B7-H4 therapeutic binding molecules for treatment of cancer

Abstract

The present disclosure relates to a method for treating cancer in a subject comprising administering to the subject an antibody-drug conjugate (ADC) that specifically binds to B7-H4. In various aspects, the ADC is administered to the subject in an amount of from about 0.8 mg / kg to about 4.8 mg / kg.

Description

[0001] Cross-reference to related applications

[0002] This application claims priority to U.S. Provisional Patent Application No. 63 / 498,998, filed April 28, 2023, which is incorporated herein by reference in its entirety for all purposes. This application also claims priority to U.S. Provisional Patent Application No. 63 / 505,727, filed June 2, 2023, which is incorporated herein by reference in its entirety for all purposes.

[0003] Materials submitted electronically are incorporated by reference

[0004] This application is incorporated by reference as a sequence list submitted with this application as an XML file named "B7H4-200-WO-PCT SL", created on April 12, 2024, and is 76 kilobytes in size. Technical Field

[0005] This disclosure provides a method for treating cancer in a subject, the method comprising administering to the subject an antibody-drug conjugate (ADC) that specifically binds to B7-H4. Background Technology

[0006] Surgery, radiation therapy, and chemotherapy, alone or in combination, are the most traditional and widely used treatments for cancer. While these treatments are effective in removing or killing cancer cells, they often cause undesirable side effects such as hair loss, anemia, severe nausea, and the death of healthy cells in treated patients. These limitations highlight the urgent need for innovative and less harmful treatments for cancer. Antibody-based cancer therapies rely on antibody-drug conjugates recognizing and binding to specific proteins on cancer cells. Antibody-drug conjugates (ADCs) utilize the specificity of the antibody moiety of the conjugate to deliver highly toxic agents directly to the cells to be killed. Using antibody or ADC cancer therapies alone, or in combination with other small molecule-based cancer therapies, can improve treatment outcomes by attacking malignant cells and tumors in one or more ways. Summary of the Invention

[0007] This disclosure provides a method for treating cancer in a subject, the method comprising administering to the subject an antibody-drug conjugate (ADC) specifically bound to B7-H4 in an amount of about 0.8 mg / kg to about 4.8 mg / kg. The ADC comprises:

[0008] i. An antibody or antigen-binding fragment thereof that binds to the B7-H4 polypeptide, the antibody or antigen-binding fragment thereof comprising:

[0009] a) heavy chain CDR1 (HCDR1), heavy chain CDR2 (HCDR2), heavy chain CDR3 (HCDR3), light chain CDR1 (LCDR1), light chain CDR2 (LCDR2), and light chain CDR3 (LCDR3) comprising the amino acid sequences of SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, respectively, or functional variants thereof;

[0010] b) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 comprising the amino acid sequences of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6, respectively, or functional variants thereof;

[0011] c) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 comprising the amino acid sequences of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, and SEQ ID NO: 18, respectively, or functional variants thereof;

[0012] (d) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 comprising the amino acid sequences of SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, and SEQ ID NO: 24, respectively, or functional variants thereof; or

[0013] (e) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 comprising the amino acid sequences of SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, and SEQ ID NO: 30, respectively, or functional variants thereof;

[0014] ii. a cleavable linker; and

[0015] iii. a cytotoxic agent.

[0016] In some aspects, the amount of ADC administered is about 0.8 mg / kg, about 1.6 mg / kg, about 2.0 mg / kg, about 2.4 mg / kg, about 3.2 mg / kg, about 3.6 mg / kg, or about 4.8 mg / kg. In some aspects, the amount of ADC administered is about 0.8 mg / kg, about 1.6 mg / kg, about 2.4 mg / kg, about 3.2 mg / kg, about 3.6 mg / kg, or about 4.8 mg / kg. In some aspects, the amount of ADC administered is about 1.6 mg / kg. In some aspects, the amount of ADC administered is about 2.0 mg / kg. In some aspects, the amount of ADC administered is about 2.4 mg / kg. In some aspects, the amount of ADC administered is about 3.2 mg / kg.

[0017] In some aspects, the cancer comprises cancer cells that express B7-H4.

[0018] In some aspects, the ADC is administered to the subject once every three weeks. In some aspects, the subject has previously received treatment with another chemotherapeutic agent. In some aspects, the subject has previously received treatment with a platinum-based chemotherapeutic agent.

[0019] In some aspects, the cancer is selected from endometrial cancer, ovarian cancer, biliary tract cancer, breast cancer, pancreatic cancer, prostate cancer, hematological cancer, cholangiocarcinoma, NSCLC (squamous carcinoma and / or adenocarcinoma), gastrointestinal cancer (such as gastric cancer and colorectal cancer), and lung cancer. In some aspects, the cancer is selected from endometrial cancer, ovarian cancer, breast cancer, pancreatic cancer, prostate cancer, hematological cancer, cholangiocarcinoma, NSCLC (squamous carcinoma and / or adenocarcinoma), gastrointestinal cancer (such as gastric cancer and colorectal cancer), and lung cancer. In some aspects, the cancer is a breast cancer selected from hormone receptor positive (HR+) breast cancer, human epidermal growth factor receptor 2 positive (HER2+) breast cancer, and triple negative breast cancer (TNBC). In some aspects, the cancer is a homologous recombination deficient (HRD) cancer. In some aspects, the cancer comprises one or more cells having a mutation in an HRD gene selected from BRCA1, BRCA2, ATM, BRIP1, BARD1, CDK12, CHEK1, CHEK2, FANCL, PALB2, PPP2R2A, RAD51B, RAD51C, RAD51D, and RAD54L. In some aspects, the mutation in the HRD gene is selected from BRCA1, BRCA2, and ATM.

[0020] In some aspects, the antibody or antigen-binding fragment thereof comprises:

[0021] i. a variable heavy (VH) chain and a variable light (VL) chain comprising the amino acid sequences of SEQ ID NO: 45 and SEQ ID NO: 34, respectively, or functional variants thereof;

[0022] ii. a variable heavy (VH) chain and a variable light (VL) chain comprising the amino acid sequences of SEQ ID NO: 33 and SEQ ID NO: 34, respectively, or functional variants thereof;

[0023] iii. a variable heavy (VH) chain and a variable light (VL) chain comprising the amino acid sequences of SEQ ID NO: 43 and SEQ ID NO: 34, respectively, or functional variants thereof;

[0024] iv. a variable heavy (VH) chain and a variable light (VL) chain comprising the amino acid sequences of SEQ ID NO: 46 and SEQ ID NO: 34, respectively, or functional variants thereof;

[0025] v. a variable heavy (VH) chain and a variable light (VL) chain comprising the amino acid sequences of SEQ ID NO: 47 and SEQ ID NO: 34, respectively, or functional variants thereof;

[0026] vi. a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 31 and SEQ ID NO: 32, respectively, or functional variants thereof;

[0027] vii. a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 35 and SEQ ID NO: 36, respectively, or functional variants thereof;

[0028] viii. a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 37 and SEQ ID NO: 38, respectively, or functional variants thereof; or

[0029] ix. a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 39 and SEQ ID NO: 40, respectively, or functional variants thereof.

[0030] In some aspects, the antibody or antigen-binding fragment thereof comprises a HCDR1, a HCDR2, a HCDR3, a LCDR1, a LCDR2, and a LCDR3 comprising the amino acid sequences of SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, respectively, or functional variants thereof.

[0031] In some aspects, the antibody or antigen-binding fragment thereof comprises a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 45 and SEQ ID NO: 34, respectively, or functional variants thereof.

[0032] In some aspects, the antibody or antigen-binding fragment thereof binds to an OVCAR4 cell line.

[0033] In some aspects, the antibody or antigen-binding fragment thereof comprises: a heavy chain constant region comprising the amino acid sequence of SEQ ID NO: 41. In some aspects, the antibody or antigen-binding fragment thereof comprises: a heavy chain constant region comprising the amino acid sequence of SEQ ID NO: 52. In some aspects, the antibody or antigen-binding fragment thereof comprises: a light chain constant region comprising the amino acid sequence of SEQ ID NO: 42.

[0034] In some aspects, the antibody or antigen-binding fragment thereof comprises: a heavy chain comprising the amino acid sequence of SEQ ID NO: 51; and a light chain comprising the amino acid sequence of SEQ ID NO: 44. In some aspects, the antibody or antigen-binding fragment thereof comprises: a heavy chain comprising the amino acid sequence of SEQ ID NO: 48; and a light chain comprising the amino acid sequence of SEQ ID NO: 44.

[0035] In some aspects, the antibody or antigen-binding fragment thereof is a monoclonal antibody. In some aspects, the antibody or antigen-binding fragment thereof is a humanized monoclonal antibody.

[0036] In some aspects, the cleavable linker is an mp-PEG8-val-ala linker.

[0037] In some aspects, the cytotoxic agent is a topoisomerase inhibitor.

[0038] In some aspects, the topoisomerase inhibitor is a compound of Formula A

[0039] .

[0040] In some aspects, ii) the cleavable linker and iii) the cytotoxic agent together are selected from the following compounds:

[0041] (SG3932); (SG4010); (SG4057); and (SG4052).

[0042] In some aspects, ii) the linker and iii) the cytotoxic agent together are compound SG3932.

[0043] In some aspects, the ADC has a drug-to-antibody ratio (DAR) of between about 1 and about 8. In some aspects, the ADC has a DAR of about 8.

[0044] ​In some aspects, the disclosure further provides a method of treating a cancer in a subject, the method comprising administering to the subject an antibody-drug conjugate (ADC) that specifically binds to B7-H4, the ADC comprising:

[0045] i. an antibody or antigen-binding fragment thereof that binds to a B7-H4 polypeptide, the antibody or antigen-binding fragment thereof comprising: a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2), a heavy chain CDR3 (HCDR3), a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), and a light chain CDR3 (LCDR3) comprising the amino acid sequences of SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, respectively, or functional variants thereof; and

[0046] ii. a cleavable linker and a cytotoxic agent conjugated to the antibody or antigen-binding fragment thereof having the following formula:

[0047] (SG3932).

[0048] In some aspects, the amount of ADC administered is about 0.8 mg / kg, about 1.6 mg / kg, about 2.0 mg / kg, about 2.4 mg / kg, about 3.2 mg / kg, about 3.6 mg / kg, or about 4.8 mg / kg. In some aspects, the amount of ADC administered is about 0.8 mg / kg, about 1.6 mg / kg, about 2.4 mg / kg, about 3.2 mg / kg, about 3.6 mg / kg, or about 4.8 mg / kg. In some aspects, the amount of ADC administered is about 1.6 mg / kg. In some aspects, the amount of ADC administered is about 2.0 mg / kg. In some aspects, the amount of ADC administered is about 2.4 mg / kg. In some aspects, the amount of ADC administered is about 3.2 mg / kg.

[0049] In some aspects, the antibody or antigen-binding fragment thereof comprises a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 45 and SEQ ID NO: 34, respectively, or functional variants thereof.

[0050] In some aspects, the cancer comprises cancer cells that express B7-H4.

[0051] In some aspects, the ADC is administered to the subject once every three weeks. In some aspects, the subject has previously received treatment with another chemotherapeutic agent. In some aspects, the subject has previously received treatment with a platinum-based chemotherapeutic agent.

[0052] In some aspects, the cancer is selected from endometrial cancer, ovarian cancer, biliary tract cancer, breast cancer, pancreatic cancer, prostate cancer, hematological cancer, cholangiocarcinoma, NSCLC (squamous carcinoma and / or adenocarcinoma), gastrointestinal cancer (such as gastric cancer and colorectal cancer), and lung cancer. In some aspects, the cancer is selected from endometrial cancer, ovarian cancer, breast cancer, pancreatic cancer, prostate cancer, hematological cancer, cholangiocarcinoma, NSCLC (squamous carcinoma and / or adenocarcinoma), gastrointestinal cancer (such as gastric cancer and colorectal cancer), and lung cancer.

[0053] In some aspects, the cancer is a breast cancer selected from hormone receptor positive (HR+) breast cancer, human epidermal growth factor receptor 2 positive (HER2+) breast cancer, and triple negative breast cancer (TNBC).

[0054] In some aspects, the cancer is a homologous recombination deficient (HRD) cancer.

[0055] In some aspects, the cancer comprises one or more cells having a mutation in an HRD gene selected from BRCA1, BRCA2, ATM, BRIP1, BARD1, CDK12, CHEK1, CHEK2, FANCL, PALB2, PPP2R2A, RAD51B, RAD51C, RAD51D, and RAD54L.

[0056] In some aspects, the mutation in the HRD gene is selected from BRCA1, BRCA2, and ATM.

[0057] In some aspects of the above methods, at least about 25% of the cancer cells in the subject are B7-H4 positive cells. In some aspects of the above methods, the B7-H4 positive cells are determined using immunohistochemistry (IHC).

[0058] In some aspects, the disclosure further provides a pharmaceutical composition comprising an antibody-drug conjugate (ADC) that specifically binds to B7-H4, the ADC being administered in an amount of about 0.8 mg / kg to about 4.8 mg / kg, the ADC comprising:

[0059] i. an antibody or antigen-binding fragment thereof that binds to a B7-H4 polypeptide, the antibody or antigen-binding fragment thereof comprising:

[0060] ii. a cleavable linker and a cytotoxic agent conjugated to an antibody or antigen-binding fragment thereof having the formula:

[0061] (SG3932).

[0062] In some aspects, the pharmaceutical composition is administered in an amount of about 0.8 mg / kg, about 1.6 mg / kg, about 2.0 mg / kg, about 2.4 mg / kg, about 3.2 mg / kg, about 3.6 mg / kg, or about 4.8 mg / kg of the ADC. In some aspects, the pharmaceutical composition is administered in an amount of about 0.8 mg / kg, about 1.6 mg / kg, about 2.4 mg / kg, about 3.2 mg / kg, about 3.6 mg / kg, or about 4.8 mg / kg of the ADC. In some aspects, the pharmaceutical composition is administered in an amount of about 1.6 mg / kg of the ADC. In some aspects, the pharmaceutical composition is administered in an amount of about 2.0 mg / kg of the ADC. In some aspects, the pharmaceutical composition is administered in an amount of about 2.4 mg / kg of the ADC. In some aspects, the pharmaceutical composition is administered in an amount of about 3.2 mg / kg of the ADC.

[0063] In some aspects, the antibody or antigen-binding fragment of the ADC comprises a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 45 and SEQ ID NO: 34, respectively, or functional variants thereof.

[0064] In some aspects, the present disclosure further provides a pharmaceutical composition comprising an antibody-drug conjugate (ADC) that specifically binds to B7-H4, the ADC being administered in an amount of about 25 mg to about 900 mg, the ADC comprising:

[0065] i. an antibody or antigen-binding fragment thereof that binds to a B7-H4 polypeptide, the antibody or antigen-binding fragment thereof comprising:

[0066] a) heavy chain CDR1 (HCDR1), heavy chain CDR2 (HCDR2), heavy chain CDR3 (HCDR3), light chain CDR1 (LCDR1), light chain CDR2 (LCDR2), and light chain CDR3 (LCDR3) comprising the amino acid sequences of SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, respectively, or functional variants thereof;

[0067] b) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 comprising the amino acid sequences of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6, respectively, or functional variants thereof;

[0068] c) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 comprising the amino acid sequences of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, and SEQ ID NO: 18, respectively, or functional variants thereof;

[0069] d) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 comprising the amino acid sequences of SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, and SEQ ID NO: 24, respectively, or functional variants thereof; or

[0070] e) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 comprising the amino acid sequences of SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, and SEQ ID NO: 30, respectively, or functional variants thereof;

[0071] ii. a cleavable linker; and

[0072] iii. a cytotoxic agent.

[0073] In some aspects of the pharmaceutical composition, the amount of ADC administered is about 25 mg, about 40 mg, about 60 mg, about 80 mg, about 100 mg, about 120 mg, about 140 mg, about 160 mg, about 180 mg, about 200 mg, about 220 mg, about 240 mg, about 260 mg, about 280 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 600 mg, about 750 mg, or about 900 mg.

[0074] In some aspects of the pharmaceutical composition, the cleavable linker and cytotoxic agent of the combination have the following formula:

[0075] (SG3932).

[0076] In some aspects of the pharmaceutical composition, the antibody or antigen-binding fragment of the ADC comprises VH and VL chains comprising the amino acid sequences of SEQ ID NO: 45 and SEQ ID NO: 34, respectively, or functional variants thereof.

[0077] The present disclosure further provides a kit comprising a pharmaceutical composition as described above and elsewhere herein. In some aspects, the kit further comprises instructions for administering the pharmaceutical composition.

[0078] The present disclosure further provides a pharmaceutical composition as described above and elsewhere herein comprising an ADC that specifically binds to B7-H4 for use in treating a cancer. The treatment can be any of the methods of treating a cancer according to the methods of using an ADC that specifically binds to B7-H4 as described herein. In related aspects, the present disclosure further provides the use of a pharmaceutical composition as described above and elsewhere herein comprising an ADC that specifically binds to B7-H4 in the manufacture of a medicament for treating a cancer. The treatment can be any of the methods of treating a cancer according to the methods of using an ADC that specifically binds to B7-H4 as described herein. In some aspects, the cancer comprises cancer cells that express B7-H4.

[0079] In some aspects, the cancer is selected from endometrial cancer, ovarian cancer, biliary tract cancer, breast cancer, pancreatic cancer, prostate cancer, hematological cancer, cholangiocarcinoma, NSCLC (squamous carcinoma and / or adenocarcinoma), gastrointestinal cancer (such as gastric cancer and colorectal cancer), and lung cancer. In some aspects, the cancer is selected from endometrial cancer, ovarian cancer, breast cancer, pancreatic cancer, prostate cancer, hematological cancer, cholangiocarcinoma, NSCLC (squamous carcinoma and / or adenocarcinoma), gastrointestinal cancer (such as gastric cancer and colorectal cancer), and lung cancer. In some aspects, the cancer is a breast cancer selected from hormone receptor positive (HR+) breast cancer, human epidermal growth factor receptor 2 positive (HER2+) breast cancer, and triple negative breast cancer (TNBC). In some aspects, the cancer is a homologous recombination deficient (HRD) cancer.

[0080] In some aspects, the cancer comprises one or more cells having a mutation in an HRD gene selected from BRCA1, BRCA2, ATM, BRIP1, BARD1, CDK12, CHEK1, CHEK2, FANCL, PALB2, PPP2R2A, RAD51B, RAD51C, RAD51D, and RAD54L. In some aspects, the mutation in the HRD gene is selected from BRCA1, BRCA2, and ATM.

[0081] In some aspects, the disclosure provides a method of treating a cancer in a subject, the method comprising administering to the subject an antibody-drug conjugate (ADC) that specifically binds to B7-H4, the ADC comprising:

[0082] i. an antibody or antigen binding fragment thereof that binds to a B7-H4 polypeptide, the antibody or antigen binding fragment thereof comprising:

[0083] a) a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2), a heavy chain CDR3 (HCDR3), a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), and a light chain CDR3 (LCDR3) comprising the amino acid sequences of SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, respectively, or functional variants thereof;

[0084] b) a HCDR1, a HCDR2, a HCDR3, a LCDR1, a LCDR2, and a LCDR3 comprising the amino acid sequences of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6, respectively, or functional variants thereof;

[0085] c) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3, respectively, comprising the amino acid sequences of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17 and SEQ ID NO: 18, or functional variants thereof;

[0086] d) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3, respectively, comprising the amino acid sequences of SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23 and SEQ ID NO: 24, or functional variants thereof; or

[0087] e) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3, respectively, comprising the amino acid sequences of SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29 and SEQ ID NO: 30, or functional variants thereof;

[0088] ii. a cleavable linker; and

[0089] iii. a cytotoxic agent;

[0090] wherein at least about 25% of the cancer cells in the subject are B7-H4 positive cells.

[0091] In some aspects, the B7-H4 positive cells are determined by measuring membrane B7-H4 expression prior to administration of the ADC, optionally wherein the B7-H4 expression is determined using immunohistochemistry (IHC), further wherein the IHC is optionally performed with an antibody reagent. The assessment can be based on measuring B7-H4 expression of a sample (e.g., a tissue sample, optionally a cancer sample) obtained from the subject. Thus, in some aspects, the disclosure relates to a method for diagnosing a cancer sample as B7-H4 positive, the method comprising: determining the cancer sample, optionally using immunohistochemistry (IHC), to determine whether at least 25% of the cells express membrane B7-H4, further wherein the IHC is optionally performed with an antibody reagent. If the cancer sample is diagnosed as B7-H4 positive, the subject can be treated with an ADC that specifically binds to B7-H4 as described herein.

[0092] In some aspects, the ADC is administered in an amount of about 0.8 mg / kg to about 4.8 mg / kg. In some aspects, the ADC is administered to the subject once every three weeks. In some aspects, the subject has previously received treatment with another chemotherapeutic agent. In some aspects, the subject has previously received treatment with a platinum-based chemotherapeutic agent. In some aspects, the cancer is selected from endometrial cancer, ovarian cancer, biliary tract cancer, breast cancer, pancreatic cancer, prostate cancer, hematological cancer, cholangiocarcinoma, NSCLC (squamous carcinoma and / or adenocarcinoma), gastrointestinal cancer (such as gastric cancer and colorectal cancer), and lung cancer. In some aspects, the cancer is selected from endometrial cancer, ovarian cancer, breast cancer, pancreatic cancer, prostate cancer, hematological cancer, cholangiocarcinoma, NSCLC (squamous carcinoma and / or adenocarcinoma), gastrointestinal cancer (such as gastric cancer and colorectal cancer), and lung cancer. In some aspects, the cancer is a breast cancer selected from hormone receptor positive (HR+) breast cancer, human epidermal growth factor receptor 2 positive (HER2+) breast cancer, and triple negative breast cancer (TNBC). In some aspects, the cancer is a homologous recombination deficiency (HRD) cancer. In some aspects, the cancer comprises one or more cells having a mutation in an HRD gene selected from BRCA1, BRCA2, ATM, BRIP1, BARD1, CDK12, CHEK1, CHEK2, FANCL, PALB2, PPP2R2A, RAD51B, RAD51C, RAD51D, and RAD54L. In some aspects, the mutation in the HRD gene is selected from BRCA1, BRCA2, and ATM. In some aspects, the antibody or antigen-binding fragment thereof comprises a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 45 and SEQ ID NO: 34, respectively, or a functional variant thereof. In some aspects, the antibody or antigen-binding fragment thereof comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 51; and a light chain comprising the amino acid sequence of SEQ ID NO: 44. In some aspects, the antibody or antigen-binding fragment thereof is a monoclonal antibody. In some aspects, the antibody or antigen-binding fragment thereof is a humanized monoclonal antibody. In some aspects, the cleavable linker is an mp-PEG8-val-ala linker. In some aspects, the cytotoxic agent is a topoisomerase inhibitor. In some aspects, the topoisomerase inhibitor is a compound of Formula A

[0093]

[0094] In some aspects, ii) the cleavable linker and iii) the cytotoxic agent are together selected from the following compounds:

[0095] (SG3932); (SG4010); (SG4057); and ​(SG4052).

[0096] In some aspects, ii) the linker and iii) the cytotoxic agent together are compound SG3932. In some aspects, the ADC has a drug to antibody ratio (DAR) of between about 1 and about 8. In some aspects, the ADC has a DAR of about 8. BRIEF DESCRIPTION OF DRAWINGS

[0097] The following drawings form part of the present specification and are included to further demonstrate certain exemplary aspects of the present disclosure.

[0098] Figure 1 is a schematic diagram showing the main protocol structure for the treatment as described in the Examples. The main protocol structure includes sub-study 1 of AZD8205 monotherapy, and one or more combination sub-studies using AZD8205 with another active pharmaceutical agent.

[0099] Figure 2 is a schematic diagram showing an exemplary treatment method including a monotherapy dose escalation phase (Part A) and a monotherapy dose expansion phase (Part B) as described in the Examples.

[0100] Figure 3 is a graph showing the response of human ovarian cancer (OVCA) subjects to AZD8205 treatment at amounts of 0.8 mg / kg, 1.6 mg / kg, 2.4 mg / kg, and 3.2 mg / kg, respectively. Response was confirmed by detecting levels of cancer antigen 125 (CA-125) in the subjects. Subjects with measurable CA-125 (>2x upper limit of normal (ULN)) are shown. Circulating tumor DNA (ctDNA) was used to detect molecular response (MR), where P-MR indicates partial molecular response (ctDNA reduction from baseline >50%), C-MR indicates complete molecular response (clearance), and N-MR indicates no molecular response (ctDNA reduction <50%, no change, or increase).

[0101] Figure 4 shows the clinical AZD8205 activity observed in breast cancer (BC) and ovarian cancer (OvCA) according to Example 8, with preliminary signs of activity in endometrial cancer (EC).

[0102] Figure 5 shows a plot of AZD8205 efficacy measured by percent change in tumor growth versus percent of cells expressing B7-H4 in mouse xenograft studies as described in Example 9.

[0103] Figure 6is a plot of the Youden statistic versus the percentage of cells expressing B7-H4 for the mouse xenograft studies as described in Example 9. As Figure 6 As can be seen in the middle, the Youden statistic starts to plateau at 25% B7-H4.

[0104] Figure 7 The objective response rate (ORR) to AZD8205 for heavily pre-treated subjects according to Example 10 is shown. DETAILED DESCRIPTION

[0105] Definitions

[0106] Unless otherwise defined herein, scientific and technical terms used in this disclosure shall have the meanings that are commonly understood by one of ordinary skill in the art. Further, unless otherwise required by context, singular terms shall include pluralities and plural terms shall include the singular.

[0107] As used herein, "a" or "an" can mean one or more. As used herein, when the word "comprise" is used in a claim, the word "comprise" and its

[0108] The use of the term "or" in the claims is used to mean "and / or" unless explicitly indicated to refer to alternatives only or the alternatives are mutually exclusive, although the disclosure supports a definition that refers to only alternatives and "and / or."

[0109] As used herein, the terms "comprising," "having," "including," "containing," or "including" (and any variations of these terms) are inclusive or open-ended and do not exclude additional, unrecited elements or method steps.

[0110] Throughout this application, the term "about" is used to indicate that a value includes inherent error variations in the method / apparatus used to determine that value, or variations present in study subjects. Generally, the term "about" is intended to cover variability of approximately or less than 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, or 20% or higher ("greater than" or "less than" indicating values), depending on the specific circumstances. In some respects, those skilled in the art will understand the level of variability represented by the term "about" due to the context in which it is used herein. It should also be understood that the use of the term "about" includes specifically enumerated values.

[0111] The use of the term "for example" and its corresponding abbreviation "eg" (whether italic or not) indicates that the specific term referenced is a representative example and aspect of this disclosure and is not intended to be limited to the specific example referenced or cited unless otherwise expressly stated.

[0112] Any range provided herein includes all values ​​within that specific range and values ​​with respect to the endpoints of that range. As used herein, “between” is a range that includes the endpoints of the range. For example, a number between x and y explicitly includes the numbers x and y, as well as any numbers falling within x and y.

[0113] As used herein, the term "antibody" refers to a protein that recognizes and specifically binds to an antigen. Common or conventional mammalian antibodies comprise tetramers, which typically consist of two pairs of identical polypeptide chains, each pair consisting of a "light" chain (typically with a molecular weight of about 25 kDa) and a "heavy" chain (typically with a molecular weight of about 50 kDa–70 kDa). As used herein, the terms "heavy chain" and "light chain" refer to any immunoglobulin polypeptide with a sufficient sequence of variable domains to confer specificity against a target antigen. The amino-terminal portion of each light and heavy chain typically includes a variable domain of about 100 to 110 or more amino acids, which is typically responsible for antigen recognition. The carboxyl-terminal portion of each chain typically defines a constant domain responsible for effector function. Thus, in naturally occurring antibodies, full-length heavy chain immunoglobulin polypeptides include a variable domain (V... H ) and three constant structural domains (C H1 C H2 and C H3 ) and C H1 and C H2 The hinge area between, where V H The domain is located at the amino terminus of the polypeptide and C H3 The domain is at the carboxyl terminus, and the full-length light chain immunoglobulin polypeptide includes a variable domain (V).L ) and a constant domain (C L ), wherein V L domains are at the amino-terminal end and C L domains are at the carboxy-terminal end. However, one of skill in the art will appreciate that the location of the domains in naturally occurring antibodies can be modified in certain antibody-like binding protein formats without loss of antigen binding ability. The classes of human light chains are designated kappa and lambda.

[0114] In some aspects, the light chain constant region is a kappa chain. In some aspects, the light chain constant region is a lambda chain.

[0115] Within full-length light and heavy chains, the variable and constant domains are typically connected by a "J" region of about 12 or more amino acids, with the heavy chain also including a "D" region of about 10 or more amino acids. The variable region of each light / heavy chain pair typically forms an antigen binding site. The variable domains of naturally occurring antibodies typically exhibit the same general structure of a relatively conserved framework region (FR) typically referred to as FR1, FR2, FR3, and FR4, connected by three high variable regions, also called complementarity determining regions or CDRs. CDRs from each of the two chains typically align to form a binding site for an epitope of a specific antigen. From amino-terminus to carboxy-terminus, both light and heavy chain variable domains typically include the domains FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4. Heavy chain complementarity determining regions can be referred to herein as "HCDR" or "CHDR". Similarly, light chain complementarity determining regions can be referred to herein as "LCDR" or "CLDR".

[0116] The term "antibody fragment" refers to a portion or a part of an intact or full-length chain or antibody, typically the target binding or variable region. Examples of antibody fragments include, but are not limited to, F ab , F ab' , F (ab')2 , and F v fragments. As used herein, the term "functional fragment" is generally synonymous with "antibody fragment" and can refer to an antibody fragment such as F v , F ab , F (ab')2 , with respect to an antibody.

[0117] References to numbering of amino acid residues described herein are made according to the EU numbering system (also described in Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD. (1991)).

[0118] As used herein, the term "human antibody" includes antibodies having variable and constant regions substantially corresponding to human germline immunoglobulin sequences. In some aspects, human antibodies are produced in non-human mammals, including but not limited to rodents (such as mice and rats) and lagomorphs (such as rabbits). In other aspects, human antibodies are produced in hybridoma cells. In yet other aspects, human antibodies are produced recombinantly. In some aspects, the bispecific binding protein is a human antibody or a humanized antibody.

[0119] As used herein, the term "antigen" or "target antigen" refers to a molecule or a portion of a molecule that is capable of being recognized and bound by a binding protein of the present disclosure. A target antigen can be used in an animal to generate antibodies that are capable of binding to an epitope of that antigen. A target antigen can have one or more epitopes.

[0120] As used herein, the term "epitope" refers to a region or structural element of an antigen that is recognized and bound by a binding protein of the present disclosure. More precisely, an epitope is the specific structure that is bound by the CDRs of a binding protein. An epitope can include portions of protein structural elements, carbohydrates, or even lipid structures found in membranes. A binding protein is said to specifically bind an antigen when it preferentially recognizes its antigen target in a complex mixture of proteins and / or macromolecules. The term "specifically binds" refers to a binding protein that specifically binds to a molecule or fragment thereof (e.g., an antigen). A binding protein that specifically binds to a molecule or fragment thereof can bind to other molecules with lower affinity, as determined by, for example, immunoassays, BIAcore, or other assays known in the art. Specifically, an antibody or fragment that specifically binds to at least one molecule or fragment thereof can compete away non-specifically bound molecules. The present disclosure specifically encompasses antibodies having multiple specificities (e.g., antibodies specific for two or more discrete antigens). For example, a bispecific antibody can bind to two adjacent epitopes on a single target antigen, or can bind to two different antigens.

[0121] As used herein, the term "native Fc" refers to a molecule comprising the sequence of a non-antigen binding fragment produced by digestion of an antibody or produced by other means, whether in monomeric or multimeric form, and can include a hinge region. The original immunoglobulin source of the native Fc is preferably of human origin, and can be any of the immunoglobulins. Native Fc molecules consist of monomeric polypeptides that can associate, either covalently (i.e., disulfide bonds) and non-covalently, into dimeric or multimeric forms. The number of intermolecular disulfide bonds between monomeric subunits of a native Fc molecule ranges from 1 to 4, depending on the class (e.g., IgG, IgA, and IgE) or subclass (e.g., IgGl, IgG2, IgG3, IgAl, and IgGA2). One example of a native Fc is a disulfide-bonded dimer produced by papain digestion of IgG. As used herein, the term "native Fc" is a generic name for molecules or sequences in monomeric, dimeric, and multimeric forms.

[0122] As used herein, the term "Fc variant" refers to a molecule or sequence that is modified from a native Fc but still includes the binding site for the salvage receptor FcRn (neonatal Fc receptor). Exemplary Fc variants, and their interactions with the salvage receptor, are known in the art. Thus, the term "Fc variant" can include a molecule or sequence that is humanized from a non-human native Fc. In addition, a native Fc includes a region of certain residues that can be removed or mutated to produce an Fc variant to alter structural features or biological activities that are not needed to provide the binding proteins of the present disclosure. Thus, the term "Fc variant" includes molecules or sequences that lack one or more native Fc sites or residues, or in which one or more Fc sites or residues have been modified that affect or are involved in: (1) disulfide bond formation, (2) incompatibility with a selected host cell, (3) N-terminal heterogeneity upon expression in a selected host cell, (4) glycosylation, (5) interaction with complement, (6) binding to Fc receptors other than the salvage receptor, or (7) antibody-dependent cellular cytotoxicity (ADCC).

[0123] As used herein, the term "Fc domain" includes native Fc and Fc variants and sequences as defined above. As with Fc variant and native Fc molecules, the term "Fc domain" includes molecules in monomeric or multimeric form, whether digested from an intact antibody or produced by other means.

[0124] The term "treatment" refers to the administration of a compound or pharmaceutical composition to a subject in order to effect a change or improvement in a disease, disorder, or condition in the subject. The term "treatment" as used herein can refer to therapeutic treatment and prophylactic or preventative measures. Those in need of treatment include those who are suffering from a disease or condition, as well as those in which a disease or condition is to be prevented or prevented.

[0125] The term "dose" means a specified amount of a compound or pharmaceutical agent provided in a single administration or over a specified period of time. In some aspects, a dose can be administered in two or more boluses, tablets, or injections. For example, in some aspects, where subcutaneous administration is required, the desired dose can not be easily accommodated in a single injection. In such aspects, two or more injections can be used to achieve the desired dose. In some aspects, a dose can be administered in two or more injections to minimize injection site reactions in the individual. In other aspects, a compound or pharmaceutical agent is administered by infusion or continuous infusion over an extended period of time. A dose can be expressed as an amount of pharmaceutical agent per hour, day, week, or month.

[0126] The terms "subject," "individual," and "patient" are used interchangeably herein to refer to a mammalian subject. In one aspect, a "subject" is a human, domestic animal, farm animal, sport animal, and zoo animal, such as a human, non-human primate, dog, cat, guinea pig, rabbit, rat, mouse, horse, cow, and the like. In one aspect, the subject is a cynomolgus monkey (Macaca fascicularis). In a preferred aspect, the subject is a human. In the methods of the application, the subject can not have been previously diagnosed with a cancer. Alternatively, the subject can have been previously diagnosed with a cancer. The subject can also be a subject who exhibits risk factors for a disease, or a subject who is asymptomatic for a cancer. The subject can also be a subject who has a cancer or is at risk for a cancer. Thus, in one aspect, the methods of the application can be used to confirm the presence of a cancer in a subject. For example, the subject can have been previously diagnosed with a cancer by alternative means. In one aspect, the subject has been previously administered a cancer therapy.

[0127] The term "therapeutic effect" means the ability to produce a desired effect. A "therapeutically effective dose" or "therapeutic dose" is an amount sufficient to effect a desired clinical outcome (i.e., to obtain therapeutic efficacy). A therapeutically effective dose can be administered in one or more administrations.

[0128] The term“side effect” means a physiological illness and / or condition attributable to treatment other than the desired effect. In some aspects, side effects include injection site reactions, abnormal liver function tests, abnormal kidney function, hepatotoxicity, nephrotoxicity, central nervous system abnormalities, myopathy, and malaise. For example, elevated transaminase levels in serum can indicate hepatotoxicity or abnormal liver function. For example, elevated bilirubin can indicate hepatotoxicity or abnormal liver function. A“disease” or“condition” refers to any condition from which a subject would benefit from treatment using the methods of the disclosure. “Disease” and“condition” are used interchangeably herein and include chronic and acute disorders or diseases, including those pathological conditions that predispose a patient to the disorder in question. In some aspects, the disease is a tumor. In some aspects, the disease is a solid tumor. In some aspects, the disease is a cancer. In some aspects, the cancer is one or more of endometrial cancer, ovarian cancer, breast cancer, biliary tract cancer, colorectal cancer, prostate cancer, cervical cancer, uterine cancer, testicular cancer, bladder cancer, head and neck cancer, melanoma, pancreatic cancer, renal cell carcinoma, and lung cancer.

[0129] The term“administering” as used herein refers to providing, contacting, and / or delivering one or more compounds by any suitable route to obtain a desired effect. Administering can include, but is not limited to, oral, sublingual, parenteral (e.g., intravenous, subcutaneous, intradermal, intramuscular, intraarticular, intraarterial, intrasynovial, intrasternal, intrathecal, intralesional, or intracranial injection), transdermal, topical, buccal, rectal, vaginal, nasal, ocular, via inhalation, and implantation.

[0130] The term“pharmaceutical composition” or“therapeutic composition” as used herein refers to a compound or composition that is capable of inducing a desired therapeutic effect when suitably administered to a subject. In some aspects, the disclosure provides a pharmaceutical composition comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of a binding protein of the disclosure.

[0131] The term“pharmaceutically acceptable carrier” or“physiologically acceptable carrier” as used herein refers to one or more formulation materials suitable for use in effecting or enhancing delivery of one or more binding proteins of the disclosure.

[0132] Methods of treating cancer

[0133] The disclosure relates to a method of treating cancer in a subject comprising administering to the subject an antibody-drug conjugate (ADC) that specifically binds to B7-H4 in an amount of about 0.8 mg / kg to about 4.8 mg / kg. In some aspects, the amount of ADC administered is about 25 mg to about 900 mg per subject. The disclosure also provides compositions (including pharmaceutical compositions) and kits comprising such ADCs.

[0134] In some aspects, the ADC comprises:

[0135] i. an antibody or antigen-binding fragment thereof that binds to a B7-H4 polypeptide, the antibody or antigen-binding fragment thereof comprising:

[0136] a) a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2), a heavy chain CDR3 (HCDR3), a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), and a light chain CDR3 (LCDR3) comprising the amino acid sequences of SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, respectively, or functional variants thereof;

[0137] b) a HCDR1, a HCDR2, a HCDR3, a LCDR1, a LCDR2, and a LCDR3 comprising the amino acid sequences of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6, respectively, or functional variants thereof;

[0138] c) a HCDR1, a HCDR2, a HCDR3, a LCDR1, a LCDR2, and a LCDR3 comprising the amino acid sequences of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, and SEQ ID NO: 18, respectively, or functional variants thereof;

[0139] (d) a HCDR1, a HCDR2, a HCDR3, a LCDR1, a LCDR2, and a LCDR3 comprising the amino acid sequences of SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, and SEQ ID NO: 24, respectively, or functional variants thereof; or

[0140] (e) a HCDR1, a HCDR2, a HCDR3, a LCDR1, a LCDR2, and a LCDR3 comprising the amino acid sequences of SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, and SEQ ID NO: 30, respectively, or functional variants thereof;

[0141] ii. a cleavable linker; and

[0142] iii. a cytotoxic agent.

[0143] B7-H4 (also known as V-set domain-containing T cell activation inhibitor 1, encoded by the VTCN1 gene) is a transmembrane polypeptide of the B7 family of co-stimulatory proteins. B7-H4 is understood to be expressed on the surface of antigen presenting cells for interaction with ligands of immune cells (e.g., T lymphocytes, CD28 is a potential ligand). B7-H4 has been observed to be highly expressed on cells of various cancer types and is considered a tumor-associated antigen. Moreover, B7-H4 expression is not limited to specific cancer types, making it a target antigen for treating a broad spectrum of cancer types.

[0144] A "B7-H4 polypeptide" can comprise the full-length polypeptide sequence of B7-H4 (e.g., SEQ ID NO: 55), or can comprise a fragment of B7-H4 of any length of the full-length polypeptide sequence of B7-H4 (e.g., a polypeptide sequence comprising 5%, 15%, 25%, 35%, 45%, 55%, 65%, 75%, 85%, or 95% of the full-length polypeptide sequence of B7-H4) that comprises an epitope that can bind to (e.g., is bound by) an antibody or antigen-binding fragment of the present disclosure. A B7-H4 polypeptide can comprise a sequence having 75%, 80%, 85%, 90%, or 90% sequence identity to the sequence of SEQ ID NO: 55. Preferably, a B7-H4 polypeptide comprises the sequence of SEQ ID NO: 55.

[0145] In some aspects, an antibody or antigen-binding fragment thereof can bind to a B7-H4 molecule across species, e.g., the antibody or fragment can bind to mouse B7-H4, rat B7-H4, rabbit, human B7-H4, and / or cynomolgus B7-H4. In some aspects, the antibody or fragment can bind to human B7-H4 and cynomolgus B7-H4. In some aspects, the antibody or antigen-binding fragment can also bind to mouse B7-H4.

[0146] In some aspects, an antibody or antigen-binding fragment thereof can specifically bind to B7-H4, e.g., human B7-H4 and cynomolgus B7-H4, but not specifically to human B7-H1, B7-H2, and / or B7-H3.

[0147] In some aspects, the antibody or antigen-binding fragment thereof of the ADC comprises a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2), a heavy chain CDR3 (HCDR3), a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), and a light chain CDR3 (LCDR3) comprising the amino acid sequences of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6, respectively, or functional variants thereof. An antibody or antigen-binding fragment thereof comprising the sequences can be referred to herein as “ZY0EPQ-E02” or “EPQ-E02.”

[0148] In some aspects, the antibody or antigen-binding fragment thereof comprises: a HCDR1, a HCDR2, a HCDR3, a LCDR1, a LCDR2, and a LCDR3 comprising the amino acid sequences of SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, respectively, or functional variants thereof. An antibody or antigen-binding fragment thereof comprising the sequences can be referred to herein as “ZY0EQD-E02” or “EQD-E02.”

[0149] In some aspects, the antibody or antigen-binding fragment thereof of the ADC comprises a HCDR1, a HCDR2, a HCDR3, a LCDR1, a LCDR2, and a LCDR3 comprising the amino acid sequences of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, and SEQ ID NO: 18, respectively, or functional variants thereof. An antibody or antigen-binding fragment thereof comprising the sequences can be referred to herein as “ZY0EOB-F05” or “EOB-F05.”

[0150] In some aspects, the antibody or antigen-binding fragment thereof of the ADC comprises a HCDR1, a HCDR2, a HCDR3, a LCDR1, a LCDR2, and a LCDR3 comprising the amino acid sequences of SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, and SEQ ID NO: 24, respectively, or functional variants thereof. An antibody or antigen-binding fragment thereof comprising the sequences can be referred to herein as “ZY0EO5-E07” or “EO5-E07.”

[0151] In some aspects, the antibody or antigen-binding fragment thereof of the ADC comprises HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 comprising the amino acid sequences of SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, and SEQ ID NO: 30, respectively, or functional variants thereof. The antibody or antigen-binding fragment thereof comprising the sequences can be referred to herein as“ZY0EP0-C07” or“EP0-C07.”

[0152] In some aspects, the antibody or antigen-binding fragment thereof having CDRs with the above disclosed amino acid sequences has advantageous benefits, such as high targeting specificity or binding activity to B7-H4 peptides on the surface of cancer cells.

[0153] Additionally or alternatively, the antibody or antigen-binding fragment thereof described herein can be described by way of its variable heavy chain (VH) and variable light chain (VL).

[0154] In some aspects, the antibody or antigen-binding fragment thereof comprises: a variable heavy (VH) chain and a variable light (VL) chain comprising the amino acid sequences of SEQ ID NO: 45 and SEQ ID NO: 34, respectively, or functional variants thereof; a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 33 and SEQ ID NO: 34, respectively, or functional variants thereof; a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 43 and SEQ ID NO: 34, respectively, or functional variants thereof; a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 46 and SEQ ID NO: 34, respectively, or functional variants thereof; a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 47 and SEQ ID NO: 34, respectively, or functional variants thereof; a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 31 and SEQ ID NO: 32, respectively, or functional variants thereof; a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 35 and SEQ ID NO: 36, respectively, or functional variants thereof; a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 37 and SEQ ID NO: 38, respectively, or functional variants thereof; or a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 39 and SEQ ID NO: 40, respectively, or functional variants thereof.

[0155] For example, in one aspect, the antibody or antigen-binding fragment thereof of the ADC comprises: (i) a variable heavy chain comprising an amino acid sequence having at least 70%, 75%, 80%, 90%, 95%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 31, 33, 35, 37, 39, 43, 45, 46, or 47, or a functional variant thereof; and (ii) a variable light chain comprising an amino acid sequence having at least 70%, 75%, 80%, 90%, 95%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 32, 34, 36, 38, or 40, or a functional variant thereof.

[0156] In some aspects, the antibody or antigen-binding fragment thereof comprises HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 comprising the amino acid sequences of SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, respectively, or functional variants thereof.

[0157] In some aspects, the antibody or antigen-binding fragment thereof comprises a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 45 and SEQ ID NO: 34, respectively, or functional variants thereof. In some aspects, the antibody or antigen-binding fragment thereof comprises a VH chain having an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 45. In some aspects, the antibody or antigen-binding fragment thereof comprises a VL chain having an amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 34.

[0158] In some aspects, the antibody or antigen-binding fragment thereof can include a light chain constant region or fragment thereof in addition to the VH and VL and optional heavy chain constant region or fragment thereof. In some aspects, the light chain constant region is a kappa lambda light chain constant region, e.g., a human kappa constant region or a human lambda constant region.

[0159] In some aspects, the antibody or antigen-binding fragment thereof comprises a heavy chain constant region comprising the amino acid sequence of SEQ ID NO: 41. In some aspects, the antibody or antigen-binding fragment thereof comprises a heavy chain constant region that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 41.

[0160] In some aspects, the antibody or antigen-binding fragment thereof comprises a heavy chain constant region comprising the amino acid sequence of SEQ ID NO: 52. In some aspects, the antibody or antigen-binding fragment thereof comprises a heavy chain constant region that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 52.

[0161] In some aspects, the antibody or antigen-binding fragment thereof comprises a light chain constant region comprising the amino acid sequence of SEQ ID NO: 42. In some aspects, the antibody or antigen-binding fragment thereof comprises a light chain constant region that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 42.

[0162] In some aspects, the antibody or antigen-binding fragment thereof comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 51 and a light chain comprising the amino acid sequence of SEQ ID NO: 44. In some aspects, the antibody or antigen-binding fragment thereof comprises a heavy chain that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 51. In some aspects, the antibody or antigen-binding fragment thereof comprises a light chain that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 44.

[0163] In some aspects, the antibody or antigen-binding fragment thereof comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 48 and a light chain comprising the amino acid sequence of SEQ ID NO: 44. In some aspects, the antibody or antigen-binding fragment thereof comprises a heavy chain that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 48. In some aspects, the antibody or antigen-binding fragment thereof comprises a light chain that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 44.

[0164] In some aspects, the antibody or antigen-binding fragment thereof is a monoclonal antibody. In some aspects, the antibody or antigen-binding fragment thereof is a humanized monoclonal antibody.

[0165] Advantageously, the present disclosure has demonstrated that the antibodies or antigen-binding fragments described above can target a broader spectrum of B7-H4-expressing cells when compared to existing (commercially) available antibodies that target B7-H4. Thus, the present disclosure not only provides antibodies (or antigen-binding fragments) that have affinity and specificity for clinically relevant targets, but also demonstrates unique advantages associated therewith (e.g., unexpected technical effects).

[0166] In some aspects, the cancer is selected from ovarian cancer, biliary tract cancer, breast cancer, pancreatic cancer, prostate cancer, hematological cancer, endometrial cancer, cholangiocarcinoma, NSCLC (squamous carcinoma and / or adenocarcinoma), gastrointestinal cancer (such as gastric cancer and colorectal cancer), and lung cancer.

[0167] In some aspects, the cancer is endometrial cancer.

[0168] In some aspects, the cancer is ovarian cancer.

[0169] In some aspects, the cancer is biliary tract cancer.

[0170] In some aspects, the cancer is breast cancer.

[0171] In some aspects, the cancer is breast cancer selected from hormone receptor positive (HR+) breast cancer, human epidermal growth factor receptor 2 positive (HER2+) breast cancer, and triple negative breast cancer (TNBC).

[0172] In some aspects, the cancer is homologous recombination deficient (HRD) cancer.

[0173] In some aspects, the cancer comprises one or more cells having a mutation in an HRD gene selected from BRCA1, BRCA2, ATM, BRIP1, BARD1, CDK12, CHEK1, CHEK2, FANCL, PALB2, PPP2R2A, RAD51B, RAD51C, RAD51D, and RAD54L. In some aspects, the mutation in the HRD gene is selected from BRCA1, BRCA2, and ATM.

[0174] In some aspects, the antibody or antigen-binding fragment thereof of the ADC binds to an OVCAR4 cell line. In some aspects, the antibody or antigen-binding fragment thereof described herein can bind to an OVCAR4 cell line and / or a CHO cell line (e.g., which can lack an exogenous nucleic acid encoding B7-H4). For example, the antibody or antigen-binding fragment thereof binds to B7-H4 (e.g., a B7-H4 epitope) of an OVCAR4 cell line and / or a CHO cell line (e.g., which can lack an exogenous nucleic acid encoding B7-H4).

[0175] In some aspects, the antibody or antigen-binding fragment thereof binds to an OVCAR4 cell line and / or a CHO cell line (e.g., which can lack an exogenous nucleic acid encoding B7-H4) with higher affinity when compared to one or more antibodies selected from E Biosciences 14-5949 anti-human B7H4 mouse IgG, US biological B0000-35B anti-human B7H4 mouse IgG, R and D systems AF2514 anti-mouse B7H4 goat IgGl, Sigma SAB2500141 anti-B7H4 goat IgGl, Isotype 1 CAT004 SP06-003, Isotype 2 R and D normal goat IgG control (AB-108C), AdD serotec MCA2632, Epitomics 2516-1, eBiosciences 145972-82, eBioscience 145970-85, or a combination thereof. Affinity (e.g., binding affinity) can be measured by any suitable method of measuring binding affinity described herein.

[0176] The OVCAR4 cell line is a human ovarian carcinoma cell line, and the CHO cell line is an epithelial cell line derived from Chinese hamster ovary, and are widely available.

[0177] In some aspects, at least about 25% of the cancer cells are B7-H4 positive cells. In some aspects, at least about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, or 75% of the cancer cells are B7-H4 positive cells. In some aspects, a B7-H4 positive cell is a cell that expresses B7-H4 on its cell surface. In some aspects, the percentage of B7-H4 positive cells is determined by analyzing a sample (e.g., a biopsy of cancer tissue from a subject). In some aspects, the percentage of B7-H4 positive cells is determined using an antibody for detecting B7-H4.

[0178] In some aspects, the percentage of cells that are B7-H4 positive in a cancer is determined by immunohistochemistry (IHC). IHC is a laboratory method that uses antibodies to detect antigens (biomarkers) in a tissue sample. In aspects, the antibodies are attached to enzymes or fluorescent dyes. After the antibodies bind to antigens in the tissue sample, the enzymes or dyes are activated, and then the antigens can be detected using a microscope or other visualization device.

[0179] In some aspects, the antibody or antigen-binding fragment thereof is linked to the cytotoxic agent by a linker. In some aspects, the antibody or antigen-binding fragment thereof is conjugated to the cytotoxic agent by a linker. As used herein, “conjugated” means linked via a covalent or ionic bond. In some aspects, the linker is attached (e.g., conjugated) to an amino residue, e.g., an amino acid of the antibody or antigen-binding fragment of the ADC, in a cleavable manner. In some aspects, the cleavable linker of the ADC is an mp-PEG8-val-ala linker.

[0180] The cytotoxic agent can be referred to herein as an “agent” or “active agent.” In some aspects, the cytotoxic agent is a drug. The cytotoxic agent or cytotoxin can be any molecule known in the art to be capable of inhibiting or preventing the function of cells and / or causing destruction of cells (cell death) and / or exerting an anti-tumor / anti-proliferative effect. Many classes of cytotoxic agents are known to have potential utility in ADC molecules. These include, but are not limited to, topoisomerase I inhibitors, amanitins, auristatins, daunomycin, doxorubicin, duocarmycins, dolastatins, enediynes, lexitropsins, taxanes, puromycins, maytansinoids, vinca alkaloids, tubulysins, and pyrrolobenzo-diazepines (PBDs). Examples of such cytotoxic agents are AFP, MMAF, MMAE, AEB, AEVB, auristatin E, paclitaxel, docetaxel, CC-1065, SN-38, topotecan, morpholino-doxorubicin, rhizoxin, cyano-morpholino-doxorubicin, dolastatin-10, echinomycin, combretatstatin, chalicheamicin, maytansine, DM-1, vinblastine, methotrexate, and netropsin, as well as derivatives and analogs thereof.

[0181] In some aspects, the cytotoxic agent is a drug. In some aspects, the cytotoxic agent of the ADC is a topoisomerase inhibitor, a tubulin inhibitor derivative, a pyrrolobenzo-diazepine, or a combination thereof. In some aspects, the cytotoxic agent is one of the cytotoxic agents disclosed in U.S. Patent Application Publication Nos. US 2022-0211863 and US 2017-0291955, both of which are incorporated by reference herein in their entireties.

[0182] In preferred aspects, the cytotoxic agent is a topoisomerase inhibitor. Topoisomerase inhibitors are compounds that block the action of topoisomerase enzymes (topoisomerase I and II), which are enzymes that control structural changes in DNA by catalyzing the breakage and rejoining of phosphodiester bonds in the DNA strand during normal cell cycle. The topoisomerase I inhibitor can be, for example, SG3932, SG4010, SG4057, or SG4052 provided below.

[0183] The present disclosure provides a cytotoxic agent (drug unit) of an ADC that is linked to a ligand unit of the ADC via a linker (linker unit) described above. The ligand unit is preferably an antibody or antigen-binding fragment thereof. In some aspects, the present disclosure provides a conjugate comprising the following topoisomerase inhibitor derivative (A , drug unit):

[0184]

[0185] In some aspects, the cytotoxic agent is a topoisomerase inhibitor, and wherein the cleavable linker of the ADC and the cytotoxic agent together comprise a compound of Formula I:

[0186]

[0187] and salts and solvates thereof. R L is a cleavable linker described above.

[0188] In some aspects, the linker and the cytotoxic agent together comprise one of the following compounds:

[0189] (SG3932); (SG4010); (SG4057); or (SG4052).

[0190] In some aspects, the linker and the cytotoxic agent together are compound SG3932.

[0191] The cytotoxic agent is typically linked to or “loaded onto” the antibody or antigen-binding fragment. The drug loading (p) is the average number of drug agents per antibody or antigen-binding fragment (e.g., ligand unit).

[0192] The average number of drugs per antibody in an ADC preparation from a conjugation reaction can be characterized by routine means such as UV, reverse phase HPLC, HIC, mass spectrometry, ELISA assays, and electrophoresis. The quantitative distribution of p for an ADC can also be determined. By ELISA, the average value of p in a particular ADC preparation can be determined (Hamblett et al., (2004) Clin. Cancer Res. 10:7063-7070; Sanderson et al., (2005) Clin. Cancer Res. 11 :843-852). However, the distribution of p (drug) values cannot be resolved by the antibody-antigen binding and detection limitations of ELISA. In addition, ELISA assays for detecting antibody-drug conjugates cannot determine where the drug moiety is attached to the antibody, such as the heavy or light chain fragment, or specific amino acid residues. In some cases, separation, purification, and characterization of homogenous ADCs, where p is a specific value from ADCs with other drug loads, can be achieved by means such as reverse phase HPLC or electrophoresis. Such techniques are also applicable to other types of conjugates.

[0193] Typically, less than the theoretical maximum amount of drug moieties are conjugated to the antibody during the conjugation reaction. The antibody can contain, for example, a number of lysine residues that do not react with the drug linker. Only the most reactive lysine groups can react with the amine-reactive linker reagent. In addition, only the most reactive cysteine thiol groups can react with the thiol-reactive linker reagent. Generally, the antibody does not contain many, if any, free and reactive cysteine thiol groups that can be attached to drug moieties. Most of the cysteine thiol residues in the compound antibody exist as disulfide bridges and must be reduced with a reducing agent such as dithiothreitol (DTT) or TCEP under partial or complete reducing conditions. The loading of the ADC (drug / antibody ratio) can be controlled in several different ways, including: (i) limiting the molar excess of drug linker relative to the antibody, (ii) limiting the conjugation reaction time or temperature, and (iii) partial or limiting reducing conditions for cysteine thiol modification.

[0194] Certain antibodies have reducing interchain disulfides, i.e., cysteine bridges. By treatment with a reducing agent such as DTT (dithiothreitol), the antibody can be rendered reactive for conjugation with linker reagents. Thus, in theory, each cysteine bridge will form two reactive thiol nucleophiles. Additional nucleophilic groups can be introduced into the antibody by reaction of lysines with 2-iminothiolane (Traut's reagent), resulting in the conversion of amines to thiols. Reactive thiol groups can be introduced into an antibody (or fragment thereof) by engineering one, two, three, four, or more cysteine residues (e.g., making a mutant antibody comprising one or more non-native cysteine residues). US 7521541 teaches engineering antibodies by introducing reactive cysteine amino acids.

[0195] Cysteine amino acids can be engineered at the reactive site of the antibody and do not form intra- or intermolecular disulfide bonds (Junutula et al., 2008b Nature Biotech., 26(8):925-932; Dornan et al., (2009) Blood 114(13):2721-2729; US 7521541; US 7723485; WO2009 / 052249). The engineered cysteine thiols can react with drug linkers of the present application having thiol-reactive electrophilic groups such as maleimides or a-haloamides to form ADCs with the cysteine engineered antibodies. Thus, the location of the drug units can be designed, controlled, and known. Drug loading can be controlled because the engineered cysteine thiol groups typically react with drug-linker reagents in high yield. IgG antibodies are engineered to introduce cysteine amino acids by substitution at a single site on either the heavy chain or light chain, resulting in two new cysteines on a symmetric antibody.

[0196] In some aspects, the average number of agents per antibody (or antigen binding fragment thereof) is in the range of 1 to 20. In some aspects, the range is selected from 1 to 10, 2 to 10, 2 to 8, 2 to 6, and 4 to 10. In some aspects, there is one agent per antibody (or antigen binding fragment thereof). In some aspects, the number of agents per antibody (or antigen binding fragment thereof) can be expressed as a ratio of agents (i.e., drugs) to antibody. This ratio is referred to as the drug-to-antibody ratio (DAR). The DAR is the average number of drugs (i.e., agents) attached to each antibody. In some aspects of the disclosure, the DAR is in the range of 1 to 20. In some aspects, the range of the DAR is selected from 1 to 10, 2 to 10, 2 to 8, 2 to 6, and 4 to 10. In some aspects, the DAR is between about 1 and about 8. In particular aspects of the disclosure, the DAR is about 8. In particular aspects of the disclosure, the DAR is 8.

[0197] In some aspects, the ADC is AZD8205. AZD8205 is an anti-B7H4 Ab conjugated to a topoisomerase inhibitor (TOPO) payload. Specifically, AZD8205 has a heavy chain of SEQ ID NO: 51 and a light chain of SEQ ID NO: 44, and is conjugated to SG3932. Typically, the DAR is about 8. In some aspects, AZD8205 targets ovarian cancer and cholangiocarcinoma (CCA). In some aspects, AZD8205 targets triple negative breast cancer (TNBC). In some aspects, AZD8205 targets endometrial cancer.

[0198] In some aspects, the amount of ADC administered is about 0.5 mg / kg to about 8 mg / kg. In some aspects, the amount of ADC administered is about 0.8 mg / kg to about 4.8 mg / kg. In some aspects, the amount of ADC administered is about 1.2 mg / kg to about 3.6 mg / kg. In some aspects, the amount of ADC administered is about 1.4 mg / kg to about 3.2 mg / kg. In some aspects, the amount of ADC administered is about 1.6 mg / kg to about 2.4 mg / kg. In some aspects, the amount of ADC administered is about 1.8 mg / kg to about 2.2 mg / kg.

[0199] In some aspects, the amount of ADC administered is about 0.5 mg / kg, about 0.8 mg / kg, about 1.0 mg / kg, about 1.2 mg / kg, about 1.4 mg / kg, about 1.6 mg / kg, about 1.8 mg / kg, about 2.0 mg / kg, about 2.2 mg / kg, about 2.4 mg / kg, about 2.6 mg / kg, about 2.8 mg / kg, about 3.0 mg / kg, about 3.2 mg / kg, about 3.6 mg / kg, about 4.0 mg / kg, about 4.4 mg / kg, about 4.8 mg / kg, about 5.5 mg / kg, about 6.0 mg / kg, about 7.0 mg / kg, or about 8 mg / kg.

[0200] In some aspects, the amount of ADC administered is about 1.2 mg / kg, about 1.6 mg / kg, about 1.8 mg / kg, about 2.0 mg / kg, about 2.2 mg / kg, about 2.4 mg / kg, or about 3.2 mg / kg.

[0201] In some aspects, the amount of ADC administered is about 1.6 mg / kg. In some aspects, the amount of ADC administered is about 2.0 mg / kg. In some aspects, the amount of ADC administered is about 2.4 mg / kg. In some aspects, the amount of ADC administered is about 3.2 mg / kg.

[0202] In some aspects, the amount of ADC administered is about 25 mg to about 900 mg. In some aspects, the amount of ADC administered is about 40 mg to about 600 mg. In some aspects, the amount of ADC administered is about 60 mg to about 450 mg. In some aspects, the amount of ADC administered is about 80 mg to about 300 mg. In some aspects, the amount of ADC administered is about 100 mg to about 240 mg. In some aspects, the amount of ADC administered is about 120 mg to about 200 mg. In some aspects, the amount of ADC administered is about 140 mg to about 180 mg.

[0203] In some aspects, the amount of ADC administered is about 25 mg, about 40 mg, about 60 mg, about 80 mg, about 100 mg, about 120 mg, about 140 mg, about 160 mg, about 180 mg, about 200 mg, about 220 mg, about 240 mg, about 260 mg, about 280 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 600 mg, about 750 mg, or about 900 mg.

[0204] In some aspects, the amount of ADC administered is about 80 mg, about 100 mg, about 120 mg, about 140 mg, about 160 mg, about 180 mg, about 200 mg, about 240 mg, about 280 mg, or about 320 mg.

[0205] In some aspects, the amount of ADC administered is about 120 mg. In some aspects, the amount of ADC administered is about 160 mg. In some aspects, the amount of ADC administered is about 200 mg.

[0206] In some aspects, the ADC is administered to the subject once every three weeks (Q3W). In some aspects, the ADC is administered to the subject once a week (Q1W), once every two weeks (Q2W), once every four weeks (Q4W), once every five weeks (Q5W), or once every six weeks (Q6W).

[0207] In some aspects, the disclosure further provides a method of treating a cancer in a subject, the method comprising administering to the subject an antibody-drug conjugate (ADC) that specifically binds to B7-H4 in an amount of about 0.8 mg / kg to about 4.8 mg / kg. In some aspects, the ADC comprises: i. an antibody or antigen-binding fragment thereof that binds to a B7-H4 polypeptide, the antibody or antigen-binding fragment thereof comprising: a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2), a heavy chain CDR3 (HCDR3), a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), and a light chain CDR3 (LCDR3) comprising the amino acid sequences of SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, respectively, or functional variants thereof; and ii. a cleavable linker and a cytotoxic agent conjugated to the antibody or antigen-binding fragment thereof having the following formula:

[0208] (SG3932).

[0209] In some aspects, the antibody or antigen-binding fragment thereof comprises a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 45 and SEQ ID NO: 34, respectively, or functional variants thereof.

[0210] In some aspects, the cancer is selected from ovarian cancer, biliary tract cancer, breast cancer, pancreatic cancer, prostate cancer, hematological cancer, endometrial cancer, cholangiocarcinoma, NSCLC (squamous carcinoma and / or adenocarcinoma), gastrointestinal cancer (such as gastric cancer and colorectal cancer), and lung cancer.

[0211] In some aspects, the cancer is endometrial cancer.

[0212] In some aspects, the cancer is ovarian cancer.

[0213] In some aspects, the cancer is biliary tract cancer.

[0214] In some aspects, the cancer is breast cancer.

[0215] In some aspects, the cancer is breast cancer selected from hormone receptor positive (HR+) breast cancer, human epidermal growth factor receptor 2 positive (HER2+) breast cancer, and triple negative breast cancer (TNBC). In some aspects, the cancer is a homologous recombination deficient (HRD) cancer.

[0216] In some aspects, the cancer comprises one or more cells having a mutation in an HRD gene selected from BRCA1, BRCA2, ATM, BRIP1, BARD1, CDK12, CHEK1, CHEK2, FANCL, PALB2, PPP2R2A, RAD51B, RAD51C, RAD51D, and RAD54L. In some aspects, the mutation in the HRD gene is selected from BRCA1, BRCA2, and ATM.

[0217] In some aspects, the amount of ADC administered is about 0.5 mg / kg to about 8 mg / kg. In some aspects, the amount of ADC administered is about 0.8 mg / kg to about 4.8 mg / kg. In some aspects, the amount of ADC administered is about 1.2 mg / kg to about 3.6 mg / kg. In some aspects, the amount of ADC administered is about 1.4 mg / kg to about 3.2 mg / kg. In some aspects, the amount of ADC administered is about 1.6 mg / kg to about 2.4 mg / kg. In some aspects, the amount of ADC administered is about 1.8 mg / kg to about 2.2 mg / kg.

[0218] In some aspects, the amount of ADC administered is about 0.5 mg / kg, about 0.8 mg / kg, about 1.0 mg / kg, about 1.2 mg / kg, about 1.4 mg / kg, about 1.6 mg / kg, about 1.8 mg / kg, about 2.0 mg / kg, about 2.2 mg / kg, about 2.4 mg / kg, about 2.6 mg / kg, about 2.8 mg / kg, about 3.0 mg / kg, about 3.2 mg / kg, about 3.6 mg / kg, about 4.0 mg / kg, about 4.4 mg / kg, about 4.8 mg / kg, about 5.5 mg / kg, about 6.0 mg / kg, about 7.0 mg / kg, or about 8 mg / kg.

[0219] In some aspects, the amount of ADC administered is about 1.2 mg / kg, about 1.6 mg / kg, about 1.8 mg / kg, about 2.0 mg / kg, about 2.2 mg / kg, about 2.4 mg / kg, or about 3.2 mg / kg.

[0220] In some aspects, the amount of ADC administered is about 1.6 mg / kg. In some aspects, the amount of ADC administered is about 2.0 mg / kg. In some aspects, the amount of ADC administered is about 2.4 mg / kg. In some aspects, the amount of ADC administered is about 3.2 mg / kg.

[0221] In some aspects, the amount of ADC administered is about 25 mg to about 900 mg. In some aspects, the amount of ADC administered is about 40 mg to about 600 mg. In some aspects, the amount of ADC administered is about 60 mg to about 450 mg. In some aspects, the amount of ADC administered is about 80 mg to about 300 mg. In some aspects, the amount of ADC administered is about 100 mg to about 240 mg. In some aspects, the amount of ADC administered is about 120 mg to about 200 mg. In some aspects, the amount of ADC administered is about 140 mg to about 180 mg.

[0222] In some aspects, the amount of ADC administered is about 25 mg, about 40 mg, about 60 mg, about 80 mg, about 100 mg, about 120 mg, about 140 mg, about 160 mg, about 180 mg, about 200 mg, about 220 mg, about 240 mg, about 260 mg, about 280 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 600 mg, about 750 mg, or about 900 mg.

[0223] In some aspects, the amount of ADC administered is about 80 mg, about 100 mg, about 120 mg, about 140 mg, about 160 mg, about 180 mg, about 200 mg, about 240 mg, about 280 mg, or about 320 mg.

[0224] In some aspects, the amount of ADC administered is about 120 mg. In some aspects, the amount of ADC administered is about 160 mg. In some aspects, the amount of ADC administered is about 200 mg.

[0225] In some aspects, the ADC is administered to the subject once every three weeks (Q3W). In some aspects, the ADC is administered to the subject once every week (Q1W), once every two weeks (Q2W), once every four weeks (Q4W), once every five weeks (Q5W), or once every six weeks (Q6W).

[0226] In some aspects, the disclosure further provides a method of treating a cancer in a subject, the method comprising administering to the subject an antibody-drug conjugate (ADC) that specifically binds to B7-H4, the ADC comprising:

[0227] i. an antibody or antigen binding fragment thereof that binds to a B7-H4 polypeptide, the antibody or antigen binding fragment thereof comprising:

[0228] f) comprise HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, respectively, comprising the amino acid sequences of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, or functional variants thereof;

[0229] g) comprise HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, respectively, comprising the amino acid sequences of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO:4, SEQ ID NO: 5, and SEQ ID NO: 6, or functional variants thereof;

[0230] h) comprise HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, respectively, comprising the amino acid sequences of SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, and SEQ ID NO: 18, or functional variants thereof;

[0231] i) comprise HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, respectively, comprising the amino acid sequences of SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, and SEQ ID NO: 24, or functional variants thereof; or

[0232] j) comprise HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, respectively, comprising the amino acid sequences of SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, and SEQ ID NO: 30, or functional variants thereof;

[0233] ii. a cleavable linker; and

[0234] iii. a cytotoxic agent;

[0235] wherein at least about 25% of the cancer cells in the subject are B7-H4 positive cells.

[0236] Described herein are cleavable linkers and cytotoxic agents for use in the ADCs in the methods. Described herein are cancers that can be treated by the methods.

[0237] In some aspects, at least about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, or 75% of the cancer cells are B7-H4 positive cells. In some aspects, a B7-H4 positive cell is a cell that expresses B7-H4 on its cell surface. In some aspects, the percentage of B7-H4 positive cells is determined by analyzing a sample (e.g., a biopsy of cancer tissue from a subject). In some aspects, an antibody for detecting B7-H4 is used to determine the percentage of B7-H4 positive cells.

[0238] In aspects of the methods, the ADC is administered in an amount of about 0.8 mg / kg to about 4.8 mg / kg. In aspects, the amount of ADC administered is about 0.8 mg / kg, about 1.6 mg / kg, about 2.4 mg / kg, about 3.2 mg / kg, about 3.6 mg / kg, or about 4.8 mg / kg. In aspects, the amount of ADC administered is about 1.6 mg / kg. In aspects, the amount of ADC administered is about 2.4 mg / kg. In aspects, the amount of ADC administered is about 3.2 mg / kg.

[0239] In other aspects of the methods, the ADC is administered in an amount as disclosed herein.

[0240] In some aspects, the ADC is administered to the subject once every three weeks (Q3W). In some aspects, the ADC is administered to the subject once every week (Q1W), once every two weeks (Q2W), once every four weeks (Q4W), once every five weeks (Q5W), or once every six weeks (Q6W).

[0241] In aspects of any of the methods or uses described herein, the subject has previously received treatment with another chemotherapeutic agent. In some aspects, the subject has previously received treatment with a platinum-based chemotherapeutic agent. In some aspects, the platinum-based chemotherapeutic agent is cisplatin, carboplatin, oxaliplatin, nedaplatin, lobaplatin, or heptaplatin.

[0242] Pharmaceutical compositions

[0243] In some aspects, the disclosure further provides a pharmaceutical composition comprising an antibody-drug conjugate (ADC) that specifically binds to B7-H4, the ADC being administered in an amount of about 0.8 mg / kg to about 4.8 mg / kg. In some aspects, the ADC comprises: i. an antibody or antigen-binding fragment thereof that binds to a B7-H4 polypeptide, the antibody or antigen-binding fragment thereof comprising: a heavy chain CDR1 (HCDR1), a heavy chain CDR2 (HCDR2), a heavy chain CDR3 (HCDR3), a light chain CDR1 (LCDR1), a light chain CDR2 (LCDR2), and a light chain CDR3 (LCDR3) comprising the amino acid sequences of SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, respectively, or functional variants thereof; and ii. a cleavable linker and a cytotoxic agent conjugated to the antibody or antigen-binding fragment thereof having the following formula:

[0244] (SG3932).

[0245] In some aspects, the amount of ADC administered is about 0.5 mg / kg to about 8 mg / kg. In some aspects, the amount of ADC administered is about 0.8 mg / kg to about 4.8 mg / kg. In some aspects, the amount of ADC administered is about 1.2 mg / kg to about 3.6 mg / kg. In some aspects, the amount of ADC administered is about 1.4 mg / kg to about 3.2 mg / kg. In some aspects, the amount of ADC administered is about 1.6 mg / kg to about 2.4 mg / kg. In some aspects, the amount of ADC administered is about 1.8 mg / kg to about 2.2 mg / kg.

[0246] In some aspects, the amount of ADC administered is about 0.5 mg / kg, about 0.8 mg / kg, about 1.0 mg / kg, about 1.2 mg / kg, about 1.4 mg / kg, about 1.6 mg / kg, about 1.8 mg / kg, about 2.0 mg / kg, about 2.2 mg / kg, about 2.4 mg / kg, about 2.6 mg / kg, about 2.8 mg / kg, about 3.0 mg / kg, about 3.2 mg / kg, about 3.6 mg / kg, about 4.0 mg / kg, about 4.4 mg / kg, about 4.8 mg / kg, about 5.5 mg / kg, about 6.0 mg / kg, about 7.0 mg / kg, or about 8 mg / kg.

[0247] In some aspects, the amount of ADC administered is about 1.2 mg / kg, about 1.6 mg / kg, about 1.8 mg / kg, about 2.0 mg / kg, about 2.2 mg / kg, about 2.4 mg / kg, or about 3.2 mg / kg.

[0248] In some aspects, the amount of ADC administered is about 1.6 mg / kg. In some aspects, the amount of ADC administered is about 2.0 mg / kg. In some aspects, the amount of ADC administered is about 2.4 mg / kg. In some aspects, the amount of ADC administered is about 3.2 mg / kg.

[0249] In some aspects, the amount of ADC administered is about 25 mg to about 900 mg. In some aspects, the amount of ADC administered is about 40 mg to about 600 mg. In some aspects, the amount of ADC administered is about 60 mg to about 450 mg. In some aspects, the amount of ADC administered is about 80 mg to about 300 mg. In some aspects, the amount of ADC administered is about 100 mg to about 240 mg. In some aspects, the amount of ADC administered is about 120 mg to about 200 mg. In some aspects, the amount of ADC administered is about 140 mg to about 180 mg.

[0250] In some aspects, the amount of ADC administered is about 25 mg, about 40 mg, about 60 mg, about 80 mg, about 100 mg, about 120 mg, about 140 mg, about 160 mg, about 180 mg, about 200 mg, about 220 mg, about 240 mg, about 260 mg, about 280 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 600 mg, about 750 mg, or about 900 mg.

[0251] In some aspects, the amount of ADC administered is about 80 mg, about 100 mg, about 120 mg, about 140 mg, about 160 mg, about 180 mg, about 200 mg, about 240 mg, about 280 mg, or about 320 mg.

[0252] In some aspects, the amount of ADC administered is about 120 mg. In some aspects, the amount of ADC administered is about 160 mg. In some aspects, the amount of ADC administered is about 200 mg.

[0253] In some aspects of the use of the pharmaceutical compositions disclosed herein to treat cancer, the ADC is administered to the subject once every three weeks (Q3W). In some aspects of the use of the pharmaceutical compositions disclosed herein to treat cancer, the ADC is administered to the subject once a week (Q1W), once every two weeks (Q2W), once every four weeks (Q4W), once every five weeks (Q5W), or once every six weeks (Q6W).

[0254] In some aspects of the use of the pharmaceutical compositions disclosed herein to treat cancer, the subject having cancer has previously received treatment with another chemotherapeutic agent. In some aspects, the subject has previously received treatment with a platinum-based chemotherapeutic agent. In some aspects, the platinum-based chemotherapeutic agent is cisplatin, carboplatin, oxaliplatin, nedaplatin, lobaplatin, or heptaplatin.

[0255] In some aspects, the antibody or antigen-binding fragment of the ADC comprises a VH chain and a VL chain comprising the amino acid sequences of SEQ ID NO: 45 and SEQ ID NO: 34, respectively, or a functional variant thereof. In some aspects, the antibody or antigen-binding fragment of the ADC comprises a VH chain comprising an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO: 45, and a VL chain comprising an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO: 34.

[0256] In some aspects, the pharmaceutical compositions disclosed herein can be formulated with a pharmaceutically acceptable carrier, excipient, or stabilizer. In certain aspects, such pharmaceutical compositions are suitable for administration to a human or non-human animal via any one or more of the administration routes using methods known in the art. The term "pharmaceutically acceptable carrier" means one or more nontoxic materials that do not interfere with the effectiveness of the biological activity of the active ingredients. Such preparations can routinely contain salts, buffering agents, preservatives, compatible carriers, and optionally other therapeutic agents. Such pharmaceutically acceptable formulations can also include compatible solid or liquid fillers, diluents, or encapsulating substances which are suitable for administration to humans. Other contemplated carriers, excipients, and / or additives that can be utilized in the formulations described herein include, for example, flavoring agents, antimicrobial agents, sweeteners, antioxidants, antistatic agents, lipids, protein excipients such as serum albumin, gelatin, casein, salt-forming counterions such as sodium, and the like. These and additional known pharmaceutical carriers, excipients, and / or additives suitable for use in the formulations described herein are known in the art, for example, as described in "Remington: The Science & Practice of Pharmacy", 21stEd., Lippincott Williams & Wilkins, (2005) and "Physician's Desk Reference", 60thEd., Medical Economics, Montvale, N.J. (2005). The pharmaceutically acceptable carrier can be selected for the desired or required mode of administration, solubility, and / or stability.

[0257] The present disclosure further provides the pharmaceutical compositions as described above for use in treating cancer. In some aspects, the cancer comprises cancer cells that express B7-H4. In some aspects, the cancer is selected from the group consisting of ovarian cancer, biliary tract cancer, breast cancer, pancreatic cancer, prostate cancer, hematological cancer, endometrial cancer, cholangiocarcinoma, NSCLC (squamous carcinoma and / or adenocarcinoma), gastrointestinal cancer (such as gastric cancer and colorectal cancer), and lung cancer.

[0258] In some aspects, the cancer is cholangiocarcinoma.

[0259] In some aspects, the cancer is endometrial cancer.

[0260] In some aspects, the cancer is ovarian cancer.

[0261] In some aspects, the cancer is breast cancer. In some aspects, the cancer is a breast cancer selected from the group consisting of hormone receptor positive (HR+) breast cancer, human epidermal growth factor receptor 2 positive (HER2+) breast cancer, and triple negative breast cancer (TNBC). In some aspects, the cancer is a homologous recombination deficient (HRD) cancer. In some aspects, the cancer comprises one or more cells having a mutation in an HRD gene selected from the group consisting of BRCA1, BRCA2, ATM, BRIP1, BARD1, CDK12, CHEK1, CHEK2, FANCL, PALB2, PPP2R2A, RAD51B, RAD51C, RAD51D, and RAD54L. In some aspects, the mutation in the HRD gene is selected from the group consisting of BRCA1, BRCA2, and ATM.

[0262] Kits

[0263] In some aspects, the present disclosure further provides a kit comprising any of the above pharmaceutical compositions. In some aspects, the kit comprises instructions for administering the pharmaceutical composition. In some aspects, the kit comprises an additional anti-cancer agent as described herein.

[0264] Antibody or antigen-binding fragment sequences of the ADC

[0265] Aspects of the present disclosure include antibodies or antigen-binding fragments in the form of DuetMabs that bind to B7-H4 polypeptides created using the sequences in Table 1 below. The CDRs in Table 1 were determined based on the system in Kabat.

[0266] Table 1. Antibody or antigen-binding fragment sequences of the ADC

[0267]

[0268]

[0269]

[0270]

[0271]

[0272]

[0273]

[0274]

[0275] All references cited herein, including patents, patent applications, papers, textbooks, and the like, and references cited therein, and their documents not yet cited, are hereby incorporated by reference in their entireties.

[0276] Many aspects of the present disclosure are described for purposes of illustration and not by way of limitation.

[0277] Examples

[0278] The following examples are illustrative of specific aspects of the present disclosure and various uses thereof. They are set forth to illustrate, but not to limit, the scope of the present disclosure.

[0279] AZD8205 is provided in embodiments for use in therapy. AZD8205 is an ADC targeting B7-H4 (VTCN1) consisting of a human anti-B7-H4 antibody (“INT016”) conjugated to a TOP1 i warhead (“AZ14170132”) via a cleavable maleimide-PEG8-valine-alanine linker. AZD8205 has a DAR of about 8. The antibody component of AZD8205 binds to B7-H4 on the cell surface and facilitates internalization of the ADC and its trafficking to the lysosome. Enzymatic cleavage of the linker releases the TOP1 i warhead, which traps TOP1 on DNA and leads to the formation of double-stranded DNA breaks. If un-repaired, the double-stranded breaks ultimately lead to programmed cell death.

[0280] Preclinical pharmacology studies indicate that the primary mechanism of action of AZD8205 is the intracellular delivery of the TOP1 i warhead into B7-H4 positive tumor cells. In vitro assays demonstrate that AZD8205 specifically binds to B7-H4 expressing tumor cells but not to B7-H4 negative cells and is cytotoxic to B7-H4 expressing tumor cells. In vivo pharmacodynamic studies using human tumor xenograft mouse models show that AZD8205 accumulates in a dose-dependent manner and there is an increase in positive staining for γH2AX foci (an indicator of DNA double-strand breaks), elevated cleaved caspase-3 and a general decrease in epithelial cell density over time, which is consistent with the mechanism of action of its TOP1 i warhead, binding to B7-H4 on tumor cells resulting in DNA damage and apoptotic cell death.

[0281] Subsequent in vivo efficacy studies of AZD8205 in triple negative breast cancer human tumor cell line-derived xenograft models and patient-derived xenograft models further demonstrated that single IV administration of AZD8205 produced target engagement and dose-dependent anti-tumor efficacy. In a study evaluating patient-derived xenograft models of triple negative breast cancer (N=26), a single IV dose of 1.25 mg / kg or 3.5 mg / kg resulted in effective tumor regression (30% or greater reduction in tumor volume from baseline) in 12 of 26 models (46%) and 18 of 26 models (69%), respectively. To elucidate the relationship between B7-H4 expression and AZD8205 response, immunohistochemistry and image analysis techniques were used to quantify B7-H4 expression in each patient-derived xenograft model. A statistically significant correlation between B7-H4 expression and anti-tumor efficacy was observed following treatment with 1.25 mg / kg AZD8205, indicating that elevated B7-H4 levels are associated with AZD8205 response.

[0282] Example 1: Starting dose adjustment

[0283] AZD8205 monotherapy is being investigated at dose levels ranging from 0.8 mg / kg to 3.2 mg / kg Q3W intravenously as described in the examples below. The recommended starting dose of 0.8 mg / kg for the single agent AZD8205 is based on the results of a good laboratory practice (GLP) toxicology study in cynomolgus monkeys, scaling of pharmacokinetics (PK) from cynomolgus monkey to human, and FDA regulatory guidelines. Dose increments and a clinically effective dose range (0.8 mg / kg to 2.4 mg / kg) were determined and modelled exposures were estimated. In a GLP study in cynomolgus monkeys, the highest non-severely toxic dose (HNSTD) of AZD8205 was identified as 15 mg / kg every 3 weeks (Q3W) for a total of 2 doses, and the human equivalent dose (HED) based on "US FDA Guidance for Industry: estimating the maximum safe starting dose in initial clinical trials for therapeutics in adult healthy volunteers" was calculated to be 4.8 mg / kg. The clinical starting dose of 0.8 mg / kg Q3W for the first-in-human (FTIH) study was based on one sixth of the HED of 4.8 mg / kg. At a dose of 0.8 mg / kg AZD8205 Q3W, with administration by 1 hour intravenous infusion, the steady state plasma exposure of AZD8205 is predicted to be Cmaxof 16 μg / mL and AUC of 77 μg.d / mL. max 0-21d

[0284] This approach to identifying a starting dose for an ADC has generally shown an acceptable balance of safety and effective dose escalation in FTIH trials. The human starting dose of 0.8 mg / kg is predicted to have an acceptable safety margin based on the HNSTD: 6.05 based on HED, 20.4 based on Cmax, and 11.2 based on AUC. max

[0285] Example 2: Study design and definitions

[0286] ​​​This first-in-human (FTIH) study is conducted in the context of a main protocol to evaluate the safety, tolerability, and preliminary efficacy of AZD8205 initially as monotherapy and in combination with other anti-cancer agents in subjects with advanced or metastatic solid malignancies expressing B7-H4, after the definition of a recommended Phase 2 dose (RP2D). The protocol also explores the potential biological activity of AZD8205 by characterizing pharmacokinetics (PK), pharmacodynamics (PDx), exploratory biomarkers, and anti-tumor activity.

[0287] Figure 1 The main protocol design is shown. As shown in Figure 1 The information relevant for the whole study, including study objectives, rationale, general inclusion and exclusion criteria, safety assessments, and adverse event (AE) reporting common to all sub-studies, can be found in the main protocol. Sub-study specific information, including planned dose levels and rationale, toxicity management, and dose modifications, can be found in the relevant sub-studies.

[0288] The main protocol consists of several sub-studies evaluating AZD8205. After the definition of a recommended Phase 2 dose (RP2D) based on emerging supportive preclinical and / or clinical data and scientific rationale, sub-studies of combination therapies are added. Each sub-study contains a dose escalation (e.g., Part A) and a dose expansion (e.g., Part B). The objective of the dose escalation (Part A) of each sub-study is to determine the safety, tolerability, RP2D, and / or maximum tolerated dose (MTD) of AZD8205 in subjects with tumors expressing B7-H4. As described in Example 9, all subjects selected for this study have a tumor in which at least 25% of the cells in a tumor biopsy are positive for B7-H4. The RP2D is selected by pooling and evaluating all available data, including appropriate PK, PD, efficacy, safety, and tolerability. The objective of the dose expansion (Part B) of each sub-study is to evaluate the anti-tumor activity of AZD8205. The initiation of Part B of each sub-study depends on the evaluation of safety results in Part A, and the determination of the RP2D for AZD8205 monotherapy and each combination.

[0289] Figure 2 The study flow for sub-study 1, outlined in the following examples, is shown. Key aspects of this study, such as the pre-screening selection process and starting dose of AZD8205 in sub-study 1, dose escalation, stopping criteria, and cohort size for Part A of all sub-studies, are based on accepted methods for Phase I / II oncology studies. In addition, blood samples are collected to enable characterization of AZD8205 PK, and, where appropriate, to enable investigation of additional metabolites.

[0290] As part of the AZD8205 clinical drug development program, PDx and exploratory biomarker (DNA, RNA, protein, or metabolite) profiles were investigated and their relationship to drug action was investigated. Potential benefits of this exploratory investigation include: (1) identification of subjects most likely to benefit from treatment; (2) prediction of which subjects are likely to be non-responders to treatment; (3) identification of adverse reactions associated with drug exposure; and (4) characterization of toxicities.

[0291] In summary, the examples provide the information and understanding needed to deliver a safe and effective new medicine to subjects with tumors expressing B7-H4.

[0292] Definitions The following terms were used in the studies described in the following examples.

[0293] Dose-limiting toxicities (DLT). DLT was evaluated during Part A (dose escalation). The DLT evaluation period was 21 days from the first dose of AZD8205 on Cycle 1, Day 1, or until and including the planned end of Cycle 1 if an alternative dosing schedule was explored. A DLT was defined as any Grade >3 treatment-induced AE that occurred during the DLT evaluation period that could not be attributed to the underlying disease or disease-related processes or concurrent illness with modification or exception.

[0294] Maximum tolerated dose (MTD). The MTD was defined as the highest dose with a target DLT probability of 30%. The MTD was determined by isobolographic analysis applied to the DLT rates observed during the dose escalation phase using the modified Toxicity Probability Interval-2 (mTPI-2) method (Guo et al. 2017).

[0295] Efficacy assessments Tumor response was assessed by RECIST vl. l (Eisenhauer et al. 2009) per schedule.

[0296] Tumor assessments Tumor assessments used computed tomography (CT, preferred) or magnetic resonance imaging (MRI) images of the chest, abdomen, and pelvis, supplemented with intravenous contrast, and additionally based on individual subject signs and symptoms collected at screening / baseline and at periodic (follow-up) intervals during the study intervention. In this study, a brain scan (preferred MRI) was mandatory for all subjects at screening / baseline. Only subjects with brain metastases at baseline required a post-baseline MRI of the brain, while subjects without brain metastases did not require additional brain scans for subsequent tumor assessments unless clinically indicated. If possible, the imaging modality used for baseline tumor assessment was kept the same at each subsequent follow-up assessment throughout the study.

[0297] Screening / baseline imaging is performed no more than 28 days prior to study intervention initiation and, ideally, as close to study intervention initiation as possible. Scans obtained prior to informed consent but within the 28-day period are acceptable as part of standard clinical practice. Tumor assessments are performed per schedule in each sub-study until objective disease progression as defined by RECIST vl. l and assessed by the investigator or withdrawal of consent. Confirmation of response (complete response (CR) or partial response (PR)) is made by repeating the consecutive scan at least 4 weeks after the first response documentation.

[0298] Tumor markers are not used for tumor response evaluation according to RECIST vl. l. Tumor markers are collected for separate analysis. However, based on the RECIST vl. l evaluation, results will not contribute to tumor response.

[0299] Eastern Cooperative Oncology Group (ECOG) performance status (PS). The performance status of subjects is assessed using the ECOG PS scale as detailed in the schedule of each sub-study.

[0300] Adverse events An adverse event (AE) is any untoward medical occurrence in a subject administered a medicinal product or clinical study subject that does not necessarily have a causal relationship with treatment. Thus, an AE can be any unfavorable and unintended sign (e.g., abnormal laboratory finding), symptom (e.g., nausea, chest pain), or disease associated temporally with the use of a medicinal product whether or not considered related to the medicinal product.

[0301] The term AE is used to include serious AEs and non-serious AEs and can include a worsening of a pre-existing medical condition. AEs can occur at any time, including during the run-in or washout period, even if there is no administration of study intervention.

[0302] Serious adverse events A serious adverse event (SAE) is an AE that occurs during any study phase (i.e., run-in, treatment, washout, follow-up) that meets one or more of the following criteria: (1) results in death; (2) immediately life-threatening; (3) requires subject hospitalization or prolongation of existing hospitalization; (4) results in a persistent or significant disability or incapacity; (5) is a congenital anomaly or birth defect; (6) is an important medical event that can jeopardize the subject or can require medical intervention to prevent one of the outcomes listed above.

[0303] Disease progressionDisease progression can be considered as worsening of the subject's condition due to the disease for which the study product is intended. It can be an increase in severity of the disease under study and / or an increase in symptoms of the disease. The occurrence of a new metastasis from a primary cancer under study or progression of an existing metastasis should be considered disease progression and not an AE. Events that are clearly attributable to disease progression should not be reported as an AE during the study.

[0304] New cancers Development of a new cancer should be considered an SAE. New primary cancers refer to those cancers that are not the primary reason for undergoing study intervention and that are identified after the subject has been enrolled in the study. They do not include metastases of the original cancer.

[0305] Example 3: Formulation and administration of AZD8205

[0306] This example shows the formulation and administration of AZD8205. AZD8205 is supplied in Part A and Part B as either a lyophilized product or a liquid product.

[0307] Lyophilized AZD8205 is supplied in 20R amber vials containing 100 mg AZD8205 (nominal) as a sterile lyophilized solid. AZD8205 investigational product (IP) must be stored at 2°C to 8°C (refrigerated) prior to use and protected from light. AZD8205 vials must not be frozen.

[0308] Liquid AZD8205 is supplied as an intravenous bag protector (IVBP) solution to ensure compatibility of AZD8205 with IV infusion components and dilution solutions. IVBP is stored at 2°C to 8°C (refrigerated). Lyophilized AZD8205 cannot be reconstituted with IVBP solution. Reconstitution is performed using sterile water for injection.

[0309] All IP should be kept in a safe and dry place. Vials should be stored at 2°C to 8°C in the dark (refrigerated) and must not be frozen.

[0310] The body weight-based dose is calculated using the following formula:

[0311]

[0312] Example 4: Initial risk / benefit assessment

[0313] The study is conducted to assess the risk of AZD8205 administration and to provide appropriate AZD8205 doses for Part A and Part B.

[0314] Preclinical toxicology studies to assess the safety of AZD8205 showed that cynomolgus monkeys exposed to AZD8205 at levels greater than the anticipated clinical effective range caused changes consistent with other marketed TOP1 i ADCs. The key target organs for toxicity were those with rapid cell turnover (e.g., bone marrow and testes).

[0315] In a non-GLP dose ranging discovery study, male cynomolgus monkeys were exposed to AZD8205 at dose levels of 15 mg / kg, 20 mg / kg, 22.5 mg / kg, and 25 mg / kg (IV Q3W x 2). The findings were consistent with expectations and consistent with cytotoxic TOP1 inhibitors. Dose levels > 20 mg / kg were not tolerated. At dose levels > 20 mg / kg, there were moderate to marked effects on the hematopoietic system (most notably lymphocyte, neutrophil, and reticulocyte reductions) in addition to signs of moribundity, with moderate to marked bone marrow cell reductions and cell reductions in other lymphoid organs correlating with histology; gastrointestinal toxicities (minimal to marked degeneration and regeneration) were noted in both large and small intestinal mucosa and were considered a key contributor to the moribund condition of the animals.

[0316] Other organs that showed treatment-related pharmacologic effects of AZD8205 included reversible mild to moderate histopathology findings in the kidney, liver, pancreatic islets, and testes. Some of these treatment-related findings also indicated stress / adverse and worsening clinical conditions.

[0317] Clinical chemistry changes were noted in the liver and kidney, including mild transient increases in liver enzymes (2-fold over control ALT / AST levels only on days 4-6, but without histopathology correlation) and marked increases in kidney enzymes associated with clinically observed dehydration. All changes were recoverable prior to the scheduled necropsy on day 43. The effects on the hematopoietic system were also only minimal to mild in severity in surviving animals (2 / 3 at 22.5 mg / kg and 3 / 3 at 15 mg / kg). The only treatment-related microscopic finding noted in animals dosed at 22.5 mg / kg IV Q3W x 2 was again consistent with the mechanism of action (MOA) of TOP1 inhibitors. There were no histopathology findings in any of the organs examined at 15 mg / kg.

[0318] In a pivotal GLP 6-week repeat-dose (Q3Wx2) IV study in cynomolgus monkeys with a 6-week recovery period, dose levels up to and including 15 mg / kg did not reveal any adverse findings (maximum dose tested and HNSTD). Assessments were made on all major organs and functions (respiratory, cardiovascular, renal, hepatic and nervous system), and no AZD 8205-related clinical signs or effects were found on survival, local (skin) tolerability, body weight, ECG, blood pressure, respiratory rate, ophthalmoscopy, neurological endpoints, clinical chemistry, urinalysis, systemic cytokine release or coagulation parameters. The only treatment-related changes observed were at dose levels > 10 mg / kg, and were related to the expected pharmacological activity of AZD 8205 on the hematopoietic system, and included minimal and transient effects on some red blood cell parameters, which were reversible during the 6-week recovery period.

[0319] A pivotal GLP AZD8205 toxicity study was found in male and female cynomolgus monkeys at dose levels up to and including 15 mg / kg, with no evidence of liver enzyme elevations, hypersensitivity / allergic reactions, or infusion-related reactions (mean integrated gender AUC systemic exposure was 963 μg.d / mL, and C max max was 323 μg / mL). These values are expected to provide approximately 13-fold and 20-fold safety margins over the minimum predicted clinically effective AUC (77 μg.d / mL) and C max max (16 μg / mL) levels, respectively, at the starting dose in this study. In the case of liver enzyme changes noted in the dose ranging study, these were minimal (approximately 2-fold), transient (noted in some animals for 1 or 2 days), not associated with hepatocyte necrosis, and fully reversible at the end of dosing animals with 15 mg / kg AZD8205.

[0320] Example 5: Study subjects

[0321] This example provides the criteria for selecting subjects for Part A and Part B studies of Substudy 1.

[0322] Inclusion criteria for Part A and Part B:

[0323] 1. Must be 18 years of age at the time of study participation.

[0324] 2. Have an Eastern Cooperative Oncology Group (ECOG) performance status of 0 or 1 at the time of enrollment.

[0325] 3. Subjects with recurrent / metastatic solid tumors must have previously received appropriate SoC therapy for their tumor type and disease stage, or where a clinical trial is the best option for next line treatment based on response and / or tolerability to prior therapy.

[0326] 4. According to RECIST vl. l, subjects must have measurable disease as defined by at least 1 lesion that can be accurately measured in longest diameter at baseline as >10 mm by CT or MRI (except for lymph nodes, the short axis must be >15 mm), which is suitable for accurate repeated measurements. Previously irradiated lesions or lesions in a field of radiation are not to be used as target lesions unless the lesion has demonstrated unequivocal disease progression by RECIST vl. l. Target lesions are not to be used for baseline tumor biopsies unless there are no other lesions suitable for biopsy and they meet the pre-defined requirements.

[0327] 5. Predicted life expectancy of >12 weeks.

[0328] 6. Adequate organ function as defined in Table 2.

[0329] Table 2. Criteria for adequate organ and bone marrow function

[0330]

[0331] aPTT = activated partial thromboplastin time; CrCl = creatinine clearance; D1C1 = Day 1 of Cycle 1; INR = international normalized ratio; ULN = upper limit of normal.

[0332] Example 6: Part A dose escalation

[0333] This example describes the Part A dose escalation study to determine the AZD8205 doses for Part B.

[0334] This sub-study is ongoing to explore the safety and tolerability of AZD8205 as monotherapy and to assess the anti-tumor activity of AZD8205 in breast, ovarian, endometrial, and cholangiocarcinoma cancers. AZD8205 is administered intravenously Q3W. The study is conducted in the following phases, applicable to both Part A and Part B:

[0335] 1. Pre-screening phase: All subjects provide a baseline tissue sample for B7-H4 analysis. Subjects with tumors expressing B7-H4 progress to the next phase.

[0336] 2. Screening phase: Subjects are assessed for study intervention eligibility. Only subjects who meet all inclusion criteria and do not meet any exclusion criteria are assigned to the study.

[0337] 3. Treatment phase: Subjects continue to receive the study intervention until one of the discontinuation criteria is met, including any AE contraindication (further dosing or AE) or radiological progression.

[0338] 4. Follow-up phase: All subjects are followed for survival.

[0339] Part A (dose escalation) will evaluate AZD8205 in approximately 60 subjects with positive B7-H4 tumor expression levels to determine the MTD and / or RP2D. The RP2D will be selected by pooling and evaluating all available data, including appropriate PK, PD, efficacy, safety, and tolerability. Based on safety, PK, and PDx data, the SRC can recommend an intermediate dose level and / or an alternative dosing schedule and longer treatment intervals. Any dose level not exceeding the MTD can be expanded by up to an additional 18 subjects (referred to as a PDx Supplemental Cohort, for a total of 36 subjects) based on emerging data. Subjects enrolled into the PDx Supplemental Cohort will not influence dosing decisions made by the SRC based on the mTBI-2 algorithm. The PDx Supplemental Cohort can be enrolled for specific tumor types with mandatory pre-treatment and on-treatment tumor biopsies. These PDx Supplemental Cohorts provide additional PDx and safety data that can inform optimal dose level selection for expansion based on available data.

[0340] In Part A, four dose levels of 0.8 mg / kg, 1.6 mg / kg, 2.4 mg / kg, and 3.2 mg / kg will be administered via intravenous (IV) infusion at Q3W (every three weeks).

[0341] Following an initial screening period of up to 28 days, the Part A intervention period is scheduled. The Part A intervention period includes up to or exceeding 20 cycles, each of 21 days or 28 days, and administration of AZD8205 every three weeks (Q3W), i.e., once every 21 -day cycle or once every 28-day cycle.

[0342] Eligible subjects receive AZD8205 administered via intravenous (IV) infusion at the selected dose Q3W, starting on Cycle 1 Day 1, for up to or exceeding 20 cycles. Subjects are treated with study intervention until disease progression, unacceptable toxicity, investigator’s decision, completion of the maximum number of treatment cycles, or withdrawal of consent. All subjects are followed for survival until the end of the study.

[0343] Following the invention period, subjects are further evaluated for disease progression, end-of-treatment status, and follow-up status.

[0344] Example 7: Part B dose expansion

[0345] The Part B study phase in this example is conducted with a 2.4 mg / kg dose of AZD8205 for a dose expansion.

[0346] Objectives and endpoints are described above in Example 6.

[0347] Part B (dose expansion) is initiated to investigate preliminary efficacy and to establish safety data from approximately 220 subjects from specific populations. Four expansion cohorts are planned prior to review of emerging data from Part A:

[0348] Approximately 40 subjects with BTC (cohort B1)

[0349] Approximately 60 subjects with ovarian cancer (cohort B2)

[0350] Approximately 30 subjects with PRR (cohort B2A)

[0351] Approximately 30 subjects with PSR (cohort B2B)

[0352] Approximately 90 subjects with HER2-negative (IHC: 0, 1+, 2+ / ISH-, as defined by the most recent ASCO / CAP guidelines) breast cancer (cohort B3):

[0353] Approximately 30 subjects with HR+, HER2-negative breast cancer (cohort B3A)

[0354] Approximately 30 subjects with TNBC (cohort B3B)

[0355] Approximately 30 subjects with HR±, HER2-negative breast cancer who have previously received treatment with a TOP1i ADC (cohort B3C)

[0356] Approximately 30 subjects with endometrial cancer (cohort B4).

[0357] Each expansion cohort can be initiated at more than one dose level or schedule depending on emerging data. Expansion cohorts can be opened simultaneously or independently in a parallel or staggered approach. If a new dosing schedule is explored, a dose modification will be agreed upon and this dose modification will be based on the previously established safe exposure range. If a decision is made to investigate two dose levels / schedules within an expansion cohort, the total sample size for Part B can be increased to 440 subjects. In this case, subjects can be randomized to the selected dose level / schedule; allocation will not be blinded. Randomization will occur on Day 1.

[0358] Safety and tolerability data will be assessed regularly for all subjects, and subjects will be classified in terms of their clinical response status according to RECIST vl.l.

[0359] Example 8: Preliminary Part A and Part B results

[0360] Approximately 40 subjects with BTC (cohort B1) Approximately 60 subjects with ovarian cancer (cohort B2) Approximately 30 subjects with PRR (cohort B2A) Approximately 30 subjects with PSR (cohort B2B) Approximately 90 subjects with HER2-negative (IHC: 0, 1+, 2+ / ISH-, as defined by the most recent ASCO / CAP guidelines) breast cancer (cohort B3): Approximately 30 subjects with HR+, HER2-negative breast cancer (cohort B3A) Approximately 30 subjects with TNBC (cohort B3B) Approximately 30 subjects with HR±, HER2-negative breast cancer who have previously received treatment with a TOP1i ADC (cohort B3C) Approximately 30 subjects with endometrial cancer (cohort B4).

[0361] Table 3 shows the demographics of the subjects evaluated in Part A and Part B.

[0362] Table 3. Demographics of subjects evaluated

[0363]

[0364] Preliminary PK data were provided. Data were available from n=3 patients (0.8 mg / kg), n=9 patients (1.6 mg / kg), n=9 (2.4 mg / Kg), and n=5 patients (3.2 mg / Kg). Based on the PK data, total AZD8205 ADC and total antibody concentrations were comparable across all dose levels tested, indicating that the warhead conjugation was stable in subjects. Specifically, AZD8205 exposure was consistent with preclinical predictions, increasing proportionally with dose, with a half-life of =4.9-7.2 days. In addition, AZD8205 exposure (AUC(0-21) of 143.5 ± 39.2 days pg / mL and 190.9 ± 10.1 days pg / mL at 1.6 mg / kg and 2.4 mg / kg, respectively, were within the predicted efficacious range (80-230 days pg / mL). Unconjugated warhead exposure in plasma was within 1.2-fold of the predicted AUC.

[0365] Computed tomography data indicated that AZD8205 was active in breast cancer (BC), ovarian cancer (OC), and endometrial cancer (EC). Specifically, 4 subjects showed both reduction + shrinkage (>30%) of tumor markers in target lesions (1 uPR and 3 cPR). By RECIST, 18 subjects showed SD as BoR (including 4 ongoing subjects with >20% tumor shrinkage). In OC, BC, and EC, tumor markers were reduced by >50% in 10 / 24 evaluable patients at all dose levels. ctDNA molecular measurements showed responses in N=14 / 21 evaluable subjects (about 67%) - N=5 / 8 subjects (63%) in Dose Level 2 (DL2), N=6 / 9 subjects (67%) in Dose Level 3 (DL3), 3 / 4 subjects (75%) in Dose Level 4 (DL4) - with patterns largely consistent with clinical response and tumor markers.

[0366] OvCa subjects with measurable CA125 (>2xULN) showed a reduction in CA125 in Figure 3Plots were generated in which it was confirmed the response to AZD8205 at 1.6 mg / kg, 2.4 mg / kg and 3.2 mg / kg. At 1.6 mg / kg, 2.4 mg / kg and 3.2 mg / kg, ctDNA molecular response was observed in approximately 67% of evaluable subjects (n=14 / 21). ctDNA changes largely mirrored clinical response, with greater ctDNA reduction observed in subjects who derived benefit. Figure 4 Emerging preclinical / clinical efficacy results are shown.

[0367] Example 9: Determining B7-H4 cutoff values for selected subjects

[0368] In the early phase of the first-in-human study described in the above example, it was observed that a higher overall response rate (ORR) was seen in subjects with tumors that had higher levels of B7-H4 expression, they were more responsive to treatment with AZD8205. The ORR for all subjects and subjects expressing B7-H4 is summarized in Table 7 below.

[0369] Table 7. ORR for all subjects compared to B7-H4 expressers

[0370]

[0371] To determine a B7-H4 expression cut-off for continuation of the human study, mouse xenograft studies were performed in CCA, ovarian cancer and TNBC. Athymic nude (Fox1 nu ) or NOD-SCID mice were xenografted with tumors 21 days prior to AZD8205 administration. AZD8205 was administered as a single i.v. dose on day 0. Changes in tumor growth were monitored 28 days post-treatment. A plot of the percent change in tumor growth versus the percent of cells expressing B7-H4 is shown in Figure 5

[0372] The receiver operator characteristic (ROC) curve was determined for each dose and these curves were used to calculate the Youden statistic for each dose. As shown in Figure 6 the Youden statistic was plotted versus the percent of B7-H4 cells. A Youden statistic of 1 represents a perfect biomarker, while a statistic of 0 means that the biomarker is indistinguishable. As can be seen in Figure 6 the Youden statistic plateaus at a B7-H4 percent of 25% or greater. Therefore, a 25% B7-H4 expression cut-off was used to evaluate the doses used.

[0373] Example 10: Overall response rate for heavily pretreated subjects .

[0374] ​The objective response rate (ORR) was evaluated in a total of 44 severely pretreated subjects from the studies described in the above examples. Figure 7 As shown, as a basis Figure 4 The second set of data, comprising 44 participants, included 18 with ovarian cancer, 9 with cholangiocarcinoma (CCA), 12 with breast cancer (BC), and 5 with endometrial cancer (EC). Of these 44 participants, 40 were evaluable for ORR. Figure 7 As indicated, some subjects had previously received platinum-based chemotherapy. (For example...) Figure 7 Materials submitted electronically are incorporated by reference Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 6 Figure 7 Definitions Methods of treating cancer Pharmaceutical compositions Kits Antibody or antigen-binding fragment sequences of the ADC Table 1. Antibody or antigen-binding fragment sequences of the ADC Examples Example 1: Starting dose adjustment Example 2: Study design and definitions Figure 1 Figure 1 Figure 2 Definitions Dose-limiting toxicities Maximum tolerated dose Efficacy assessments Tumor assessments Adverse events Serious adverse events Disease progression New cancers Example 3: Formulation and administration of AZD8205 Example 4: Initial risk / benefit assessment Example 5: Study subjects Table 2. Criteria for adequate organ and bone marrow function Example 6: Part A dose escalation Example 7: Part B dose expansion Example 8: Preliminary Part A and Part B results Table 3. Demographics of subjects evaluated Figure 3 Figure 4 Example 9: Determining B7-H4 cutoff values for selected subjects Table 7. ORR for all subjects compared to B7-H4 expressers Figure 5 Figure 6 Figure 6 Example 10: Overall response rate for heavily pretreated subjects Figure 7 Figure 4 Figure 7 Figure 7 Materials submitted electronically are incorporated by reference Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 6 Figure 7 Definitions Methods of treating cancer Pharmaceutical compositions Kits Antibody or antigen-binding fragment sequences of As shown, the mean ORR was approximately 25% for those subjects with significant reductions in tumor markers and / or ctDNA.

Claims

1. A method of treating a subject with cancer, the method comprising administering to the subject an antibody-drug conjugate (ADC) specifically bound to B7-H4 in an amount of about 0.8 mg / kg to about 4.8 mg / kg, said ADC comprising: i. An antibody or antigen-binding fragment thereof that binds to the B7-H4 polypeptide, said antibody or antigen-binding fragment comprising: a) Heavy chain CDR1 (HCDR1), heavy chain CDR2 (HCDR2), heavy chain CDR3 (HCDR3), light chain CDR1 (LCDR1), light chain CDR2 (LCDR2), and light chain CDR3 (LCDR3), or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, respectively; b) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6, respectively; c) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, and SEQ ID NO: 18, respectively; d) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, and SEQ ID NO: 24, respectively; or e) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, and SEQ ID NO: 30, respectively; ii. Cuttable connectors; and iii. Cytotoxic agents.

2. The method according to claim 1, wherein the amount of ADC applied is about 0.8 mg / kg, about 1.6 mg / kg, about 2.4 mg / kg, about 3.2 mg / kg, about 3.6 mg / kg, or about 4.8 mg / kg.

3. The method of claim 1, wherein the amount of ADC applied is about 1.6 mg / kg.

4. The method of claim 1, wherein the amount of ADC applied is about 2.4 mg / kg.

5. The method of claim 1, wherein the amount of ADC applied is about 3.2 mg / kg.

6. The method according to any one of claims 1 to 5, wherein the cancer comprises cancer cells expressing B7-H4.

7. The method according to any one of claims 1 to 6, wherein the ADC is administered to the subject once every three weeks.

8. The method according to any one of claims 1 to 7, wherein the cancer is selected from ovarian cancer, biliary tract cancer, breast cancer, pancreatic cancer, prostate cancer, hematologic malignancies, endometrial cancer, bile duct cancer, NSCLC (squamous cell carcinoma and / or adenocarcinoma), gastrointestinal cancers such as gastric cancer and colorectal cancer, and lung cancer.

9. The method according to any one of claims 1 to 8, wherein the cancer is selected from hormone receptor-positive (HR+) breast cancer, human epidermal growth factor receptor 2-positive (HER2+) breast cancer, and triple-negative breast cancer (TNBC).

10. The method according to any one of claims 1 to 9, wherein the cancer is homologous recombination defective (HRD) cancer.

11. The method of claim 10, wherein the cancer comprises one or more cells having a mutation in one or more HRD genes selected from BRCA1, BRCA2, ATM, BRIP1, BARD1, CDK12, CHEK1, CHEK2, FANCL, PALB2, PPP2R2A, RAD51B, RAD51C, RAD51D, and RAD54L.

12. The method of claim 11, wherein the mutation in the HRD gene is selected from BRCA1, BRCA2, and ATM.

13. The method according to any one of claims 1 to 12, wherein the antibody or its antigen-binding fragment comprises: i. Variable heavy (VH) chains and variable light (VL) chains, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 45 and SEQ ID NO: 34, respectively; ii. VH and VL chains, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 33 and SEQ ID NO: 34, respectively; iii. VH and VL chains, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 43 and SEQ ID NO: 34, respectively; iv. VH and VL chains, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 46 and SEQ ID NO: 34, respectively; v. VH and VL chains, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 47 and SEQ ID NO: 34, respectively; vi. VH and VL chains, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 31 and SEQ ID NO: 32, respectively; vii. VH and VL chains, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 35 and SEQ ID NO: 36, respectively; viii. VH and VL chains, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 37 and SEQ ID NO: 38, respectively; or ix. VH and VL chains, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 39 and SEQ ID NO: 40, respectively.

14. The method according to any one of claims 1 to 12, wherein the antibody or its antigen-binding fragment comprises: HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11 and SEQ ID NO: 12, respectively.

15. The method according to any one of claims 1 to 13, wherein the antibody or its antigen-binding fragment comprises a VH chain and a VL chain comprising the amino acid sequences SEQ ID NO: 45 and SEQ ID NO: 34, respectively, or functional variants thereof.

16. The method according to any one of claims 1 to 15, wherein the antibody or its antigen-binding fragment binds to the OVCAR4 cell line.

17. The method according to any one of claims 1 to 16, wherein the antibody or antigen-binding fragment thereof comprises: a heavy chain constant region comprising the amino acid sequence SEQ ID NO:

41.

18. The method according to any one of claims 1 to 16, wherein the antibody or its antigen-binding fragment comprises: a heavy chain constant region comprising the amino acid sequence SEQ ID NO:

52.

19. The method according to any one of claims 1 to 16, wherein the antibody or its antigen-binding fragment comprises: a light chain constant region comprising the amino acid sequence SEQ ID NO:

42.

20. The method according to any one of claims 1 to 16, wherein the antibody or antigen-binding fragment thereof comprises: a heavy chain comprising the amino acid sequence SEQ ID NO: 51; and a light chain comprising the amino acid sequence SEQ ID NO:

44.

21. The method according to any one of claims 1 to 16, wherein the antibody or antigen-binding fragment thereof comprises: a heavy chain comprising the amino acid sequence SEQ ID NO: 48; and a light chain comprising the amino acid sequence SEQ ID NO:

44.

22. The method according to any one of claims 1 to 21, wherein the antibody or its antigen-binding fragment is a monoclonal antibody.

23. The method according to any one of claims 1 to 22, wherein the antibody or its antigen-binding fragment is a humanized monoclonal antibody.

24. The method according to any one of claims 1 to 23, wherein the severable connector is an mp-PEG8-val-ala connector.

25. The method according to any one of claims 1 to 24, wherein the cytotoxic agent is a topoisomerase inhibitor.

26. The method of claim 25, wherein the topoisomerase inhibitor is of formula A. Compounds: 。 27. The method according to any one of claims 1 to 26, wherein the cleavable linker (ii) and the cytotoxic agent (iii) are selected from the group consisting of: (SG3932); (SG4010); (SG4057); and (SG4052).

28. The method of claim 27, wherein the linker in ii) and the cytotoxic agent in iii) together are compound SG3932.

29. The method according to any one of claims 1 to 28, wherein the ADC has a drug-to-antibody ratio (DAR) between about 1 and about 8.

30. The method according to any one of claims 1 to 28, wherein the ADC has a DAR of about 8.

31. A method of treating a subject with cancer, the method comprising administering to the subject an antibody-drug conjugate (ADC) specifically bound to B7-H4 in an amount of about 0.8 mg / kg to about 4.8 mg / kg, said ADC comprising: i. An antibody or antigen-binding fragment thereof binding to the B7-H4 polypeptide, said antibody or antigen-binding fragment comprising: heavy chain CDR1 (HCDR1), heavy chain CDR2 (HCDR2), heavy chain CDR3 (HCDR3), light chain CDR1 (LCDR1), light chain CDR2 (LCDR2), and light chain CDR3 (LCDR3), or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, respectively; and ii. A cleavable linker and a cytotoxic agent, said cleavable linker and cytotoxic agent being conjugated to an antibody or an antigen-binding fragment thereof having the following formula: (SG3932) 32. The method of claim 31, wherein the amount of ADC applied is about 0.8 mg / kg, about 1.6 mg / kg, about 2.4 mg / kg, about 3.2 mg / kg, about 3.6 mg / kg, or about 4.8 mg / kg.

33. The method of claim 31, wherein the amount of ADC applied is about 1.6 mg / kg.

34. The method of claim 31, wherein the amount of ADC applied is about 2.4 mg / kg.

35. The method of claim 31, wherein the amount of ADC applied is about 3.2 mg / kg.

36. The method according to any one of claims 31 to 35, wherein the antibody or its antigen-binding fragment comprises a VH chain and a VL chain comprising the amino acid sequences SEQ ID NO: 45 and SEQ ID NO: 34, respectively, or functional variants thereof.

37. The method according to any one of claims 31 to 36, wherein the cancer comprises cancer cells expressing B7-H4.

38. The method according to any one of claims 31 to 37, wherein the ADC is administered to the subject once every three weeks.

39. The method according to any one of claims 31 to 38, wherein the cancer is selected from ovarian cancer, biliary tract cancer, breast cancer, pancreatic cancer, prostate cancer, hematologic malignancies, endometrial cancer, bile duct cancer, NSCLC (squamous cell carcinoma and / or adenocarcinoma), gastrointestinal cancers such as gastric cancer and colorectal cancer, and lung cancer.

40. The method according to any one of claims 31 to 39, wherein the cancer is selected from hormone receptor-positive (HR+) breast cancer, human epidermal growth factor receptor 2-positive (HER2+) breast cancer, and triple-negative breast cancer (TNBC).

41. The method according to any one of claims 31 to 40, wherein the cancer is homologous recombination defective (HRD) cancer.

42. The method of claim 41, wherein the cancer comprises one or more cells having a mutation in one or more HRD genes selected from BRCA1, BRCA2, ATM, BRIP1, BARD1, CDK12, CHEK1, CHEK2, FANCL, PALB2, PPP2R2A, RAD51B, RAD51C, RAD51D, and RAD54L.

43. The method of claim 42, wherein the mutation in the HRD gene is selected from BRCA1, BRCA2, and ATM.

44. The method according to any one of claims 1 to 43, wherein at least about 25% of the cancer cells in the subject are B7-H4 positive cells, and optionally wherein the cancer cells are determined using immunohistochemistry (IHC).

45. A pharmaceutical composition comprising an antibody-drug conjugate (ADC) specifically bound to B7-H4, said ADC being administered in an amount from about 0.8 mg / kg to about 4.8 mg / kg, said ADC comprising: i. An antibody or antigen-binding fragment thereof binding to the B7-H4 polypeptide, said antibody or antigen-binding fragment comprising: heavy chain CDR1 (HCDR1), heavy chain CDR2 (HCDR2), heavy chain CDR3 (HCDR3), light chain CDR1 (LCDR1), light chain CDR2 (LCDR2), and light chain CDR3 (LCDR3), or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, respectively; and ii. A cleavable linker and a cytotoxic agent, said cleavable linker and cytotoxic agent being conjugated to an antibody or an antigen-binding fragment thereof having the following formula: (SG3932) 46. ​​The pharmaceutical composition of claim 45, wherein the pharmaceutical composition is administered in an amount of about 0.8 mg / kg, about 1.6 mg / kg, about 2.4 mg / kg, about 3.2 mg / kg, about 3.6 mg / kg or about 4.8 mg / kg ADC.

47. The pharmaceutical composition of claim 45, wherein the pharmaceutical composition is administered at an amount of about 1.6 mg / kg ADC.

48. The pharmaceutical composition of claim 45, wherein the pharmaceutical composition is administered at an amount of about 2.4 mg / kg ADC.

49. The pharmaceutical composition of claim 45, wherein the pharmaceutical composition is administered at an amount of about 3.2 mg / kg ADC.

50. The pharmaceutical composition according to any one of claims 45 to 49, wherein the antibody or antigen-binding fragment of the ADC comprises a VH chain and a VL chain, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 45 and SEQ ID NO: 34, respectively.

51. A pharmaceutical composition comprising an antibody-drug conjugate (ADC) specifically bound to B7-H4, said ADC being administered in an amount of about 25 mg to about 900 mg, said ADC comprising: i. An antibody or antigen-binding fragment thereof that binds to the B7-H4 polypeptide, said antibody or antigen-binding fragment comprising: f) Heavy chain CDR1 (HCDR1), heavy chain CDR2 (HCDR2), heavy chain CDR3 (HCDR3), light chain CDR1 (LCDR1), light chain CDR2 (LCDR2), and light chain CDR3 (LCDR3), or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, respectively; g) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6, respectively; h) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, and SEQ ID NO: 18, respectively; i) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, and SEQ ID NO: 24, respectively; or j) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, and SEQ ID NO: 30, respectively; ii. Cuttable connectors; and iii. Cytotoxic agents.

52. The pharmaceutical composition according to claim 51, wherein the amount of ADC applied is about 25 mg, about 40 mg, about 60 mg, about 80 mg, about 100 mg, about 120 mg, about 140 mg, about 160 mg, about 180 mg, about 200 mg, about 220 mg, about 240 mg, about 260 mg, about 280 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 600 mg, about 750 mg, or about 900 mg.

53. The pharmaceutical composition according to claim 51 or 52, wherein the combined cleavable linker and cytotoxic agent have the following formula: (SG3932) 54. The pharmaceutical composition according to any one of claims 51 to 53, wherein the antibody or antigen-binding fragment of the ADC comprises a VH chain and a VL chain, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 45 and SEQ ID NO: 34, respectively.

55. A kit comprising a pharmaceutical composition according to any one of claims 45 to 54.

56. The kit according to claim 55, further comprising instructions for use of the pharmaceutical composition.

57. The pharmaceutical composition according to any one of claims 45 to 54, wherein the pharmaceutical composition is used in the treatment of cancer.

58. The pharmaceutical composition according to claim 57, wherein the cancer comprises cancer cells expressing B7-H4.

59. The pharmaceutical composition for use according to claim 57 or 58, wherein the ADC is administered once every three weeks.

60. The pharmaceutical composition for use according to any one of claims 57 to 59, wherein the cancer is selected from ovarian cancer, biliary tract cancer, breast cancer, pancreatic cancer, prostate cancer, hematologic malignancies, endometrial cancer, bile duct cancer, NSCLC (squamous cell carcinoma and / or adenocarcinoma), gastrointestinal cancers such as gastric cancer and colorectal cancer, and lung cancer.

61. The pharmaceutical composition for use according to any one of claims 57 to 60, wherein the cancer is breast cancer selected from hormone receptor-positive (HR+) breast cancer, human epidermal growth factor receptor 2-positive (HER2+) breast cancer, and triple-negative breast cancer (TNBC).

62. The pharmaceutical composition for use according to any one of claims 57 to 61, wherein the cancer is homologous recombination defective (HRD) cancer.

63. The pharmaceutical composition for use according to any one of claims 57 to 62, wherein the cancer comprises one or more cells having a mutation in the HRD gene selected from BRCA1, BRCA2, ATM, BRIP1, BARD1, CDK12, CHEK1, CHEK2, FANCL, PALB2, PPP2R2A, RAD51B, RAD51C, RAD51D, and RAD54L.

64. The pharmaceutical composition for use according to claim 63, wherein the mutation in the HRD gene is selected from BRCA1, BRCA2, and ATM.

65. A method of treating a subject's cancer, the method comprising administering to the subject an antibody-drug conjugate (ADC) specifically bound to B7-H4, the ADC comprising: i. An antibody or antigen-binding fragment thereof that binds to the B7-H4 polypeptide, said antibody or antigen-binding fragment comprising: k) comprising the heavy chain CDR1 (HCDR1), heavy chain CDR2 (HCDR2), heavy chain CDR3 (HCDR3), light chain CDR1 (LCDR1), light chain CDR2 (LCDR2), and light chain CDR3 (LCDR3) of the amino acid sequences SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12, respectively; or functional variants thereof. l) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6, respectively; m) contains HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, or functional variants thereof, of the amino acid sequences SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, and SEQ ID NO: 18, respectively; n) comprising HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, or functional variants thereof, of the amino acid sequences SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, and SEQ ID NO: 24, respectively; or o) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, or functional variants thereof, comprising the amino acid sequences SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, and SEQ ID NO: 30, respectively; ii. Cuttable connectors; and iii. Cytotoxic agents; The subjects in the study had at least approximately 25% of their cancer cells as B7-H4 positive cells.

66. A method for diagnosing a cancer sample as B7-H4 positive, the method comprising: The cancer sample may optionally be analyzed using immunohistochemistry (IHC) to determine whether at least 25% of the cells express membrane-bound B7-H4, further wherein the IHC may optionally be performed using an antibody reagent.

67. The method of claim 65, wherein the B7-H4 positive cells are determined by measuring membrane B7-H4 expression prior to administration of the ADC, optionally wherein the B7-H4 expression is determined using immunohistochemistry (IHC), further wherein the IHC is optionally performed using an antibody reagent.

68. The method of claim 65, wherein the ADC is administered in an amount of about 0.8 mg / kg to about 4.8 mg / kg.

69. The method of any one of claims 65 or 67 to 68, wherein the ADC is administered to the subject once every three weeks.

70. The method of claim 69, wherein the subject has previously received treatment with a platinum-based chemotherapy agent.

71. The method according to any one of claims 65 to 70, wherein the cancer is selected from ovarian cancer, biliary tract cancer, breast cancer, pancreatic cancer, prostate cancer, hematologic malignancies, endometrial cancer, bile duct cancer, NSCLC (squamous cell carcinoma and / or adenocarcinoma), gastrointestinal cancers such as gastric cancer and colorectal cancer, and lung cancer.

72. The method according to any one of claims 65 to 71, wherein the cancer is selected from hormone receptor-positive (HR+) breast cancer, human epidermal growth factor receptor 2-positive (HER2+) breast cancer, and triple-negative breast cancer (TNBC).

73. The method according to any one of claims 65 to 72, wherein the cancer is homologous recombination defective (HRD) cancer.

74. The method of claim 73, wherein the cancer comprises one or more cells having a mutation in one or more HRD genes selected from BRCA1, BRCA2, ATM, BRIP1, BARD1, CDK12, CHEK1, CHEK2, FANCL, PALB2, PPP2R2A, RAD51B, RAD51C, RAD51D, and RAD54L.

75. The method of claim 73, wherein the mutation in the HRD gene is selected from BRCA1, BRCA2, and ATM.

76. The method according to any one of claims 65 or 67 to 75, wherein the antibody or its antigen-binding fragment comprises a VH chain and a VL chain comprising the amino acid sequences SEQ ID NO: 45 and SEQ ID NO: 34, respectively, or functional variants thereof.

77. The method according to any one of claims 65 or 67 to 76, wherein the antibody or antigen-binding fragment thereof comprises: a heavy chain comprising the amino acid sequence SEQ ID NO: 51; and a light chain comprising the amino acid sequence SEQ ID NO:

44.

78. The method according to any one of claims 65 to 77, wherein the antibody or its antigen-binding fragment is a monoclonal antibody.

79. The method according to any one of claims 65 to 77, wherein the antibody or its antigen-binding fragment is a humanized monoclonal antibody.

80. The method according to any one of claims 65 to 79, wherein the severable connector is an mp-PEG8-val-ala connector.

81. The method according to any one of claims 65 to 80, wherein the cytotoxic agent is a topoisomerase inhibitor.

82. The method according to claim 81, wherein the topoisomerase inhibitor is of formula A. Compounds: 。 83. The method according to any one of claims 65 to 81, wherein the cleavable linker (ii) and the cytotoxic agent (iii) together are selected from the following compounds: (SG3932); (SG4010); (SG4057); and (SG4052).

84. The method according to claim 83, wherein the linker in ii) and the cytotoxic agent in iii) together are compound SG3932.

85. The method according to any one of claims 65 to 84, wherein the ADC has a drug-to-antibody ratio (DAR) between about 1 and about 8.

86. The method according to any one of claims 65 to 84, wherein the ADC has a DAR of about 8.

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