WO2024173376A2 - Combination therapy comprising multispecific gamma delta tcr antibodies - Google Patents

Abstract

The present disclosure relates to a method for the treatment of cancer comprising administration to a subject in need thereof, of a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vγ9Vδ2 T cell receptor, in combination with a common gamma chain cytokine and/or an immune checkpoint inhibitor.

Description

COMBINATION THERAPY COMPRISING MULTISPECIFIC GAMMA DELTA TCR ANTIBODIES

CROSS-REFERENCE TO RELATED APPLICATIONS

[1] The present application claims priority to U.S. Provisional Application 63/445,171 , filed February 13, 2023; U.S. Provisional Application 63/471 ,647, filed June 7, 2023; U.S. Provisional Application 63/547,960, filed November 9, 2023; U.S. Provisional Application 63/624,652, filed January 24, 2024; U.S. Provisional Application 63/543,364, filed October 10, 2023; U.S. Provisional Application 63/618,073, filed January 5, 2024; the contents each of which are incorporated herein by reference in their entireties.

REFERENCE TO SEQUENCE LISTING

[2] The contents of the electronic sequence listing (LVAT_013_04WO_SeqList_ST26.xml; Size: 306,129 bytes; and Date of Creation: February 13, 2024) are herein incorporated by reference in its entirety.

FIELD

[3] The present disclosure relates to methods and medical uses for the treatment of cancer comprising administration of a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vy9V52 T cell receptor in combination with a common gamma chain cytokine and/or an immune checkpoint inhibitor.

BACKGROUND

[4] Gamma-delta (yb) T cells are T cells that express a T cell receptor (TOR) that is made up of a gamma chain and a delta chain. The majority of y6 T cells express TCRs comprising Vy9 and V<52 chains. VY9V<52 T cells can react against a wide array of pathogens and tumor cells. This broad reactivity is understood to be conferred by phosphoantigens that are able to specifically activate this T-cell subset in a TOR dependent fashion. The broad antimicrobial and anti-tumor reactivity of VY9V<52 T-cells suggest a direct involvement in immune control of cancers and infections.

[5] Agents that can activate VY9V<52 T cells can be useful in the treatment of infections or cancer as these may promote VY9V<52 T cell reactivity towards the pathogen or infected cells or cancer cell. WO 2015/156673 describes antibodies that bind VY9V<52 TCRS and are capable of activating VY9V<52 T cells. WO 2020/060405 and WO 2022/008646 describe multispecific antibodies that bind both VY9V<52 T cells and a human cancer antigen and thus have the potential to recruit VY9V<52 T cells to cancer cells and thus stimulate a therapeutic effect. [6] There is still a need for improved methods for the treatment of cancer using such multispecific antibodies wherein the methods achieve an efficacious and long-lasting response in a broad population of cancer patients while minimizing adverse effects.

SUMMARY

[7] In some embodiments, the present disclosure provides a method of treating cancer in a subject in need thereof comprising administering (i) a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigenbinding region capable of binding a human Vy9V52 T cell receptor; and (ii) a dose of a common gamma chain cytokine. In some embodiments, the common gamma chain cytokine is IL-2. In some embodiments, the IL-2 is administered at a dose of less than 3 MIU per day. In some embodiments, the common gamma chain cytokine is IL-15. In some embodiments, the first antigen-binding region binds to a human cancer antigen selected from CD1d, PSMA, CD40, CD123, 5T4, Nectin-4, EGFR, and CD33. In some embodiments, the first antigen binding region binds to human PSMA.

[8] In some embodiments, the present disclosure provides a method of treating cancer in a subject in need thereof comprising administering (i) a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigenbinding region capable of binding a human Vy9V<52 T cell receptor; and (ii) an anti-PD1 or an anti-PD-L1 antibody. In some embodiments, the method comprises administration of pembrolizumab. In some embodiments, the first antigen-binding region binds to a human cancer antigen selected from CD1d, PSMA, CD40, CD123, 5T4, Nectin-4, EGFR, and CD33. In some embodiments, the first antigen binding region binds to human PSMA.

[9] In some embodiments, the present disclosure provides a method of treating cancer in a subject in need thereof comprising administering (i) a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigenbinding region capable of binding a human Vy9V<52 T cell receptor; (ii) an anti-PD1 or an anti- PD-L1 antibody; and (iii) a common gamma chain cytokine. In some embodiments, the common gamma chain cytokine is IL-2. In some embodiments, the IL-2 is administered at a dose of less than 3 MIU per day. In some embodiments, the common gamma chain cytokine is IL-15. In some embodiments, the method comprises administration of pembrolizumab. In some embodiments, the first antigen-binding region binds to a human cancer antigen selected from CD1d, PSMA, CD40, CD123, 5T4, Nectin-4, EGFR, and CD33. In some embodiments, the first antigen binding region binds to human PSMA.

[10] In some embodiments, the present disclosure provides a kit for the treatment of cancer comprising: (a) a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vy9V<52 T cell receptor; and (b) a common gamma chain cytokine, wherein the kit optionally comprises instructions for use. In some embodiments, the common gamma chain cytokine is IL-2. In some embodiments, the dose of IL-2 is less than 3 MIU per day. In some embodiments, the common gamma chain cytokine is IL-15. In some embodiments, the first antigen-binding region binds to a human cancer antigen selected from CD1d, PSMA, CD40, CD123, 5T4, Nectin-4, EGFR, and CD33. In some embodiments, the first antigen binding region binds to human PSMA.

[11] In some embodiments, the present disclosure provides a kit for the treatment of cancer comprising: (a) a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vy9V<52 T cell receptor; and (b) an anti-PD1 or anti-PD-L1 antibody, wherein the kit optionally comprises instructions for use. In some embodiments, the anti-PD1 antibody is pembrolizumab. In some embodiments, the first antigen-binding region binds to a human cancer antigen selected from CD1d, PSMA, CD40, CD123, 5T4, Nectin-4, EGFR, and CD33. In some embodiments, the first antigen binding region binds to human PSMA.

[12] In some embodiments, the present disclosure provides a kit for the treatment of cancer comprising: (a) a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vy9V<52 T cell receptor; (b) an anti-PD1 or anti-PD-L1 antibody; and (c) a common gamma chain cytokine, wherein the kit optionally comprises instructions for use. In some embodiments, the common gamma chain cytokine is IL-2. In some embodiments, the dose of IL-2 is less than 3 MIU per day. In some embodiments, the common gamma chain cytokine is IL-15. In some embodiments, the first antigen-binding region binds to a human cancer antigen selected from CD1d, PSMA, CD40, CD123, 5T4, Nectin-4, EGFR, and CD33. In some embodiments, the first antigen binding region binds to human PSMA. In some embodiments, the anti-PD1 antibody is pembrolizumab.

BRIEF DESCRIPTION OF FIGURES

[13] Fig. 1 shows Prostate Specific Antigen (PSA) responses as % change from baseline. Shown is best PSA response and, for those patients with no PSA decline, PSA value at Cycle 5 Day 1 . Dose Levels (DL) for each patient are indicated.

[14] Fig. 2 shows PSA levels over time in Pt N304.

[15] Fig. 3A - Fig. 3D show changes in Vy9V<52 T cell-related pharmacodynamic parameters over time after administration of LAVA-1207.

[16] Fig. 4A - Fig. 4B show various pharmacokinetic parameters of LAVA- 1207.

[17] Fig. 5A - Fig. 5B show a summary of adverse events during dose escalation trial of

LAVA- 1207. [18] Fig. 6A - Fig. 6E show changes in PD1 expression and frequency of Vy9V<52 T cells over time and maximum observed receptor occupancy of LAVA-1207. Fig. 6A shows relative change in PD1+ Vy9V<52 T cells after administration of LAVA-1207 compared to baseline for cohort 4. Fig. 6B shows relative change in MFI of PD1 on Vy9V<52 T cells compared to baseline for cohort 4. Fig. 6C shows relative change of Vy9V52 T cells (compared to baseline, set at 100) from the completed cohort 6A2; dashed lines indicate administration of IL-2. Fig. 6D shows Vy9V<52 T cells per ml of blood over time from completed cohort 6A2; dashed lines indicate administration of IL-2. Fig. 6E shows the increase in the percentage of LAVA-1207 receptor occupancy on Vy9V<52 T cells in the completed cohorts.

[19] Fig. 7A - Fig. 7C shows percentages of Vy9V<52 T cells (Fig. 7A), of PD1+ Vy9V<52 T cells (Fig. 7B), and of CD3+ T cells (Fig. 7C) in prostate cancer patient PBMCs, malignant (m), and non-malignant (n.m.) areas of the patient prostate.

[20] Fig. 8 shows upregulation of PD1 expression on Vy9V<52 T cells activated with a V62- bsTCE.

[21] Fig. 9 shows an exemplary dosing schedule of the disclosed embodiments.

[22] Fig. 10 shows exemplary dose cohorts of LAVA-1207 and LAVA-1207 + IL-2.

[23] Fig. 11A-FIG. 11 B show the effect of a multispecific binding agent on Vy9V<52 T cell expansion. FIG. 11 A shows the increase in Vy9V<52 T cell frequency as a percentage of CD3⁺ T cells after incubation with IL-2 and either medium alone, pamidronate or the multispecific binding agent. FIG. 11 B shows the fold expansion of Vy9V<52 T cells over baseline after incubation with IL-2 and either medium alone, pamidronate or the multispecific binding agent.

[24] Fig. 12A-Fig. 12E show the Vy9V52-T cell frequency and phenotype of patient-derived tumor and non-tumor tissue and the cytolytic activity of Vy9V52-T cells against patient-derived tumor and non-tumor tissue in the presence of LAVA-1207. Fig. 12A shows the frequency of Vy9V52-T cells as a percentage of T cells in patient PBMC, patient-derived malignant, and non-malignant tissue. Fig. 12B shows the phenotype of PBMC-, malignant-, and non- malignant tissue-derived Vy9V52-T cells and CD3+ cells, Fig. 12C shows LAVA-1207- mediated increase in Vy9V52-T cell degranulation (as assessed by CD107a expression) when malignant tissue is incubated with LAVA-1207 and PBMC. Fig. 12D shows the lack of LAVA- 1207-mediated increase in Vy9V52-T cell degranulation when non-malignant tissue is incubated with LAVA-1207 and PBMC. Fig. 12E shows the increase in tumor cell lysis when malignant tissue is incubated with LAVA-1207 and PBMCs as opposed to non-malignant cells.

[25] Fig. 13A-Fig. 13C show the phenotypical analysis of ligands and matched coreceptors on patient-derived tumor and non-tumor tissue and Vy9V52-T cells. Fig. 13A shows differences in PSMA, BTN3A, and BTN2A1 expression. Fig. 13B shows differences in Nectin- 2, PVR, MIC-A/MIC-B, ULPB1 , ULBP3, ULBP2/5/6, and HLA-E expression. Fig. 13C shows the frequency of Vy9V<52 T cells that express the DNAM-1 , NKG2D, or NKG2A receptors in PBMC, malignant, and non-malignant tissue.

[26] Fig. 14A-Fig.14H show the effect of blocking the DNAM-1 or NKG2D receptors on LAVA-1207-mediated Vy9V52-T cell cytokine production and degranulation as well as lysis of prostate cancer cell lines. Effects on IFNy (Fig. 14A), TNF (Fig. 14B), IL-2 (Fig. 14C), IL-4 (Fig. 14D) are shown, as well as effects on Vy9V<52 T cell degranulation (Fig. 14E), tumor cell lysis (Fig. 14F) in the presence or absence of DNAM-1 blocking antibodies. Fig. 14G shows the effect of blocking the NKG2D receptor on Vy9V<52 T cell degranulation. Fig. 14H shows the effect of blocking the NKG2D receptor on tumor cell lysis.

[27] Fig. 15A-Fig. 15B show the effects of blocking NKG2A (Fig. 15A) or BTN3A (Fig. 15B) on LAVA-1207-mediated T cell activation.

[28] Fig. 16 shows the effect of either anti-DNAM-1 or anti-NKG2D blocking antibodies on LAVA-1207-mediated Vy9V<52 T cell degranulation using patient prostate tumor tissue.

[29] Fig. 17A-Fig. 17B show that NCG mice implanted with prostate cancer cells alone or admixed with PBMCs and treated with LAVA- 1207 had a reduction in tumor growth volume (Fig. 17A) and an increased survival (Fig. 17B) as compared to mice that were only implanted with prostate cancer cells and treated with either PBMCs or PBS or PSMA-V52-Fc bsTCE.

[30] Fig. 18A-Fig. 18B show that labeling LAVA-1207 with FITC does not impair its binding to either Vy9V52-T cells (Fig. 18A) or prostate cancer cells (Fig. 18B).

[31] Fig. 19A-Fig. 19B show exemplary immunohistochemistry staining showing tissues positive for PSMA and binding by LAVA- 1207 (Fig. 19A) and tissues negative for PSMA and an absence of binding by LAVA-1207 (Fig. 19B). Arrows indicate potential V52+ T cells (positive binding of LAVA-1207).

[32] Fig. 20 illustrates the different CDR numbering systems (Kabat, Chothia, IMGT, and Combined) for the 6H4 VHH.

DETAILED DESCRIPTION

Overview

[33] Vy9V52-T cells represent a conserved T cell subset (approximately 1-5% of all cluster of differentiation [CD]3+ T cells) that can induce cell death in a wide range of malignant cells in an H LA-peptide-independent fashion (Lo Presti et al. 2017; de Weerdt et al. 2018; Kunzmann, Bauer, and Wilhelm 1999; Gertner-Dardenne et al. 2012).

[34] Because Vy9V52-T cells have properties of both the innate and adaptive immune systems, they serve as a functional bridge between these two critical systems to impact tumor killing. As such, not only do they have the capability to be activated for immediate and potent killing of tumor cells, they also have the potential to contribute to a cascade response in which downstream activation of innate immune cells such as natural killer (NK) cells and adaptive immune cells such as classical (alpha-beta) T cells can result in additional tumor killing. Finally, activated Vy9V52-T cells can take up, process and present antigen, which may lead to priming of the adaptive immune system resulting in immunological memory with the potential to provide deep and durable antitumor responses.

[35] The activation of Vy9V52-T cells is driven by their interaction with butyrophilin (BTN)2A1 (BTN2A1) and BTN3A1 upon binding of phosphoantigens to the intracellular B30.2 domain of BTN3A1 (Harly et al. 2012; Vantourout and Hayday 2013; Rigau et al. 2020). Phosphoantigens accumulate upon dysregulation of the mevalonate pathway and are upregulated during cellular stress, such as malignant transformation, or through pharmacological manipulation with e.g. aminobisphosphonates. In addition, the presence of other cell surface receptors such as NKG2D allow Vy9V52-T cells to interact with cells expressing NKG2D ligands MICA/B and ULBPs (Vantourout and Hayday 2013; Kong et al. 2009; Rincon-Orozco et al. 2005). Following activation, Vy9V52-T cell functions include cytotoxicity, secretion of chemokines and pro-inflammatory cytokines and antigen presentation (Vantourout and Hayday 2013; Brandes, Willimann, and Moser 2005; Devilder et al. 2006).

[36] Since the presence of Vy9V52-T cells in solid tumors strongly correlates with patient survival (Gentles et al. 2015; Tosolini et al. 2017), approaches that improve targeting and activation of Vy9V52-T cells to tumors are thought to have the potential for the development of novel, efficacious and safe cancer treatments (de Bruin et al. 2016).

[37] Interleukin (I L)-2 is a cytokine known to promote T cell proliferation and function (Nada et al. 2017; Chen et al. 2022; Li and David Pauza 2011). The rationale for exploring IL-2 as an immune modulator for LAVA-1207 is based on its known T-cell stimulatory effect. IL-2 has been shown to induce rapid expansion of Vy9V52-T cells in vitro and to also support Vy9V52- T cell expansion in both non-human primates and humans exposed to phosphoantigen stimulation (Nada et al. 2017; Sicard et al. 2005; Wilhelm et al. 2003; Dieli et al. 2007; Meraviglia et al. 2010; Bennouna et al. 2010). IL-2, and other common gamma chain cytokines such as IL-15, could therefore expand and enhance the availability of Vy9V52-T cells in patients treated with multispecific antibodies described herein (including LAVA-1207) to promote more potent responses towards tumor cells.

Definitions

[38] The term “human V52”, when used herein, refers to the rearranged 52 chain of the Vy9V52-T cell receptor (TCR). UniProtKB - A0JD36 (A0JD36_HUMAN) gives an example of a variable TRDV2 sequence. [39] The term “human VY9”, when used herein, refers to the rearranged y9 chain of the VY9V52-T cell receptor (TCR). UniProtKB - Q99603_HUMAN gives an example of a variable TRGV9 sequence.

[40] The term “PSMA”, when used herein, refers to the human Prostate-Specific Membrane Antigen protein (UniProtKB - Q04609 (FOLH1 _HUMAN)).

[41] The term “CD1d”, when used herein, refers to the human CD1d protein (UniProtKB - P15813 (CD1 D_HUMAN)).

[42] The term “CD40”, when used herein, refers to the CD40 protein, also known as tumor necrosis factor receptor superfamily member 5 (UniProtKB - P25942 (TNR5_HUMAN)), Isoform I.

[43] The term “CD123”, when used herein, refers to the human CD123 protein, also termed interleukin-3 receptor alpha chain (NCBI Reference Sequence: NP_002174.1).

[44] The terms “IL-2” or“IL2”, when used herein, refer to an interleukin-2 protein. UniProtKB - P60568 provides the WT human IL-2 amino acid sequence. The term IL-2 is inclusive of variants of the WT sequence.

[45] The terms “IL-15” or “IL15,” when used herein, refer to an interleukin-15 protein. UniProtKB-P40933 provides the WT human IL-15 amino acid sequence. The term IL-15 is inclusive of variants of the WT sequence.

[46] The term “5T4”, when used herein, refers to the human Trophoblast glycoprotein (UniProtKB - Q13641).

[47] The term “human Nectin-4”, when used herein, refers to the human Nectin-4 protein (UniProt Q96NY8 ■ NECT4_HUMAN).

[48] The term "EGFR", when used herein, refers to the human EGFR protein (UniProtKB - P00533 (EGFR_HUMAN)).

[49] The term “CD33”, when used herein, refers to the human CD33 protein, also known as Siglec3 (UniProt P20138 (CD33_HUMAN)).

[50] The term “immunoglobulin” as used herein is intended to refer to a class of structurally related glycoproteins consisting of two pairs of polypeptide chains, one pair of light (L) chains and one pair of heavy (H) chains, all four potentially inter-connected by disulfide bonds. The term “immunoglobulin heavy chain”, “heavy chain of an immunoglobulin” or “heavy chain” as used herein is intended to refer to one of the chains of an immunoglobulin. A heavy chain is typically comprised of a heavy chain variable region (abbreviated herein as VH) and a heavy chain constant region (abbreviated herein as CH) which defines the isotype of the immunoglobulin. The heavy chain constant region typically is comprised of three domains, CH1 , CH2, and CH3. The heavy chain constant region further comprises a hinge region. Within the structure of the immunoglobulin (e.g. IgG), the two heavy chains are interconnected via disulfide bonds in the hinge region. Equally to the heavy chains, each light chain is typically comprised of several regions; a light chain variable region (VL) and a light chain constant region (CL). Furthermore, the VH and VL regions may be subdivided into regions of hypervariability (or hypervariable regions which may be hypervariable in sequence and/or form structurally defined loops), also termed complementarity determining regions (CDRs), interspersed with regions that are more conserved, termed framework regions (FRs). Each VH and VL is typically composed of three CDRs and four FRs, arranged from amino-terminus to carboxy-terminus in the following order: FR1 , CDR1 , FR2, CDR2, FR3, CDR3, FR4. CDR sequences may be determined by use of various methods, e.g. the methods provided by Chothia and Lesk (1987) J. Mol. Biol. 196:901 or Kabat et al. (1991) Sequence of protein of immunological interest, fifth edition. NIH publication. Various methods for CDR determination and amino acid numbering can be compared on www.abysis.org (UCL).

[51] The term “isotype” as used herein, refers to the immunoglobulin (sub)class (for instance lgG1 , lgG2, lgG3, lgG4, IgD, IgA, IgE, or IgM) or any allotype thereof, such as lgG1m(za) and lgG1 m(f) that is encoded by heavy chain constant region genes. Each heavy chain isotype can be combined with either a kappa (K) or lambda (A) light chain. An antibody of the present disclosure can possess any isotype.

[52] The term “antibody” is intended to refer to an immunoglobulin molecule, a fragment of an immunoglobulin molecule, or a derivative of either thereof, which has the ability to specifically bind to an antigen under typical physiological conditions with a half-life of significant periods of time, such as at least about 30 minutes, at least about 45 minutes, at least about one hour, at least about two hours, at least about four hours, at least about 8 hours, at least about 12 hours, about 24 hours or more, about 48 hours or more, about 3, 4, 5, 6, 7 or more days, etc., or any other relevant functionally-defined period (such as a time sufficient to induce, promote, enhance, and/or modulate a physiological response associated with antibody binding to the antigen and/or time sufficient for the antibody to recruit an effector activity).

[53] The terms “antigen-binding region” and “antigen-binding domain” are used interchangeably herein and refer to a portion of an antibody which interacts with an antigen. Antigen-binding domains may comprise variable regions of both the heavy and light chains of the immunoglobulin molecule or may be a single-domain antigen-binding region, e.g. a heavy chain variable region only. The constant regions of an antibody, if present, may mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system and components of the complement system such as 01 q, the first component in the classical pathway of complement activation.

[54] The Fc region of an immunoglobulin is defined as the fragment of an antibody which would be typically generated after digestion of an antibody with papain which includes the two CH2-CH3 regions of an immunoglobulin and a connecting region, e.g. a hinge region. The constant domain of an antibody heavy chain defines the antibody isotype, e.g. lgG1 , lgG2, lgG3, lgG4, lgA1 , lgA2, IgM, IgD, or IgE. The Fc-region mediates the effector functions of antibodies with cell surface receptors called Fc receptors and proteins of the complement system.

[55] The term “hinge region” as used herein is intended to refer to the hinge region of an immunoglobulin heavy chain. Thus, for example, the hinge region of a human IgG 1 antibody corresponds to amino acids 216-230 according to the Ell numbering.

[56] The term “CH2 region” or “CH2 domain” as used herein is intended to refer to the CH2 region of an immunoglobulin heavy chain. Thus, for example the CH2 region of a human IgG 1 antibody corresponds to amino acids 231-340 according to the Ell numbering. However, the CH2 region may also be any of the other subtypes as described herein.

[57] The term “CH3 region” or “CH3 domain” as used herein is intended to refer to the CH3 region of an immunoglobulin heavy chain. Thus, for example the CH3 region of a human IgG 1 antibody corresponds to amino acids 341-447 according to the Ell numbering. However, the CH3 region may also be any of the other subtypes as described herein.

[58] As indicated above, the term antibody as used herein, unless otherwise stated or clearly contradicted by context, includes fragments of an antibody that retain the ability to specifically bind to the antigen. It has been shown that the antigen-binding function of an antibody may be performed by fragments of a full-length antibody. Examples of binding fragments encompassed within the term "antibody" include (i) a Fab’ or Fab fragment, i.e. a monovalent fragment consisting of the VL, VH, CL and CH1 domains, or a monovalent antibody as described in WO 2007/059782; (ii) F(ab')2 fragments, i.e. bivalent fragments comprising two Fab fragments linked by a disulfide bridge at the hinge region; (iii) a Fd fragment consisting essentially of the VH and CH1 domains; and (iv) a Fv fragment consisting essentially of the VL and VH domains of a single arm of an antibody. Furthermore, although the two domains of the Fv fragment, VL and VH, are coded for by separate genes, they may be joined, using recombinant methods, by a synthetic linker that enables them to be made as a single protein chain in which the VL and VH regions pair to form monovalent molecules (known as single chain antibodies or single chain Fv (scFv), see for instance Bird et al., Science 242, 423-426 (1988) and Huston et al., PNAS USA 85, 5879-5883 (1988)). Such single chain antibodies are encompassed within the term antibody unless otherwise indicated by context. Although such fragments are generally included within the meaning of antibody, they collectively and each independently are unique features of the present disclosure, exhibiting different biological properties and utility. The term antibody, unless specified otherwise, also includes polyclonal antibodies, monoclonal antibodies (mAbs), chimeric antibodies and humanized antibodies, and antibody fragments provided by any known technique, such as enzymatic cleavage, peptide synthesis, and recombinant techniques. [59] In some embodiments, of the antibodies of the present disclosure, the first antigenbinding region or the second antigen-binding region, or both, is a single domain antibody. Single domain antibodies are well known to the skilled person, see e.g. Hamers-Casterman et al. (1993) Nature 363:446, Roovers et al. (2007) Curr Opin Mol Ther 9:327 and Krah et al. (2016) Immunopharmacol Immunotoxicol 38:21. Single domain antibodies comprise a single CDR1 , a single CDR2, and a single CDR3. Examples of single domain antibodies are variable fragments of heavy-chain-only antibodies, antibodies that naturally do not comprise light chains, single domain antibodies derived from conventional antibodies, and engineered antibodies. Single domain antibodies may be derived from any species. For example, single domain antibodies can be derived from antibodies raised in Camelidae species, for example in camel, dromedary, llama, alpaca and guanaco. Like a whole antibody, a single domain antibody is able to bind selectively to a specific antigen. Single domain antibodies may contain only the variable domain of an immunoglobulin chain, i.e. CDR1 , CDR2 and CDR3 and framework regions. Such antibodies are also called Nanobody®, or VHH.

[60] The term “parent antibody”, is to be understood as an antibody which is identical to an antibody according to the present disclosure, but wherein the parent antibody does not have one or more of the specified mutations. An “antibody variant” or a “variant of a parent antibody” of the present disclosure is an antibody molecule which comprises one or more mutations as compared to a “parent antibody”. Amino acid substitutions may exchange a native amino acid for another naturally- occurring amino acid, or for a non-naturally- occurring amino acid derivative. The amino acid substitution may be conservative or non-conservative. In the context of the present disclosure, conservative substitutions may be defined by substitutions within the classes of amino acids reflected in one or more of TABLE 1 , TABLE 2, and TABLE 3.

TABLE 1 : Amino acid residue classes for conservative substitutions

TABLE 2: Alternative conservative amino acid residue substitution classes

TABLE 3: Alternative Physical and Functional Classifications of Amino Acid Residues

[61] The term “parent protein”, is to be understood as a protein which is identical to a protein according to the present disclosure, but wherein the parent protein does not have one or more of the specified mutations. A “variant” or “protein variant” of the present disclosure is a protein which comprises one or more amino acid mutations as compared to a “parent protein”.

[62] In the context of the present disclosure, a substitution in a variant is indicated as: Original amino acid - position - substituted amino acid; The three-letter code, or one letter code, are used, including the codes Xaa and X to indicate amino acid residue. Accordingly, the notation “T366W” means that the variant comprises a substitution of threonine with tryptophan in the variant amino acid position corresponding to the amino acid in position 366 in the parent antibody.

[63] Furthermore, the term “a substitution” embraces a substitution into any one of the other nineteen natural amino acids, or into other amino acids, such as non-natural amino acids. For example, a substitution of amino acid T in position 366 includes each of the following substitutions: 366A, 366C, 366D, 366G, 366H, 366F, 366I, 366K, 366L, 366M, 366N, 366P, 366Q, 366R, 366S, 366E, 366V, 366W, and 366Y.

[64] The term “full-length antibody” when used herein, refers to an antibody which contains all heavy and light chain constant and variable domains corresponding to those that are normally found in a wild-type antibody of that isotype.

[65] The term “chimeric antibody” refers to an antibody wherein the variable region is derived from a non-human species (e.g. derived from rodents) and the constant region is derived from a different species, such as human. Chimeric antibodies may be generated by genetic engineering. Chimeric monoclonal antibodies for therapeutic applications are developed to reduce antibody immunogenicity.

[66] The term “humanized antibody” refers to a genetically engineered non-human antibody, which contains human antibody constant domains and non-human variable domains modified to contain a high level of sequence homology to human variable domains. This can be achieved by grafting of the six non-human antibody complementarity-determining regions (CDRs), which together form the antigen binding site, onto a homologous human acceptor framework region (FR). In order to fully reconstitute the binding affinity and specificity of the parental antibody, the substitution of framework residues from the parental antibody (i.e. the non-human antibody) into the human framework regions (back-mutations) may be required. Structural homology modeling may help to identify the amino acid residues in the framework regions that are important for the binding properties of the antibody. Thus, a humanized antibody may comprise non-human CDR sequences, primarily human framework regions optionally comprising one or more amino acid back-mutations to the non-human amino acid sequence, and, optionally, fully human constant regions. Optionally, additional amino acid modifications, which are not necessarily back-mutations, may be introduced to obtain a humanized antibody with preferred characteristics, such as affinity and biochemical properties. Humanization of non-human therapeutic antibodies is performed to minimize its immunogenicity in man while such humanized antibodies at the same time maintain the specificity and binding affinity of the antibody of non-human origin.

[67] The term “multispecific antibody” or “MSAb” refers to an antibody having specificities for at least two different, such as at least three, typically non-overlapping, epitopes. Such epitopes may be on the same or on different target antigens. If the epitopes are on different targets, such targets may be on the same cell or different cells or cell types. A multispecific antibody may comprise one or more single-domain antibodies.

[68] The term “bispecific antibody” refers to an antibody having specificities for two different, typically non-overlapping, epitopes. Such epitopes may be on the same or different targets. If the epitopes are on different targets, such targets may be on the same cell or different cells or cell types. A bispecific antibody may comprise one or two single-domain antibodies.

[69] The term “bispecific T cell engager” or “bsTCE” refers to a bispecific antibody having specificities for two different epitopes, in which one of the epitopes is expressed by a T cell.

[70] Examples of different classes of multispecific, such as bispecific, antibodies include but are not limited to (i) IgG-like molecules with complementary CH3 domains to force heterodimerization; (ii) recombinant IgG-like dual targeting molecules, wherein the two sides of the molecule each contain the Fab fragment or part of the Fab fragment of at least two different antibodies; (iii) IgG fusion molecules, wherein full length IgG antibodies are fused to extra Fab fragment or parts of Fab fragment; (iv) Fc fusion molecules, wherein single chain Fv molecules or stabilized diabodies are fused to heavy-chain constant- domains, Fc-regions or parts thereof; (v) Fab fusion molecules, wherein different Fab- fragments are fused together, fused to heavy-chain constant-domains, Fc-regions or parts thereof; and (vi) ScFv-and diabody-based and heavy chain antibodies (e.g., domain antibodies, Nanobodies®) wherein different single chain Fv molecules or different diabodies or different heavy-chain antibodies (e.g. domain antibodies, Nanobodies®) are fused to each other or to another protein or carrier molecule fused to heavy-chain constant-domains, Fc-regions or parts thereof.

[71] Examples of IgG-like molecules with complementary CH3 domains molecules include but are not limited to the Triomab® (Trion Pharma/Fresenius Biotech), the Knobs-into-Holes (Genentech), CrossMAbs (Roche) and the electrostatically-matched (Amgen, Chugai, Oncomed), the LLIZ-Y (Genentech, Wranik et al. J. Biol. Chem. 2012, 287(52): 43331-9, doi: 10.1074/jbc.M 112.397869. Epub 2012 Nov 1), DIG-body and PIG-body (Pharmabcine, WO2010134666, WO2014081202), the Strand Exchange Engineered Domain body (SEEDbody) (EMD Serono), the Biclonics (Merus, WO2013157953), FcAAdp (Regeneron), bispecific lgG1 and lgG2 (Pfizer/Rinat), Azymetric scaffold (Zymeworks/Merck), mAb-Fv (Xencor), bivalent bispecific antibodies (Roche, WO 2009/080254) and DuoBody® molecules (Genmab).

[72] Examples of recombinant IgG-like dual targeting molecules include but are not limited to Dual Targeting (DT)-lg (GSK/Domantis, WO 2009/058383), Two-in-one Antibody (Genentech, Bostrom, et al 2009. Science 323, 1610-1614), Cross-linked Mabs (Karmanos Cancer Center), mAb2 (F-Star), ZybodiesTM (Zyngenia, LaFleur et al. MAbs. 2013 Mar- Apr;5(2):208-18), approaches with common light chain, K Bodies (Novlmmune, WO 2012/023053) and CovX-body® (CovX/Pfizer, Doppalapudi, V.R., et al 2007. Bioorg. Med. Chem. Lett. 17,501-506).

[73] Examples of IgG fusion molecules include but are not limited to Dual Variable Domain (DVD)-lg (Abbott), Dual domain double head antibodies (Unilever; Sanofi Aventis), IgG-like Bispecific (ImClone/Eli Lilly, Lewis et al. Nat Biotechnol. 2014 Feb;32(2):191-8), Ts2Ab (Medlmmune/AZ, Dimasi et al. J Mol Biol. 2009 Oct 30;393(3):672-92) and BsAb (Zymogenetics, WO2010111625), HERCULES (Biogen Idee), scFv fusion (Novartis), scFv fusion (Changzhou Adam Biotech Inc) and TvAb (Roche).

[74] Examples of Fc fusion molecules include but are not limited to ScFv/Fc Fusions (Academic Institution, Pearce et al Biochem Mol Biol Int. 1997 Sep;42(6):1179), SCORPION (Emergent BioSolutions/T rubion, Blankenship JW, et al. AACR 100th Annual meeting 2009 (Abstract #5465); Zymogenetics/BMS, WO2010111625), Dual Affinity Retargeting Technology (Fc-DARTTM) (MacroGenics) and Dual(ScFv)2-Fab (National Research Center for Antibody Medicine - China). [75] Examples of Fab fusion bispecific antibodies include but are not limited to F(ab)2 (Medarex/AMGEN), Dual-Action or Bis-Fab (Genentech), Dock-and-Lock® (DNL) (ImmunoMedics), Bivalent Bispecific (Biotecnol) and Fab-Fv (UCB-Celltech).

[76] Examples of ScFv-, diabody-based and domain antibodies include but are not limited to Bispecific T Cell Engager (BiTE®) (Micromet, Tandem Diabody (Tandab) (Affimed), Dual Affinity Retargeting Technology (DARTTM) (MacroGenics), Single-chain Diabody (Academic, Lawrence FEBS Lett. 1998 Apr 3;425(3):479-84), TCR-like Antibodies (AIT, ReceptorLogics), Human Serum Albumin ScFv Fusion (Merrimack, W02010059315) and COMBODY molecules (Epigen Biotech, Zhu et al. Immunol Cell Biol. 2010 Aug;88(6):667-75), dual targeting nanobodies® (Ablynx, Hmila et al., FASEB J. 2010), dual targeting heavy chain only domain antibodies.

[77] In some embodiments, the multispecific antibody used in the present disclosure is in a VHH-Fc format, i.e. the antibody comprises two or more single-domain antigen-binding regions that are linked to each other via a human Fc region dimer. In this format, each singledomain antigen-binding region is fused to an Fc region polypeptide and the two fusion polypeptides form a dimeric bispecific antibody via disulfide bridges in the hinge region. Such constructs typically do not contain full, or any, CH1 or light chain sequences. Figure 12B of W006064136 provides an illustration of an example of this format.

[78] In the context of antibody binding to an antigen, the terms “binds”, “capable of binding” or “specifically binds” refer to the binding of an antibody to a predetermined antigen or target (e.g. human V<52 or human PSMA) to which binding typically is with an apparent affinity corresponding to a KD of about 10'⁶ M or less, e.g. 10'⁷ M or less, such as about 10'⁸ M or less, such as about 10'⁹ M or less, about 10'¹⁰ M or less, or about 10-¹¹ M or even less, e.g. when determined using flow cytometry. Alternatively, KD values can be determined using for instance surface plasmon resonance (SPR) technology in a BIAcore T200 or bio-layer interferometry (BLI) in an Octet RED96 instrument using the antigen as the ligand and the binding moiety or binding molecule as the analyte. Specific binding means that the antibody binds to the predetermined antigen with an affinity corresponding to a KD that is at least tenfold lower, such as at least 100-fold lower, for instance at least 1 ,000 fold lower, such as at least 10,000 fold lower, for instance at least 100,000 fold lower than its affinity for binding to a non-specific antigen (e.g., BSA, casein) other than the predetermined antigen or a closely- related antigen. The degree with which the affinity is lower is dependent on the KD of the binding moiety or binding molecule, so that when the KD of the binding moiety or binding molecule is very low (that is, the binding moiety or binding molecule is highly specific), then the degree with which the affinity for the antigen is lower than the affinity for a non-specific antigen may be at least 10,000-fold. The term "KD" (M), as used herein, refers to the dissociation equilibrium constant of a particular interaction between the antigen and the binding moiety or binding molecule.

[79] “Capable of binding the V<52 chain of a VY9V52-TCR” or “binding the V<52 chain of a VY9V52-TCR” or the like means that the antibody can bind the V<52 chain as a separate molecule and/or as part of a VY9V52-TCR. However, the antibody will not bind to the VY9 chain as a separate molecule.

[80] “Capable of binding the VY9 chain of a VY9V52-TCR” or “binding the VY9 chain of a VY9V52-TCR” or the like means that the antibody can bind the VY9 chain as a separate molecule and/or as part of a VY9V52-TCR. However, the antibody will not bind to the V<52 chain as a separate molecule.

[81] In the context of the present disclosure, “competition” or “able to compete” or “competes” refers to any detectably significant reduction in the propensity for a particular binding molecule (e.g. an PSMA binding antibody) to bind a particular binding partner (e.g. PSMA) in the presence of another molecule (e.g. a different PSMA antibody) that binds the binding partner. Typically, competition means an at least about 25 percent reduction, such as an at least about 50 percent, e.g. an at least about 75 percent, such as an at least 90 percent reduction in binding, caused by the presence of another molecule, such as an antibody, as determined by, e.g., ELISA analysis or flow cytometry using sufficient amounts of the two or more competing molecules, e.g. antibodies. Additional methods for determining binding specificity by competitive inhibition may be found in for instance Harlow et al., Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1988), Colligan et al., eds., Current Protocols in Immunology, Greene Publishing Assoc, and Wiley InterScience N. Y., (1992, 1993), and Muller, Meth. Enzymol. 92, 589-601 (1983)).

[82] The method or use of the present disclosure may involve the use of an antibody that binds to the same epitope on a target (e.g. V<52 or PSMA) as antibody described herein. There are several methods available for mapping antibody epitopes on target antigens known in the art, including but not limited to: crosslinking coupled mass spectrometry, allowing identification of peptides that are part of the epitope, and X-ray crystallography identifying individual residues on the antigen that form the epitope. Epitope residues can be determined as being all amino acid residues with at least one atom less than or equal to 5 A from the antibody. 5 A was chosen as the epitope cutoff distance to allow for atoms within a van der Waals radius plus a possible water- mediated hydrogen bond. Next, epitope residues can be determined as being all amino acid residues with at least one atom less than or equal to 8 A. Less than or equal to 8 A is chosen as the epitope cutoff distance to allow for the length of an extended arginine amino acid. Crosslinking coupled mass spectrometry begins by binding the antibody and the antigen with a mass labeled chemical crosslinker. Next the presence of the complex is confirmed using high mass MALDI detection. Because after crosslinking chemistry the Ab/Ag complex is extremely stable, many various enzymes and digestion conditions can be applied to the complex to provide many different overlapping peptides. Identification of these peptides is performed using high resolution mass spectrometry and MS/MS techniques. Identification of the crosslinked peptides is determined using mass tag linked to the crosslinking reagents. After MS/MS fragmentation and data analysis, peptides that are crosslinked and are derived from the antigen are part of the epitope, while peptides derived from the antibody are part of the paratope. All residues between the most N- and C-terminal crosslinked residue from the individual crosslinked peptides found are considered to be part of the epitope or paratope.

[83] The terms “first” and “second” antigen-binding regions when used herein do not refer to their orientation I position in the antibody, i.e. they have no meaning with regard to the N- or C-terminus. The terms “first” and “second” only serve to correctly and consistently refer to the two different antigen-binding regions in the claims and the description.

[84] “ % sequence identity”, when used herein, refers to the number of identical nucleotide or amino acid positions shared by different sequences (i.e., % identity = # of identical positions/total # of positions x 100), taking into account the number of gaps, and the length of each gap, which need to be introduced for optimal alignment. The percent identity between two nucleotide or amino acid sequences may e.g. be determined using the algorithm of E. Meyers and W. Miller, Comput. Appl. Biosci 4, 11-17 (1988) which has been incorporated into the ALIGN program (version 2.0), using a PAM 120 weight residue table, a gap length penalty of 12 and a gap penalty of 4.

[85] “ Treatment” or “treating” refers to the administration of an effective amount of an antibody, common gamma chain cytokine, and/or immune checkpoint inhibitor according to the present disclosure with the purpose of easing, ameliorating, arresting, eradicating (curing) or preventing symptoms or disease states.

[86] An "effective amount" or “an amount effective to treat” refer to an amount of an agent described herein (e.g., a multispecific antibody, common gamma chain cytokine, and/or immune checkpoint inhibitor, either alone or in combination) effective, at dosages and for periods of time necessary, to achieve a desired therapeutic result (e.g. , prevent or treat cancer in an individual). An effective amount of a polypeptide, such as an antibody, may vary according to factors such as the disease stage, age, sex, and weight of the individual, the timing of administration, route of administration; the duration of the treatment; drugs used in combination; the judgment of the prescribing physician; and like factors known in the medical arts. An effective amount is also one in which any toxic or detrimental effects of the antibody are outweighed by the therapeutically beneficial effects. Administration may be carried out by any suitable route, but will typically be parenteral, such as intravenous, intramuscular, or subcutaneous. The size of the dose will also be determined by the existence, nature, and extent of any adverse side- effects that might accompany the administration of a particular active, and the desired physiological effect. It will be appreciated by one of skill in the art that various diseases or disorders could require prolonged treatment involving multiple administrations, perhaps using various rounds of administration.

[87] Herein, a “dosing interval” refers to the frequency of administration of a particular agent (e.g., the frequency of administration of a multispecific antibody, common gamma chain cytokine, or immune checkpoint inhibitor) during the course of treatment.

Combination therapy

[88] In some embodiments, the present disclosure provides methods of treating a cancer in a subject in need thereof comprising administering to the subject (i) a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vy9V<52 T cell receptor and (ii) a common gamma chain cytokine.

[89] The term “common gamma chain cytokine” as used herein refers to a cytokine that binds the common gamma chain receptor (common yc), also known as CD132. Common gamma chain cytokines include IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21. In some embodiments, the methods and kits described herein utilize IL-2 or IL-15. IL-2 and IL-15 bind a common heterodimeric receptor composed of the I L-2/15Rp (CD122) and the common y chains. The specificity of action of both cytokines is conferred by their corresponding a receptor chains, IL- 2Ra (CD25) and IL-15Ra (CD215).

IL-2 Combination Therapy

[90] In some embodiments, the present disclosure provides methods of treating a cancer in a subject in need thereof comprising administering to the subject (i) a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vy9V<52 T cell receptor and (ii) IL- 2.

[91] The IL-2 used in the method of the present disclosure may be recombinant IL-2. IL-2 may be Aldesleukin (Proleukin®) for example presented in a 5 mL vial with 1.3 mg Aldesleukin/Proleukin (22 x 10⁶ IU (international units) per vial), wherein the product can be reconstituted using sterile water for injection. Other suitable forms of IL-2 include NKTR-214 (Bempegaldesleukin/Nektar), ALK 4230 (nemvaleukin/Aalkermes-Reliant), SAR444245 - THOR 707/Synthorx/Sanofi) and XTX-202 (Xilio). In some embodiments, the IL-2 is a variant IL-2. In some embodiments, the variant IL-2 binds to the IL-2Rbeta and gamma chains, but does not bind or demonstrates reduced/abrogated binding to the IL-2Ralpha chain (e.g. NL201 from Neoleukin, Silva et al. De novo design of potent and selective mimics of IL-2 and IL-15. Nature 565, 186-191 (2019), and MDNA11 from Medicenna). In some embodiments, the IL- 2 and/or variants thereof is pegylated. IL-2 may be administered via any suitable administration route, for example subcutaneously.

[92] In some embodiments, the multispecific antibody is administered prior to the administration of IL-2. In some embodiments, IL-2 is administered at least once, at least twice, at least three time, at least four time, at least five times, at least six times, or at least seven times after the administration of the multispecific antibody. For example, in some embodiments, the method comprises administration of a multispecific antibody followed by 1 , 2, 3, 4, 5, 6, 7, or more daily doses of IL-2. In some embodiments, the multispecific antibody is administered concurrently with the administration of IL-2. In some embodiments, the IL-2 is administered prior to the administration of the multispecific antibody.

IL-2 dosing

[93] IL-2 or a variant thereof may be administered at any suitable dose. IL-2 may for example be given at a dose of between 0.1 and 5 MIU (0.1 and 5x10⁶ IU), between 0.1 and 2.5 MIU/day, between 0.1 and 3.5 MIU/day, between 0.1 and 4.5 MIU, between 0.5 and 3.5 MIU/day, between 0.5 and 2.5 MIU/day, or between 0.5 and 1.5 MIU/day. In some embodiments, IL-2 is given at a dose of 0.1 , 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, or 1.0 MIU. In some embodiments, IL-2 is given at a daily dose of 1.0 MIU. In some embodiments, IL-2 is given at a dose of 1.1 , 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1 , 2.2, 2.3, 2.4, 2.5, 2.6, 2.7,

2.8, 2.9, or 3.0 MIU. In some embodiments, IL-2 is given at a dose of 2.0 MIU. In some embodiments, IL-2 is given at a dose of 3.0 MIU.

[94] Low dose subcutaneous interleukin-2 (LDSC IL-2) is currently used and studied for the management of immune mediated diseases, and well tolerated, when administered at low doses (< 1 MIU/day (Mahmoudpour et al. 2019; de Bono et al. 2022). In an ongoing study of ICT01 , a BTN3A specific monoclonal antibody that stimulates Vy9V52-T cell activation in patients with advanced solid tumors (EVICTION-2 trial), the addition of LDSC IL-2 at a dose of 1 MIU/m2 (administered on days 1 , 2, 3, 4 and 5 following ICT01) was reported to be safe and to induce Vy9V52-T cell expansion in 6/6 evaluable patients (de Bono et al. 2022). As such, in some embodiments, IL-2 is given as a “low-dose”. A low dose of IL-2 refers to a dose that is less than 3MIU. In some embodiments, IL-2 is given at a dose of less than 3.0 MIU. In some embodiments, IL-2 is given at a dose of less than 2.0 MIU. In some embodiments, IL-2 is given at a dose of less than 1.0 MIU. IL-2 may also be given multiple times daily, for example twice daily at a dose of between 0.1 and 5 MIU (0.1 and 5x10⁶ IU), such as a dose of between 0.2 and 2 MIU, e.g. 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1 , 1.1 , 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8,

1.9, 2.0 MIU. In some embodiments, in IL-2 is dosed at different amounts depending on the treatment cycle. In some embodiments, IL-2 is dosed at a lower amount during the first treatment cycle and the dose is increased during subsequent treatment cycles. [95] IL-2 may be administered with any suitable dosing interval, for example a dosing interval of 1 day to 1 month, for example a dosing interval of 1 to 21 days, such as 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20 or 21 days or a dose interval of between 7 and 21 days, such as 7 or 14 days. In some embodiments, the dosing of the IL-2 is multiple times daily, for example the dose of IL-2 can be dosed 1 , 2, 3, 4, 5, or 6 times daily. In some embodiments, the IL-2 is dosed once daily. In some embodiments, the IL-2 is dosed twice daily.

IL-15 Combination Therapy

[96] In some embodiments, the present disclosure provides methods of treating a cancer in a subject in need thereof comprising administering to the subject (i) a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vy9V<52 T cell receptor and (ii) IL- 15.

[97] Suitable forms of IL-15 are known in the art. See e.g., Int J Mol Sci. 2022 Jul; 23(13): 7311 , which is herein incorporated by reference in its entirety. In some embodiments, the IL- 15 is a heterodimer of IL-15 and IL-15 receptor alpha (hetlL-15), which is also referred to as NIZ985. In some embodiments, the IL-15 is N-803, formerly known as ALT-803, which is an IL-15 variant complexed with a human IL-15Ra sushi domain-Fc fusion protein. In some embodiments, the IL-15 is SOT101 , also referred to as Nanrilkefusp alfa, which is a human fusion protein comprising the cytokine IL-15 and the high-affinity binding sushi+ domain of IL- 15 receptor alpha (IL-15Ra). In some embodiments, the IL-15 is NKTR-255, which is a polyethylene glycol-conjugate of rhlL-15. In some embodiments, NKTR-255 is dosed at 1.5 pg/kg, 3.0 pg/kg, or 3.0/6.0 pg/kg. In some embodiments, the IL-15 and/or variants thereof is pegylated.

[98] In some embodiments, the multispecific antibody is administered prior to the administration of IL-15. In some embodiments, IL-15 is administered at least once, at least twice, at least three times, at least four times, at least five times, at least six times, or at least seven times after the administration of the multispecific antibody. For example, in some embodiments, the method comprises administration of a multispecific antibody followed by 1 , 2, 3, 4, 5, 6, 7, or more daily doses of IL-15. In some embodiments, the multispecific antibody is administered concurrently with the administration of IL-15. In some embodiments, the IL-15 is administered prior to the administration of the multispecific antibody.

IL-15 dosing

[99] IL-15 may be administered at any suitable dose. In some embodiments, the IL-15 is given at a dose of between 0.001-1000 MIU (0.001 and 1000x10⁶ IU)). In some embodiments, the dose of IL-15 is between 0.01-100 MIU. In some embodiments, the dose of IL-15 is between 0.1-10 Mill. In some embodiments, the dose of IL-15 is between 0.5-5 Mill. In some embodiments, the dose of IL-15 is between 1-5 MIU. In some embodiments, the dose of IL-15 is between 2-5 MIU. In some embodiments, the IL-15 is given at a dose of between 0.5 and 1.5 MIU/day, e.g. 0.5, 0.6, 0.7, 0.8, 0.9, 1 , 1.1 , 1.2, 1.3, 1 ,4 or 1.5 MIU/day. IL-15 may also be given multiple times daily, for example twice daily at a dose of between 0.1 and 5 MIU (0.1 and 5 10⁶ IU), such as a dose of between 0.2 and 2MIU/day, such as between 0.5 and 1.5 MIU, e.g. 0.5, 0.6, 0.7, 0.8, 0.9, 1 , 1.1 , 1.2, 1.3, 1 ,4 or 1.5 MIU.

[100] In some embodiments, the IL-15 is given at a dose of between 0.01-40 pg/kg. In some embodiments, the dose of IL-15 is between 0.05-20 pg/kg. In some embodiments, the dose of IL-15 is between 0.1-10 pg/kg. In some embodiments, the dose of IL-15 is between 1-15 pg/kg. In some embodiments, the dose is any dose between 1 and 15 pg/kg, such as 1 pg/kg, 1.5 pg/kg, 2 pg/kg, 2.5 pg/kg, 3 pg/kg, 3.5 pg/kg, 4 pg/kg, 4.5 pg/kg, 5 pg/kg, 5.5 pg/kg, 6 pg/kg, 6.5 pg/kg, 7 pg/kg, 7.5 pg/kg, 8 pg/kg, 8.5 pg/kg, 9 pg/kg, 9.5 pg/kg, 10 pg/kg, 10.5 pg/kg, 11 pg/kg, 11.5 pg/kg, 12 pg/kg, 12.5 pg/kg, 13 pg/kg, 13.5 pg/kg, 14 pg/kg, 14.5 pg/kg, or 15 pg/kg. In some embodiments, in IL-15 is dosed at different amounts depending on the treatment cycle. In some embodiments, IL-15 is dosed at a lower amount during treatment cycle one and the dose is increased during subsequent treatment cycles.

[101] IL-15 may be administered with any suitable dosing interval, for example a dosing interval of 1 day to 1 month, for example a dosing interval of 1 to 21 days, such as 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20 or 21 days or a dose interval of between 7 and 21 days, such as 7 or 14 days. In some embodiments, the dosing of the IL-15 is multiple times daily, for example the dose of IL-15 can be dosed 1 , 2, 3, 4, 5, or 6 times daily. In some embodiments, the IL-15 is dosed once daily. In some embodiments, the IL-15 is dosed twice daily.

Immune checkpoint inhibitor Combination Therapy

[102] In some embodiments, the present disclosure provides methods of treating a cancer in a subject in need thereof comprising administering to the subject (i) a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vy9V<52 T cell receptor and (ii) an immune checkpoint inhibitor.

[103] Immune checkpoint inhibitors are known in the art, including agents that target PD-1 , PD-L1 , CTLA4, LAG-3, TIM-3, TIGIT, and/or NKG2A. In some embodiments, the immune checkpoint inhibitors target PD-1 or PD-L1

[104] In some embodiments, the immune checkpoint inhibitor is an anti-PD1 antibody or an anti-PD-L1 antibody. In some embodiments, the anti-PD1 antibody is selected from pembrolizumab (Keytruda®), nivolumab (Opdivo®), Zynz™ (retifanlimab), Jemperli™ (dostarlimuab), cemiplimab (Libtayo®), Tislelizumab (BGB-A317), and Sintilimab (IBI308). In some embodiments, the anti-PD-L1 antibody is selected from atezolizumab (Tecentriq®), durvalumab (Imfinzi®), and avelumab (Bavencio®). In some embodiments, the anti-PD1 antibody is pembrolizumab.

[105] In some embodiments, the immune checkpoint inhibitor is an anti-CTLA4 antibody. In some embodiments, the anti-CTLA4 antibody is ipilimumab (Yervoy®). In some embodiments, the immune checkpoint inhibitor is an inhibitor of LAG3. In some embodiments, the inhibitor is an anti-LAG3 antibody selected from relatlimab, Tebotelimab, or Favezelimab. Additional LAG-3 antibody and inhibitors are summarized in Huo et al., The promising immune checkpoint LAG-3 in cancer immunotherapy: from basic research to clinical application. Front Immunol. 2022 Jul 26;13:956090, incorporated herein by reference. In some embodiments, the immune checkpoint inhibitor is an inhibitor of TIM3 or TIGIT. Antibodies and inhibitors of TIM3 or TIGIT are descried in Cai et al. Targeting LAG-3, TIM-3, and TIGIT for cancer immunotherapy. J Hematol Oncol 16, 101 (2023), incorporated herein by reference. In some embodiments, the immune checkpoint inhibitor is an inhibitor of NKG2A. In some embodiments, the inhibitor of NKG2A is Monalizumab.

[106] In some embodiments, the multispecific antibody is administered prior to the administration of the immune checkpoint inhibitor (e.g., an anti-PD1 antibody). In some embodiments, the multispecific antibody is administered concurrently with the administration of immune checkpoint inhibitor (e.g., an anti-PD1 antibody). In some embodiments, the immune checkpoint inhibitor (e.g., an anti-PD1 antibody) is administered prior to the administration of the multispecific antibody.

Immune checkpoint inhibitor dosing

[107] In some embodiments, the immune checkpoint inhibitor is administered at any suitable dose and dosing interval according to the agent in use. In some embodiments, the immune checkpoint inhibitor is an antibody and is given at a dose of between 100 and 1000, such as a dose of between 200-800 mg, such as dose between 300-500 mg, such as a dose of 400 mg. In some embodiments, the dosing is every 6 weeks. For example, in some embodiments, the immune checkpoint inhibitor is an anti-PD1 antibody. In some embodiments, the immune checkpoint inhibitor antibody is administered according to the instructions on the approved label.

[108] In some embodiments, the anti-PD1 antibody is pembrolizumab. In some embodiments, pembrolizumab is administered at a dose specified on the drug label for the appropriate indication. The pembrolizumab label is available at accessdata.fda.gov/drugsatfda_docs/label/2021/125514s096lbl.pdf. In some embodiments, pembrolizumab is dosed at 400 mg IV over 30 minutes every 6 weeks. The nivolumab label is available at accessdata.fda.gov/drugsatfda_docs/label/2018/125554s058lbl.pdf. Additional drug labels for approved products are available at the FDA website.

[109] The anti-PD1 or anti-PD-L1 antibody may be administered with any suitable dosing interval, for example a dosing interval of 1 day to 2 month, for example a dosing interval of 1 to 50 days, such as 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , 28, 35, 42, or 49 days or a dose interval of between 7 and 49 days, such as 7, 14, 21 , 28, 35, or 42 days. In some embodiments, the anti-PD1 antibody is administered every 42 days {e.g., every 6 weeks).

[110] In some embodiments, the multispecific antibody is administered prior to the administration of the immune checkpoint inhibitor (e.g., an anti-PD1 antibody). In some embodiments, the multispecific antibody is administered concurrently with the administration of immune checkpoint inhibitor (e.g., an anti-PD1 antibody). In some embodiments, the immune checkpoint inhibitor (e.g., an anti-PD1 antibody) is administered prior to the administration of the multispecific antibody.

[111] In some embodiments, the immune checkpoint inhibitor (e.g., an anti-PD1 antibody) is used in combination with a common gamma chain cytokine (e.g., IL-2 or IL-15) described above. In some embodiments, the multispecific antibody is administered prior to the administration of the immune checkpoint inhibitor (e.g., an anti-PD1 antibody) and IL-2 or IL- 15. In some embodiments, the multispecific antibody is administered concurrently with the administration of immune checkpoint inhibitor (e.g., an anti-PD1 antibody) and IL-2 or IL-15. In some embodiments, the immune checkpoint inhibitor (e.g., an anti-PD1 antibody) and IL-2 or IL-15 are administered prior to the administration of the multispecific antibody. For example, IL-2 or IL-15 and/or anti-PD1 or anti-PD-L1 antibody may be administered daily for at least 1 day, such as 2, 3, 4, 5, 6, 7 or more days after administration of the multispecific antibody, for example starting the day after administration of the multispecific antibody. For example, the multispecific antibody may be administered on day 1 and IL-2 or IL-15 and/or anti-PD1 or anti- PD-L1 antibody may be administered on day 2 or on day 2, 3 and 4. Such a cycle may be repeated at least 1 time, such as 2, 3, 4, 5, 6, 7, 8 or more times, e.g. with intervals between the multispecific antibody administration of 14 days.

Multispecific antibody dosing

[112] The multispecific antibody may be administered with any suitable dosing interval, for example a dosing interval of 1 day to 1 month, for example a dosing interval of 1 to 21 days, such as 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20 or 21 days or a dose interval of between 7 and 21 days, such as 7 or 14 days. In some embodiments, the multispecific antibody is administered every 14 days. [113] The multispecific antibody may be administered at any suitable dose. In some embodiments, the multispecific antibody is administered at a dose of at least 1 pg, at least 2 pg, at least 3 pg, at least 4 pg, at least 5 pg, at least 6 pg, at least 7 pg, at least 8 pg, at least 9 pg, at least 10 pg, at least 11 pg, at least 12 pg, at least 13 pg, at least 14 pg, at least 15 pg, at least 16 pg, at least 17 pg, at least 18 pg, at least 19 pg, at least 20 pg, at least 25 pg, at least 30 pg, at least 35 pg, at least 40 pg, or at least 45 pg. In some embodiments, the multispecific antibody is administered at a dose of at least 100 pg, at least 110 pg, at least 120 pg, or at least 130 pg. In some embodiments, the multispecific antibody is administered at a dose of at least 300 pg, at least 310 pg, at least 320 pg, at least 330 pg, at least 340 pg, at least 350 pg, at least 360 pg, at least 370 pg, at least 380 pg, at least 390 pg, or at least 400 pg. In some embodiments, the multispecific antibody is administered at a dose of at least 500 pg, at least 510 pg, at least 520 pg, at least 530 pg, at least 540 pg, at least 550 pg, at least 560 pg, at least 570 pg, at least 580 pg, at least 590 pg, or at least 600 pg. In some embodiments, the multispecific antibody is administered at a dose of at least 750 pg, at least 760 pg, at least 770 pg, at least 780 pg, at least 790 pg, at least 800 pg, at least 810 pg, at least 820 pg, at least 830 pg, at least 840 pg, or at least 850 pg. In some embodiments, the multispecific antibody is administered at a dose of at least 1000 pg, at least 1.1 mg, at least 1.2 mg, at least 1.3 mg, at least 1.4 mg, at least 1.5 mg, at least 1.6 mg, at least 1.7 mg, at least 1.8 mg, at least 1.9 mg, or at least 2 mg. In some embodiments, the multispecific antibody is administered at a dose of at least 2.5 mg, at least 3 mg, at least 3.5 mg, at least 4 mg, at least 4.5 mg, at least 5 mg, at least 5.5 mg, at least 6 mg, at least 6.5 mg, at least 7 mg, at least 7.5 mg, at least 8 mg, at least 8.5 mg, at least 9 mg, at least 9.5 mg, or at least 10 mg. In some embodiments, the multispecific antibody is administered at a dose of at least 20 mg, at least 30 mg, at least 40 mg, at least 50 mg, at least 60 mg, at least 70 mg, at least 80 mg, at least 90 mg, or at least 100 mg. In some embodiments, he multispecific antibody is administered at a dose of at least 110 mg, at least 120 mg, at least 130 mg, at least 140 mg, at least 150 mg, at least 160 mg, at least 170 mg, at least 180 mg, at least 190 mg, or at least 200 mg. In some embodiments, the multispecific antibody is administered at a dose between 360 micrograms and 10 mg per administration. In some embodiments, the multispecific antibody is administered at a dose between 10 mg and 100 mg per administration.

[114] In some embodiments, the multispecific antibody is administered once every 14 days at a dose of at least 1 pg, at least 2 pg, at least 3 pg, at least 4 pg, at least 5 pg, at least 6 pg, at least 7 pg, at least 8 pg, at least 9 pg, at least 10 pg, at least 11 pg, at least 12 pg, at least 13 pg, at least 14 pg, at least 15 pg, at least 16 pg, at least 17 pg, at least 18 pg, at least 19 pg, at least 20 pg, at least 25 pg, at least 30 pg, at least 35 pg, at least 40 pg, or at least 45 pg. In some embodiments, the multispecific antibody is administered once every 14 days at a dose of at least 100 pg, at least 110 pg, at least 120 pg, or at least 130 pg. In some embodiments, the multispecific antibody is administered once every 14 days at a dose of at least 300 pg, at least 310 pg, at least 320 pg, at least 330 pg, at least 340 pg, at least 350 pg, at least 360 pg, at least 370 pg, at least 380 pg, at least 390 pg, or at least 400 pg. In some embodiments, the multispecific antibody is administered once every 14 days at a dose of at least 500 pg, at least 510 pg, at least 520 pg, at least 530 pg, at least 540 pg, at least 550 pg, at least 560 pg, at least 570 pg, at least 580 pg, at least 590 pg, or at least 600 pg. In some embodiments, the multispecific antibody is once every 14 days administered at a dose of at least 750 pg, at least 760 pg, at least 770 pg, at least 780 pg, at least 790 pg, at least 800 pg, at least 810 pg, at least 820 pg, at least 830 pg, at least 840 pg, or at least 850 pg. In some embodiments, the multispecific antibody is administered once every 14 days at a dose of at least 1000 pg, at least 1.1 mg, at least 1.2 mg, at least 1.3 mg, at least 1.4 mg, at least 1.5 mg, at least 1 .6 mg, at least 1.7 mg, at least 1.8 mg, at least 1.9 mg, or at least 2 mg. In some embodiments, the multispecific antibody is administered once every 14 days at a dose of at least 2.5 mg, at least 3 mg, at least 3.5 mg, at least 4 mg, at least 4.5 mg, at least 5 mg, at least 5.5 mg, at least 6 mg, at least 6.5 mg, at least 7 mg, at least 7.5 mg, at least 8 mg, at least 8.5 mg, at least 9 mg, at least 9.5 mg, or at least 10 mg. In some embodiments, the multispecific antibody is administered once every 14 days at a dose between 360 micrograms and 10 mg per administration.

[115] In some embodiments, the multispecific antibody is administered twice every 14 days at a dose of at least 1 pg, at least 2 pg, at least 3 pg, at least 4 pg, at least 5 pg, at least 6 pg, at least 7 pg, at least 8 pg, at least 9 pg, at least 10 pg, at least 11 pg, at least 12 pg, at least 13 pg, at least 14 pg, at least 15 pg, at least 16 pg, at least 17 pg, at least 18 pg, at least 19 pg, at least 20 pg, at least 25 pg, at least 30 pg, at least 35 pg, at least 40 pg, or at least 45 pg. In some embodiments, the multispecific antibody is administered twice every 14 days at a dose of at least 100 pg, at least 110 pg, at least 120 pg, or at least 130 pg. In some embodiments, the multispecific antibody is administered twice every 14 days at a dose of at least 300 pg, at least 310 pg, at least 320 pg, at least 330 pg, at least 340 pg, at least 350 pg, at least 360 pg, at least 370 pg, at least 380 pg, at least 390 pg, or at least 400 pg. In some embodiments, the multispecific antibody is administered twice every 14 days at a dose of at least 500 pg, at least 510 pg, at least 520 pg, at least 530 pg, at least 540 pg, at least 550 pg, at least 560 pg, at least 570 pg, at least 580 pg, at least 590 pg, or at least 600 pg. In some embodiments, the multispecific antibody is twice every 14 days administered at a dose of at least 750 pg, at least 760 pg, at least 770 pg, at least 780 pg, at least 790 pg, at least 800 pg, at least 810 pg, at least 820 pg, at least 830 pg, at least 840 pg, or at least 850 pg. In some embodiments, the multispecific antibody is administered twice every 14 days at a dose of at least 1000 pg, at least 1.1 mg, at least 1.2 mg, at least 1.3 mg, at least 1.4 mg, at least 1.5 mg, at least 1 .6 mg, at least 1.7 mg, at least 1.8 mg, at least 1.9 mg, or at least 2 mg. In some embodiments, the multispecific antibody is administered twice every 14 days at a dose of at least 2.5 mg, at least 3 mg, at least 3.5 mg, at least 4 mg, at least 4.5 mg, at least 5 mg, at least 5.5 mg, at least 6 mg, at least 6.5 mg, at least 7 mg, at least 7.5 mg, at least 8 mg, at least 8.5 mg, at least 9 mg, at least 9.5 mg, or at least 10 mg. In some embodiments, the multispecific antibody is administered twice every 14 days at a dose between 360 micrograms and 10 mg per administration.

[116] In some embodiments, the multispecific antibody is administered at a dose of at least

I pg/kg, at least 2 pg/kg, at least 3 pg/kg, at least 4 pg/kg, at least 5 pg/kg, at least 6 pg/kg, at least 7 pg/kg, at least 8 pg/kg, at least 9 pg/kg, at least 10 pg/kg, at least 11 pg/kg, at least 12 pg/kg, at least 13 pg/kg, at least 14 pg/kg, at least 15 pg/kg, at least 16 pg/kg, at least 17 pg/kg, at least 18 pg/kg, at least 19 pg/kg, at least 20 pg/kg, at least 25 pg/kg, at least 30 pg/kg, at least 35 pg/kg, at least 40 pg/kg, or at least 45 pg/kg. In some embodiments, the multispecific antibody is administered at a dose of at least 100 pg/kg, at least 110 pg/kg, at least 120 pg/kg, or at least 130 pg/kg. In some embodiments, the multispecific antibody is administered at a dose of at least 300 pg/kg, at least 310 pg/kg, at least 320 pg/kg, at least 330 pg/kg, at least 340 pg/kg, at least 350 pg/kg, at least 360 pg/kg, at least 370 pg/kg, at least 380 pg/kg, at least 390 pg/kg, or at least 400 pg/kg. In some embodiments, the multispecific antibody is administered at a dose of at least 500 pg/kg, at least 510 pg/kg, at least 520 pg/kg, at least 530 pg/kg, at least 540 pg/kg, at least 550 pg/kg, at least 560 pg/kg, at least 570 pg/kg, at least 580 pg/kg, at least 590 pg/kg, or at least 600 pg/kg. In some embodiments, the multispecific antibody is administered at a dose of at least 750 pg/kg, at least 760 pg/kg, at least 770 pg/kg, at least 780 pg/kg, at least 790 pg/kg, at least 800 pg/kg, at least 810 pg/kg, at least 820 pg/kg, at least 830 pg/kg, at least 840 pg/kg, or at least 850 pg/kg. In some embodiments, the multispecific antibody is administered at a dose of at least 1000 pg/kg, at least 1.1 mg/kg, at least 1.2 mg/kg, at least 1.3 mg/kg, at least 1.4 mg/kg, at least 1.5 mg/kg, at least 1.6 mg/kg, at least 1.7 mg/kg, at least 1.8 mg/kg, at least 1.9 mg/kg, or at least 2 mg/kg. In some embodiments, the multispecific antibody is administered at a dose of at least 2.5 mg/kg, at least 3 mg/kg, at least 3.5 mg/kg, at least 4 mg/kg, at least 4.5 mg/kg, at least 5 mg/kg, at least 5.5 mg/kg, at least 6 mg/kg, at least 6.5 mg/kg, at least 7 mg/kg, at least 7.5 mg/kg, at least 8 mg/kg, at least 8.5 mg/kg, at least 9 mg/kg, at least 9.5 mg/kg, or at least 10 mg/kg.

[117] In some embodiments, the multispecific antibody is administered once every 14 days at a dose of at least 1 pg/kg, at least 2 pg/kg, at least 3 pg/kg, at least 4 pg/kg, at least 5 pg/kg, at least 6 pg/kg, at least 7 pg/kg, at least 8 pg/kg, at least 9 pg/kg, at least 10 pg/kg, at least

I I pg/kg, at least 12 pg/kg, at least 13 pg/kg, at least 14 pg/kg, at least 15 pg/kg, at least 16 pg/kg, at least 17 pg/kg, at least 18 pg/kg, at least 19 pg/kg, at least 20 pg/kg, at least 25 pg/kg, at least 30 pg/kg, at least 35 pg/kg, at least 40 pg/kg, or at least 45 pg/kg. In some embodiments, the multispecific antibody is administered once every 14 days at a dose of at least 100 pg/kg, at least 110 pg/kg, at least 120 pg/kg, or at least 130 pg/kg. In some embodiments, the multispecific antibody is administered once every 14 days at a dose of at least 300 pg/kg, at least 310 pg/kg, at least 320 pg/kg, at least 330 pg/kg, at least 340 pg/kg, at least 350 pg/kg, at least 360 pg/kg, at least 370 pg/kg, at least 380 pg/kg, at least 390 pg/kg, or at least 400 pg/kg. In some embodiments, the multispecific antibody is administered once every 14 days at a dose of at least 500 pg/kg, at least 510 pg/kg, at least 520 pg/kg, at least 530 pg/kg, at least 540 pg/kg, at least 550 pg/kg, at least 560 pg/kg, at least 570 pg/kg, at least 580 pg/kg, at least 590 pg/kg, or at least 600 pg/kg. In some embodiments, the multispecific antibody is administered once every 14 days at a dose of at least 750 pg/kg, at least 760 pg/kg, at least 770 pg/kg, at least 780 pg/kg, at least 790 pg/kg, at least 800 pg/kg, at least 810 pg/kg, at least 820 pg/kg, at least 830 pg/kg, at least 840 pg/kg, or at least 850 pg/kg. In some embodiments, the multispecific antibody is administered once every 14 days at a dose of at least 1000 pg/kg, at least 1.1 mg/kg, at least 1.2 mg/kg, at least 1.3 mg/kg, at least 1.4 mg/kg, at least 1.5 mg/kg, at least 1.6 mg/kg, at least 1.7 mg/kg, at least 1.8 mg/kg, at least 1.9 mg/kg, or at least 2 mg/kg. In some embodiments, the multispecific antibody is administered once every 14 days at a dose of at least 2.5 mg/kg, at least 3 mg/kg, at least 3.5 mg/kg, at least 4 mg/kg, at least 4.5 mg/kg, at least 5 mg/kg, at least 5.5 mg/kg, at least 6 mg/kg, at least 6.5 mg/kg, at least 7 mg/kg, at least 7.5 mg/kg, at least 8 mg/kg, at least 8.5 mg/kg, at least 9 mg/kg, at least 9.5 mg/kg, or at least 10 mg/kg.

[118] In some embodiments, the multispecific antibody is administered twice every 14 days at a dose of at least 1 pg/kg, at least 2 pg/kg, at least 3 pg/kg, at least 4 pg/kg, at least 5 pg/kg, at least 6 pg/kg, at least 7 pg/kg, at least 8 pg/kg, at least 9 pg/kg, at least 10 pg/kg, at least 11 pg/kg, at least 12 pg/kg, at least 13 pg/kg, at least 14 pg/kg, at least 15 pg/kg, at least 16 pg/kg, at least 17 pg/kg, at least 18 pg/kg, at least 19 pg/kg, at least 20 pg/kg, at least 25 pg/kg, at least 30 pg/kg, at least 35 pg/kg, at least 40 pg/kg, or at least 45 pg/kg. In some embodiments, the multispecific antibody is administered twice every 14 days at a dose of at least 100 pg/kg, at least 110 pg/kg, at least 120 pg/kg, or at least 130 pg/kg. In some embodiments, the multispecific antibody is administered twice every 14 days at a dose of at least 300 pg/kg, at least 310 pg/kg, at least 320 pg/kg, at least 330 pg/kg, at least 340 pg/kg, at least 350 pg/kg, at least 360 pg/kg, at least 370 pg/kg, at least 380 pg/kg, at least 390 pg/kg, or at least 400 pg/kg. In some embodiments, the multispecific antibody is administered twice every 14 days at a dose of at least 500 pg/kg, at least 510 pg/kg, at least 520 pg/kg, at least 530 pg/kg, at least 540 pg/kg, at least 550 pg/kg, at least 560 pg/kg, at least 570 pg/kg, at least 580 pg/kg, at least 590 pg/kg, or at least 600 pg/kg. In some embodiments, the multispecific antibody is administered twice every 14 days at a dose of at least 750 pg/kg, at least 760 pg/kg, at least 770 pg/kg, at least 780 pg/kg, at least 790 pg/kg, at least 800 pg/kg, at least 810 pg/kg, at least 820 pg/kg, at least 830 pg/kg, at least 840 pg/kg, or at least 850 pg/kg. In some embodiments, the multispecific antibody is administered twice every 14 days at a dose of at least 1000 pg/kg, at least 1.1 mg/kg, at least 1.2 mg/kg, at least 1.3 mg/kg, at least 1.4 mg/kg, at least 1.5 mg/kg, at least 1.6 mg/kg, at least 1.7 mg/kg, at least 1.8 mg/kg, at least 1.9 mg/kg, or at least 2 mg/kg. In some embodiments, the multispecific antibody is administered twice every 14 days at a dose of at least 2.5 mg/kg, at least 3 mg/kg, at least 3.5 mg/kg, at least 4 mg/kg, at least 4.5 mg/kg, at least 5 mg/kg, at least 5.5 mg/kg, at least 6 mg/kg, at least 6.5 mg/kg, at least 7 mg/kg, at least 7.5 mg/kg, at least 8 mg/kg, at least 8.5 mg/kg, at least 9 mg/kg, at least 9.5 mg/kg, or at least 10 mg/kg.

Step-dosing

[119] Although multispecific antibodies (including bispecific T cell engagers) hold great promise for cancer therapy, their use can be limited by therapy-related toxicities, including dose-related cytokine response syndrome (CRS). Cytokine release syndrome (CRS) is a systemic inflammatory response that can be triggered by immunotherapeutic agents, such as T-cell-engaging bispecific antibodies. CRS events typically occur in the first treatment cycle with the greatest level of cytokine production often observed after the first dose of the antibody. For example, compounds in clinical development, such as AMG 160 and HPN424, aim to direct cytotoxic T cells to prostate tumors by targeting CD3 and PSMA simultaneously. Interim results of Phase 1 clinical trials of these molecules have shown first indications of efficacy (Tran 2020.; Bendell et al. 2020). In line with other CD3-based T cell engagers (Khadka et al. 2019), CRS was observed as most frequent adverse event (AE) for AMG 160 and HPN424, and prophylactic risk mitigations for CRS were implemented.

[120] In order to mitigate the risk of CRS, a step dosing approach can be used. Step dosing involves administering one or more priming doses of a multispecific antibody prior to the target dose. The priming dose is a lower dose of the multispecific antibody than the target dose. This allows the immune system to be primed in a stepwise manner in an effort to prevent an uncontrolled inflammatory response at the target dose (Ball et al, 2023).

[121] Accordingly, the term “priming dose” or as used herein refers to a dose of a multispecific antibody described herein that primes a subject for administration of a target dose of the multispecific antibody such that the target dose does not result in one or more therapy- related toxicities such as CRS. A “target dose” as used herein refers to the therapeutically effective dose of a multispecific antibody described herein or the dose of a multispecific antibody that is being evaluated for use as a therapeutically effective dose (e.g., in the dose escalation studies described herein).

[122] In some embodiments, the priming dose is 25% or less of the target dose. For example, if a target dose of a multispecific antibody is 100 mg, the priming dose would be 25 mg. In some embodiments, the priming dose is 25% or less, 20% or less, 15% or less, 10% or less, or 5% or less of the target dose. In some embodiments, the priming dose is 5%, 6%, 7%, 8%, 9%, 10%, 11 %, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% of the target dose. In some embodiments, the priming dose is 15% of the target dose. In some embodiments, the priming dose is 50% of the target dose.

[123] In some embodiments, the priming dose is 1 pg, 2 pg, 3 pg, 4 pg, 5 pg, 10 pg, 15 pg, 20 pg, 25 pg, 30 pg, 35 pg, 40 pg, 45 pg, 50 pg, 60 pg, 70 pg, 80 pg, 90 pg, 100 pg, 110 pg, 120 pg, 130 pg, 140 pg, 150 pg, 160 pg, 170 pg, 180 pg, 190 pg, 200 pg, 210 pg, 220 pg, 230 pg, 240 pg, 250 pg, 260 pg, 270 pg, 280 pg, 290 pg, 300 pg, 310 pg, 320 pg, 330 pg, 340 pg, 350 pg, 360 pg, 370 pg, 380 pg, 390 pg, 400 pg, 410 pg, 420 pg, 430 pg, 440 pg, 450 pg, 460 pg, 470 pg, 480 pg, 490 pg, 500 pg, 510 pg, 520 pg, 530 pg, 540 pg, 550 pg, 560 pg, 570 pg, 580 pg, 590 pg, or 600 pg.

[124] In some embodiments, the first priming dose is 120 pg, and the second priming dose is 360 pg.

[125] In some embodiments, the target dose of the multispecific antibody (e.g., LAVA-1207) is 120 pg and the priming dose is 18 pg. In some embodiments, the target dose of the multispecific antibody (e.g., LAVA-1207) is 540 pg and the priming dose is 81 pg. In some embodiments, the target dose of the multispecific antibody (e.g., LAVA-1207) is 800 pg and the priming dose is 120 pg. In some embodiments, the target dose of the multispecific antibody (e.g., LAVA-1207) is 800 pg and the priming doses are 120 pg and 360 pg. In some embodiments, the target dose of the multispecific antibody (e.g., LAVA-1207) is 1200 pg and the priming dose is 180 pg. In some embodiments, the target dose of the multispecific antibody (e.g., LAVA-1207) is 1200 pg and the priming doses are 120 pg and 360 pg. In some embodiments, the target dose of the multispecific antibody (e.g., LAVA-1207) is 1800 pg and the priming dose is 180 pg. In some embodiments, the target dose of the multispecific antibody (e.g., LAVA-1207) is 1800 pg and the priming doses are 120 pg and 360 pg. In some embodiments, the target dose of the multispecific antibody (e.g., LAVA-1207) is 1800 pg and the priming dose is 180 pg. In some embodiments, the target dose of the multispecific antibody (e.g., LAVA-1207) is 1800 pg and the priming doses are 120 pg and 360 pg. In some embodiments, the target dose of the multispecific antibody (e.g., LAVA-1207) is 3600 pg and the priming dose is 180 pg. In some embodiments, the target dose of the multispecific antibody (e.g., LAVA-1207) is 3600 pg and the priming doses are 120 pg and 360 pg.

[126] In some embodiments, there are two or more priming doses. In some embodiments, the two or more priming doses are administered 1 day apart. In some embodiments, the two or more priming doses are administered 2 days apart. In some embodiments, the two or more priming doses are administered 3 days apart. In some embodiments, the two or more priming doses are administered 4 days apart. In some embodiments, the two or more priming doses are administered 5 days apart. In some embodiments, the two or more priming doses are administered 6 days apart.

some embodiments, the two or more priming doses are administered 7 days apart. In some embodiments, the two or more priming doses are administered 8 days apart. In some embodiments, the two or more priming doses are administered 9 days apart. In some embodiments, the two or more priming doses are administered 10 days apart. In some embodiments, the two or more priming doses are administered 11 days apart. In some embodiments, the two or more priming doses are administered 12 days apart. In some embodiments, the two or more priming doses are administered 13 days apart. In some embodiments, the two or more priming doses are administered 14 days apart. In some embodiments, the first priming dose is administered on day 1 , and the second priming dose is administered on day 4. In some embodiments, the first priming does is administered on day 1 , and the second priming dose is administered on day 8.

[127] In some embodiments, the last priming dose is administered 1 day before the target dose. In some embodiments, the last priming dose is administered 2 days before the target dose. In some embodiments, the last priming dose is administered 3 days before the target dose. In some embodiments, the last priming dose is administered 4 days before the target dose. In some embodiments, the last priming dose is administered 5 days before the target dose. In some embodiments, the last priming dose is administered 6 days before the target dose. In some embodiments, the last priming dose is administered 7 days before the target dose. In some embodiments, the last priming dose is administered 8 days before the target dose. In some embodiments, the last priming dose is administered 9 days before the target dose. In some embodiments, the last priming dose is administered 10 days before the target dose. In some embodiments, the last priming dose is administered 11 days before the target dose. In some embodiments, the last priming dose is administered 12 days before the target dose. In some embodiments, the last priming dose is administered 13 days before the target dose. In some embodiments, the last priming dose is administered 14 days before the target dose.

[128] In some embodiments, the first priming dose is administered on day 1 , the second priming dose is administered on day 4, and the target dose is administered on day 8. In some embodiments, the first priming does is administered on day 1 , the second priming dose is administered on day 8, and the target dose is administered on day 15.

[129] In some embodiments, the multispecific antibody is LAVA- 1207 and the first treatment cycle comprises administration of one or more priming doses and one or more target doses.

In some embodiments, the first treatment cycle comprises administration of one priming dose and one or more target doses. In some embodiments, the first treatment cycle comprises administration of two priming doses and one or more target doses. In some embodiments, the first treatment cycle comprises administration of three priming doses and one or more target doses. In some embodiments, the first treatment cycle comprises administration of one priming dose and one target dose. In some embodiments, the first treatment cycle comprises administration of two priming doses and one target dose. In some embodiments, the first treatment cycle comprises administration of three priming doses and one target dose.

[130] In some embodiments, the multispecific antibody is LAVA-1207 and the second treatment cycle comprises administration of one or more priming doses and one or more target doses. In some embodiments, the second treatment cycle comprises administration of one priming dose and one or more target doses. In some embodiments, the second treatment cycle comprises administration of two priming doses and one or more target doses. In some embodiments, the second treatment cycle comprises administration of three priming doses and one or more target doses. In some embodiments, the second treatment cycle comprises administration of one priming dose and one target dose. In some embodiments, the second treatment cycle comprises administration of two priming doses and one target dose. In some embodiments, the second treatment cycle comprises administration of three priming doses and one target dose.

In some embodiments, the multispecific antibody is LAVA- 1207 and the first and second treatment cycles comprise administration of one or more priming doses and one or more target doses. In some embodiments, the first and second treatment cycles comprise administration of one priming dose and one or more target doses. In some embodiments, the first and second treatment cycles comprise administration of two priming doses and one or more target doses. In some embodiments, the first and second treatment cycles comprise administration of three priming doses and one or more target doses. In some embodiments, the first and second treatment cycles comprise administration of one priming dose and one target dose. In some embodiments, the first and second treatment cycles comprise administration of two priming doses and one target dose. In some embodiments, the first and second treatment cycles comprise administration of three priming doses and one target dose.

Treatment regimens

[131] TABLE 4 below provides exemplary administration regimens for the multispecific antibody, IL-2 or IL-15, and immune checkpoint inhibitor within a particular treatment cycle. One of skill in the art will understand that the following regimens can be combined. For example, Regimen A can be utilized for a first treatment cycle, followed by Regimen B in a second treatment cycle, followed by Regimen C in a third treatment cycle. In some embodiments, any of the Regimens described below may be repeated. For example, Regimen A can be utilized for a first, a second, a third, or more treatment cycles, followed by Regimen B for one or more treatment cycles, followed by Regimen C for one or more treatment cycles. [132] Further still, while TABLE 4 below refers to a “1^st agent”, “2^nd agent”, and “3^rd agent”, one of skill in the art will understand that each agent may be administered one or more times. For example, in Regimen A, the administration of the multispecific antibody (MSAb) as the 1^st agent may be administration of a single dose of a MSAb or multiple (e.g., 2, 3, 4, 5, 6, or more) doses of the MSAb over the course of one or more days, the administration of the cytokine as the 2^nd agent may be administration of a single dose of a cytokine or multiple (e.g., 2, 3, 4, 5, 6, or more) doses of the cytokine over the course of one or more days. Reference to “cytokine” in TABLE 4 is intended to refer to a common gamma chain cytokine (e.g., IL-2 or IL-15). Reference to a MSAb includes bispecific T cell engagers (bsTCE). Inclusion of two agents as either the first or second agent is intended to refer to the concurrent administration of the listed agents

TABLE 4: Exemplary administration regimens

MSAb = multispecific antibody ICI = immune checkpoint inhibitor

[133] The agents described herein may be administered over the course of one or more treatment cycles. Herein, a “treatment cycle” refers to a period of treatment (e.g., administration of a multispecific antibody, common gamma chain cytokine, or immune checkpoint inhibitor) optionally followed by a period of rest (e.g., wherein no therapeutic agents are administered). A full course of treatment may comprise one or more treatment cycles, e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, or more treatment cycle. A treatment cycle may be 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 or more days long.

[134] Thus, an exemplary course of treatment may comprise the following:

(a) administration of a multispecific antibody as an intravenous infusion on day 1 of a 14-day treatment cycle; and

(b) administration of a common gamma chain cytokine (e.g., IL-2 or IL- 15) once daily on day 2, 3, and 4 of the 14-day treatment cycle;

(c) wherein the full course of treatment comprises at least 4 {e.g., 4, 5, 6, 7, 8, 9, 10, or more) treatment cycles.

[135] In this exemplary embodiment, the dosing interval of the multispecific antibody over the full course of treatment is every 14 days.

[136] Exemplary treatment cycles and full courses of treatment for LAVA-1207 are provided in TABLE 5.

[137] Exemplary treatment regimens comprising a multispecific antibody that binds to human PSMA (e.g., LAVA-1207) are provided below in TABLE 5. Although not shown in TABLE 5, an immune checkpoint inhibitor (e.g., an anti-PD1 antibody) can be administered at a variety of points throughout the course of treatment. For example, an anti-PD1 antibody can be administered before, concurrent with, or after a dose of LAVA-1207. In some embodiments, an anti-PD1 antibody can be administered before, concurrent with, or after a dose of IL-2 and/or IL-15.

[138] In some embodiments, the treatment regimen for LAVA- 1207 comprises administration of one or more premedications. The premedications are typically administered prior to administration of a target dose of LAVA- 1207. In some embodiments, the premedications are administered with a priming dose of LAVA-1207. Predmedications can include one or more of an antipyretic (e.g., paracetamol or acetaminophen), an antihistamine (e.g._s diphenhydramine), and a corticosteroid (e.g., dexamethasone or methylprednisolone). In some embodiments, the premedications include each of paracetamol or acetaminophen, diphenhydramine, and dexamethasone or methylprednisolone. In some embodiments, the premedications are given in combination with hydration (e.g. 1000 mL or more of fluids).

[139] In some embodiments, methylprednisolone is administered at a dose of 1 mg, 2 mg, 3 mg, 4 mg, 5 mg, 6 mg, 7 mg, 8 mg, 9 mg, 10 mg, 15 mg, 20 mg, 25 mg, 30 mg, 35 mg, 40 mg, 45 mg, or 50 mg. In some embodiments, methylprednisolone is administered at a dose of 20 mg. In some embodiments, dexamethasone is administered at a dose of 0.5 mg, 1 mg, 2 mg, 3 mg, 4 mg, 5 mg, 6 mg, 7 mg, 8 mg, 9 mg, or 10 mg. In some embodiments, dexamethasone is administered at a dose of 4 mg. In some embodiments, acetaminophen is administered at a dose of 100 mg, 200 mg, 300 mg, 400 mg, 500 mg, 600 mg, 700 mg, 800 mg, 900 mg, 1000 mg, 1100 mg, 1200 mg, 1300 mg, 1400 mg, 1500 mg, 1600 mg, 1700 mg, 1800 mg, 1900 mg, or 2000 mg. In some embodiments, diphenhydramine is administered at a dose of 5 mg, 10 mg, 15 mg, 20 mg, 25 mg, 30 mg, 35 mg, 40 mg, 45 mg, 50 mg, 55 mg, 60 mg, 65 mg, 70 mg, 75 mg, 80 mg, 85 mg, 90 mg, 95 mg, 100 mg, 110 mg, 120 mg, 130 mg, 140 mg, or 150 mg. [140] In some embodiments, premedications are administered 4, 3, 2, or 1 hours before the first dose of LAVA-1207. One or more of the premedications may be administered after LAVA- 1207 administrations have begun based on physician opinion. In some embodiments, premedications are administered 4, 3, 2, or 1 hours before any of the LAVA-1207 doses (priming doses or target doses) shown in TABLE 5.

TABLE 5: Exemplary treatment regimens for LAVA-1207

PrD - Priming dose TD = Target dose

[141] A subject’s response to treatment with LAVA- 1207 can be monitored by subject sample collection prior to and during treatment with LAVA-1207. For example, blood samples are taken from a subject prior to the start of treatment with LAVA-1207 and additional blood samples are taken at various timepoints after treatment has been initiated. These blood samples can be analyzed for pharmacodynamic characteristics including LAVA-1207 binding to Vy9V62 T cells, Vy9V<52 T-cell activation status, general immune cell analysis (e.g., activation status and frequency of B cells, T cells, NK cells, and monocytes), cytokine levels, and anti-drug antibodies. In some embodiments, additional blood samples are taken for use in cell-free DNA analysis of tumor mutation burden, tumor load, and/or circulating tumor cells.

[142] In some embodiments, the methods of the present disclosure comprise administration of LAVA-1207 at a dose of 1.5 pg, 4.5 pg, 13.5 pg, 40 pg, 120 pg, 360 pg, 540 pg, 800 pg, 1.2 mg, 1.8 mg, 3.6 mg, 5.4 mg, 7.2 mg, 8.1 mg, 12 mg, 21 mg, 65 mg, or 200 mg once per 2 week treatment cycle for a total of 12 cycles.

[143] In some embodiments, the methods of the present disclosure comprise 12 2-week treatment cycles, wherein the first and second treatment cycle comprise administration of LAVA- 1207 at a dose of 1.5 pg, 4.5 pg, 13.5 pg, 40 pg, 120 pg, 360 pg, 540 pg, 800 pg, 1.2 mg, 1.8 mg, 3.6 mg, 5.4 mg, 7.2 mg, 8.1 mg, 12 mg, 21 mg, 65 mg, or 200 mg on the first day of the treatment cycle followed by 1 or 3 days of once daily doses of IL-2 at 1 MIU for a total of 12 cycles. Cycles 5-12 comprise administration of LAVA-1207 at a dose of 1.5 pg, 4.5 pg, 13.5 pg, 40 pg, 120 pg, 360 pg, 540 pg, 800 pg, 1.2 mg, 1.8 mg, 3.6 mg, 5.4 mg, 7.2 mg, 8.1 mg, 12 mg, 21 mg, 65 mg, or 200mg, on the first day of the treatment cycle.

[144] In some embodiments, the methods of the present disclosure comprise 12 2-week treatment cycles, wherein the first treatment cycle comprises administration of LAVA- 1207 at a priming dose of 120 pg on the first and second day of the treatment cycle, followed by administration of a target dose of 800 pg on the third day of the treatment cycle, followed by 1 or 3 days of once daily doses of IL-2 at 1 MIU. The second, third, and fourth treatment cycles comprise administration of LAVA-1207 at a target dose of 800 pg on the first day of the treatment cycle followed by 3 days of once daily doses of IL-2 at 1 MIU. Cycles 5-12 comprise administration of LAVA-1207 at a target dose of a target dose of 800 pg on the first day of the treatment cycle.

[145] In some embodiments, the methods of the present disclosure comprise 12 2-week treatment cycles, wherein the first treatment cycle comprises administration of LAVA- 1207 at a priming dose of 120 pg on the first day of the treatment cycle, and a priming dose of 360 pg on the second day of the treatment cycle, followed by administration of a target dose of 800 pg on the third day of the treatment cycle, followed by 1 or 3 days of once daily doses of IL-2 at 1 Mill. The second, third, and fourth treatment cycles comprise administration of LAVA-1207 at a target dose of 800 pg on the first day of the treatment cycle followed by 3 days of once daily doses of IL-2 at 1 MIU. Cycles 5-12 comprise administration of LAVA-1207 at a target dose of 800 pg on the first day of the treatment cycle.

[146] In some embodiments, the methods of the present disclosure comprise 12 2-week treatment cycles, wherein the first treatment cycle comprises administration of LAVA- 1207 at a priming dose on the first and second day of the treatment cycle, followed by administration of a target dose of 1200 pg on the third day of the treatment cycle, followed by 1 or 3 days of once daily doses of IL-2 at 1 MIU. The second, third, and fourth treatment cycles comprise administration of LAVA- 1207 at a target dose of 1200 pg on the first day of the treatment cycle followed by 3 days of once daily doses of IL-2 at 1 MIU. Cycles 5-12 comprise administration of LAVA-1207 at a target dose of 1200 pg on the first day of the treatment cycle.

[147] In some embodiments, the methods of the present disclosure comprise 12 2-week treatment cycles, wherein the first treatment cycle comprises administration of LAVA- 1207 at a priming dose of 120 pg on the first day of the treatment cycle, and a priming dose of 360 pg on the second day of the treatment cycle, followed by administration of a target dose of 1200 pg on the third day of the treatment cycle, followed by 1 or 3 days of once daily doses of IL-2 at 1 MIU. The second, third, and fourth treatment cycles comprise administration of LAVA-1207 at a target dose of 1200 pg on the first day of the treatment cycle followed by 3 days of once daily doses of IL-2 at 1 MIU. Cycles 5-12 comprise administration of LAVA-1207 at a target dose of 1200 pg on the first day of the treatment cycle.

[148] In some embodiments, the methods of the present disclosure comprise 12 2-week treatment cycles, wherein the first treatment cycle comprises administration of LAVA- 1207 at a priming dose on the first and second day of the treatment cycle, followed by administration of a target dose of 1800 pg on the third day of the treatment cycle, followed by 1 or 3 days of once daily doses of IL-2 at 1 MIU. The second, third, and fourth treatment cycles comprise administration of LAVA- 1207 at a target dose of 1800 pg on the first day of the treatment cycle followed by 3 days of once daily doses of IL-2 at 1 MIU. Cycles 5-12 comprise administration of LAVA-1207 at a target dose of a target dose of 1800 pg on the first day of the treatment cycle.

[149] In some embodiments, the methods of the present disclosure comprise 12 2-week treatment cycles, wherein the first treatment cycle comprises administration of LAVA- 1207 at a priming dose on the first and second day of the treatment cycle, followed by administration of a target dose of 3600 pg on the third day of the treatment cycle, followed by 1 or 3 days of once daily doses of IL-2 at 1 MIU. The second, third, and fourth treatment cycles comprise administration of LAVA- 1207 at a target dose of 3600 pg on the first day of the treatment cycle followed by 3 days of once daily doses of IL-2 at 1 MIU. Cycles 5-12 comprise administration of LAVA-1207 at a target dose of a target dose of 3600 pg on the first day of the treatment cycle.

[150] In some embodiments, the methods of the present disclosure comprise 12 2-week treatment cycles, wherein the first treatment cycle comprises administration of LAVA- 1207 at a priming dose on the first and second day of the treatment cycle, followed by administration of a target dose of 5400 pg on the third day of the treatment cycle, followed by 1 or 3 days of once daily doses of IL-2 at 1 MIU. The second, third, and fourth treatment cycles comprise administration of LAVA- 1207 at a target dose of 5400 pg on the first day of the treatment cycle followed by 3 days of once daily doses of IL-2 at 1 MIU. Cycles 5-12 comprise administration of LAVA-1207 at a target dose of a target dose of 5400 pg on the first day of the treatment cycle.

[151] In some embodiments, the methods of the present disclosure comprise 12 2-week treatment cycles, wherein the first treatment cycle comprises administration of LAVA- 1207 at a priming dose on the first and second day of the treatment cycle, followed by administration of a target dose of 7200 pg on the third day of the treatment cycle, followed by 1 or 3 days of once daily doses of IL-2 at 1 MIU. The second, third, and fourth treatment cycles comprise administration of LAVA- 1207 at a target dose of 7200 pg on the first day of the treatment cycle followed by 3 days of once daily doses of IL-2 at 1 MIU. Cycles 5-12 comprise administration of LAVA-1207 at a target dose of a target dose of 7200 pg on the first day of the treatment cycle.

[152] In some embodiments, the methods of the present disclosure comprise 12 2-week treatment cycles, wherein the first treatment cycle comprises administration of LAVA- 1207 at a priming dose on the first and second day of the treatment cycle, followed by administration of a target dose of 8100 pg on the third day of the treatment cycle, followed by 1 or 3 days of once daily doses of IL-2 at 1 MIU. The second, third, and fourth treatment cycles comprise administration of LAVA- 1207 at a target dose of 8100 pg on the first day of the treatment cycle followed by 3 days of once daily doses of IL-2 at 1 MIU. Cycles 5-12 comprise administration of LAVA-1207 at a target dose of a target dose of 8100 pg on the first day of the treatment cycle.

[153] In some embodiments, the methods of the present disclosure comprise 12 2-week treatment cycles, wherein the first treatment cycle comprises administration of LAVA- 1207 at a priming dose on the first and second day of the treatment cycle, followed by administration of a target dose of 12000 pg on the third day of the treatment cycle, followed by 1 or 3 days of once daily doses of IL-2 at 1 MIU. The second, third, and fourth treatment cycles comprise administration of LAVA-1207 at a target dose of 12000 pg on the first day of the treatment cycle followed by 3 days of once daily doses of IL-2 at 1 Mill. Cycles 5-12 comprise administration of LAVA-1207 at a target dose of a target dose of 12000 pg on the first day of the treatment cycle.

[154] In some embodiments, the methods of the present disclosure comprise 12 2-week treatment cycles, wherein the first treatment cycle comprises administration of LAVA- 1207 at a priming dose on the first and second day of the treatment cycle, followed by administration of a target dose of 65000 pg on the third day of the treatment cycle, followed by 1 or 3 days of once daily doses of IL-2 at 1 MIU. The second, third, and fourth treatment cycles comprise administration of LAVA-1207 at a target dose of 65000 pg on the first day of the treatment cycle followed by 3 days of once daily doses of IL-2 at 1 MIU. Cycles 5-12 comprise administration of LAVA-1207 at a target dose of a target dose of 65000 pg on the first day of the treatment cycle.

[155] In some embodiments, the methods of the present disclosure comprise 12 2-week treatment cycles, wherein the first treatment cycle comprises administration of LAVA- 1207 at a priming dose on the first and second day of the treatment cycle, followed by administration of a target dose of 200000 pg on the third day of the treatment cycle, followed by 1 or 3 days of once daily doses of IL-2 at 1 MIU. The second, third, and fourth treatment cycles comprise administration of LAVA-1207 at a target dose of 200000 pg on the first day of the treatment cycle followed by 3 days of once daily doses of IL-2 at 1 MIU. Cycles 5-12 comprise administration of LAVA-1207 at a target dose of a target dose of 200000 pg on the first day of the treatment cycle.

[156] In some embodiments, the methods of the present disclosure comprise at least 4 treatment cycles, wherein the first cycle is 4 weeks and the second - fourth cycles are two weeks each. In some such embodiments, the first cycle comprises administration of a first priming dose of 120 pg LAVA-1207 on day 1 and administration of a target dose of 800 pg LAVA-1207 on day 15, wherein the target dose is administered on day 1 of each of cycles 2-4. In some such embodiments, the first cycle comprises administration of a first priming dose of 120 pg LAVA- 1207 on day 1 , a second priming dose of 360 pg LAVA-1207 on day 8, and a target dose of 800 pg LAVA- 1207 on day 15, wherein the target dose is administered on day 1 of each of cycles 2- 4. In some such embodiments, the first cycle comprises administration of a first priming dose of 120 pg LAVA-1207 on day 1 , a second priming dose of 360 pg LAVA-1207 on day 8, and a target dose of 1200 pg LAVA- 1207 on day 15, wherein the target dose is administered on day 1 of each of cycles 2-4.

[157] In some such embodiments, the first cycle comprises administration of a first priming dose of 120 pg LAVA-1207 on day 1 , a second priming dose of 360 pg LAVA-1207 on day 8, and a target dose of 800 pg LAVA-1207 on day 15, followed by 1 MIU of IL-2 on day 16, wherein the target dose is administered on day 1 of each of cycles 2-4, followed by 1 MIU of IL-2 on day 2 of each of cycles 2-4. In some such embodiments, the first cycle comprises administration of a first priming dose of 120 pg LAVA-1207 on day 1, a second priming dose of 360 pg LAVA-1207 on day 8, and a target dose of 800 pg LAVA-1207 on day 15, followed by 1 MIU of IL-2 on day 16, 17, and 18, wherein the target dose is administered on day 1 of each of cycles 2-4, followed by 1 MIU of IL-2 on day 2, 3, and 4 of each of cycles 2-4.

[158] In some such embodiments, the first cycle comprises administration of a first priming dose of 120 pg LAVA-1207 on day 1 , a second priming dose of 360 pg LAVA-1207 on day 8, and a target dose of 1200 pg LAVA-1207 on day 15, followed by 1 MIU of IL-2 on day 16, wherein the target dose is administered on day 1 of each of cycles 2-4, followed by 1 MIU of IL-2 on day 2 of each of cycles 2-4. In some such embodiments, the first cycle comprises administration of a first priming dose of 120 pg LAVA-1207 on day 1 , a second priming dose of 360 pg LAVA-1207 on day 8, and a target dose of 1200 pg LAVA-1207 on day 15, followed by 1 MIU of IL-2 on day 16, 17, and 18, wherein the target dose is administered on day 1 of each of cycles 2-4, followed by 1 MIU of IL-2 on day 2, 3, and 4 of each of cycles 2-4.

Indications and Methods of Treatment

[159] In some embodiments, the present disclosure provides a method of treating cancer in a subject in need thereof comprising administering a multispecific binding agent descried herein or a composition comprising the same and (i) a common gamma chain cytokine (e.g., IL-2 or IL-15) and/or (ii) an immune checkpoint inhibitor (e.g., an anti-PD1 or anti-PDL1 antibody).

[160] In some embodiments, treating refers to the treatment of a cancer in a mammal, e.g., in a human, including (a) inhibiting the cancer, i.e., arresting cancer development or preventing cancer progression; (b) relieving the cancer, i.e., causing regression of the cancer state or relieving one or more symptoms of the cancer; and (c) curing the cancer, i.e., remission of one or more cancer symptoms. In some embodiments, treatment may refer to a short-term (e.g., temporary and/or acute) and/or a long-term (e.g., sustained) reduction in one or more cancer symptoms. In some embodiments, treatment results in an improvement or remediation of the symptoms of the cancer. The improvement is an observable or measurable improvement or may be an improvement in the general feeling of well-being of the subject.

[161] In some embodiments, the subject may be a neonate, a juvenile, or an adult. Of particular interest are mammalian subjects. Mammalian species that may be treated with the present methods include canines and felines; equines; bovines; ovines; etc. and primates, particularly humans. Animal models, particularly small mammals (e.g. mice, rats, guinea pigs, hamsters, rabbits, etc.) may be used for experimental investigations. In some embodiments, the subject is a human.

[162] In some embodiments, the cancer is one in which the target antigen is expressed on the tumor neo-vasculature or tumor-associated endothelial cells of primary or metastatic tumors, including colorectal cancer, lung cancer, non-small cell lung cancer, endometrial and ovarian cancer, uterine cancer, uterine corpus endometrial carcinoma, gastric cancer, urothelial carcinomas, hepatocellular carcinoma, oral squamous cancer, thyroid tumors and glioblastomas, or adenoid cystic carcinoma of the head and neck, head and neck squamous cell carcinoma, prostate cancer, non-metastatic prostate cancer, metastatic prostate cancer, therapy refractory metastatic castration resistant prostate cancer, glioblastoma multiform, or blastic plasmacytoid dendritic neoplasm.

[163] In some embodiments, the cancer is a hematological malignancy such as T cell lymphoma, multiple myeloma, acute myeloid leukemia, chronic myeloid leukemia, acute lymphoblastic leukemia, chronic lymphocytic leukemia, mantle cell lymphoma, myelodysplastic syndrome pre-B acute lymphoblastic leukemia, B cell lymphoma, smoldering myeloma, myelomonocytic leukemias, lymphoplasmacytic lymphoma, hairy cell leukemia, splenic marginal zone lymphoma, non-Hodgkin's lymphoma, Hodgkin's lymphoma, follicular lymphoma, pancreatic cancer, colon cancer, B-cell acute lymphoblastic leukemia, B-cell lymphoma/leukemia, B-cell chronic lymphoproliferative disorders, Burkitt lymphoma or B acute lymphoblastic leukemia.

[164] In some embodiments, the cancer is a solid tumor cancer, such as renal cell carcinoma, melanoma, breast cancer (including triple-negative breast cancer), gastro-esophageal cancer, small bowel carcinoma, central nervous system tumors, medulloblastomas, hepatocellular carcinoma, glioma, neuroblastoma, bladder cancer, sarcoma, penile cancer, basal cell carcinoma, merkel cell carcinoma, neuroendocrine carcinoma, neuroendocrine tumors, carcinoma of unknown primary (CUP), thymoma, vulvar cancer, cervical carcinoma, testicular cancer, cholangiocarcinoma, appendicular carcinoma, mesothelioma, ampullary carcinoma, anal cancer, or choriocarcinoma.

[165] Administration of the agents described herein (e.g., a multispecific antibody, common gamma chain cytokine, and/or immune checkpoint inhibitor, either alone or in combination) can occur by injection, irrigation, inhalation, consumption, electro-osmosis, hemodialysis, iontophoresis, and other methods known in the art. In some embodiments, administration route is local or systemic. In some embodiments administration route is intraarterial, intracranial, intradermal, intraduodenal, intrammamary, intrameningeal, intraperitoneal, intrathecal, intramuscular, intratumoral, intravenous, intravitreal, ophthalmic, parenteral, spinal, subcutaneous, ureteral, urethral, vaginal, or intrauterine.

[166] In some embodiments, the administration route is by infusion (e.g., continuous or bolus). Examples of methods for local administration, that is, delivery to the site of injury or disease, include through an Ommaya reservoir, e.g. for intrathecal delivery (See e.g., US Patent Nos. 5,222,982 and 5,385,582, incorporated herein by reference); by bolus injection, e.g. by a syringe, e.g. into a joint; by continuous infusion, e.g. by cannulation, such as with convection (See e.g., US Patent Application Publication No. 2007-0254842, incorporated herein by reference); or by implanting a device upon which the agents have been reversibly affixed (see e.g. US Patent Application Publication Nos. 2008-0081064 and 2009-0196903, incorporated herein by reference). In some embodiments, the administration route is by topical administration or direct injection.

[167] In some embodiments, subjects are selected for treatment according to the methods described herein by one or more exclusion and/or inclusion criteria. Inclusion/exclusion criteria may include: age, sex, infection status (e.g., hepatis B or C), prior treatment regimens (e.g., immunosuppressants or aminobisphosphonates), presence of malignancies other than the cancer intended for treatment, and general health status.

[168] In some embodiments, subjects are selected for treatment with LAVA-1207 by one or more of the following inclusion criteria:

(a) males with metastatic castration-resistant prostate cancer (mCRPC) as defined by PCWG3 criteria (histologically confirmed adenocarcinoma; adenocarcinoma with <10% small-cell or neuroendocrine features is allowed but brain metastasis are allowable as long as the subject’s symptoms are well controlled);

(b) failure on at least one line of taxane-based chemotherapy or deemed medically unsuitable to be treated with a taxane regimen;

(c) receipt of a 2^nd generation or later androgen receptor targeted therapy/androgen biosynthesis inhibitor (e.g. abiraterone, enzalutamide, and/or apalutamide) or deemed unsuitable for such treatment;

(d) evidence of a progressive disease as defined by one or more of a PSA level > 1 ng/mL that has increased on at least 2 successive occasions at least 1 week apart;

(e) computed tomography (CT) or magnetic resonance imaging (MRI) scan showing nodal or visceral progression as defined by RECIST 1.1 ; and/or

(f) bone scintigraphy showing 2 or more new metastatic lesions. [169] In some embodiments, subjects are excluded from treat if any one or more of the following criteria apply:

(a) diagnosis of other malignancies within the last 2 years except adequately treated carcinoma in situ, basal or squamous cell skin carcinoma;

(b) an uncontrolled or severe intercurrent medical condition

(c) positive serological test for human immunodeficiency virus (HIV) antibodies;

(d) positive serological test for hepatitis B surface antigen (HBsAg). Subjects who are positive for anti-HBc or hepatitis C antibody may be included if they have a negative polymerase chain reaction (PCR) within 6 weeks prior to initial multispecific antibody administration. A subject who is PCR positive will be excluded.

(e) active-, uncontrolled-, or suspected infection;

(f) known clinically relevant immunodeficiency disorders;

(g) significant history of renal, neurologic, psychiatric, pulmonary, endocrinologic, metabolic, immunologic, cardiovascular, or hepatic disease that in the opinion of the investigator would adversely affect participation in this trial. Unstable cardiovascular function is defined as: (a) symptomatic ischemia, or (b) uncontrolled clinically significant conduction abnormalities (i.e. ventricular tachycardia on antiarrhythmic agents are excluded; 1st degree atrioventricular block or asymptomatic left anterior fascicular block/right bundle branch block is not excluded), or (c) congestive heart failure New York Heart Association Class > 3, or (d) myocardial infarction within 3 months;

(h) previous treatment with antitumor therapies 2 weeks prior to initial multispecific antibody for radiotherapy and androgen receptor targeted therapy/ androgen biosynthesis inhibitor, and 4 weeks for systemic chemotherapy or targeted-/ immunotherapy;

(i) previous treatment with live or live attenuated vaccines within 2 weeks prior to initial multispecific antibody administration (new types of vaccines need to be evaluated as to their mode of action);

(j) treatment with other investigational agents in the 4 weeks prior to initial multispecific antibody;

(k) major surgery within 4 weeks prior to initial multispecific antibody administration;

(l) hypersensitivity to any of the excipients present in LAVA-1207, IL-2, IL-15, or pembrolizumab (if applicable); (m) previous treatment with any systemic immunosuppressant within 4 weeks prior to initial multispecific antibody administration, with the exception of systemic corticosteroid use up to oral dose of 10 mg prednisone daily (or equivalent for other steroids);

(n) previous treatment with an aminobisphosphonate IV (e.g. ibandronate, pamidronate, zoledronate etc) within 4 weeks or 1 year prior to initial IMP; and/or

(o) known ongoing drug and alcohol abuse in the opinion of the investigator.

Multispecific Antibodies

[170] Multispecific antibodies suitable for use according to the methods described herein are described below. In some embodiments, the multispecific antibody comprises a first antigenbinding domain capable of binding a human cancer antigen and a second antigen-binding domain capable of binding a human Vy9V<52 T cell receptor by binding either the Vy9 or V02 chain (e.g., a bispecific antibody). In some embodiments, the multispecific antibody comprises a first antigenbinding domain that capable of binding a human cancer antigen, a second antigen-binding domain capable of binding a human Vy9V52 T cell receptor, and a third antigen-binding domain capable of binding to a third target antigen or epitope. In some embodiments, the multispecific antibody further comprises a fourth, fifth, or sixth antigen-binding domain.

[171] In some embodiments, any of the first, second, third, fourth, fifth, or sixth antigen-binding domains of the multispecific, antibody may be a single-domain antibody, such as a VHH. In some embodiments, the first and second antigen binding domains of the multispecific antibody are VHHs.

[172] Antibodies may be formulated with pharmaceutically-acceptable excipients in accordance with conventional techniques such as those disclosed in Rowe et al. 2012 Handbook of Pharmaceutical Excipients, ISBN 9780857110275). The pharmaceutically-acceptable excipient as well as any other carriers, diluents or adjuvants should be suitable for the antibodies and the chosen mode of administration. Suitability for excipients and other components of pharmaceutical compositions is determined based on the lack of significant negative impact on the desired biological properties of the chosen antibody or pharmaceutical composition of the present disclosure (e.g., less than a substantial impact (10% or less relative inhibition, 5% or less relative inhibition, etc.) upon antigen binding). A pharmaceutical composition may include diluents, fillers, salts, buffers, detergents (e.g., a nonionic detergent, such as Tween-20 or Tween-80), stabilizers (e.g., sugars or protein-free amino acids), preservatives, tissue fixatives, solubilizers, and/or other materials suitable for inclusion in a pharmaceutical composition. Further pharmaceutically- acceptable excipients include any and all suitable solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonicity agents, antioxidants and absorption-delaying agents and the like that are physiologically compatible with an antibody of the present disclosure.

[173] Suitable formulations for multispecific antibodies for use in the present disclosure have been described in WO 2022/008646. In some embodiments, the multispecific antibody may be formulated at a strength of about 1 mg/mL. In some embodiments, the formulation includes 1-20 mM histidine. In some embodiments, the formulation includes 0.1-10 mM methionine. In some embodiments, the formulation includes 50-500 mM sucrose. In some embodiments, the formation includes 0.01-0.05% polysorbate 80. In some embodiments, the formulation is at a pH 5.0-7.5. In some embodiments, the multispecific antibody is formulated at a strength of 1 mg/mL. In some embodiments, the formulation includes 10 mM histidine. In some embodiments, the formulation includes 1 mM methionine. In some embodiments, the formulation includes 280 mM sucrose. In some embodiments, the formulation includes 0.02 % polysorbate 80. In some embodiments, the formulation is at pH 6.0. For example, the multispecific antibody may be formulated at a strength of 1 mg/mL in 10 mM histidine, 1 mM methionine, 280 mM sucrose, 0.02 % polysorbate 80, pH 6.0. The multispecific antibody may be administered via any suitable administration route, for example intravenously or subcutaneously.

[174] Multispecific antibodies for use in the present disclosure are typically produced recombinantly, i.e. by expression of nucleic acid constructs encoding the antibodies in suitable host cells, followed by purification of the produced recombinant antibody from the cell culture. Nucleic acid constructs can be produced by standard molecular biological techniques well-known in the art. The constructs are typically introduced into the host cell using an expression vector. Suitable nucleic acid constructs and expression vectors are known in the art. Host cells suitable for the recombinant expression of antibodies are well-known in the art, and include CHO, H EK- 293, Expi293F, PER-C6, NS/0 and Sp2/0 cells.

[175] In some embodiments, any of the multispecific antibodies of this disclosure are modified to reduce or eliminate the formation of pyroglutamic acid at the N-terminus of the modified multispecific antibodies as compared to the unmodified multispecific antibodies of this disclosure. In some embodiments, the glutamate residue (E) or glutamine (Q) residue is modified to an aspartic acid residue (D). In some embodiments, any of the multispecific antibodies of the present disclosure may comprise an aspartic acid (D) residue in the first position.

[176] In some embodiments, the first antigen-binding region and second antigen-binding region of the multispecific antibody used in the present disclosure may be covalently linked via a peptide linker, e.g. a linker having a length of from 1 to 20 amino acids, e.g. from 1 to 10 amino acids, such as 2, 3, 4, 5, 6, 7, 8 or 10 amino acids. The peptide linker may comprise or consist of four glycine residues followed by a serine residue. The first antigen-binding region may be located N- terminally or C-terminally of the second antigen-binding region. In some embodiments, the first and second antigen-binding regions may be linked or directly fused to Fc domain monomers.

[177] The multispecific or bispecific antibody used in the present disclosure is capable of binding a human Vy9V52 T cell receptor. In some embodiments, the multispecific antibody is capable of binding to human V52. The term “human V02”, when used herein, refers to the rearranged 52 chain of the Vy9V62-T cell receptor (TOR). UniProtKB - A0JD36 (A0JD36_HUMAN) gives an example of a variable TRDV2 sequence. Antigen-binding domains that specifically bind to V52+ T cell receptors are known in the art. See e.g., WO 2015/156673, WO 2022/008646, WO 2022/122973, and WO 2023/242319, each disclosing antigen binding domains that specifically bind to V52 and each incorporated herein by reference. Exemplary V52 antigen-binding domain sequences are shown in TABLE 6A, TABLE 6B, TABLE 6C, and TABLE 6D.

Table 6A: Exemplary V52 TCR-specific CDR (Kabat) and VHH Sequences

Table 6B: Exemplary V52 TCR-specific CDR Sequences (IMGT)

Table 6C: Exemplary V52 TCR-specific CDR Sequences (Chothia)

Table 6D: Exemplary V52 TCR-specific CDR Sequences (Combined)

[178] One of skill in the art will recognize that there are many numbering systems known in the art to define the CDR sequences of an antibody, including AbM, Kabat, Chothia, and IMGT systems. An overview of these systems is provided in Dondelinger et al., Understanding the Significance and Implications of Antibody Numbering and Antigen-Binding Surface/Residue Definition. Front Immunol. 2018 Oct 16;9:2278. CDR sequences of V62 antigen-binding domains as defined by Kabat, IMGT, and Chothia are provided in TABLE 6A, TABLE 6B, and TABLE 6C above. The description of the binding agents herein refers to the CDRs as defined by the Kabat numbering system. However, one of skill in the art is able to correlate a CDR sequence defined by one numbering system to the same CDR sequence as defined by another system.

[179] In some embodiments, a combination of CDR numbering systems may be used. In such embodiments, CDR sequences of a given binding agent as determined by multiple numbering systems (e.g., a CDR1 sequence as determined by the Kabat, IMGT, and Chothia numbering systems) are compiled into a single sequence that encompasses the entirety of each of the CDR amino acid ranges in the variable region. As an illustrative example, CDRs for the 6H4 VHH described in Tables 6A, 6B, and 6C are shown in FIG. 20. The N- and C-terminal ranges for each CDR region are indicated by dashed lines. The combined CDR sequences covering the entirety of these ranges are shown in the last row. Illustrative combined CDRs for V52 antigen-binding domains are shown in Table 6D.

[180] As an illustrative example, a 6H4 VHH comprising a CDR1 amino acid sequence comprising SEQ ID NO: 26, a CDR2 amino acid sequence comprising SEQ ID NO: 27, and a CDR3 amino acid sequence comprising SEQ ID NO: 28 as defined by the Kabat numbering system will be understood by one of skill in the art to be the functional equivalent of (1) a 6H4 VHH comprising a CDR1 amino acid sequence comprising SEQ ID NO: 45, a CDR2 amino acid sequence comprising SEQ ID NO: 46, and a CDR3 amino acid sequence comprising SEQ ID NO: 47 as defined by the IMGT numbering system; (2) a 6H4 VHH comprising a CDR1 amino acid sequence comprising SEQ ID NO: 63, a CDR2 amino acid sequence comprising SEQ ID NO: 64, and a CDR3 amino acid sequence comprising SEQ ID NO: 65 as defined by the Chothia numbering system; and (3) a 6H4 VHH comprising a CDR1 amino acid sequence comprising SEQ ID NO: 84, a CDR2 amino acid sequence comprising SEQ ID NO: 85, and a CDR3 amino acid sequence comprising SEQ ID NO: 86.

[181] In some embodiments, the multispecific antibody is capable of binding to human Vy9. The term “human Vy9”, when used herein, refers to the rearranged Vy9 chain of the Vy9V62-T cell receptor (TOR). UniProtKB - Q99603 (TRGV9_HUMAN) gives an example of a variable Vy9 sequence. Exemplary Vy9V<52 TCR antigen-binding domain sequences are known in the art, see e.g. WO 2015/156673.

[182] In some embodiments, the (second) antigen-binding domain capable of binding the human Vy9V<52 T cell receptor may comprise the VH CDR1 sequence of SEQ ID NO:5, the VH CDR2 sequence of SEQ ID NO: 6 and the VH CDR3 sequence of SEQ ID NO: 7. In some embodiments, the second antigen-binding region may comprise an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 8. In some embodiments, the second antigen-binding region may comprise an amino acid sequence having at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 9. In some embodiments, the second antigen-binding domain comprises or consists of SEQ ID NO: 8. In some embodiments, the second antigen-binding domain comprises or consists of SEQ ID NO: 9.

[183] In some embodiments, the (second) antigen-binding domain capable of binding the human Vy9V52 T cell receptor may comprise the VH CDR1 sequence of SEQ ID NO: 10, the VH CDR2 sequence of SEQ ID NO: 11 and the VH CDR3 sequence of SEQ ID NO: 12. In some embodiments, the second antigen-binding region may comprise an amino acid sequence having at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 13. In some embodiments, the second antigen-binding domain comprises or consists of SEQ ID NO: 13. In some embodiments, the (second) antigen-binding domain capable of binding the human Vy9V52 T cell receptor may comprise the VH CDR1 sequence of SEQ ID NO: 14, the VH CDR2 sequence of SEQ ID NO: 15 and the VH CDR3 sequence of SEQ ID NO: 16. In some embodiments, the second antigen-binding region may comprise an amino acid sequence having at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 17. In some embodiments, the second antigen-binding domain comprises or consists of SEQ ID NO: 17.

[184] In some embodiments, the (second) antigen-binding domain capable of binding the human Vy9V<52 T cell receptor may comprise the VH CDR1 sequence of SEQ ID NO: 18, the VH CDR2 sequence of SEQ ID NO: 19 and the VH CDR3 sequence of SEQ ID NO: 20. In some embodiments, the second antigen-binding region may comprise an amino acid sequence having at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 21. In some embodiments, the second antigen-binding domain comprises or consists of SEQ ID NO: 21.

[185] In some embodiments, the (second) antigen-binding domain capable of binding the human Vy9V<52 T cell receptor may comprise the VH CDR1 sequence of SEQ ID NO: 22, the VH CDR2 sequence of SEQ ID NO: 23 and the VH CDR3 sequence of SEQ ID NO: 24. In some embodiments, the second antigen-binding region may comprise an amino acid sequence having at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 25. In some embodiments, the second antigen-binding domain comprises or consists of SEQ ID NO: 25. [186] In some embodiments, the multispecific antibody used in the method of the present disclosure competes (i.e. is able to compete) for binding to V<52 with an antibody having a sequence selected from SEQ ID NOs: 4, 8, 9, 13, 17, 21 , 25, and 29. The multispecific antibody used in the method of the present disclosure may bind to the same epitope on human V52 as an antibody having a sequence selected from SEQ ID NOs: 4, 8, 9, 13, 17, 21 , 25, and 29.

[187] The multispecific or bispecific antibody used in the present disclosure is capable of binding a human cancer antigen. The human cancer antigen may for example be an antigen associated with solid tumors or an antigen associated with hematological cancer disease. Human cancer antigens may be exclusively expressed on malignant cells or overexpressed on malignant cells relative to healthy cells.

[188] In some embodiments, the human cancer antigen is selected from HER2/neu, EGFR, CA- 125, PSA, CD44, EpCAM, CEACAM5, GD2, CD1d, CD40, CD123, 5T4, Nectin-4, PSMA, B7-H3, RAGE, CA72-4, HE4, Glypican-3, PSMA, MUC1 , CD133, EGFRvlll, CAIX, CD24, CD166, CD47, CD70, CD133, CD276 (B7-H3) , CD271 , CD146, CD164, CD123, CD38, CD166, CD24, CD29, CD49f, CD56, CD20, CD71 , CD98, CD99, CD147, CD166, CD200, CD184 (CXCR4), CD44v6, CD271 , CD276, CD304 (BDCA-4), CD326 (EpCAM), and CD338 (ABCB5). In some embodiments, the human cancer antigen is selected from the group consisting of PSMA, CD1d, CD40, CD123, 5T4, and Nectin-4. In some embodiments, the human cancer antigen is selected from the group consisting of PSMA, CD1d, CD40, CD123, 5T4, EGFR, and Nectin-4. In some embodiments, the human cancer antigen is selected from PSMA, CD1d, CD40, CD123, 5T4, EGFR, CD33, and Nectin-4. Exemplary cancer antigen-specific CDR and VHH sequences are provided in TABLE 7A, TABLE 7B, TABLE 7C, and TABLE 7D below.

Table 7 A: Exemplary cancer antigen-specific CDR (Kabat) and VHH Sequences

Table 7B: Exemplary cancer antigen-specific CDR Sequences (Chothia)

Table 7C: Exemplary cancer antigen-specific CDR Sequences (IMGT)

Table 7D: Exemplary cancer antigen-specific CDR Sequences (Combined)

PSMA Multispecific Antibodies

[189] In some embodiments, the human cancer antigen may be PSMA. Multispecific antibodies having antigen-binding regions capable of binding a human PSMA suitable for use in the present disclosure have for example been described in WO 2022/008646 (herein incorporated by reference).

[190] In some embodiments, the (first) antigen-binding region of the multispecific antibody used in the present disclosure may comprise the VH CDR1 sequence of SEQ ID NO: 87, the VH CDR2 sequence of SEQ ID NO: 88 and the VH CDR3 sequence of SEQ ID NO: 89. In some embodiments, the first antigen-binding region may comprise an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 90. In some embodiments, the first antigen-binding region comprises or consists of SEQ ID NO: 90.

[191] The multispecific antibody used in the method of the present disclosure may compete (i.e. be able to compete) for binding to PSMA with an antibody having the sequence set forth in SEQ ID NO:90. The multispecific antibody may bind the same epitope on human PSMA as an antibody having the sequence set forth in SEQ ID NO:90.

[192] In some embodiments, the multispecific antibody used in the present disclosure may comprise a first antigen-binding region and a second antigen-binding region, wherein the first antigen-binding region comprises the CDR1 sequence set forth in SEQ ID NO: 87 , the CDR2 sequence set forth in SEQ ID NO:88 and the CDR3 sequence set forth in SEQ ID NO:89 and wherein the second antigen-binding region comprises the CDR1 sequence set forth in SEQ ID NO:5, the CDR2 sequence set forth in SEQ ID NO:6 and the CDR3 sequence set forth in SEQ ID NO:7. In some embodiments, the multispecific antibody used in the present disclosure may comprise a first antigen-binding region and a second antigen-binding region, wherein the first antigen-binding region comprises the sequence set forth in SEQ ID NO:90 and the second antigen-binding region comprises the sequence set forth in SEQ ID NO:8. In some embodiments, the second antigen-binding domain comprises any of the CDR combinations recited in Tables 6A-6D.

[193] The multispecific antibody may comprise or consist of the sequences set forth in SEQ ID NO:334 and SEQ ID NO:335. Such a multispecific antibody is referred to herein as LAVA-1207 or PSMA-V62-Fc. Sequences for LAVA-1207 are provided below in TABLE 8. Antigen binding domains are shown in underlined text; hinge regions are in bold an italicized text; DEL sequence is shown in boxed text; Fc domains are in regular text with mutations relative to WT I gG 1 in bold and underlined text.

TABLE 8: LAVA-1207 Sequences

[194] The multispecific antibody of the present disclosure may be capable of mediating killing of PSMA-expressing cells, e.g. LNCaP cells, 22Rv1 cells or VCaP cells through activation of Vy9V32 T cells. For example, the multispecific antibody may be capable of inducing killing of LNCaP cells through activation of Vy9V62 T cells with an EC50 value of 50 pM or less, such as 25 pM or less, e.g. 20 pM or less, such as 15 pM or less, e.g. 10 pM or less, or even 5 pM or less, such as 2 pM or less or 1 pM or less when tested as described in Example 6 of WO 2022/008646. [195] The multispecific antibody may be capable of inducing killing of LNCaP, 22Rv1 or VCaP cells through activation of Vy9V52 T cells with an EC50 value of 50 pM or less, such as 25 pM or less, e.g. 20 pM or less, such as 15 pM or less when tested after 24 hours as described in Example 13 of WO 2022/008646, preferably both at a 1 :1 and a 1 :10 effector to target cell ratio.

[196] The multispecific antibody of the present disclosure may be capable of binding to the PSMA positive prostate cancer cell line LNCaP with an EC50 of 50 nM or less, such as 20 nM or less, e.g. 10 nM of less, when tested as described in Example 7 of WO 2022/008646. The multispecific antibody of the present disclosure may further or alternatively be capable of binding to Vy9V52 T cells with an EC50 of 10 nM or less, such as 5 nM or less, e.g. 2 nM of less, when tested as described in Example 7 of WO 2022/008646.

[197] The multispecific antibody may be capable of binding to recombinant human PSMA protein with a KD value of 100 nM or less, such as 50 nM or less, when tested as described in Example 11 of WO 2022/008646. The multispecific antibody of the present disclosure may further or alternatively be capable of binding to human Vy9V52-Fcwith a KD value of 10 nM or less, such as 5 nM or less, e.g. 2 nM or less, such as 1 nM or less when tested as described in Example 11 of WO 2022/008646.

[198] The multispecific antibody may be capable of mediating killing of human PSMA- expressing cells from a prostate cancer patient. Killing of human PSMA-expressing cells from a prostate cancer patient may e.g. be determined as described in Example 10 of WO 2022/008646. The multispecific antibody of the present disclosure may further or alternatively be capable of mediating specific cell death of more than 25%, such as more than 50%, at a concentration of 50 nM, as determined in the assays described in Example 10 or Example 14 of WO 2022/008646.

[199] The multispecific antibody may not be capable of mediating killing of PSMA-negative cells, such as PSMA negative human cells. The multispecific antibody may not induce IL-2, IL-4, IL-6, IL- 10 or TN Fa in whole blood from healthy donors at concentrations up to 280 nM, when tested as described in Example 16 of WO 2022/008646. The multispecific antibody may alternatively or in addition induce more than 10-fold less IL-8 and/or more than 50-fold less IFNy than Campath® in whole blood from healthy donors when tested as described in Example 16 of WO 2022/008646.

[200] The multispecific antibody used in the present disclosure may comprise a first antigenbinding region capable of binding PSMA and be for use in the treatment of prostate cancer, such as non-metastatic or metastatic prostate cancer, for example metastatic castration resistant prostate cancer, such as therapy refractory metastatic castration resistant prostate cancer. For example, the multispecific antibody may comprise a first antigen-binding region capable of binding PSMA and be used for the treatment of prostate cancer in patients that have previously received at least one or more treatments for prostate cancer. In some embodiments, the one or more treatments are selected from taxane-based chemotherapy, androgen receptor targeted therapy treatment, with an androgen biosynthesis inhibitor (e.g. abiraterone, enzalutamide, and/or apalutamide), treatment with a PARP inhibitor, and treatment with lutetium/radiolabeled PSMA.

[201] Furthermore, the multispecific antibody capable of binding PSMA may be for use in the treatment of cancers in which PSMA is expressed on the tumor neo-vasculature or tumor- associated endothelial cells of primary or metastatic tumors including from colorectal cancer, lung cancer, breast cancer, endometrial and ovarian cancer, gastric cancer, renal cell cancer, urothelial cancer, hepatocellular cancer, oral squamous cancer, thyroid tumors and glioblastomas. Moreover, the multispecific antibody capable of binding PSMA may be for use in the treatment of adenoid cystic carcinoma of the head and neck.

CD1d Multispecific Antibodies

[202] In some embodiments, the human cancer antigen is CD1d. Antibodies having antigenbinding regions capable of binding a human CD1d suitable for incorporation into a multispecific antibody to be used in the present disclosure have for example been described in WO 2016/122320 and WO 2020/060405 (both herein incorporated by reference).

[203] The (first) antigen-binding region of the multispecific antibody used in the present disclosure may comprise the VH CDR1 sequence of SEQ ID NO:95, the VH CDR2 sequence of SEQ ID NO: 96 and the VH CDR3 sequence of SEQ ID NO: 97. In some embodiments, the first antigen-binding region comprises an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 98. In some embodiments, the first antigen-binding region comprises an amino acid sequence comprising or consisting of SEQ ID NO: 98.

[204] The multispecific antibody used in the method of the present disclosure may compete (i.e. be able to compete) for binding to CD1d with an antibody having the sequence set forth in SEQ ID NO: 98. The multispecific antibody may bind the same epitope on human CD1d as an antibody having the sequence set forth in SEQ ID NO: 98.

[205] The multispecific antibody used in the present disclosure may comprise a first antigenbinding region and a second antigen-binding region, wherein the first antigen-binding region comprises the CDR1 sequence set forth in SEQ ID NO: 95, the CDR2 sequence set forth in SEQ ID NO: 96 and the CDR3 sequence set forth in SEQ ID NO: 97 and wherein the second antigenbinding region comprises the CDR1 sequence set forth in SEQ ID NO: 5, the CDR2 sequence set forth in SEQ ID NO: 6 and the CDR3 sequence set forth in SEQ ID NO: 7. In some embodiments, the second antigen-binding domain comprises any of the CDR combinations recited in Tables 6A-6D.

[206] The multispecific antibody used in the disclosure may comprise a first antigen-binding region capable of binding CD1d and be for use in the treatment of hematological malignancies such as T cell lymphoma, multiple myeloma, acute myeloid leukemia, acute lymphoblastic leukemia, chronic lymphocytic leukemia, mantle cell lymphoma, B cell lymphoma, smoldering myeloma, Hodgkin lymphoma, myelomonocytic leukemias, lymphoplasmacytic lymphoma, hairy cell leukemia, or splenic marginal zone lymphoma, or solid tumors, such as renal cell carcinoma, melanoma, colorectal carcinoma, head and neck cancer, breast cancer, prostate cancer, lung cancer, pancreatic cancer, gastro-esophageal cancer, small bowel carcinoma, central nervous system tumors, medulloblastomas, hepatocellular carcinoma, ovarian cancer, glioma, neuroblastoma, urothelial carcinomas, bladder cancer, sarcoma, penile cancer, basal cell carcinoma, merkel cell carcinoma, neuroendocrine carcinoma, neuroendocrine tumors, carcinoma of unknown primary (CUP), thymoma, vulvar cancer, cervical carcinoma, testicular cancer, cholangiocarcinoma, appendicular carcinoma, mesothelioma, ampullary carcinoma, anal cancer or choriocarcinoma.

CD40 Multispecific Antibodies

[207] In another embodiment, the human cancer antigen is CD40. Antibodies having antigenbinding regions capable of binding a human CD40 suitable for incorporation into a multispecific antibody to be used in the present disclosure have for example been described in WO 2020/159368 (herein incorporated by reference).

[208] The (first) antigen-binding region of the multispecific antibody used in the present disclosure may comprise the VH CDR1 sequence of SEQ ID NO: 99, the VH CDR2 sequence of SEQ ID NO: 100, and the VH CDR3 sequence of SEQ ID NO: 101. In some embodiments, the first antigen-binding region comprises an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 102. In some embodiments, the first antigen-binding region comprises an amino acid sequence comprising or consisting of SEQ ID NO: 102.

[209] In some embodiments, the multispecific antibody used in the method of the present disclosure may compete (i.e. be able to compete) for binding to CD40 with an antibody having the sequence set forth in SEQ ID NO: 102. The multispecific antibody may bind the same epitope on human CD40 as an antibody having the sequence set forth in SEQ ID NO: 102. [210] The multispecific antibody used in the present disclosure may comprise a first antigenbinding region and a second antigen-binding region, wherein the first antigen-binding region comprises the CDR1 sequence set forth in SEQ ID NO: 99, the CDR2 sequence set forth in SEQ ID NO: 100 and the CDR3 sequence set forth in SEQ ID NO: 101 and wherein the second antigenbinding region comprises the CDR1 sequence set forth in SEQ ID NO: 5, the CDR2 sequence set forth in SEQ ID NO: 6 and the CDR3 sequence set forth in SEQ ID NO: 7. In some embodiments, the second antigen-binding domain comprises any of the CDR combinations recited in Tables 6A-6D.

[211] The multispecific antibody used in the present disclosure may comprise a first antigenbinding region capable of binding CD40 and be for use in the treatment of chronic lymphocytic leukemia, multiple myeloma, non-Hodgkin's lymphoma, Hodgkin's lymphoma, follicular lymphoma, head and neck cancer, pancreatic cancer, ovarian cancer, lung cancer, breast cancer, colon cancer, prostate cancer, B-cell lymphoma/leukemia, Burkitt lymphoma or B acute lymphoblastic leukemia.

CD123 Multispecific Antibodies

[212] In another embodiment, the human cancer antigen is CD123. Antibodies having antigenbinding regions capable of binding a human CD123 suitable for incorporation into a multispecific antibody to be used in the present disclosure have for example been described in WO 2022/180271 (herein incorporated by reference).

[213] The (first) antigen-binding region of the multispecific antibody used in the present disclosure may comprise the VH CDR1 sequence of SEQ ID NO: 103, the VH CDR2 sequence of SEQ ID NO:104 and the VH CDR3 sequence of SEQ ID NO:105. In some embodiments, the first antigen-binding region comprises an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 106. In some embodiments, the first antigen-binding region comprises an amino acid sequence comprising or consisting of SEQ ID NO: 106.

[214] In some embodiments, the multispecific antibody used in the method of the present disclosure may compete (i.e. be able to compete) for binding to CD123 with an antibody having the sequence set forth in SEQ ID NO: 106. The multispecific antibody may bind the same epitope on human CD123 as an antibody having the sequence set forth in SEQ ID NO: 106.

[215] The multispecific antibody used in the present disclosure may comprise a first antigenbinding region and a second antigen-binding region, wherein the first antigen-binding region comprises the CDR1 sequence set forth in SEQ ID NO: 103, the CDR2 sequence set forth in SEQ ID NO: 104 and the CDR3 sequence set forth in SEQ ID NO: 105 and wherein the second antigenbinding region comprises the CDR1 sequence set forth in SEQ ID NO: 5, the CDR2 sequence set forth in SEQ ID NO: 6 and the CDR3 sequence set forth in SEQ ID NO: 7. In some embodiments, the second antigen-binding domain comprises any of the CDR combinations recited in Tables 6A-6D.

[216] The multispecific antibody used in the present disclosure may comprise a first antigenbinding region capable of binding CD123 and be for use in the treatment of acute myeloid leukemia, B-cell acute lymphoblastic leukemia, hairy cell leukemia, Hodgkin lymphoma, blastic plasmacytoid dendritic cell neoplasm, chronic myeloid leukemia, chronic lymphocytic leukemia, B-cell chronic lymphoproliferative disorders or myelodysplastic syndrome.

Nectin 4 Multispecific Antibodies

[217] In another embodiment, the human cancer antigen is Nectin-4. The (first) antigen-binding region of the multispecific antibody used in the present disclosure may comprise the VH CDR1 sequence selected from SEQ ID NOs: 107, 11 , 115, 119, 123, 127, and 131 , the VH CDR2 sequence selected from SEQ ID NOs: 108, 112, 116, 120, 124, 128, and 132 and the VH CDR3 sequence selected from SEQ ID NOs: 109, 113, 117, 121 , 125, 129, 133. In some embodiments, the first antigen-binding region comprises an amino acid sequence having at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to a sequence selected from SEQ ID NO: 110, 114, 118, 122, 126, 130, and 134. In some embodiments, the first antigenbinding region comprises an amino acid sequence comprising or consisting of a sequence selected from SEQ ID NO: 110, 114, 118, 122, 126, 130, and 134.

[218] In some embodiments, the multispecific antibody used in the method of the present disclosure may competes (i.e. be able to compete) for binding to Nectin-4 with an antibody having a sequence selected from SEQ ID NO: 110, 114, 118, 122, 126, 130, and 134. The multispecific antibody may bind the same epitope on human Nectin-4 as an antibody having a sequence selected from SEQ ID NO: 110, 114, 118, 122, 126, 130, and 134.

[219] The multispecific antibody used in the present disclosure may comprise a first antigenbinding region capable of binding Nectin-4 and be for use in the treatment of bladder cancer, breast cancer, renal cancer, prostate cancer, ovarian cancer, esophageal cancer, head and neck cancer, lung cancer, pancreatic cancer, gastric cancer, thyroid cancer, colorectal cancer, cholangiocarcinoma, or uterine corpus endometrial carcinoma. 5T4 Multispecific Antibodies

[220] In another embodiment, the human cancer antigen is 5T4. The (first) antigen-binding region of the multispecific antibody used in the present disclosure may comprise the VH CDR1 sequence selected from SEQ ID NOs: 135, 139, 143, 147, 151, and 159, the VH CDR2 sequence selected from SEQ ID NOs: 136, 140, 144, 148, 152, 156, and 160, and the VH CDR3 sequence selected from SEQ ID NOs: 137, 141 , 145, 149, 153, 157, and 161. In some embodiments, the first antigen-binding region comprises an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to a sequence selected from SEQ ID NO: 138, 142, 146, 150, 154, 158, and 162. In some embodiments, the first antigen-binding region comprises an amino acid sequence comprising or consisting a sequence selected from SEQ ID NO: 138, 142, 146, 150, 154, 158, and 162.

[221] In some embodiments, the multispecific antibody used in the method of the present disclosure may compete (i.e. be able to compete) for binding to 5T4 with an antibody having a sequence selected from SEQ ID NO: 138, 142, 146, 150, 154, 158, and 162. The multispecific antibody may bind the same epitope on human 5T4 as an antibody having a sequence selected from SEQ ID NO: 138, 142, 146, 150, 154, 158, and 162.

[222] The multispecific antibody used in the methods of present disclosure may comprise a first antigen-binding region capable of binding 5T4 and be for use in the treatment bladder cancer, cervical cancer, non-small cell lung cancer, mesothelioma, head and neck squamous cell carcinoma, glioblastoma multiform, esophageal cancer, pancreatic cancer, breast cancer, including triple-negative breast cancer, colorectal cancer, gastric cancer, ovarian cancer, uterine cancer, prostate cancer, renal cancer, esophageal cancer or pre-B acute lymphoblastic leukemia.

EGFR Multispecific Antibodies

[223] In some embodiments, the human cancer antigen is EGFR. The (first) antigen-binding region of the multispecific antibody used in the present disclosure may comprise the VH CDR1 sequence of SEQ ID NO: 91 , the VH CDR2 sequence of SEQ ID NO: 92, and the VH CDR3 sequence of SEQ ID NO: 93. In some embodiments, the first antigen-binding region comprises an amino acid sequence having at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to a sequence selected from SEQ ID NO: 94. In some embodiments, the first antigen-binding region comprises an amino acid sequence comprising or consisting of SEQ ID NO: 94.

[224] In some embodiments, the multispecific antibody used in the method of the present disclosure my i.e. be able to compete) for binding to EGFR with an antibody having the sequence set forth in SEQ ID NO: 94. The multispecific antibody may bind the same epitope on human EGFR as an antibody having the sequence set forth in SEQ ID NO: 94.

[225] The multispecific antibody used in the present disclosure may comprise a first antigenbinding region and a second antigen-binding region, wherein the first antigen-binding region comprises the CDR1 sequence set forth in SEQ ID NO: 91, the CDR2 sequence set forth in SEQ ID NO: 92 and the CDR3 sequence set forth in SEQ ID NO: 93 and wherein the second antigenbinding region comprises the CDR1 sequence set forth in SEQ ID NO: 5, the CDR2 sequence set forth in SEQ ID NO: 6 and the CDR3 sequence set forth in SEQ ID NO: 7.. In some embodiments, the second antigen-binding region comprises a CDR combination selected from Tabled 6A-6D.

[226] The multispecific antibody used in the methods of present disclosure may comprise a first antigen-binding region capable of binding EGFR and be for use in the treatment of primary or metastatic colon or colorectal cancer, peritoneal cancer, liver cancer, head and neck squamous cell carcinoma (HNSCC), non-small cell lung carcinoma (NSCLC), squamous cell carcinoma of the skin.

[227] The EGFR multispecific antibody may be able to activate human Vy9\/52 T cells. Activation of Vy9V52 T cells may be measured through measuring alterations in gene-expression and/or (surface) marker expression (e.g., activation markers, such as CD25, CD69, or CD107a) and/or secretory protein (e.g., cytokines or chemokines) profiles. The EGFR multispecific antibody may be able to increase the number of cells positive for CD107a at least 2-fold, such as at least 5-fold, for example when tested as described in Example 9 in WO 2022/122973 (herein incorporated by reference), e.g. at a concentration of 1 nM, preferably 100pM, preferably 10pM, preferably 1pM, even more preferably 100fM. The EGFR multispecific antibody of the present disclosure may have an EC50 value for increasing the percentage of CD107a positive cells of 100 pM or less, such as 50 pM or less, e.g. 25 pM or less, such as 20 pM or less, e.g. 15 pM or less when tested for example using Vy9V52 T cells and A431 target cells as described in Example 9 of WO 2022/122973.

Half-life extension domains

[228] In some embodiments, the multispecific antibody used in the present disclosure may further comprise a half-life extension domain that extends the serum half-life of the multispecific antibody. Examples for means to extend serum half-life of the binding agents of the disclosure include peptides, proteins or domains of proteins, which are fused or otherwise attached to the multispecific antibodies. The group of peptides, proteins or protein domains includes peptides binding to other proteins with preferred pharmacokinetic profile in the human body such as serum albumin (see WO 2009/127691). As used herein, the term "human serum albumin" refers to the albumin protein present in human blood plasma. Human serum albumin is the most abundant protein in the blood. It constitutes about half of the blood serum protein. In some embodiments, a human serum albumin has the sequence of UniProt ID NO: P02768. In some embodiments, the multispecific antibody may have a terminal half-life that is longer than about 168 hours when administered to a human subject. The terminal half-life may be 336 hours or longer. The “terminal half-life” of an antibody, when used herein refers to the time taken for the serum concentration of the polypeptide to be reduced by 50%, in vivo in the final phase of elimination.

[229] Half-life extension domains include larger domains of proteins or complete proteins includes e.g. the fusion of human serum albumin, variants or mutants of human serum albumin (see WO 2011/051489, WO 2012/059486, WO 2012/150319, WO 2013/135896, WO 2014/072481 , WO 2013/075066) or domains thereof as well as the fusion of constant region of immunoglobulins (Fc domains) and variants thereof, as described herein. Such variants of Fc domains may be optimized/modified in order to allow the desired pairing of dimers or multimers, to abolish Fc receptor binding (e.g., the Fcg receptor), to enhance binding to FcRn, or for other reasons. A further concept known in the art to extend the half-life of small protein compounds in the human body is the pegylation of those compounds such as the polypeptide or binding agent of the present disclosure.

[230] In some embodiments, the half-life extension domain may be an Fc domain (also referred to as an “Fc region”). As used herein, “Fc domain” describes the minimum region (in the context of a larger polypeptide) or smallest protein folded structure (in the context of an isolated protein) that can bind to or be bound by an Fc receptor (FcR). As used herein, “Fc domain monomer” describes the single chain protein that, when associated with another Fc domain monomer, forms a functional Fc domain. The association of two Fc domain monomers creates one Fc domain. When two Fc domain monomers associate, the resulting Fc domain has Fc receptor binding activity. Thus, an Fc domain is a dimeric structure that can bind an Fc receptor. Unless otherwise noted, all references herein to a “variant Fc domain” are to be understood as referring to a dimeric Fc domain, in which each Fc domain monomer comprises the referenced mutation.

[231] It will be understood that Fc domain as used herein includes the polypeptides comprising the constant region of an antibody excluding the first constant region immunoglobulin domain. Thus, Fc domain refers to the last two constant region immunoglobulin domains (CH2, CH3) of IgG and optionally the flexible hinge N-terminal to these domains. Unless otherwise noted, all references to amino acid positions in Fc domains and Fc domain monomers are according to the EU index as set forth in Kabat (1991 , NIH Publication 91-3242, National Technical Information Service, Springfield, Va.). It is noted that polymorphisms have been observed at a number of Fc domain positions, including but not limited to Kabat 270, 272, 312, 315, 356, and 358, and thus slight differences between the sequences provided herein and sequences in the art may exist.

[232] The Fc domain may be derived from any of a variety of different antibody isotypes, including but not limited to, a wild-type or modified lgG1 , lgG2, lgG3, lgG4. IgA, IgE, or IgM. In some embodiments, the Fc domain is derived from a human lgG1.

[233] There are many known polymorphs for the lgG1 Fc domain, including the “DEL” polymorph and the “EEM” polymorph. The DEL polymorph comprises the amino acids D-E-L at positions 356, 357, and 358, respectively (e.g., SEQ ID NO: 337). The EEM polymorph comprises the amino acids E-E-M at positions 356, 357, and 358, respectively (e.g., SEQ ID NO: 336). Two binding agents that are otherwise identical except for the presence of a DEL Fc domain or an EEM Fc domain are expected to demonstrate similar properties in terms of ligand binding and therapeutic efficacy. In some embodiments, the Fc domain is a DEL Fc domain. In some embodiments, the Fc domain is an EEM Fc domain.

[234] In some embodiments, the Fc domain is a variant Fc domain that forms a variant Fc domain with a desirable property, such as increased half-life, compared to naturally occurring (wild-type) Fc sequences. As used herein, a “variant Fc domain” refers to a non-naturally occurring Fc domain, for example an Fc domain comprising one or more non-naturally occurring amino acid residues, one or more amino acid substitutions relative to a wild-type human constant domain, or one or more amino acid deletion, addition and/or modification.

[235] The serum half-life of binding agents comprising Fc domains may be increased by increasing the binding affinity of the Fc domain for FcRn. In some embodiments, the Fc domain variant has enhanced serum half-life relative to a comparable molecule. In a particular embodiment, the Fc domain variant comprises at least one amino acid substitution at one or more positions selected from the group consisting of M252Y, S254T and T256E (referred to herein as “YTE”; e.g., SEQ ID NO: 338 and 339). In another embodiment, the Fc domain variant comprises a Y at position 252 (referred to herein as “Y”, e.g., SEQ ID NO: 340 and 341). Optionally, the Fc domain variant may comprise a non-naturally occurring amino acid residue at additional and/or alternative positions known to one skilled in the art (see, e.g., U.S. Pat. Nos. 5,624,821 ; 6,277,375; 6,737,056; PCT Patent Publications WO 01/58957; WO 02/06919; WO 04/016750; WO 04/029207; WO 04/035752; WO 04/074455; WO 04/099249; WO 04/063351 ; WO 05/070963; WO 05/040217, WO 05/092925, and WO 06/020114).

[236] In an exemplary embodiment, the Fc domain monomer allows assembly of two or more polypeptide chains in a covalent manner, for example by disulfide linking between cysteine residues. In this way the Fc domain monomer acts as a dimerization domain, allowing assembly of two polypeptide chains to form a dimer. In some embodiments, such dimers comprise two polypeptides, each polypeptide including an antigen-binding domain described herein linked to an Fc domain monomer, thereby forming a bivalent binding agent. In some embodiments, such dimers comprise two polypeptides, one polypeptide comprising an antigen-binding domain described herein linked to an Fc domain monomer, and one polypeptide comprising an Fc domain monomer, thereby forming a monovalent binding agent.

[237] The Fc domain may be a heterodimer comprising two Fc monomers, wherein the first antigen-binding domain is fused to the first Fc monomer and the second antigen-binding region is fused to the second Fc monomer and wherein the first and second Fc monomers comprise asymmetric amino acid mutations that favor the formation of heterodimers over the formation of homodimers (see e.g., Ridgway et al. (1996) 'Knobs-into-holes' engineering of antibody CH3 domains for heavy chain heterodimerization. Protein Eng 9:617). The CH3 regions of the Fc monomers may comprise said asymmetric amino acid mutations, for example the first Fc polypeptide may comprise a T366W substitution (e.g., “knob” mutations), and the second Fc polypeptide may comprise T366S, L368A and Y407V substitutions (e.g., “hole” mutations), or vice versa, wherein the amino acid positions correspond to human lgG1 according to the EU numbering system. Furthermore, the cysteine residues at position 220 in the first and second Fc polypeptides may have been deleted or substituted, wherein the amino acid position corresponds to human lgG1 according to the EU numbering system. See e.g., SEQ ID NOs: 343 and 344.

[238] Furthermore, the first and/or second Fc monomers may contain mutations that render the antibody inert, i.e. unable to, or having reduced ability to, mediate Fc effector functions. The inert Fc domain may in addition not be able to bind C1q. The first and second Fc monomers may comprise a mutation at position 234 and/or 235, for example the first and second Fc monomer may comprise an L234F and an L235E substitution, wherein the amino acid positions correspond to human lgG1 according to the EU numbering system. See e.g., SEQ ID NO: 342.

[239] Exemplary Fc domain monomer sequences are shown in TABLE 9. Hinge sequences are shown in bold and italicized text; DEL and EEM polymorphisms are shown in boxed text; mutations relative to WT IgG 1 are shown in bold and underlined text. Table 9: Fc domain sequences*

[240] In some embodiments, the Fc domains described in TABLE 9 comprise a wild type lgG1 hinge (e.g., SEQ ID NO: 347 - EPKSCDKTHTCPPCP). In some embodiments, the Fc domains described in TABLE 9 comprise a modified lgG1 hinge (e.g., SEQ ID NO: 348 - AAASDKTHTCPPCP).

Kits

[241] In some embodiments, the present disclosure provides a kit for the treatment of cancer comprising: (a) a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vy9V62 T cell receptor, as described herein; and (b) a dose of a common gamma cytokine (e.g., IL-2, IL-15, or a variant thereof), optionally comprising instructions for use.

[242] In some embodiments, the present disclosure provides a kit for the treatment of cancer comprising: (a) a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vy9V<52 T cell receptor, as described herein; and (b) an immune checkpoint inhibitor, optionally comprising instructions for use. In some embodiments, the immune checkpoint inhibitor is an anti- PD1 antibody (e.g., pembrolizumab).

[243] In a further aspect, the present disclosure relates to a kit for the treatment of cancer comprising: (a) a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vy9V<52 T cell receptor, as described herein; (b) an immune checkpoint inhibitor (e.g., an anti- PD1 antibody such as pembrolizumab); and (c) a common gamma cytokine (e.g., IL-2 or IL-15), optionally comprising instructions for use.

[244] The kit may further comprise a solvent for the dilution of the multispecific antibody, the common gamma chain cytokine, and/or the immune checkpoint inhibitor.

INCORPORATION BY REFERENCE

[245] All references, articles, publications, patents, patent publications, and patent applications cited herein are incorporated by reference in their entireties for all purposes. However, mention of any reference, article, publication, patent, patent publication, and patent application herein is not, and should not be, taken as acknowledgment or any form of suggestion that they constitute valid prior art or form part of the common general knowledge in any country in the world.

EXAMPLES

Example 1 - Dose escalation of LAVA-1207 in metastatic castration resistant prostate cancer (mCRPC) patients

Background

[246] LAVA-1207 is a bispecific humanized antibody of 78 kDa that engages Vy9V52-T cells and PSMA+ tumor cells. LAVA-1207 is a disulfide-linked heterodimer of two fusion proteins set forth in SEQ ID NO: 334 and SEQ ID NO:335, further described and characterized in WO 2022/008646 (incorporated herein by reference).

Trial Design and Objectives

[247] This study was an open-label, multi-center, Phase 1/2a dose escalation study in patients with therapy-refractory, metastatic, castration-resistant prostate cancer (mCRPC).

[248] LAVA-1207 is administered via intravenous infusion every 2 weeks. Dose levels are as follows: 1.5 pg (starting dose) (Dose Level (DL) 1), 4.5 pg (DL2), 13.5 pg (DL3), 40 pg (DL4), 120 pg (DL5), 360 pg (DL6), 540 pg (DL7), and 800 pg (DL8)

[249] The objectives of the study were to investigate safety and tolerability, evaluate PK, PD, immunogenicity, and preliminary antitumor activity of LAVA-1207. Exemplary baseline patient characteristics are provided in Table 10 below. Patients were treated with LAVA-1207 with treatment duration ranging from 4 to 38 weeks. Table 10: Exemplary Patient Baseline Characteristics

Results

[250] Safety: Up to and including a dose of 120 pg, the frequency and severity of adverse effects did not appear to be dose-dependent. Most observed adverse effects were not suspected to be treatment-related.

[251] Antitumor activity: Anti-tumour activity was determined by evaluating prostate-specific antigen (PSA) changes in blood samples of patients upon treatment with LAVA-1207. Several patients showed stable or reduced PSA levels after LAVA-1207 treatment (Fig. 1). Furthermore, the baseline Vy9V62-T cell frequencies as percentage of total T cells were determined for each patient (not shown). At least in some instances, PSA decline was observed patients with high Vy9V52-T cells as a percentage of total T cells. See N304 PSA levels in Fig. 2 and N304 Vy9V52- T cell relative changes in Fig. 3A. Fig. 3A shows a pronounced drop in Vy9V52-T cell frequency in the peripheral blood 2 hr after dosing, likely reflective of Vy9V62-T cell migration/re-distribution with subsequent recovery. Fig. 3C and Fig. 3D also show Vy9V62-T cell activation markers (CD25 - Fig. 3C and CD69 - Fig. 3D) upregulated following dosing and that Receptor occupancy (RO - Fig. 3B) was detectable up to day 14 after Eol, with peak levels ranging from 6.1% to 12.6% in dose level 4 (40pg). Example 2 - Evaluation of the safety, tolerability, pharmacokinetics, pharmacodynamics, immunogenicity, and antitumor activity of a PSMA targeting bispecific y5-T cell engager, in patients with therapy refractory metastatic castration resistant prostate cancer

Study Design:

[252] This trial is an open-label, multi-center, Phase 1 and 2a dose escalation trial with an expansion cohort to investigate the safety, tolerability, pharmacokinetics, pharmacodynamics, immunogenicity and preliminary antitumor activity of LAVA-1207 in patients with therapy refractory metastatic castration resistant prostate cancer (mCRPC).

[253] The trial starts with an open-label, dose-escalation part (Part 1) to determine the recommended Phase 2 dose (RP2D). The second part of the trial (Part 2) is an open-label expansion cohort at the RP2D and schedule, in which the number of patients will be expanded to confirm safety in a patient population with therapy refractory mCRPC with measurable disease. LAVA-1207 will be administered with or without low dose subcutaneous IL-2 (referred to as LDSC IL-2), and optionally pembrolizumab may be administered.

Part 1- dose escalation

Dose cohorts and dosing regimen

[254] In the present trial, escalating dose levels are envisaged, including cohorts that include priming doses. Eligible patients will receive sequentially higher doses of LAVA-1207 in subsequent dose cohorts and will continue receiving LAVA-1207 until 24 weeks or up to disease progression, unacceptable toxicity or withdrawal of consent or otherwise as specified in the investigational medicinal product (IMP) discontinuation criteria.

[255] Patients in a dose cohort will receive LAVA- 1207 as intravenous (IV) infusions at the same dose in a dosing interval of 2 weeks (equal to one cycle) consecutively without interruption, except when necessary to manage adverse events (AEs). The infusion duration for this biologic product will initially be 2 hours in the first cycle, 1 hour in the 2^nd cycle and 30 minutes in subsequent cycles.

[256] In all cohorts, at least 3 patients will be enrolled at the same dose level unless 1 patient develops a dose-limiting-toxicity (DLT) during the first 28 days of treatment (defined as DLT period).

Dose escalation and RP2D determination

[257] The planned dose cohorts for Part 1 of the trial are presented in Table 11 below: Table 11 : Planned dose cohorts for Part 1

[258] Dose escalation will continue until the RP2D can be defined by an optimal biological active dose of LAVA-1207 or until a maximum tolerated dose (MTD) has been determined, whichever is reached first. An optimal biological active dose is defined as a safe dose that demonstrates the greatest pharmacological activity of LAVA-1207. Considerations for pharmacological activity include the evaluation of effects on Vy9V52-T cells such as binding of LAVA-1207 to Vy9V52-T cells and changes in the frequency or activation status of Vy9V52-T cells as assessed in peripheral blood. For the final RP2D determination, safety (including AEs that may have occurred during the whole treatment period, including any late-onset effects during the posttreatment phase), preliminary antitumor activity, pharmacokinetic and pharmacodynamic data will be considered. [259] Preliminary data on Adverse events is provided in Figure 5. Treatment emergent AEs (TEAEs) that were suspected to be related were grade 1 or 2. No increase in severity or frequency of TEAEs with increasing doses and no patient discontinued treatment due to AE.

Exploratory dose cohorts with LAVA-1207 and low dose SC administration of IL-2 (LDSC IL-2)

[260] For dose level 6 of LAVA-1207, two exploratory cohorts (n=3 per cohort) will be initiated in which IL-2 will be administered subcutaneously (SC) as a single dose (cohort A1) or as 3 doses (cohort A2) starting at 24 hours after the start of LAVA-1207 infusion (see Table 12).

Table 12: Exploratory dose cohorts with LAVA-1207 and LDSC IL-2

a) LDSC IL-2 administration of 1 x 10⁶ IU (1 MIU/day) - one day - at 24 hours after LAVA-1207 start of infusion (Sol). Note: the allowed time window is -1 hour before - and +3 hours at 24 hours after LAVA-1207 Sol. b) LDSC IL-2 administration of 1 x 10^e IU (1 MIU/day) - every day for 3 days - at 24 -, 48-, and 72 hours after LAVA-1207 start of infusion. Note: the allowed time window is -1 hour before and +3 hours after each specified timepoint.

[261] Administration of LDSC IL-2 will occur during the first 4 cycles of LAVA-1207 treatment only (e.g. up to 8 weeks). IL-2 is expected to increase the number of Vy9V52-T cells, thereby increasing the total number of Vy9V52 -T cells available for LAVA-1207 mediated lysis of PSMA expressing prostate cancer cells.

[262] Starting from a dose level which is determined as safe and well tolerated by the DEC, cohorts will be initiated in with an anti-PD-L1 antibody at 400 mg intravenously over 30 minutes every 6 weeks [Q6W] in addition to the target dose of the multispecific antibody starting at Cycle 2 (second target dose). Treatment will be administered for a period not to exceed 2 years (18 cycles of the anti-PD-L1 antibody or until one or more discontinuation criteria have been met.

[263] The dose of pembrolizumab will remain constant at 400 mg Q6W for each dose level of the multispecific antibody.

Part 2 - Expansion cohort

[264] This trial will comprise additional cohorts to further investigate the safety, tolerability, immunogenicity, pharmacokinetics and pharmacodynamics of LAVA-1207 in combination with IL- 2 and to investigate the preliminary antitumor activity in patients with therapy refractory mCRPC in measurable disease.

[265] Primary objectives of the dose escalation cohort will include: (a) Investigate the safety and tolerability of LAVA-1207 in patients with therapy refractory mCRPC.

(b) Determine the RP2D of LAVA- 1207 in patients with therapy refractory mCRPC for LAVA-1207 monotherapy, for LAVA-1207 + LDSC IL-2, and for LAVA-1207 + pembrolizumab.

[266] Primary objectives of the expansion cohort will include:

(a) Investigate the safety and tolerability of LAVA-1207 at the RP2D in therapy refractory mCRPC patients with measurable and non-measurable disease.

[267] Secondary objectives of both cohorts will include:

(a) Explore the preliminary antitumor activity of LAVA-1207.

(b) Evaluate the pharmacokinetics of LAVA-1207.

(c) Evaluate the pharmacodynamics of LAVA- 1207.

(d) Evaluate the immunogenicity of LAVA-1207.

[268] Exploratory objectives of both cohorts will include:

(a) Investigate the safety and tolerability of LAVA-1207 and LDSC IL-2 in patients with therapy refractory mCRPC.

(b) Explore the preliminary pharmacokinetics of LAVA-1207 and LDSC IL-2.

(c) Explore the preliminary pharmacodynamics of LAVA-1207 and LDSC IL-2.

(d) Explore the preliminary immunogenicity of LAVA-1207 and LDSC IL-2

(e) Explore the preliminary antitumor activity of LAVA-1207 and LDSC IL-2.

(f) To evaluate the effect of study treatment on circulating tumor cells (CTCs) and circulating tumor DNA (ctDNA)

[269] Primary endpoints for the dose escalation for LAVA-1207 alone, LAVA-1207 plus LDSC IL-2, and LAVA-1207 plus pembrolizumab will include:

(a) Frequency and severity of AEs using the Common Terminology Criteria for Adverse Events (CTCAE) version 5.0 and ASTCT grading for CRS.

(b) Frequency and type of DLT.

[270] Primary endpoints for the expansion cohort for LAVA-1207 alone, LAVA-1207 plus LDSC IL-2, and LAVA-1207 plus pembrolizumab will include:

(a) Frequency and severity of AEs using the CTCAE version 5.0 and ASTCT grading of CRS at the RP2D.

[271] Secondary endpoints for both cohorts will include:

(a) Number of participants with an antitumor response according to immune response evaluation criteria in solid tumors (iRECIST) in patients with measurable disease. (b) Duration of response.

(c) Disease control rate (DCR) for patients with measurable disease at 8, 16 and 24 weeks.

(d) Number of participants who experience any prostate specific antigen (PSA) decrease, and number of participants who experience a PSA decrease of > 50% from baseline.

(e) Progression free survival (using Prostate Cancer Working Group (PCWG3) for bone lesions and/or iRECIST criteria for soft-tissue lesions).

(f) Pharmacokinetic parameters.

(g) Pharmacodynamic markers.

(h) Incidence and prevalence of anti-LAVA-1207 antibodies.

[272] Exploratory endpoints for both cohorts include:

(a) Frequency and severity of AEs using the CTCAE version 5.0 and ASTCT grading for CRS.

(b) Pharmacokinetic parameters.

(c) Pharmacodynamic markers.

(d) Incidence and prevalence of anti-LAVA-1207 antibodies.

(e) Number of participants with an antitumor response according to immune response evaluation criteria in solid tumors (iRECIST) in patients with measurable disease.

(f) Number of participants who experience any PSA decrease from baseline, and number of participants who experience a decrease of > 50% from baseline.

(g) DCR for patients with measurable disease at 8, 16 and 24 weeks.

(h) Determine any relationships between CTC/ctDNA changes versus baseline and treatment dose and clinical response.

Inclusion Criteria:

[273] Patients are eligible to be included in the trial only if all of the following criteria apply:

(a) Patient must be 18 years of age inclusive or above, at the time of signing the informed consent.

(b) Male patient with mCRPC as defined by PCWG3 criteria (histologically confirmed adenocarcinoma; adenocarcinoma with <10% small-cell or neuroendocrine features is allowed). Brain metastasis are allowed as long as the patient’s symptoms are well controlled.

(c) Patient should have failed at least 1 line of taxane-based chemotherapy or is deemed medically unsuitable to be treated with a taxane regimen. (d) Patient should have received a 2nd generation or later androgen receptor targeted therapy/ androgen biosynthesis inhibitor (e.g. abiraterone, enzalutamide, and/or apalutamide). Progression on novel antiandrogen therapy may have occurred in the non- metastatic CRPC setting.

(e) Patient is unlikely to tolerate or derive clinically meaningful benefit from other available therapy.

(f) Patients for which any drug related toxicity adverse effects of any prior cancer therapy should have resolved to Grade 1 or less according CTCAE v5.0 or to baseline severity level.

(g) Patients with evidence of progressive disease, defined as 1 or more criteria:

(i) PSA level > 1 ng/mL that has increased on at least 2 successive occasions at least 1 week apart.

(ii) Computed tomography (CT) or magnetic resonance imaging (MRI) scan: nodal or visceral progression as defined by RECIST 1.1.

(iii) Bone scintigraphy: appearance of 2 or more new metastatic lesions.

(h) Patient should have undergone bilateral orchiectomy or should be on continuous androgen-deprivation therapy (ADT) with a gonadotropin-releasing hormone agonist or antagonist (surgical or medical castration).

(i) Total serum testosterone < 50 ng/dL or 1 .73 nmol/L.

(j) Evaluable (measurable or non-measurable) disease for prostate cancer.

(k) Predicted life-expectancy of > 6 months.

(l) Eastern Cooperative Oncology Group (ECOG) performance status of 0 or 1.

(m) Adequate renal (estimated glomerular filtration rate [eGFR] per local laboratory> 40 ml_/min/1.73m2), hepatic (bilirubine < 2 times upper limit of normal (ULN), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) < 3.0 times ULN). In case of liver metastases AST and ALT < 5.0 times ULN is allowed. Adequate hematological function (neutrophils > 1 x10⁹/L, platelet count > 75x10⁹/L, Hb>9g/dL) with no prior transfusions within 2 weeks.

(n) Males who are:

(i) Surgically sterile (bilateral orchiectomy, vasectomy).

(ii) Compliant with an effective contraceptive regimen (i.e. use of male condom with female partner and assuring use of an additional highly effective contraceptive method with a failure rate of <1% per year when having sexual intercourse with a woman of childbearing potential who is not currently pregnant) following from signing of the informed consent form (ICF) through 90 days after the last IMP administration. Abstinence is not considered an adequate contraceptive regimen.

(iii) Refraining from donating sperm following from signing of the ICF through 90 days after the last IMP administration.

(o) Capable of giving signed and dated informed consent prior to initiation of any trial-related procedures that are not considered Standard of Care which includes compliance with the requirements and restrictions listed in the ICF and in the protocol.

Exclusion Criteria:

[274] Patients are excluded from the trial if any of the following criteria apply:

(a) Other malignancies within the last 2 years except adequately treated carcinoma in situ, basal or squamous cell skin carcinoma.

(b) Uncontrolled or severe intercurrent medical condition.

(c) Positive serological testing for human immunodeficiency virus (HIV) antibody.

(d) Positive serological hepatitis B surface antigen [HBsAg] and hepatitis B core antibody (anti-HBc) negative, and hepatitis C virus antibody. Patients who are positive for anti-HBc or hepatitis C antibody may be included if they have a negative polymerase chain reaction (PCR) within 6 weeks prior to initial IMP administration. Those who are PCR positive will be excluded.

(e) Patient has any active-, uncontrolled-, or suspected infection.

(f) Known clinically relevant immunodeficiency disorders.

(g) A significant history of renal, neurologic, psychiatric, pulmonary, endocrinologic, metabolic, immunologic, cardiovascular, or hepatic disease that in the opinion of the investigator would adversely affect participation in this trial.

(h) Unstable cardiovascular function defined as: (a) symptomatic ischemia, or (b) uncontrolled clinically significant conduction abnormalities (i.e. ventricular tachycardia on antiarrhythmic agents are excluded; 1 st degree atrioventricular block or asymptomatic left anterior fascicular block/right bundle branch block is not excluded), or (c) congestive heart failure New York Heart Association Class > 3, or (d) myocardial infarction within 3 months. (i) Previous treatment with antitumor therapies 2 weeks prior to initial IMP for radiotherapy and androgen receptor targeted therapy/ androgen biosynthesis inhibitor, and 4 weeks for systemic chemotherapy or targeted-/ immunotherapy.

(j) Previous treatment with live or live attenuated vaccines within 2 weeks prior to initial IMP administration. New types of vaccines need to be evaluated as to their mode of action.

(k) Treatment with other investigational agents in the 4 weeks prior to initial IMP.

(l) Major surgery within 4 weeks prior to initial IMP administration.

(m) Hypersensitivity to any of the excipients present in LAVA-1207, IL-2, or pembrolizumab (if applicable).

(n) Previous treatment with any systemic immunosuppressant within 4 weeks prior to initial IMP administration, with the exception of systemic corticosteroid use up to oral dose of 10 mg prednisone daily (or equivalent for other steroids).

(o) Previous treatment with an aminobisphosphonate IV (e.g. ibandronate, pamidronate, zoledronate etc) within 4 weeks or 1 year prior to initial IMP.

(p) Known ongoing drug and alcohol abuse in the opinion of the investigator.

Additional Exclusion Criteria for the pembrolizumab trial:

[275] Patients are excluded from the LAVA-1207 + pembrolizumab trial if any of the following criteria apply:

(a) Active infection requiring systemic therapy.

(b) Has a history of (non-infectious) pneumonitis/interstitial lung disease that required steroids or has current pneumonitis/interstitial lung disease.

(c) Has a diagnosis of immunodeficiency or is receiving chronic systemic steroid therapy (in dosing exceeding 10 mg daily of prednisone equivalent) or any other form of immunosuppressive therapy within 7 days prior to the first dose of study drug.

(d) Has an active autoimmune disease that has required systemic treatment in past 2 years (i.e. , with use of disease modifying agents, corticosteroids or immunosuppressive drugs). Replacement therapy (e.g., thyroxine, insulin, or physiologic corticosteroid replacement therapy for adrenal or pituitary insufficiency) is not considered a form of systemic treatment and is allowed.

(e) Has received prior therapy with an anti-programmed cell death 1 (PD 1), anti-programmed cell death ligand 1 (PD-L1), or anti-programmed cell death ligand 2 (PD-L2) agent or with an agent directed to another stimulatory or co-inhibitory T-cell receptor (e.g., CTLA- 4, OX 40, CD137), and was discontinued from that treatment due to a Grade 3 or higher immune related AE.

(f) A significant history of renal, neurologic, psychiatric, pulmonary, endocrinologic, metabolic, immunologic, cardiovascular, or hepatic disease that in the opinion of the investigator would adversely affect participation in this trial.

(g) Has known active central nervous system (CNS) metastases and/or carcinomatous meningitis. Participants with previously treated brain metastases may participate provided they are radiologically stable, i.e. , without evidence of progression for at least 4 weeks by repeat imaging (note that the repeat imaging should be performed during study screening), clinically stable and without requirement of steroid treatment for at least 14 days prior to first dose of study treatment.

(h) Has received prior systemic anti-cancer therapy including investigational agents within 4 weeks (could consider shorter interval for kinase inhibitors or other short half-life drugs) prior to treatment. Note: Participants must have recovered from all AEs due to previous therapies to <Grade 1 or baseline. Participants with <Grade 2 neuropathy may be eligible. Participants with endocrine-related AEs <Grade 2 requiring treatment or hormone replacement may be eligible. Note: If the participant had major surgery, the participant must have recovered adequately from the procedure and/or any complications from the surgery prior to starting study intervention.

(i) Has received prior radiotherapy within 2 weeks of start of study treatment or have had a history of radiation pneumonitis. Note: Participants must have recovered from all radiation-related toxicities and not require corticosteroids. A 1-week washout is permitted for palliative radiation (<2 weeks of radiotherapy) to non-CNS disease.

0) Has received a live or live-attenuated vaccine within 30 days prior to the first dose of study intervention. Note: Administration of killed vaccines are allowed.

(k) Is currently participating in or has participated in a study of an investigational agent or has used an investigational device within 4 weeks prior to the first dose of study treatment. Note: Participants who have entered the follow-up phase of an investigational study may participate as long as it has been 4 weeks after the last dose of the previous investigational agent.

(l) Has a history or current evidence of any condition, therapy, or laboratory abnormality, or other circumstance that might confound the results of the study or interfere with the participant’s participation for the full duration of the study, such that it is not in the best interest of the participant to participate, in the opinion of the treating investigator.

(m) Has a known psychiatric or substance abuse disorder that would interfere with the participant’s ability to cooperate with the requirements of the study.

Duration Patient Participation:

[276] Screening phase: within 28 days before initial IMP.

[277] Treatment phase: planned treatment duration of 24 weeks. At the request of the treating physician and in consultation with the sponsor, continued access to IMP, alone or in combination with pembrolizumab, might be offered beyond the planned treatment phase for individual patients with ongoing disease control.

[278] For patients on LAVA-1207 plus pembrolizumab arm, treatment duration will be for up to 18 cycles of pembrolizumab (approximately 24 months) or until any discontinuation criteria are met.

[279] Post-treatment phase: A follow-up phase of 120 days after the last dose of IMP for each patient with an end of treatment (EoT) visit between 14 and 42 days after the last dose of IMP for each patient.

IMP, dose, mode of administration and dose regimen:

[280] LAVA-1207 is a concentrate for solution for infusion and will be administered as IV infusion with a 14-day dosing interval. The infusion duration will be 2 hours (+15 minutes) for any priming dose(s) and for the first target dose, followed by 1 hour for the second target dose and 30 minutes in subsequent cycles. A step-dosing regimen during Cycle 1 will start at Expanded Cohort 8 onwards.

[281] Patients in the dose escalation arm with pembrolizumab will receive pembrolizumab 400 mg IV Q6W starting at the second target LAVA-1207 dose. Patients who are dosed with LAVA- 1207 and pembrolizumab on the same day will wait at least 30 minutes post end of infusion of pembrolizumab to be dosed with LAVA-1207.

[282] LDSC IL-2 will be administered as a single dose or as 3 daily doses starting at 24 hours (-1 hr- and +2 hrs) after the start of LAVA-1207 infusion for a total duration of 4 cycles.

[283] In Part 2, patients will receive IV infusions of LAVA 1207 at the RP2D (dose and schedule) as established in the dose escalation part of the trial (with- or without LDSC IL-2).

[284] In Part 2 expansion arm with pembrolizumab, patients will receive IV infusions of LAVA- 1207 plus pembrolizumab at the RP2D (dose[s] and schedule) as established in the dose escalation part of the trial. Pembrolizumab will be administered at least 30 minutes before LAVA- 1207 administration.

[285] In Part 1 of this trial, a patient experiencing a DLT per protocol or Grade 3 non-hematologic toxicity may require a dose reduction or dose delay.

[286] For Part 2, specific toxicities requiring dose interruptions or reductions will be added to the protocol prior to initiation of a dose expansion cohort.

[287] In general, patients may continue treatment upon resolution of a (serious) (S)AE to a Grade 1 AE or be withdrawn from the trial.

Criteria for Evaluation and Analyses:

Safety of LAVA-1207 alone. LAVA-1207 + LDSC IL-2, and LAVA-1207 + pembrolizumab

[288] Vital signs, physical examination, clinical chemistry, hematology, urinalysis, electrocardiograms (ECG) and documentation of any reported AEs will be recorded. The severity of AEs will be graded according to the National Cancer Institute’s CTCAE version 5.0 (NCI CTCAE) Grading Scale (see the NCI CTCAE web page at http://ctep.cancer.gov for details) with the exception of CRS, which is evaluated and graded according to the ASTCT consensus criteria.

[289] LAVA-1207 has safely reached a dose of 1200 pg in therapy refractory mCRPC patients.

Pharmacodynamics of LAVA-1207 alone, LAVA-1207 + LDSC IL-2, and LAVA-1207 + pembrolizumab

[290] Blood samples will be collected to determine LAVA-1207 concentrations and evaluate the following pharmacodynamic markers: binding of LAVA- 1207 to Vy9V52-T cells, Vy9V52-T cells (activation status and frequency) and general immune cell analysis (activation status and frequency of B-cells, T-cell subsets (CD4+ , CD8+ and Treg), natural killer (NK) cells and monocytes) and induction of cytokines (e.g. interleukin [IL]-1P, IL-2, IL 6, IL-8, tumor necrosis factor [TNF]-a, interferon [IFN]-y and Granulocyte-macrophage colony-stimulating factor [GM- CSF]).

[291] Biopsies (optional and only for patients with metastatic sites that are accessible for biopsy) will be collected to evaluate the following pharmacodynamic markers: the infiltration and activation status of Vy9V52- T-cells / PSMA expression I Butyrophilin expression/ Tumor cell viability. Preliminary pharmacokinetics results of LAVA-1207 are shown in Fig. 4. Fig. 4 shows that the pharmacokinetics of LAVA-1207 appears linear.

Immunogenicity

[292] Blood samples will be collected to determine potential development of anti-drug antibodies directed against LAVA-1207. Antitumor activity of LAVA-1207 alone, LAVA-1207 + LDSC IL-2, and LAVA-1207 + pembrolizumab

[293] Blood samples will be collected to determine:

(a) PSA assessment.

(b) CTC and/or ctDNA assessments.

[294] Imaging will be performed for:

(a) PSMA-Positron emission tomography (PET).

(b) Tumor assessments by (contrast-enhanced) CT scan of chest, abdomen and pelvis.

(c) Bone scintigraphy.

Statistical Considerations:

[295] In general, data will be summarized by dose level (and separately with or without LDSC IL-2 with or without pembrolizumab) as well as for the entire population. Baseline and safety data will be presented based on the Safety Set, which will include all patients who received at least one dose of IMP. Summaries of antitumor data will be based on the Evaluable Set, which will include all patients who received at least one dose of IMP and who have disease evaluations performed to assess response, and the intent- to- treat analysis set (all enrolled patients). Pharmacokinetic and pharmacodynamic analyses will include all patients who received at least one dose of IMP and have at least one post-dose measurement available.

[296] Safety: All safety outcomes will be summarized using descriptive statistics. AEs starting or worsening at or after the first dose of IMP (treatment-emergent AEs [TEAEs]) will be presented separately from those starting prior to the first dose of IMP or those observed beyond the EoT visit during the post-treatment phase. The number and percentage of patients with TEAEs will be summarized by primary Medical Dictionary for Regulatory Activities (MedDRA) System Organ Class (SOC) and Preferred Term (PT). Separate tables showing all TEAEs, all serious TEAEs and all TEAEs suspected to be related to IMP will be generated. Number and percentage of patients with DLTs will be summarized; DLTs will be presented in by-patient listings.

[297] Vital signs, ECGs, physical examination findings, and safety laboratory parameters will be analyzed using descriptive statistics.

[298] Laboratory abnormalities will be summarized by parameter in frequency tables and in shift tables relative to the baseline value.

[299] Pharmacokinetics: Individual patient serum LAVA-1207 concentrations will be used to derive pharmacokinetic parameters using standard non-compartmental methods. [300] Pharmacodynamics: Pharmacodynamic markers and possible induction of cytokines will be analyzed descriptively.

[301] Immunogenicity: The incidence and individual serum titer and isotype of anti-drug antibodies directed against LAVA-1207 will be summarized.

Antitumor activity and disease evaluation

[302] Tumor response according to iRECIST for patients with measurable disease (in Part 1 and 2) will be summarized.

[303] DCR (complete response (CR)+partial response (PR)+ stable disease (SD)) for patients with measurable disease (in Part 1 and 2) at 8, 16 and 24 weeks.

[304] Any PSA reduction (defined as any decline of PSA level compared to baseline) and PSA response (defined as > 50% decline of PSA level compared to baseline) for patients evaluable for PSA response will be analyzed descriptively.

[305] Progression free survival will be descriptively analyzed with using a Kaplan-Meier approach. Progression free survival is defined as the time from start of treatment until progression (soft-tissue disease progression [by iRECIST], bone lesion progression [by PCWG3 criteria]), or death.

[306] Exploratory endpoints (e.g. CTC/ctDNA changes versus baseline) will be analyzed descriptively.

Sample Size Justification:

[307] The sample size for Part 1 of this trial is determined by clinical assessment and predefined dose escalations rather than statistical considerations.

[308] The sample size for Part 2 (n=30 for LAVA-1207 or LAVA-1207 plus LDSC IL-2 and n=40 for LAVA-1207 plus pembrolizumab arm) will allow for an overall safety evaluation of LAVA-1207 at the recommended dose and schedule. At least 20 patients with measurable disease will be enrolled to receive LAVA-1207 or LAVA-1207 plus LDSC IL-2 allowing for a preliminary efficacy evaluation of LAVA-1207 in 2 relatively homogeneous populations of patients with measurable and non-measurable disease.

Dose Escalation Committee (DEC):

[309] The DEC consists of minimally the medical monitor, minimally one of the principal investigators and a representative of the sponsor. [310] The trial design outlined in Example 2 can be repeated, wherein IL-15 is substituted for IL-2.

Example 3: Anti-PD1 combination therapy

[311] Fig. 6A-Fig. 6B shows increased PD-1 expression on Vy9V52-T cells after treatment with LAVA-1207. Fig. 7A shows percentages of Vy9V32 T cells in untreated non-malignant (n.m), malignant (m) areas of the prostate of patients with prostate cancer, and prostate cancer patient PBMC samples. Fig. 7B and Fig. 7C shows PD-1 expression on Vy9V62 T cells and CD3+ T cells from the same prostate cancer patient samples. Fig. 8 shows that Vy9V<52 T cells activated in vitro with a V62-bsTCE (bispecific T cell engager) upregulate PD-1. Each of these data suggest a potential for benefit with combination therapy with anti-PD1 or anti-PD-L1 antibodies.

Example 4: Exemplary LAVA-1207 and IL-2 dosing regimen

[312] Fig. 9 and Fig. 10 show exemplary dosing regimen of LAVA-1207 and IL-2. In each of these exemplary regimens, pembrolizumab may also be included.

[313] In the exemplary dosing regimen of Fig. 9, in cycle one, the patient is administered a priming dose of LAVA-1207 at day -14 and at day -7. Then at day 0 the patient is administered a target dose of LAVA-1207. Then at days 1 , 2, and 3 the patient is administered a low dose of IL- 2. This cycle is repeated without the priming dose 3 more time every 2 weeks, and then the patient is administered only a target dose of LAVA-1207 for each subsequent cycle.

[314] The rationale for including IL-2 in the clinical protocol can be seen in Fig.11A and Fig. 11 B showing that the expansion of Vy9V52 T cells over baseline by an exemplary bsTCE or pamidronate is highly IL-2-dependent. Little to no expansion was observed in the absence of IL- 2.

[315] A similar exemplary dosing regimen may be performed, using IL-15 instead of IL-2.

Example 5: LAVA-1207 triggers activation of Vy9V52-T cells in patient-derived tumor tissue and PBMCs and induces selective prostate tumor cell lysis

[316] Peripheral blood mononuclear cells (PBMC) were derived from healthy donor- and prostate cancer patient blood and isolated using Lymphoprep™ (AXI-1114547, Fresenius) density gradient centrifugation. Blood samples were obtained from Sanquin (Amsterdam, The Netherlands) in case of healthy donors or from prostate cancer patients under written informed consent from the Amsterdam UMC (location VUmc, Amsterdam, The Netherlands). PBMC were processed for phenotypic analysis using flow cytometry or resuspended in RPMI medium complete for functional experiments. [317] Healthy donor-derived Vy9V62-T cells were isolated from PBMC and expanded. If purity of Vy9V62-T cells was >95% of total cells (referred to as expanded Vy9V52-T cells) then they were used in experiments. Alternatively, Vy9V62-T cells were isolated from PBMC using the EasySep™ human gamma/delta T cell isolation kit (19255, STEMCELL Technologies) and directly used in functional experiments (referred to as non-expanded Vy9V62-T cells).

[318] Non-malignant and malignant tissue samples were collected from radical prostatectomies of patients with non-metastatic prostate cancer. In combination with clinical and radiological findings provided by treating physicians, the pathologist assessed whether the tissue pieces were non-malignant or malignant by macroscopic analysis. Tissues were cut into small pieces, resuspended in dissociation medium composed of Iscove’s Modified Dulbecco’s Medium (IMDM, 12440053, Gibco) supplemented with 0.1% DNAse I (10104159001 , Roche), 0.14% Collagenase A (10103586001, Roche), 100 lU/mL PSG and 5% FCS, transferred to a sterile flask, and then incubated with a magnetic stirrer for 45 minutes at 37° C. Cell suspensions were subsequently strained with a 100 pM cell strainer (352360, BD Falcon).

[319] Dissociation steps were repeated for 1-3 rounds. Single cell suspensions derived from these tissues were processed for phenotypic analysis using flow cytometry or resuspended in RPMI medium complete for functional experiments.

[320] LNCaP (PSMA+ clone FGC, CRL-1740) and PC3 (PSMA-, CRL-1435) were obtained from American Type Culture Collection (ATCC), and VCaP (PSMA+, 06020201-1VL) and 22Rv1 (PSMA+, 5092802) were obtained from the European Collection of Authenticated Cell Cultures (ECACC). Cell lines were maintained in Roswell Park Memorial Institute (RPMI) medium (22400089, Gibco) supplemented with 10% (v/v) fetal calf serum (FCS, 04-007-1 A, Biological Industries), 0.05 mM [3-mercaptoethanol (200-646-6, Merck), 100 lU/mL sodium penicillin, 100 pg/mL streptomycin sulfate and 2.0 mM L-glutamine (PSG, 10378-016, Life Technologies), referred to as RPMI medium complete. Cell lines were kept at 37°C in a humidified atmosphere containing 5% CO2.

[321] FIG. 12A shows the frequency of Vy9V<52 T cells of all CD3+ cells in PBMCs (2.6 ± 0.7% (mean ± SEM; n=40)), non-malignant tissue (1.4 ± 0.4% (mean ± SEM; n=31)), and malignant tissue (1.6 ± 0.5% (mean ± SEM; n=31)).

[322] Phenotypical analysis of Vy9VS2-T cells and CD3+ non-Vy9VS2-T cells showed that CD69, an early activation and tissue-residency marker, trended towards higher expression in non- malignant and malignant prostate tissue as compared to PBMCs (68.1% ± 4.5% of Vy9V52-T cells in PBMC; CD69 mean ± SEM: 81.4% ± 8.9% of Vy9V62-T cells in non-malignant prostate tissue; and 87.7% ± 6.0% of Vy9V52-T cells in malignant prostate tissue); malignant prostate tissue VY9V62-T cells also expressed higher levels of the costimulatory marker 4-1 BB and the terminal differentiation marker CD57. However, PD-1 expression did not differ across Vy9V52-T cells in PBMC, non-malignant and malignant prostate tissue. Comparatively, the percentage of all CD3+ cells that expressed CD69, 4-1 BB and PD-1 was significantly higher in non-malignant and malignant prostate tissue as compared to PBMC. See FIG. 12B.

[323] In order to test the responsiveness of Vy9V52-T cells in prostate cancer patients, LAVA- 1207 (PSMA-V62-Fc) was incubated with malignant (PSMA expression MFI mean ± SEM; 5.7 ± 2.4) and non-malignant prostate tissue (PSMA expression MFI mean ± SEM; 0.7 ± 0.3), alone or co-cultured with autologous patient PBMCs for 24 hours. When incubated with the co-culture of malignant prostate tissue and PBMCs, 50 nM LAVA- 1207 increased the degranulation in both the malignant prostate tissue (Tl tissue-infiltrated) and PBMCs (FIG. 12C). In contrast, LAVA-1207 did not enhance the degranulation in non-malignant prostate tissue alone or when co-cultured with PBMCs (see FIG. 12D). Furthermore, in 24 hour co-cultures of patient-derived PBMCs, prostate tissue (PBMC:target cell ratio = 10:1 ; Vy9V52-T celktarget cell ratio = 1 :25 (donor 1), 1 :9 (donor 2) and 1 :5 (donor 3)), and LAVA-1207, there was an increase in tumor cell lysis (as defined by CD45-EpCAM^dim/+ cells) as compared to non-tumor cell lysis (CD45’EpCAM‘ cells) See FIG. 12E. In 2 of the 3 donors tumor lysis was observed in the LAVA-1207-only conditions showing that the tumor cell lysis was being mediated by intratumoral Vy9V02-T cells.

Example 6: NKG2D and DNAM-1 receptor ligand interactions contribute to LAVA-1207- induced Vy9V52-T cell degranulation and tumor cell lysis

[324] Since LAVA- 1207 induced Vy9V52-T cell degranulation and lysis in the presence of malignant but not non-malignant tissue, the factors that could potentially contribute to this tumor preferential activity were analyzed. PSMA (unpaired and paired), BTN3A, BTN2A1 , Nectin-2 (DNAM-1 ligand), PVR (DNAM-1 ligand), MIC-A/B (NKG2D ligand), ULBP-3 (NKG2D ligand), ULBP-2/5/6 (NKG2D ligand), and HLA-E (NKG2A ligand) were all expressed at higher levels in malignant prostate tissue as compared to non-malignant prostate tissue. Only ULBP-1 (NKG2D ligand) was expressed at similar levels between the malignant and non-malignant prostate tissue. However, the frequency of the NKG2D and NKG2A receptor expression on Vy9V52-T cells did vary significantly between malignant and non-malignant prostate tissue, and the percentage of Vy9V62-T cells positive for DNAM-1 receptor was lower in malignant prostate tissue than in non- malignant prostate tissue. See FIG. 13A-FIG. 13C.

[325] Expanded healthy donor-derived Vy9V52-T cells were cultured with prostate cell lines (LNCaP, VCaP, or 22Rv1) for 4 hours after a 30 minute pre-incubation with 10 pg/ml Fc receptor block (130-059-901 , Miltenyi Biotec) and a-DNAM-1 (clone DX11 , 559787, BD Bioscience), a- NKG2D (clone 149810, MAB139-100, R&D Systems) or a-NKG2A (Monalizumab, PX-TA1392- 100UG, Proteogenics) blocking antibodies were added 30 minutes before incubation with 0.001- 5 nM LAVA-1207.

[326] Vy9V52-T cell degranulation (as measured by CD107a expression), cytokine production (IFN-y, TNF, IL-2, and IL-4), and tumor cell lysis all increased with increasing concentration of LAVA-1207. However, in the presence of the neutralizing antibodies directed towards the DNAM- 1 or NKG2D receptors this increase could be partially mitigated except at the highest (saturating) concentrations of LAVA-1207. See FIG. 14A- FIG. 14H. Neutralizing antibodies to NKG2A and BTN3A did not have a significant impact on the LAVA-1207-mediated Vy9V52-T cell degranulation. See FIG. 15A-FIG. 15B.

[327] Next, to assess if these receptor interactions also impact Vy9V52-T cell reactivity using patient tumor samples, dissociated prostate tumor samples from 3 patients were cultured for 4 hours in the presence of 0.05 nM LAVA-1207 and either DNAM-1 or NKG2D blocking antibodies. LAVA-1207 induced an increase in Vy9V52-T cell degranulation, but this increase was significantly reduced in the presence of the DNAM-1 blocking antibody. While the presence of the NKG2D blocking antibody only reduced degranulation in 1 of the 3 donors. See FIG. 16.

[328] In conclusion, DNAM-1 and NKG2D receptor interactions can contribute to Vy9V52-T cell reactivity and prostate cancer lysis at non-saturating LAVA- 1207 concentrations.

Example 7: LAVA-1207 triggers in vivo antitumor activity in a xenograft prostate cancer model using human PBMC

[329] In order to evaluate the in vivo therapeutic efficacy of LAVA-1207, male immunodeficient NCG mice were inoculated subcutaneously with 22Rv1 5x10⁶ tumor cells mixed with human healthy donor derived PBMC in a ratio of 2:1 (22Rv1 :PBMC, n=2 PBMC donors: Vy9V52-T cell frequencies of respectively 21.9% and 8.8% of total CD3+ cells resulting in 22Rv1 :Vy9V52-T ratios of 22:1 and 60:1). Mice were treated with the LAVA-1207 (0.2 and 2.0 mg/kg) or PBS IV weekly starting at the day of tumor cell + PBMC inoculation on days 7, 14 and 21. Tumor volumes were measured twice per week using a caliper, and the volume was expressed in mm³ using the formula: “V = (L x W x W)/2, where V was tumor volume, L was tumor length (the longest tumor dimension) and Wwas tumor width (the longest tumor dimension perpendicular to L). At the end of the follow-up period, tumor growth inhibition (TGI) was calculated as: ((tumor volume of LAVA- 1207-treated mice in mm³ - tumor volume of PBMC treated mice in mm³)/tumor volume of PBMC treated mice in mm³)*100%. The animals were sacrificed when the mean tumor volume of a group exceeded 2.000 mm³.

[330] As seen in FIG. 17A-FIG. 17B, inoculation with PBMCs did not influence tumor growth, but when combined with LAVA-1207 at either 0.2 or 2.0 mg/kg there was significant inhibition of tumor growth (TGI) (0.2 mg/kg: P=<0.0001 , TGI 61% and 2.0 mg/kg: P=<0.0001 and TGI 60%) increase in overall survival (0.2 mg/kg: P=<0.0001 and 2.0 mg/kg: P=<0.0001). The median overall survival was 51 days in both of the LA A-1207+PBMC-treated groups and 37 days in the PBMC treated group.

Example 8: LAVA-1207 Tissue Cross-reactivity

[331] In order to ascertain tissue cross-reactivity of LAVA-1207 in humans, LAVA-1207 was FITC-labeled by Squarix GmbH (Marl, Germany) using a controlled reaction with NHS-coupled FITC. The average number of fluorochrome molecules conjugated per protein (F/P) was 4.3. Tissue-reactivity was assessed by Charles River Laboratories (Evreux, France) by immunohistochemistry (IHC) using the FITC-labeled-LAVA-1207 in a panel of 41 different frozen normal human tissues (adrenal, bone marrow, breast/ mammary gland, cecum, cerebellum, cerebral cortex, colon, duodenum, endothelium [vessels], eye, esophagus, fallopian tube [oviduct], gall bladder, heart [ventricle], ileum, jejunum, kidney [cortex], liver, lung, lymph node, muscle [striated, skeletal], peripheral nerve, ovary, pancreas, parotid, parathyroid pituitary, placenta, prostate, rectum, skin, spinal cord, spleen, stomach, testis, thymus, thyroid, tonsil, ureter, urinary bladder, uterus [cervix] and uterus [endometrium]) and peripheral blood smears (three donors per tissue or blood smear). Frozen sections were air-dried, fixed in zinc formalin (4087236, Microm Microtech), and rinsed using Milipore water. Test samples were incubated with 0, 3 or 20 pg/ml FITC-labeled-LAVA-1207 followed by 2 pg/ml goat a-FITC antibody (01-40-01 , KPL) after which antibody block (760-4204, Discovery) was applied. The ChromoMab DAB kit (760-159, Discovery) was used for detection according to manufacturer recommendations. Test samples were then stained with hematoxylin II (790-2208, Ventana) and bluing reagent (760- 2037, Ventana), washed, dehydrated and mounted. Slides were evaluated by a pathologist using a light microscope (Olympus BX51).

[332] Labeling of LAVA-1207 with FITC did not affect the affinity of LAVA-1207 to either Vy9V52- T cells (Vy9V62-T cell binding EC50= 1.41 nM (FITC-labeled LAVA-1207) versus 0.97 nM (LAVA- 1207)) (FIG. 18A) or PSMA-expressing LNCaP cells (LNCaP binding EC50 6.00 nM (FITC- labeled LAVA-1207) versus 6.28 nM (LAVA-1207)) (FIG. 18B). [333] Next, the reactivity of LAVA-1207 to a panel of 42 different frozen normal human tissues and blood smears (three donors per tissue/ blood smear) was tested.

[334] Moderate to marked membrane staining (with some variable cytoplasmic staining) was seen in acinar cells of the prostate. Low, mainly cytoplasmatic, intensity staining was seen in acinar cells of the parotid gland in three out of three donors. Minimal staining in fusiform cells of blood vessels and placenta villi, which are considered to represent endothelial cells, was seen in the placenta of one out of three donors (representative images are depicted in FIG. 19A). No reactivity was detected in a wide range of other tissue samples including: adrenal, blood smears, breast, small and large intestines, gall bladder, kidney, liver, lung, lymph node, nerve, ovary, oviduct, spleen, testis, thymus, tonsil, ureter, urinary bladder, cervix and uterus endometrium, with the exception of incidental reactivity to membranes of lymphocyte like cells, which are most probably Vy9V52-T cells (representative images shown in FIG. 19B). This tissue reactivity analysis of the LAVA-1207 was in line with the known tissue distribution of PSMA expression (i.e. prostate, parotid glands and placenta) and Vy9V52-T cells and did not reveal any unexpected reactivity.

Claims

1. A method of treating a cancer in a subject in need thereof comprising administering to the subject a. a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vy9V52 T cell receptor; and b. an immune checkpoint inhibitor.

2. A method of treating a cancer in a subject in need thereof comprising administering to the subject a. a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vy9V52 T cell receptor; and b. a common gamma chain cytokine.

3. The method of claim 2, further comprising administering an immune checkpoint inhibitor.

4. The method of claim 1 or claim 3, wherein the immune checkpoint inhibitor is an anti-PD1 antibody or an anti-PDL1 antibody.

5. The method of any one of claims 1-3, wherein the common gamma chain cytokine is IL-2 or a variant thereof.

6. The method of claim 5, wherein the IL-2 is administered a dose of less than 3 MIU/day or less than 2 MIU/day.

7. The method of any one of claims 1-3, wherein the common gamma chain cytokine is IL- 15.

8. The method of claim 7, wherein the IL-15 is administered at a dose of 0.5-5 MIU/ day.

9. The method according to claim 8, wherein the IL-2 or IL- 15 is pegylated.

10. The method according to any of the preceding claims, wherein the multispecific antibody is administered intravenously or subcutaneously.

11. The method according to any of the preceding claims, wherein IL-2 or IL-15 is administered subcutaneously.

12. The method according to any of the preceding claims, wherein the multispecific antibody is administered in a dosing interval of 7 to 21 days.

13. The method according to any of the preceding claims, wherein the multispecific antibody is administered in a dosing interval of 14 days.

14. The method according to any of the preceding claims, wherein administering the multispecific antibody comprises: a. administering one or more priming doses of the multispecific antibody and b. administering one or more target doses of the multispecific antibody.

15. The method according to any of the preceding claims, wherein the IL-2 or IL-15 is administered daily for at least 1 day after administration of the multispecific antibody.

16. The method of any of the preceding claims, wherein the IL-2 or IL-15 is administered daily for at least 3 days after administration of the multispecific antibody.

17. The method according to any of the preceding claims, wherein the IL-2 or IL-15 is administered at a dose of between 0.5 and 1.5 MIU/day.

18. The method of any one of the preceding claims, wherein the IL-2 or IL-15 is administered at a dose of 1 MIU/day.

19. The method according to any of the preceding claims, wherein the multispecific antibody is administered at a dose of at least 1 .5, 4.5, 13.5, 120, 360, 540, 800, or 1200 micrograms.

20. The method of any one of the preceding claims, wherein the human cancer antigen is selected from the group consisting of: PSMA, CD1d, CD40, CD123, 5T4, and Nectin-4.

21. The method according to any one of the preceding claims, wherein the multispecific antibody is capable of binding to human V62.

22. The method according to any one of the preceding claims, wherein the second antigenbinding region comprises: a. the VH CDR1 sequence of SEQ ID NO: 1 , the VH CDR2 sequence of SEQ ID NO: 2 and the VH CDR3 sequence of SEQ ID NO: 3; b. the VH CDR1 sequence of SEQ ID NO: 5, the VH CDR2 sequence of SEQ ID NO: 6 and the VH CDR3 sequence of SEQ ID NO: 7; c. the VH CDR1 sequence of SEQ ID NO: 10, the VH CDR2 sequence of SEQ ID NO: 11 and the VH CDR3 sequence of SEQ ID NO: 12; d. the VH CDR1 sequence of SEQ ID NO: 14, the VH CDR2 sequence of SEQ ID NO: 15 and the VH CDR3 sequence of SEQ ID NO: 16; e. the VH CDR1 sequence of SEQ ID NO: 18, the VH CDR2 sequence of SEQ ID NO: 19 and the VH CDR3 sequence of SEQ ID NO: 20; f. the VH CDR1 sequence of SEQ ID NO: 22, the VH CDR2 sequence of SEQ ID NO: 23 and the VH CDR3 sequence of SEQ ID NO: 24; or g. the VH CDR1 sequence of SEQ ID NO: 26, the VH CDR2 sequence of SEQ ID NO: 27 and the VH CDR3 sequence of SEQ ID NO: 28.

23. The method according to any one of the preceding claims, wherein the second antigenbinding region comprises an amino acid sequence comprising at least 90% sequence identity to an amino acid sequence selected from SEQ ID NOs: 4, 8, 9, 13, 17, 21 , 25, and 29.

24. The method according to any one of the preceding claims, wherein: a. the human cancer antigen is PSMA and wherein the first antigen-binding region comprises the VH CDR1 sequence of SEQ ID NO: 87, the VH CDR2 sequence of SEQ ID NO: 88 and the VH CDR3 sequence of SEQ ID NO: 89; b. the human cancer antigen is CD1d and wherein the first antigen-binding region comprises the VH CDR1 sequence of SEQ ID NO: 95, the VH CDR2 sequence of SEQ ID NO: 96 and the VH CDR3 sequence of SEQ ID NO: 97; c. the human cancer antigen is CD40 and wherein the first antigen-binding region comprises the VH CDR1 sequence of SEQ ID NO: 99, the VH CDR2 sequence of SEQ ID NO: 100 and the VH CDR3 sequence of SEQ ID NO: 101 ; d. the human cancer antigen is CD123 and wherein the first antigen-binding region comprises the VH CDR1 sequence of SEQ ID NO: 103, the VH CDR2 sequence of SEQ ID NO: 104 and the VH CDR3 sequence of SEQ ID NO: 105; e. the human cancer antigen is Nectin-4 and wherein the first antigen-binding region comprises the VH CDR1 sequence selected from SEQ ID NO: 107, 11 , 115, 119, 123, 127, and 131 , the VH CDR2 sequence selected from SEQ ID NO: 108, 112, 116, 120, 124, 128, and 132 and the VH CDR3 sequence selected from SEQ ID NO: 109, 113, 117, 121 , 125, 129, and 133; or f. the human cancer antigen is 5T4 and wherein the first antigen-binding region comprises the VH CDR1 sequence of SEQ ID NO: 135, 139, 143, 147, 151 , 155, and 159, the VH CDR2 sequence of SEQ ID NO: 136, 140, 144, 148, 152, 156, and 160 and the VH CDR3 sequence of SEQ ID NO: 137, 141, 145, 149, 153, 157, and 161.

25. The method according to any one of the preceding claims, wherein a. the human cancer antigen is PSMA and the first antigen-binding region comprises an amino acid sequence having at least 90% sequence identity to SEQ ID NO:90; b. the human cancer antigen is CD1d and the first antigen-binding region comprises an amino acid sequence having at least 90% sequence identity to SEQ ID NO:98; c. the human cancer antigen is CD40 and the first antigen-binding region comprises an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 102; d. the human cancer antigen is CD123 and the first antigen-binding region comprises an amino acid sequence having at least 90% sequence identity to the sequence set forth in SEQ ID NO: 106; e. the human cancer antigen is Nectin-4 and the first antigen-binding region comprises an amino acid sequence having at least 90% sequence identity to a sequence selected from SEQ ID NOs: 110, 114, 118, 122, 126, 130, and 134; or f. the human cancer antigen is 5T4 and the first antigen-binding region comprises an amino acid sequence having at least 90% sequence identity a sequence selected from SEQ ID NOs: 138, 142, 146, 150, 154, 158, and 162.

26. The method according to any one of the preceding claims, wherein the multispecific antibody comprises an Fc region.

27. The method of claim 26, wherein the Fc region is a heterodimeric Fc region comprising a first and a second Fc monomer polypeptide.

28. The method of claim 27, wherein (a) the first Fc polypeptide comprises the sequence set forth in SEQ ID NO: 345 and the second Fc polypeptide comprises the sequence set forth in SEQ ID NO: 346, or (b) the first Fc polypeptide comprises the sequence set forth in SEQ ID NO: 346 and the second Fc polypeptide comprises the sequence set forth in SEQ ID NO: 345.

29. The method according to any of the preceding claims, wherein the multispecific antibody is formulated at a strength of 1 mg/mL in 10 mM histidine, 1 mM methionine, 280 mM sucrose, 0.02 % polysorbate 80, pH 6.0.

30. The method of any one of the preceding claims, wherein the cancer is (i) prostate cancer, such as non-metastatic or metastatic prostate cancer, for example metastatic castration resistant prostate cancer, such as therapy refractory metastatic castration resistant prostate cancer, (ii) a cancer in which PSMA is expressed on the tumor neo-vasculature or tumor-associated endothelial cells of primary or metastatic tumors, including from colorectal cancer, lung cancer, breast cancer, endometrial and ovarian cancer, gastric cancer, renal cell cancer, urothelial cancer, hepatocellular cancer, oral squamous cancer, thyroid tumors and glioblastomas, or (iii) adenoid cystic carcinoma of the head and neck.

31. The method of any one of the preceding claims, wherein the cancer is (i) a hematological malignancy such as T cell lymphoma, multiple myeloma, acute myeloid leukemia, acute lymphoblastic leukemia, chronic lymphocytic leukemia, mantle cell lymphoma, B cell lymphoma, smoldering myeloma, Hodgkin lymphoma, myelomonocytic leukemias, lymphoplasmacytic lymphoma, hairy cell leukemia, or splenic marginal zone lymphoma, or (ii) a solid tumor cancer, such as renal cell carcinoma, melanoma, colorectal carcinoma, head and neck cancer, lung cancer, pancreatic cancer, gastro-esophageal cancer, small bowel carcinoma, central nervous system tumors, medulloblastomas, hepatocellular carcinoma, glioma, neuroblastoma, urothelial carcinomas, bladder cancer, sarcoma, penile cancer, basal cell carcinoma, merkel cell carcinoma, neuroendocrine carcinoma, neuroendocrine tumors, carcinoma of unknown primary (CUP), thymoma, vulvar cancer, cervical carcinoma, testicular cancer, cholangiocarcinoma, appendicular carcinoma, mesothelioma, ampullary carcinoma, anal cancer, or choriocarcinoma.

32. The method of any one of the preceding claims, wherein the cancer is non-Hodgkin's lymphoma, Hodgkin's lymphoma, follicular lymphoma, pancreatic cancer, lung cancer, colon cancer, B-cell lymphoma/leukemia, Burkitt lymphoma or B acute lymphoblastic leukemia.

33. The method of any one of the preceding claims, wherein the cancer is B-cell acute lymphoblastic leukemia, blastic plasmacytoid dendritic neoplasm, chronic myeloid leukemia, B- cell chronic lymphoproliferative disorders or myelodysplastic syndrome.

34. The method of any one of the preceding claims, wherein the cancer is bladder cancer, renal cancer, esophageal cancer, lung cancer, pancreatic cancer, thyroid cancer, colorectal cancer, cholangiocarcinoma, or uterine corpus endometrial carcinoma.

35. The method of any one of the preceding claims, wherein the cancer is bladder cancer, cervical cancer, non-small cell lung cancer, mesothelioma, head and neck squamous cell carcinoma, glioblastoma multiform, esophageal cancer, pancreatic cancer, colorectal cancer, uterine cancer, renal cancer, esophageal cancer, or pre-B acute lymphoblastic leukemia.

36. A kit for the treatment of cancer comprising: (a) a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vy9V62 T cell receptor; and (b) a dose of IL-2 less than 3 Ml U and/or an anti-PD1 or anti-PD-L1 antibody, optionally comprising instructions for use.

37. A kit for the treatment of cancer comprising: (a) a multispecific antibody comprising a first antigen-binding region capable of binding a human cancer antigen and a second antigen-binding region capable of binding a human Vy9V62 T cell receptor; and (b) a dose of IL-2 less than 3 MIU and/or an anti-PD1 or anti-PD-L1 antibody, optionally comprising instructions for use.

38. The kit of claim 37, further comprising a solvent for the dilution of the multispecific antibody, the dose of IL-2 and/or IL-15, and/or the anti-PD1 or anti-PD-L1 antibody