Compositions and methods for treating cancer

Abstract

The invention provides methods, compositions, and combinations for treating cancer or a non-malignant disease or condition in which αv and / or α5 (e.g., α5β1 ) integrin is expressed or implicated in pathogenesis or progression. The methods include administration of a bispecific antibody that specifically binds αv integrin and α5 (e.g., α5β1 ) integrin and a Src inhibitor to provide treatment for the cancer or non-malignant disease or condition.

Description

[0001] ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700

[0002] COMPOSITIONS AND METHODS FOR TREATING CANCER

[0003] Statement as to Federally Funded Research

[0004] This invention was made with government support under grant number 1 R01 CA245864-01 A1 awarded by the National Institutes of Health. The government has certain rights in the invention.

[0005] Sequence Listing

[0006] The instant application contains a Sequence Listing which has been submitted electronically in XML format and is hereby incorporated by reference in its entirety. Said XML copy, created on June 17, 2025, is named 00398-571 WO2_Sequence_Listing_6_17_25 and is 41 ,785 bytes in size.

[0007] Background of the Invention

[0008] Integrin av, integrin a5, and Src signaling have been independently implicated in cancer biology. However, there has been limited success in the development of approved therapies against these targets. There remains a need for treatment options for patients having cancers characterized by signaling through integrin av, integrin a5, and Src.

[0009] Summary of the Invention

[0010] In one aspect, the invention provides methods of treating a subject having cancer or a non- malignant condition or disease in which av and / or a5 (e.g., a5p1 ) integrin is expressed or implicated in pathogenesis or progression, the method comprising administering to the subject therapeutically effective amounts of (i) a bispecific antibody that specifically binds av integrin and a5 (e.g., a5p1 ) integrin and (ii) a Src inhibitor.

[0011] In some embodiments, the bispecific antibody comprises (i) a first paratope that specifically binds an av integrin; and (ii) a second paratope that specifically binds an a5 (e.g., a5p1 ) integrin.

[0012] In some embodiments, the second paratope specifically binds an epitope residing wholly within the a5 integrin.

[0013] In some embodiments, the second paratope specifically binds an epitope comprising both amino acid residues with the a5 integrin and amino acid residues within a pi integrin dimerized to the a5 integrin.

[0014] In some embodiments, (i) the first paratope comprises a light chain variable domain comprising CDR amino acid sequences of SEQ ID NOs: 1 -3 and a heavy chain variable domain comprising CDR amino acid sequences of SEQ ID NOs: 4-6; and (ii) the second paratope comprises a light chain variable domain comprising CDR amino acid sequences of SEQ ID NOs: 7-9 and a heavy chain variable domain comprising CDR amino acid sequences of SEQ ID NOs: 10-12.

[0015] In some embodiments, (i) the light chain variable domain of the first paratope further comprises at least one framework region (FR) comprising the amino acid sequence of one of SEQ ID NOs: 13-16 or an amino acid sequence that is at least 80% identical to any one of SEQ ID NOs: 13-16; (ii) the heavy chain variable domain of the first paratope further comprises at least one FR comprising the amino acid sequence of one of SEQ ID NOs: 17-20 or an amino acid sequence that is at least 80% identical to any ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700 one of SEQ ID NOs: 17-20; (iii) the light chain variable domain of the second paratope further comprises at least one FR comprising the amino acid sequence of one of SEQ ID NOs: 21 -24 or an amino acid sequence that is at least 80% identical to any one of SEQ ID NOs: 21 -24; or (iv) the heavy chain variable domain of the second paratope further comprises at least one FR comprising the amino acid sequence of one of SEQ ID NOs: 25-28 or an amino acid sequence that is at least 80% identical to any one of SEQ ID NOs: 25-28.

[0016] In some embodiments, the bispecific antibody comprises (i) an antigen binding fragment portion that comprises one of the first or second paratopes; and (ii) a single-chain Fv portion that comprises the other of the first or second paratopes.

[0017] In some embodiments, the bispecific antibody further comprises an Fc region.

[0018] In some embodiments, the bispecific antibody comprises (i) two polypeptides, each comprising an amino acid sequence that is at least 80% identical to SEQ ID NO: 41 ; and (ii) two additional polypeptides, each comprising an amino acid sequence that is at least 80% identical to SEQ ID NO: 42.

[0019] In some embodiments, the bispecific antibody is chimeric.

[0020] In some embodiments, the bispecific antibody is at least partially humanized or de-immunized.

[0021] In some embodiments, the bispecific antibody is divalent, trivalent, or tetravalent.

[0022] In some embodiments, the Src inhibitor is a dual Src-Abl inhibitor.

[0023] In some embodiments, the Src inhibitor is selected from the group consisting of dasatinib, dasatinib monohydrate, bafetinib, bosutinib, danusertib, ponatinib, repotrectinib, saracatinib, tirbanibulin, AP24149, AP24163, AZD0424, CCT196969, DC2036, NVP-BHG712, PD173955, PP1 , PP2, PP121 , SGX393, WH-4-023, XL228, and 3,4-Methylenedioxy-p-nitrostyrene.

[0024] In some embodiments, the Src inhibitor is dasatinib.

[0025] In some embodiments, the bispecific antibody and Src inhibitor are administered to the subject at the same time.

[0026] In some embodiments, the bispecific antibody and Src inhibitor are administered to the subject sequentially.

[0027] In some embodiments, the bispecific antibody is administered to the subject before the Src inhibitor is administered to the subject.

[0028] In some embodiments, the bispecific antibody is administered to the subject after the Src inhibitor is administered to the subject.

[0029] In some embodiments, the cancer is selected from the group consisting of prostate cancer, kidney cancer, breast cancer, melanoma, pancreatic cancer, ovarian cancer, colonic cancer, cervical cancer, head and neck cancer, lung cancer, gastric cancer, endometrial cancer, bone and / or brain metastases, soft-tissue sarcoma, osteosarcoma, hepatoma, basal cell carcinoma, glioblastoma, angiosarcoma, T-cell lymphoma, and multiple myeloma.

[0030] In some embodiments, the cancer is prostate cancer.

[0031] In some embodiments, the cancer is breast cancer.

[0032] In some embodiments, the non-malignant condition is selected from the group consisting of non- malignant fibrotic disease involving lung, liver, heart, kidney, bone marrow, or skin, scleroderma, chronic graft-vs-host disease, liver fibrosis, interstitial lung disease. ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700

[0033] In some embodiments, the non-malignant disease is selected from the group consisting of pathological angiogenesis, proliferative retinopathy, wet macular degeneration, arteriovenous malformation, and cavernous hemangioma.

[0034] In some embodiments, the subject is resistant to monotherapy with an av integrin inhibitor.

[0035] In some embodiments, the subject is resistant to monotherapy with an a5 (e.g., a5p1 ) integrin inhibitor.

[0036] In some embodiments, the subject is resistant to monotherapy with a Src inhibitor.

[0037] In some embodiments, the bispecific antibody and the Src inhibitor are present within a single pharmaceutical composition.

[0038] In another aspect, the invention provides pharmaceutical compositions comprising (i) a bispecific antibody that specifically binds av integrin and a5 (e.g., a5p1 ) integrin and (ii) a Src inhibitor.

[0039] In some embodiments, the pharmaceutical composition further comprises a pharmaceutically acceptable carrier.

[0040] In another aspect, the invention provides combinations of (i) a bispecific antibody that specifically binds av integrin and a5 (e.g., a5p1 ) integrin and (ii) a Src inhibitor for use in a method of treating a subject having cancer or non-malignant condition or disease.

[0041] Other features and advantages of the invention are within the scope of the following detailed description, the drawings, and the claims.

[0042] Definitions

[0043] To facilitate the understanding of the invention, a number of terms are defined below. Terms defined herein have meanings as commonly understood by those of ordinary skill in the areas relevant to the invention.

[0044] As used herein, the terms “administration” and “administering” refer to providing or giving a subject one or more therapeutic agents including, for example, one or more bispecific antibodies that specifically bind av and a5 (e.g., a5p1 ) integrins and one or more Src inhibitors, or a pharmaceutical composition that includes these agents, by any effective route. Exemplary routes of administration are described herein and include systemic administration routes, such as intravenous injection and oral tablets, among others.

[0045] As used herein, the terms “bispecific antibody construct” and “bispecific antibody” refer to an antibody construct that comprises or consists of a protein or a complex of proteins having a sequence or sequences that specifically and simultaneously bind two molecular targets. For the bispecific antibody construct described herein, the first target is av integrin and the second target is a5 integrin (alone or when heterodimerized with pi ). The bispecific antibody can be further configured and modified as described herein (e.g., modified to include a detectable label). The term “construct” is used, in part, because the compositions can be constructed using genetic engineering and can be non-naturally occurring. Accordingly, any of the bispecific antibody constructs described herein, nucleic acids that encode them, related expression vectors, and host cells can be non-naturally occurring.

[0046] As is known in the art, naturally occurring immunoglobulins include a heavy chain and a light chain, each of which is further divided into a constant region and a variable region. Within each variable region, there are three hypervariable regions (HVRs), also known as “complementarity determining ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700 regions” or “CDRs,” interspersed within four “framework regions” or “FRs”. It is the CDRs that are primarily responsible for binding target molecules, which can be antigens. The sequences within the antibody that bind the target molecule comprise the “paratope.” The sequences within the target molecule that are bound comprise the “epitope.” The CDRs of each chain are typically referred to as CDR1 , CDR2, and CDR3, numbered sequentially from the N-terminus, and the variable regions of the heavy and light chains can be abbreviated as “VH” and “VL”, respectively. As the constructs described herein are bispecific, they will include two paratopes.

[0047] As used herein, the terms "effective amount," “therapeutically effective amount,” and a “sufficient amount” of a pharmaceutical composition described herein refer to amounts that can result in a beneficial or desired physiological change in a subject, such as a human subject. As such, an “effective amount” or synonym thereof depends upon the context in which it is being applied. In the context of treating cancer (e.g., prostate cancer and breast cancer), it is an amount of a pharmaceutical composition that achieves a treatment response, e.g., in the context of the presently claimed combinations, as compared to a response obtained without such treatment. The amount will vary depending upon various factors, such as the pharmaceutical formulation, the route of administration, the type of disease or disorder, the identity of the subject (e.g., age, sex, weight) or host being treated, and the like, but can nevertheless be determined by those skilled in the art. The amount needs not fully prevent or cure the disease or the condition but can rather provide a partial benefit, such as a delay of onset or an alleviation or an improvement of at least one symptom of the disease or the condition. The amount can be readily determined by those of ordinary skill in the art by methods that are known in the art. Dosage regimes can be adjusted to provide an optimum therapeutic response.

[0048] As used herein, the term “framework” or “FR” refers to variable domain residues other than HVR residues. The FR of a variable domain generally consists of four FR domains: FR1 , FR2, FR3, and FR4. Accordingly, the HVR and FR sequences generally appear in the following sequence in VH (or VL): FR1 - CDRH1 (CDRL1 )-FR2-CDRH2(CDRL2)-FR3-CDRH3(CDRL3)-FR4.

[0049] As used herein, the term “humanized antibody” refers to a chimeric antibody comprising amino acid residues from non-human HVRs and amino acid residues from human FRs. In certain embodiments, a humanized antibody will comprise substantially all of at least one, and typically two, variable domains, in which all or substantially all of the HVRs (e.g., CDRs) correspond to those of a non-human antibody, and all or substantially all of the FRs correspond to those of a human antibody. A humanized antibody optionally can comprise at least a portion of an antibody constant region derived from a human antibody. A ’’humanized form” of an antibody, e.g., a non-human antibody, refers to an antibody that has undergone humanization. Immunoglobulins generated in a non-human animal (e.g., a rodent or non-human primate) can be humanized and / or de-immunized according to standard methods. In some instances, the humanized and / or de-immunized antibody constructs will be chimeric, in that they will include sequences (e.g., variable domain sequences) found in the non-human immunoglobulins from which they were generated as well as sequences (e.g., constant domain sequences) found in human immunoglobulins. In other embodiments, the antibody constructs described herein can include CDRs and FRs from a fully human immunoglobulin. ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700

[0050] As used herein, the term “monotherapy” refers to the use of a single therapeutic agent to treat a disease or condition in a subject. In contrast, the term “combination therapy” refers to the use of at least two therapeutic agents in a treatment regimen.

[0051] “Percent (%) sequence identity” with respect to a reference polynucleotide or polypeptide (or peptide) sequence is defined as the percentage of nucleic acids or amino acids in a candidate sequence that are identical to the nucleic acids or amino acids in the reference polynucleotide or polypeptide (or peptide) sequence, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity. Alignment for purposes of determining percent nucleic acid or amino acid sequence identity can be achieved in various ways that are within the capabilities of those of skill in the art, for instance, using publicly available computer software such as BLAST, BLAST-2, or Megalign software. Those skilled in the art can determine appropriate parameters for aligning sequences, including any algorithms needed to achieve maximal alignment over the full length of the sequences being compared. For example, percent sequence identity values can be generated using the sequence comparison computer program BLAST. As an illustration, the percent sequence identity of a given nucleic acid or amino acid sequence, A, to, with, or against a given nucleic acid or amino acid sequence, B, (which can alternatively be phrased as a given nucleic acid or amino acid sequence, A that has a certain percent sequence identity to, with, or against a given nucleic acid or amino acid sequence, B) is calculated as follows:

[0052] 100 multiplied by (the fraction X / Y) where X is the number of nucleotides or amino acids scored as identical matches by a sequence alignment program (e.g., BLAST) in that program’s alignment of A and B, and where Y is the total number of nucleic acids in B. It will be appreciated that where the length of nucleic acid or amino acid sequence A is not equal to the length of nucleic acid or amino acid sequence B, the percent sequence identity of A to B will not equal the percent sequence identity of B to A.

[0053] As used herein, the term “pharmaceutically acceptable” is used to describe compounds, materials, compositions, and / or dosage forms that are suitable for contact with the tissues of a subject, such as a mammal (e.g., a human) without excessive toxicity, irritation, allergic response, and other problems or complications commensurate with a reasonable benefit / risk ratio.

[0054] As used herein, the term “single-chain Fv” or “scFv” refers to a binding entity in which the variable regions of VH and VL of a conventional antibody have been engineered to form one chain. A linker sequence is typically inserted between the VH and VL regions of an scFv to facilitate proper folding and creation of an active paratope.

[0055] As used herein, the term “Src inhibitor” refers to an agent that inhibits or decreases signaling through the Src tyrosine kinase. In some embodiments, the Src inhibitor is dasatinib, dasatinib monohydrate, bafetinib, bosutinib, danusertib, ponatinib, repotrectinib, saracatinib, tirbanibulin, AP24149, AP24163, AZD0424, CCT196969, DC2036, NVP-BHG712, PD173955, PP1 , PP2, PP121 , SGX393, WH-4-023, XL228, or 3,4-Methylenedioxy-p-nitrostyrene. In one embodiment, the Src inhibitor is dasatinib.

[0056] As used herein, the terms “subject” and “patient” refer to an organism that receives treatment for a particular disease or condition as described herein (such as cancer, e.g., prostate cancer and breast ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700 cancer). Examples of subjects and patients include mammals, such as humans, receiving treatment for a disease or condition described herein.

[0057] As used herein, the terms “treat,” “treatment,” and “treating” refer to an approach for obtaining beneficial or desired results, e.g., clinical results. Beneficial or desired clinical results include, but are not limited to, alleviation of one or more symptoms or conditions, diminishment of extent of disease or condition, stabilized (i.e. , not worsening) state of disease, disorder, or condition, preventing spread of disease or condition, delay or slowing the progression of disease or condition, amelioration or palliation of the disease or condition, and remission (whether partial or total), whether detectable or undetectable. “Ameliorating” or “palliating” a disease or condition means that the extent and / or undesirable clinical manifestations of the disease, disorder, or condition are lessened and / or time course of the progression is slowed or lengthened, as compared to the extent or time course in the absence of treatment. “Treatment” can also mean prolonging survival as compared to expected survival if not receiving treatment. Those in need of treatment include those already with the condition or disorder, as well as those prone to have the condition or disorder or those in which the condition or disorder is to be prevented.

[0058] As used herein, the term "therapeutic treatment" refers to a treatment administered to a subject who displays symptoms or signs of a disease or condition, and the therapeutic treatment is administered to the subject for the purpose of diminishing or eliminating the symptoms or the signs of the disease or the condition.

[0059] Throughout the description and claims of this specification, the words “comprise” and “contain” and variations of them mean “including but not limited to,” and they are not intended to (and do not) exclude other moieties, additives, components, integers, or steps. Throughout the description and claims of this specification, the singular encompasses the plural unless the context otherwise requires. In particular, where the indefinite article is used, the specification is to be understood as contemplating plurality as well as singularity, unless the context requires otherwise.

[0060] Brief Description of the Drawings

[0061] FIG. 1 is a set of western blot images showing the effects of BsAba5p1 / av on the levels of phosphorylated Src at Tyr 416, total Src, and p-actin in prostate cancer cell lines (PC-3, DU-145, VCAP, C4-2B, and LnCAP) and triple negative breast cancer (TNBC) cell lines (BT-20, MDA-MB-231 , and MDA- MB-468). Con: vehicle control; BiSp: BsAba5p1 / av

[0062] FIG. 2 is a set of western blot images showing the effects of dasatinib, BsAba5p1 / av, PF-431396, and combinations thereof on the expression levels of active p-catenin, YAP, and lamin A / C in cytosolic and nuclear fractions from DU-145 cells (DU-145 Parental; FIG. 2A) and DU-145 cells in which the PTEN tumor suppressor had been knocked out (DU-145 PTEN-mt; FIG. 2B). BsAb: BsAba5p1 / av.

[0063] FIG. 3 is a set of flow cytometry graphs showing the effects of vehicle alone, BsAba5p1 / av alone, dasatinib alone, or the combination of BsAba5p1 / av and dasatinib on apoptotic cell death as annotated by annexin V-PI staining in DU-145 parental and DU-145 PTEN-mt cell lines. Vehicle: vehicle alone; BsAb: BsAba5p1 / av alone; Dasatinib: dasatinib alone; Combination: BsAba5p1 / av and dasatinib.

[0064] FIG. 4 is a set of western blot images showing the effects of BsAba5p1 / av alone, dasatinib alone, and the combination of BsAba5p1 / av and dasatinib on the expression levels of cleaved PARP, PTEN, and GAPDH in DU-145 parental and DU-145 PTEN-mt cell lines. BsAb: BsAba5p1 / av. ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700

[0065] FIG. 5A and FIG. 5B are a set of histograms (FIG. 5A) and associated bar plot quantifications (FIG. 5B) showing the effects of dasatinib, monospecific antibody against a5pi integrin, monospecific antibody against av integrin, BsAba5p1 / av, and combinations thereof on the percent cell viability as assessed by annexin V-PI staining in DU-145 parental and DU-145 PTEN-mt cell lines. MsAb A5B1 : monospecific antibody against a5pi integrin; MsAb Av: monospecific antibody against av integrin; BsAb A5B1 / Av: BsAba5p1 / av.

[0066] FIG. 6 is a set of flow cytometry graphs showing the effects of vehicle alone, combination of dasatinib and BsAba5p1 / av, and combination of dasatinib and PF-431396 on apoptotic cell death as annotated by annexin V-PI staining in DU-145 parental and DU-145 PTEN-mt cell lines.

[0067] FIG. 7 is a set of western blot images showing the expression levels of phosphorylated Src at Tyr 416, total Src, and GAPDH in DU-145 cells (DU-145-WT) and DU-145 cells in which Src had been genetically knocked out (DU-145-SRC-KD).

[0068] FIG. 8 is a set of flow cytometry graphs showing the effects of vehicle alone, BsAba5p1 / av alone, and combination of dasatinib and BsAba5p1 / av on apoptotic cell death as annotated by annexin V-PI staining in DU-145-WT and DU-145-SRC-KD cells. Control: vehicle alone.

[0069] FIG. 9 is a set of flow cytometry graphs showing the effects of vehicle alone, BsAba5p1 / av alone, dasatinib alone, PF-431396 alone, combination of dasatinib and BsAba5p1 / av, and combination of dasatinib and PF-431396 on apoptotic cell death as annotated by annexin V-PI staining in PC-3, C42B, MDA-MB-231 , and BT-20 cell lines.

[0070] Detailed Description of the Invention

[0071] The invention provides methods of treating a subject having cancer or a non-malignant condition or disease in which av and / or a5 (e.g., a5p1 ) integrins are expressed or implicated in pathogenesis or progression. The methods comprise administration of therapeutically effective amounts of (i) a bispecific antibody that specifically binds av and a5 (e.g., a5p1 ) integrins and (ii) a Src inhibitor. Also provided are compositions and kits comprising the bispecific antibody and a Src inhibitor and combinations of the bispecific antibody and a Src inhibitor for use in a method of treating a subject having cancer or non- malignant condition or disease. The invention is based, in part, on the inventor’s discovery that the combination of the bispecific antibody and a Src inhibitor has synergistic effects on apoptotic cell death in cancer cells. The methods, bispecific antibodies, Src inhibitors, compositions, and combinations are described in more detail below.

[0072] Bispecific antibody targeting the av and a5 integrins

[0073] The av and a5 integrins are implicated in a range of cancer-specific behaviors, as well as many non-cancer pathologies that are related to fibrosis or angiogenesis. A bispecific antibody that can be used in the methods, compositions, and combinations described herein simultaneously targets av and a5 (e.g., a5p1 ) integrins. Such bispecific antibodies can be significantly superior to combinations of individual av and a5 monospecific antibodies. The full structure of a bispecific antibody that can be used in the invention, ITGA5B1 Xav, is described in U.S. Patent No. 11 ,339,221 , the entire content of which is incorporated herein by reference. Additional antibodies that can be used in the present invention, as well ATTORNEY DOCKET NO. 00398-571 WO2

[0074] TMC 700 as sources of additional antibodies against the present targets that can be used as sources for sequences in generating bispecific antibodies for use in the present methods, are also described in U.S. Patent No. 11 ,339,221 .

[0075] The bispecific antibody used in the methods, compositions, and combinations described herein can be conjugated to a chemotherapeutic or cytotoxic agent and specifically bind av and a5 (e.g., a5p1 ) integrins with or without inhibiting the activity of either or both targets. The bispecific antibody can also be unconjugated to a chemotherapeutic or cytotoxic agent, specifically bind av and a5 (e.g., a5p1 ) integrins, and inhibit their biological activities to a clinically beneficial extent.

[0076] Accordingly, in some embodiments, the bispecific antibody for use in methods, compositions, and combinations described herein can include (i) a first paratope that specifically binds an aV integrin; and (ii) a second paratope that specifically binds an a5 (e.g., a5p1 ) integrin. The epitope on the a5 integrin can be wholly contained within that protein. Alternatively, the epitope can reside at least partially within the p1 integrin that, in living systems, forms a heterodimer with a5 integrin. Any type of bispecific antibody can be used. Non-limiting examples are shown in Fig. 11 of U.S. Patent No. 11 ,339,221 . More information about bispecific antibodies can be found in a review by Brinkmann et al, mAbs., 9(2) :182-212, 2017. In particular, Fig. 2 of the review illustrates many bispecific antibody formats that have been used in the art and that can be utilized for the bispecific antibody construct that can be used for the methods, compositions, and combinations described herein.

[0077] In some embodiments, the bispecific antibody comprises an antigen binding fragment (Fab) portion that contains the first paratope and a single-chain variable fragment (scFv) portion that contains the second paratope. Either paratope can be from an anti-av antibody, such as abituzumab, with the other paratope being from an anti-a5 antibody, such as volociximab. The bispecific construct can be bivalent, trivalent, tetravalent, or of a higher valency. Additional sources of antibodies for use in construction of antibodies used in the invention are provided in U.S. Patent No. 11 ,339,221 . Furthermore, newly generated antibodies having the requisite specificity and / or activity can be used. In some embodiments, the antibody is ITGA5B1xAV, as described in U.S. Patent No. 11 ,339,221 .

[0078] The bispecific antibody construct that can be used in the methods, compositions, and combinations described herein can include, for example, one or more of the following CDR amino acid sequences:

[0079] (i) within a light chain variable domain of the first paratope: RASQDISNYLA (P1 VLCDR1 ; SEQ ID NO: 1 ); YTSKIHS (P1 VICDR2; SEQ ID NO: 2); QQGNTFPYT (P1 VLCDR3; SEQ ID NO: 3); or an amino acid sequence at least 80% identical to any of SEQ ID NOs: 1 -3 or wherein there are zero, one, or two amino acid deletions, additions, or substitutions in any of them;

[0080] (ii) within a heavy chain variable domain of the first paratope: SFWMH (P1 VHCDR1 ; SEQ ID NO: 4); YINPRSGYTEYNEIFRD (P1 VHCDR2; SEQ ID NO: 5); FLGRGAMDY (P1 VHCDR3; SEQ ID NO: 6); or an amino acid sequence at least 80% identical to any of SEQ ID NOs: 4-6, or wherein there are zero, one, or two amino acid deletions, additions, or substitutions in any of them;

[0081] (iii) within a light chain variable domain of the second paratope, TASSSVSSNYLH (P2VLCDR1 ; SEQ ID NO: 7); STSNLAS (P2VLCDR2; SEQ ID NO: 8); HQYLRSPPT (P2VLCDR3; SEQ ID NO: 9); or an amino acid sequence at least 80% identical to any of SEQ ID NOs: 7-9, or wherein there are zero, one, or two amino acid deletions, additions, or substitutions in any of them; or ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700

[0082] (iv) within a heavy chain variable domain of the second paratope, GFSLTDYGVH (P2VHCDR1 ; SEQ ID NO: 10); VIWSDGSSTYNSALKS (P2VHCDR33; SEQ ID NO: 11 ); HGTYYGMTTTGDALDY (P2VHCDR3; SEQ ID NO: 12); or an amino acid sequence at least 80% identical to any of SEQ ID NOs: 10-12, or wherein there are zero, one, or two amino acid deletions, additions, or substitutions in any of them.

[0083] Additionally, the bispecific antibody construct can include one or more of the following FR amino acid sequences:

[0084] (i) within a light chain variable domain contributing to the first paratope: DIQMTQSPSSLSASVGDRVTITC (P1 VLFR1 ; SEQ ID NO: 13); WYQQKPGKAPKLLIY (P1 VLFR2; SEQ ID NO: 14); GVPSRFSGSGSGTDYTFTISSLQPEDIATYYC (P1 VLFR3; SEQ ID NO: 15); FGQGTKVEIK (P1 VLFR4; SEQ ID NO: 16); or an amino acid sequence at least 80% identical to any of SEQ ID NOs: 13- 16 (e.g., there may be zero, one, or two amino acid deletions, insertions, or substitutions in any of them);

[0085] (ii) within a heavy chain variable domain contributing to the first paratope, QVQLQQSGAELAEPGASVKMSCKASGYTFS (P1 VHFR1 ; SEQ ID NO: 17); WVRQAPGQGLEWIG (P1 VHFR2; SEQ ID NO: 18); KATMTTDTSTSTAYMELSSLRSEDTAVYYCAS (P1 VHFR3; SEQ ID NO: 19); WGQGTTVTVSS (P1 VHFR4; SEQ ID NO: 20); or an amino acid sequence at least 80% identical to SEQ ID NOs: 17-20;

[0086] (Hi) within a light chain variable domain contributing to the second paratope, QIVLTQSPAIMSASLGERVTMTC (P2VLFR1 ; SEQ ID NO: 21 ); WYQQKPGSAPNLWIY (P2VLFR2; SEQ ID NO: 22); GVPARFSGSGSGTSYSLTISSMEAEDAATYYC (P2VLFR3; SEQ ID NO: 23); FGGGTKLEIK (P2VLFR4; SEQ ID NO: 24); or an amino acid sequence at least 80% identical to any of SEQ ID NOs: 21 - 24 (e.g., there may be zero, one, or two amino acid deletion, additions, or substitutions in any of them); or

[0087] (iv) within a heavy chain variable domain contributing to the second paratope, QVQLKESGPGLVAPSQSLSITCTIS (P2VHFR1 ; SEQ ID NO: 25); WVRQPPGKGLEWLV (P2VHFR2; SEQ ID NO: 26); RMTIRKDNSKSQVFLIMNSLQTDDSAMYYCAR (P2VHFR3; SEQ ID NO: 27); WGQGTSVTVSS (P2VHFR4; SEQ ID NO: 28); or an amino acid sequence at least 80% identical to any of SEQ ID NOs: 25-28 (e.g., there may be zero, one, or two amino acid deletions, additions, or substitutions in any of them).

[0088] In some embodiments, the bispecific antibody that can be used in the methods, compositions, and combinations described herein can include a second paratope that binds and may inhibit the activity (e.g., at least partially neutralize the activity) of a5 integrin or a5pi integrin. Such a bispecific antibody can include one or more of the following CDRs:

[0089] (i) within a light chain variable domain of the second paratope, RASQSVSSYLA (P1 VLCDR1 ; SEQ ID NO: 29); DASNRAT (P1 VLCDR2; SEQ ID NO: 30); QQRSNWPLT (P1 VLCDR3; SEQ ID NO: 31); or an amino acid sequence at least 80% identical to any of SEQ ID NOs: 29-31 , or where there are zero, one, or two amino acid deletions, additions, or substitutions in any of them; and

[0090] (ii) within a heavy chain variable domain of the second paratope, SSSYWG (P1 VHCDR1 ; SEQ ID NO: 32); SIYYSGRNYNNPSLKS (P1 VHCDR2; SEQ ID NO: 33); and HYYGSGSSYYYYDLD

[0091] (P1 VHCDR3; SEQ ID NO: 34); or an amino acid sequence at least 80% identical to any of SEQ ID NOs: 32-34, or where there are zero, one, or two amino acid deletions, additions, or substitutions in any of them. ATTORNEY DOCKET NO. 00398-571 WO2

[0092] TMC 700

[0093] In some embodiments, the bispecific antibody that can be used in the methods, compositions, and combinations described herein can include a second paratope that binds and may inhibit at least partially neutralize the activity of a5 (e.g., a5p1 ) integrin. Such a bispecific antibody can include one or more of the following CDRs:

[0094] (i) within a light chain variable domain of the second paratope: SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31 or an amino acid sequence at least 80% identical to SEQ ID NOs: 29-31 (CDRL1 -3), or where there are zero, one, or two amino acid deletions, additions, or substitutions in any of them; and

[0095] (ii) within a heavy chain variable domain of the second paratope: SYAMH (CDRH1 ; SEQ ID NO: 35); VISFDGSNKNYADSVKG (CDRH2; SEQ ID NO: 36); and EYWGTYYYGMDV (CDRH3; SEQ ID NO:

[0096] 37); or an amino acid sequence at least 80% identical to any of SEQ ID NOs: 35-37, or where there are zero, one, or two amino acid deletions, additions, or substitutions in any of them.

[0097] In some embodiments, the bispecific antibody that can be used in the methods, compositions, and combinations described herein can include a second paratope that binds and may inhibit at least partially neutralize the activity of a5 (e.g., a5p1 ) integrin. Such bispecific antibody can include one or more of the following CDRs:

[0098] (i) within a light chain variable domain of the second paratope: SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31 or an amino acid sequence at least 80% identical to SEQ ID NOs: 29-31 (CDRL1 -3), or an amino acid sequence at least 80% identical to any of SEQ ID NOs: 29-31 , or where there are zero, one, or two amino acid deletions, additions, or substitutions in any of them; and

[0099] (ii) within a heavy chain variable domain of the second paratope: STYAMH (CDRH1 ; SEQ ID NO:

[0100] 38); VISYDGSNKYYADSVKG (CDRH2; SEQ ID NO: 39); and RESPPIYYYYGMDV (CDRH3; SEQ ID NO: 40); or an amino acid sequence at least 80% identical to any of SEQ ID NOs: 38-40.

[0101] CDRs, FRs, and biologically active fragments or other variants thereof described above are from previously generated or newly generated immunoglobulins. For example, the CDRs within a paratope that binds av integrin can include those described in U.S. Patent No. 8,562,986, the entire content of which is hereby incorporated by reference herein. The CDRs within a paratope that binds a5 or a5p1 integrin can include those described in U.S. Patent No. 7,276,589 and U.S. Patent No. 8,039,596, the contents of which are incorporated herein by reference.

[0102] A bispecific antibody construct described herein can also have one or more of the following characteristics: it can further include an Fc region, such as one derived from a human IgA or human IgG Fc region; it can be a chimera (with one or all variable domain sequences derived from a non-human animal, and the Fc region derived from a human); it can be fully human, humanized, or de-immunized; it can be divalent, trivalent, or tetravalent; and / or it can include a detectable marker. For example, an antibody construct can include an Fc portion and be chimeric; can be human and tetravalent; can be humanized and include a detectable marker; or have any other compatible combination of the features just mentioned or set out below.

[0103] The bispecific antibody construct for use in the methods, compositions, and combinations described herein can comprise a first polypeptide comprising a first light chain variable domain linked to a light chain constant region, and a second polypeptide chain comprising, in order from amino to carboxy terminus, a first antibody heavy chain variable domain linked to a heavy chain constant region linked to an scFv, where the scFv contains, in either order, a second heavy chain variable domain that is different ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700 from the first heavy chain variable domain and is linked to a second light chain variable domain that is different from the first light chain variable domain. The first light and heavy chain variable domains form a paratope that binds to either av integrin or a5 (e.g., a5p1) integrin. The second light and heavy chain variable domains form a second paratope that binds to the other integrin. An example of this construct with signal sequences still attached to the respective amino termini is shown in Figs. 12A and 12B of U.S. Patent No 11 ,339, 221 , and described therein. As is known in the art, the signal sequences are cleaved when the antibody is secreted from the host cell in which it is generated but are included to facilitate production (see U.S. Patent No. 11 ,339,221 , for details).

[0104] In some embodiments, the antibody constructs described herein comprise linker sequences. In some embodiments, a single chain antibody described herein comprises a linker sequence between the heavy chain variable sequences and the light chain variable region sequences. In some embodiments, an IgG heavy chain sequence of a bispecific antibody described herein is linked to a single chain antibody described herein by a linker sequence. In some embodiments, a linker used herein is a glycine rich linker. In some embodiments, the glycine-rich linker comprises GGGG (SEQ ID NO: 43) or GGGGS (SEQ ID NO: 44) sequences, for example, 1 , 2, 3, 4, or more instances thereof (or combinations thereof). In some embodiments, the linker comprises the sequence GGGGSGGGGS (SEQ ID NO: 45). In some embodiments, the sequence of SEQ ID NO: 43, 44, 45, or 46 is used to link an IgG heavy chain sequence of a bispecific antibody described herein to a single chain antibody described herein. In some embodiments, the linker comprises the sequence GGGGSGGGGSGGGGS (SEQ ID NO: 46). In some embodiments, the sequence of SEQ ID NO: 43, 44, 45, or 46 is used to link the heavy chain variable region sequences of a single chain antibody to the light chain variable region sequences of the single chain antibody. In some embodiments, the linker comprises 1 , 2, 3, 4, 5, 6, 7, 8, or more deletions, insertions, or substitutions of linker sequences as set forth herein.

[0105] The bispecific antibody construct for use in the methods, compositions, and combinations described herein can comprise an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identical to the amino acid sequence set forth in SEQ ID NO: 41 . In certain instances, the bispecific antibody construct comprises an amino acid sequence set forth in SEQ ID NO: 41 except having 1 to 20 (e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, or 20) amino acid substitutions (e.g., conservative amino acid substitutions). In certain instances, the substitutions are in the framework, constant, and / or linker region. In certain instances, the substitutions are in one or more of the CDRs.

[0106] In some embodiments, the bispecific antibody construct for use in the methods, compositions, and combinations described herein can comprise an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identical to the amino acid sequence set forth in SEQ ID NO: 42. In certain instances, the bispecific antibody construct comprises an amino acid sequence set forth in SEQ ID NO: 42 except having 1 to 20 (e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, or 20) amino acid substitutions (e.g., conservative amino acid substitutions). In certain instances, the substitutions are in the framework and / or constant region. In certain instances, the substitutions are in one or more of the CDRs.

[0107] In some embodiments, the bispecific antibody construct for use in the methods, compositions, and combinations described herein can comprise: (i) an amino acid sequence that is at least 70%, at ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700 least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identical to the amino acid sequence set forth in SEQ ID NO: 41 ; and (ii) an amino acid sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% identical to the amino acid sequence set forth in SEQ ID NO: 42. In certain instances, the disclosure features a bispecific construct comprising an amino acid sequence set forth in SEQ ID NO: 41 and / or 42 except having 1 to 20 (e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, or 20) amino acid substitutions (e.g., conservative amino acid substitutions). In certain instances, the substitutions are in the framework, constant, and / or linker region. In certain instances, the substitutions are in one or more of the CDRs.

[0108] Nucleic acid molecules, expression vectors, host cells, and expression vectors that can be used to generate the bispecific antibodies described herein are known in the art and described, e.g., in U.S. Patent No. 11 ,339,221 .

[0109] The bispecific antibody described herein is capable of binding av integrin and a5 (e.g., a5p1 ) integrin with sufficient affinity to be useful as diagnostic and / or therapeutic agents. In some embodiments, the extent of binding of an antibody construct of the invention to an unrelated protein (i.e. , a protein other than av integrin or a5 (e.g., a5p1 ) integrin) is less than about 10% of the binding of the antibody to av integrin or a5 (e.g., a5p1 ) integrin as measured, e.g., by a radioimmunoassay. In certain embodiments, an antibody that binds to av integrin or a5 (e.g., a5p1 ) integrin has a dissociation constant (KD) of < 1 pM, < 100 nM, < 10 nM, < 1 nM, < 0.1 nM, < 0.01 nM, or < 0.001 nM (e.g. 10-8M or less, e.g. from 10-8M to 10-13M, e.g., from 10-9M to 10-13M), where KD is determined using a surface plasmon resonance technique (e.g., with a Biacore™ instrument) in which the antigen is immobilized, the antibody serves as analyte, and the following conditions are used: 37 °C, 0.05% Tween® 20 polyethylene glycol sorbitan monolaurate, 20 mM N-(2-acetamido)-2-aminoethanesulfonic acid (ACES buffer), 150 mM NaCI, pH 7.4. In certain embodiments, an anti-av integrin or anti-a5 (e.g., a5p1 ) integrin antibody binds to an epitope of av integrin or a5 (e.g., a5p1 ) integrin that is conserved among those integrins from different species (e.g., human and non-human primate or human and murine).

[0110] The bispecific antibody constructs described herein can also include one or more additional moieties. For example, the constructs can include a detectable label and / or a moiety that extends the construct's circulating half-life (e.g., modified human serum albumin). A “detectable label” is a composition detectable by spectroscopic, photochemical, biochemical, immunochemical, chemical, or other physical means. For example, a detectable label within a bispecific antibody construct of the present invention can be a fluorescent dye, an electron-dense reagent, a radioisotope, an enzyme (e.g., an enzyme commonly used in an ELISA), biotin, digoxigenin, a hapten, or any other protein or non-protein entity that can be made detectable. The radioisotope can be, for example,3H,14C,32P,35S, or125l. In some cases, radioisotopes can be employed not only because they are detectable, but also because they can be toxic to cells in their vicinity.

[0111] Detectable labels can be incorporated into the bispecific antibody constructs at a number of positions, and any method known in the art for conjugating the construct to the label can be employed. For example, one could use a method described by Hunter et al. {Nature, 144:945, 1962); David et al. {Biochemistry, 13:1014, 1974); Pain et al. {J. Immunol. Meth., 40:219, 1981 ); or Nygren {J. Histochem. and Cytochem., 30:407, 1982). The lifetime of radiolabeled antibody constructs can be extended by adding a substance that stabilizes the radiolabeled construct by, for example, protecting it from ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700 degradation. Any substance or combination of substances that stabilizes the construct can be used, including those disclosed in U.S. Patent No. 5,961 ,955.

[0112] Src inhibitors

[0113] Src is a non-receptor tyrosine kinase the activation of which is typically downstream of integrin activation. As a regulator of tumor cell survival, angiogenesis, proliferation, and invasion, Src has been identified as a target for cancer therapies. Src inhibitors have variable inhibitory effects on Abl, another tyrosine kinase that is implicated in cancer cell invasion.

[0114] The Src inhibitor that can be used in the methods, compositions, and combinations described herein can be a dual Src-Abl inhibitor. In some embodiments, the Src inhibitor is selected from the group consisting of dasatinib, dasatinib monohydrate, bafetinib, bosutinib, danusertib, ponatinib, repotrectinib, saracatinib, tirbanibulin, AP24149, AP24163, AZD0424, CCT196969, DC2036, NVP-BHG712, PD173955, PP1 , PP2, PP121 , SGX393, WH-4-023, XL228, and 3,4-Methylenedioxy-p-nitrostyrene. In some embodiments, the Src inhibitor is dasatinib, which has the chemical formula C22H26CIN7O2S and is a dual Src-Abl inhibitor.

[0115] Pharmaceutical compositions

[0116] The bispecific antibody and Src inhibitor described herein can be formulated as pharmaceutical compositions for administration to a subject, such as a human subject with cancer (e.g., prostate cancer or breast cancer; also see above) or a non-malignant disease or condition in which av and / or a5 (e.g., a5p1 ) integrin is expressed or implicated in pathogenesis or progression in a biologically compatible form suitable for administration in vivo.

[0117] In some embodiments, the pharmaceutical composition containing the bispecific antibody binding to av and a5 (e.g., a5p1 ) integrins and a Src inhibitor include a pharmaceutically acceptable carrier, which does not produce significantly adverse, allergic, or other untoward reactions that can outweigh the benefit of administration, whether for research, prophylactic, and / or therapeutic treatments. Exemplary pharmaceutically acceptable carriers and formulations are disclosed in Remington's Pharmaceutical Sciences, 18th Ed., Mack Printing Company, 1990, which is incorporated by reference herein for its teachings regarding the same.

[0118] Moreover, formulations can be prepared to meet sterility, pyrogenicity, general safety, and purity standards as required by the United States FDA's Division of Biological Standards and Quality Control and / or other relevant U.S. and foreign regulatory agencies.

[0119] Exemplary, generally used, pharmaceutically acceptable carriers can optionally include bulking agents or fillers, solvents or co-solvents, dispersion media, coatings, surfactants, antioxidants (e.g., ascorbic acid, methionine, and vitamin E), preservatives, isotonic agents, absorption delaying agents, salts, stabilizers, buffering agents, chelating agents (e.g., EDTA), gels, binders, disintegration agents, and / or lubricants.

[0120] Exemplary buffering agents include citrate buffers, succinate buffers, tartrate buffers, fumarate buffers, gluconate buffers, oxalate buffers, lactate buffers, acetate buffers, phosphate buffers, histidine buffers, and / or trimethylamine salts. ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700

[0121] Exemplary preservatives include phenol, benzyl alcohol, meta-cresol, methylparaben, propyl paraben, octadecyldimethylbenzyl ammonium chloride, benzalkonium halides, hexamethonium chloride, alkyl parabens (such as methyl or propyl paraben), catechol, resorcinol, cyclohexanol, and / or 3-pentanol.

[0122] Exemplary isotonic agents include polyhydric sugar alcohols comprising, but not limited to, trihydric or higher sugar alcohols, (e.g., glycerin, erythritol, arabitol, xylitol, sorbitol, and / or mannitol).

[0123] Exemplary stabilizers include organic sugars, polyhydric sugar alcohols, polyethylene glycol, sulfur-containing reducing agents, amino acids, low molecular weight polypeptides, proteins, immunoglobulins, hydrophilic polymers, and / or polysaccharides.

[0124] Methods of treatment

[0125] Patient selection

[0126] Subjects that can be treated using the methods, compositions, or combinations described herein are subjects, such as human subjects, having cancer (e.g., prostate cancer or breast cancer) in which either av or a5 (e.g., a5p1 ) integrin is implicated in the pathogenesis or progression of the disease. Examples of cancers that can be treated include, e.g., prostate, kidney, breast, melanoma, pancreatic, ovarian, colonic, cervical, head and neck, lung, gastric, endometrial, bone and brain metastases, soft- tissue sarcoma, osteosarcoma, hepatoma, basal cell carcinoma, glioblastoma, angiosarcoma, T-cell lymphoma, and multiple myeloma.

[0127] The methods, compositions, and combinations described herein can also be used to treat a non- malignant disease or condition in subjects, such as human subjects, in which either av or a5 (e.g., a5p1) integrin is expressed and implicated in the pathogenesis or progression of the disease. Such diseases include, e.g., non-malignant fibrotic diseases involving lung, liver, heart, kidney, bone marrow, or skin, such as scleroderma, chronic graft-vs-host disease, liver fibrosis, and interstitial lung disease. The general category of non-malignant diseases treatable with the present antibodies also includes, e.g., conditions involving pathological angiogenesis, such as proliferative retinopathy, wet macular degeneration, arteriovenous malformations, and cavernous hemangiomas. The method of treatment can include administering a therapeutically effective amount of a bispecific antibody construct described herein to a patient in identified need thereof.

[0128] The methods, compositions, and combinations described herein can be used to treat cancer or a non-malignant disease or condition as described herein. In some embodiments, the methods, compositions, and combinations can be used with subjects who do not respond to monotherapy with integrin-targeting agents (e.g., anti-av antibodies, anti-a5 (e.g., a5p1 ) antibodies) or with Src inhibitors. In some embodiments, the subjects do not respond to an initial or repeat treatment with anti-av antibodies. In some embodiments, the subjects do not respond to an initial or repeat treatment with anti-a5 (e.g., a5p1 ) antibodies. In some embodiments, the subjects do not respond to an initial or repeat treatment with Src inhibitors. In some embodiments, the subjects have developed resistance to monotherapy with integrin-targeting agents (e.g., anti-av antibodies, anti-a5 (e.g., a5p1 ) antibodies) or with Src inhibitors. In some embodiments, the subjects have developed resistance to treatment with anti-av antibodies. In some embodiments, the subjects have developed resistance to anti-a5 (e.g., a5p1 ) antibodies. In some embodiments, the subjects have developed resistance to Src inhibitors. ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700

[0129] Routes of administration

[0130] The bispecific antibodies, Src inhibitors, compositions, and combinations described herein can be administered to subjects with cancer (e.g., prostate cancer or breast cancer; also see above) or a non- malignant disease or condition as described elsewhere herein by any of a variety of routes selected to be appropriate by those of skill in the art. Thus, for example, the bispecific antibody, Src inhibitor, compositions comprising the bispecific antibody and Src inhibitor, or combinations of the bispecific antibody and Src inhibitor can be administered intravenously, intradermally, intraarterially, intraperitoneally, intralesionally, intracranially, intraarticularly, intraprostaticly, intrapleurally, intratracheally, intranasally, intravitreally, intravaginally, intrarectally, topically, intratumorally, intramuscularly, intravesicularly, intrapericardially, intraumbilically, intraocularly, mucosally, nasally, orally, subcutaneously, or subconjunctivally. In some embodiments, the bispecific antibody described herein is administered to a subject intravenously. In some embodiments, the Src inhibitor described herein is administered to a subject orally (e.g., in the form of a tablet or a capsule, or in liquid form).

[0131] The most suitable route for administration in any given case will depend on the particular therapeutic agent administered, the subject, pharmaceutical formulation methods, administration methods (e.g., administration time and administration route), the subject’s age, body weight, sex, severity of the cancer being treated, the subject’s diet, and the subject’s excretion rate. Multiple routes of administration can be used to treat a single subject, e.g., intravenous injection and oral tablets. Multiple routes of administration can be used to treat a single subject at one time, or the subject can receive treatment via one route of administration first and receive treatment via another route of administration during a second appointment, e.g., 1 week later, 2 weeks later, 1 month later, 6 months later, or 1 year later.

[0132] The bispecific antibodies and Src inhibitors described herein can be co-administered to a subject. In some embodiments, the bispecific antibody and Src inhibitor are administered to the subject at the same time. In some embodiments, the bispecific antibody and Src inhibitor are administered to the subject sequentially. In some embodiments, the bispecific antibody is administered to the subject before the Src inhibitor is administered to the subject. In some embodiments, the bispecific antibody is administered to the subject after the Src inhibitor is administered to the subject.

[0133] Dosing

[0134] For administration, effective amounts and therapeutically effective amounts (also referred to herein as doses) can be initially estimated based on results from in vitro assays or animal model studies. For example, a dose can be formulated in animal models to achieve a circulating concentration range that includes the IC50 as determined in cell culture. Such information can be used to determine useful doses in subjects of interest more accurately. Clinical studies can further be carried out to determine dosages.

[0135] The actual dose amount administered to a particular subject can be determined by a physician, a veterinarian, or a researcher, taking into account parameters such as physical and physiological factors including body weight, severity of condition, type of disease, previous or concurrent therapeutic interventions, idiopathy of the subject, and / or route of administration.

[0136] In some embodiments, the bispecific antibody described herein is administered to a subject in a dose of from about 0.1 mg / kg to 50 mg / kg, 0.2 mg / kg to 25 mg / kg, 0.4 mg / kg to 20 mg / kg, or 0.5 mg / kg to about 15 mg / kg (e.g., a dose of about 0.5 mg / kg, 0.6 mg / kg, 0.7 mg / kg, 0.8 mg / kg, 0.9 mg / kg, 1 mg / kg, 2 ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700 mg / kg, 3 mg / kg, 4 mg / kg, 5 mg / kg, 6 mg / kg, 7 mg / kg, 8 mg / kg, 9 mg / kg, 10 mg / kg, 11 mg / kg, 12 mg / kg,

[0137] 13 mg / kg, 14 mg / kg, or 15 mg / kg, or within a range between any of these amounts).

[0138] In some embodiments, the Src inhibitor described herein is administered to a subject in a dose of from about 1 mg to 500 mg, 5 mg to 300 mg, 10 mg to 250 mg, or 15 mg to 200 mg, or 20 mg to 140 mg (e.g., a dose of about 20 mg, 30 mg, 40 mg, 50 mg, 60 mg, 70 mg, 80 mg, 90 mg, 100 mg, 110 mg, 120 mg, 130 mg, or 140 mg, or within a range between any of these amounts).

[0139] Therapeutically effective amounts can be achieved by administering single or multiple doses during the course of a treatment regimen (i.e. , days, weeks, months, etc.). Sequences ATTORNEY DOCKET NO. 00398-571 WO2

[0140] TMC 700

[0141] Examples

[0142] The following examples are put forth so as to provide those of ordinary skill in the art with a description of how the compositions and methods described herein can be used and evaluated and are intended to be purely exemplary of the invention and are not intended to limit the scope of what the inventors regard as their invention. ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700

[0143] Example 1. Reciprocal relationship between integrin signaling and Src signaling

[0144] Objective

[0145] The objective of this study was to explore the relationship between integrin activation and Src activation in cancer cell lines.

[0146] Materials and Methods

[0147] Approximately 1 x 105of cells from various prostate cancer cell lines (PC-3, DU-145, VCAP, C4- 2B, and LnCAP) and triple negative breast cancer (TNBC) cell lines (BT-20, MDA-MB-231 , and MDA-MB- 468) were seeded into 6-well plates that are pre-coated with collagen and were treated with control or 10 ug / mL of BsAba5p1 / av. After 48 hours, cell lysates were prepared and assessed for expression of total Src, phosphorylated Src (Phospho-Src), and p-actin as loading control by western blot.

[0148] Next, approximately 1 x 105of DU-145 cells (DU-145 parental) and DU-145 cells in which PTEN had been knocked out (DU-145 PTEN-mt) were seeded into 6-well plates and treated with 100 nM of dasatinib (Src inhibitor), 10 ug / mL of BsAba5p1 / av, 100 nM of PF-431396 (focal adhesion kinase (FAK) inhibitor), both dasatinib and BsAba5p1 / av, or both dasatinib and PF-431396. After 48 hours, cells were collected and subjected to subcellular fractionation. Total, cytosolic, membranous, and nuclear fractions were generated by sequential centrifugation methods in appropriate buffers. Cytosolic and nuclear fractions were assessed for expression of active p-catenin (ABC) and yes-associated protein (YAP). Densitometric corrections were generated by comparison to nuclear lamin A / C as loading control.

[0149] Results

[0150] Phospho-Src expression is induced in basal-type cancer cells following Bs Aba 5 / 31 / av treatment

[0151] BsAba5p1 / av treatment upregulates phospho-Src specifically in basal-type prostate cancer cells (PC-3 and DU-145) and TNBC cells (BT-20, MDA-MB-231 , and MDA-MB-468), but not in luminal-type cancer cells (VCAP, C4-2B, and LnCAP) (FIG. 1). The greatest increase in phospho-Src following BsAba5p1 / av treatment is observed in DU-145 prostate cancer cells.

[0152] Reciprocal control of basal transcription factors in basal-type cancer cell lines

[0153] In DU-145 cells, BsAba5p1 / av treatment results in marked reduction of nuclear ABC and YAP, transcription factors which are markers of epithelial-mesenchymal transition and pathological mechanotransduction associated with basal-type of prostate cancer, whereas dasatinib treatment strongly upregulates nuclear ABC and YAP (FIG. 2A). In DU-145 PTEN-mt cells, BsAba5p1 / av treatment strongly induces nuclear ABC and YAP and dasatinib strongly reduces nuclear ABC and YAP (FIG. 2B). In both DU-145 and DU-145 PTEN-mt cells, the combination of BsAba5p1 / av and dasatinib reduces the expression of nuclear ABC and YAP (FIG. 2A and 2B). PF-431396 alone does not alter the expression of nuclear ABC or YAP in either cell line but does reduce nuclear ABC and YAP in both cells lines when combined with dasatinib (FIG. 2A and 2B). ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700

[0154] Conclusion

[0155] These results suggest that in addition to the canonical signaling pathway, which puts Src activation downstream of integrin activation, there exists a non-canonical model in which integrin- blockade leads to Src activation, in basal-type cancer cells specifically. Further, while there is a dynamic reciprocity of integrin and Src signaling in the nucleus, the combination of BsAba5p1 / av and Src inhibitor has potential synergistic action. Together, these data suggest that while markers of epithelial- mesenchymal transition and pathological mechanotransduction are reversed by BsAba5p1 / av, a pathway driven by Src signaling may mediate tumor cell survival and resistance to therapy. It is hypothesized that combined targeting of a5 and av integrins (with BsAba5p1 / av) and the Src / Abl pathway (with dasatinib) may overcome this resistance to induce cell death.

[0156] Example 2. Effects of the combined treatment with BsAba5|31 / av and a Src inhibitor on cell death

[0157] Objective

[0158] The objective of this study was to assess the effects of integrin inhibition and Src inhibition on apoptotic cell death.

[0159] Materials and Methods

[0160] DU-145 parental and DU-145 PTEN-mt cells were treated with vehicle control, 10 ug / mL of BsAba5p1 / av, 100 nM of dasatinib, or the combination of BsAba5p1 / av and dasatinib for 48 hours. Following treatment, apoptotic cell death was annotated by annexin V-PI staining using flow cytometry or by assessing the expression level of cleaved PARP using western blot. GAPDH was used as loading control.

[0161] Next, DU-145 parental and DU-145 PTEN-mt cells were treated with 10 ug / mL of monospecific a5pi integrin antibody (MsAb A5B1 ), 10 ug / mL of monospecific av integrin antibody (MsAb Av), 10 ug / mL of BsAba5p1 / av (BsAb A5B1 / Av), or 100 nM of dasatinib, alone or in combination, for 48 hours. Cell viability from treatment was assessed by annexin V-PI staining and normalized to the viability of vehicle control.

[0162] Results

[0163] Synergistic effects of the combined Bs Aba 5 / 31 / av and dasatinib treatment

[0164] In both DU-145 parental and DU-145 PTEN-mt cells, the combined BsAba5p1 / av and dasatinib treatment induces synthetic lethality (FIG. 3) and increases the expression of cleaved PARP (FIG. 4). In both DU-145 parental and DU-145 PTEN-mt cells, BsAba5p1 / av treatment leads to markedly superior induction of cell death over monospecific antibodies when combined with dasatinib (FIG. 5).

[0165] Conclusion

[0166] These results show that simultaneous inhibition of integrin and Src has synergic effects on apoptosis. Inhibiting all three targets - a5 integrin, av integrin, and Src - leads to greater effects on cell viability than inhibiting Src and either of the two integrins. Moreover, the inhibition of both a501 and av integrins with the bispecific antibody results in greater effects on cell viability than the combined use of the integrin-targeting monospecific antibodies, especially when combined with a Src inhibitor. This ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700 suggests that while simultaneous inhibition of a5 integrin, av integrin, and Src is necessary, the maximal synergistic effects of integrin and Src inhibition on cell viability is obtained with the bispecific antibody.

[0167] Example 3. Mechanism of action of the combined treatment with BsAba5|31 / av and a Src inhibitor

[0168] Objective

[0169] The objective of this study was to examine the mechanism by which the simultaneous inhibition of integrins and Src results in synergistic effects on apoptotic cell death. First, the involvement of FAK was interrogated, because targeting FAK, which is downstream of integrin activation, is an alternative strategy for modulating integrin-regulated signaling. Then, the effects of Src knockdown was examined.

[0170] Materials and Methods

[0171] FAK inhibition

[0172] DU-145 parental and DU-145 PTEN-mt cells were treated with vehicle control, 100 nM of dasatinib combined with 10 ug / mL of BsAba5p1 / av, or 100 nM of dasatinib combined with 100 nM of PF- 431396 for 48 hours. Cytotoxicity was monitored by annexin V-PI staining using flow cytometry.

[0173] Genetic knockdown of Src

[0174] Src was knocked down in DU-145 cells by introducing a dominant-negative kinase-dead mutant into a stable Src-knockdown cell line. DU-145 parental (DU-145-WT) and DU-145 with Src-knockdown (DU-145-SRC-KD) cells were treated with vehicle control, 10 ug / mL of BsAba5p1 / av, or 10 ug / mL of BsAba5p1 / av and 100 nM of dasatinib. After 48 hours of treatment, apoptotic cell death was assessed.

[0175] Results

[0176] Effects of FAK inhibition

[0177] In both DU-145 parental and DU-145 PTEN-mt cells, the combination of dasatinib and PF- 431396 leads to markedly reduced or absent cytotoxicity when compared to vehicle or the combination of dasatinib and BsAba5p1 / av (FIG. 6).

[0178] Effects of genetic knockdown of Src

[0179] Src knockdown in DU-145-SRC-KD is confirmed by reduced expression of SRC and phospho-Src in DU-145-SRC-KD compared to DU-145-WT (FIG. 7). While BsAba5p1 / av treatment of DU-145 parental cells does not result in cell death, BsAba5p1 / av treatment of DU-145-SRC-KD cells results in comparable levels of apoptotic cell death as induced by BsAba5p1 / av and dasatinib in DU-145 parental cells (FIG. 8).

[0180] Conclusion

[0181] Combined with the lack of effects of FAK inhibition on modulation of ABC and YAP (FIG. 2), these results suggest that FAK inhibition does not phenocopy the cytotoxic effects of integrin inhibition when combined with Src inhibition. These indicate that FAK is not a part of the mechanism of action of integrin inhibition. ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700

[0182] In addition, the ability of integrin inhibition to induce cell death even when Src is knocked down provides supportive evidence for the distinct and non-canonical role of Src signaling in the co-regulation of cell survival pathways with the integrins.

[0183] Example 4. Effects of the combined treatment with BsAba5pi / av and a Src inhibitor on cell death across prostate and breast cancer cell lines

[0184] Objective

[0185] The objective of this study was to assess the effects of the combined integrin and Src inhibition on cell death in additional prostate and TNBC cell lines.

[0186] Materials and Methods

[0187] Prostate cancer cell lines (PC-3 and C4-2B) and TNBC cell lines (MDA-MB-231 and BT-20) were treated with vehicle, 10 ug / mL of BsAba5p1 / av, 100 nM of dasatinib, 100 nM of PF-431396, combination of 100 nM of dasatinib and 10 ug / mL of BsAba5p1 / av, or combination of 100 nM of dasatinib and 100 nM of PF-431396. Treatment was for 120 hours in PC-3 cells and 48 hours in C4-2B, MDA-MB-231 , and BT- 20 cells. After treatment, cell death was annotated by annexin V-PI staining.

[0188] Results

[0189] The combination of dasatinib and BsAba5p1 / av induces synthetic lethality, and the combination of dasatinib and PF-431396 reduces synthetic lethality in all four cell lines (FIG. 9).

[0190] Conclusion

[0191] These results show that the synthetic lethal activity of the combination of integrin inhibition and Src inhibition is not wholly restricted to basal-type cancer cell lines.

[0192] Other Embodiments

[0193] While the invention has been described in connection with specific embodiments thereof, it will be understood that it is capable of further modifications and this application is intended to cover any variations, uses, or adaptations of the invention following, in general, the principles of the invention and including such departures from the invention that come within known or customary practice within the art to which the invention pertains and can be applied to the essential features hereinbefore set forth, and follows in the scope of the claims.

[0194] 1 . A method of treating a subject having cancer or a non-malignant condition or disease in which av and / or a5 (e.g., a5p1 ) integrin is expressed or implicated in pathogenesis or progression, the method comprising administering to the subject therapeutically effective amounts of (i) a bispecific antibody that specifically binds av integrin and a5 (e.g., a5p1 ) integrin and (ii) a Src inhibitor.

[0195] 2. The method of paragraph 1 , wherein the bispecific antibody comprises: (i) a first paratope that specifically binds an av integrin; and (ii) a second paratope that specifically binds an a5 (e.g., a5p1 ) integrin.

[0196] 3. The method of paragraph 2, wherein the second paratope specifically binds an epitope residing wholly within the a5 integrin. ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700

[0197] 4. The method of paragraph 2, wherein the second paratope specifically binds an epitope comprising both amino acid residues within the a5 integrin and amino acid residues within a pi integrin dimerized to the a5 integrin.

[0198] 5. The method of any one of paragraphs 2-4, wherein: (i) the first paratope comprises a light chain variable domain comprising CDR amino acid sequences of SEQ ID NOs: 1 -3 and a heavy chain variable domain comprising CDR amino acid sequences of SEQ ID NOs: 4-6; and (ii) the second paratope comprises a light chain variable domain comprising CDR amino acid sequences of SEQ ID NOs: 7-9 and a heavy chain variable domain comprising CDR amino acid sequences of SEQ ID NOs: 10-12.

[0199] 6. The method of paragraph 5, wherein: (i) the light chain variable domain of the first paratope further comprises at least one framework region (FR) comprising the amino acid sequence of one of SEQ ID NOs: 13-16 or an amino acid sequence that is at least 80% identical to any one of SEQ ID NOs: 13-16; (ii) the heavy chain variable domain of the first paratope further comprises at least one FR comprising the amino acid sequence of one of SEQ ID NOs: 17-20 or an amino acid sequence that is at least 80% identical to any one of SEQ ID NOs: 17-20; (Hi) the light chain variable domain of the second paratope further comprises at least one FR comprising the amino acid sequence of one of SEQ ID NOs: 21 -24 or an amino acid sequence that is at least 80% identical to any one of SEQ ID NOs: 21 -24; or (iv) the heavy chain variable domain of the second paratope further comprises at least one FR comprising the amino acid sequence of one of SEQ ID NOs: 25-28 or an amino acid sequence that is at least 80% identical to any one of SEQ ID NOs: 25-28.

[0200] 7. The method of any one of paragraphs 1 -6, wherein the bispecific antibody comprises: (i) an antigen binding fragment portion that comprises one of the first or second paratopes; and (ii) a singlechain Fv portion that comprises the other of the first or second paratopes.

[0201] 8. The method of any one of paragraphs 1 -7, wherein the bispecific antibody further comprises an Fc region.

[0202] 9. The method of any one of paragraphs 1 -8, wherein the bispecific antibody comprises: (i) one or two polypeptides, each comprising an amino acid sequence that is at least 80% identical to SEQ ID NO: 41 ; and (ii) one or two additional polypeptides, each comprising an amino acid sequence that is at least 80% identical to SEQ ID NO: 42. In some embodiments, the sequences herein are at least 85%, 90%, 95%, 98%, 99%, or 100% to the reference sequences.

[0203] 10. The method of any one of paragraphs 1 -9, wherein the bispecific antibody is chimeric.

[0204] 11 . The method of paragraph 10, wherein the bispecific antibody is at least partially humanized or de-immunized.

[0205] 12. The method of any one of paragraphs 1 -11 , wherein the bispecific antibody is divalent, trivalent, or tetravalent.

[0206] 13. The method of any one of paragraphs 1 -12, wherein the Src inhibitor is a dual Src-Abl inhibitor.

[0207] 14. The method of any one of paragraphs 1 -13, wherein the Src inhibitor is selected from the group consisting of dasatinib, dasatinib monohydrate, bafetinib, bosutinib, danusertib, ponatinib, repotrectinib, saracatinib, tirbanibulin, AP24149, AP24163, AZD0424, CCT196969, DC2036, NVP- BHG712, PD173955, PP1 , PP2, PP121 , SGX393, WH-4-023, XL228, and 3,4-Methylenedioxy-p- nitrostyrene. ATTORNEY DOCKET NO. 00398-571 WO2 TMC 700

[0208] 15. The method of paragraph 14, wherein the Src inhibitor is dasatinib.

[0209] 16. The method of any one of paragraphs 1 -15, wherein the bispecific antibody and Src inhibitor are administered to the subject at the same time.

[0210] 17. The method of any one of paragraphs 1 -15, wherein the bispecific antibody and Src inhibitor are administered to the subject sequentially.

[0211] 18. The method of paragraph 17, wherein the bispecific antibody is administered to the subject before the Src inhibitor is administered to the subject.

[0212] 19. The method of paragraph 17, wherein the bispecific antibody is administered to the subject after the Src inhibitor is administered to the subject.

[0213] 20. The method of any one of paragraphs 1 -19, wherein the cancer is selected from the group consisting of prostate cancer, kidney cancer, breast cancer, melanoma, pancreatic cancer, ovarian cancer, colonic cancer, cervical cancer, head and neck cancer, lung cancer, gastric cancer, endometrial cancer, bone and / or brain metastases, soft-tissue sarcoma, osteosarcoma, hepatoma, basal cell carcinoma, glioblastoma, angiosarcoma, T-cell lymphoma, and multiple myeloma.

[0214] 21 . The method of paragraph 20, wherein the cancer is prostate cancer.

[0215] 22. The method of paragraph 20, wherein the cancer is breast cancer.

[0216] 23. The method of any one of paragraphs 1 -22, wherein the non-malignant condition is selected from the group consisting of non-malignant fibrotic disease involving lung, liver, heart, kidney, bone marrow, or skin, scleroderma, chronic graft-vs-host disease, liver fibrosis, interstitial lung disease.

[0217] 24. The method of any one of paragraphs 1 -22, wherein the non-malignant disease is selected from the group consisting of pathological angiogenesis, proliferative retinopathy, wet macular degeneration, arteriovenous malformation, and cavernous hemangioma.

[0218] 25. The method of any one of paragraphs 1 -24, wherein the subject does not respond to or is resistant to monotherapy with an av integrin inhibitor.

[0219] 26. The method of any one of paragraphs 1 -24, wherein the subject does not respond to or is resistant to monotherapy with an a5 (e.g., a5p1 ) integrin inhibitor.

[0220] 27. The method of any one of paragraphs 1 -24, wherein the subject does not respond to or is resistant to monotherapy with a Src inhibitor.

[0221] 28. The method of any one of paragraphs 1 -27, wherein the bispecific antibody and the Src inhibitor are present within a single pharmaceutical composition.

[0222] 29. A pharmaceutical composition comprising (i) a bispecific antibody that specifically binds av integrin and a5 (e.g., a5p1 ) integrin (e.g., as described herein) and (ii) a Src inhibitor (e.g., as described herein).

[0223] 30. The pharmaceutical composition of paragraph 29, further comprising a pharmaceutically acceptable carrier.

[0224] 31 . A combination of (i) a bispecific antibody that specifically binds av integrin and a5 (e.g., a5p1 ) integrin (e.g., as described herein) and (ii) a Src inhibitor (e.g., as described herein) for use in a method of treating a subject having cancer or non-malignant condition or disease in which av and / or a5 (e.g., a5p1 ) integrin is expressed or implicated in pathogenesis or progression.

[0225] Other embodiments are within the claims.

Claims

ATTORNEY DOCKET NO. 00398-571 WO2TMC 700CLAIMS1 . A method of treating a subject having cancer or a non-malignant condition or disease in which av and / or a5 integrin is expressed or implicated in pathogenesis or progression, the method comprising administering to the subject therapeutically effective amounts of (i) a bispecific antibody that specifically binds av integrin and a5 integrin and (ii) a Src inhibitor.

2. The method of claim 1 , wherein the bispecific antibody comprises:(i) a first paratope that specifically binds an av integrin; and(ii) a second paratope that specifically binds an a5 integrin.

3. The method of claim 2, wherein the second paratope specifically binds an epitope residing wholly within the a5 integrin.

4. The method of claim 2, wherein the second paratope specifically binds an epitope comprising both amino acid residues within the a5 integrin and amino acid residues within a pi integrin dimerized to the a5 integrin.

5. The method of claim 2, wherein:(i) the first paratope comprises a light chain variable domain comprising CDR amino acid sequences of SEQ ID NOs: 1 -3 and a heavy chain variable domain comprising CDR amino acid sequences of SEQ ID NOs: 4-6; and(ii) the second paratope comprises a light chain variable domain comprising CDR amino acid sequences of SEQ ID NOs: 7-9 and a heavy chain variable domain comprising CDR amino acid sequences of SEQ ID NOs: 10-12.

6. The method of claim 5, wherein:(i) the light chain variable domain of the first paratope further comprises at least one framework region (FR) comprising the amino acid sequence of one of SEQ ID NOs: 13-16 or an amino acid sequence that is at least 80% identical to any one of SEQ ID NOs: 13-16;(ii) the heavy chain variable domain of the first paratope further comprises at least one FR comprising the amino acid sequence of one of SEQ ID NOs: 17-20 or an amino acid sequence that is at least 80% identical to any one of SEQ ID NOs: 17-20;(Hi) the light chain variable domain of the second paratope further comprises at least one FR comprising the amino acid sequence of one of SEQ ID NOs: 21 -24 or an amino acid sequence that is at least 80% identical to any one of SEQ ID NOs: 21 -24; or(iv) the heavy chain variable domain of the second paratope further comprises at least one FR comprising the amino acid sequence of one of SEQ ID NOs: 25-28 or an amino acid sequence that is at least 80% identical to any one of SEQ ID NOs: 25-28.

7. The method of claim 1 , wherein the bispecific antibody comprises:(i) an antigen binding fragment portion that comprises one of the first or second paratopes; andATTORNEY DOCKET NO. 00398-571 WO2 TMC 700(ii) a single-chain Fv portion that comprises the other of the first or second paratopes.

8. The method of claim 1 , wherein the bispecific antibody further comprises an Fc region.

9. The method of claim 1 , wherein the bispecific antibody comprises:(i) two polypeptides, each comprising an amino acid sequence that is at least 80% identical to SEQ ID NO: 41 ; and(ii) two additional polypeptides, each comprising an amino acid sequence that is at least 80% identical to SEQ ID NO: 42.

10. The method of claim 1 , wherein the bispecific antibody is chimeric.11 . The method of claim 10, wherein the bispecific antibody is at least partially humanized or deimmunized.

12. The method of claim 1 , wherein the bispecific antibody is divalent, trivalent, or tetravalent.

13. The method of claim 1 , wherein the Src inhibitor is a dual Src-Abl inhibitor.

14. The method of claim 1 , wherein the Src inhibitor is selected from the group consisting of dasatinib, dasatinib monohydrate, bafetinib, bosutinib, danusertib, ponatinib, repotrectinib, saracatinib, tirbanibulin, AP24149, AP24163, AZD0424, CCT196969, DC2036, NVP-BHG712, PD173955, PP1 , PP2, PP121 , SGX393, WH-4-023, XL228, and 3,4-Methylenedioxy-p-nitrostyrene.

15. The method of claim 14, wherein the Src inhibitor is dasatinib.

16. The method of claim 1 , wherein the bispecific antibody and Src inhibitor are administered to the subject at the same time.

17. The method of claim 1 , wherein the bispecific antibody and Src inhibitor are administered to the subject sequentially.

18. The method of claim 17, wherein the bispecific antibody is administered to the subject before the Src inhibitor is administered to the subject.

19. The method of claim 17, wherein the bispecific antibody is administered to the subject after the Src inhibitor is administered to the subject.

20. The method of claim 1 , wherein the cancer is selected from the group consisting of prostate cancer, kidney cancer, breast cancer, melanoma, pancreatic cancer, ovarian cancer, colonic cancer, cervical cancer, head and neck cancer, lung cancer, gastric cancer, endometrial cancer, bone and / or brainATTORNEY DOCKET NO. 00398-571 WO2 TMC 700 metastases, soft-tissue sarcoma, osteosarcoma, hepatoma, basal cell carcinoma, glioblastoma, angiosarcoma, T-cell lymphoma, and multiple myeloma.21 . The method of claim 20, wherein the cancer is prostate cancer.

22. The method of claim 20, wherein the cancer is breast cancer.

23. The method of claim 1 , wherein the non-malignant condition is selected from the group consisting of non-malignant fibrotic disease involving lung, liver, heart, kidney, bone marrow, or skin, scleroderma, chronic graft-vs-host disease, liver fibrosis, interstitial lung disease.

24. The method of claim 1 , wherein the non-malignant disease is selected from the group consisting of pathological angiogenesis, proliferative retinopathy, wet macular degeneration, arteriovenous malformation, and cavernous hemangioma.

25. The method of claim 1 , wherein the subject does not respond to or is resistant to monotherapy with an av integrin inhibitor.

26. The method of claim 1 , wherein the subject does not respond to or is resistant to monotherapy with an a5 (e.g., a5p1 ) integrin inhibitor.

27. The method of claim 1 , wherein the subject does not respond to or is resistant to monotherapy with a Src inhibitor.

28. The method of claim 1 , wherein the bispecific antibody and the Src inhibitor are present within a single pharmaceutical composition.

29. A pharmaceutical composition comprising (i) a bispecific antibody that specifically binds av integrin and a5 (e.g., a5p1 ) integrin and (ii) a Src inhibitor.

30. The pharmaceutical composition of claim 29, further comprising a pharmaceutically acceptable carrier.31 . A combination of (i) a bispecific antibody that specifically binds av integrin and a5 (e.g., a5p1 ) integrin and (ii) a Src inhibitor for use in a method of treating a subject having cancer or non-malignant condition or disease in which av and / or a5 (e.g., a5p1 ) integrin is expressed or implicated in pathogenesis or progression.

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