antibodies described herein is according to the EU Index, as in Kabat et al. (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, U.S. Department of Health and Human Services, NIH Publication No. 91-3242. In certain embodiments, the variable region is a human variable region. In certain embodiments, the variable region comprises murine (e.g., mouse or rat) CDRs and human framework regions (FRs). In particular embodiments, the variable region is a primate (e.g., human or non-human primate) variable region. In certain embodiments, the variable region comprises murine (e.g., mouse or rat) CDRs and primate (e.g., human or non-human primate) framework regions (FRs). As a non-limiting example, a variable region described herein is obtained from assembling two or more fragments of human sequences into a composite human sequence. [0101] The term “pharmaceutically acceptable carrier” or “pharmaceutically acceptable excipient” includes any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents and the like. 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 ingredient, its use in the therapeutic compositions is contemplated. In addition, various adjuvants such as are commonly used in the art may be included. Considerations for the inclusion of various components in pharmaceutical compositions are described, e.g., in Gilman et al. (Eds.) (1990); Goodman and Gilman’s: The Pharmacological Basis of Therapeutics, 8th Ed., Pergamon Press, which is incorporated herein by reference in its entirety. The pharmaceutically acceptable excipient can be a monosaccharide or monosaccharide derivative.

[0102] The term “subject” as used herein, means a human or a non-human mammal, e.g., a dog, a cat, a mouse, a rat, a cow, a sheep, a pig, a goat, a non-human primate or a bird, e.g., a chicken, as well as any other vertebrate or invertebrate.

[0103] The term “mammal” is used in its usual biological sense. Thus, it specifically includes, but is not limited to, primates, including simians (chimpanzees, apes, monkeys) and humans, cattle, horses, sheep, goats, swine, rabbits, dogs, cats, rodents, rats, mice, guinea pigs, or the like.

[0104] The terms “effective amount” or a “therapeutically effective amount” as used herein refers to an amount of a therapeutic agent that is effective to relieve, to some extent, or to reduce the likelihood of onset of, one or more of the symptoms of a disease or condition, and can include curing a disease or condition.

[0105] The terms “treat,” “treatment,” or “treating,” as used herein refers to administering a compound or pharmaceutical composition to a subject for prophylactic and/or therapeutic purposes. The term “prophylactic treatment” refers to treating a subject who does not yet exhibit symptoms of a disease or condition, but who is susceptible to, or otherwise at risk of, a particular disease or condition, whereby the treatment reduces the likelihood that the patient will develop the disease or condition. The term “therapeutic treatment” refers to administering treatment to a subject already suffering from a disease or condition. [0106] As used herein, the term “chemotherapeutic agent” refers to an agent that reduces, prevents, mitigates, limits, and/or delays the growth of metastases or neoplasms, or kills neoplastic cells directly by necrosis or apoptosis of neoplasms or any other mechanism, or that can be otherwise used, in a pharmaceutically-effective amount, to reduce, prevent, mitigate, limit, and/or delay the growth of metastases or neoplasms in a subject with neoplastic disease. Chemotherapeutic agents include but are not limited to, for example, fluoropyrimidines; pyrimidine nucleosides; purine nucleosides; anti-folates, platinum-based agents; anthracyclines/anthracenediones; epipodophyllotoxins; camptothecins; hormones; hormonal complexes; antihormonals; enzymes, proteins, peptides and polyclonal and/or monoclonal antibodies; vinca alkaloids; taxanes; epothilones; antimicrotubule agents; alkylating agents; antimetabolites; topoisomerase inhibitors; antivirals; and various other cytotoxic and cytostatic agents.

Compounds

[0107] In some embodiments, the compounds described herein are a dehydrophenylahistin represented by Formula (I):

[0108] In some embodiments of Formula (I), Ri, R4, and Re, are each separately selected from the group consisting of a hydrogen atom, a deuterium atom, a halogen atom, and saturated Ci,-C24 alkyl, unsaturated C1-C24 alkenyl, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, azido, substituted nitro, phenyl, and substituted phenyl groups, hydroxy, carboxy, — CO — O — R7, cyano, alkylthio, halogenated alkyl including polyhalogenated alkyl, halogenated carbonyl, and carbonyl — CH2CO — R7, wherein R7 is selected from a hydrogen atom, a halogen atom, and saturated C1-C24 alkyl, unsaturated C1-C24 alkenyl, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, azido, substituted nitro, phenyl, and substituted phenyl groups.

[0109] In some embodiments of Formula (I), Ri' and Ri" are each independently selected from the group consisting of a hydrogen atom, a deuterium atom, a halogen atom, and saturated C1-C24 alkyl, unsaturated C1-C24 alkenyl, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, azido, substituted nitro, phenyl, and substituted phenyl groups, hydroxy, carboxy, — CO — O — R7, cyano, alkylthio, halogenated alkyl including polyhalogenated alkyl, halogenated carbonyl, and carbonyl — CH2CO — R7, wherein R? is selected from a hydrogen atom, a halogen atom, and saturated C1-C24 alkyl, unsaturated C1-C24 alkenyl, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, azido, substituted nitro, phenyl, and substituted phenyl groups.

[0110] In some embodiments of Formula (I), R, Ri' and Ri" are either covalently bound to one another or are not covalently bound to one another;

[0111] In some embodiments of Formula (I), R2, R3, and R5 are each separately selected from the group consisting of a hydrogen atom, a deuterium atom, a halogen atom, and saturated C1-C12 alkyl, unsaturated Ci-C 12 alkenyl, acyl, cycloalkyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, and substituted nitro groups, sulfonyl and substituted sulfonyl groups.

[0112] In some embodiments of Formula (I), m is an integer equal to zero, one or two.

[0113] In some embodiments of Formula (I), Xi and X2 are separately selected from the group consisting of an oxygen atom, a nitrogen atom and a sulfur atom.

[0114] In some embodiments of Formula (I), Y is selected from the group consisting of a nitrogen atom, a substituted nitrogen atom with a R5 group from above, an oxygen atom, a sulfur atom, a oxidized sulfur atom, a methylene group and a substituted methylene group. [0115] In some embodiments of Formula (I), Z, for each separate n, if non-zero, and Zi, Z2, Za and Z4 are each separately selected from a carbon atom, a sulfur atom, a nitrogen atom or an oxygen atom; and the dashed bonds may be either single or double bonds.

[0116] In some embodiments, the compounds described herein are a dehydrophenylahistin represented by Formula (II):

[0117] In some embodiments of Formula (II), R2 and Ra arc each separately selected from the group consisting of a hydrogen atom; a halogen atom; mono-substituted; poly-substituted or unsubstituted, straight or branched chain variants of the following residues: C1-C12 alkyl, Ci-C 12 alkenyl, acyl, and alkoxy; and mono-substituted, polysubstituted or unsubstituted variants of the following residues: cycloalkyl, cycloalkoxy, aryl, heteroaryl, amino, nitro, and sulfonyl; or R2 is a bond to Ar.

[0118] In some embodiments of Formula (II), R4 and Re are each separately selected from the group consisting of hydrogen; halogen; hydroxyl; mono-substituted, polysubstituted or unsubstituted, straight or branched chain variants of the following residues: Ci- C24 alkyl, C2-C24 alkenyl, C2-C24 alkynyl, alkoxy, acyl, arylalkyl, heteroarylalkyl, alkyloxycarbonyloxy, ester, arylalkoxy, alkoxy, and alkylthio; mono-substituted, polysubstituted or unsubstituted variants of the following residues: acyloxy, aryloxycarbonyloxy, cycloalkyl, cycloalkenyl, cycloalkoxy, aryl, heteroaryl, aryloxy, arylcarbonyl, heterocycloalkyl, carbonyl, amino, aminocarbonyl, amide, aminocarbonyloxy, nitro, azido, phenyl, hydroxyl, thio, alkylthio, arylthio, thiooxysulfonyl, thiophene, carboxy, and cyano.

[0119] In some embodiments of Formula (II), Xi and X2 are separately selected from the group consisting of an oxygen atom, a sulfur atom, and a nitrogen atom substituted with a R5 group; [0120] In some embodiments of Formula (II), Rs is selected from the group consisting of a hydrogen atom, a halogen atom, and saturated C1-C12 alkyl, unsaturated Ci- C12 alkenyl, acyl, cycloalkyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, and substituted nitro groups, sulfonyl and substituted sulfonyl groups.

[0121] In some embodiments of Formula (II), Y is selected from the group consisting of a nitrogen atom substituted with R5, an oxygen atom, a sulfur atom, a oxidized sulfur atom, a methylene group, and a substituted methylene group.

[0122] In some embodiments of Formula (II), n is 0, 1, 2, 3, or 4.

[0123] In some embodiments of Formula (II), Ar is a cyclic or polycyclic aryl or heteroaryl ring system comprising between one and three rings. In some embodiments, each ring in said system is separately a 5, 6, 7, or 8 membered ring. In some embodiments, each ring in said system separately comprises 0, 1, 2, 3, or 4 heteroatoms selected from the group consisting of oxygen, sulfur, and nitrogen. In some embodiments, each ring in the system is optionally substituted with one or more subtituents selected from the group consisting of hydrogen; halogen; hydroxyl; mono-substituted, poly-substituted or unsubstituted, straight or branched chain variants of the following residues: C1-C24 alkyl, C2-C24 alkenyl, C2- C24 alkynyl, alkoxy, acyl, arylalkyl, heteroarylalkyl, alkyloxycarbonyloxy, ester, arylalkoxy, alkoxy, and alkylthio; mono-substituted, poly-substituted or unsubstituted variants of the following residues: acyloxy, aryloxycarbonyloxy, cycloalkyl, cycloalkenyl, cycloalkoxy, aryl, heteroaryl, aryloxy, arylcarbonyl, heterocycloalkyl, carbonyl, amino, aminocarbonyl, amide, aminocarbonyloxy, nitro, azido, phenyl, hydroxyl, thio, alkylthio, arylthio, thiophene, oxysulfonyl, sulfonyl, carboxy, and cyano; and an optionally substituted fused ring selected from the group consisting of dioxole, dithiole, oxathiole, dioxine, dithiine, and oxathiine.

[0124] In some embodiments, a compound of Formula (I) is selected from plinabulin, (3Z,6Z)-3-(phenyl-2,3,4,5,6-d5)-methylene-6-((5-(tert-butyl)-lH-imidazol-4- yl)methylene)piperazine-2, 5-dione; (3Z, 6Z)-3-(phenyl-2, 3,4,5, 6-d5)-methylene-d-6-((5-(tert- butyl)-lH-imidazol-4-yl)methylene)piperazine-2, 5-dione; (3Z,6Z)-3-(phenylmethylene-d)-6- ((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(phenyl- 2,3,4,5,6-d5)-methylene-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2,5- dione; (3Z,6Z)-3-(phenylmethylene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene- d)piperazine-2, 5-dione; (3Z,6Z)-3-(phenyl-2,3,4,5,6-d5)-methylene-d-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(4-Fluoro-(phenyl-2,3,5,6-d4))- methylene-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene)piperazine-2, 5-dione; (3Z,6Z)-3-

(trifluoromethyenzydene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2,5- dione. In some embodiments, a compound of Formula (I) is plinabulin.

[0125] The compounds described above may be made using methods described in U.S. Patent No. 7,919,497, which is incorporated herein by reference in its entirety.

Anti-CD47 Antibodies

[0126] In aspects, provided herein are antibodies which specifically bind to CD47. In some embodiments, the CD47 antibodies comprise modifications in one or more amino acid residues. For example in the heavy chain variable region. In some embodiments, the CD47 antibodies inhibit SIRPa interaction with CD47. In some embodiments, the CD47 antibodies have low or no Fc effector function. Antibodies can include, for example, monoclonal antibodies, recombinantly produced antibodies, monospecific antibodies, multispecific antibodies (including bispecific antibodies), human antibodies, humanized antibodies, murine antibodies (e.g., mouse or rat antibodies), chimeric antibodies, synthetic antibodies, and tetrameric antibodies comprising two heavy chain and two light chain molecules. In specific embodiments, antibodies can include, but are not limited to an antibody light chain monomer, an antibody heavy chain monomer, an antibody light chain dimer, an antibody heavy chain dimer, an antibody light chain- antibody heavy chain pair, intrabodies, heteroconjugate antibodies, single domain antibodies, and monovalent antibodies. In a specific embodiment, antibodies can include antigen-binding fragments or epitope binding fragments such as, but not limited to, single chain antibodies or single-chain Fvs (scFv) (e.g., including monospecific, bispecific, etc.), camelized antibodies, affybodies, Fab fragments, F(ab') fragments, F(ab')2 fragments, and disulfide-linked Fvs (sdFv). In certain embodiments, antibodies described herein refer to polyclonal antibody populations. In some embodiments, an anti-CD47 antibody or antibodies is an anti-CD47 agent. In some embodiments, an anti-CD47 agent does not include an anti-CD47 antibody or antibodies.

[0127] Antibodies can be of any type (e.g., IgG, IgE, IgM, IgD, IgA or IgY), any class, (e.g., IgGl, IgG2, IgG3, IgG4, IgAl or IgA2), or any subclass (e.g., IgG2a or IgG2b) of immunoglobulin molecule. In certain embodiments, antibodies described herein are IgG antibodies, or a class (e.g., human IgGl, IgG2, IgG3 or IgG4) or subclass thereof. In certain embodiments, antibodies described herein are IgGl antibodies (e.g., human IgGl) or a subclass thereof. In certain embodiments, IgGl antibodies described herein comprise one or more amino acid substitutions and/or deletions in the constant region. In certain embodiments, antibodies described herein are IgG4 antibodies (e.g., human IgG4) or a subclass thereof. In certain embodiments, IgG4 antibodies described herein comprise one or more amino acid substitutions and/or deletions in the constant region.

[0128] In some embodiments, the CD47 antibodies of the disclosure are monoclonal antibodies. Monoclonal antibodies that modulate, block, inhibit, reduce, antagonize, neutralize or otherwise interfere with CD47- and/or CD47/SIRPa-mediated cell signaling are generated, e.g., by immunizing an animal with membrane bound and/or soluble CD47, such as, for example, human CD47 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 CD47 such that CD47 is expressed and associated with the surface of the transfected cells. Alternatively, the antibodies are obtained by screening a library that contains antibody or antigen binding domain sequences for binding to CD47. 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 the phage particles (i.e., “phage displayed library”). Hybridomas resulting from myeloma/B cell fusions are then screened for reactivity to CD47. [0129] In some embodiments, the monoclonal antibodies of the disclosure that bind CD47, as well as fragments of these monospecific antibodies that are immunologically active and still bind CD47, include the exemplary antibodies described herein, e.g., the 5A3 antibody, the 5A3M4 antibody, the 5A3M3 antibody, the 5A3M5 antibody, the KE8 antibody, the KE8-P6H5 antibody (also referred to herein as KE8H5), the KE8-P3B2 antibody (also referred to herein as KE8B2), the KE8-P2A2 antibody (also referred to herein as KE8A25), the KE8F2 antibody, the KE8G2 antibody, the KE84G9 antibody, the KE81G9 antibody, the KE81A3 antibody, the KE8E8 antibody, the KE8G6 antibody, the KE8H3 antibody, the KE8C7 antibody, the KE8A4 antibody, the KE8A8 antibody, the KE8G11 antibody, the KE8B7 antibody, the KE8F1 antibody, the KE8C4 antibody, the KE8A3 antibody, the KE86G9 antibody, the KE8H6 antibody, the KA3 antibody, the KA3-P5G2 antibody (also referred to herein as KA3G2), the KA3-P1A3 antibody (also referred to herein as KA3A3), the KA3-P5C5 antibody (also referred to herein as KA3C5), the KA3H8 antibody, the KA3B2 antibody, the KA3A2 antibody, the KA3D3 antibody, the KA3H3 antibody, the KC4 antibody, the KC4-P1G11KC4-P4C11 antibody, the KC4-P6B1KC4- P4F4 antibody, and the KC4-P2E2 antibody (also referred to herein as KC4E2), the KC4 antibody, the KC4F4 antibody, the KC4A1 antibody, the KC4C11 antibody, the KC4E10 antibody, the KC4B1 antibody, the KC4C3 antibody, the KC4A4 antibody, the KC4G11 antibody, the KC4G9 antibody and fragments thereof.

[0130] In some embodiments, the one or more anti-CD47 agent includes isolated bispecific antibodies having a first arm that includes a first amino acid sequence that binds CD47 and a second arm that includes a second amino acid sequence that does not bind CD47, wherein the bispecific antibody inhibits interaction between CD47 and signal- regulatory protein alpha (SIRPa). In some embodiments, the second amino acid sequence binds a tumor associated antigen (TAA). In some embodiments, the bispecific antibody inhibits interaction between human CD47 and human SIRPa.

[0131] In some embodiments, the one or more anti-CD47 agents may include one or more bispecific antibodies. The bispecific antibodies of the disclosure allow for simultaneous binding of the two antibody arms to two antigens on the surface of the cell (termed co -engagement), which results in additive or synergistic increase of affinity due to avidity mechanism. As a consequence, co-engagement confers high selectivity towards cells expressing both antigens as compared to cells that express just one single antigen. In addition, the affinities of the two arms of a bispecific antibody to their respective targets can be set up in a way that binding to target cells is principally driven by one of the antibody arms. In some embodiments, the bispecific antibody includes a first arm that binds CD47 and a second arm that binds a tumor associated antigen (TAA), where the second arm binds to the TAA with high affinity, and the first arm binds to CD47 with low affinity, i.e., an affinity that is sufficient to inhibit CD47/SIRPa upon TAA co-engagement. This design allows the bispecific antibodies of the invention to preferentially inhibit CD47 in cancer versus normal cells. In the examples provided herein, a bispecific antibody with a first arm that binds CD47 with low affinity and a second arm that binds CD 19 with high affinity (termed a CD47xCD19 bispecific) allow preferential inhibition of CD47 in cancer versus normal cells. Besides the two antigen-binding arms, the CD47 x TAA bispecific antibody requires a functional Fc portion to recruit macrophages and/or other immune effector cells. A fully human bispecific IgG format (such as the KA-body format described herein) is well suited for the generation of dual targeting CD47 x TAA bispecific antibodies. As shown in the examples provided herein, the ability of dual targeting bispecific antibodies to coengage CD47 and CD 19 results in a significant increase in the affinity of binding to CD 19- positive cells and in CD19-dependent neutralization of the CD47-SIRPa interaction. In some embodiments, the bispecific antibody inhibits interaction between human CD47 and human SIRPa at a level that is at least ten times more potent than a corresponding level of inhibition of human CD47/human SIRPa interaction exhibited by a monovalent anti-CD47 antibody that includes the first amino acid sequence that binds CD47 and a second amino acid sequence that does not bind a human protein.

[0132] In some embodiments, the bispecific antibody inhibits interaction between human CD47 and human SIRPa at a level that is at least 100 times more potent than a corresponding level of inhibition of human CD47/human SIRPa interaction exhibited by a monovalent anti-CD47 antibody that includes the first amino acid sequence that binds CD47 and a second amino acid sequence that does not bind a human protein.

[0133] In some embodiments, the bispecific antibody inhibits interaction between human CD47 and human SIRPa at a level that is at least 1,000 times more potent than a corresponding level of inhibition of human CD47/human SIRPa interaction exhibited by a monovalent anti-CD47 antibody that includes the first amino acid sequence that binds CD47 and a second amino acid sequence that does not bind a human protein.

[0134] In some embodiments, the antibody comprises light chain and heavy chain segments. With respect to the light chain, in a specific embodiment, the light chain of an antibody described herein is a kappa (K) light chain. In another specific embodiment, the light chain of an antibody described herein is a lambda ( ) light chain. In another embodiment, light chain is a mixed sequence, e.g., the variable portion of the light chain comprises kappa light chain sequences and the constant region of the light chain comprises lambda light chain sequences, or vice versa. In certain embodiments, the light chain of an antibody described herein is a human kappa light chain or a human lambda light chain. Non-limiting examples of human constant region sequences have been described in the art, e.g., see U.S. Pat. No. 5,693,780 and Kabat et al. (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, U.S. Department of Health and Human Services, NIH Publication No. 91-3242. In some embodiments, the bispecific antibody includes two copies of a single heavy chain polypeptide and a first light chain and a second light chain, wherein the first and second light chains are different.

[0135] In some embodiments, the an antibody, e.g. a monoclonal antibody, which specifically binds to human CD47, wherein such an anti-CD47 antibody is a variant of a parental anti-CD47 antibody, wherein the anti-CD47 antibody, when produced using a cell- free (CF) expression system, has a higher antibody expression titer or yield compared to that of the parental anti-CD47 antibody when expressed in the CF system, and wherein the anti- CD47 antibody comprises one or more amino acid modifications, for example, 1-15 amino acid modifications, relative to the parental anti-CD47 antibody. In some embodiments, one or more amino acid modifications are within the heavy chain or VH. In some embodiments, the anti-CD47 antibody provided herein may be a variant of a parental anti-CD47 antibody comprising one or more CDRs of the parental anti-CD47 antibody.

[0136] In a specific aspect, provided herein is an antibody, e.g. a monoclonal antibody, which specifically binds to human CD47, wherein such an anti-CD47 antibody is a variant of a parental anti-CD47 antibody, wherein the anti-CD47 antibody, when produced using a cell-free (CF) expression system, has a higher antibody expression titer or yield compared to that of the parental anti-CD47 antibody when expressed in the CF system, and wherein the anti-CD47 antibody comprising one or more amino acid modifications, for example, 1-15 amino acid modifications, relative to the parental anti-CD47 antibody. In a particular aspect, the one or more amino acid modifications, for example, 5 or 14 amino acid modifications, are within the heavy chain or VH. In a particular aspect, the one or more amino acid modifications, for example, 5, 10, 13 or 14 amino acid modifications, are within the framework region of a VH. In a particular aspect, the one or more amino acid modifications, for example, 5, 13 or 14 amino acid modifications are within the framework region of a VH (e.g., SEQ ID NO: 1). In a certain aspect, the anti-CD47 antibody provided herein which is a variant of a parental anti-CD47 antibody comprising the CDRs (e.g., Kabat CDRs) of the parental anti-CD47 antibody. In certain aspects, such anti-CD47 antibody is an IgGl, IgG2, IgG3, or IgG4 isotype antibody. In certain aspects, such anti-CD47 antibody is an IgGl isotype antibody. In certain aspects, such anti-CD47 antibody is an IgGl Z allotype isotype antibody. In certain aspects, such anti-CD47 antibody is an IgG4, such as an IgG4P or IgG4PE, isotype antibody.

[0137] In aspects provided herein, is an antibody, e.g. a monoclonal antibody, which specifically binds to human CD47, wherein such an anti-CD47 antibody is a variant of a parental anti-CD47 antibody, wherein the anti-CD47 antibody, when produced using a cell- free (CF) expression system, has a higher antibody expression titer or yield compared to that of the parental anti-CD47 antibody when expressed in the CF system. In specific embodiments, the parental anti-CD47 antibody is antibody AB6.12 (see, e.g., U.S. Application Publication No. US 2014/0140989 Al or U.S. Application Publication No. US 2021/0054070, which is incorporated herein by reference in its entirety). The amino acid sequences of the heavy chain variable region (VH) and light chain variable region (VU) of antibody AB6.12 are provided below, wherein the Kabat CDRs are underlined. In a certain aspect, the anti-CD47 antibody provided herein is a variant of parental antibody AB6.12, and comprises the CDRs (e.g., Kabat CDRs) of parental antibody AB6.12. In certain aspects, such anti-CD47 antibody is an IgGl, IgG2, IgG3, or IgG4 isotype antibody. In certain aspects, such anti-CD47 antibody is an IgGl isotype antibody. In certain aspects, such anti- CD47 antibody is an IgGl Z allotype isotype antibody. In certain aspects, such anti- CD47 antibody is an IgG4, such as an IgG4P or IgG4PE, isotype antibody. [0138] In some embodiments, the monoclonal antibodies may include fully human antibodies or humanized antibodies. In some embodiments, the monoclonal antibodies are suitable for administration to humans without engendering an immune response by the human against the administered immunoglobulin.

[0139] Monoclonal antibodies can also be made by recombinant DNA methods, such as those described in U.S. Pat. No. 4,816,567, which is incorporated herein by reference in its entirety. 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).

[0140] Monoclonal antibodies of the disclosure may have the ability to bind CD47, to inhibit the binding of SIRPa to CD47, decrease CD47-SIRPa-mediated signaling, promote phagocytosis, and to inhibit tumor growth and/or migration.

[0141] Exemplary antibodies of the invention include the 2A1 antibody, the chimeric version of 2A1, and humanized variants of 2A1. Exemplary antibodies of the invention include an antibody having a variable heavy (VH) chain selected from SEQ ID NOs: 305-330, and having a variable light (VL) chain selected from SEQ ID NOs: 331-347. Specifically, exemplary antibodies include those provided in Table 1.

Table 1

[0142] These anti-CD47 antibodies, monospecific anti-CD47 antibodies, monovalent anti-CD47 antibodies, and/or bispecific antibodies in which at least one binding site is specific for CD47 contain a variable heavy chain amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to the amino acid sequence of SEQ ID NO: 114 and a variable light chain amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to an amino acid sequence selected from SEQ ID NO: 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122, 124, 126, 128, 130, 132, 134, 136, 138, 140, 142, 144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174, 176, 178, 180, 182, 184 and 186.

[0143] The disclosure also provides monovalent antibodies that bind CD47. These antibodies are collectively referred to herein as anti-CD47 monovalent antibodies or anti-CD47 mono mAbs. The monovalent antibodies include one arm that specific recognizes CD47, and a second arm referred to herein as a dummy arm. The dummy arm includes an amino acid sequence that does not bind or otherwise cross-react with a human protein. In some embodiments, the dummy arm includes an amino acid sequence that does not bind or otherwise cross-react with a human protein that is found in whole blood. Those of ordinary skill in the art will appreciate that human proteins found in the blood are a proxy that represent all, or substantially all, antigens present in system circulation. In some embodiments, the dummy arm includes an amino acid sequence that does not bind or otherwise cross-react with a human protein that is found in solid tissue. Preferably, the monovalent antibodies are specific for at least human CD47. In some embodiments, the monovalent antibodies that recognize human CD47 are also cross -reactive for at least one other non-human CD47 protein, such as, by way of non-limiting example, non-human primate CD47, e.g., cynomolgus monkey CD47, and/or rodent CD47.

[0144] In some embodiments, the one or more anti-CD47 agent includes an immunizing agent. In some embodiments, an immunizing agent may include a 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.

[0145] In some embodiments, an immortalized cell lines are those that fuse efficiently, support stable high level expression of antibody by the selected antibodyproducing 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, Calif, and the American Type Culture Collection, Manassas, Va. 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)).

[0146] Also included in the disclosure are antibodies that bind to the same epitope as the CD47 antibodies described herein. For example, antibodies of the invention specifically bind to an epitope that includes one or more amino acid residues on human CD47 (see e.g., GenBank Accession No. Q08722.1). The amino acid sequence of an exemplary human CD47 is provided below (GenBank Accession No. Q08722.1 (GI: 1171879), incorporated herein by reference)

[0147] 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 amino 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 arginine, 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; (iii) 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 with 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 in the art can recognize sequence motifs and structural conformations that may be used to define structural and functional domains in accordance with the invention.

[0148] Preferred amino 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 substitution 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, N.Y. (1991)); and Thornton et al. Nature 354:105 (1991).

[0149] 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 The Scientist, Inc., Philadelphia Pa., Vol. 14, No. 8 (Apr. 17, 2000), pp. 25-28).

[0150] Screening of monoclonal antibodies can be carried out, e.g., by measuring CD47- and/or CD47/SIRPa- mediated signaling, and determining whether the test monoclonal antibody is able to modulate, block, inhibit, reduce, antagonize, neutralize or otherwise interfere with CD47- and/or CD47/SIRPa-mediated signaling. These assays can include competitive binding assays. Additionally, these assays can measure a biologic readout, for example the ability to promote phagocytosis of a CD47 expressing cell by a macrophage. In some embodiments, CD47 antibodies are identified using a modified RIMMS (Repetitive Immunization Multiple Sites) immunization strategy in mice and subsequent hybridoma generation.

[0151] Various procedures known within the art may be used for the production of monoclonal antibodies directed against CD47, 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, N.Y., incorporated herein by reference). 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 “fully human antibodies” herein. Human monoclonal antibodies are prepared, for example, using the procedures described in the Examples provided below. Human monoclonal antibodies can be also 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). Antibodies may also be produced by Chinese hamster ovary (CHO) cells (see Dhara, et ah, 2018. BioDrugs 32, 571- 584)

[0152] An alternative method for determining whether a monoclonal antibody has the specificity of monoclonal antibody is to pre-incubate the monoclonal antibody of the invention with soluble CD47 protein (with which it is normally reactive), and then add the monoclonal antibody being tested to determine if the monoclonal antibody being tested is inhibited in its ability to bind CD47. If the monoclonal antibody being tested is inhibited then, in all likelihood, it has the same, or functionally equivalent, epitope specificity as the monoclonal antibody.

[0153] In other, alternative methods, a CD47 antibody is developed, for example, using phage-display methods using antibodies containing only human sequences. Such approaches are well-known in the art, e.g., in W092/01047 and U.S. Pat. No. 6,521,404, which are hereby incorporated by reference. In this approach, a combinatorial library of phage carrying random pairs of light and heavy chains are screened using natural or recombinant source of cd47 or fragments thereof. In another approach, a CD47 antibody can be produced by a process wherein at least one step of the process includes immunizing a transgenic, non-human animal with human CD47 protein. In this approach, some of the endogenous heavy and/or kappa light chain loci of this xenogenic non-human animal have been disabled and are incapable of the rearrangement required to generate genes encoding immunoglobulins in response to an antigen. In addition, at least one human heavy chain locus and at least one human light chain locus have been stably transfected into the animal. Thus, in response to an administered antigen, the human loci rearrange to provide genes encoding human variable regions immuno specific for the antigen. Upon immunization, therefore, the xenomouse produces B-cells that secrete fully human immunoglobulins.

[0154] The present disclosure also provides isolated, recombinant and/or synthetic anti-CD47 human, primate, rodent, mammalian, chimeric, humanized and/or CDR- grafted antibodies as well as compositions and encoding nucleic acid molecules comprising at least one polynucleotide encoding at least a portion of one anti-CD47 antibody molecule. The present disclosure further includes, but is not limited to, methods of making and using such nucleic acids and antibodies including diagnostic and therapeutic compositions, methods and devices. [0155] An exemplary anti-CD47 monovalent and bispecific antibodies include a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ

ID NO: 1.

[0156] Common-HC-NT (SEQ ID NO: 1)

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTC CCTG AGACTCTCCTGTGCAGCCTCTGGATTCACCTTTAGCAGCTATGCCA TGAGCTGGG TCCGCCAGGCTCCAGGGAAGGGGCTGGAGTGGGTCTCAGCT ATTAGTGGTAGTG GTGGTAGCACATACTACGCAGACTCCGTGAAGGGCCG GTTCACCATCTCCAGAG ACAATTCCAAGAACACGCTGTATCTGCAAATGA ACAGCCTGAGAGCCGAGGAC ACGGCCGTATATTACTGTGCGAAAAGTTAT GGTGCTTTTGACTACTGGGGCCAG GGAACCCTGGTCACAGTCTCGAGCGC CTCCACCAAGGGCCCATCGGTCTTCCCC CTGGCACCCTCCTCCAAGAGCA CCTCTGGGGGCACAGCGGCCCTGGGCTGCCTG GTCAAGGACTACTTCCCC GAACCGGTGACAGTCTCGTGGAACTCAGGAGCCCTG ACCAGCGGCGTGCA CACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCC TCAGCAGCG TGGTGACTGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCT GCAAC GTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAGAGTTGAGCCC AA ATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCC TG GGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTC ATGATC TCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCA CGAAGACCCT GAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGC ATAATGCCAAGAC AAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGT GTGGTCAGCGTCCTCA CCGTCCTGCACCAGGACTGGCTGAATGGCAAGGA GTACAAGTGCAAGGTCTCCA ACAAAGCCCTCCCAGCCCCCATCGAGAAAA CCATCTCCAAAGCCAAAGGGCAG CCCCGAGAACCACAGGTGTATACCCTG CCCCCATCTCGGGAGGAGATGACCAAG AACCAGGTCAGCCTGACTTGCCT GGTCAAAGGCTTCTATCCCAGCGACATCGCC GTGGAGTGGGAGAGCAACG GGCAGCCGGAGAACAACTACAAGACCACGCCTCC CGTGCTGGACTCCGAC GGCTCCTTCTTCCTCTATAGCAAGCTCACCGTGGACAAG TCCAGGTGGCA GCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTG CACAACC ACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTTAA

[0157] COMMON-HC-AA (SEQ ID NO: 2)

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSA ISGSGG STYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSY GAFDYWGQGTL VTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFP EPVTVSWNSGALTSGVHT FPAVLQSSGLYSLSSVVTVPSSSLGTQTYICN VNHKPSNTKVDKRVEPKSCDKTHTC PPCPAPELLGGPSVFLFPPKPKDTL MISRTPEVTCVVVDVSHEDPEVKFNWYVDGVE VHNAKTKPREEQYNSTYR VVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKA KGQPREPQVYTL PPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTP PVLDSD GSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG

[0158] The 5A3 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 4) encoded by the nucleic acid sequence shown in SEQ ID NO: 3. [0159] 5A3-LC-NT (SEQ ID NO: 3)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGGACATTAATAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTACGGT GCATCCAGGTTG GAAACAGGGGTCCCATCAAGGTTCAGTGGAAGTGGATC TGGGACAGATTTTACT TTCACCATCAGCAGCCTGCAGCCTGAAGATATTG CAACATATTACTGTCAGCAG AAGCACCCCCGGGGGCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0160] 5A3-LC-AA (SEQ ID NO: 4)

DIQMTQSPSSLSASVGDRVTITCQASQDINKYLNWYQQKPGKAPKLLIYG ASRLETG VPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQKHPRGPRTFG QGTKVEIKRTVAAPS VFIFPPSDEQEKSGTASVVCEENNFYPREAKVQWK VDNAEQSGNSQESVTEQDSKD STYSESSTETESKADYEKHKVYACEVTHQ GESSPVTKSFNRGEC

[0161] The 5A3 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 96) encoded by the nucleic acid sequence shown in SEQ ID NO: 95.

[0162] 5A3-VL-NT (SEQ ID NO: 95)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGGACATTAATAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTACGGT GCATCCAGGTTG GAAACAGGGGTCCCATCAAGGTTCAGTGGAAGTGGATC TGGGACAGATTTTACT TTCACCATCAGCAGCCTGCAGCCTGAAGATATTG CAACATATTACTGTCAGCAG AAGCACCCCCGGGGGCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0163] 5A3-VL-AA (SEQ ID NO: 96)

DIQMTQSPSSLSASVGDRVTITCQASQDINKYLNWYQQKPGKAPKLLIYG ASRLETG VPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQKHPRGPRTFG QGTKVEIK

[0164] The 5A3-M4 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 6) encoded by the nucleic acid sequence shown in SEQ ID NO: 5.

[0165] 5A3-M4-LC-NT (SEQ ID NO: 5) GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGGACATTAATAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTACGGT GCATCCAGGTTG GAAACAGGGGTCCCATCAAGGTTCAGTGGAAGTGGATC TGGGACAGATTTTACT TTCACCATCAGCAGCCTGCAGCCTGAAGATATTG CAACATATTACTGTCAGCAG AAGCACCCCCGGAACCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0166] 5A3-M4-LC-AA (SEQ ID NO: 6)

DIQMTQSPSSLSASVGDRVTITCQASQDINKYLNWYQQKPGKAPKLLIYG ASRLETG VPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQKHPRNPRTFG QGTKVEIKRTVAAPS VFIFPPSDEQEKSGTASVVCEENNFYPREAKVQWK VDNAEQSGNSQESVTEQDSKD STYSESSTETESKADYEKHKVYACEVTHQ GESSPVTKSFNRGEC

[0167] The 5A3-M4 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 98) encoded by the nucleic acid sequence shown in SEQ ID NO: 97.

[0168] 5A3-M4-VL-NT (SEQ ID NO: 97)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGGACATTAATAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTACGGT GCATCCAGGTTG GAAACAGGGGTCCCATCAAGGTTCAGTGGAAGTGGATC TGGGACAGATTTTACT TTCACCATCAGCAGCCTGCAGCCTGAAGATATTG CAACATATTACTGTCAGCAG AAGCACCCCCGGAACCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0169] 5A3-M4-VL-AA (SEQ ID NO: 98)

DIQMTQSPSSLSASVGDRVTITCQASQDINKYLNWYQQKPGKAPKLLIYG ASRLETG VPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQKHPRNPRTFG QGTKVEIK

[0170] The 5A3-M3 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 8) encoded by the nucleic acid sequence shown in SEQ ID NO: 7.

[0171] 5A3-M3-LC-NT (SEQ ID NO: 7)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGTCCATTAGTAGTTATTTAA ATTGGTATC AGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTACGCT GCATCCTCGTTGG AAACAGGGGTCCCATCAAGGTTCAGTGGAAGTGGATC TGGGACAGATTTTACTT TCACCATCAGCAGCCTGCAGCCTGAAGATATTG CAACATATTACTGTCAGCAGA AGCACCCCCGGGGGCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA CGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTGA AATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGGC CAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGA GTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTG ACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCAC CCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTA A

[0172] 5A3-M3-LC-AA (SEQ ID NO: 8)

DIQMTQSPSSLSASVGDRVTITCQASQSISSYLNWYQQKPGKAPKLLIYA ASSLETGV PSRFSGSGSGTDFTFTISSLQPEDIATYYCQQKHPRGPRTFG QGTKVEIKRTVAAPSVF IFPPSDEQEKSGTASVVCEENNFYPREAKVQWK VDNAEQSGNSQESVTEQDSKDST YSESSTETESKADYEKHKVYACEVTHQ GESSPVTKSFNRGEC

[0173] The 5A3-M3 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 100) encoded by the nucleic acid sequence shown in SEQ ID NO: 99.

[0174] 5A3-M3-VL-NT (SEQ ID NO: 99)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGTCCATTAGTAGTTATTTAA ATTGGTATC AGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTACGCT GCATCCTCGTTGG AAACAGGGGTCCCATCAAGGTTCAGTGGAAGTGGATC TGGGACAGATTTTACTT TCACCATCAGCAGCCTGCAGCCTGAAGATATTG CAACATATTACTGTCAGCAGA AGCACCCCCGGGGGCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA [0175] 5A3-M3-VL-AA (SEQ ID NO: 100)

DIQMTQSPSSLSASVGDRVTITCQASQSISSYLNWYQQKPGKAPKLLIYA ASSLETGV PSRFSGSGSGTDFTFTISSLQPEDIATYYCQQKHPRGPRTFG QGTKVEIK

[0176] The 5A3-M5 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 10) encoded by the nucleic acid sequence shown in SEQ ID NO: 9.

[0177] 5A3-M5-LC-NT (SEQ ID NO: 9)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGGACATTAATAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTACGGT GCATCCAGGTTG GAAACAGGGGTCCCATCAAGGTTCAGTGGAAGTGGATC TGGGACAGATTTTACT TTCACCATCAGCAGCCTGCAGCCTGAAGATATTG CAACATATTACTGTCAGCAG AAGCACCCCCGGTACCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0178] 5A3-M5-LC-AA (SEQ ID NO: 10)

DIQMTQSPSSLSASVGDRVTITCQASQDINKYLNWYQQKPGKAPKLLIYG ASRLETG VPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQKHPRYPRTFG QGTKVEIKRTVAAPS VFIFPPSDEQEKSGTASVVCEENNFYPREAKVQWK VDNAEQSGNSQESVTEQDSKD STYSESSTETESKADYEKHKVYACEVTHQ GESSPVTKSFNRGEC

[0179] The 5A3-M5 antibody includes a common variable heavy domain (SEQ

ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 102) encoded by the nucleic acid sequence shown in SEQ ID NO: 101.

[0180] 5A3-M5-VL-NT (SEQ ID NO: 101)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGGACATTAATAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTACGGT GCATCCAGGTTG GAAACAGGGGTCCCATCAAGGTTCAGTGGAAGTGGATC TGGGACAGATTTTACT TTCACCATCAGCAGCCTGCAGCCTGAAGATATTG CAACATATTACTGTCAGCAG AAGCACCCCCGGTACCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0181] 5A3-M5-VL-AA (SEQ ID NO: 102)

DIQMTQSPSSLSASVGDRVTITCQASQDINKYLNWYQQKPGKAPKLLIYG ASRLETG VPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQKHPRYPRTFG QGTKVEIK

[0182] The Ke8 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 12) encoded by the nucleic acid sequence shown in SEQ ID NO: 11.

[0183] Ke8-LC-NT (SEQ ID NO: 11)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGCAGCTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAGT TCCACAAGCGGCGGCCGCAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA CGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTGA AATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGGC CAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGA GTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTG ACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCAC CCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTA A

[0184] KE8-LC-AA (SEQ ID NO: 12)

DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYA ASSLQSGV PSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFHKRRPQTFG QGTKVEIKRTVAAPSV FIFPPSDEQEKSGTASVVCEENNFYPREAKVQWK VDNAEQSGNSQESVTEQDSKDS TYSESSTETESKADYEKHKVYACEVTHQ GESSPVTKSFNRGEC

[0185] The Ke8 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 104) encoded by the nucleic acid sequence shown in SEQ ID NO: 103.

[0186] Ke8-VL-NT (SEQ ID NO: 103)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGCAGCTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAGT TCCACAAGCGGCGGCCGCAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA [0187] KE8-VL-AA (SEQ ID NO: 104)

DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYA ASSLQSGV PSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFHKRRPQTFG QGTKVEIK

[0188] The Ke8H5 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 14) encoded by the nucleic acid sequence shown in SEQ ID NO: 13.

[0189] KE8H5-LC-NT (SEQ ID NO: 13)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGCGAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAGT TCCATAAGCGTGCGCCGCAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA CGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTGA AATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGGC CAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGA GTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTG ACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCAC CCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTA A

[0190] KE8H5-LC-AA (SEQ ID NO: 14) DIQMTQSPSSLSASVGDRVTITCRASQSIARYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFHKRAPQTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0191] The Ke8H5 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 106) encoded by the nucleic acid sequence shown in SEQ ID NO: 105.

[0192] KE8H5-VL-NT (SEQ ID NO: 105)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGCGAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAGT TCCATAAGCGTGCGCCGCAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA [0193] KE8H5-VL-AA (SEQ ID NO: 106)

DIQMTQSPSSLSASVGDRVTITCRASQSIARYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFHKRAPQTFG QGTKVEIK

[0194] The Ke8B2 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 16) encoded by the nucleic acid sequence shown in SEQ ID NO: 15.

[0195] KE8B2-LC-NT (SEQ ID NO: 15)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCG GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AAGCACCCGCGTGCCCCGCGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0196] KE8B2-LC-AA (SEQ ID NO: 16)

DIQMTQSPSSLSASVGDRVTITCRASQSIGKYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQKHPRAPRTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC [0197] The Ke8B2 antibody includes a common variable heavy domain (SEQ ID

NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 108) encoded by the nucleic acid sequence shown in SEQ ID NO: 107.

[0198] KE8B2-VL-NT (SEQ ID NO: 107)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCG GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AAGCACCCGCGTGCCCCGCGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0199] KE8B2-VL-AA (SEQ ID NO: 108)

DIQMTQSPSSLSASVGDRVTITCRASQSIGKYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQKHPRAPRTFG QGTKVEIK

[0200] The Ke8A2 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 18) encoded by the nucleic acid sequence shown in SEQ ID NO: 17.

[0201] KE8A2-LC-NT (SEQ ID NO: 17)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGATAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AAGCATCCCCGTGGGCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0202] KE8A2-LC-AA (SEQ ID NO: 18)

DIQMTQSPSSLSASVGDRVTITCRASQSIDRYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQKHPRGPRTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC [0203] The Ke8A2 antibody includes a common variable heavy domain (SEQ ID

NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 110) encoded by the nucleic acid sequence shown in SEQ ID NO: 109.

[0204] KE8A2-VL-NT (SEQ ID NO: 109)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGATAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AAGCATCCCCGTGGGCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0205] KE8A2-VL-AA (SEQ ID NO: 110)

DIQMTQSPSSLSASVGDRVTITCRASQSIDRYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQKHPRGPRTFG QGTKVEIK

[0206] The Ke8E8 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 20) encoded by the nucleic acid sequence shown in SEQ ID NO: 19.

[0207] KE8E8-LC-NT (SEQ ID NO: 19)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGGACATTAATAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AAGCATCCCCGTGGCCCGCGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0208] KE8E8-LC-AA (SEQ ID NO: 20)

DIQMTQSPSSLSASVGDRVTITCQASQDINKYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQKHPRGPRTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0209] The Ke8E8 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 112) encoded by the nucleic acid sequence shown in SEQ ID NO: 111.

[0210] KE8E8-VL-NT (SEQ ID NO: 111)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGGACATTAATAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AAGCATCCCCGTGGCCCGCGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0211] KE8E8-VL-AA (SEQ ID NO: 112)

DIQMTQSPSSLSASVGDRVTITCQASQDINKYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQKHPRGPRTFG QGTKVEIK

[0212] The Ke8H3 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 22) encoded by the nucleic acid sequence shown in SEQ ID NO: 21.

[0213] KE8H3-LC-NT (SEQ ID NO: 21)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAATAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AAGCATCCGCGTGGGCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0214] KE8H3-LC-AA (SEQ ID NO: 22)

DIQMTQSPSSLSASVGDRVTITCRASQSINRYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQKHPRGPRTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0215] The Ke8H3 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113. [0216] KE8H3-VL-NT (SEQ ID NO: 113)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAATAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AAGCATCCGCGTGGGCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0217] KE8H3-VL-AA (SEQ ID NO: 114)

DIQMTQSPSSLSASVGDRVTITCRASQSINRYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQKHPRGPRTFG QGTKVEIK

[0218] The Ke8G6 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 24) encoded by the nucleic acid sequence shown in SEQ ID NO: 23.

[0219] KE8G6-LC-NT (SEQ ID NO: 23)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGGTAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCGCGTGCGCCGAAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0220] KE8G6-LC-AA (SEQ ID NO: 24)

DIQMTQSPSSLSASVGDRVTITCRASQSIGRYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRAPKTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0221] The Ke8G6 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 116) encoded by the nucleic acid sequence shown in SEQ ID NO: 115.

[0222] KE8G6-VL-NT (SEQ ID NO: 115)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGGTAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCGCGTGCGCCGAAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0223] KE8G6-VL-AA (SEQ ID NO: 116)

DIQMTQSPSSLSASVGDRVTITCRASQSIGRYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRAPKTFG QGTKVEIK

[0224] The Ke8A3 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 26) encoded by the nucleic acid sequence shown in SEQ ID NO: 25.

[0225] KE8A3-LC-NT (SEQ ID NO: 25)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGTAAGTCAGAGCATTAGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AGGCATCCCCGTGGGCCGAGCACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0226] KE8A3-LC-AA (SEQ ID NO: 26)

DIQMTQSPSSLSASVGDRVTITCRVSQSISKYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQRHPRGPSTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0227] The Ke8A3 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 118) encoded by the nucleic acid sequence shown in SEQ ID NO: 117.

[0228] KE8A3-VL-NT (SEQ ID NO: 117)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGTAAGTCAGAGCATTAGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AGGCATCCCCGTGGGCCGAGCACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0229] KE8A3-VL-AA (SEQ ID NO: 118)

DIQMTQSPSSLSASVGDRVTITCRVSQSISKYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQRHPRGPSTFG QGTKVEIK

[0230] The Ke81A3 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 28) encoded by the nucleic acid sequence shown in SEQ ID NO: 27.

[0231] KE81 A3-LC-NT (SEQ ID NO: 27)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGGACATTAATAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AGGCATCCGCGTGCCCCGCGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0232] KE81 A3-LC-AA (SEQ ID NO: 28)

DIQMTQSPSSLSASVGDRVTITCQASQDINRYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQRHPRAPRTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0233] The Ke81A3 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 120) encoded by the nucleic acid sequence shown in SEQ ID NO: 119.

[0234] KE81A3-VL-NT (SEQ ID NO: 119)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGGACATTAATAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AGGCATCCGCGTGCCCCGCGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0235] KE81A3-VL-AA (SEQ ID NO: 120) DIQMTQSPSSLSASVGDRVTITCQASQDINRYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQRHPRAPRTFG QGTKVEIK

[0236] The Ke8A8 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 30) encoded by the nucleic acid sequence shown in SEQ ID NO: 29.

[0237] KE8A8-LC-NT (SEQ ID NO: 29)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCACTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCGCGTGCGCCGAAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0238] KE8A8-LC-AA (SEQ ID NO: 30)

DIQMTQSPSSLSASVGDRVTITCRASQSISKYLNWYQQKPGKAPKLLIYA ASTLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRAPKTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0239] The Ke8A8 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 122) encoded by the nucleic acid sequence shown in SEQ ID NO: 121.

[0240] KE8A8-VL-NT (SEQ ID NO: 121)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCACTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCGCGTGCGCCGAAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0241] KE8A8-VL-AA (SEQ ID NO: 122)

DIQMTQSPSSLSASVGDRVTITCRASQSISKYLNWYQQKPGKAPKLLIYA ASTLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRAPKTFG QGTKVEIK [0242] The Ke8C7 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 32) encoded by the nucleic acid sequence shown in SEQ ID NO: 31.

[0243] KE8C7-LC-NT (SEQ ID NO: 31)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGGACATTAATAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAGC GCCATCCGCGTGGCCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA CGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTGA AATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGGC CAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGA GTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTG ACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCAC CCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTA A

[0244] KE8C7-LC-AA (SEQ ID NO: 32)

DIQMTQSPSSLSASVGDRVTITCQASQDINRYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQRHPRGPRTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0245] The Ke8C7 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 124) encoded by the nucleic acid sequence shown in SEQ ID NO: 123.

[0246] KE8C7-VL-NT (SEQ ID NO: 123)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGGACATTAATAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAGC GCCATCCGCGTGGCCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA [0247] KE8C7-VL-AA (SEQ ID NO: 124)

DIQMTQSPSSLSASVGDRVTITCQASQDINRYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQRHPRGPRTFG QGTKVEIK

[0248] The Ke8G2 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 34) encoded by the nucleic acid sequence shown in SEQ ID NO: 33. [0249] KE8G2-LC-NT (SEQ ID NO: 33)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGGTAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCG GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAACAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AAGCATCCCCGTGCGCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0250] KE8G2-LC-AA (SEQ ID NO: 34)

DIQMTQSPSSLSASVGDRVTITCRASQSIGRYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTINSLQPEDFATYYCQQKHPRAPRTFG QGTKVEIKRTVAAPS VFIFPPSDEQEKSGTASVVCEENNFYPREAKVQWK VDNAEQSGNSQESVTEQDSKD STYSESSTETESKADYEKHKVYACEVTHQ GESSPVTKSFNRGEC

[0251] The Ke8G2 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 126) encoded by the nucleic acid sequence shown in SEQ ID NO: 125.

[0252] KE8G2-VL-NT (SEQ ID NO: 125)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGGTAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCG GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAACAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AAGCATCCCCGTGCGCCGAGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0253] KE8G2-VL-AA (SEQ ID NO: 126)

DIQMTQSPSSLSASVGDRVTITCRASQSIGRYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTINSLQPEDFATYYCQQKHPRAPRTFG QGTKVEIK

[0254] The Ke81G9 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 36) encoded by the nucleic acid sequence shown in SEQ ID NO: 35.

[0255] KE81G9-LC-NT (SEQ ID NO: 35)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGATAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCG GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAGC GGCATAAGCGTTCCCCGCAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA CGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTGA AATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGGC CAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGA GTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTG ACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCAC CCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTA A

[0256] KE81G9-LC-AA (SEQ ID NO: 36)

DIQMTQSPSSLSASVGDRVTITCRASQSIDKYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQRHKRSPQTFG QGTKVEIKRTVAAPS VFIFPPSDEQEKSGTASVVCEENNFYPREAKVQWK VDNAEQSGNSQESVTEQDSKD STYSESSTETESKADYEKHKVYACEVTHQ GESSPVTKSFNRGEC

[0257] The Ke81G9 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 128) encoded by the nucleic acid sequence shown in SEQ ID NO: 127.

[0258] KE81G9-VL-NT (SEQ ID NO: 127)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGATAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCG GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAGC GGCATAAGCGTTCCCCGCAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA [0259] KE81G9-VL-AA (SEQ ID NO: 128)

DIQMTQSPSSLSASVGDRVTITCRASQSIDKYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQRHKRSPQTFG QGTKVEIK

[0260] The Ke8F2 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 38) encoded by the nucleic acid sequence shown in SEQ ID NO: 37.

[0261] KE8F2-LC-NT (SEQ ID NO: 37)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGATAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AAGCATCCGCGTGCGCCGCGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0262] KE8F2-LC-AA (SEQ ID NO: 38)

DIQMTQSPSSLSASVGDRVTITCRASQSIDKYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQKHPRAPRTFG QGTKVEIKRTVAAPS VFIFPPSDEQEKSGTASVVCEENNFYPREAKVQWK VDNAEQSGNSQESVTEQDSKD STYSESSTETESKADYEKHKVYACEVTHQ GESSPVTKSFNRGEC

[0263] The Ke8F2 antibody includes a common variable heavy domain (SEQ ID

NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 130) encoded by the nucleic acid sequence shown in SEQ ID NO: 129.

[0264] KE8F2-VL-NT (SEQ ID NO: 129)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGATAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AAGCATCCGCGTGCGCCGCGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0265] KE8F2-VL-AA (SEQ ID NO: 130)

DIQMTQSPSSLSASVGDRVTITCRASQSIDKYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQKHPRAPRTFG QGTKVEIK

[0266] The Ke8B7 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 40) encoded by the nucleic acid sequence shown in SEQ ID NO: 39.

[0267] KE8B7-LC-NT (SEQ ID NO: 39)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGGGAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCGCGTAGCCCGAAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0268] KE8B7-LC-AA (SEQ ID NO: 40)

DIQMTQSPSSLSASVGDRVTITCRASQSIGKYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRSPKTFG QGTKVEIKRTVAAPS VFIFPPSDEQEKSGTASVVCEENNFYPREAKVQWK VDNAEQSGNSQESVTEQDSKD STYSESSTETESKADYEKHKVYACEVTHQ GESSPVTKSFNRGEC

[0269] The Ke8B7 antibody includes a common variable heavy domain (SEQ ID

NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 132) encoded by the nucleic acid sequence shown in SEQ ID NO: 131.

[0270] KE8B7-VL-NT (SEQ ID NO: 131)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGGGAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCGCGTAGCCCGAAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0271] KE8B7-VL-AA (SEQ ID NO: 132)

DIQMTQSPSSLSASVGDRVTITCRASQSIGKYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRSPKTFG QGTKVEIK

[0272] The Ke8C4 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 42) encoded by the nucleic acid sequence shown in SEQ ID NO: 41.

[0273] KE8C4-LC-NT (SEQ ID NO: 41)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGTAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAATTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCGCGTGGGCCGAAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0274] KE8C4-LC-AA (SEQ ID NO: 42)

DIQMTQSPSSLSASVGDRVTITCRASQSISRYLNWYQQKPGKAPKLLIYA ASNLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRGPKTFG QGTKVEIKRTVAAPS VFIFPPSDEQEKSGTASVVCEENNFYPREAKVQWK VDNAEQSGNSQESVTEQDSKD STYSESSTETESKADYEKHKVYACEVTHQ GESSPVTKSFNRGEC

[0275] The Ke8C4 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 134) encoded by the nucleic acid sequence shown in SEQ ID NO: 133.

[0276] KE8C4-VL-NT (SEQ ID NO: 133)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGTAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAATTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCGCGTGGGCCGAAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0277] KE8C4-VL-AA (SEQ ID NO: 134)

DIQMTQSPSSLSASVGDRVTITCRASQSISRYLNWYQQKPGKAPKLLIYA ASNLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRGPKTFG QGTKVEIK

[0278] The Ke8Fl antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 44) encoded by the nucleic acid sequence shown in SEQ ID NO: 43.

[0279] KE8F1-LC-NT (SEQ ID NO: 43)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGCTTCTTATGTAA ATTGGTATC AGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCGGTTTGC AAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACTC TCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAGTT CCATAAGCGTCGGCCGCAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAAAC GTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTGAA ATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCC AAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAG TGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGA CGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACC CATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAA

[0280] KE8F1-LC-AA (SEQ ID NO: 44) DIQMTQSPSSLSASVGDRVTITCRASQSIASYVNWYQQKPGKAPKLLIYA ASGLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFHKRRPQTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0281] The Ke8Fl antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 136) encoded by the nucleic acid sequence shown in SEQ ID NO: 135.

[0282] KE8F1-VL-NT (SEQ ID NO: 135)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGCTTCTTATGTAA ATTGGTATC AGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCGGTTTGC AAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACTC TCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAGTT CCATAAGCGTCGGCCGCAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA [0283] KE8F1-VL-AA (SEQ ID NO: 136)

DIQMTQSPSSLSASVGDRVTITCRASQSIASYVNWYQQKPGKAPKLLIYA ASGLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFHKRRPQTFG QGTKVEIK

[0284] The Ke8Gll antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 46) encoded by the nucleic acid sequence shown in SEQ ID NO: 45.

[0285] KE8G11-LC-NT (SEQ ID NO: 45)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGGGAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCGCGTGGGCCGAAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0286] KE8G11-LC-AA (SEQ ID NO: 46)

DIQMTQSPSSLSASVGDRVTITCRASQSIGRYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRGPKTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC [0287] The Ke8Gll antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 138) encoded by the nucleic acid sequence shown in SEQ ID NO: 137.

[0288] KE8G11-VL-NT (SEQ ID NO: 137)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGGGAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCGCGTGGGCCGAAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0289] KE8G11-VL-AA (SEQ ID NO: 138)

DIQMTQSPSSLSASVGDRVTITCRASQSIGRYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRGPKTFG QGTKVEIK

[0290] The Ke8H6 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 48) encoded by the nucleic acid sequence shown in SEQ ID NO: 47.

[0291] KE8H6-LC-NT (SEQ ID NO: 47)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATAAT GCATCCACTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AGGCATCCGCGTGGGCCGCGCACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0292] KE8H6-LC-AA (SEQ ID NO: 48)

DIQMTQSPSSLSASVGDRVTITCRASQSISKYLNWYQQKPGKAPKLLIYN ASTLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQRHPRGPRTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0293] The Ke8H6 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 140) encoded by the nucleic acid sequence shown in SEQ ID NO: 139.

[0294] KE8H6-VL-NT (SEQ ID NO: 139)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATAAT GCATCCACTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AGGCATCCGCGTGGGCCGCGCACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0295] KE8H6-VL-AA (SEQ ID NO: 140)

DIQMTQSPSSLSASVGDRVTITCRASQSISKYLNWYQQKPGKAPKLLIYN ASTLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQRHPRGPRTFG QGTKVEIK

[0296] The Ke84G9 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 50) encoded by the nucleic acid sequence shown in SEQ ID NO: 49.

[0297] KE84G9-LC-NT (SEQ ID NO: 49)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AAGCATCCGCGTAGCCCGCGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0298] KE84G9-LC-AA (SEQ ID NO: 50)

DIQMTQSPSSLSASVGDRVTITCRASQSISKYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQKHPRSPRTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0299] The Ke84G9 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 142) encoded by the nucleic acid sequence shown in SEQ ID NO: 141. [0300] KE84G9-VL-NT (SEQ ID NO: 141)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AAGCATCCGCGTAGCCCGCGGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0301] KE84G9-VL-AA (SEQ ID NO: 142)

DIQMTQSPSSLSASVGDRVTITCRASQSISKYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQKHPRSPRTFG QGTKVEIK

[0302] The Ke8A4 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 52) encoded by the nucleic acid sequence shown in SEQ ID NO: 51.

[0303] KE8A4-LC-NT (SEQ ID NO: 51)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGCTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAGT TCCATAAGCGTAGCCCGCAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA CGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTGA AATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGGC CAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGA GTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTG ACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCAC CCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTA A

[0304] KE8A4-LC-AA (SEQ ID NO: 52)

DIQMTQSPSSLSASVGDRVTITCRASQSIAKYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFHKRSPQTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0305] The Ke8A4 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 144) encoded by the nucleic acid sequence shown in SEQ ID NO: 143.

[0306] KE8A4-VL-NT (SEQ ID NO: 143)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGCTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAGT TCCATAAGCGTAGCCCGCAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA [0307] KE8A4-VL-AA (SEQ ID NO: 144)

DIQMTQSPSSLSASVGDRVTITCRASQSIAKYLNWYQQKPGKAPKLLIYA ASSLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFHKRSPQTFG QGTKVEIK

[0308] The Ke86G9 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 54) encoded by the nucleic acid sequence shown in SEQ ID NO: 53.

[0309] KE86G9-LC-NT (SEQ ID NO: 53)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATAAT GCATCCAATTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AGGCATCCGCGTGGGCCGACCACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0310] KE86G9-LC-AA (SEQ ID NO: 54)

DIQMTQSPSSLSASVGDRVTITCRASQSISKYLNWYQQKPGKAPKLLIYN ASNLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQRHPRGPTTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0311] The Ke86G9 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 146) encoded by the nucleic acid sequence shown in SEQ ID NO: 145.

[0312] KE86G9-VL-NT (SEQ ID NO: 145)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATAAT GCATCCAATTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG AGGCATCCGCGTGGGCCGACCACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0313] KE86G9-VL-AA (SEQ ID NO: 146)

DIQMTQSPSSLSASVGDRVTITCRASQSISKYLNWYQQKPGKAPKLLIYN ASNLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQRHPRGPTTFG QGTKVEIK

[0314] The Ka3 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 56) encoded by the nucleic acid sequence shown in SEQ ID NO: 55.

[0315] KA3-LC-NT (SEQ ID NO: 55)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGCAGCTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCACCCGCGCGCCCCGAAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0316] KA3-LC-AA (SEQ ID NO: 56)

DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYA ASSLQSGV PSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRAPKTFG QGTKVEIKRTVAAPSV FIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKDS TYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0317] The Ka3 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 148) encoded by the nucleic acid sequence shown in SEQ ID NO: 147.

[0318] KA3-VL-NT (SEQ ID NO: 147)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGCAGCTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCACCCGCGCGCCCCGAAGACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0319] KA3-VL-AA (SEQ ID NO: 148) DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYA ASSLQSGV PSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRAPKTFG QGTKVEIK

[0320] The Ka3A2 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 58) encoded by the nucleic acid sequence shown in SEQ ID NO: 57.

[0321] KA3A2-LC-NT (SEQ ID NO: 57)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCTCGCTCGCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA CGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTGA AATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGGC CAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGA GTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTG ACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCAC CCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTA A

[0322] KA3A2-LC-AA (SEQ ID NO: 58)

DIQMTQSPSSLSASVGDRVTITCRASQSISKYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRSPKTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0323] The Ka3A2 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 150) encoded by the nucleic acid sequence shown in SEQ ID NO: 149.

[0324] KA3A2-VL-NT (SEQ ID NO: 149)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCTCGCTCGCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA [0325] KA3A2-VL-AA (SEQ ID NO: 150)

DIQMTQSPSSLSASVGDRVTITCRASQSISKYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRSPKTFG QGTKVEIK [0326] The Ka3H3 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 60) encoded by the nucleic acid sequence shown in SEQ ID NO: 59.

[0327] KA3H3-LC-NT (SEQ ID NO: 59)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGGACATTGCTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCGCTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCTCGCTCGCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA CGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTGA AATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGGC CAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGA GTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTG ACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCAC CCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTA A

[0328] KA3H3-LC-AA (SEQ ID NO: 60)

DIQMTQSPSSLSASVGDRVTITCQASQDIAKYLNWYQQKPGKAPKLLIYA ASALQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRSPKTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0329] The Ka3H3 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 152) encoded by the nucleic acid sequence shown in SEQ ID NO: 151.

[0330] KA3H3-VL-NT (SEQ ID NO: 151)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCAGGCGAGTCAGGACATTGCTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCGCTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCTCGCTCGCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA

[0331] KA3H3-VL-AA (SEQ ID NO: 152)

DIQMTQSPSSLSASVGDRVTITCQASQDIAKYLNWYQQKPGKAPKLLIYA ASALQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRSPKTFG QGTKVEIK [0332] The Ka3A3 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 62) encoded by the nucleic acid sequence shown in SEQ ID NO: 61.

[0333] KA3A3-LC-NT (SEQ ID NO: 61)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGCTAGTTATTTAA ATTGGTATC AGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCG GCATCCAGGTTGC AAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACTC TCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAGAT GCATCCTCGCGCGCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAAAC GTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTGAA ATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCC AAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAG TGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGA CGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACC CATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAA

[0334] KA3A3-LC-AA (SEQ ID NO: 62)

DIQMTQSPSSLSASVGDRVTITCRASQSIASYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRAPKTFG QGTKVEIKRTVAAPS VFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK VDNALQSGNSQESVTEQDSKD STYSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0335] The Ka3A3 antibody includes a common variable heavy domain (SEQ ID

NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 154) encoded by the nucleic acid sequence shown in SEQ ID NO: 153.

[0336] KA3A3-VL-NT (SEQ ID NO: 153)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGCTAGTTATTTAA ATTGGTATC AGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCG GCATCCAGGTTGC AAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACTC TCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAGAT GCATCCTCGCGCGCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA [0337] KA3A3-VL-AA (SEQ ID NO: 154)

DIQMTQSPSSLSASVGDRVTITCRASQSIASYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRAPKTFG QGTKVEIK

[0338] The Ka3H8 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 64) encoded by the nucleic acid sequence shown in SEQ ID NO: 63. [0339] KA3H8-LC-NT (SEQ ID NO: 63)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGCGAGTTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCG GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCTCGCTCGCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA CGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTGA AATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGGC CAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGA GTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTG ACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCAC CCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTA A

[0340] KA3H8-LC-AA (SEQ ID NO: 64)

DIQMTQSPSSLSASVGDRVTITCRASQSIASYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRSPKTFG QGTKVEIKRTVAAPS VFIFPPSDEQEKSGTASVVCEENNFYPREAKVQWK VDNAEQSGNSQESVTEQDSKD STYSESSTETESKADYEKHKVYACEVTHQ GESSPVTKSFNRGEC

[0341] The Ka3H8 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 156) encoded by the nucleic acid sequence shown in SEQ ID NO: 155.

[0342] KA3H8-VL-NT (SEQ ID NO: 155)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGCGAGTTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCG GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCTCGCTCGCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAAA

[0343] KA3H8-VL-AA (SEQ ID NO: 156)

DIQMTQSPSSLSASVGDRVTITCRASQSIASYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRSPKTFG QGTKVEIK

[0344] The Ka3B2 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 66) encoded by the nucleic acid sequence shown in SEQ ID NO: 65.

[0345] KA3B2-LC-NT (SEQ ID NO: 65)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAACATTGGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATAGT GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCTCGCGCGCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0346] KA3B2-LC-AA (SEQ ID NO: 66)

DIQMTQSPSSLSASVGDRVTITCRASQNIGKYLNWYQQKPGKAPKLLIYS ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRAPKTFG QGTKVEIKRTVAAPS VFIFPPSDEQEKSGTASVVCEENNFYPREAKVQWK VDNAEQSGNSQESVTEQDSKD STYSESSTETESKADYEKHKVYACEVTHQ GESSPVTKSFNRGEC

[0347] The Ka3B2 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 158) encoded by the nucleic acid sequence shown in SEQ ID NO: 157.

[0348] KA3B2-VL-NT (SEQ ID NO: 157)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAACATTGGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATAGT GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCTCGCGCGCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0349] KA3B2-VL-AA (SEQ ID NO: 158)

DIQMTQSPSSLSASVGDRVTITCRASQNIGKYLNWYQQKPGKAPKLLIYS ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRAPKTFG QGTKVEIK

[0350] The Ka3C5 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 68) encoded by the nucleic acid sequence shown in SEQ ID NO: 67.

[0351] KA3C5-LC-NT (SEQ ID NO: 67)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGTAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATTCT GCATCCTCTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCTCGCGCCCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0352] KA3C5-LC-AA (SEQ ID NO: 68)

DIQMTQSPSSLSASVGDRVTITCRASQSISRYLNWYQQKPGKAPKLLIYS ASSLQSGV PSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRAPKTFG QGTKVEIKRTVAAPSV FIFPPSDEQEKSGTASVVCEENNFYPREAKVQWK VDNAEQSGNSQESVTEQDSKDS TYSESSTETESKADYEKHKVYACEVTHQ GESSPVTKSFNRGEC

[0353] The Ka3C5 antibody includes a common variable heavy domain (SEQ ID

NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 160) encoded by the nucleic acid sequence shown in SEQ ID NO: 159.

[0354] KA3C5-VL-NT (SEQ ID NO: 159)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTAGTAGGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATTCT GCATCCTCTTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCTCGCGCCCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0355] KA3C5-VL-AA (SEQ ID NO: 160)

DIQMTQSPSSLSASVGDRVTITCRASQSISRYLNWYQQKPGKAPKLLIYS ASSLQSGV PSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRAPKTFG QGTKVEIK

[0356] The Ka3G2 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 70) encoded by the nucleic acid sequence shown in SEQ ID NO: 69.

[0357] KA3G2-LC-NT (SEQ ID NO: 69)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGATAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCTCGCGGGCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0358] KA3G2-LC-AA (SEQ ID NO: 70)

DIQMTQSPSSLSASVGDRVTITCRASQSIDKYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRGPKTFG QGTKVEIKRTVAAPS VFIFPPSDEQEKSGTASVVCEENNFYPREAKVQWK VDNAEQSGNSQESVTEQDSKD STYSESSTETESKADYEKHKVYACEVTHQ GESSPVTKSFNRGEC

[0359] The Ka3G2 antibody includes a common variable heavy domain (SEQ ID

NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 162) encoded by the nucleic acid sequence shown in SEQ ID NO: 161.

[0360] KA3G2-VL-NT (SEQ ID NO: 181)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGATAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCTCGCGGGCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0361] KA3G2-VL-AA (SEQ ID NO: 182)

DIQMTQSPSSLSASVGDRVTITCRASQSIDKYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRGPKTFG QGTKVEIK

[0362] The Ka3D3 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a kappa light chain (SEQ ID NO: 72) encoded by the nucleic acid sequence shown in SEQ ID NO: 71.

[0363] KA3D3-LC-NT (SEQ ID NO: 71)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCTCGCGGGCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAA ACGTACGGTGGCTGCACCATCTGTCTT CATCTTCCCGCCATCTGATGAGCAGTTG AAATCTGGAACTGCCTCTGTTG TGTGCCTGCTGAATAACTTCTATCCCAGAGAGG CCAAAGTACAGTGGAAG GTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAG AGTGTCACAGAGCA GGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCT GACGCTGAGCA AAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCA CCCATCAG GGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTT AA

[0364] KA3D3-LC-AA (SEQ ID NO: 72)

DIQMTQSPSSLSASVGDRVTITCRASQSIGKYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRAPKTFG QGTKVEIKRTVAAPS VFIFPPSDEQEKSGTASVVCEENNFYPREAKVQWK VDNAEQSGNSQESVTEQDSKD STYSESSTETESKADYEKHKVYACEVTHQ GESSPVTKSFNRGEC

[0365] The Ka3D3 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a kappa variable light domain (SEQ ID NO: 164) encoded by the nucleic acid sequence shown in SEQ ID NO: 163.

[0366] KA3D3-VL-NT (SEQ ID NO: 163)

GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGA CAGA GTCACCATCACTTGCCGGGCAAGTCAGAGCATTGGTAAGTATTTAA ATTGGTAT CAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGATCTATGCT GCATCCAGGTTG CAAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTGGATC TGGGACAGATTTCACT CTCACCATCAGCAGTCTGCAACCTGAAGATTTTG CAACTTACTACTGTCAGCAG ATGCATCCTCGCGCGCCGAAAACCTTCGGC CAAGGGACCAAGGTGGAAATCAA A

[0367] KA3D3-VL-AA (SEQ ID NO: 164)

DIQMTQSPSSLSASVGDRVTITCRASQSIGKYLNWYQQKPGKAPKLLIYA ASRLQSG VPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQMHPRAPKTFG QGTKVEIK

[0368] The Kc4 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a lambda light chain (SEQ ID NO: 74) encoded by the nucleic acid sequence shown in SEQ ID NO: 73.

[0369] KC4-LC-NT (SEQ ID NO: 73)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTGGTGGTTATAACT ATGTCTCCTG GTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATGAGGTCAGTAA TCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGCC TCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAGC TCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGACC GTCCTAGGTCAGCCCAAGGCTGC CCCCTCGGTCACTCTGTTCCCGCCCTCCTCTG AGGAGCTTCAAGCCAACA AGGCCACACTGGTGTGTCTCATAAGTGACTTCTACC CGGGAGCCGTGACA GTGGCTTGGAAAGCAGATAGCAGCCCCGTCAAGGCGGGA GTGGAGACCAC CACACCCTCCAAACAAAGCAACAACAAGTACGCGGCCAGCAG CTATCTGA GCCTGACGCCTGAGCAGTGGAAGTCCCACAGAAGCTACAGCTGCCA GGTC ACGCATGAAGGGAGCACCGTGGAGAAGACAGTGGCCCCTACAGAATGTT C ATAA

[0370] KC4-LC-AA (SEQ ID NO: 74) QSALTQPASVSGSPGQSITISCTGTSSDVGGYNYVSWYQQHPGKAPKLMI YEVSNRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVLGQ PKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVT VAWKADSSPVKAGVETTTPS KQSNNKYAASSYLSLTPEQWKSHRSYSCQV THEGSTVEKTVAPTECS

[0371] The Kc4 antibody includes a common variable heavy domain (SEQ ID

NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a lambda variable light domain (SEQ ID NO: 166) encoded by the nucleic acid sequence shown in SEQ ID NO: 165.

[0372] KC4-VL-NT (SEQ ID NO: 165)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTGGTGGTTATAACT ATGTCTCCTG GTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATGAGGTCAGTAA TCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGCC TCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAGC TCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGACC GTCCTA

[0373] KC4-VL-AA (SEQ ID NO: 166)

QSALTQPASVSGSPGQSITISCTGTSSDVGGYNYVSWYQQHPGKAPKLMI YEVSNRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVL

[0374] The Kc4Gll antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a lambda light chain (SEQ ID NO: 76) encoded by the nucleic acid sequence shown in SEQ ID NO: 75.

[0375] KC4G11-LC-NT (SEQ ID NO: 75)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTGGGAAGGCGAACT ATGTCTCCT GGTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATAAGGATAGTG ATCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGC CTCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAG CTCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGAC CGTCCTAGGTCAGCCCAAGGCTGC CCCCTCGGTCACTCTGTTCCCGCCCTCCTCT GAGGAGCTTCAAGCCAACA AGGCCACACTGGTGTGTCTCATAAGTGACTTCTAC CCGGGAGCCGTGACA GTGGCTTGGAAAGCAGATAGCAGCCCCGTCAAGGCGGG AGTGGAGACCAC CACACCCTCCAAACAAAGCAACAACAAGTACGCGGCCAGCA GCTATCTGA GCCTGACGCCTGAGCAGTGGAAGTCCCACAGAAGCTACAGCTGCC AGGTC ACGCATGAAGGGAGCACCGTGGAGAAGACAGTGGCCCCTACAGAATGT TC ATAA

[0376] KC4G11-LC-AA (SEQ ID NO: 76)

QSALTQPASVSGSPGQSITISCTGTSSDVGKANYVSWYQQHPGKAPKLMI YKDSDRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVLGQ PKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVT VAWKADSSPVKAGVETTTPS KQSNNKYAASSYLSLTPEQWKSHRSYSCQV THEGSTVEKTVAPTECS

[0377] The Kc4Gll antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a lambda variable light domain (SEQ ID NO: 168) encoded by the nucleic acid sequence shown in SEQ ID NO: 167.

[0378] KC4G11-VL-NT (SEQ ID NO: 167)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTGGGAAGGCGAACT ATGTCTCCT GGTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATAAGGATAGTG ATCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGC CTCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAG CTCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGAC CGTCCTA

[0379] KC4G11-VL-AA (SEQ ID NO: 168)

QSALTQPASVSGSPGQSITISCTGTSSDVGKANYVSWYQQHPGKAPKLMI YKDSDRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVL

[0380] The Kc4Cll antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a lambda light chain (SEQ ID NO: 78) encoded by the nucleic acid sequence shown in SEQ ID NO: 77.

[0381] KC4C11-LC-NT (SEQ ID NO: 77)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTAGGGGGAATAACT ATGTCTCCT GGTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATGAGAATAGTA AGCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGG CCTCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCA GCTCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGA CCGTCCTAGGTCAGCCCAAGGCTGC CCCCTCGGTCACTCTGTTCCCGCCCTCCTC TGAGGAGCTTCAAGCCAACA AGGCCACACTGGTGTGTCTCATAAGTGACTTCTA CCCGGGAGCCGTGACA GTGGCTTGGAAAGCAGATAGCAGCCCCGTCAAGGCGG GAGTGGAGACCAC CACACCCTCCAAACAAAGCAACAACAAGTACGCGGCCAGC AGCTATCTGA GCCTGACGCCTGAGCAGTGGAAGTCCCACAGAAGCTACAGCTGC CAGGTC ACGCATGAAGGGAGCACCGTGGAGAAGACAGTGGCCCCTACAGAATG TTC ATAA

[0382] KC4C11-LC-AA (SEQ ID NO: 78)

QSALTQPASVSGSPGQSITISCTGTSSDVRGNNYVSWYQQHPGKAPKLMI YENSKRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVLGQ PKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVT VAWKADSSPVKAGVETTTPS KQSNNKYAASSYLSLTPEQWKSHRSYSCQV THEGSTVEKTVAPTECS [0383] The Kc4Cll antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a lambda variable light domain (SEQ ID NO: 170) encoded by the nucleic acid sequence shown in SEQ ID NO: 169.

[0384] KC4C11-VL-NT (SEQ ID NO: 169)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTAGGGGGAATAACT ATGTCTCCT GGTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATGAGAATAGTA AGCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGG CCTCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCA GCTCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGA CCGTCCTA

[0385] KC4C11-VL-AA (SEQ ID NO: 170)

QSALTQPASVSGSPGQSITISCTGTSSDVRGNNYVSWYQQHPGKAPKLMI YENSKRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVL

[0386] The Kc4Al antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a lambda light chain (SEQ ID NO: 80) encoded by the nucleic acid sequence shown in SEQ ID NO: 79.

[0387] KC4A1-LC-NT (SEQ ID NO: 79)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTAGTGCGAGGAACT ATGTCTCCT GGTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATGAGAGTAGTA AGCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGG CCTCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCA GCTCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGA CCGTCCTAGGTCAGCCCAAGGCTGC CCCCTCGGTCACTCTGTTCCCGCCCTCCTC TGAGGAGCTTCAAGCCAACA AGGCCACACTGGTGTGTCTCATAAGTGACTTCTA CCCGGGAGCCGTGACA GTGGCTTGGAAAGCAGATAGCAGCCCCGTCAAGGCGG GAGTGGAGACCAC CACACCCTCCAAACAAAGCAACAACAAGTACGCGGCCAGC AGCTATCTGA GCCTGACGCCTGAGCAGTGGAAGTCCCACAGAAGCTACAGCTGC CAGGTC ACGCATGAAGGGAGCACCGTGGAGAAGACAGTGGCCCCTACAGAATG TTC ATAA

[0388] KC4A1-LC-AA (SEQ ID NO: 80)

QSALTQPASVSGSPGQSITISCTGTSSDVSARNYVSWYQQHPGKAPKLMI YESSKRPS GVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVLGQP KAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVT VAWKADSSPVKAGVETTTPSK QSNNKYAASSYLSLTPEQWKSHRSYSCQV THEGSTVEKTVAPTECS

[0389] The Kc4Al antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a lambda variable light domain (SEQ ID NO: 172) encoded by the nucleic acid sequence shown in SEQ ID NO: 171.

[0390] KC4A1-VL-NT (SEQ ID NO: 171)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTAGTGCGAGGAACT ATGTCTCCT GGTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATGAGAGTAGTA AGCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGG CCTCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCA GCTCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGA CCGTCCTA

[0391] KC4A1-VL-AA (SEQ ID NO: 172)

QSALTQPASVSGSPGQSITISCTGTSSDVSARNYVSWYQQHPGKAPKLMI YESSKRPS GVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVL

[0392] The Kc4A4 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a lambda light chain (SEQ ID NO: 82) encoded by the nucleic acid sequence shown in SEQ ID NO: 81.

[0393] KC4A4-LC-NT (SEQ ID NO: 81)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTAGAACCAGCAGTGACGTTAATAATACTAACT ATGTCTCCTG GTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATAAGACTAGTGG TCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGCC TCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAGC TCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGACC GTCCTAGGTCAGCCCAAGGCTGC CCCCTCGGTCACTCTGTTCCCGCCCTCCTCTG AGGAGCTTCAAGCCAACA AGGCCACACTGGTGTGTCTCATAAGTGACTTCTACC CGGGAGCCGTGACA GTGGCTTGGAAAGCAGATAGCAGCCCCGTCAAGGCGGGA GTGGAGACCAC CACACCCTCCAAACAAAGCAACAACAAGTACGCGGCCAGCAG CTATCTGA GCCTGACGCCTGAGCAGTGGAAGTCCCACAGAAGCTACAGCTGCCA GGTC ACGCATGAAGGGAGCACCGTGGAGAAGACAGTGGCCCCTACAGAATGTT C ATAA

[0394] KC4A4-LC-AA (SEQ ID NO: 82)

QSALTQPASVSGSPGQSITISCTRTSSDVNNTNYVSWYQQHPGKAPKLMI YKTSGRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVLGQ PKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVT VAWKADSSPVKAGVETTTPS KQSNNKYAASSYLSLTPEQWKSHRSYSCQV THEGSTVEKTVAPTECS

[0395] The Kc4A4 antibody includes a common variable heavy domain (SEQ ID

NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a lambda variable light domain (SEQ ID NO: 174) encoded by the nucleic acid sequence shown in SEQ ID NO: 173. [0396] KC4A4-VL-NT (SEQ ID NO: 173)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTAGAACCAGCAGTGACGTTAATAATACTAACT ATGTCTCCTG GTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATAAGACTAGTGG TCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGCC TCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAGC TCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGACC GTCCTA

[0397] KC4A4-VL-AA (SEQ ID NO: 174)

QSALTQPASVSGSPGQSITISCTRTSSDVNNTNYVSWYQQHPGKAPKLMI YKTSGRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVL

[0398] The Kc4E10 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a lambda light chain (SEQ ID NO: 84) encoded by the nucleic acid sequence shown in SEQ ID NO: 83.

[0399] KC4E10-LC-NT (SEQ ID NO: 83)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTAATTCTGCTAACT ATGTCTCCTG GTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATAAGAGTAGTAG TCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGCC TCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAGC TCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGACC GTCCTAGGTCAGCCCAAGGCTGC CCCCTCGGTCACTCTGTTCCCGCCCTCCTCTG AGGAGCTTCAAGCCAACA AGGCCACACTGGTGTGTCTCATAAGTGACTTCTACC CGGGAGCCGTGACA GTGGCTTGGAAAGCAGATAGCAGCCCCGTCAAGGCGGGA GTGGAGACCAC CACACCCTCCAAACAAAGCAACAACAAGTACGCGGCCAGCAG CTATCTGA GCCTGACGCCTGAGCAGTGGAAGTCCCACAGAAGCTACAGCTGCCA GGTC ACGCATGAAGGGAGCACCGTGGAGAAGACAGTGGCCCCTACAGAATGTT C ATAA

[0400] KC4E10-LC-AA (SEQ ID NO: 84)

QSALTQPASVSGSPGQSITISCTGTSSDVNSANYVSWYQQHPGKAPKLMI YKSSSRPS GVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVLGQP KAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVT VAWKADSSPVKAGVETTTPSK QSNNKYAASSYLSLTPEQWKSHRSYSCQV THEGSTVEKTVAPTECS

[0401] The Kc4E10 antibody includes a common variable heavy domain (SEQ ID

NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a lambda variable light domain (SEQ ID NO: 176) encoded by the nucleic acid sequence shown in SEQ ID NO: 175.

[0402] KC4E10-VL-NT (SEQ ID NO: 175)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTAATTCTGCTAACT ATGTCTCCTG GTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATAAGAGTAGTAG TCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGCC TCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAGC TCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGACC GTCCTA

[0403] KC4E10-VL-AA (SEQ ID NO: 176)

QSALTQPASVSGSPGQSITISCTGTSSDVNSANYVSWYQQHPGKAPKLMI YKSSSRPS GVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVL

[0404] The Kc4G9 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a lambda light chain (SEQ ID NO: 86) encoded by the nucleic acid sequence shown in SEQ ID NO: 85.

[0405] KC4G9-LC-NT (SEQ ID NO: 85)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCCGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTGAGAGGAAGAACT ATGTCTCCT GGTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATAAGAATAGTA CTCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGC CTCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAG CTCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGAC CGTCCTAGGTCAGCCCAAGGCTGC CCCCTCGGTCACTCTGTTCCCGCCCTCCTCT GAGGAGCTTCAAGCCAACA AGGCCACACTGGTGTGTCTCATAAGTGACTTCTAC CCGGGAGCCGTGACA GTGGCTTGGAAAGCAGATAGCAGCCCCGTCAAGGCGGG AGTGGAGACCAC CACACCCTCCAAACAAAGCAACAACAAGTACGCGGCCAGCA GCTATCTGA GCCTGACGCCTGAGCAGTGGAAGTCCCACAGAAGCTACAGCTGCC AGGTC ACGCATGAAGGGAGCACCGTGGAGAAGACAGTGGCCCCTACAGAATGT TC ATAA

[0406] KC4G9-LC-AA (SEQ ID NO: 86)

QSALTQPASVSGSPGQSITISCTGTSSDVERKNYVSWYQQHPGKAPKLMI YKNSTRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVLGQ PKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVT VAWKADSSPVKAGVETTTPS KQSNNKYAASSYLSLTPEQWKSHRSYSCQV THEGSTVEKTVAPTECS

[0407] The Kc4G9 antibody includes a common variable heavy domain (SEQ ID

NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a lambda variable light domain (SEQ ID NO: 178) encoded by the nucleic acid sequence shown in SEQ ID NO: 177.

[0408] KC4G9-VL-NT (SEQ ID NO: 177)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCCGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTGAGAGGAAGAACT ATGTCTCCT GGTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATAAGAATAGTA CTCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGC CTCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAG CTCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGAC CGTCCTA

[0409] KC4G9-VL-AA (SEQ ID NO: 178)

QSALTQPASVSGSPGQSITISCTGTSSDVERKNYVSWYQQHPGKAPKLMI YKNSTRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKETVE

[0410] The Kc4C3 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a lambda light chain (SEQ ID NO: 88) encoded by the nucleic acid sequence shown in SEQ ID NO: 87.

[0411] KC4C3-LC-NT (SEQ ID NO: 87)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTAGGGCGGCTAACT ATGTCTCCT GGTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATAAGAATAGTA CTCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGC CTCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAG CTCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGAC CGTCCTAGGTCAGCCCAAGGCTGC CCCCTCGGTCACTCTGTTCCCGCCCTCCTCT GAGGAGCTTCAAGCCAACA AGGCCACACTGGTGTGTCTCATAAGTGACTTCTAC CCGGGAGCCGTGACA GTGGCTTGGAAAGCAGATAGCAGCCCCGTCAAGGCGGG AGTGGAGACCAC CACACCCTCCAAACAAAGCAACAACAAGTACGCGGCCAGCA GCTATCTGA GCCTGACGCCTGAGCAGTGGAAGTCCCACAGAAGCTACAGCTGCC AGGTC ACGCATGAAGGGAGCACCGTGGAGAAGACAGTGGCCCCTACAGAATGT TC ATAA

[0412] KC4C3-LC-AA (SEQ ID NO: 88)

QSALTQPASVSGSPGQSITISCTGTSSDVRAANYVSWYQQHPGKAPKLMI YKNSTRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVLGQ PKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVT VAWKADSSPVKAGVETTTPS KQSNNKYAASSYLSLTPEQWKSHRSYSCQV THEGSTVEKTVAPTECS

[0413] The Kc4C3 antibody includes a common variable heavy domain (SEQ ID

NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a lambda variable light domain (SEQ ID NO: 180) encoded by the nucleic acid sequence shown in SEQ ID NO: 179.

[0414] KC4C3-VL-NT (SEQ ID NO: 179)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTAGGGCGGCTAACT ATGTCTCCT GGTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATAAGAATAGTA CTCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGC CTCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAG CTCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGAC CGTCCTA

[0415] KC4C3-VL-AA (SEQ ID NO: 180) QSALTQPASVSGSPGQSITISCTGTSSDVRAANYVSWYQQHPGKAPKLMI YKNSTRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVL

[0416] The Kc4F4 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a lambda light chain (SEQ ID NO: 90) encoded by the nucleic acid sequence shown in SEQ ID NO: 89.

[0417] KC4F4-LC-NT (SEQ ID NO: 89)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTAGGAGGGCTAACT ATGTCTCCT GGTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATCAGGATAGTA GTCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGC CTCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAG CTCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGAC CGTCCTAGGTCAGCCCAAGGCTGC CCCCTCGGTCACTCTGTTCCCGCCCTCCTCT GAGGAGCTTCAAGCCAACA AGGCCACACTGGTGTGTCTCATAAGTGACTTCTAC CCGGGAGCCGTGACA GTGGCTTGGAAAGCAGATAGCAGCCCCGTCAAGGCGGG AGTGGAGACCAC CACACCCTCCAAACAAAGCAACAACAAGTACGCGGCCAGCA GCTATCTGA GCCTGACGCCTGAGCAGTGGAAGTCCCACAGAAGCTACAGCTGCC AGGTC ACGCATGAAGGGAGCACCGTGGAGAAGACAGTGGCCCCTACAGAATGT TC ATAA

[0418] KC4F4-LC-AA (SEQ ID NO: 90)

QSALTQPASVSGSPGQSITISCTGTSSDVRRANYVSWYQQHPGKAPKLMI YQDSSRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVLGQ PKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVT VAWKADSSPVKAGVETTTPS KQSNNKYAASSYLSLTPEQWKSHRSYSCQV THEGSTVEKTVAPTECS

[0419] The Kc4F4 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a lambda variable light domain (SEQ ID NO: 182) encoded by the nucleic acid sequence shown in SEQ ID NO: 181.

[0420] KC4F4-VL-NT (SEQ ID NO: 181)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTAGGAGGGCTAACT ATGTCTCCT GGTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATCAGGATAGTA GTCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGC CTCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAG CTCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGAC CGTCCTA

[0421] KC4F4-VL-AA (SEQ ID NO: 182)

QSALTQPASVSGSPGQSITISCTGTSSDVRRANYVSWYQQHPGKAPKLMI YQDSSRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVL [0422] The Kc4Bl antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a lambda light chain (SEQ ID NO: 92) encoded by the nucleic acid sequence shown in SEQ ID NO: 91.

[0423] KC4B 1-LC-NT (SEQ ID NO: 91)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTAGGGCTAATAACT ATGTCTCCT GGTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATGAGAGTAGTG CGCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGC CTCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAG CTCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGAC CGTCCTAGGTCAGCCCAAGGCTGC CCCCTCGGTCACTCTGTTCCCGCCCTCCTCT GAGGAGCTTCAAGCCAACA AGGCCACACTGGTGTGTCTCATAAGTGACTTCTAC CCGGGAGCCGTGACA GTGGCTTGGAAAGCAGATAGCAGCCCCGTCAAGGCGGG AGTGGAGACCAC CACACCCTCCAAACAAAGCAACAACAAGTACGCGGCCAGCA GCTATCTGA GCCTGACGCCTGAGCAGTGGAAGTCCCACAGAAGCTACAGCTGCC AGGTC ACGCATGAAGGGAGCACCGTGGAGAAGACAGTGGCCCCTACAGAATGT TC ATAA

[0424] KC4B 1-LC-AA (SEQ ID NO: 92)

QSALTQPASVSGSPGQSITISCTGTSSDVRANNYVSWYQQHPGKAPKLMI YESSARP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVLGQ PKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVT VAWKADSSPVKAGVETTTPS KQSNNKYAASSYLSLTPEQWKSHRSYSCQV THEGSTVEKTVAPTECS

[0425] The Kc4B 1 antibody includes a common variable heavy domain (SEQ ID NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a lambda variable light domain (SEQ ID NO: 184) encoded by the nucleic acid sequence shown in SEQ ID NO: 183.

[0426] KC4B 1-VL-NT (SEQ ID NO: 183)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTAGGGCTAATAACT ATGTCTCCT GGTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATGAGAGTAGTG CGCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGC CTCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAG CTCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGAC CGTCCTA

[0427] KC4B 1-VL-AA (SEQ ID NO: 184)

QSALTQPASVSGSPGQSITISCTGTSSDVRANNYVSWYQQHPGKAPKLMI YESSARP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVL [0428] The Kc4E2 antibody includes a common heavy chain (SEQ ID NO: 2) encoded by the nucleic acid sequence shown in SEQ ID NO: 1 and includes a lambda light chain (SEQ ID NO: 94) encoded by the nucleic acid sequence shown in SEQ ID NO: 93.

[0429] KC4E2-LC-NT (SEQ ID NO: 93)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTTATTATAATAAGT ATGTCTCCTG GTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATGAGAGTAGTAA GCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGCC TCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAGC TCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGACC GTCCTAGGTCAGCCCAAGGCTGC CCCCTCGGTCACTCTGTTCCCGCCCTCCTCTG AGGAGCTTCAAGCCAACA AGGCCACACTGGTGTGTCTCATAAGTGACTTCTACC CGGGAGCCGTGACA GTGGCTTGGAAAGCAGATAGCAGCCCCGTCAAGGCGGGA GTGGAGACCAC CACACCCTCCAAACAAAGCAACAACAAGTACGCGGCCAGCAG CTATCTGA GCCTGACGCCTGAGCAGTGGAAGTCCCACAGAAGCTACAGCTGCCA GGTC ACGCATGAAGGGAGCACCGTGGAGAAGACAGTGGCCCCTACAGAATGTT C ATAA

[0430] KC4E2-LC-AA (SEQ ID NO: 94)

QSALTQPASVSGSPGQSITISCTGTSSDVYYNKYVSWYQQHPGKAPKLMI YESSKRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVLGQ PKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVT VAWKADSSPVKAGVETTTPS KQSNNKYAASSYLSLTPEQWKSHRSYSCQV THEGSTVEKTVAPTECS

[0431] NO: 114) encoded by the nucleic acid sequence shown in SEQ ID NO: 113 and includes a lambda variable light domain (SEQ ID NO: 186) encoded by the nucleic acid sequence shown in SEQ ID NO: 185.

[0432] KC4E2-VL-NT (SEQ ID NO: 185)

CAGTCTGCCCTGACTCAGCCTGCCTCCGTGTCTGGGTCTCCTGGACAGTC GATCA CCATCTCCTGCACTGGAACCAGCAGTGACGTTTATTATAATAAGT ATGTCTCCTG GTACCAACAGCACCCAGGCAAAGCCCCCAAACTCATGATT TATGAGAGTAGTAA GCGGCCCTCAGGGGTTTCTAATCGCTTCTCTGGCTC CAAGTCTGGCAACACGGCC TCCCTGACCATCTCTGGGCTCCAGGCTGAGG ACGAGGCTGATTATTACTGCAGC TCATATGATTGGTGGTTCCGCCCCAAG GTGTTCGGCGGAGGGACCAAGCTGACC GTCCTA

[0433] KC4E2-VL-AA (SEQ ID NO: 186)

QSALTQPASVSGSPGQSITISCTGTSSDVYYNKYVSWYQQHPGKAPKLMI YESSKRP SGVSNRFSGSKSGNTASLTISGLQAEDEADYYCSSYDWWFRPK VFGGGTKLTVL

[0434] Exemplary monoclonal antibodies of the disclosure include, for example, humanized antibodies having a variable heavy chain (VH) and/or variable light (VL) chain region shown in the sequences below.

[0435] SEQ ID NO: 187 EVQLQQSGAELVRSGASVKLSCTASQPN1KDYYLHWVKQRPEQGLEWIGWIDPDNG DTEFAPKFQGKATMTADTSSNTAYLQLSSLTSEDTAVYYCNAAYGSSSYPMDYWG QGTSVTV

[0436] SEQ ID NO: 188

EVQLVQSGAEVKKPGATVKISCKVSGFNIKDYYLHWVQQAPGKGLEWMGWIDPDN

GDTEYAEKFQGRVTITADTSTDTAYMELSSLRSEDTAVYYCNAAYGSSSYPMDYW GQGTSVTV

[0437] SEQ ID NO: 189

QMQLVQSGAEVKKTGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWIDPD

NGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0438] SEQ ID NO: 190

EVQLVQSGAEVKKTGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWIDPD

NGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0439] SEQ ID NO: 191

QMQLVQSGAEVKKTGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWWPD

NGDTEYAQKFQGRVTMTADTSSNTAYMELSSLRSEDTAMYYCNAAYGSSSYH1DY WGQGTTVTV

[0440] SEQ ID NO: 192

QMQLVQSGAEVKKTGSSVKVSCKASGPNIKDYYLHWVRQAPGQALEWMGWIDPD

NGDTEYAQKFQGRVTMTEDTSTDTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0441] SEQ ID NO: 193

QMQLVQSGAEVKKTGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWIDPD

QGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0442] SEQ ID NO: 194

QMQLVQSGAEVKKTGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWIDPD

YGDTEYAQFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDYW GQGTTVTV

[0443] SEQ ID NO: 195

QMQLVGSGAEVKKTGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWIDPD

SGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0444] SEQ ID NO: 196

QMQLVGSGAEVKKTGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWIDPD

NADTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0445] SEQ ID NO: 197 QMQLVGSGAEVKKTGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWIDPD NTDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0446] SEQ ID NO: 198

QMQLVGSGAEVKKTGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWIDPD

NTDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSPYPMDY WGQGTTVTV

[0447] SEQ ID NO: 199

QMQLVGSGAEVKKTGSSVKVSCKASGYTFTYYYLHWVRQAPGQALEWMGWIDPD

NGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSPYPMDY WGQGTTVTV

[0448] SEQ ID NO: 200

QMQLVGSGAEVKKTGSSVKVSCKASGFTFTYYYLHWVRQAPGQALEWMGWIDPD

NGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0449] SEQ ID NO: 201

QMQLVGSGAEVKKTGSSVKVSCKASGYNFTYYYLHWVRQAPGQALEWMGWIDPD

NGQTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0450] SEQ ID NO: 202

QMQLVGSGAEVKKTGSSVKVSCKASGYTITYYYLHWVRQAPGQALEWMGWIDPD

NGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0451] SEQ ID NO: 203

QMQLVGSGAEVKKTGSSVKVSCKASGYTFTDYYLHWVRQAPGQALEWMGWIDPD

NGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0452] SEQ ID NO: 204

QMQLVGSGAEVKKTGSSVKVSCKASGYTFTDYYLHWVRQAPGQALEWMGWIDPD

NGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0453] SEQ ID NO: 205

QMQLVGSGAEVKKTGSSVKVSCKASGFTFTDYYLHWVRQAPGQALEWMGWIDPD

NGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0454] SEQ ID NO: 206

QMQLVGSGAEVKKTGSSVKVSCKASGFTITDYYLHWVRQAPGQALEWMGWIDPD

NGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0455] SEQ ID NO: 207 QMQLVGSGAEVKKTGSSVKVSCKASGYTFKDYYLHWVRQAPGQALEWMGWIDPD NGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0456] SEQ ID NO: 208

QMQLVGSGAEVKKTGSSVKVSCKASGFTFKDYYLHWVRQAPGQALEWMGWIDPD

NGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0457] SEQ ID NO: 209

QMQLVGSGAEVKKTGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWIDPD

NGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0458] SEQ ID NO: 210

QMQLVGSGAEVKKTGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWIDPD

NGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0459] SEQ ID NO: 211

EVQLVGSGAEVKKTGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWIDPD

NGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0460] SEQ ID NO: 212

EVQLVGSGAEVKKTGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWIDPD

NGDTEYAQKFQDRVTITRDRSMSTAYMELSSLRSEDTAMYYCNAAYGSSSYPMDY WGQGTTVTV

[0461] In some embodiments, variable light (VL) chain regions of some CD47 antibodies are provided below:

[0462] SEQ ID NO: 213

DIKMTQSPSSLYASLGERVTITCKASQDIHRYLSWFQQKPGKSPKILIYRANRLVDGV

PSRFSGSGSGQDYSLTISSLEYEDMGIYYCLQYDEFPYTFGGGTKLEMK

[0463] SEQ ID NO: 214

DIKMTQSPSSLYASLGERVTITCKASQDIHRYLSWFQQKPGKSPKILIYRANRLVDGV

PSRFSGSGSGQDYSLTISSLEYEDMGIYYCLQYDEFPYTFGGGTKLEIK [0464] SEQ ID NO: 215

DIQMTGSPSSLSASVGDRVTITCKASQDTHRYLSWYQQKPKAPKLLIYRANPLVDGV

PSRFSGSGSGTDFTFTISSLQPEDIATYYCLQYDEFPYTFGGGTKVEIK [0465] SEQ ID NO: 216

DIQMTQSPSSLSASVGDRVTITCKASQDIHRYLSWFQQKPGKAPKSLIYRANRLVDG

VPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQYDEFPYTFGGGTKVEIK

[0466] SEQ ID NO: 217

NIQMTQSPSAMSASVGDRVTITCKASQDIHRYLSWFQQKPGKVPKHLIYRANRLVD

GVPSRFSGSGSGTEFTLTISSLQPEDFATYYCLQYDEFPYTFGGGTKVEIK [0467] SEQ ID NO: 218

DIQMTQSPSSLSASVGDRVTITCKASQDIHRYLSWYQQKPGKAPKRLIYRANRLVDG

VPSRFSGSGSGTEFTLTISSLQPEDFATYYCLQYDEFFYTFGGGTKVEIK

[0468] SEQ ID NO: 219

DIQMTQSPSSLSASVGDRVTITCRASQDIHRYLAWYQQKPGKVPKLLIYRANRLQSG

VPSRFSGSGSGTDFTLTISSLQPEDVATYYCLQYDEFPYTFGQGTKVEIK

[0469] SEQ ID NO: 220

EIVLTQSPATLSLSPGERATLSCRASQDIHRYLAWYQQKPGQAPRLLIYFANRRATGI

PARFSGSGSGTDFTLTISSLEPEDFAVYYCLQYDEFPYTGFQGTRLEIK

[0470] SEQ ID NO: 221

DIQMTQSPSAMSASVGDRVTITCKASQDIHRYLSWFQQKPGKVPKHLIYRANRLVD

GVPSRFSGSGSGTEFTLTISSLQPEDFATYYCLQYDEFPYTFGGGTKVEIK

[0471] SEQ ID NO: 222

NIQMTQSPSAMSASVGDRVTITCRARQGIHRYLSWFQQKPGKVPKHLIYRANRLVD

GVPSRFSGSGSGTEFTLTISSLQPEDFATYYCLQYDEFPYTFGGGTKVEIK

[0472] SEQ ID NO: 223

NIQMTQSPSAMSASVGDRVTITCKASQDTHRYLSWFQQKPGKVPKILIYRANRLVDG

VSRFSGSGSGTEFTLTISSLQPEDFATYYCLQYDFPYTFGGGTKVEIK

[0473] SEQ ID NO: 224

NIQMTQSPSAMSASVGDRVTITCKASQDIHRYLSWFQQKPGKVPKHLIYRANRLVSG

VPSRFSGSGSGTEFTLTISSLQPEDFATYYCLQYDEFPYTFGGGTKVEIK

[0474] SEQ ID NO: 225

NIQMTQSPSAMSASVGDRVTITCRARQGIHRYLSSWFQQKPGKVPKILIYRANRLVD

GVPSRFSGSGSGTEFTLTISSLQPEDFATYYCLQYDEFPYTFGGGTKVEIK

[0475] SEQ ID NO: 226

NIQMTQSPSAMSASVGDRVTITCRARQGIHRYLSWFQQKPGKVPKHLIYRANRLVSG

VPSRFSGSGSGTEFTLTISSLQPEDFATYYCLQYDEFPYTFGGGTKVEIK

[0476] SEQ ID NO: 227

NIQMTQSPSAMSASVGDRVTITCKASQDIHRYLSWFQQKPGKVPKLLIYRANRLVDG

CPSRFSGSGSGTEFTLTISSLQPEDFATYYCLQYDEFPYTFGGGTKVEIK

[0477] SEQ ID NO: 228

NIQMTQSPSAMSASVGDRVTITCKASQDIHRYLSWFQQKPGKVPKLLIYRANRLVSG

VPSRFSGSGSGTEFTLTISSLQPEDFATYYCLQYDEFPYTFGGGTKVEIK

[0478] SEQ ID NO: 229

NIQMTQSPSAMSASVGDRVTITCRARQGIHRYLSWFQQKPGKVPKLLIYRANRLVSG

VPSRFSGSGSGTEFTLTISSLQPEDFATYYCLQYDEFPYTFGGGTKVEIK

[0479] In some embodiments, the anti-CD47 antibody is SEQ ID NO: 230.

[0480] SEQ ID NO: 230 - B6H12.2 MWPLVAALLLGSACCGSAQLLFNKTKSVEFTFCNDTVVIPCFVTNMEAQNTTEVYV

KWKFKGRDIYTFDGALNKSTVPTDFSSAKIEVSQLLKGDASLKMDKSDAVSHTGNY

TCEVTELTREGETIIELKYRVVSWFSPNENILIVIFPIFAILLFWGQFGIKTLKYRSGGM

DEKTIALLVAGLVITVIVIVGAILFVPGEYSLKNATGLGLIVTSTGILILLHYYVFSTAI

GLTSFVIAILVIQVIAYILAVVGLSLCIAACIPMHGPLLISGLSILALAQLLGLVYMKFV

ASNQKTIQPPRKAVEEPLNAFKESKGMMNDE

[0481] In some embodiments, the anti-CD47 antibody is SEQ ID NO: 231.

[0482] SEQ ID NO: 231 - BRIC 126

MWPLVAALLLGSACCGSAQLLFNKTKSVEFTFCNDTVVIPCFVTNMEAQNTTEVYV

KWKFKGRDIYTFDGALNKSTVPTDFSSAKIEVSQLLKGDASLKMDKSDAVSHTGNY

TCEVTELTREGETIIELKYRVVSWFSPNENILIVIFPIFAILLFWGQFGIKTLKYRSGGM

DEKTIALLVAGLVITVIVIVGAILFVPGEYSLKNATGLGLIVTSTGILILLHYYVFSTAI

GLTSFVIAILVIQVIAYILAVVGLSLCIAACIPMHGPLLISGLSILALAQLLGLVYMKFV ASNQKTIQPPRKAVEEPLNAFKESKGMMNDE

[0483] In some embodiments, the anti-CD47 antibody is Hu5f9-g4. SEQ ID NO: 232.

[0484] SEQ ID NO: 232 - Hu5f9-g4

QVQLVQSGAEVKKPGASVKVSCKASGYTFTNYNMHWVRQAPGQRLEWMGTIYPG

NDDTSYNQKFKDRVTITADTSASTAYMELSSLRSEDTAVYYCARGGYRAMDYWGQ

GTLVTVSSDIVMTQSPLSLPVTGEPASISCRSSQSIVYSNGNTYLGWYLQKPGQSPQL

LIYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQGSHVPYTFGQGTK LEIK

[0485] In some embodiments, the anti-CD47 antibody is STI-6643. STI-6643 is described in U.S. Application Publication No. US 2014/0140989 Al, which is hereby incorporated by reference in its entirety.

[0486] SEQ ID NO: 233 - STI-6643

SVLTQPSSVSASPGQSITISCSGTSSDVGGHNYVSWYQQHPGKAPKLMIYDVTKRPS

GVPDRFSGSKSGNTASLTVSGLQAEDEADYYCLSYAGSRVYVFGTGTKLTVL

[0487] In some embodiments, the anti-CD47 antibody is Rituximab. SEQ ID

NO: 234.

[0488] SEQ ID NO: 234 - Rituximab

IVLSQSPAILSASPGEKVTMTCRASSSVSYIHWFQQKPGSSPKPWIYATSNLASGVPV

RFSGSGSGTSYSLTISRVEAEDAATYYCQQWTSNPPTFGGGTKLEIKRTVAAPSVFIFP

PSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSL SSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0489] In some embodiments, the anti-CD47 antibody is Obinutuzumab. SEQ ID

NO: 235.

[0490] SEQ ID NO: 235 - Obinutuzumab IVMTQTPLSLPVTPGEPASISCRSSKSLLHSNGITYLYWYLQKPGQSPQLLIYQMSNLV SGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCAQNLELPYTFGGGTKVEIKRTVAA PSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSK DSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0491] In some embodiments, the anti-CD47 antibody is Ofatumumab. SEQ ID NO: 236.

[0492] SEQ ID NO: 236 - Ofatumumab

IVLTQSPATLSLSPGERATLSCRASQSVSSYLAWYQQKPGQAPRLLIYDASNRATGIP ARFSGSGSGTDFTLTISSLEPEDFAVYYCQQRSNWPITFGQGTRLEIKRTVAAPSVFIF PPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYS LSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNR

[0493] In some embodiments, the anti-CD47 antibody is Daratumumab. SEQ ID NO: 237.

[0494] SEQ ID NO: 237 - Daratumumab

IVLTQSPATLSLSPGERATLSCRASQSVSSYLAWYQQKPGQAPRLLIYDASNRATGIP ARFSGSGSGTDFTLTISSLEPEDFAVYYCQQRSNWPPTFGQGTKVEIKRTVAAPSVFIF PPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYS LSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0495] In some embodiments, the anti-CD47 antibody is CC-90002. CC-90002 s another humanized IgG4 anti-CD47 antibody that inhibits CD47-SIRPa interaction and enabled phagocytosis in cancer cell lines, including hematological cancer cell lines, as well as in solid tumor xenografts. CC-90002 is described in Eladl et al, “Role of CD47 in Hematological Malignancies,” Journal of Hematology & Oncology, 13, 96 (2020), which is hereby incorporated by reference in its entirety.

[0496] In some embodiments, the anti-CD47 antibody is SRF231. SRF231 is a fully humanized monoclonal anti-CD47 antibody produced by phage technology. Data presented in an abstract showed it selectively blocks CD47- SIRP a, promoting phagocytosis of cancer cells and sparing T-cells and RBCs in vitro. SRF231 is described in Eladl et al, “Role of CD47 in Hematological Malignancies,” Journal of Hematology & Oncology, 13, 96 (2020), which is hereby incorporated by reference in its entirety.

[0497] In some embodiments, the anti-CD47 antibody is ALX148. ALX148 is generated by fusing an inactivated human IgGl Fc with a modified SIRPa DI domain. It blocks the CD47-SIRPa interaction and stimulates both innate and acquired immune responses, promoting DCs, macrophage, and T-cell responses. ALX148 is described in Eladl et al, “Role of CD47 in Hematological Malignancies,” Journal of Hematology & Oncology, 13, 96 (2020), which is hereby incorporated by reference in its entirety.

[0498] In some embodiments, the anti-CD47 antibody is KWAR23. KWAR23 is a blocking antibody to human SIRPa, and is inert when administered on its own, but enhances the effect of rituximab. KWAR23 is described in Eladl et al, “Role of CD47 in Hematological Malignancies,” Journal of Hematology & Oncology, 13, 96 (2020), which is hereby incorporated by reference in its entirety.

[0499] In some embodiments, the anti-CD47 antibody is C47B 116. In some embodiment, the anti-CD47 antibody is C47B91. In some embodiments the anti-CD47 antibody is C47B222. The antibodies C47B 116, C47B91, and C47B222 are described in Pietsch, et al, “Anti-leukemic activity and tolerability of anti-human CD47 monoclonal antibodies,” Blood Cancer Journal 7, e536 (2017), which is hereby incorporated by reference in its entirety.

[0500] In some embodiments, the anti-CD47 antibody is TJC4. TJC4 is a fully human anti-CD47 IgG4 antibody that shares a similar binding affinity to human and cynomolgus monkey CD47. Like other anti-CD47 antibodies, TJC4 blocks the interaction of CD47 and SIRPa, leading to the enhanced macrophage phagocytosis of various CD47+ tumor cell lines and primary AML cells. Mono-treatment of TJC4 completely eradicated tumor cells in a Raji cell xenograft model and significantly extended the overall survival of treated mice in an AML model. When combined with Rituximab, TJC4 showed a superior efficacy in a DLBCL model over the mono-treatment group. TJC4 is described in Meng et al, “TJC4, a Differentiated Anti-CD47 Antibody with Novel Epitope and RBC Sparing Properties,” Blood (2019), 134 (Supplement- 1): 4063, which is hereby incorporated by reference in its entirety.

[0501] In aspects provided herein is an antibody, e.g. a monoclonal antibody, which specifically binds to human CD47, wherein such an anti-CD47 antibody is a variant of a parental anti-CD47 antibody, wherein the anti-CD47 antibody, when produced using a cell- free (CF) expression system, has a higher antibody expression titer or yield compared to that of the parental anti-CD47 antibody when expressed in the CF system, and wherein the anti- CD47 antibody comprises a VH comprising SEQ ID NO: 22. In certain aspects, such anti- CD47 antibody is an IgGl, IgG2, IgG3, or IgG4 isotype antibody. In certain aspects, such anti-CD47 antibody is an IgGl isotype antibody. In certain aspects, such anti-CD47 antibody is an IgGl Z allotype isotype antibody. In certain aspects, such anti-CD47 antibody is an IgG4, such as an IgG4P or IgG4PE, isotype antibody.

[0502] SEQ ID NO: 238 - C47B161

QLLFNKTKSVEFTFCNDTVVIPCFVTNMEAQNTTEVYVKWKFKGRDIYTFDGALNK STVPTDFSSAKIEVSQLLKGDASLKMDKSDAVSHTGNYTCEVTELTREGETIIELKYR VVSWFSPNE

[0503] SEQ ID NO: 239 - C47B222

QLLFNKTKSVEFTFCNDTVVIPCFVTNMEAQNTTEVYVKWKFKGRDIYTFDGALNK STVPTDFSSAKIEVSQLLKGDASLKMDKDAVSHTGNYTCEVTELTREGETIIELKYRV VSWFSPNE

[0504] In a particular aspect, an anti-CD47 antibody (IgGl- 13m) provided herein comprises an IgGl heavy chain comprising the amino acid sequence as set forth below:

[0505] SEQ ID NO: 240

QVQLVQSGAEVKKPGASVKVSCKASGFNIKDYYLHWVRQAPGQGLEWMGWIDPD QGDTEYAQKLQGRVTMTTDTSTSTAYMELRSLRSDDTAVYYCNAAYGSSSYPMDY WGQGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGAL TSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSC DKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW YVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPI EKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPEN NYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSP GK

[0506] In a particular aspect, an anti-CD47 antibody (IgGl-13mZ) provided herein comprises an IgGl-Z allotype heavy chain comprising the amino acid sequence as set forth below:

[0507] SEQ ID NO: 241

MQVQLVQSGAEVKKPGASVKVSCKASGFNIKDYYLHWVRQAPGQGLEWMGWIDP DQGDTEYAQKLQGRVTMTTDTSTSTAYMELRSLRSDDTAVYYCNAAYGSSSYPMD YWGQGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSG ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKS CDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFN WYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPA PIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPE NNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLS PGK

[0508] In a particular aspect, an anti-CD47 antibody (IgGl -5m) provided herein comprises an IgGl heavy chain comprising the amino acid sequence as set forth below:

[0509] SEQ ID NO: 242 QMQLVQSGAEVKKPGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWIDPD

QGDTEYAQKFQGRVTITRDRSTSTAYMELRSLRSEDTAVYYCNAAYGSSSYPMDY

WGQGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGAL

TSGVHTFPAVEQSSGEYSESSVVTVPSSSEGTQTYICNVNHKPSNTKVDKKVEPKSC

DKTHTCPPCPAPEEEGGPSVFEFPPKPKDTEMISRTPEVTCVVVDVSHEDPEVKFNW

YVDGVEVHNAKTKPREEQYNSTYRVVSVETVEHQDWENGKEYKCKVSNKAEPAPI

EKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPEN

NYKTTPPVEDSDGSFFEYSKETVDKSRWQQGNVFSCSVMHEAEHNHYTQKSESESP GK

[0510] In a particular aspect, an anti-CD47 antibody (IgG4P-13m) provided herein comprises an IgG4P antibody comprising the amino acid sequence as set forth below:

[0511] SEQ ID NO: 243

QVQLVQSGAEVKKPGASVKVSCKASGFNIKDYYLHWVRQAPGQGLEWMGWIDPD

QGDTEYAQKEQGRVTMTTDTSTSTAYMEERSERSDDTAVYYCNAAYGSSSYPMDY

WGQGTTVTVSSASTKGPSVFPEAPCSRSTSESTAAEGCEVKDYFPEPVTVSWNSGAE

TSGVHTFPAVEQSSGEYSESSVVTVPSSSEGTKTYTCNVDHKPSNTKVDKRVESKYG

PPCPPCPAPEFEGGPSVFEFPPKPKDTEMISRTPEVTCVVVDVSQEDPEVQFNWYVDG

VEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISK

AKGQPREPQVYTEPPSQEEMTKNQVSETCEVKGFYPSDIAVEWESNGQPENNYKTT

PPVEDSDGSFFEYSRETVDKSRWQEGNVFSCSVMHEAEHNHYTQKSESESEGK

[0512] In a particular aspect, an anti-CD47 antibody (IgG4P-5m) provided herein comprises an IgG4P heavy chain comprising the amino acid sequence as set forth below:

[0513] SEQ ID NO: 244

QMQLVQSGAEVKKPGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWIDPD

QGDTEYAQKFQGRVTITRDRSTSTAYMELRSLRSEDTAVYYCNAAYGSSSYPMDY

WGQGTTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGAL

TSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYG

PPCPPCPAPEFLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDG

VEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISK

AKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTT

PPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGK

[0514] In a particular aspect, an anti-CD47 antibody (IgG4PE-13m) provided herein comprises an IgG4PE heavy chain comprising the amino acid sequence as set forth below:

[0515] SEQ ID NO: 245

QVQLVQSGAEVKKPGASVKVSCKASGFNIKDYYLHWVRQAPGQGLEWMGWIDPD

QGDTEYAQKLQGRVTMTTDTSTSTAYMELRSLRSDDTAVYYCNAAYGSSSYPMDY

WGQGTTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGAL

TSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYG

PPCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQNWYVDG

VEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISK AKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTT PPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGK

[0516] In a particular aspect, an anti-CD47 antibody (IgG4PE-5m) provided herein comprises an IgG4PE heavy chain comprising the amino acid sequence as set forth below:

[0517] SEQ ID NO: 246

QMQLVQSGAEVKKPGSSVKVSCKASGFNIKDYYLHWVRQAPGQALEWMGWIDPD QGDTEYAQKFQGRVTITRDRSTSTAYMELRSLRSEDTAVYYCNAAYGSSSYPMDY WGQGTTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGAL TSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYG PPCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDG VEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISK AKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTT PPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGK

[0518] In a specific aspect, provided herein is an antibody, e.g. a monoclonal antibody, which specifically binds to human CD47, wherein such an anti-CD47 antibody is a variant of a parental anti-CD47 antibody, wherein the anti-CD47 antibody, when produced using a cell-free (CF) expression system, has a higher antibody expression titer or yield compared to that of the parental anti-CD47 antibody when expressed in the CF system, and wherein the anti-CD47 antibody comprises a light chain comprising a kappa or lambda light chain constant region (e.g., human kappa or lambda light chain constant region), for example SEQ ID NO: 507.

[0519] In a specific aspect, provided herein is an antibody, e.g. a monoclonal antibody, which specifically binds to human CD47, wherein such an anti-CD47 antibody is a variant of a parental anti-CD47 antibody, wherein the anti-CD47 antibody, when produced using a cell-free (CF) expression system, has a higher antibody expression titer or yield compared to that of the parental anti-CD47 antibody when expressed in the CF system, and wherein the anti-CD47 antibody comprises (i) a VH described herein or a heavy chain described herein, and (ii) a light chain comprising a kappa or lambda light chain constant region (e.g., human kappa or lambda light chain constant region), for example SEQ ID NO: 507, e.g., as set forth below (anti-CD47 antibody light chain (Ig K)), or SEQ ID NO: 507 without the amino acid M at the N-terminus:

[0520] SEQ ID NO: 247 NIQMTQSPSAMSASVGDRVTITCKASQDIHRYLSWFQQKPGKVPKHLIYRANRLVSG VPSRFSGSGSGTEFTLTISSLQPEDFATYYCLQYDEFPYTFGGGTKVEIKRTVAAPSVF IFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST YSLSSTLTLSKADYEKHKVYACEVTHQ GLSSPVTKSFNRGEC

[0521] In a specific embodiment, an anti-CD47 described herein is an anti- CD47 antibody described in U.S. Application Publication No. US 2014/0140989 Al, which is hereby incorporated by reference in its entirety, for example any one of anti- CD47 antibodies in Table 1 of the publication (e.g., anti-CD47 antibody 2A1, AB2.03, AB2.04, AB2.05, AB2.06, AB2.07, AB2.08, AB2.09, AB2.13, AB3.09, AB6.12, AB6.13, AB6.14, AB6.17, AB10.13, AB10.14, AB11.05, AB 12.05, AB15.05, AB16.05, AB 17.05, AB22.05, AB23.05, AB24.05, and AB25.05), or any antibody comprising any of SEQ ID NOS: 5-30 of the publication.

[0522] In some embodiments, the human IgG4 constant region is modified within the hinge region to prevent or reduce strand exchange, e.g., Ser228Pro (S228P). In other embodiments, the human IgG4 constant region is modified at amino acid 235 to alter Fc receptor interactions, e.g., Leu235Glu (L235E). In some embodiments, the human IgG4 constant region is modified within the hinge and at amino acid 235, e.g., Ser228Pro and Leu235Glu (S228P/L235E). In some embodiments, the human IgG4 constant region is modified at amino acid Asn297 (Kabat Numbering) to prevent to glycosylation of the antibody, e.g., Asn297Ala (N297A). In some embodiments of the invention, the human IgG4 constant region is modified at amino acid positions Ser228, Leu235, and Asn297 (e.g., S228P/L235E/N297A). (EU index of Kabat et al 1991 Sequences of Proteins of Immunological Interest). In other embodiments of the invention, the antibody is of human IgG4 subclass and lacks glycosylation. In these embodiments the glycosylation can be eliminated by mutation at position 297 (Kabat numbering), for example N297A. In other embodiments, the glycosylation can be eliminated by production of the antibody in a host cell that lacks the ability for post-translational glycosylation, for example a bacterial or yeast derived system or a modified mammalian cell expression system.

[0523] In some embodiments, the human IgG constant region is modified to alter antibody-dependent cellular cytotoxicity (ADCC) and/or complement-dependent cytotoxicity (CDC), e.g., the amino acid modifications described in Natsume et al., 2008 Cancer Res, 68(10): 3863-72; Idusogie et al., 2001 J Immunol, 166(4): 2571-5; Moore et al., 2010 mAbs, 2(2): 181-189; Lazar et al., 2006 PNAS, 103(11): 4005-4010, Shields et al., 2001 JBC, 276(9): 6591-6604; Stavenhagen et al., 2007 Cancer Res, 67(18): 8882-8890; Stavenhagen et al., 2008 Advan. Enzyme Regul., 48: 152-164; Alegre et al, 1992 J Immunol, 148: 3461- 3468; Reviewed in Kaneko and Niwa, 2011 Biodrugs, 25(1): 1-11.

[0524] In certain aspects, anti-CD47 antibodies provided herein exhibit one or more desirable characteristics, such as, by way of non-limiting example, blocking of the interaction between CD47 and its ligand SIRPa and/or promoting (e.g., inducing or increasing) phagocytosis, without promoting (e.g., inducing or increasing) hemagglutination of erythrocytes, as well as anti-tumor activity. For example, anti-CD47 antibodies provided herein block at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 95%, or at least 99% of the interaction between CD47 and SIRPa as compared to the level of interaction between CD47 and SIRPa in the absence of the anti-CD47 antibody described herein.

[0525] In specific aspects, anti-CD47 antibodies described herein promote (e.g., induce or increase) phagocytosis of cells, e.g., CD47-expressing cells (e.g., CCRF-CEM cells), for example, by macrophages. In one aspect, the level of phagocytosois in the presence of anti-CD47 antibodies described herein is increased by at least 5%, at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 99%, at least 150%, at least 200%, compared to the level of phagocytosis in the presence of anti-CD47 antibodies described herein.

[0526] In specific aspects, anti-CD47 antibodies described herein do not promote (e.g, induce or increase), or cause a significant level of, agglutination of cells, e.g., anti- CD47 antibodies described herein do not promote (e.g, induce or increase), or cause a significant level of, hemagglutination of red blood cells. In one aspect, the level of agglutination in the presence of anti-CD47 antibodies described herein is reduced by at least 5%, at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 99% compared to the level of agglutination in the presence of anti-CD47 antibodies known to induce agglutination, such as MCA911 mouse anti-human CD47 antibody (BRIC126). In some aspects, anti-CD47 antibodies described herein do not promote (e.g., induce or increase), or cause a significant level of, agglutination if the level of agglutination in the presence of anti-CD47 antibodies described herein is reduced by at least 5%, at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 99% compared to the level of agglutination in the presence of existing anti-CD47 antibodies known to induce agglutination, such as MCA911 mouse anti-human CD47 antibody (BRIC126).

Pharmaceutical Compositions

[0527] Standard pharmaceutical formulation techniques can be used to make the pharmaceutical compositions described herein, such as those disclosed in Remington's The Science and Practice of Pharmacy, 21st Ed., Lippincott Williams & Wilkins (2005), incorporated herein by reference in its entirety. Accordingly, some embodiments include pharmaceutical compositions comprising: (a) a safe and therapeutically effective amount of a compound of Formula (I) or pharmaceutically acceptable salts thereof; (b) one or more monoclonal antibodies that bind to CD47 or an immunologically active fragment of one or more monoclonal antibodies that bind to CD47 and (c) a pharmaceutically acceptable carrier, diluent, excipient or combination thereof. In some embodiments, a compound of Formula (I) is selected from plinabulin, (3Z,6Z)-3-(phenyl-2, 3,4,5, 6-d5)-methylene-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene)piperazine-2, 5-dione; (3Z, 6Z)-3-(phenyl-2, 3,4,5, 6-ds)-methylene-d- 6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene)piperazine-2, 5-dione; (3Z,6Z)-3-

(phenylmethylene-d)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(phenyl-2,3,4,5,6-d5)-methylene-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene- d)piperazine-2, 5-dione; (3Z,6Z)-3-(phenylmethylene)-6-((5-(tert-butyl)-lH-imidazol-4- yl)methylene-d)piperazine-2, 5-dione; (3Z, 6Z)-3-(phenyl-2, 3,4,5, 6-ds)-methylene-d-6-((5- (tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(4-Fluoro- (phenyl-2,3,5,6-d4))-methylene-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene)piperazine- 2, 5-dione; (3Z,6Z)-3-(4-Fluoro-(phenyl-2,3,5,6-d4))-methylene-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-fluorobenzylidene)-6-((5- ( tert-butyl)- lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3- benzoylbenzylidene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-(4-fluorobenzoyl)benzylidene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene- d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-(4-methoxybenzoyl)benzylidene)-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-methoxybenzylidene)-6-((5- ( tert-butyl)- lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; or (3Z,6Z)-3-(3- (trifluoromethyenzydene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2,5- dione

[0528] The term “pharmaceutically acceptable carrier” or “pharmaceutically acceptable excipient” includes any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents and the like. 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 ingredient, its use in the therapeutic compositions is contemplated. In addition, various adjuvants such as are commonly used in the art may be included. Considerations for the inclusion of various components in pharmaceutical compositions are described, e.g., in Gilman et al. (Eds.) (1990); Goodman and Gilman’s: The Pharmacological Basis of Therapeutics, 8th Ed., Pergamon Press, which is incorporated herein by reference in its entirety.

[0529] Some examples of substances, which can serve as pharmaceutically- acceptable carriers or components thereof, are sugars, such as lactose, glucose and sucrose; starches, such as com starch and potato starch; cellulose and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose, and methyl cellulose; powdered tragacanth; malt; gelatin; talc; solid lubricants, such as stearic acid and magnesium stearate; calcium sulfate; vegetable oils, such as peanut oil, cottonseed oil, sesame oil, olive oil, corn oil and oil of theobroma; polyols such as propylene glycol, glycerine, sorbitol, mannitol, and polyethylene glycol; alginic acid; emulsifiers, such as the TWEENS; wetting agents, such sodium lauryl sulfate; coloring agents; flavoring agents; tableting agents, stabilizers; antioxidants; preservatives; pyrogen-free water; isotonic saline; and phosphate buffer solutions.

Method of Treatment

[0530] In aspects, methods for treating, managing, reducing, ameliorating, or preventing a disease or condition are provided herein. Some embodiments relate to a method for treating, managing, reducing, ameliorating, or preventing a disease or condition by coadministering a compound of Formula (I) and one or more anti-CD47 agents to a subject in need thereof. The term “co-administering” does not restrict the order in which therapies are administered. The therapies may be administered, e.g., serially, sequentially, concurrently, or concomitantly. As used herein, the terms “manage,” “managing,” and “management” refer to the beneficial effects that a subject derives from a therapy (e.g., a prophylactic or therapeutic agent), which does not result in a cure of a condition associated with CD47. In certain embodiments, a subject is administered one or more therapies (e.g., prophylactic or therapeutic agents, such as an antibody described herein) to “manage” a condition or disorder described herein, one or more symptoms thereof, so as to prevent the progression or worsening of the condition or disorder. As used herein, the terms “reduce” or “reducing” in the context of a condition or disorder provided herein (e.g., autoimmune disorder, immunological disorder, cancer, or inflammation) refer to the total or partial inhibition (e.g., less than 100%, 95%, 90%, 80%, 70%, 60%, 50%, 40%, 30%, 20%, 10%, or 5%) or blockage of the development, recurrence, onset or spread of a condition or disorder provided herein (e.g., cancer, metastasis, or angiogenesis) and/or symptom related thereto, resulting from the administration of a therapy or combination of therapies provided herein (e.g., a combination of prophylactic or therapeutic agents, such as an antibody described herein).

[0531] Some embodiments relate to a method for treating cancer using the pharmaceutical composition described herein to a subject in need thereof. Some embodiments relate to a method for treating cancer, comprising co-administering a compound of Formula (I) and one or more anti-CD47 agents. In some embodiments, the compound of Formula (I) is selected from plinabulin, (3Z, 6Z)-3-(phenyl-2, 3,4,5, 6-ds)- methylene-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene)piperazine-2, 5-dione; (3Z,6Z)-3- (phenyl-2,3,4,5,6-d5)-methylene-d-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene)piperazine- 2, 5-dione; (3Z,6Z)-3-(phenylmethylene-d)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene- d)piperazine-2, 5-dione; (3Z,6Z)-3-(phenyl-2,3,4,5,6-d5)-methylene-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(phenylmethylene)-6-((5-(tert- butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z, 6Z)-3-(phenyl-2, 3,4,5, 6-ds)- methylene-d-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)- 3-(4-Fluoro-(phenyl-2,3,5,6-d4))-methylene-6-((5-(tert-butyl)-lH-imidazol-4- yl)methylene)piperazine-2, 5-dione; (3Z,6Z)-3-(4-Fluoro-(phenyl-2,3,5,6-d4))-methylene-6- ((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3- fluorobenzylidene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-benzoylbenzylidene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene- d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-(4-fluorobenzoyl)benzylidene)-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-(4- methoxybenzoyl)benzylidene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-

2, 5-dione; (3Z,6Z)-3-(3-methoxybenzylidene)-6-((5-(tert-butyl)-lH-imidazol-4- yl)methylene-d)piperazine-2, 5-dione; or (3Z,6Z)-3-(3-(trifluoromethyenzydene)-6-((5-(tert- butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione. In some embodiments, the one or more anti-CD47 agents include one or more monoclonal antibodies that bind to CD47 to a subject in need thereof. In some embodiments, the subject can be an animal, e.g., a mammal, a human. In some embodiments, the subject is a human. In some embodiments, the monoclonal antibody specifically binds to CD47 is an isolated monoclonal antibody. In some embodiments, the one or more anti-CD47 agent is an antibody or antigen-binding fragment that specifically binds to SIRPa. In some embodiments, the monoclonal antibody that specifically binds to SIRPa is an isolated monoclonal antibody. In some embodiments, the monoclonal antibody promotes macrophage-mediated phagocytosis of a CD47- expressing cell. In some embodiments, the one or more anti-CD47 agents is a soluble CD-47 binding SIRPa fragment. In some embodiments, the one or more anti-CD47 agents prevents interaction between CD47 and SIRPa. In some embodiments, the anti-CD47 agent is an IgG isotype selected from the group consisting of IgG 1 isotype, IgG2 isotype, IgG3 isotype, IgG4 isotype, IG-G1-N297Q, IG4-S228P, and IG64 PE. In some embodiments, the cancer is selected from non-Hodgkin lymphoma. In some embodiments, plinabulin is administered at a dose from about 5 mg/m² to 150 mg/m². In some embodiments, the plinabulin is administered orally, sublingually, buccally, subcutaneously, intravenously, intranasally, intratumorally, topically, transdermally, intradermally, intraperitoneally, intramuscularly, intrapulmonarilly, vaginally, rectally, or intraocularly. In some embodiments, plinabulin is administered in combination with radiation. In some embodiments, plinabulin is administering in combination with a chemotherapy agent. In some embodiments, the plinabulin is administered once a week. In some embodiments, the plinabulin is administered once on each of day 1 and day 8 of a three-week (21 day) treatment cycle.

[0532] Some aspects of the disclosure provide for a method of treating a subject with a cancer cell expressing CD-47. In some embodiments, the method comprises detecting tumor cell expression of CD-47 in a subject, and providing the subject a therapeutic amount of plinabulin and an anti-CD47 agent. In some embodiments, detecting a tumor cell expression of CD-47 comprises at least one of immunocytochemistry, proteomics, mRNA quantification, or a combination thereof, used to detect tumor cell expression of CD-47 in a subject.

[0533] In some embodiments, a disease or disorder associated with aberrant CD47 expression, activity, and/or signaling in a subject in need thereof by co-administering an anti-CD47 antibody and a compound of Formula (I) to a subject in need thereof. Diseases or disorders related to aberrant CD47 expression, activity and/or signaling include, by way of non-limiting example, hematological cancer and/or solid tumors. Hematological cancers include, e.g., leukemia, lymphoma and myeloma. Certain forms of leukemia include, by way of non-limiting example, acute lymphocytic leukemia (ALL); acute myeloid leukemia (AML); chronic lymphocytic leukemia (CLL); chronic myelogenous leukemia (CML); Myeloproliferative disorder/neoplasm (MPDS); and myelodysplasia syndrome. Certain forms of lymphoma include, by way of non-limiting example, Hodgkin's lymphoma, both indolent and aggressive non-Hodgkin's lymphoma, Burkitt's lymphoma, and follicular lymphoma (small cell and large cell). Certain forms of myeloma include, by way of non-limiting example, multiple myeloma (MM), giant cell myeloma, heavy-chain myeloma, and light chain or Bence-Jones myeloma. Solid tumors include, e.g., breast tumors, ovarian tumors, lung tumors, pancreatic tumors, prostate tumors, melanoma tumors, colorectal tumors, lung tumors, head and neck tumors, bladder tumors, esophageal tumors, liver tumors, and kidney tumors. Symptoms associated with cancers and other neoplastic disorders include, for example, inflammation, fever, general malaise, fever, pain, often localized to the inflamed area, loss of appetite, weight loss, edema, headache, fatigue, rash, anemia, muscle weakness, muscle fatigue and abdominal symptoms such as, for example, abdominal pain, diarrhea or constipation.

[0534] In some embodiments, a method for treating, delaying the progression of, impeding, or alleviating a symptom of cancer in a subject in need thereof by coadministering a compound of Formula (I) and an anti-CD47 antibody to the subject in need thereof. For example, the CD47 antibodies described herein are useful in treating hematological malignancies and/or tumors, e.g., hematological malignancies and/or tumors. For example, the CD47 antibodies described herein are useful in treating CD47+ tumors. By way of non-limiting example, the CD47 antibodies described herein are useful in treating non-Hodgkin's lymphoma (NHL), acute lymphocytic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), chronic myelogenous leukemia (CML), multiple myeloma (MM), breast cancer, ovarian cancer, head and neck cancer, bladder cancer, melanoma, colorectal cancer, pancreatic cancer, lung cancer, leiomyoma, leiomyosarcoma, glioma, glioblastoma, and so on. Solid tumors include, e.g., breast tumors, ovarian tumors, lung tumors (e.g., NSCLC), pancreatic tumors, prostate tumors, melanoma tumors, colorectal tumors, lung tumors, head and neck tumors, bladder tumors, esophageal tumors, liver tumors (e.g., hepatocellular carcinoma), sarcoma, and kidney tumors.

[0535] In an embodiment, a method of treating cancer (e.g., a hematological disorder/cancer or solid cancer) in a subject comprises administering (e.g., administering concurrently or sequentially) to a subject in need thereof (i) an anti-CD47 antibody described herein or antigen-binding fragment thereof which specifically binds to CD47 such as human CD47, (ii) a compound of Formula (I); and (iii) another anti-cancer agent. In certain embodiments, the anti-cancer agent is a chermotherapeutic agent (e.g., microtubule disassembly blocker, antimetabolite, topisomerase inhibitor, and DNA crosslinker or damaging agent). In certain embodiments, the anti-cancer agent is a tyrosine kinase inhibitor (e.g., GLEEVEC® (imatinib mesylate) or SUTENT® (SU11248 or Sunitinib)). Other nonlimiting examples of tyrosine kinse inhibitors include 706 and AMNI07 (nilotinib). RADOOI, PKC412, gefitinib (IRESSA™), erlotinib (TARCEVA®), sorafenib (NEXAVAR®), pazopanib (VOTRIENT™), axitinib, bosutinib, cediranib (RECENTIN®), SPRYCEL® (dasatinib), lapatinib (TYKERB®), lestaurtinib, neratinib, nilotinib (TASIGNA®), semaxanib, toceranib (PALLADIA™), vandetanib (ZACTIMA™), and vatalanib.

[0536] In a specific aspect, provided herein is a method of treating cancer (e.g., a hematological disorder/cancer or solid cancer) in a subject comprising administering (e.g., administering concurrently or sequentially) to a subject in need thereof (i) an anti- CD47 antibody described herein or antigen-binding fragment thereof which specifically binds to CD47 such as human CD47, (ii) a compound of Formula (I) and (iii) radiation therapy. [0537] In a particular aspect, provided herein is a method of reducing tumor volume, comprising contacting an effective amount of an anti-CD47 antibody described herein which specifically binds to human CD47 with the tumor and a compound of Formula (I). Also provided herein is a method of reducing tumor volume in a subject in need thereof (e.g., a subject with a tumor, such as a CD47 expressing tumor), comprising administering to the subject an effective amount of an anti-CD47 antibody described herein which specifically binds to human CD47 and a compound of Formula (I).

[0538] In a particular aspect, provided herein is a method of inhibiting cancer cell growth or proliferation, comprising contacting an effective amount of an anti-CD47 antibody described herein which specifically binds to human CD47 with cancer cells and a compound of Formula (I). Also provided herein is a method of inhibiting cancer cell growth or proliferation in a subject in need thereof (e.g., a subject with cancer cells, such as CD47 expressing cancer cells), comprising administering to the subject an effective amount of an anti-CD47 antibody described herein which specifically binds to human CD47 and a compound of Formula (I).

[0539] In some embodiments, plinabulin and an anti-CD47 agent are administered in the same formulation. Other embodiments include co-administering a compound of Formula (I) and one or more monoclonal antibodies that bind to CD47 in separate compositions. Thus, some embodiments include a first pharmaceutical compositions comprising: (a) a safe and therapeutically effective amount of a compound of Formula (I) or pharmaceutically acceptable salts thereof and (b) a pharmaceutically acceptable carrier, diluent, excipient or combination thereof; and a second pharmaceutical composition comprising: (a) one or more monoclonal antibodies that bind to CD47 and (b) a pharmaceutically acceptable carrier, diluent, excipient or combination thereof.

[0540] In some embodiments, cancer is head and neck cancer, lung cancer, stomach cancer, colon cancer, pancreatic cancer, prostate cancer, breast cancer, kidney cancer, bladder cancer, ovary cancer, cervical cancer, melanoma, glioblastoma, myeloma, lymphoma, hematological cancer, or leukemia. In some embodiments, the cancer is renal cell carcinoma, malignant melanoma, non-small cell lung cancer (NSCLC), ovarian cancer, Hodgkin's lymphoma or squamous cell carcinoma. In some embodiments, the cancer is selected from breast cancer, colon cancer, rectal cancer, lung cancer, prostate cancer, melanoma, leukemia, ovarian cancer, gastric cancer, renal cell carcinoma, liver cancer, pancreatic cancer, lymphomas and myeloma. In some embodiments, the cancer is a solid tumor or hematological cancer.

[0541] Some embodiments include using the compositions described herein to diagnose, prognose, monitor, treat, alleviate, and/or prevent a disease or pathology associated with aberrant CD47 expression, activity and/or signaling in a subject. A therapeutic regimen is carried out by identifying a subject, e.g., a human patient suffering from (or at risk of developing) a disease or disorder associated with aberrant CD47 expression, activity and/or signaling, e.g., a cancer or other neoplastic disorder, using standard methods. In some embodiments, the diseases or disorders related to aberrant CD47 expression, activity and/or signaling include, by way of non-limiting example, hematological cancer and/or solid tumors. Hematological cancers include, e.g., leukemia, lymphoma and myeloma. Certain forms of leukemia include, by way of non-limiting example, acute lymphocytic leukemia (ALL); acute myeloid leukemia (AML); chronic lymphocytic leukemia (CLL); chronic myelogenous leukemia (CML); Myeloproliferative disorder/neoplasm (MPDS); and myelodysplasia syndrome. Certain forms of lymphoma include, by way of non-limiting example, Hodgkin's lymphoma, both indolent and aggressive non-Hodgkin's lymphoma, Burkitt's lymphoma, and follicular lymphoma (small cell and large cell). Certain forms of myeloma include, by way of non-limiting example, multiple myeloma (MM), giant cell myeloma, heavy-chain myeloma, and light chain or Bence-Jones myeloma. Solid tumors include, e.g., breast tumors, ovarian tumors, lung tumors, pancreatic tumors, prostate tumors, melanoma tumors, colorectal tumors, lung tumors, head and neck tumors, bladder tumors, esophageal tumors, liver tumors, and kidney tumors.

[0542] Symptoms associated with cancers and other neoplastic disorders include, for example, inflammation, fever, general malaise, fever, pain, often localized to the inflamed area, loss of appetite, weight loss, edema, headache, fatigue, rash, anemia, muscle weakness, muscle fatigue and abdominal symptoms such as, for example, abdominal pain, diarrhea or constipation.

[0543] A therapeutically effective amount of a composition 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 100 mg/kg body weight. Common dosing frequencies may range, for example, from twice daily to once a week.

Administration

[0544] Administration of the pharmaceutical compositions described herein can be via any of the accepted modes of administration for agents that serve similar utilities including, but not limited to, orally, sublingually, buccally, subcutaneously, intravenously, intranasally, intratumorally, topically, transdermally, intradermally, intraperitoneally, intramuscularly, intrapulmonarilly, vaginally, rectally, or intraocularly. Oral and parenteral administrations are customary in treating the indications that are the subject of the preferred embodiments.

[0545] The compositions described herein may be provided in unit dosage form. As used herein, a "unit dosage form" is a composition containing an amount of a compound or composition that is suitable for administration to an animal, preferably mammal subject, in a single dose, according to good medical practice. The preparation of a single or unit dosage form however, does not imply that the dosage form is administered once per day or once per course of therapy. Such dosage forms are contemplated to be administered once, twice, thrice or more per day and may be administered as infusion over a period of time (e.g., from about 30 minutes to about 2-6 hours), or administered as a continuous infusion, and may be given more than once during a course of therapy, although a single administration is not specifically excluded. The skilled artisan will recognize that the formulation does not specifically contemplate the entire course of therapy and such decisions are left for those skilled in the art of treatment rather than formulation.

[0546] The compositions useful as described above may be in any of a variety of suitable forms for a variety of routes for administration, for example, for oral, sublingual, buccal, nasal, rectal, topical (including transdermal and intradermal), ocular, intracerebral, intracranial, intrathecal, intra-arterial, intravenous, intramuscular, or other parental routes of administration. The skilled artisan will appreciate that oral and nasal compositions include compositions that are administered by inhalation, and made using available methodologies. Depending upon the particular route of administration desired, a variety of pharmaceutically- acceptable carriers well-known in the art may be used. Pharmaceutically-acceptable carriers include, for example, solid or liquid fillers, diluents, hydrotropies, surface-active agents, and encapsulating substances. Optional pharmaceutically-active materials may be included, which do not substantially interfere with the inhibitory activity of the compound or composition. The amount of carrier employed in conjunction with the compound or composition is sufficient to provide a practical quantity of material for administration per unit dose of the compound. Techniques and compositions for making dosage forms useful in the methods described herein are described in the following references, all incorporated by reference herein: Modern Pharmaceutics, 4th Ed., Chapters 9 and 10 (Banker & Rhodes, editors, 2002); Lieberman et al., Pharmaceutical Dosage Forms: Tablets (1989); and Ansel, Introduction to Pharmaceutical Dosage Forms 8th Edition (2004).

[0547] Various oral dosage forms can be used, including such solid forms as tablets, capsules (e.g. solid gel capsules and liquid gel capsules), granules and bulk powders. Tablets can be compressed, tablet triturates, enteric-coated, sugar-coated, film-coated, or multiple-compressed, containing suitable binders, lubricants, diluents, disintegrating agents, coloring agents, flavoring agents, flow-inducing agents, and melting agents. Liquid oral dosage forms include aqueous solutions, emulsions, suspensions, solutions and/or suspensions reconstituted from non-effervescent granules, and effervescent preparations reconstituted from effervescent granules, containing suitable solvents, preservatives, emulsifying agents, suspending agents, diluents, sweeteners, melting agents, coloring agents and flavoring agents.

[0548] The pharmaceutically-acceptable carriers suitable for the preparation of unit dosage forms for peroral administration is well-known in the art. Tablets typically comprise conventional pharmaceutically-compatible adjuvants as inert diluents, such as calcium carbonate, sodium carbonate, mannitol, lactose and cellulose; binders such as starch, gelatin and sucrose; disintegrants such as starch, alginic acid and croscarmelose; lubricants such as magnesium stearate, stearic acid and talc. Glidants such as silicon dioxide can be used to improve flow characteristics of the powder mixture. Coloring agents, such as the FD&C dyes, can be added for appearance. Sweeteners and flavoring agents, such as aspartame, saccharin, menthol, peppermint, and fruit flavors, are useful adjuvants for chewable tablets. Capsules typically comprise one or more solid diluents disclosed above. The selection of carrier components depends on secondary considerations like taste, cost, and shelf stability, which are not critical, and can be readily made by a person skilled in the art.

[0549] Peroral compositions also include liquid solutions, emulsions, suspensions, and the like. The pharmaceutically-acceptable carriers suitable for preparation of such compositions are well known in the art. Typical components of carriers for syrups, elixirs, emulsions and suspensions include ethanol, glycerol, propylene glycol, polyethylene glycol, liquid sucrose, sorbitol and water. For a suspension, typical suspending agents include methyl cellulose, sodium carboxymethyl cellulose, AVICEL RC-591, tragacanth and sodium alginate; typical wetting agents include lecithin and polysorbate 80; and typical preservatives include methyl paraben and sodium benzoate. Peroral liquid compositions may also contain one or more components such as sweeteners, flavoring agents and colorants disclosed above.

[0550] Such compositions may also be coated by conventional methods, typically with pH or time-dependent coatings, such that the subject composition is released in the gastrointestinal tract in the vicinity of the desired topical application, or at various times to extend the desired action. Such dosage forms typically include, but are not limited to, one or more of cellulose acetate phthalate, polyvinylacetate phthalate, hydroxypropyl methyl cellulose phthalate, ethyl cellulose, Eudragit coatings, waxes and shellac.

[0551] Compositions described herein may optionally include additional drug actives.

[0552] Other compositions useful for attaining systemic delivery of the subject compounds include sublingual, buccal and nasal dosage forms. Such compositions typically comprise one or more of soluble filler substances such as sucrose, sorbitol and mannitol; and binders such as acacia, microcrystalline cellulose, carboxymethyl cellulose and hydroxypropyl methyl cellulose. Glidants, lubricants, sweeteners, colorants, antioxidants and flavoring agents disclosed above may also be included. [0553] A liquid composition, which is formulated for topical ophthalmic use, is formulated such that it can be administered topically to the eye. The comfort may be maximized as much as possible, although sometimes formulation considerations (e.g. drug stability) may necessitate less than optimal comfort. In the case that comfort cannot be maximized, the liquid may be formulated such that the liquid is tolerable to the patient for topical ophthalmic use. Additionally, an ophthalmically acceptable liquid may either be packaged for single use, or contain a preservative to prevent contamination over multiple uses.

[0554] For ophthalmic application, solutions or medicaments are often prepared using a physiological saline solution as a major vehicle. Ophthalmic solutions may preferably be maintained at a comfortable pH with an appropriate buffer system. The formulations may also contain conventional, pharmaceutically acceptable preservatives, stabilizers and surfactants.

[0555] Preservatives that may be used in the pharmaceutical compositions disclosed herein include, but are not limited to, benzalkonium chloride, PHMB, chlorobutanol, thimerosal, phenylmercuric, acetate and phenylmercuric nitrate. A useful surfactant is, for example, Tween 80. Likewise, various useful vehicles may be used in the ophthalmic preparations disclosed herein. These vehicles include, but are not limited to, polyvinyl alcohol, povidone, hydroxypropyl methyl cellulose, poloxamers, carboxymethyl cellulose, hydroxyethyl cellulose and purified water.

[0556] Tonicity adjustors may be added as needed or convenient. They include, but are not limited to, salts, particularly sodium chloride, potassium chloride, mannitol and glycerin, or any other suitable ophthalmically acceptable tonicity adjustor.

[0557] Various buffers and means for adjusting pH may be used so long as the resulting preparation is ophthalmically acceptable. For many compositions, the pH will be between 4 and 9. Accordingly, buffers include acetate buffers, citrate buffers, phosphate buffers and borate buffers. Acids or bases may be used to adjust the pH of these formulations as needed.

[0558] Ophthalmically acceptable antioxidants include, but are not limited to, sodium metabisulfite, sodium thiosulfate, acetylcysteine, butylated hydroxyanisole and butylated hydroxy toluene. [0559] Other excipient components, which may be included in the ophthalmic preparations, are chelating agents. A useful chelating agent is edetate disodium, although other chelating agents may also be used in place or in conjunction with it.

[0560] For topical use, creams, ointments, gels, solutions or suspensions, etc., containing the composition disclosed herein are employed. Topical formulations may generally be comprised of a pharmaceutical carrier, co-solvent, emulsifier, penetration enhancer, preservative system, and emollient.

[0561] For intravenous administration, the compositions described herein may be dissolved or dispersed in a pharmaceutically acceptable diluent, such as a saline or dextrose solution. Suitable excipients may be included to achieve the desired pH, including but not limited to NaOH, sodium carbonate, sodium acetate, HC1, and citric acid. In various embodiments, the pH of the final composition ranges from 2 to 8, or preferably from 4 to 7. Antioxidant excipients may include sodium bisulfite, acetone sodium bisulfite, sodium formaldehyde, sulfoxylate, thiourea, and EDTA. Other non-limiting examples of suitable excipients found in the final intravenous composition may include sodium or potassium phosphates, citric acid, tartaric acid, gelatin, and carbohydrates such as dextrose, mannitol, and dextran. Further acceptable excipients are described in Powell, et al., Compendium of Excipients for Parenteral Formulations, PDA J Pharm Sci and Tech 1998, 52 238-311 and Nema et al., Excipients and Their Role in Approved Injectable Products: Current Usage and Future Directions, PDA J Pharm Sci and Tech 2011, 65 287-332, both of which are incorporated herein by reference in their entirety. Antimicrobial agents may also be included to achieve a bacteriostatic or fungistatic solution, including but not limited to phenylmercuric nitrate, thimerosal, benzethonium chloride, benzalkonium chloride, phenol, cresol, and chlorobutanol.

[0562] The compositions for intravenous administration may be provided to caregivers in the form of one more solids that are reconstituted with a suitable diluent such as sterile water, saline or dextrose in water shortly prior to administration. In other embodiments, the compositions are provided in solution ready to administer parenterally. In still other embodiments, the compositions are provided in a solution that is further diluted prior to administration. In embodiments that include administering a combination of a compound described herein and another agent, the combination may be provided to caregivers as a mixture, or the caregivers may mix the two agents prior to administration, or the two agents may be administered separately.

[0563] In some embodiments where plinabulin and the anti-CD47 agent are administered in separate compositions, the plinabulin composition and the anti-CD47 composition are administered using the same route of administration. In other embodiments, the plinabulin composition and the anti-CD47 composition are administered using different routes of administration. In some embodiments, both the plinabulin and anti-CD47 compositions are administered intravenously. In some embodiments, both the plinabulin and anti-CD47 compositions are administered orally. In some embodiments, both the plinabulin and anti-CD47 compositions are administered intratumorally. In some embodiments, the plinabulin composition is administered intravenously and the anti-CD47 composition is administered orally. In some embodiments, the plinabulin composition is administered intravenously and the anti-CD47 composition is administered intratumorally. In some embodiments, the plinabulin composition is administered orally and the anti-CD47 composition is administered intravenously. In some embodiments, the plinabulin composition is administered orally and the anti-CD47 composition is administered intratumorally. In some embodiments, the plinabulin composition is administered intratumorally and the anti-CD47 composition is administered intravenously. In some embodiments, the plinabulin composition is administered intratumorally and the anti-CD47 composition is administered orally.

[0564] The actual dose of the active compounds described herein depends on the specific compound, and on the condition to be treated; the selection of the appropriate dose is well within the knowledge of the skilled artisan. In some embodiments, plinabulin may be administered at a dose in the range of about 1 mg/m² to about 50 mg/m². In some embodiments, the plinabulin is administered at a dose in the range of about 1-50 mg/m² of the body surface area. In some embodiments, the plinabulin is administered at a dose in the range of about 1-2, 1-3, 1-4, 1-5, 1-6, 1-7, 1-8, 1-9, 1-10, 1-11, 1-12, 1-13, 1-13.75, 1-14, 1-15, 1- 16, 1-17, 1-18, 1-19, 1-20, 1-22.5, 1-25, 1-27.5, 1-30, 1.5-2, 1.5-3, 1.5-4, 1.5-5, 1.5-6, 1.5-7,

1.5-8, 1.5-9, 1.5-10, 1.5-11, 1.5-12, 1.5-13, 1.5-13.75, 1.5-14, 1.5-15, 1.5-16, 1.5-17, 1.5-18,

1.5-19, 1.5-20, 1.5-22.5, 1.5-25, 1.5-27.5, 1.5-30, 2.5-2, 2.5-3, 2.5-4, 2.5-5, 2.5-6, 2.5-7, 2.5- 8, 2.5-9, 2.5-10, 2.5-11, 2.5-12, 2.5-13, 2.5-13.75, 2.5-14, 2.5-15, 2.5-16, 2.5-17, 2.5-18, 2.5- 19, 2.5-20, 2.5-22.5, 2.5-25, 2.5-27.5, 2.5-30, 2.5-7.5, 3-4, 3-5, 3-6, 3-7, 3-8, 3-9, 3-10, 3-11,

3-12, 3-13, 3-13.75, 3-14, 3-15, 3-16, 3-17, 3-18, 3-19, 3-20, 3-22.5, 3-25, 3-27.5, 3-30, 3.5-

6.5, 3.5-13.75, 3.5-15, 2.5-17.5, 4-5, 4-6, 4-7, 4-8, 4-9, 4-10, 4-11, 4-12, 4-13, 4-13.75, 4-14,

4-15, 4-16, 4-17, 4-18, 4-19, 4-20, 4-22.5, 4-25, 4-27.5, 4-30, 5-6, 5-7, 5-8, 5-9, 5-10, 5-11,

5-12, 5-13, 5-13.75, 5-14, 5-15, 5-16, 5-17, 5-18, 5-19, 5-20, 5-22.5, 5-25, 5-27.5, 5-30, 6-7,

6-8, 6-9, 6-10, 6-11, 6-12, 6-13, 6-13.75, 6-14, 6-15, 6-16, 6-17, 6-18, 6-19, 6-20, 6-22.5, 6- 25, 6-27.5, 6-30, 7-8, 7-9, 7-10, 7-11, 7-12, 7-13, 7-13.75, 7-14, 7-15, 7-16, 7-17, 7-18, 7-19,

7-20, 7-22.5, 7-25, 7-27.5, 7-30, 7.5-12.5, 7.5-13.5, 7.5-15, 8-9, 8-10, 8-11, 8-12, 8-13, 8- 13.75, 8-14, 8-15, 8-16, 8-17, 8-18, 8-19, 8-20, 8-22.5, 8-25, 8-27.5, 8-30, 9-10, 9-11, 9-12, 9-13, 9-13.75, 9-14, 9-15, 9-16, 9-17, 9-18, 9-19, 9-20, 9-22.5, 9-25, 9-27.5, 9-30, 10-11, 10- 12, 10-13, 10-13.75, 10-14, 10-15, 10-16, 10-17, 10-18, 10-19, 10-20, 10-22.5, 10-25, 10-

27.5, 10-30, 11.5-15.5, 12.5-14.5, 7.5-22.5, 8.5-32.5, 9.5-15.5, 15.5-24.5, 5-35, 17.5-22.5, 22.5-32.5, 25-35, 25.5-24.5, 27.5-32.5, 2-20, 2.5-22.5, or 9.5-21.5 mg/m², of the body surface area. In some embodiments, the plinabulin is administered at a dose of about 0.5, 1,

1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10, 10.5, 11, 11.5, 12, 12.5, 13,

13.5, 14, 14.5, 15, 15.5, 16, 16.5, 17, 17.5, 18, 18.5, 19, 19.5, 20, 20.5, 21, 21.5, 22, 22.5, 23,

23.5, 24, 24.5, 25, 25.5, 26, 26.5, 27, 27.5, 28, 28.5, 29, 29.5, 30, 30.5, 31, 32, 33, 34, 35, 36,

37, 38, 39, 40 mg/m² of the body surface area. In some embodiments, the plinabulin is administered at a dose less than about 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8,

8.5, 9, 9.5, 10, 10.5, 11, 11.5, 12, 12.5, 13, 13.5, 14, 14.5, 15, 15.5, 16, 16.5, 17, 17.5, 18,

18.5, 19, 19.5, 20, 20.5, 21, 21.5, 22, 22.5, 23, 23.5, 24, 24.5, 25, 25.5, 26, 26.5, 27, 27.5, 28,

28.5, 29, 29.5, 30, 30.5, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40 mg/m² of the body surface area. In some embodiments, the plinabulin is administered at a dose greater than about 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10, 10.5, 11, 11.5, 12, 12.5, 13, 13.5, 14, 14.5, 15, 15.5, 16, 16.5, 17, 17.5, 18, 18.5, 19, 19.5, 20, 20.5, 21, 21.5, 22, 22.5, 23, 23.5, 24, 24.5, 25, 25.5, 26, 26.5, 27, 27.5, 28, 28.5, 29, 29.5, 30, 30.5, 31, 32, 33, 34, 35, 36, 37,

38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50 mg/m² of the body surface area.

[0565] In some embodiments, the plinabulin dose is about 5 mg - 300 mg, 5 mg - 200 mg, 7.5 mg - 200 mg, 10 mg - 100 mg, 15 mg - 100 mg, 20 mg - 100 mg, 30 mg - 100 mg, 40 mg - 100 mg, 10 mg - 80 mg, 15 mg - 80 mg, 20 mg - 80 mg, 30 mg - 80 mg, 40 mg - 80 mg, 10 mg - 60 mg, 15 mg - 60 mg, 20 mg - 60 mg, 30 mg - 60 mg, or about 40 mg - 60 mg. In some embodiments, the plinabulin administered is about 20 mg - 60 mg, 27 mg - 60 mg, 20 mg - 45 mg, or 27 mg - 45 mg. In some embodiments, the plinabulin administered is about 5 mg-7.5 mg, 5 mg-9 mg, 5 mg-10 mg, 5 mg-12mg, 5mg-14mg, 5mg-15 mg, 5 mg-16 mg, 5 mg- 18 mg, 5 mg-20 mg, 5 mg-22 mg, 5 mg-24 mg, 5 mg-26 mg, 5 mg-28mg, 5mg- 30mg, 5mg-32mg, 5mg-34mg, 5mg-36mg, 5mg-38mg, 5mg-40mg, 5mg-42mg, 5mg-44mg, 5mg-46mg, 5mg-48mg, 5mg-50mg, 5mg-52mg, 5mg-54mg, 5mg-56mg, 5mg-58mg, 5mg- 60mg, 7 mg-7.7 mg, 7 mg-9 mg, 7 mg-10 mg, 7 mg-12mg, 7mg-14mg, 7mg-15 mg, 7 mg-16 mg, 7 mg- 18 mg, 7 mg-20 mg, 7 mg-22 mg, 7 mg-24 mg, 7 mg-26 mg, 7 mg-28mg, 7mg- 30mg, 7mg-32mg, 7mg-34mg, 7mg-36mg, 7mg-38mg, 7mg-40mg, 7mg-42mg, 7mg-44mg, 7mg-46mg, 7mg-48mg, 7mg-50mg, 7mg-52mg, 7mg-54mg, 7mg-56mg, 7mg-58mg, 7mg- 60mg, 9 mg-10 mg, 9 mg-12mg, 9mg-14mg, 9mg-15 mg, 9 mg-16 mg, 9 mg-18 mg, 9 mg-20 mg, 9 mg-22 mg, 9 mg-24 mg, 9 mg-26 mg, 9 mg-28mg, 9mg-30mg, 9mg-32mg, 9mg-34mg, 9mg-36mg, 9mg-38mg, 9mg-40mg, 9mg-42mg, 9mg-44mg, 9mg-46mg, 9mg-48mg, 9mg- 50mg, 9mg-52mg, 9mg-54mg, 9mg-56mg, 9mg-58mg, 9mg-60mg, 10 mg-12mg, lOmg- 14mg, 10mg-15 mg, 10 mg-16 mg, 10 mg-18 mg, 10 mg-20 mg, 10 mg-22 mg, 10 mg-24 mg, 10 mg-26 mg, 10 mg-28mg, 10mg-30mg, 10mg-32mg, 10mg-34mg, 10mg-36mg, lOmg- 38mg, 10mg-40mg, 10mg-42mg, 10mg-44mg, 10mg-46mg, 10mg-48mg, 10mg-50mg, 10mg-52mg, 10mg-54mg, 10mg-56mg, 10mg-58mg, 10mg-60mg, 12mg-14mg, 12mg-15 mg, 12 mg-16 mg, 12 mg-18 mg, 12 mg-20 mg, 12 mg-22 mg, 12 mg-24 mg, 12 mg-26 mg, 12 mg-28mg, 12mg-30mg, 12mg-32mg, 12mg-34mg, 12mg-36mg, 12mg-38mg, 12mg- 40mg, 12mg-42mg, 12mg-44mg, 12mg-46mg, 12mg-48mg, 12mg-50mg, 12mg-52mg, 12mg-54mg, 12mg-56mg, 12mg-58mg, 12mg-60mg, 15 mg-16 mg, 15 mg-18 mg, 15 mg-20 mg, 15 mg-22 mg, 15 mg-24 mg, 15 mg-26 mg, 15 mg-28mg, 15mg-30mg, 15mg-32mg, 15mg-34mg, 15mg-36mg, 15mg-38mg, 15mg-40mg, 15mg-42mg, 15mg-44mg, 15mg-46mg, 15mg-48mg, 15mg-50mg, 15mg-52mg, 15mg-54mg, 15mg-56mg, 15mg-58mg, 15mg-60mg, 17 mg-18 mg, 17 mg-20 mg, 17 mg-22 mg, 17 mg-24 mg, 17 mg-26 mg, 17 mg-28mg, 17mg-30mg, 17mg-32mg, 17mg-34mg, 17mg-36mg, 17mg-38mg, 17mg-40mg, 17mg-42mg, 17mg-44mg, 17mg-46mg, 17mg-48mg, 17mg-50mg, 17mg-52mg, 17mg-54mg, 17mg-56mg, 17mg-58mg, 17mg-60mg, 20 mg-22 mg, 20 mg-24 mg, 20 mg-26 mg, 20 mg-28mg, 20mg- 30mg, 20mg-32mg, 20mg-34mg, 20mg-36mg, 20mg-38mg, 20mg-40mg, 20mg-42mg, 20mg-44mg, 20mg-46mg, 20mg-48mg, 20mg-50mg, 20mg-52mg, 20mg-54mg, 20mg-56mg, 20mg-58mg, 20mg-60mg, 22 mg-24 mg, 22 mg-26 mg, 22 mg-28mg, 22mg-30mg, 22mg- 32mg, 22mg-34mg, 22mg-36mg, 22mg-38mg, 22mg-40mg, 22mg-42mg, 22mg-44mg, 22mg-46mg, 22mg-48mg, 22mg-50mg, 22mg-52mg, 22mg-54mg, 22mg-56mg, 22mg-58mg, 22mg-60mg, 25 mg-26 mg, 25 mg-28mg, 25mg-30mg, 25mg-32mg, 25mg-34mg, 25mg- 36mg, 25mg-38mg, 25mg-40mg, 25mg-42mg, 25mg-44mg, 25mg-46mg, 25mg-48mg, 25mg-50mg, 25mg-52mg, 25mg-54mg, 25mg-56mg, 25mg-58mg, 25mg-60mg, 27 mg- 28mg, 27mg-30mg, 27mg-32mg, 27mg-34mg, 27mg-36mg, 27mg-38mg, 27mg-40mg, 27mg-42mg, 27mg-44mg, 27mg-46mg, 27mg-48mg, 27mg-50mg, 27mg-52mg, 27mg-54mg, 27mg-56mg, 27mg-58mg, 27mg-60mg, 30mg-32mg, 30mg-34mg, 30mg-36mg, 30mg-38mg, 30mg-40mg, 30mg-42mg, 30mg-44mg, 30mg-46mg, 30mg-48mg, 30mg-50mg, 30mg-52mg, 30mg-54mg, 30mg-56mg, 30mg-58mg, 30mg-60mg, 33mg-34mg, 33mg-36mg, 33mg-38mg, 33mg-40mg, 33mg-42mg, 33mg-44mg, 33mg-46mg, 33mg-48mg, 33mg-50mg, 33mg-52mg, 33mg-54mg, 33mg-56mg, 33mg-58mg, 33mg-60mg, 36mg-38mg, 36mg-40mg, 36mg-42mg, 36mg-44mg, 36mg-46mg, 36mg-48mg, 36mg-50mg, 36mg-52mg, 36mg-54mg, 36mg-56mg, 36mg-58mg, 36mg-60mg, 40mg-42mg, 40mg-44mg, 40mg-46mg, 40mg-48mg, 40mg-50mg, 40mg-52mg, 40mg-54mg, 40mg-56mg, 40mg-58mg, 40mg-60mg, 43mg-46mg, 43mg-48mg, 43mg-50mg, 43mg-52mg, 43mg-54mg, 43mg-56mg, 43mg-58mg, 42mg-60mg, 45mg-48mg, 45mg-50mg, 45mg-52mg, 45mg-54mg, 45mg-56mg, 45mg-58mg, 45mg-60mg, 48mg-50mg, 48mg-52mg, 48mg-54mg, 48mg-56mg, 48mg-58mg, 48mg-60mg, 50mg-52mg, 50mg-54mg, 50mg-56mg, 50mg-58mg, 50mg-60mg, 52mg-54mg, 52mg-56mg, 52mg-58mg, or 52mg- 60mg. In some embodiments, the plinabulin dose is greater than about 5 mg, about 10 mg, about 12.5 mg, about 13.5 mg, about 15 mg, about 17.5 mg, about 20 mg, about 22.5 mg, about 25 mg, about 27 mg, about 30 mg, about 40 mg, about 50 mg, about 60 mg, about 70 mg, about 80 mg, about 90 mg, about 100 mg, about 125 mg, about 150mg, or about 200 mg. In some embodiments, the plinabulin dose is about less than about 5 mg, about 10 mg, about 12.5 mg, about 13.5 mg, about 15 mg, about 17.5 mg, about 20 mg, about 22.5 mg, about 25 mg, about 27 mg, about 30 mg, about 40 mg, about 50 mg, about 60 mg, about 70 mg, about 80 mg, about 90 mg, about 100 mg, about 125 mg, about 150mg, or about 200 mg.

[0566] In some embodiments, a daily dose of one or more monoclonal antibodies that bind to CD47 may be from about 100 pg/kg to about 100 mg/kg or more of body weight, from about 0.5 pg/kg or less to about 1 mg/kg, from about 1.0 mg/kg to about 100 mg/kg of body weight, or from about 1.0 mg/kg to about 50 mg/kg of body weight. In some embodiments, a daily dose of a one or more monoclonal antibodies that bind to CD47 may be from about 0.1 mg to about 150 mg per dose, from about 0.5 mg or less to about 100 mg, from about 1.0 mg to about 75 mg per dose, or from about 35 mg to about 50 mg per dose.

[0567] In some embodiments, anti-CD47 agent may be administered at a dose in the range of about 1 mg/kg to about 50 mg/kg. In some embodiments, the anti-CD47 is administered at a dose in the range of about 1-50 mg/kg. In some embodiments, the anti- CD47 agent is administered at a dose in the range of about 1-2, 1-3, 1-4, 1-5, 1-6, 1-7, 1-8,

1-9, 1-10, 1-11, 1-12, 1-13, 1-13.75, 1-14, 1-15, 1-16, 1-17, 1-18, 1-19, 1-20, 1-22.5, 1-25, 1-

27.5, 1-30, 1.5-2, 1.5-3, 1.5-4, 1.5-5, 1.5-6, 1.5-7, 1.5-8, 1.5-9, 1.5-10, 1.5-11, 1.5-12, 1.5-13,

1.5-13.75, 1.5-14, 1.5-15, 1.5-16, 1.5-17, 1.5-18, 1.5-19, 1.5-20, 1.5-22.5, 1.5-25, 1.5-27.5,

1.5-30, 2.5-2, 2.5-3, 2.5-4, 2.5-5, 2.5-6, 2.5-7, 2.5-8, 2.5-9, 2.5-10, 2.5-11, 2.5-12, 2.5-13,

2.5-13.75, 2.5-14, 2.5-15, 2.5-16, 2.5-17, 2.5-18, 2.5-19, 2.5-20, 2.5-22.5, 2.5-25, 2.5-27.5,

2.5-30, 2.5-7.5, 3-4, 3-5, 3-6, 3-7, 3-8, 3-9, 3-10, 3-11, 3-12, 3-13, 3-13.75, 3-14, 3-15, 3-16,

3-17, 3-18, 3-19, 3-20, 3-22.5, 3-25, 3-27.5, 3-30, 3.5- 6.5, 3.5-13.75, 3.5-15, 2.5-17.5, 4-5,

4-6, 4-7, 4-8, 4-9, 4-10, 4-11, 4-12, 4-13, 4-13.75, 4-14, 4-15, 4-16, 4-17, 4-18, 4-19, 4-20, 4-

22.5, 4-25, 4-27.5, 4-30, 5-6, 5-7, 5-8, 5-9, 5-10, 5-11, 5-12, 5-13, 5-13.75, 5-14, 5-15, 5-16,

5-17, 5-18, 5-19, 5-20, 5-22.5, 5-25, 5-27.5, 5-30, 6-7, 6-8, 6-9, 6-10, 6-11, 6-12, 6-13, 6- 13.75, 6-14, 6-15, 6-16, 6-17, 6-18, 6-19, 6-20, 6-22.5, 6-25, 6-27.5, 6-30, 7-8, 7-9, 7-10, 7- 11, 7-12, 7-13, 7-13.75, 7-14, 7-15, 7-16, 7-17, 7-18, 7-19, 7-20, 7-22.5, 7-25, 7-27.5, 7-30,

7.5-12.5, 7.5-13.5, 7.5-15, 8-9, 8-10, 8-11, 8-12, 8-13, 8-13.75, 8-14, 8-15, 8-16, 8-17, 8-18,

8-19, 8-20, 8-22.5, 8-25, 8-27.5, 8-30, 9-10, 9-11, 9-12, 9-13, 9-13.75, 9-14, 9-15, 9-16, 9-17,

9-18, 9-19, 9-20, 9-22.5, 9-25, 9-27.5, 9-30, 10-11, 10-12, 10-13, 10-13.75, 10-14, 10-15, 10- 16, 10-17, 10-18, 10-19, 10-20, 10-22.5, 10-25, 10-27.5, 10-30, 11.5-15.5, 12.5-14.5, 7.5-

22.5, 8.5-32.5, 9.5-15.5, 15.5-24.5, 5-35, 17.5-22.5, 22.5-32.5, 25-35, 25.5-24.5, 27.5-32.5,

2-20, t 2.5-22.5, or 9.5-21.5 mg/kg. In some embodiments, the anti-CD47 agent is administered at a dose of about 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9,

9.5, 10, 10.5, 11, 11.5, 12, 12.5, 13, 13.5, 14, 14.5, 15, 15.5, 16, 16.5, 17, 17.5, 18, 18.5, 19,

19.5, 20, 20.5, 21, 21.5, 22, 22.5, 23, 23.5, 24, 24.5, 25, 25.5, 26, 26.5, 27, 27.5, 28, 28.5, 29,

29.5, 30, 30.5, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 45 mg/kg. In some embodiments, the anti-CD47 agent is administered at a dose less than about 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10, 10.5, 11, 11.5, 12, 12.5, 13, 13.5, 14, 14.5, 15, 15.5, 16,

16.5, 17, 17.5, 18, 18.5, 19, 19.5, 20, 20.5, 21, 21.5, 22, 22.5, 23, 23.5, 24, 24.5, 25, 25.5, 26,

26.5, 27, 27.5, 28, 28.5, 29, 29.5, 30, 30.5, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40 mg/kg. In some embodiments, the anti-CD47 agent is administered at a dose greater than about 0.5, 1,

23.5, 24, 24.5, 25, 25.5, 26, 26.5, 27, 27.5, 28, 28.5, 29, 29.5, 30, 30.5, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50 mg/kg.

[0568] In some embodiments, the anti-CD47 agent dose is about 0.5 mg - 3000 mg, 0.5 mg - 2500 mg, 0.5 mg - 2000 mg, 0.5 mg - 1500 mg, 0.5 mg - 1000 mg, 0.5 mg - 500 mg, 0.5 mg -200 mg, 0.75 mg - 200 mg, 1.0 mg - 100 mg, 1.5 mg - 100 mg, 2.0 mg - 100 mg, 3.0 mg - 100 mg, 4.0 mg - 100 mg, 1.0 mg - 80 mg, 1.5 mg - 80 mg, 2.0 mg - 80 mg, 3.0 mg - 80 mg, 4.0 mg - 80 mg, 1.0 mg - 60 mg, 1.5 mg - 60 mg, 2.0 mg - 60 mg, 3.0 mg - 60 mg, or about 4.0 mg - 60 mg. In some embodiments, the anti-CD47 agent administered is about 20 mg - 60 mg, 27 mg - 60 mg, 20 mg - 45 mg, or 27 mg - 45 mg. In some embodiments, the anti-CD47 agent administered is about 5 mg-7.5 mg, 5 mg-9 mg, 5 mg- 10 mg, 5 mg-12mg, 5mg-14mg, 5mg-15 mg, 5 mg-16 mg, 5 mg-18 mg, 5 mg-20 mg, 5 mg-22 mg, 5 mg-24 mg, 5 mg-26 mg, 5 mg-28mg, 5mg-30mg, 5mg-32mg, 5mg-34mg, 5mg-36mg, 5mg-38mg, 5mg-40mg, 5mg-42mg, 5mg-44mg, 5mg-46mg, 5mg-48mg, 5mg-50mg, 5mg- 52mg, 5mg-54mg, 5mg-56mg, 5mg-58mg, 5mg-60mg, 7 mg-7.7 mg, 7 mg-9 mg, 7 mg-10 mg, 7 mg-12mg, 7mg-14mg, 7mg-15 mg, 7 mg-16 mg, 7 mg-18 mg, 7 mg-20 mg, 7 mg-22 mg, 7 mg-24 mg, 7 mg-26 mg, 7 mg-28mg, 7mg-30mg, 7mg-32mg, 7mg-34mg, 7mg-36mg, 7mg-38mg, 7mg-40mg, 7mg-42mg, 7mg-44mg, 7mg-46mg, 7mg-48mg, 7mg-50mg, 7mg- 52mg, 7mg-54mg, 7mg-56mg, 7mg-58mg, 7mg-60mg, 9 mg-10 mg, 9 mg-12mg, 9mg-14mg, 9mg-15 mg, 9 mg-16 mg, 9 mg-18 mg, 9 mg-20 mg, 9 mg-22 mg, 9 mg-24 mg, 9 mg-26 mg, 9 mg-28mg, 9mg-30mg, 9mg-32mg, 9mg-34mg, 9mg-36mg, 9mg-38mg, 9mg-40mg, 9mg- 42mg, 9mg-44mg, 9mg-46mg, 9mg-48mg, 9mg-50mg, 9mg-52mg, 9mg-54mg, 9mg-56mg, 9mg-58mg, 9mg-60mg, 10 mg-12mg, 10mg-14mg, 10mg-15 mg, 10 mg-16 mg, 10 mg-18 mg, 10 mg-20 mg, 10 mg-22 mg, 10 mg-24 mg, 10 mg-26 mg, 10 mg-28mg, 10mg-30mg, 10mg-32mg, 10mg-34mg, 10mg-36mg, 10mg-38mg, 10mg-40mg, 10mg-42mg, 10mg-44mg, 10mg-46mg, 10mg-48mg, 10mg-50mg, 10mg-52mg, 10mg-54mg, 10mg-56mg, 10mg-58mg, 10mg-60mg, 12mg-14mg, 12mg-15 mg, 12 mg-16 mg, 12 mg-18 mg, 12 mg-20 mg, 12 mg- 22 mg, 12 mg-24 mg, 12 mg-26 mg, 12 mg-28mg, 12mg-30mg, 12mg-32mg, 12mg-34mg, 12mg-36mg, 12mg-38mg, 12mg-40mg, 12mg-42mg, 12mg-44mg, 12mg-46mg, 12mg-48mg, 12mg-50mg, 12mg-52mg, 12mg-54mg, 12mg-56mg, 12mg-58mg, 12mg-60mg, 15 mg-16 mg, 15 mg-18 mg, 15 mg-20 mg, 15 mg-22 mg, 15 mg-24 mg, 15 mg-26 mg, 15 mg-28mg, 15mg-30mg, 15mg-32mg, 15mg-34mg, 15mg-36mg, 15mg-38mg, 15mg-40mg, 15mg-42mg, 15mg-44mg, 15mg-46mg, 15mg-48mg, 15mg-50mg, 15mg-52mg, 15mg-54mg, 15mg-56mg, 15mg-58mg, 15mg-60mg, 17 mg-18 mg, 17 mg-20 mg, 17 mg-22 mg, 17 mg-24 mg, 17 mg- 26 mg, 17 mg-28mg, 17mg-30mg, 17mg-32mg, 17mg-34mg, 17mg-36mg, 17mg-38mg, 17mg-40mg, 17mg-42mg, 17mg-44mg, 17mg-46mg, 17mg-48mg, 17mg-50mg, 17mg-52mg, 17mg-54mg, 17mg-56mg, 17mg-58mg, 17mg-60mg, 20 mg-22 mg, 20 mg-24 mg, 20 mg-26 mg, 20 mg-28mg, 20mg-30mg, 20mg-32mg, 20mg-34mg, 20mg-36mg, 20mg-38mg, 20mg- 40mg, 20mg-42mg, 20mg-44mg, 20mg-46mg, 20mg-48mg, 20mg-50mg, 20mg-52mg, 20mg-54mg, 20mg-56mg, 20mg-58mg, 20mg-60mg, 22 mg-24 mg, 22 mg-26 mg, 22 mg- 28mg, 22mg-30mg, 22mg-32mg, 22mg-34mg, 22mg-36mg, 22mg-38mg, 22mg-40mg, 22mg-42mg, 22mg-44mg, 22mg-46mg, 22mg-48mg, 22mg-50mg, 22mg-52mg, 22mg-54mg, 22mg-56mg, 22mg-58mg, 22mg-60mg, 25 mg-26 mg, 25 mg-28mg, 25mg-30mg, 25mg- 32mg, 25mg-34mg, 25mg-36mg, 25mg-38mg, 25mg-40mg, 25mg-42mg, 25mg-44mg, 25mg-46mg, 25mg-48mg, 25mg-50mg, 25mg-52mg, 25mg-54mg, 25mg-56mg, 25mg-58mg, 25mg-60mg, 27 mg-28mg, 27mg-30mg, 27mg-32mg, 27mg-34mg, 27mg-36mg, 27mg- 38mg, 27mg-40mg, 27mg-42mg, 27mg-44mg, 27mg-46mg, 27mg-48mg, 27mg-50mg, 27mg-52mg, 27mg-54mg, 27mg-56mg, 27mg-58mg, 27mg-60mg, 30mg-32mg, 30mg-34mg, 30mg-36mg, 30mg-38mg, 30mg-40mg, 30mg-42mg, 30mg-44mg, 30mg-46mg, 30mg-48mg, 30mg-50mg, 30mg-52mg, 30mg-54mg, 30mg-56mg, 30mg-58mg, 30mg-60mg, 33mg-34mg, 33mg-36mg, 33mg-38mg, 33mg-40mg, 33mg-42mg, 33mg-44mg, 33mg-46mg, 33mg-48mg, 33mg-50mg, 33mg-52mg, 33mg-54mg, 33mg-56mg, 33mg-58mg, 33mg-60mg, 36mg-38mg, 36mg-40mg, 36mg-42mg, 36mg-44mg, 36mg-46mg, 36mg-48mg, 36mg-50mg, 36mg-52mg, 36mg-54mg, 36mg-56mg, 36mg-58mg, 36mg-60mg, 40mg-42mg, 40mg-44mg, 40mg-46mg, 40mg-48mg, 40mg-50mg, 40mg-52mg, 40mg-54mg, 40mg-56mg, 40mg-58mg, 40mg-60mg, 43mg-46mg, 43mg-48mg, 43mg-50mg, 43mg-52mg, 43mg-54mg, 43mg-56mg, 43mg-58mg, 42mg-60mg, 45mg-48mg, 45mg-50mg, 45mg-52mg, 45mg-54mg, 45mg-56mg, 45mg-58mg, 45mg-60mg, 48mg-50mg, 48mg-52mg, 48mg-54mg, 48mg-56mg, 48mg-58mg, 48mg-60mg, 50mg-52mg, 50mg-54mg, 50mg-56mg, 50mg-58mg, 50mg-60mg, 52mg-54mg, 52mg-56mg, 52mg-58mg, 52mg-60mg, 100mg-200mg, 200mg-300mg, 300mg-400mg, 400mg-500mg, 500mg-1000mg, 1000mg-2000mg, or 1000mg-3000mg . In some embodiments, the anti- CD47 agent dose is greater than about 5 mg, about 10 mg, about 12.5 mg, about 13.5 mg, about 15 mg, about 17.5 mg, about 20 mg, about 22.5 mg, about 25 mg, about 27 mg, about 30 mg, about 40 mg, about 50 mg, about 60 mg, about 70 mg, about 80 mg, about 90 mg, about 100 mg, about 125 mg, about 150mg, or about 200 mg. In some embodiments, the anti- CD47 agent dose is about less than about 5 mg, about 10 mg, about 12.5 mg, about 13.5 mg, about 15 mg, about 17.5 mg, about 20 mg, about 22.5 mg, about 25 mg, about 27 mg, about 30 mg, about 40 mg, about 50 mg, about 60 mg, about 70 mg, about 80 mg, about 90 mg, about 100 mg, about 125 mg, about 150mg, or about 200 mg, about 300 mg, about 400 mg, about 500 mg, about 1000 mg, about 2000 mg, or about 3000 mg.

[0569] In some embodiments, the initial dose of the anti-CD47 agent is 1 mg/kg on day 1 followed by a weekly dose of 45 mg/kg.

[0570] In some embodiments, the plinabulin is administered prior to the administration of the anti-CD47 agent or anti-CD47 antibody composition. In some embodiments, the plinabulin is administered concurrently with the anti-CD47 agent or anti- CD47 antibody composition. In some embodiments, the plinabulin is administered after the anti-CD47 agent or anti-CD47 antibody composition.

[0571] In some embodiments, the plinabulin is administered about 1 min, 5min, 10 min, 15 min, 20 min, 25 min, 30 min, Ih, 1.5h, 2h, 2.5h, 3h, 4h, 5h, 6h, 7h, 8h, 9h, lOh, l lh, 12h, 13h, 14h, 15h, 16h, 17h, 18h, 19h, 20h, 24h, 30h, 36h, 40h, or 48h after the administration of the anti-CD47 agent or anti-CD47 antibody composition. In some embodiments, the plinabulin is administered in less than about 1 min, 5min, 10 min, 15 min, 20 min, 25 min, 30 min, Ih, 1.5h, 2h, 2.5h, 3h, 4h, 5h, 6h, 7h, 8h, 9h, lOh, l lh, 12h, 13h, 14h, 15h, 16h, 17h, 18h, 19h, 20h, 21h, 22h, 23h, 24h, 30h, 36h, 40h, or 48h after the administration of a the anti-CD47 agent or anti-CD47 antibody composition. In some embodiments, the plinabulin is administered in more than about 1 min, 5min, 10 min, 15 min, 20 min, 25 min, 30 min, Ih, 1.5h, 2h, 2.5h, 3h, 4h, 5h, 6h, 7h, 8h, 9h, lOh, l lh, 12h, 13h, 14h, 15h, 16h, 17h, 18h, 19h, 20h, 21h, 22h, 23h, 24h30h, 36h, 40h, or 48h after the administration of the anti-CD47 agent or anti-CD47 antibody composition. In some embodiments, the plinabulin is administered in about lmin-5min, Imin-lOmin, lmin-15min, lmin-20min, 1 min-25min, 1 min-30min, 0.25h-0.5h, 0.25-0.75h, 0.25-lh,0.5h-lh, 0.5h-2h, 0.5h-2.5h, lh-2h, lh-3h, lh-5h, lh-24h, lmin-24h, or 1 min-2h, 1 day- 2days, Iday - 3days, 1 day-4 days, 1 day-5 days, or 1 day-6 days after the administration of the anti-CD47 agent or anti-CD47 antibody composition.

[0572] In some embodiments, when plinabulin is administered prior to the anti- CD47 agent or anti-CD47 antibody composition administration, the plinabulin is administered about lmin-5min, Imin-lOmin, lmin-15min, lmin-20min, 1 min-25min, 1 min- 30min, 0.25h-0.5h, 0.25-0.75h, 0.25-lh,0.5h-lh, 0.5h-2h, 0.5h-2.5h, lh-2h, lh-3h, lh-5h, Ih- 24h, Imin-lh, lmin-2h, lmin-5h, lmin-24h, 1 day- 2days, Iday - 3days, 1 day-4 days, 1 day- 5 days, or 1 day-6 days before the administration of the anti-CD47 agent or anti-CD47 antibody composition. In some embodiments, the plinabulin is administered about 1 min, 5min, 10 min, 15 min, 20 min, 25 min, 30 min, Ih, 1.5h, 2h, 2.5h, 3h, 4h, 5h, 6h, 7h, 8h, 9h, lOh, l lh, 12h, 30h, 36h, 40h, 48h, 4 days, 5 days, 6 days, or 7 days before the administration of the anti-CD47 agent or anti-CD47 antibody composition. In some embodiments, the plinabulin is administered in less than about 1 min, 5min, 10 min, 15 min, 20 min, 25 min, 30 min, Ih, 1.5h, 2h, 2.5h, 3h, 4h, 5h, 6h, 7h, 8h, 9h, lOh, l lh, 12h, 13h, 14h, 15h, 16h, 17h, 18h, 19h, 20h, 21h, 22h, 23h, 24h, 30h, 36h, 40h, 48h, 4 days, 5 days, 6 days, or 7 days before the administration of the anti-CD47 agent or anti-CD47 antibody composition. In some embodiments, the plinabulin is administered in more than about 1 min, 5min, 10 min, 15 min, 20 min, 25 min, 30 min, Ih, 1.5h, 2h, 2.5h, 3h, 4h, 5h, 6h, 7h, 8h, 9h, lOh, l lh, 12h, 13h, 14h, 15h, 16h, 17h, 18h, 19h, 20h, 21h, 22h, 23h, 24h, 30h, 36h, 40h, 48h, 3 days, 4 days, 5 days, 6 days, or 7 days before the administration of the anti-CD47 agent or anti-CD47 antibody composition.

[0573] In some embodiments, the treatment schedule includes co-administration of the anti-CD47 agent or anti-CD47 antibody composition and plinabulin once every 3 weeks. In some embodiments, the treatment schedule includes co-administration of the anti- CD47 agent or anti-CD47 antibody composition and plinabulin once every 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, or 8 weeks. In some embodiments, the treatment schedule includes co-administration of the anti-CD47 agent or anti-CD47 antibody and plinabulin two times every 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, or 8 weeks. In some embodiments, the treatment schedule includes co-administration of the anti-CD47 agent or anti-CD47 antibody composition and plinabulin once every 1 week in a treatment cycle of 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, or 8 weeks. In some embodiments, the treatment schedule includes co-administration of the anti- CD47 agent or anti-CD47 antibody composition and plinabulin twice every 1 week in a treatment cycle of 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, or 8 weeks. In some embodiments, the treatment schedule includes co-administration of the anti- CD47 agent or anti-CD47 antibody composition and plinabulin on day 1, day 8, and day 15 of a 21 -day treatment cycle. In some embodiments, co-administration of the anti-CD47 agent or anti-CD47 antibody composition and plinabulin includes administering the anti-CD47 agent or anti-CD47 antibody prior to administering plinabulin. In some embodiments, co- administration of the anti-CD47 agent or anti-CD47 antibody composition and plinabulin includes administering the anti-CD47 agent or anti-CD47 antibody after administering plinabulin. In some embodiments, co-administration of the anti-CD47 agent or anti-CD47 antibody composition and plinabulin includes administering the anti-CD47 agent or anti- CD47 antibody concurrently with plinabulin. In some embodiments, the anti-CD47 agent or anti-CD47 antibody composition described in this paragraph can independently be a first, second, third, fourth, fifth, sixth, seventh, or eighth anti-CD47 agent or anti-CD47 antibody. In some embodiments, the treatment schedule includes co-administration of the anti-CD47 agent or anti-CD47 antibody composition and plinabulin every day of the week for a week. In some embodiments, the treatment schedule includes co-administration of the anti-CD47 agent or anti-CD47 antibody composition and plinabulin every day of the week for 2 weeks, 3 weeks, or 4 weeks. In some embodiments, the treatment schedule includes co- administration of the anti-CD47 agent or anti-CD47 antibody composition and plinabulin on day 1 in weekly treatment. In some embodiments, the treatment schedule includes co- administration of the anti-CD47 agent or anti-CD47 antibody composition and plinabulin on day 1 and day 2 in weekly treatment. In some embodiments, the treatment schedule includes co-administration of the anti-CD47 agent or anti-CD47 antibody composition and plinabulin on day 1, day 2, and day 3 in weekly treatment. In some embodiments, the treatment schedule includes co-administration of the anti-CD47 agent or anti-CD47 antibody composition and plinabulin on day 1, day 2, day 3 in weekly treatment. In some embodiments, the treatment schedule includes co-administration of the anti-CD47 agent or anti-CD47 antibody composition and plinabulin on day 1, day 2, day 3, and day 4 in weekly treatment. In some embodiments, the treatment schedule includes co-administration of the anti-CD47 agent or anti-CD47 antibody composition and plinabulin on day 1, day 2, day 3, day 4, and day 5 in weekly treatment. In some embodiments, the treatment schedule includes co-administration of the anti-CD47 agent or anti-CD47 antibody composition and plinabulin on day 1, day 2, day 3, day 4, day 5, and day 6 in weekly treatment. In some embodiments, the treatment schedule includes co-administration of the anti-CD47 agent or anti-CD47 antibody composition and plinabulin on day 1, day 3, and day 5 in weekly treatment. In some embodiments, the anti-CD47 agent or anti-CD47 antibody used on each administration day can be the same or different. In some embodiments the anti-CD47 agent or anti-CD47 antibody used on the first administration day is different from the anti-CD47 agent or anti- CD47 antibody used on the rest of the administration days. In some embodiments, the anti- CD47 agent or anti-CD47 antibody used on the first administration day is the same as or different from the anti-CD47 agent or anti-CD47 antibody used on the second administration day. In some embodiments, the anti-CD47 agent or anti-CD47 antibody used on the first administration day is the same as or different from the anti-CD47 agent or anti-CD47 antibody composition used on the third administration day. In some embodiments, the anti- CD47 agent or anti-CD47 antibody composition used on the first administration day is the same as or different from the anti-CD47 agent or anti-CD47 antibody composition used on the fourth administration day. In some embodiments, the anti-CD47 agent or anti-CD47 antibody composition used on the first administration day is the same as or different from the anti-CD47 agent or anti-CD47 antibody composition used on the fifth administration day. In some embodiments, the anti-CD47 agent or anti-CD47 antibody used on the first administration day is the same as or different from the anti-CD47 agent or anti-CD47 antibody composition used on the sixth administration day. In some embodiments, the anti- CD47 agent or anti-CD47 antibody composition used on the first administration day is the same as or different from the anti-CD47 agent or anti-CD47 antibody composition used on the seventh administration day. [0574] In some embodiments, the treatment schedule includes administration of the anti-CD47 agent or anti-CD47 antibody composition (e.g., the first, the second, the third, the fourth, the fifth, the sixth, the seventh, or the eighth) once every 3 weeks. In some embodiments, the treatment schedule includes administration of the anti-CD47 agent or anti- CD47 antibody composition once every 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, or 8 weeks. In some embodiments, the treatment schedule includes administration of the anti-CD47 agent or anti-CD47 antibody composition two times every 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, or 8 weeks. In some embodiments, the treatment schedule includes administration of the anti-CD47 agent or anti- CD47 antibody composition once every 1 week in a treatment cycle of 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, or 8 weeks. In some embodiments, the treatment schedule includes administration of the anti-CD47 agent or anti-CD47 antibody composition twice every 1 week in a treatment cycle of 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, or 8 weeks. In some embodiments, the treatment schedule includes administration of the anti-CD47 agent or anti-CD47 antibody composition three times (e.g., day 1, 2, 3, or day 1, 3, 5) every week in a treatment cycle of 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, or 8 weeks. In some embodiments, the treatment schedule includes administration of the anti-CD47 agent or anti-CD47 antibody composition day 1, day 8, and day 15 of a 21-day treatment cycle. The anti-CD47 agent or anti-CD47 antibody composition described in this paragraph can independently be the first, second, third, fourth, fifth, sixth, seventh, or eighth the anti-CD47 agent or anti-CD47 antibody. In some embodiments, the treatment schedule includes administration of the anti-CD47 agent or anti- CD47 antibody composition every day of the week for a week. In some embodiments, the treatment schedule includes administration of the anti-CD47 agent or anti-CD47 antibody composition every day of the week for 2 weeks, 3 weeks, or 4 weeks. In some embodiments, the treatment schedule includes administration of the anti-CD47 agent or anti-CD47 antibody composition on day 1 in weekly treatment. In some embodiments, the treatment schedule includes administration of the anti-CD47 agent or anti-CD47 antibody composition on day 1 and day 2 in weekly treatment. In some embodiments the treatment schedule includes administration of the anti-CD47 agent or anti-CD47 antibody composition on day 1, day 2, and day 3 in weekly treatment. In some embodiments, the treatment schedule includes administration of the anti-CD47 agent or anti-CD47 antibody composition on day 1, day 3, day 5 in weekly treatment. In some embodiments, the treatment schedule includes administration of the anti-CD47 agent or anti-CD47 antibody composition on day 1, day 2, day 3, and day 4 in weekly treatment. In some embodiments, the treatment schedule includes administration of the anti-CD47 agent or anti-CD47 antibody composition on day 1, day 2, day 3, day 4, and day 5 in weekly treatment. In some embodiments, the treatment schedule includes administration of the anti-CD47 agent or anti-CD47 antibody composition on day 1, day 2, day 3, day 4, day 5, and day 6 in weekly treatment.

[0575] In some embodiments, the treatment schedule includes administration of plinabulin once every 3 weeks. In some embodiments, the treatment schedule includes administration of plinabulin once every 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, or 8 weeks. In some embodiments, the treatment schedule includes administration of plinabulin two times every 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, or 8 weeks. In some embodiments, the treatment schedule includes administration of plinabulin once every 1 week in a treatment cycle of 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, or 8 weeks. In some embodiments, the treatment schedule includes administration of plinabulin twice every 1 week in a treatment cycle of 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, or 8 weeks. In some embodiments, the treatment schedule includes administration of plinabulin three times (e.g., day 1, 2, 3, or day 1, 3, 5) every 1 week in a treatment cycle of 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, or 8 weeks. In some embodiments, the treatment schedule includes administration of plinabulin day 1, day 8, and day 15 of a 21-day treatment cycle. The anti-CD47 agent or anti-CD47 antibody composition described in this paragraph can independently be the first, second, third, fourth, fifth, sixth, seventh, or eighth the anti-CD47 agent or anti-CD47 antibody composition. In some embodiments, the treatment schedule includes administration of plinabulin every day of the week for a week. In some embodiments, the treatment schedule includes administration of plinabulin every day of the week for 2 weeks, 3 weeks, or 4 weeks. In some embodiments, the treatment schedule includes administration of plinabulin on day 1 in weekly treatment. In some embodiments, the treatment schedule includes administration of plinabulin on day 1 and day 2 in weekly treatment. In some embodiments the treatment schedule includes administration of plinabulin on day 1, day 2, and day 3 in weekly treatment. In some embodiments, the treatment schedule includes administration of plinabulin on day 1, day 3, day 5 in weekly treatment. In some embodiments, the treatment schedule includes administration of plinabulin on day 1, day 2, day 3, and day 4 in weekly treatment. In some embodiments, the treatment schedule includes administration of plinabulin on day 1, day 2, day 3, day 4, and day 5 in weekly treatment, the treatment schedule includes administration of plinabulin on day 1, day 2, day 3, day 4, day 5, and day 6 in weekly treatment.

[0576] The treatment cycle can be repeated as long as the regimen is clinically tolerated. In some embodiments, the treatment cycle for the anti-CD47 agent or anti-CD47 antibody and plinabulin is repeated for n times, wherein n is an integer in the range of 2 to 30. In some embodiments, n is 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, a new treatment cycle can occur immediately after the completion of the previous treatment cycle. In some embodiments, a new treatment cycle can occur a period of time after the completion of the previous treatment cycle. In some embodiments, a new treatment cycle can occur after 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, or 7 weeks after the completion of the previous treatment cycle.

[0577] Administration of the composition disclosed herein can be via any of the accepted modes of administration for agents that serve similar utilities including, but not limited to, orally, subcutaneously, buccally, subcutaneously, intravenously, intranasally, intratumorally, topically, transdermally, intraperitoneally, intramuscularly, intrapulmonarilly, vaginally, rectally, intragastrically, or intraocularly. Oral and parenteral administrations are customary in treating the indications that are the subject of the some embodiments.

[0578] In some embodiments, the compositions described herein can be used in combination with other therapeutic agents. In some embodiments, the compositions described herein can be administered or used in combination with treatments such as chemotherapy, radiation, and biologic therapies.

[0579] To further illustrate this invention, the following examples are included. The examples should not, of course, be construed as specifically limiting the invention. Variations of these examples within the scope of the claims are within the purview of one skilled in the art and are considered to fall within the scope of the invention as described, and claimed herein. The reader will recognize that the skilled artisan, armed with the present disclosure, and skill in the art is able to prepare and use the invention without exhaustive examples.

EXAMPLES

Example 1 - Anti-CD47 / Plinabulin Synergism

[0580] Plinabulin amplifies the presence and function of Ml anticancer macrophages, including their phagocytic activity. Blocking the activation of signal regulatory proteins alpha (SIRPoc) on macrophages by binding to CD47 on cancer cells with antibodies to CD47 (anti-CD47) increases the phagocytosis and killing of cancer cells by macrophages. In the study, the synergy of plinabulin and anti-CD47 is demonstrated with regard to tumor cell phagocytosis by macrophages, with and without antibodies to tumor cell antigens that activate the Fc receptor.

[0581] Synergy between plinabulin and anti-CD47, with or without antibodies recognizing tumor cell antigens, is evaluated in in vitro phagocytosis assays utilizing human primary non-hodgkin lymphoma (NHL) cells or NHL cell lines that are CFSE-labeled. These CFSE-labeled cells are incubated with either mouse or human macrophages that are pretreated with vehicle control or plinabulin (possibly in a 4:1 targeteffector cell ratio). Cancer cells are incubated with pre-treated macrophages in the presence of IgGl isotype control or an anti-CD47 antibody (likely clones B6H12.2, BRIC 126, or 2D3), with or without rituximab for 2 hr, as previously described (Majeti et al., 2009 Cell 138:286-299, incorporated herein by reference in its entirety). Mouse and human macrophages are also prepared as previously described. BALB/c or Fcyr7 (Taconic, Hudson, NY, USA) mouse bone marrow mononuclear cells are used to differentiate into mouse macrophages. At the end of the experiment, cells are analyzed by fluorescence microscopy to determine the phagocytic index (number of cells ingested per 100 macrophages).

[0582] More specifically (as described in Chao et al., 2010 Cell 142:699-713, incorporated herein by reference in its entirety), in vitro phagocytosis assays is conducted with NHL cells incubated with different concentration or doses of anti-CD47 antibody (B6H12.2) (from 1 pg/ ml to 10 pg/ml), and with or without a fixed concentration of anti- CD20 IgG2a, or rituximab. The incubation of NHL cells with these antibodies incubated with mouse or human macrophages is pretreated for 24-48 hours with plinabulin vehicle or plinabulin at different concentrations, from 10 nM to 1 pg/ml. The concentration of the anti- CD47 antibody and plinabulin required to produce a defined single-agent effect versus negative control (phagocytic index) is determined for each cell type. Concentrations of the agents combined to achieve this same phagocytic index are plotted on an isobologram and the combination index (CI) determined. The CI is calculated from the formula CI = (dl/Dl) + (d2/D2), whereby dl = dose of drug 1 in combination to achieve the phagocytic index, d2 = dose of drug 2 in combination to achieve the phagocytic index, Dl = dose of drug 1 alone to achieve the phagocytic index, D2 = dose of drug 2 alone to achieve the phagocytic index. A CI of less than, equal to, and greater than 1 indicates synergy, additivity, and antagonism, respectively.

[0583] In separate but similar studies, trastuzumab is used in place of rituximab, and SKBR3 Her2 overexpressing breast cancer cells is utilized instead of NHL cells to further evaluate the ability of plinabulin to boost the effects of anti-CD47 plus a tumor targeting antibody on macrophage phagocytic activity.

Example 2 - Effect of Plinabulin on Macrophages

[0584] The tumor growth promoting M2 phenotype was dominant in tumor associated macrophages (TAM) and correlates to poor prognosis. As Ml polarized macrophages process anti-tumor functions, in vivo reprogramming of macrophages was believed to a show a promising strategy for cancer treatment. The alternate polarization of TAMs from M2 to Ml was to elicit tumor- specific immune responses. In this study, the effect of plinabulin was investigated on macrophage function.

[0585] C57BF6 mice were injected subcutaneously with IxlO⁶ MC38 tumor cells and treated with 3 i.v. injections of plinabulin (15 mg/kg) every other day when the tumors were in the range of 50-80 mm³. Tumors were dissected 7 days after plinabulin first dose and analyzed on FACS using following mouse markers: CD45, CDl lb, Ly6G, ly6C and CD206. Macrophages were characterized as: CD45+ CD1 lb+Ly6G-Ly6C-L/D-. The results are depicted in FIG. 1A and show that plinabulin increased the M1/M2 ratio compared to untreated cells. C57BF6 and Rag -I- mice were injected subcutaneously with IxlO⁵ MC38 tumor cells and treated with plinabulin (7 mg/kg) or vehicle when the tumors were in the range of 50-80 mm³. Overall survival is depicted in FIG. IB, which indicates that plinabulin- treated wild-type mice had the highest survival.

[0586] Tumor-associated macrophages (TAMs) isolated from MC38 tumorbearing mice, bone marrow-derived macrophages (BMDMs) isolated from C57BI/6 mice and human monocyte-derived macrophages isolated from PBMCs. were treated with 10, 100, 200 and 1000 nM plinabulin. Expression of Ml and M2 markers were analyzed by flow cytometry following 48, 72 and 96 hours of treatment for human macrophages and 72 hours for murine TAMs and BMDMs. Mouse-anti CD206 and human-anti CD163 antibodies were used to distinguish M2 and mouse anti-CD86 or mouse anti-CD80 and human anti-CD86 antibodies were used for Ml. Mouse and human IFN y + LPS and IL-4 were applied to polarize macrophages to Ml or M2, respectively. FIG. 2A shows the M1/M2 ratio for TAM and BMDM macrophages and demonstrate the effectiveness of plinabulin to polarize macrophages to Ml. FIG. 2B shows the M1/M2 ratio for monocyte-derived macrophages and demonstrate the effectiveness of plinabulin to polarize macrophages to Ml.

[0587] Monocyte-derived macrophages were treated with 25ng IL-4 (M2 Control), 50ng IFN y + lOng LPS (Ml control), 10, 200 and lOOOnM plinabulin. Following 72 hours, treated macrophages were co-cultured with calcein-labeled NA8-mel tumor cells in an E:T ratio 1:2. Phagocytosis was analyzed subsequently after 2 hours via flow cytometry using anti-human CD14-APC and CDl lb-PE antibodies. Anti-human CD47 (lOpg/ml) was incubated with macrophages and tumors as a positive control to block “don’t eat me” signal. CD14+CDl lb+: macrophages; Calcein+CDl lb+: phagocytosis. Plinabulin treatment of macrophages induces phagocytosis of tumor cells. The results are shown in FIG. 3.

[0588] Human monocyte-derived macrophages were treated with 200nM plinabulin or LPS+ INF y for 4, 6, 8, 18, 20, 24 and 48 hours. Supernatant was collected for analysis of the cytokines by Legend plex and mRNA from cells was collected for qPCR analyses. An increase in IL-lb, IL-6 and IL-12b40 (Ml markers) protein accumulation was observed following 48h of plinabulin treatment, while IL-lb and iNOS mRNA were induced as early as 6h. Among M2 markers a decrease in Egr2, Tgfb, IL-4, CCL17 and IL- 10 was observed in mRNA level up to 20 hours following treatment with plinabulin. The results are shown in FIG. 4. Example 3 - h vitro differentiation and treatment of murine and human macrophages

[0589] MC38 murine cell line was provided by Thomas Wirth, Medizinischen Hochschule Hannover. EMT6 murine cell line and HuT 78 human cell line were purchased from the ATCC. The murine cell lines were cultured in DMEM medium containing L- glutamine (Sigma-Aldrich, D6429) and supplemented with 10% FCS (Pan Biotech, P30- 5500), IX MEM on essential amino acids (Sigma- Aldrich, M7145) and lx penicillin/streptomycin (Sigma-Aldrich, P4333). The HuT 78 human cell line was maintained in RPMI medium containing L-glutamine (R8758, Sigma- Aldrich) supplemented with penicillin/streptomycin (100 ng/ml, Sigma-Aldrich) and 10% FBS (Sigma- Aldrich).

[0590] C57BL/6N wild-type mice were obtained from the breeding facility of the Department Biomedicine or from Janvier Labs (France). Rag2% C57BL/6N mice were bred in-house at the Department Biomedicine. All animals were bred and housed according to institutional guidelines and all experiments were performed in accordance with Swiss federal regulations (Basel Kantonal license numbers: 2370 and 2408). All experiments were carried out in mice between 8-16 weeks old and both males and females were used with no influence on results. All animals were maintained under a 12 hours light cycle and given food and water available ad libitum.

[0591] Human peripheral blood mononuclear cells (PBMCs) were isolated by density gradient centrifugation, using Histopaque (Sigma- Aldrich, 10771), from buffy coats obtained from healthy blood donors (Blood Bank, University Hospital Basel, Switzerland). Fresh tumor samples were obtained from two ovarian cancer patients undergoing tumor resections at University Hospital Basel, Switzerland. Patient characteristics are summarized in Table 2. The study was approved by the local Ethical Review Board (Ethikkommission Nordwestschweiz) and University Hospital Basel, Switzerland. Written consent to use their tumor samples for research purposes was obtained from all patients. Fresh tumor samples were mechanically dissociated and digested using accutase (Innovative Cell Technologies, AT- 104), collagenase IV (Worthington, LS004188), hyaluronidase (Sigma- Aldrich, H6254), and DNAse type IV (Sigma-Aldrich, D5025), directly after excision. Single-cell suspensions were prepared and samples were stored in liquid nitrogen until further use. In the following assays, single-cell suspensions derived from ovarian cancer samples were maintained in RPMI medium containing L-glutamine (Sigma-Aldrich, R8758) supplemented with lx penicillin/streptomycin (Sigma-Aldrich, P4333) and 10% FBS (Pan Biotech, P30-5500).

Table 2

[0592] C57BL/6N WT or C57BL/6N Rag2% mice were injected subcutaneously into the right flank with 500.000 MC38 cells suspended in phenol red-free DMEM (without additives). After 18 days (named day 0), mice bearing established MC38 tumors (tumors ranging between 40-80 mm3) received peri tumoural injection of plinabulin (7.0 mg/kg, Beyondspring), vehicle (DMSO) or were left untreated on days 0, 1, 2, 4, 7, 9 and 11. Tumor volume was calculated according to the formula: D /2*d*d, with D and d being the longest and shortest tumor diameter in mm, respectively. In some experiments, mice were sacrificed on day 7, after injections of 7.5 mg/kg plinabulin (two doses per day, three times per week), the tumors were harvested and analyzed by flow cytometry.

[0593] For murine macrophage differentiation, bone-marrow cells were isolated from C57BL/6N WT mice and differentiated into macrophages by culturing the cells in complete RPMI supplemented with 20 ng/mL murine macrophage colony stimulating factor (M-CSF, Peprotech, 315-02) at 37° C for 7 days. On day 4 of the culture additional 4mL of RPMI complete medium, supplemented with MCSF (20ng/ml final) was added. On day 7, cells were washed with cold PBS and incubated with PBS containing EDTA (2mM, Sigma- Aldrich). 100,000 cells were then cultured in a 96 well plate in complete RPMI supplemented with 20 ng/mL M-CSF at 37° C overnight. Next, cells were treated with plinabulin (Beyondspring) for 48 or 72 hours at different concentrations or vehicle (0.1% DMSO),

[0594] Alternatively, tumor infiltrating macrophages were isolated from established MC38 tumors by harvesting the tumor, mechanically and enzymatically digesting it as detailed below and FACS sorting for CDl lb⁺ F4/80+ cells. TAMs were then treated with 200 nM or 1000 nM plinabulin for 48 hours ex vivo.

[0595] For human macrophage differentiation, CD 14+ cells were isolated from healthy human-derived PBMCs by positive selection using a MACS separation kit (Miltenyi Biotec, 130-050-201) - following the manufacturer’s instructions - and treated with 50 ng/ml of human M-CSF, (Peprotech, 300-25) for 6 days. At day 3 of the culture, medium was replaced with fresh medium containing 50 ng/mL M-CSF. Cells were then counted, seeded at the density of 200,000 cells per well in a 96-well plate with fresh medium (with M-CSF) and rested overnight prior to plinabulin treatment. Cells were treated with varying doses of plinabulin (Beyondspring) in the presence or absence of JNK inhibitor SP600125 (20 pM or 40 pM dose, Selleckchem) at 37°C for 48 or 72 hours.

[0596] Primary human macrophages were obtained from ovarian cancer samples by thawing the tumor single cell suspensions, obtained as described above, and FACS sorting for CDl lb+CD14+ cells using a BD Fortessa. Cells were then counted and 200,000 cells per condition were seeded in 96-well plates in RPMI medium with 50 ng/ml M-CSF and treated with plinabulin or control treatments for 48 hours at 37 °C.

[0597] In all cases, as control conditions, murine or human macrophages were treated with a combination of LPS (In Vivo Gen - 20 ng/mL for murine and 10 ng/mL for human experiments) and IFNy (50 ng/mL, Peptrotech) or IL-4 (20 ng/mL for murine and 25 ng/mL for human experiments; Peprotech), to induce either an Ml -like or M2-like polarization, respectively. Medium in these cultures was also supplemtented with M-CSF (20 ng/mL for murine cells and 50 ng/mL for human cells). Proliferation and cell viability of macrophages was assessed on BD Fortessa. Additionally, absolute cell counts were calculated by flow cytometry using Precision Count Beads (Biolegend, 424902) following the manufacturer’s instructions.

[0598] For co-culture experiments, HuT 78 lymphoma cells were pre-stained using CTV (ThermoFisher, C34557), following the manufacturer’s instructions and seeded at a density of 5,000 cells per well of a 96-well plate. Differentiated human macrophages were then added to the wells of the 96-well plate containing HuT 78 cells, at a density of 25,000 or 50,000 cells per well. The cells were co-cultured for 48 or 72 hours at 37°C. Proliferation and cell viability of tumor cells was assessed on BD Fortessa. Additionally, absolute cell counts were calculated by flow cytometry using Precision Count Beads (Biolegend, 424902) following the manufacturer’s instructions.

[0599] Human macrophages were isolated and differentiated as described previously. Total RNA was extracted using QIAGEN RNeasy kit (74104) and converted into cDNA using the iScript cDNA synthesis kit (Bio-Rad, 1708890). Next, SsoAdvanced Universal SYBR Green supermix kit (Bio-Rad, 1725270) was used for real-time qPCR according to the manufacturer’s specifications, using a Thermo Fisher ABI ViiA7 machine. GAPDH was used as housekeeping gene. The primers sequences are reported in Table 3.

Table 3

[0600] For measurement of cytokine release, human macrophages were obtained as described above and treated with either plinabulin (200 nM) or combination of LPS (10 ng/ml) and IFNy (50 ng/ml). Following 24 or 48 hours of treatment, the 96-well plates were centrifuged at 1500 rpm for 5 minutes and the cell culture supernatant was collected. Cytokine release was measured using a bead-based multiplex kit on a human M1/M2 macrophage panel (Biolegend, 740509), following the manufacturer’s instructions.

[0601] For analysis of immune populations isolated from MC38 tumors, following protocol was used. Harvested MC38 tumors were cut into smaller fragments with razor blades and then placed in 2-4 mL digestion mix (containing accutase (PAA), collagenase IV (Worthington), hyaluronidase (Sigma), and DNAse type IV (Sigma)) at 37°C for 45 min with constant shaking. Tumour suspensions were then filtered via 70 pM nylon mesh and washed with PBS containing EDTA (5mM). Cell suspensions were subjected to red blood cell (RBC) lysis by adding 1 mL of RBC lysis buffer for 1 min, followed by the wash with PBS containing 2mM EDTA. As a final step, cell suspensions were filtered via 70 pM nylon mesh and either stored in -80°C until further analysis or used directly for flow cytometry staining. [0602] Single cell suspensions were washed with PBS and stained with the fixable live/dead UV Zombie dye (BioLegend). Cells were then blocked with Fc receptorblocking anti-CD16/32 antibody (clone 2.4G2; 1: 100) or with a human Fc Receptor Binding Inhibitor (Invitrogen, 1:100) for 20 minutes at 4°C. Next, cells were stained for cell surface antigens using the fluorophore-conjugated anti-murine or anti-human antibodies listed in Table S3 for 20 minutes at 4°C. For intranuclear staining, cells were permeabilized and fixed using Invitrogen Fixation/Perm diluent (00-5223-56). Washing and antibody incubations were performed in FACS buffer (PBS, 0.5 mM EDTA, 2% FCS, 10% NaN3). Cells were either fixed with IC fix buffer (eBioscience, 00-8222-49) for 20 minutes or directly acquired on LSR Fortessa or FACS Aria III (BD Bioscience).

Plinabulin treatment leads to shrinkage of MC38 tumors and intratumoral accumulation of Ml-like TAMs

[0603] It was first investigated that the anti-tumor activity of plinabulin in vivo in mice bearing subcutaneous MC38 colon cancer tumors. Mice with established tumors (50- 100 mm³) were treated with seven doses of plinabulin (7.0 mg/kg; given peri-tumorally) spread over 11 days (FIG. 5A), and tumor growth was measured. Plinabulin-treated mice had significantly smaller tumor volumes after 14 days of treatment compared to vehicle- treated mice (FIG. 5B, left). Similarly, the percentage of live non-immune cells (CD45" cells; tumor cells) was significantly reduced in the treated group (FIG. 5B, right). Plinabulin- treated mice also had significantly prolonged survival (p=0.0081, log-rank test) to the end point (FIG. 5C). Similar findings were observed in the EMT6 breast cancer murine model (Fig. 11A and 11B).

[0604] Upon analysis of the tumor microenvironment by flow cytometry 7 days post plinabulin treatment (as outlined in Fig. 5D), no change in CD4+ (FIG. 5E, left) and CD8+ T cell (FIG. 5E, right) frequency was observed in plinabulin or control-treated mice. Plinabulin-treated MC38 tumors had significantly lower frequency of tumor infiltrating Tregs when compared to control tumors (FIG. 5F). The tumor infiltrating T cells (both CD4+ and CD8+) of plinabulin-treated mice were functional, with increased capacity to produce intracellular IFN-y, compared to control (FIG. 5G), upon ex vivo re- stimulation with anti- CD3 and anti-CD28 monoclonal antibodies (mAbs). TNF-oc expression remained unchanged in treated and control T cells (Fig. 11C). To assess the role of T cells in this system, MC38 tumor growth was monitored after plinabulin treatment in C57BL/6 Rag2"/^_ mice, which lack T cells and B cells. Both Rag2"/^_ and WT mice were equally sensitive to plinabulin, as tumor growth delay in response to plinabulin was observed in mice with and without T cells (FIG. 5H). This suggests that T cells may not be the primary driver of efficacy to plinabulin in the MC38 tumor model.

[0605] Of note, it was observed that an overall reduction in CDl lb⁺CDl lc+ myeloid cells (FIG. 51, left) and particularly of F4/80+ TAMs (FIG. 51, right) in plinabulin- treated MC38 tumors. It was also noted that a significant increase in the Ml -like to M2-like TAM ratio (defined as the ratio between CD80+ and CD206+ TAMs, FIG. 5 J), indicating an increased presence of anti-tumor Ml-like TAMs. The effects of plinabulin was subsequentially characterized on macrophage polarization and function.

Plinabulin induces Ml polarization of murine and human macrophages

[0606] In order to elucidate the direct effect of plinabulin treatment on murine macrophages, CDl lb⁺F4/80⁺ TAMs was isolated from established MC38 tumors by FACS and treated them ex vivo with plinabulin for 48h (FIG. 6A). Murine TAMs treated with a combination of LPS (20 ng/ml) and IFNy (50 ng/ml) or with IL-4 (20 ng/ml) alone served as Ml- or M2-TAM controls, respectively. As expected, treatment with LPS and IFNy led to an increase in CD80 expression in TAMs, while a decreased CD80 expression was observed in the IL-4 condition (FIG 6B and 6C). Strikingly, treatment with either dose (200 nM or 1000 nM) of plinabulin resulted in a significant increase in the M1/M2 ratio (Ml marker: CD80 and M2 marker: CD206), similar to the LPS/IFNy treatment (FIG 6C and 6D). Importantly, plinabulin did not alter the viability of TAMs. Similar to the control conditions, more than 90% of all cells were found to be alive after plinabulin treatment (Fig 12A). Next, it was assessed that the effects of plinabulin on bone marrow derived macrophages (BMDM, FIG. 6E). Similar to TAMs, plinabulin treatment of BMDMs led to a significant, dose-dependent increase in Ml markers CD80 and CD86 (FIG. 6F, left and Suppl. Fig. 12B, respectively) and a concomitant decrease in expression of CD206 (FIG. 6F, right). Accordingly, it was observed that a dose-dependent increase in the M1/M2 ratio upon plinabulin treatment, similar to LPS/IFN-y Ml control (FIG. 6G and Fig. 12C). [0607] As a next step, it was investigated that healthy human monocyte-derived macrophages (FIG. 6H). Following 48 hours of treatment with increasing doses of plinabulin or with control treatments (10 ng/ml LPS and 50 ng/ml IFN-y or 25 ng/ml IL-4 - alone in the presence of M-CSF), the expression of human Ml marker CD80 and M2 markers CD206 and CD163 were assessed by flow cytometry. Similar to murine BMDMs, treatment of human macrophages with plinabulin resulted in a dose-dependent increase in expression of CD80 (FIG. 61, left), a decrease of the M2 marker expression (FIG. 61, right and Fig. 12D) and a significant increase in the M1/M2 ratio (FIG. 6J). This phenotype was sustained also at 72 and 96 hours of culture (Fig. 12E-F).

Plinabulin induces Mi-macrophage proliferation and pro-inflammatory cytokine release

[0608] It was next assessed that whether the plinabulin-induced shift towards the Ml phenotype was accompanied by changes in macrophage proliferation and effector functions. Macrophages were differentiated from healthy human CD14+ monocytes (FIG. 7A). Proliferation was measured as a dilution of the cell tracking violet (CTV) dye in the treated macrophages. Plinabulin treatment resulted in an increase in the proliferation of CD86+ macrophages (Ml) as indicated in the histograms in FIG. 7B (left). Interestingly, no such proliferation in CD86+ macrophages was observed in untreated and the Ml control (LPS/IFN-y) and IL-4-treated groups. CD163⁺ macrophage (M2) proliferation remained unaltered in all conditions (FIG. 7B, right). Dilution of the CTV was also quantified as mean fluorescence intensity (MFI, FIG. 7C). AnnexinV was stained with to analyze potential cell death induction in the different subsets. There were no significant changes in Annexin V⁺ cells with plinabulin treatment when the total macrophage population or Ml and M2 subsets were analyzed (Fig. 3D). These findings indicate that plinabulin-induced microtubule destabilization specifically and preferentially triggers Ml -like macrophage proliferation, without inducing M2-like macrophage cell death.

[0609] Next, protein and gene expression of Ml- or M2-specific cytokines was assessed. Macrophages treated with plinabulin or the LPS/IFN-y control over 24 or 48 hours showed increased capacity to produce Ml-associated cytokines IL-ip, IL-6 and IL-12p40 (FIG. 7E). Additionally, it was detected that increased gene expression of iNOS as early as 4 hours after plinabulin treatment (FIG. 7F) and of IL-ip as early as 6 hours (Fig. 13A). Conversely, mRNAs for M2-associated genes Egr2, Tgfbl, 114, and Ccll7 remained downregulated up to 20 hours post plinabulin treatment (Fig. 13B).

JNK pathway is involved in plinabulin-induced Ml-macrophaqe polarization and proliferation

[0610] Given the recent finding that microtubule destabilization triggers the activation of JNK in murine dendritic cells, it was investigated that the involvement of JNK in plinabulin-induced Ml polarization and proliferation of human macrophages in vitro, by combining plinabulin treatment (FIG. 8A). JNK inhibition with SP600125, a potent and specific JNK inhibitor, decreased CD86 expression in plinabulin-treated macrophages and thus resulted in partial reversion of the plinabulin effect on Ml polarization (FIG. 8B, left). In contrast, JNK inhibition did not affect expression of CD163 (FIG. 8B, right). Accordingly, upon assessing the total counts by FACS, Ml (CD86+) events were higher upon plinabulin treatment and this effect was partially lost upon JNK inhibition; a similar effect was not observed on CD163+ events (FIG. 8C). Of note, JNK inhibition did not significantly alter macrophage viability (FIG. 8D). These findings suggest a role for plinabulin in directly inducing polarization of macrophages towards an Ml -like phenotype and potentially the proliferation of these Ml -like macrophages in a JNK-dependent fashion.

Plinabulin-polarized Ml-like human macrophages induce Fas-dependent direct killing of tumor cells

[0611] Next it was sought to determine the effector functions of plinabulin- polarized Ml-like macrophages. CD 14+ monocyte-derived macrophages were treated with plinabulin for 48 hours. Upon removal of plinabulin by changing the culture medium, macrophages were co-cultured for further 48 hours with HuT 78 tumor cells, pre-stained with CTV, at 5:1 and 10:1 macrophage to tumor cell ratio (FIG. 9A). Tumor cell viability was measured by live/dead stain and tumor cell proliferation was determined by the dilution of the CTV dye.

[0612] Flow cytometry analysis showed a decrease in HuT 78 viability upon coculture with 1000 nM plinabulin-treated macrophages at either cell ratio compared to the untreated and the HuT 78 cells only controls (FIG. 9B). A similar result in decreased tumor cell viability was seen in the LPS/IFN-y Ml treatment control (FIG. 9B). Similarly, HuT 78 proliferation was strongly inhibited when co-cultured with plinabulin (1000 nM)-polarized human macrophages (FIG 9C and 9D), while there was a significantly increased proportion of live macrophages at this dose (Fig 14).

[0613] Interestingly, it was observed than an increase in Fas+ tumor cells in coculture with plinabulin-treated macrophages, compared to untreated cells and positive control (LPS/IFN-y treatment, FIG. 9E). Similarly, it was observed that a dose-dependent increase in Fas-L expression on human macrophages treated with plinabulin (FIG. 9F). Taken together, the data demonstrate that plinabulin-polarized macrophages suppress tumor cell proliferation and increase tumor cell death, which is potentially mediated through Fas/Fas-L interaction.

Plinabulin triggers functional polarization of ovarian cancer patient tumor infiltrating mgcrophgges

[0614] In order to investigate whether plinabulin induces Ml polarization of human TAMs, FACS sorted CDl lb+CD14+ cells from two tumor digests derived from ovarian cancer patients (predominantly of the M2 phenotype, Fig. 15), and treated them with plinabulin prior to staining with CTV (FIG. 10A). Strikingly, after 48 hours of treatment with increasing doses of plinabulin, a dose dependent increase in percentage of CD86+ TAMs was observed (FIG. 10B, left). A similar increase was noted in the LPS/IFNy Ml control treatment (FIG. 10B, left). Conversely, expression of CD206 was significantly lower in plinabulin-treated cells compared to the untreated and IL-4 M2 control treatment (FIG. 10B, right).

As with healthy human CD 14+ derived macrophages, plinabulin treatment of intratumoral TAMs led to a preferential increase in the proliferation of Ml (CD86+) TAMs (measured by CTV dilution) as shown in the histograms in FIG. 10C (left) and MFI quantified in FIG. 10D (left), which was not observed for M2 TAMs (FIG. 10C and 10D, right). Altogether, these data demonstrate that plinabulin treatment can re-polarize TAMs derived from cancer patients towards an Ml -like phenotype as well as preferentially induce Ml proliferation. [0615] The anti-tumor mode of action of microtubule-targeting agents (MTAs) is predominantly through mitotic spindle arrest and subsequent activation of apoptotic pathways and tumor cell death.

[0616] However, MT A Plinabulin binds to the colchicine pocket of P-tubulin, in oc, P-tubulin heterodimers, at a distinct site and with predicted kinetics that differ from colchicine and other tubulin-targeting agents. Plinabulin-bound tubulin heterodimers are prevented from polymerizing into microtubules, which ultimately affects cellular functions in a cell-type specific manner. In line with this, while plinabulin selectively decreases tumor blood flow by eliminating endothelial cells in tumor blood vessels, it has also been shown to increase the maturation of dendritic cells and to induce apoptosis of cancer cells. Potentially relevant to the ability of plinabulin to alleviate chemotherapy -induced neutropenia, - an indication for which plinabulin is in Phase 3 clinical testing (NCT03294577) - targeting tubulin with plinabulin boosted the number of hematopoietic stem/progenitor cells in the bone marrow of tumor bearing mice.

[0617] Nevertheless, the effects of plinabulin on the immune tumor microenvironment in tumor models and cancer patients remains relatively unexplored. This is particularly important as the prospect to enhance current cancer immunotherapies via rational combinations and by modulating distinct steps in the cancer-immunity cycle offers great clinical potential [22, 23].

[0618] In the study, it was confirmed that plinabulin exhibits single agent efficacy in vivo in syngeneic MC38 and EMT6 murine tumor models, which are both myeloid- dominated and therefore appropriate models to study myeloid cells. Indeed, multiple studies show that depletion of macrophages with an anti-CSFRl antibody leads to lower tumor burden in the MC38 model, supporting the functional role of myeloid cells and particularly macrophages in altering tumor rejection.

[0619] Given the known role of plinabulin, in inducing maturation and strong activation of DCs and subsequently enhancing anti-tumor immunity, it was hypothesized that other myeloid cells are involved in the anti-tumor efficacy of plinabulin. Indeed, immunophenotyping revealed a macrophage dominant phenotype, with increased proportion of Ml -like TAMs in plinabulin-treated tumors. T cells were found to be non-critical in driving monotherapy efficacy of plinabulin, as suggested by the studies in T cell deficient Rag2-/- animals. In fact, in vitro plinabulin treatment of TAMs isolated from MC38 tumors, murine BMDM and human monocyte-derived macrophages led to a dose-dependent increase in phenotypic and functional polarization towards the Ml phenotype. Of note, the dose of plinabulin used for the in vitro studies was demonstrated to be achievable in cancer patients treated with the drug. The plinabulin-induced Ml phenotype was characterised by upregulation of Ml surface markers as well as enhanced pro -inflammatory cytokine secretion, similar to classical Ml-polarizing agents such as LPS and IFN-y. However, plinabulin-induced Ml-like polarization displayed unique characteristics not observed for classical in vitro polarized Ml macrophages, such as increased proliferation of Ml-like macrophages as well as direct tumor killing upon exposure to plinabulin. This suggests that plinabulin may exhibit Ml-like immunomodulatory properties which are mechanistically and functionally distinct from classic Ml polarization observed after LPS and IFN-y stimulation in vitro. An unbiased approach, such as differential gene expression analysis, could perspectively help in determining the extent of overlap between classical and plinabulin- induced macrophage polarization.

[0620] Furthermore, the work shows that a crucial functional consequence of plinabulin treatment on human macrophages is their enhanced tumor cell killing capacity. Experimentally, the macrophages were treated with plinabulin and subsequently incubated them with tumor cells in the absence of the compound, to exclude any direct cytotoxic effects of plinabulin on tumor cells. Thus it was considered that the observed increase in tumor cell death as a direct consequence of plinabulin-induced immunomodulation of human macrophages. Macrophages are capable of regulating self-apoptosis in an autocrine and paracrine way via the Fas-Fas-L axis. Additionally, macrophages are known to express both Fas and Fas-L and to upregulate and release soluble Fas-L upon activation by stimulation with immune complexes, PHA, or superantigen. In this work it was hypothesized that the plinabulin-enhanced tumor cell killing by macrophages is at least partly dependent on the observed increased Fas-L expression on polarized TAMs. Additionally, it was observed that an increased Fas expression on Hu T78 tumor cells co-cultured with plinabulin-polarized macrophages. Fas-L deficiency in murine tumor models was shown to skew tumorinfiltrating myeloid cell populations towards an immunosuppressive phenotype and led to enhanced tumor burden. Another potential mechanism by which increased tumor cell death is observed in the study could be the increase in the secretion of pro-inflammatory cytokines such as IL-ip, which was previously shown to induce tumor cell death in the presence of IFN-y. It remains to be explored whether the enhanced tumor-killing capacity of plinabulin- polarized TAMs is exclusively contact-dependent and/or mediated by soluble factors such as IL- Ip.

[0621] Plinabulin is known to bind to tubulin in a differentiated manner, resulting in direct cytotoxic effects on tumor cells. It was therefore striking to find that in immune cells, namley macrophages and dendritic cells, plinabulin does not induce immune cell apoptosis and on the contrary enhances macrophage activation function and proliferation. Mechanistically, plinabulin-induced macrophage polarisation and proliferation was shown to be largely dependent on JNK signalling.

[0622] Similarly, plinabulin-induced tumor cell apoptosis observed in previous studies was shown to be dependent on JNK signalling within tumor cells. Thus, treatment with plinabulin seems to present either pro- or anti-proliferative effects depending on the cell type, which may in turn depend on cell- specific JNK downstream signaling.

[0623] Similarly to what is shown here for macrophages, it has been previously shown that the activation of the JNK pathway is critical for DC maturation in response to plinabulin treatment. In particular, GEF-H1 is released upon microtubule destabilization by plinabulin and is necessary for plinabulin-induced JNK pathway activation and subsequent DC activation.

[0624] A high-infiltration of TAMs correlates with poor prognosis in most solid tumors. In ovarian cancer, TAMs constitute the main population of immune cells within the tumor microenvironment. These cells are strongly implicated in the progression, metastasis and chemoresistance of ovarian cancer and are therefore a predictor of poor clinical outcome. While platinum derivative chemotherapeutic compounds, such as cisplatin and carboplatin, may rather favor an increase in tumor-promoting M2 macrophages, here it is shown that exposure to plinabulin, a microtubule destabilizing drug, is able to re-polarize TAMs derived from ovarian cancer patients towards an Ml -like phenotype and induce Ml proliferation. This suggests that plinabulin treatment is a viable therapeutic option for re-polarizing macrophages in human tumors. [0625] Plinabulin therapy may be particularly attractive in combination with radiotherapy, which leads to increased influx of monocytes and conversion to M2 TAMs. Indeed, plinabulin application prior to radiotherapy was shown to increase treatment efficacy. Additionally, the findings provide a rationale to design combinations of plinabulin with immunotherapies targeting myeloid cells, in a synergistic effort to achieve enhanced antitumor immunity. In conclusion, the study supports the further development of plinabulin in clinical trials and offers key novel insights into the mechanism of action of MDAs in triggering TAM polarization towards an anti-tumoral phenotype.

Example 4

[0626] The aim of this study was to evaluate the effect of Plinabulin and anti- CD47 Ab monotherapy on the ADCP effect on Raji cells by flow cytometry. The fresh PBMCs from one healthy donor were ordered from the local vendor Allcells. Tumor cell lines were grown in RPMI1640+10%FBS. Cells were cultured with corresponding growth medium in 37°C incubator with 5% CO2. Cells were passaged 2-3 times for one week after revival from the frozen state.

Day 0: Monocyte isolation and macrophage differentiation

[0627] Fresh PBMC cell number were determined with Vi-cell, which produced live cell number and viability with trypan blue method. Cell suspension was centrifuged at 1500 rpm for 10 minutes. The supernatant was completely aspirated. Cell pellet was resuspended in 80 pF of buffer per 10⁷ total cells. Twenty microliters of CD14 MicroBeads were added per 10⁷ total cells. The solution was mixed well and incubated for 15 minutes in the refrigerator (2-8 °C). The cells were washed by adding 1-2 mF of buffer per IO7 cells and centrifuge at 1500 rpm for lOmin. The cells were resuspended up to 10⁸ cells in 500 pF of buffer. The column was placed in the magnetic field of a MidiMACS Separator. The column was prepared by rinsing with 3 mF of buffer. The cell suspension was applied onto column. Unlabeled cells were collected that pass through and wash column with 3x3 mF of buffer. Total effluent was collected, this was the unlabeled cell fraction. Washing steps were performed by adding buffer three times. Only add new buffer when the column reservoir is empty. The column was removed from the separator and placed on a suitable collection tube. The 5 mL of buffer was pipetted onto the column. Immediately flushed out the magnetically labeled cells by firmly pushing the plunger into the column. This fraction represented the Monocytes. The enriched monocyte population was counted and re-suspended in complete RPMI medium to a concentration of 1 x 10⁶ cells/mL. Recombinant human M-CSF were added into the cells with a final concentration of 50 ng/mL. Ten milliliter of cell suspensions with M-CSF were seeded into 10 cm tissue culture dish.

Day 3: medium renewal

[0628] Removed the medium. Added equal volume of fresh culture medium containing M-CSF (50 ng/mL) to the dish and incubate for another 3 days at 37°C.

Day 6: re-seed macrophages

[0629] The supernatant was transferred into one tube, dissociate cells with TrypLE for 10 min and gentle scraping, collect the cells into the supernatant. The tube was centrifuged at 1200 rpm for 10 min, remove the supernatant and add appropriate volume of freshly prepared complete RPMI containing 50 ng/mL M-CSF. Count viable cells. Cell density was adjusted to 3 x 10⁶ cells/mL with complete RPMI containing 50 ng/mL M-CSF, seed 6 x 10⁶ cells/2 mL per well in 6- well plate, culture the cells overnight.

Day 7: treatment with Plinabulin

[0630] In the morning, prepared serial diluted Plinabulin in DMSO then 1000- fold dilution into the 6-well plate by adding 2 pL of compound solution into 2 mL of cell solution, mix well by gentle tapping the plate, to make the final concentration of Plinabulin as 10 nM, 30 nM, 100 nM, 300 nM, 1000 nM, 3000 nM and 10000 nM, the final concentration of DMSO is 0.1%.

Day 9: Co-culture

[0631] Target cell treatment

[0632] Collected Raji cells and washed once with HBSS, resuspended in prewarmed HBSS at a density of 1 x 10⁶ cells/mL. Centrifuged Calcein- AM tube, added 50 pL of DMSO into 50 pg of powder to make 1 mM stock solution, made 5 pL/tube aliquot and store at -20°C. Added 1 pL calcein-DMSO solution for every 5 x 10⁶ cells, mix well. Incubated the cells at 37°C for 20 minutes. Centrifuged cells at 1,250 rpm for 5 minutes, washed once with HBSS, and resuspended in serum free RPMI 1640 at a density of 12 x 10⁶ cells/mL. Seeded 25 pL of target cells into an ultra-low attachment 96 well plate at a density of 300,000 cells/well. Prepared 4x anti-CD47 solution in serum free RPMI 1640. Added 25 pL 4x drug solution to indicated target cell wells, to make the final concentration of each drug as shown in plate map. Incubated at 37°C for 30 minutes.

[0633] Seed macrophages

[0634] Removed the medium of macrophage cells, wash once with HBSS, incubate macrophages with TrypLE at 37°C for 8 minutes, gently dissociated cells in dish and pipet into a 50 mF tube, and centrifuged at 1,250 rpm for 5 minutes at 4°C. Washed once with HBSS, counted and resuspended in serum free RPMI 1640 at a density of 3 x 10⁶ cells/mL. Added macrophage cell solution into the target cells at a density of 150,000 cells/50 pL/well, so that the E:T ratio is 1:2, resuspend all the cells by gentle pipetting and incubated at 37°C for 2 hours and transferred to biomarker group for FACS analysis.

Day 9: FACS analysis

[0635] Transferred cell suspension into FACS tubes. Incubated cells with 20 pF FcR Blocking Reagent (human, Miltenyi, CAT# 130-059-901) at 4°C for 15 minutes in dark. Added the anti-human CD14antibody diluted in Fc blocking reagent to each sample. The final staining volume is 100 pL. Pulse vortex gently to mix. Stain at 4°C for 30 minutes in dark. Added 2 mL FACS Wash Buffer to each tube and re-suspend the cells. Centrifuged the tubes at 300 g for 5 min and discarded the supernatant. Repeated the wash step. Added 5 pL 7-AAD so that the final staining volume is 100 pL. Gently vortexed the cells and stain at RT for 10 minutes in dark. Analyzed the samples on cytometer.

[0636] The Raji cells were co-cultured with isolated macrophage from fresh human PBMC, and treated with different concentration of Plinabulin and anti-CD47 Ab respectively, then analyzed phagocytosis by flow cytometry, the results indicated plinabulin and anti- CD47 dose-dependently increased phagocytosis. Table 4 and FIG. 16 illustrate the percentage of phagocytosis index of Plinabulin.

Table 4

[0637] Table 5 and FIG. 17 illustrate the percentage of phagocytosis index of anti-CD47.

Table 5

Example 5

[0638] Using the mouse J774 cell system and following the procedures described in Anderson et al., Melanoma Research, 2020, 30: 147-158, synergistic effects between anti- CD47 and plinabulin to boost macrophage induced phagocytosis is evaluated. J774 macrophages are treated with plinabulin for up to 3 days and then activated 24 hours before phagocytosis assays using 100 ng/ml recombinant mlFNy (eBioscience), with and without plinabulin still present. In addition, human monocytes or macrophages are used in place of J7774 cells. Cancer cells, including Raji cells, are either GFP+ or labelled with carboxyfluorescein succinimidyl ester (CFSE) and are then be incubated with 10 ng/ml of CD47 blocking reagents, isotype controls, or tumor-targeting antibodies for 30 minutes. Macrophages are then co-cultured in non-adherent plates with cancer cells at approximately a 1:1 (J774) ratio, with or without plinabulin present. Phagocytosis is analyzed by flow cytometry after 2 hours. Phagocytosis is quantified as the percentage of macrophages that engulfed CFSE+loaded/GFP+ tumor cells. The concentrations tested for monotherapies and combination allow for an analysis of synergism between anti-CD47 and plinabulin.

Claims

WHAT IS CLAIMED IS:

1. A method of treating a subject with cancer, the method comprising: administering a therapeutic dose of a compound of Formula (I):

wherein

Ri' and Ri" are each independently selected from the group consisting of a hydrogen atom, a deuterium atom, a halogen atom, and saturated C1-C24 alkyl, unsaturated Ci- C24 alkenyl, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, azido, substituted nitro, phenyl, and substituted phenyl groups, hydroxy, carboxy, — CO — O — R7, cyano, alkylthio, halogenated alkyl including polyhalogenated alkyl, halogenated carbonyl, and carbonyl — CH2CO — R7, wherein R7 is selected from a hydrogen atom, a halogen atom, and saturated C1-C24 alkyl, unsaturated C1-C24 alkenyl, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, azido, substituted nitro, phenyl, and substituted phenyl groups; R, Ri' and Ri" are either covalently bound to one another or are not covalently bound to one another;

Y is selected from the group consisting of a nitrogen atom, a substituted nitrogen atom with a R5 group from above, an oxygen atom, a sulfur atom, a oxidized sulfur atom, a methylene group and a substituted methylene group;

Z, for each separate n, if non-zero, and Zi, Z2, Za and Z4 are each separately selected from a carbon atom, a sulfur atom, a nitrogen atom or an oxygen atom; the dashed bonds may be either single or double bonds; and an anti-CD47 agent.

2. The method of claim 1, wherein the anti-CD47 agent is an antibody or antigenbinding fragment that specifically binds to CD47.

3. The method of claim 2, wherein the antibody that specifically binds to CD47 is an isolated monoclonal antibody.

4. The method of claim 1, wherein the anti-CD47 agent is an antibody or antigenbinding fragment that specifically binds to SIRPa.

5. The method of claim 4, wherein the antibody that specifically binds to SIRPa is an isolated monoclonal antibody.

6. The method of claim 4, wherein the antibody promotes macrophage-mediated phagocytosis of a CD47-expressing cell.

7. The method of claim 1, wherein the anti-CD47 agent is a soluble CD47 binding SIRPa fragment.

8. The method of claim 1, wherein the anti-CD47 agent prevents interaction between CD47 and SIRPa.

9. The method of claim 1, wherein the anti-CD47 agent is an IgG isotype selected from the group consisting of IgGl isotype, IgG2 isotype, IgG3 isotype, IgG4 isotype, IG-G1- N297Q, IG4-S228P, and IG64 PE.

10. The method of claim 1, wherein the cancer is selected from non-Hodgkin lymphoma, acute lymphoblastic leukemia (ALL), T-ALL, B-ALL, acute myelogenous leukemia (AML), B -lymphoblastic leukemia/lymphoma; diffuse large B cell lymphoma (DLBCL); B-cell chronic lymphocytic leukemia/small lymphocytic lymphoma, chronic lymphocytic leukemia (CLL), chronic myelocytic leukemia (CML), Burkitt's lymphoma, follicular lymphoma, SLL, marginal zone lymphoma, CNS lymphoma, Richter's Syndrome, multiple myeloma, myelofibrosis, polycythemia vera, cutaneous T-cell lymphoma, MGUS, myelodysplastic syndrome (MDS), immunoblastic large cell lymphoma, precursor B- lymphoblastic lymphoma and anaplastic large cell lymphoma.

11. The method of claim 1, wherein the compound of formula (I) is administered at a dose from about 5 mg/m² to 150 mg/m².

12. The method of claim 1, wherein the compound of formula (I) is administered orally, sublingually, buccally, subcutaneously, intravenously, intranasally, intratumorally, topically, transdermally, intradermally, intraperitoneally, intramuscularly, intrapulmonarilly, vaginally, rectally, or intraocularly.

13. The method of claim 1, wherein the compound of formula (I) is administered in combination with radiation.

14. The method of claim 1, wherein the compound of formula (I) is administered once a week.

-146-

15. The method of claim 1, wherein the compound of formula (I) is administered once on each of day 1 and day 8 of a three- week (21 day) treatment cycle.

16. The method of claim 1, wherein the cancer is non-Hodgkin’s lymphoma.

17. The method of any one of claims 1 to 16, wherein the compound of Formula (I) is selected from plinabulin, (3Z,6Z)-3-(phenyl-2,3,4,5,6-d5)-methylene-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene)piperazine-2, 5-dione; (3Z, 6Z)-3-(phenyl-2, 3,4,5, 6-ds)-methylene-d- 6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene)piperazine-2, 5-dione; (3Z,6Z)-3-

(phenylmethylene-d)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(phenyl-2,3,4,5,6-d5)-methylene-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene- d)piperazine-2, 5-dione; (3Z,6Z)-3-(phenylmethylene)-6-((5-(tert-butyl)-lH-imidazol-4- yl)methylene-d)piperazine-2, 5-dione; (3Z, 6Z)-3-(phenyl-2, 3,4,5, 6-ds)-methylene-d-6-((5- (tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(4-Fluoro- (phenyl-2,3,5,6-d4))-methylene-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene)piperazine-

2, 5-dione; (3Z,6Z)-3-(4-Fluoro-(phenyl-2,3,5,6-d4))-methylene-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-fluorobenzylidene)-6-((5- ( tert-butyl)- lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3- benzoylbenzylidene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-(4-fluorobenzoyl)benzylidene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene- d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-(4-methoxybenzoyl)benzylidene)-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-methoxybenzylidene)-6-((5- ( tert-butyl)- lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; or (3Z,6Z)-3-(3-

18. A method of treating cancer in a subject in need thereof, wherein the cancer comprises cells that express CD47, the method comprising: administering to the subject an effective amount of one or more anti-CD47 agent and a compound of Formula (I):

wherein

R, Ri' and Ri" are either covalently bound to one another or are not covalently bound to one another; R2, R3, and R5 are each separately selected from the group consisting of a hydrogen atom, a deuterium atom, a halogen atom, and saturated C1-C12 alkyl, unsaturated Ci- C12 alkenyl, acyl, cycloalkyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, and substituted nitro groups, sulfonyl and substituted sulfonyl groups; m is an integer equal to zero, one or two;

Y is selected from the group consisting of a nitrogen atom, a substituted nitrogen atom with a R5 group from above, an oxygen atom, a sulfur atom, a oxidized sulfur atom, a methylene group and a substituted methylene group; and

Z, for each separate n, if non-zero, and Zi, Z2, Za and Z4 are each separately selected from a carbon atom, a sulfur atom, a nitrogen atom or an oxygen atom; and the dashed bonds may be either single or double bonds.

19. The method of claim 18, wherein the one or more anti-CD47 agent is a monoclonal antibody that inhibits the interaction between human CD47 and SIRPa.

20. The method of claim 19, wherein the monoclonal antibody is a human antibody.

21. The method of claim 19 or 20, wherein the monoclonal antibody comprises a human kappa constant region.

22. The method of any one of claims 19 to 21, wherein the monoclonal antibody is administered prior to the administration of plinabulin.

23. The method of any one of claims 19 to 21, wherein the monoclonal antibody is administered after the administration of plinabulin.

24. The method of claim 18, wherein the monoclonal antibody is an optimized antibody molecule.

-149-

25. The method of claim 18, wherein the one or more anti-CD47 agent is selected from rituximab or daratumumab.

26. The method of any one of claims 18 to 25, wherein the cancer is selected from acute lymphoblastic leukemia (ALL), T-ALL, B-ALL, acute myelogenous leukemia (AML), Non-Hodgkin lymphoma, B -lymphoblastic leukemia/lymphoma; diffuse large B cell lymphoma (DLBCL); B-cell chronic lymphocytic leukemia/small lymphocytic lymphoma, chronic lymphocytic leukemia (CLL), chronic myelocytic leukemia (CML), Burkitt's lymphoma, follicular lymphoma, SLL, marginal zone lymphoma, CNS lymphoma, Richter's Syndrome, multiple myeloma, myelofibrosis, polycythemia vera, cutaneous T-cell lymphoma, MGUS, myelodysplastic syndrome (MDS), immunoblastic large cell lymphoma, precursor B -lymphoblastic lymphoma and anaplastic large cell lymphoma.

27. The method of any one of claims 18 to 25, wherein the cancer is a cancer of a tissue selected from the group consisting of: lung, pancreas, breast, liver, ovary, testicle, kidney, bladder, spine, brain, cervix, endometrium, colon/rectum, anus, esophagus, gallbladder, gastrointestinal tract, skin, prostate, pituitary, stomach, uterus, vagina, and thyroid.

28. The method of claim 18, wherein the one or more anti-CD47 agent is administered in combination with a pharmaceutically acceptable carrier or diluent.

29. The method of claim 18, wherein the one or more anti-CD47 agent is administered subcutaneously.

30. The method of claim 18, wherein the one or more anti-CD47 agent is administered intravenously.

31. The method of any one of claims 18 to 30, wherein the compound of Formula (I) is selected from plinabulin, (3Z,6Z)-3-(phenyl-2, 3,4,5, 6-d5)-methylene-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene)piperazine-2, 5-dione; (3Z, 6Z)-3-(phenyl-2, 3,4,5, 6-ds)-methylene-d- 6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene)piperazine-2, 5-dione; (3Z,6Z)-3-

(phenylmethylene-d)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione;

- ISO- (3Z,6Z)-3-(phenyl-2,3,4,5,6-d5)-methylene-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene- d)piperazine-2, 5-dione; (3Z,6Z)-3-(phenylmethylene)-6-((5-(tert-butyl)-lH-imidazol-4- yl)methylene-d)piperazine-2, 5-dione; (3Z, 6Z)-3-(phenyl-2, 3,4,5, 6-ds)-methylene-d-6-((5- (tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(4-Fluoro- (phenyl-2,3,5,6-d4))-methylene-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene)piperazine-

2, 5-dione; (3Z,6Z)-3-(4-Fluoro-(phenyl-2,3,5,6-d4))-methylene-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-fluorobenzylidene)-6-((5- ( tert-butyl)- lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3- benzoylbenzylidene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione;

(3Z,6Z)-3-(3-(4-fluorobenzoyl)benzylidene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene- d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-(4-methoxybenzoyl)benzylidene)-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-methoxybenzylidene)-6-((5- ( tert-butyl)- lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; or (3Z,6Z)-3-(3-

32. A pharmaceutical composition comprising a compound of Formula (I)

wherein

Ri, R4, and Re, are each separately selected from the group consisting of a hydrogen atom, a deuterium atom, a halogen atom, and saturated Ci,-C24 alkyl, unsaturated Ci- C24 alkenyl, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, azido, substituted nitro, phenyl, and substituted phenyl groups, hydroxy, carboxy, — CO — O — R7, cyano, alkylthio, halogenated

-151- alkyl including polyhalogenated alkyl, halogenated carbonyl, and carbonyl — CH2CO — R7, wherein R7 is selected from a hydrogen atom, a halogen atom, and saturated C1-C24 alkyl, unsaturated C1-C24 alkenyl, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, azido, substituted nitro, phenyl, and substituted phenyl groups;

Z, for each separate n, if non-zero, and Zi, Z2, Za and Z4 are each separately selected from a carbon atom, a sulfur atom, a nitrogen atom or an oxygen atom; and the dashed bonds may be either single or double bonds; and

-152- one or more anti-CD47 agent.

33. The pharmaceutical composition of claim 32, wherein the anti-CD47 agent is an antibody or antigen-binding fragment that specifically binds to CD47.

34. The pharmaceutical composition of claim 33, wherein the antibody that specifically binds to CD47 is an isolated monoclonal antibody.

35. The pharmaceutical composition of claim 32, wherein the anti-CD47 agent is an antibody or antigen-binding fragment that specifically binds to SIRPa.

36. The pharmaceutical composition of claim 35, wherein the antibody that specifically binds to SIRPa is an isolated monoclonal antibody.

37. The pharmaceutical composition of claim 35, wherein the antibody promotes macrophage-mediated phagocytosis of a CD47-expressing cell.

38. The pharmaceutical composition of claim 32, wherein the anti-CD47 agent is a soluble CD-47 binding SIRPa fragment.

39. The pharmaceutical composition of claim 32, wherein the anti-CD47 agent prevents interaction between CD47 and SIRPa.

40. The pharmaceutical composition of claim 32, wherein the anti-CD47 agent is an IgG isotype selected from the group consisting of IgGl isotype, IgG2 isotype, IgG3 isotype, IgG4 isotype, IG-G1-N297Q, IG4-S228P, and IG64 PE.

41. The pharmaceutical composition of claim 32, wherein the compound of formula (I) is in an amount from about 5 mg to 150 mg.

42. The pharmaceutical composition of any one of claims 32 to 41, wherein the compound of Formula (I) is selected from plinabulin, (3Z, 6Z)-3-(phenyl-2, 3,4,5, 6-ds)- methylene-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene)piperazine-2, 5-dione; (3Z,6Z)-3- (phenyl-2,3,4,5,6-d5)-methylene-d-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene)piperazine-

-153- 2,5-dione; (3Z,6Z)-3-(phenylmethylene-d)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene- d)piperazine-2, 5-dione; (3Z,6Z)-3-(phenyl-2,3,4,5,6-d5)-methylene-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(phenylmethylene)-6-((5-(tert- butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z, 6Z)-3-(phenyl-2, 3,4,5, 6-ds)- methylene-d-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)- 3-(4-Fluoro-(phenyl-2,3,5,6-d4))-methylene-6-((5-(tert-butyl)-lH-imidazol-4- yl)methylene)piperazine-2, 5-dione; (3Z,6Z)-3-(4-Fluoro-(phenyl-2,3,5,6-d4))-methylene-6- ((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3- fluorobenzylidene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-benzoylbenzylidene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene- d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-(4-fluorobenzoyl)benzylidene)-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(3-(4- methoxybenzoyl)benzylidene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine- 2, 5-dione; (3Z,6Z)-3-(3-methoxybenzylidene)-6-((5-(tert-butyl)-lH-imidazol-4- yl)methylene-d)piperazine-2, 5-dione; or (3Z,6Z)-3-(3-(trifluoromethyenzydene)-6-((5-(tert- butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione.

43. A method of treating a subject with a cancer cell expressing CD-47 detecting tumor cell expression of CD-47 in a subject; and providing the subject a therapeutic amount of a compound of Formula (I):

wherein

Ri, R4, and Re, are each separately selected from the group consisting of a hydrogen atom, a deuterium atom, a halogen atom, and saturated Ci,-C24 alkyl, unsaturated Ci-

-154- C24 alkenyl, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, azido, substituted nitro, phenyl, and substituted phenyl groups, hydroxy, carboxy, — CO — O — R7, cyano, alkylthio, halogenated alkyl including polyhalogenated alkyl, halogenated carbonyl, and carbonyl — CH2CO — R7, wherein R7 is selected from a hydrogen atom, a halogen atom, and saturated C1-C24 alkyl, unsaturated C1-C24 alkenyl, cycloalkyl, cycloalkenyl, alkoxy, cycloalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, amino, substituted amino, nitro, azido, substituted nitro, phenyl, and substituted phenyl groups;

-155- Z, for each separate n, if non-zero, and Zi, Z2, Za and Z4are each separately selected from a carbon atom, a sulfur atom, a nitrogen atom or an oxygen atom; and the dashed bonds may be either single or double bonds; and an anti-CD47 agent.

44. The method of claim 43, wherein detecting a tumor cell expression of CD-47 comprises at least one of immunocytochemistry, proteomics, mRNA quantification, or a combination thereof.

45. The method of claim 43 or 44, wherein the compound of Formula (I) is selected from plinabulin, (3Z,6Z)-3-(phenyl-2,3,4,5,6-d5)-methylene-6-((5-(tert-butyl)-lH-imidazol- 4-yl)methylene)piperazine-2, 5-dione; (3Z, 6Z)-3-(phenyl-2, 3,4,5, 6-ds)-methylene-d-6-((5-

(tert-butyl)-lH-imidazol-4-yl)methylene)piperazine-2, 5-dione; (3Z,6Z)-3-(phenylmethylene- d)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(phenyl- 2,3,4,5,6-d5)-methylene-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene-d)piperazine-2,5- dione; (3Z,6Z)-3-(phenylmethylene)-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene- d)piperazine-2, 5-dione; (3Z,6Z)-3-(phenyl-2,3,4,5,6-d5)-methylene-d-6-((5-(tert-butyl)-lH- imidazol-4-yl)methylene-d)piperazine-2, 5-dione; (3Z,6Z)-3-(4-Fluoro-(phenyl-2,3,5,6-d4))- methylene-6-((5-(tert-butyl)-lH-imidazol-4-yl)methylene)piperazine-2, 5-dione; (3Z,6Z)-3-

-156-