ANTICORPO ANTI-HER3 / MUC1 OU FRAGMENTO DE LIGAÇÃO AO ANTÍGENO DO MESMO, ÁCIDO NUCLEICO, VETOR, CÉLULA, MÉTODO DE PRODUÇÃO DE UM ANTICORPO ANTI-HER3 / MUC1 OU UM FRAGMENTO DE LIGAÇÃO AO ANTÍGENO DO MESMO, CONJUGADOS ANTICORPO ANTI-HER3 / MUC1-FÁRMACO (ADC), USOS DO ANTICORPO ANTI-HER3 / MUC1 OU FRAGMENTO DE LIGAÇÃO AO ANTÍGENO DO MESMO, MÉTODO DE DIMINUIÇÃO DA TAXA DE CRESCIMENTO TUMORAL E COMPOSIÇÃO FARMACÊUTICA

Abstract

This disclosure relates to anti-HER3 / MUC1 antibodies, and antibody drug conjugates derived therefrom, and the use thereof.

Description

1 / 126 “ANTI-HER3 / MUC1 ANTIBODY OR ANTIGEN-BINDING FRAGMENT THEREOF, NUCLEIC ACID, VECTOR, CELL, METHOD OF PRODUCING AN ANTI-HER3 / MUC1 ANTIBODY OR AN ANTIGEN-BINDING FRAGMENT THEREOF, ANTI-HER3 / MUC1-DRUG CONJUGATES (ADCs), USES OF THE ANTI-HER3 / MUC1 ANTIBODY OR ANTIGEN-BINDING FRAGMENT THEREOF, METHOD OF REDUCING TUMOR GROWTH RATE AND PHARMACEUTICAL COMPOSITION” Related Patent Application

[0001] This application claims priority over PCT / CN2023 / 082375, filed on 18 March 2023, and PCT / CN2023 / 120377, filed on 21 September 2023. The entire content of the previous applications is incorporated herein by reference. Field of Invention

[0002] This invention relates to multispecific antiHER3 (human epidermal growth factor receptor 3) / MUC1 (mucin 1) antibodies (e.g., bispecific antibodies or antigen-binding fragments thereof) and antibody-drug conjugates derived therefrom. Background of the Invention

[0003] A bispecific antibody is an artificial protein that can simultaneously bind to two different types of antigens or two different epitopes. This dual specificity opens up a wide range of applications, including redirecting T cells to tumor cells, dual targeting of different disease mediators, and delivering payloads to target sites. The approval of catumaxomab (anti-EpCAM and anti-CD3) and blinatumomab (anti-CD19 and anti-CD3) became an important milestone in the development of bispecific antibodies. Petition 870250084299, dated 09 / 18 / 2025, pp. 272 / 410 2 / 126

[0004] Because bispecific antibodies have multiple applications, there is a need to continue the development of various therapies based on bispecific antibodies. Brief Description of the Invention

[0005] The present invention relates to antiHER3 / MUC1 antibodies or antigen-binding fragments thereof, wherein the antibodies or antigen-binding fragments thereof bind specifically to HER3 and MUC1. In some embodiments, the antibodies or antigen-binding fragments thereof have identical variable light chain regions. In some embodiments, the antibodies or antigen-binding fragments thereof have a common light chain. The invention also relates to antibody-drug conjugates derived from these antiHER3 / MUC1 antibodies.

[0006] In one aspect, the invention relates to an anti-HER3 / MUC1 antibody or antigen-binding fragment thereof comprising: a first antigen-binding domain that specifically binds to HER3 and a second antigen-binding domain that specifically binds to MUC1.

[0007] In some embodiments, the first antigen-binding domain comprises a first variable heavy chain region (VH1) and a first variable light chain region (VL1); and the second antigen-binding domain comprises a second variable heavy chain region (VH2) and a second variable light chain region (VL2).

[0008] In some embodiments, the first variable heavy chain region (VH1) comprises complementarity-determining regions (CDRs) 1, 2, and 3, wherein the CDR1 region of VH1 comprises an amino acid sequence that is at least 80% identical to a selected CDR1 amino acid sequence of VH1, and the CDR2 region of VH1 comprises a Petition 870250084299, dated 09 / 18 / 2025, pp. 273 / 410 3 / 126 amino acid sequence that is at least 80% identical to a selected VH1 CDR2 amino acid sequence, and the VH1 CDR3 region comprises an amino acid sequence that is at least 80% identical to a selected VH1 CDR3 amino acid sequence; and the first variable light chain region (VL1) comprises CDRs 1, 2, and 3, wherein the CDR1 region of VL1 comprises an amino acid sequence that is at least 80% identical to a selected CDR1 amino acid sequence of VL1, the CDR2 region of VL1 comprises an amino acid sequence that is at least 80% identical to a selected CDR2 amino acid sequence of VL1, and the CDR3 region of VL1 comprises an amino acid sequence that is at least 80% identical to a selected CDR3 amino acid sequence of VL1, wherein the amino acid sequences of selected CDRs 1, 2, and 3 of VL1 and the amino acid sequences of selected CDRs 1, 2, and 3 of VL1 are one of the following: The amino acid sequences of selected VH1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 4 to 6, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 1 to 3, respectively; The amino acid sequences of selected VH1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 7 to 9, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 1 to 3, respectively; The amino acid sequences of selected VH1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 7, 42, and 43, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 1 to 3, respectively; the amino acid sequences of CDRs 1, 2 and 3 of VH1 Petition 870250084299, dated 09 / 18 / 2025, pp. 274 / 410 4 / 126 selected are presented in SEQ ID NOs: 16 to 18, respectively, and the amino acid sequences of CDRs 1, 2 and 3 of VL1 selected are presented in SEQ ID NOs: 13 to 15, respectively; The amino acid sequences of selected VH1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 19 to 21, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 13 to 15, respectively; and the amino acid sequences of selected VH1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 19, 20, 44, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 1 to 3, respectively.

[0009] In some embodiments, the second variable heavy chain region (VH2) comprises CDRs 1, 2 and 3, wherein the CDR1 region of VH2 comprises an amino acid sequence that is at least 80% identical to a selected CDR1 amino acid sequence of VH2, the CDR2 region of VH2 comprises an amino acid sequence that is at least 80% identical to a selected CDR2 amino acid sequence of VH2, and the CDR3 region of VH2 comprises an amino acid sequence that is at least 80% identical to a selected CDR3 amino acid sequence of VH2;and the second variable light chain region (VL2) comprises CDRs 1, 2 and 3, wherein the CDR1 region of VL2 comprises an amino acid sequence that is at least 80% identical to a selected CDR1 amino acid sequence of VL2, the CDR2 region of VL2 comprises an amino acid sequence that is at least 80% identical to a selected CDR2 amino acid sequence of VL2, and the CDR3 region of VL2 comprises an amino acid sequence that is at least 80% identical to a selected CDR3 amino acid sequence of VL2; Petition 870250084299, dated 09 / 18 / 2025, pp. 275 / 410 5 / 126 where the amino acid sequences of selected VH2 CDRs 1, 2, and 3 and the amino acid sequences of selected VL2 CDRs 1, 2, and 3 are one of the following: The amino acid sequences of selected VH2 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 10 to 12, respectively, and the amino acid sequences of selected VL2 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 1 to 3, respectively; and the amino acid sequences of selected VH2 CDRs 1, 2, and 3 are shown in SEQ ID NOs: 22 to 24, respectively, and the amino acid sequences of selected VL2 CDRs 1, 2, and 3 are shown in SEQ ID NOs: 13 to 15, respectively.

[0010] In some embodiments, 1 the amino acid sequences of selected VH1 CDRs 1, 2 and 3 are presented in SEQ ID NOs: 4 to 6, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2 and 3 are presented in SEQ ID NOs: 1 to 3, respectively, and the amino acid sequences of selected VH2 CDRs 1, 2 and 3 are presented in SEQ ID NOs: 10 to 12, respectively, and the amino acid sequences of selected VL2 CDRs 1, 2 and 3 are shown in SEQ ID NOs: 1 to 3, respectively; The amino acid sequences of selected VH1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 16 to 18, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 13 to 15, respectively, and the amino acid sequences of selected VH2 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 22 to 24, respectively, and the amino acid sequences of selected VL2 CDRs 1, 2, and 3 are shown in SEQ ID NOs: 13 to 15, respectively; the amino acid sequences of CDRs 1, 2 and 3 of VH1 Petition 870250084299, dated 09 / 18 / 2025, pp. 276 / 410 6 / 126 selected are presented in SEQ ID NOs: 7 to 9, respectively, and the amino acid sequences of CDRs 1, 2 and 3 of VL1 selected are presented in SEQ ID NOs: 1 to 3, respectively, and the amino acid sequences of CDRs 1, 2 and 3 of VH2 selected are presented in SEQ ID NOs: 10 to 12, respectively, and the amino acid sequences of CDRs 1, 2 and 3 of VL2 selected are presented in SEQ ID NOs: 1 to 3, respectively; The amino acid sequences of selected VH1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 19 to 21, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 13 to 15, respectively, and the amino acid sequences of selected VH2 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 22 to 24, respectively, and the amino acid sequences of selected VL2 CDRs 1, 2, and 3 are shown in SEQ ID NOs: 13 to 15, respectively; The amino acid sequences of selected VH1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 7, 42, and 43, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 1 to 3, respectively, and the amino acid sequences of selected VH2 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 10 to 12, respectively, and the amino acid sequences of selected VL2 CDRs 1, 2, and 3 are shown in SEQ ID NOs: 1 to 3, respectively; or the amino acid sequences of the selected VH1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 19, 20, 44, respectively, and the amino acid sequences of the selected VL1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 13 to 15, respectively, and the amino acid sequences of the selected VH2 CDRs 1, 2, and 3 are presented in Petition 870250084299, dated 09 / 18 / 2025, pp. 277 / 410 7 / 126 SEQ ID NOs: 22 to 24, respectively, and the amino acid sequences of the selected VL2 CDRs 1, 2, and 3 are shown in SEQ ID NOs: 13 to 15, respectively.

[0011] In some embodiments, the first variable heavy chain region comprises a sequence that is at least 80%, 85%, 90%, 95%, 99%, or 100% identical to SEQ ID NO: 26, the first variable light chain region comprises a sequence that is at least 80%, 85%, 90%, 95%, 99%, or 100% identical to SEQ ID NO: 25, the second variable heavy chain region comprises a sequence that is at least 80%, 85%, 90%, 95%, 99%, or 100% identical to SEQ ID NO: 28, and the second variable light chain region comprises a sequence that is at least 80%, 85%, 90%, 95%, 99%, or 100% identical SEQ ID NO: 25.

[0012] In some embodiments, the first variable heavy chain region comprises a sequence that is at least 80%, 85%, 90%, 95%, 99%, or 100% identical to SEQ ID NO: 27, the first variable light chain region comprises a sequence that is at least 80%, 85%, 90%, 95%, 99%, or 100% identical to SEQ ID NO: 25, the second variable heavy chain region comprises a sequence that is at least 80%, 85%, 90%, 95%, 99%, or 100% identical to SEQ ID NO: 28, and the second variable light chain region comprises a sequence that is at least 80%, 85%, 90%, 95%, 99%, or 100% identical SEQ ID NO: 25.

[0013] In some embodiments, the first variable heavy chain region comprises a sequence that is at least 80%, 85%, 90%, 95%, 99%, or 100% identical to SEQ ID NO: 45, the first variable light chain region comprises a sequence that is at least 80%, 85%, 90%, 95%, 99%, or 100% identical to SEQ ID NO: 25, the second variable heavy chain region comprises a sequence that is at least 80%, 85%, 90%, 95%, 99%, or 100% identical to SEQ ID NO: 28, and the second variable region Petition 870250084299, dated 09 / 18 / 2025, pp. 278 / 410 8 / 126 of light chain comprises a sequence that is at least 80%, 85%, 90%, 95%, 99%, or 100% identical to SEQ ID NO: 25.

[0014] In some embodiments, VH1 comprises an amino acid sequence that is at least 90% identical to a selected VH sequence, and VL1 comprises an amino acid sequence that is at least 90% identical to a selected VL sequence, wherein the selected VH sequence and the selected VL sequence are one of the following: The selected VH sequence is SEQ ID NO: 26 and the selected VL sequence is SEQ ID NO: 25; The selected VH sequence is SEQ ID NO: 27 and the selected VL sequence is SEQ ID NO: 25; and the selected VH sequence is SEQ ID NO: 45 and the selected VL sequence is SEQ ID NO: 25.

[0015] In some embodiments, VH1 comprises CDR1 of VH1, CDR2 of VH1 and CDR3 of VH1 that are identical to CDR1 of VH, CDR2 of VH and CDR3 of VH of a selected VH sequence; and VL1 comprises CDR1 of VL1, CDR2 of VL1 and CDR3 of VL1 that are identical to CDR1 of VL, CDR2 of VL and CDR3 of VL of a selected VL sequence, wherein the selected VH sequence and the selected VL sequence are one of the following: The selected VH sequence is SEQ ID NO: 26 and the selected VL sequence is SEQ ID NO: 25; The selected VH sequence is SEQ ID NO: 27 and the selected VL sequence is SEQ ID NO: 25; and the selected VH sequence is SEQ ID NO: 45 and the selected VL sequence is SEQ ID NO: 25.

[0016] In some embodiments, VH2 comprises an amino acid sequence that is at least 90% identical to a sequence of Petition 870250084299, dated 09 / 18 / 2025, pp. 279 / 410 9 / 126 VH is selected, and VL2 comprises an amino acid sequence that is at least 90% identical to a selected VL sequence, wherein the selected VH sequence is SEQ ID NO: 28 and the selected VL sequence is SEQ ID NO: 25.

[0017] In some embodiments, VH2 comprises CDR1 of VH2, CDR2 of VH2 and CDR3 of VH2 that are identical to CDR1 of VH, CDR2 of VH and CDR3 of VH of a selected VH sequence; and VL2 comprises CDR1 of VL2, CDR2 of VL2 and CDR3 of VL2 that are identical to CDR1 of VL, CDR2 of VL and CDR3 of VL of a selected VL sequence, wherein the selected VH sequence is SEQ ID NO: 28 and the selected VL sequence is SEQ ID NO: 25.

[0018] In some embodiments, VH1 comprises the sequence with SEQ ID NO: 26 and VL1 comprises the sequence with SEQ ID NO: 25.

[0019] In some embodiments, VH1 comprises the sequence with SEQ ID NO: 27 and VL1 comprises the sequence with SEQ ID NO: 25.

[0020] In some embodiments, VH1 comprises the sequence with SEQ ID NO: 45 and VL1 comprises the sequence with SEQ ID NO: 25.

[0021] In some embodiments, VH2 comprises the sequence with SEQ ID NO: 28 and VL2 comprises the sequence with SEQ ID NO: 25.

[0022] In some embodiments, the first antigen-binding domain binds specifically to human or monkey HER3 and / or the second antigen-binding domain binds specifically to human or monkey MUC1.

[0023] In some embodiments, the first antigen-binding domain is human or humanized and / or the second antigen-binding domain is human or humanized.

[0024] In some embodiments, the antibody is a multispecific antibody (e.g., a bispecific antibody). Petition 870250084299, dated 09 / 18 / 2025, pp. 280 / 410 10 / 126

[0025] In some embodiments, the first antigen-binding domain is a single-strand variable fragment (scFv) and / or the second antigen-binding domain is an scFv.

[0026] In some embodiments, the first variable light chain region and the second variable light chain region are identical.

[0027] In one aspect, the invention relates to an anti-HER3 / MUC1 antibody or antigen-binding fragment thereof that cross-competes with the anti-HER3 / MUC1 antibody or antigen-binding fragment thereof described herein.

[0028] In one aspect, the invention relates to a nucleic acid comprising a polynucleotide encoding the antiHER3 / MUC1 antibody or antigen-binding fragment thereof described herein.

[0029] In one aspect, the invention relates to a vector comprising the nucleic acid described herein.

[0030] In one aspect, the invention relates to a cell comprising the vector described in this document.

[0031] In some forms, the cell is a CHO cell.

[0032] In one aspect, the invention relates to a cell comprising the nucleic acid described in this document.

[0033] In one aspect, the invention relates to a method of producing an anti-HER3 / MUC1 antibody or an antigen-binding fragment thereof, the method comprising cultivating the cell described herein under conditions sufficient for the cell to produce the anti-HER3 / MUC1 antibody or the antigen-binding fragment thereof; and collecting the anti-HER3 / MUC1 antibody or the antigen-binding fragment thereof produced by the cell. Petition 870250084299, dated 09 / 18 / 2025, pp. 281 / 410 11 / 126

[0034] In one aspect, the invention relates to an anti-HER3 / MUC1 antibody-drug conjugate (ADC) comprising a therapeutic agent covalently linked to the anti-HER3 / MUC1 antibody or antigen-binding fragment thereof described herein.

[0035] In some modalities, the therapeutic agent is a cytotoxic or cytostatic agent.

[0036] In some modalities, the therapeutic agent is MMAE or MMAF.

[0037] In some modalities, the therapeutic agent is selected from

[0038] In some embodiments, the therapeutic agent is linked to the antibody or antigen-binding fragment thereof by means of a ligand. In some forms, the ligand has a structure of: Petition 870250084299, dated 09 / 18 / 2025, pp. 282 / 410 12 / 126

[0039] In some embodiments, the antibody-drug conjugate has a structure of: In some embodiments, n = 1 to 8; in some embodiments, “Ab” represents the antibody or antigen-binding fragment thereof.

[0040] In some modalities, the drug-to-antibody ratio (DAR) is about 4 or 8.

[0041] In one aspect, the invention relates to a method of treating a subject who has cancer, wherein the method comprises administering a therapeutically effective amount of a composition comprising the anti-HER3 / MUC1 antibody or antigen-binding fragment thereof described herein, or the anti-HER3 / MUC1 antibody-drug conjugate described herein, to the subject.

[0042] In some forms, the subject has a cancer that expresses HER3 and / or MUC1 (for example, both HER3 and MUC1). Petition 870250084299, dated 09 / 18 / 2025, pp. 283 / 410 13 / 126

[0043] In some forms, the cancer is esophageal cancer, colorectal cancer, gastric cancer, breast cancer, endometrial carcinoma, lung cancer, melanoma, ovarian cancer, bladder cancer, stomach cancer, non-Hodgkin lymphoma, head and neck cancer, pancreatic cancer, lung adenocarcinoma, and cervical cancer.

[0044] In some forms, the subject is a human being.

[0045] In some embodiments, the method additionally comprises the administration of an anti-PD1 antibody to the subject.

[0046] In some modalities, the method additionally includes the administration of chemotherapy to the subject.

[0047] In one aspect, the invention relates to a method of decreasing the rate of tumor growth, wherein the method comprises placing a tumor cell in contact with an effective amount of a composition comprising the anti-HER3 / MUC1 antibody or antigen-binding fragment thereof as described herein, or the anti-HER3 / MUC1 antibody-drug conjugate described herein.

[0048] In one aspect, the invention relates to a method for exterminating a tumor cell, wherein the method comprises placing a tumor cell in contact with an effective amount of a composition comprising the anti-HER3 / MUC1 antibody or antigen-binding fragment thereof as described herein, or the anti-HER3 / MUC1 antibody-drug conjugate described herein.

[0049] In one aspect, the invention relates to a pharmaceutical composition comprising a pharmaceutically acceptable carrier and (a) the anti-HER3 / MUC1 antibody or antigen-binding fragment thereof described herein, and / or (b) the anti-HER3 / MUC1 antibody-drug conjugate described in Petition 870250084299, dated 09 / 18 / 2025, pp. 284 / 410 14 / 126 present document.

[0050] In one aspect, the invention relates to an anti-HER3 / MUC1 antibody-drug conjugate (ADC) comprising a therapeutic agent covalently linked to a bispecific antibody or antigen-binding fragment thereof comprising: a first antigen-binding domain that specifically binds to HER3 and a second antigen-binding domain that specifically binds to MUC1.

[0051] As used in this document, the term “antigen-binding domain” refers to one or more protein domains (e.g., formed from amino acids of a single polypeptide or formed from amino acids of two or more polypeptides (e.g., identical or different polypeptides)) that are capable of specifically binding to one or more different antigens (e.g., an effector antigen or control antigen). In some embodiments, an antigen-binding domain may bind to an antigen or epitope with specificity and affinity similar to those of naturally occurring antibodies. In some embodiments, the antigen-binding domain may be an antibody or a fragment thereof. An example of an antigen-binding domain is an antigen-binding domain formed by a VH-VL dimer. In some embodiments, an antigen-binding domain may include an alternative scaffold.In some embodiments, the antigen-binding domain is a VHH. Non-limiting examples of antigen-binding domains are described herein. Additional examples of antigen-binding domains are known in the art. In some examples, an antigen-binding domain may bind to a single antigen (e.g., one of an effector antigen and one of a control antigen). In other examples, an antigen-binding domain may bind to two different antigens (e.g., an effector antigen and a control antigen). Petition 870250084299, dated 09 / 18 / 2025, pp. 285 / 410 15 / 126 control).

[0052] The term “antibody” is used in this document in its broadest sense and includes certain types of immunoglobulin molecules that include one or more antigen-binding domains that bind specifically to an antigen or epitope. An antibody specifically includes, for example, intact antibodies (e.g., intact immunoglobulins), antibody fragments, bispecific antibodies, and multispecific antibodies. An example of an antibody is a protein complex that includes two heavy chains and two light chains. Additional examples of an antibody are described in this document.

[0053] As used in this document, the term “multispecific antibody” refers to an antibody that includes two or more different antigen-binding domains that collectively bind specifically to two or more different epitopes. The two or more different epitopes may be epitopes on the same antigen (e.g., a single polypeptide present on the surface of a cell) or on different antigens (e.g., different proteins present on the surface of the same cell or present on the surface of different cells). In some respects, a multispecific antibody binds to two different epitopes (i.e., a “bispecific antibody”). In some respects, a multispecific antibody binds to three different epitopes (i.e., a “trispecific antibody”). In some respects, a multispecific antibody binds to four different epitopes (i.e., a “quadrispecific antibody”).In some respects, a multispecific antibody binds to five different epitopes (i.e., a “pentaspecific antibody”). Each binding specificity can be present at any suitable valence. Non-limiting examples of multispecific antibodies are described in this document.

[0054] As used in this document, the term “bispecific antibody” refers to an antibody that binds to two different epitopes. The Petition 870250084299, dated 09 / 18 / 2025, pp. 286 / 410 16 / 126 epitopes can be on the same antigen or on different antigens.

[0055] As used in this document, the term “common light chain” refers to a light chain that can interact with two or more different heavy chains, forming different antigen-binding sites, wherein these different antigen-binding sites can specifically bind to different antigens or epitopes. Similarly, the term “common light chain variable region” refers to a light chain variable region that can interact with two or more different heavy chain variable regions, forming different antigen-binding sites, wherein these different antigen-binding sites can specifically bind to different antigens or epitopes. In some embodiments, the antibody or antigen-binding fragment thereof may have a common light chain. In some embodiments, the anti-HER3 / MUC1 antibody or antigen-binding fragment thereof may have a common light chain variable region.

[0056] As used in this document, the term “anti-HER3 / MUC1 antibody or antigen-binding fragment thereof” refers to an antibody or antigen-binding fragment that binds to both MUC1 and HER3.

[0057] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by a person of ordinary skill in the art to which the present invention pertains. Methods and materials are described herein for use in the present invention; other suitable methods and materials known in the art may also be used. The materials, methods, and examples are for illustrative purposes only and are not intended to be limiting. All publications, patent applications, patents, sequences, database entries, and other references mentioned herein are incorporated by reference in their entirety. In case of conflict, the Petition 870250084299, dated 09 / 18 / 2025, pp. 287 / 410 17 / 126 of this descriptive report, including the definitions, shall prevail.

[0058] Other features and advantages of the invention will be evident from the following detailed description and figures, and from the claims. Brief Description of the Figures

[0059] Figure 1 shows the mean tumor volumes in different groups of B-NDG mice that were injected with NUGC-4 cells and were treated with phosphate-buffered saline (PBS) or ADCs.

[0060] Figure 2 shows the mean tumor volumes in different groups of B-NDG mice that were grafted with patient-derived gastric tumor fragments (2 mm*2 mm*2 mm) and treated with PBS or ADCs.

[0061] Figure 3 shows the mean tumor volumes in different groups of B-NDG mice that were injected with HCC70 cells and were treated with PBS or ADCs.

[0062] Figure 4 shows the mean tumor volumes in different groups of B-NDG mice that were grafted with patient-derived pancreatic tumor fragments (2 mm*2 mm*2 mm) and were treated with PBS or ADCs.

[0063] Figure 5 shows the mean tumor volumes in different groups of B-NDG mice that were grafted with patient-derived lung tumor fragments (2 mm*2 mm*2 mm) and treated with PBS or ADCs.

[0064] Figure 6A shows the endocytosis rates of anti-HER3 antibodies, anti-MUC1 antibodies or bispecific anti-HER3 / MUC1 antibodies and ADCs in NUGC-4 cells.

[0065] Figure 6B shows antibody endocytosis rates Petition 870250084299, dated 09 / 18 / 2025, pp. 288 / 410 18 / 126 anti-HER3, anti-MUC1 antibodies or bispecific anti-HER3 / MUC1 antibodies and ADCs in NUGC-4 cells.

[0066] Figure 6C shows the endocytosis rates of anti-HER3 antibodies, anti-MUC1 antibodies or bispecific anti-HER3 / MUC1 antibodies and ADCs in NUGC-4 cells.

[0067] Figure 7 shows the mean tumor volumes in different groups of B-NDG mice that were grafted with patient-derived lung tumor fragments (2 mm x 2 mm x 2 mm) and treated with PBS or ADCs.

[0068] Figure 8 shows the mean tumor volumes in different groups of B-NDG mice that were grafted with NUGC-4 cells and treated with PBS or ADCs.

[0069] Figure 9A shows the mean tumor volumes in different groups of B-NDG mice that were grafted with HCC70 cells and were treated with PBS or ADCs.

[0070] Figure 9B shows the average body weights in different groups of B-NDG mice that were grafted with HCC70 cells and were treated with PBS or ADCs.

[0071] Figure 10 shows the mean tumor volumes in different groups of B-NDG mice that were grafted with patient-derived colorectal tumor fragments (2 mm x 2 mm x 2 mm) and treated with PBS or ADCs.

[0072] Figure 11 shows the mean tumor volumes in different groups of B-NDG mice that were grafted with patient-derived colorectal tumor fragments (2 mm x 2 mm x 2 mm) and treated with PBS or ADCs.

[0073] Figure 12A shows the mean tumor volumes in different groups of B-NDG mice that were grafted with Petition 870250084299, dated 09 / 18 / 2025, pp. 289 / 410 19 / 126 patient-derived pancreatic tumor fragments (2 mm*2 mm*2 mm) were treated with PBS or ADCs.

[0074] Figure 12B shows the mean body weights in different groups of B-NDG mice that were grafted with patient-derived pancreatic tumor fragments (2 mm*2 mm*2 mm) and were treated with PBS or ADCs.

[0075] Figure 13 shows the mean tumor volumes in different groups of B-NDG mice that were grafted with patient-derived breast tumor fragments (2 mm*2 mm*2 mm) and treated with PBS or ADCs.

[0076] Figure 14A shows the binding activities of anti-HER3 antibodies, anti-MUC1 antibodies, or bispecific anti-HER3 / MUC1 antibodies to NUGC-4 cells.

[0077] Figure 14B shows the binding activities of anti-HER3 antibodies, anti-MUC1 antibodies, or bispecific anti-HER3 / MUC1 antibodies to NCI-H226 cells.

[0078] Figure 15 shows the mean tumor volumes in different groups of B-NDG mice that were grafted with patient-derived gastric tumor fragments (2 mm*2 mm*2 mm) and were treated with PBS or ADCs.

[0079] Figure 16 shows the mean tumor volumes in different groups of B-NDG mice that were grafted with patient-derived colorectal tumor fragments (2 mm*2 mm*2 mm) and were treated with PBS or ADCs.

[0080] Figure 17 shows the mean tumor volumes in different groups of naked BALB / c mice that were grafted with patient-derived lung tumor fragments (2 mm*2 mm*2 mm) and treated with 5% glucose or ADC. Petition 870250084299, dated 09 / 18 / 2025, pages 290 / 410 20 / 126

[0081] Figure 18 shows the mean tumor volumes in different groups of naked BALB / c mice that were grafted with patient-derived ovarian tumor fragments (2 mm*2 mm*2 mm) and were treated with 5% glucose or ADC. Detailed Description of the Invention

[0082] A bispecific antibody or antigen-binding fragment of the same is an artificial protein that can simultaneously bind to two different epitopes (e.g., on two different antigens). In some embodiments, a bispecific antibody or antigen-binding fragment of the same may have two arms. Each arm may have a variable heavy chain region and a variable light chain region, forming an antigen-binding domain (or antigen-binding region). In some embodiments, the bispecific antibody has a common light chain.

[0083] The present invention relates to anti-HER3 / MUC1 antibodies (for example, bispecific antibodies or antigen-binding fragments thereof) that bind specifically to HER3 and MUC1, and antibody-drug conjugates derived from these anti-HER3 / MUC1 antibodies. Anti-HER3 / MUC1 antibody

[0084] HER3 is a pseudokinase member of the EGFR family that plays a role in both tumor progression and drug resistance. HER3 is an exceptional member of the EGFR family that is not oncogenic in isolation, but can cooperate with other receptors to induce tumorigenesis, metastatic events, and drug resistance. HER3 is a compelling therapeutic target against cancer. In contrast to EGFR and HER2, which have been widely targeted by TKIs, HER3 has been primarily targeted by monoclonal or bispecific antibodies due to minimal kinase activity, either by blocking ligand binding or heterodimerization with other receptors.

[0085] Ubiquitous expression of HER3 is detected in several types Petition 870250084299, dated 09 / 18 / 2025, pp. 291 / 410 21 / 126 of cancers, including breast, ovarian, colon, gastric, lung, skin, and pancreatic cancer. High HER3 expression is also associated with disease progression and / or poor prognosis in many cancer types. Although HER3 does not cause tumorigenesis on its own, HER2:HER3 heterodimers have the highest transformation capacity among all possible dimers in the EGFR family. The superior oncogenic capacity of the dimer pair makes HER3 critical for HER2-mediated tumorigenesis in several tumor types. In breast cancer cell lines, HER3 has been shown to be essential for maintaining cell viability, while EGFR has been shown to be dispensable.

[0086] HER3 expression acts as a bypass mechanism for various targeted therapies, and elevated HER3 signaling confers resistance to multiple therapeutic agents. For example, because HER3 dimerizes with receptors other than EGFR, including HER2 and the MET receptor, HER3 can confer resistance to EGFR-targeted therapies through dimerization with non-EGFR partners. HER3 expression is also connected to resistance to hormonal therapies. HER3 plays a critical role in HER2 phosphorylation in breast cancer cells, and downregulation of HER3 has reversed tamoxifen resistance of the antiestrogen receptor (ER) in breast cancer cell lines.

[0087] A detailed review of HER3 and its functions can be found in Haikala, Heidi M. and Pasi A. Jãnne. Thirty Years of HER3: From Basic Biology to Therapeutic Interventions. Cancer Clinical Research 27.13 (2021): 3528-3539; and Mishra, Rosalin, et al. HER3 Signaling and Targeted Therapy in Cancer. Oncology Reviews 12.1 (2018); Liu, Xiaolong, et al. Development of Effective HER3-Targeted Therapy for Cancer Treatment. Biological Procedures Online 21.1 (2019): 1-10, which is incorporated as a reference in its entirety.

[0088] Mucin 1 (MUC1; also known as episialin, PEM, Petition 870250084299, dated 09 / 18 / 2025, pp. 292 / 410 22 / 126 Mucinous cellulose (MUC1) is a type I single-pass transmembrane protein with a highly glycosylated extracellular domain extending 200–500 nm from the cell surface. MUC1 is normally expressed in glandular or luminal epithelial cells of the mammary gland, esophagus, stomach, duodenum, pancreas, uterus, prostate, and lungs, and to a lesser extent in hematopoietic cells. It is absent in skin epithelium and mesenchymal cells. In healthy tissues, MUC1 provides protection to the underlying epithelia. The extended, negatively charged sugar branches of MUC1 create a physical barrier and confer an anti-adhesive property to MUC1, which limits accessibility and prevents pathogenic colonization. The sugar chains oligomerize to form a mucinous gel that lubricates and protects the underlying epithelia from desiccation, pH changes, pollutants, and microbes.The abnormally glycosylated MUC1 is overexpressed in most human epithelial cancers and has gained considerable attention as an oncogenic molecule.

[0089] MUC1 is overexpressed in cancer cells, and the loss of cell polarity causes TA-MUC1 to be redistributed across the cell surface and into the cytoplasm. The lack of cell polarity also causes the redistribution of cell surface growth factors that are normally restricted to the basolateral surface of epithelial cells. Growth factors juxtaposed to MUC1 and intracellular kinases, such as ZAP-70, PKC-γ, GSK-3β, and c-Src, phosphorylate serine, tyrosine, and threonine residues in the MUC1 TC. Hypoglycosylation is also believed to unmask the peptide core of TA-MUC1, allowing cleavage and release of MUC1-N by extracellular proteases. The release of MUC1-N induces conformational changes in MUC1-C that alter its ligand status and subsequently activate downstream cell signaling pathways, such as mitogen-activated protein kinase (MAPK), phosphatidylinositol 3-kinase (P13K / Akt), and... Petition 870250084299, dated 09 / 18 / 2025, pp. 293 / 410 23 / 126 as wingless (Wnt) type pathways. As a result, MUC1-positive pancreatic, breast, lung, and colon cancer cells generally exhibit hyperactivation of these critical signaling pathways. MUC1-C also associates with several transcription factors (STAT3, NF-kB, p53, and β-catenin) and binds to the promoter region of the target gene to drive its expression. Several studies have indicated that MUC1 plays a critical role in the transcriptional regulation of genes associated with tumor invasion, metastasis, angiogenesis, proliferation, apoptosis, drug resistance, inflammation, and immune regulation.

[0090] A detailed review of MUC1 and its functions can be found in Nath, Sritama and Pinku Mukherjee. MUC1: a multifaceted oncoprotein with a key role in cancer progression. Trends in molecular medicine 20.6 (2014): 332-342, which is incorporated by reference in its entirety.

[0091] In some embodiments, the bispecific anti-HER3 / MUC1 antibody described herein may be designed to have an IgG1 subtype structure with knob-into-hole (KIH) mutations, which may promote heterodimerization and prevent mismatch between the two heavy chains. In some embodiments, the bispecific anti-HER3 / MUC1 antibody has a higher endocytosis rate than the corresponding monoclonal antibodies or the bispecific control antibodies.

[0092] In some embodiments, the bispecific antiHER3 / MUC1 antibody described in this document can be conjugated with a therapeutic agent, forming an antibody-drug conjugate (ADC). In some embodiments, the drug-to-antibody ratio (DAR) of the ADCs described in this document is about 3.8, about 3.9, about 4.0, about 4.1, about 4.2, about 4.3, about 4.4, about 4.5, about 4.6, or about 4.7. In some embodiments, the DAR of the ADCs described in this document is about 3.5 to about 4.5, about 3.6 to about Petition 870250084299, dated 09 / 18 / 2025, pp. 294 / 410 24 / 126 4.5, about 3.7 to about 4.5, about 3.8 to about 4.5, about 3.9 to about 4.5, about 4.0 to about 4.5, about 4.1 to about 4.5, about 4.2 to about 4.5, about 4.3 to about 4.5, about 4.4 to about 4.5, about 3.5 to about 4.4, about 3.6 to about 4.4, about 3.7 to about 4.4, about 3.8 to about 4.4, about 3.9 to about 4.4, about 4.0 to about 4.4, about 4.1 to about 4.4, about 4.2 to about 4.4, about 4.3 to about 4.4, about 3.5 to about 4.3, about 3.6 to about 4.3, about 3.7 to about 4.3, about 3.8 to about 4.3, about 3.9 to about 4.3, about 4.0 to about 4.3, about 4.1 to about 4.3, about 4.2 to about 4.3, about 3.5 to about 4.2, about 3.6 to about 4.2, about 3.7 to about 4.2, about 3.8 to about 4.2, about 3.9 to about 4.2, about 4.0 to about 4.2, about 4.1 to about 4.2, about 3.5 to about 4.1, about 3.6 to about 4.1approximately 3.7 to approximately 4.1, approximately 3.8 to approximately 4.1, approximately 3.9 to approximately 4.1, approximately 4.0 to approximately 4.1, approximately 3.5 to approximately 4.0, approximately 3.6 to approximately 4.0, approximately 3.7 to approximately 4.0, approximately 3.8 to approximately 4.0, approximately 3.9 to approximately 4.0, approximately 3.5 to approximately 3.9, approximately 3.6 to approximately 3.9, approximately 3.7 to approximately 3.9, approximately 3.8 to approximately 3.9, approximately 3.5 to approximately 3.8, approximately 3.6 to approximately 3.8, approximately 3.7 to approximately 3.8, approximately 3.5 to approximately 3.7, approximately 3.6 to approximately of 3.7 or about 3.5 to about 3.6. In some modalities, the DAR of the ADCs described in this document is about 7.5 to about 8.5, about 7.6 to about 8.5, about 7.7 to about 8.5, about 7.8 to about 8.5, about 7.9 to about 8.5, about 8.0 to about 8.5, about 8.1 to about 8.5, about 8.2 to about 8.5, about 8.3 to about 8.5, about 8.4 to about 8.5, about 7.5 to about 8.4, about 7.6 to about 8.4, about 7,7 to about 8.4, about 7.8 to about 8.4, about 7.9 to about 8.4, about 8.0 to about 8.4, about 8.1 to about 8.4, about 8.2 to about 8.4, about 8.3 to about 8.4, about 7.5 to about 8.3, about 7.6 to about 8.3, about 7.7 to about 8.3, about 7.8 to about 8.3, Petition 870250084299, dated 09 / 18 / 2025, pages 295 / 410 25 / 126 near 7.9 to near 8.3, near 8.0 to near 8.3, near 8.1 to near 8.3, near 8.2 to near 8.3, near 7.5 to near 8.2, near 7.6 to near 8.2, near 7.7 to near 8.2, near 7.8 to near 8.2, near 7.9 to near 8.2, near 8.0 to near 8.2, near 8.1 to near 8.2, near 7.5 to near 8.1, near 7.6 to near 8.1, near 7.7 to near 8.1, near 7.8 to near 8.1, near 7.9 to near 8.1, near 8.0 to near 8.1, near from 7.5 to about 8.0, about 7.6 to about 8.0, about 7.7 to about 8.0, about 7.8 to about 8.0, about 7.9 to about 8.0, about 7.5 to about 7.9, about 7.6 to about 7.9, about 7.7 to about 7.9, about 7.8 to about 7.9, about 7.5 to about 7.8, about 7.6 to about 7.8, about 7.7 to about 7.8, about 7.5 to about 7.7, about 7.6 to about 7.7 or about 7.5 to about 7.6.

[0093] In some embodiments, the anti-HER3 / MUC1 ADC described herein can effectively inhibit the growth of cancer cells in vitro at a concentration of less than 10 μg / ml, less than 3.33 μg / ml, less than 1.11 μg / ml, less than 0.37 μg / ml, less than 0.12 μg / ml, less than 0.04 μg / ml or less than 0.01 μg / ml. In some embodiments, the antiHER3 / MUC1 ADC described herein can inhibit the growth of cancer cells in vivo (e.g., lung cancer, gastric cancer, or skin cancer) in a mouse xenograft model at a dose level lower than 10 mg / kg, 9 mg / kg, 8 mg / kg, 7 mg / kg, 6 mg / kg, 5 mg / kg, 4 mg / kg, 3 mg / kg, 2 mg / kg, or 1 mg / kg.

[0094] In some embodiments, the anti-HER3 / MUC1 antibody described herein has a common light chain. In some embodiments, the anti-HER3 / MUC1 antibody includes an anti-HER3 antigen-binding domain (e.g., 1B2, 3G6, 3E1) or an anti-MUC1 antigen-binding domain (e.g., 10D1). In some embodiments, the anti-HER3 / MUC1 antibodies have a variable heavy chain region. Petition 870250084299, dated 09 / 18 / 2025, pp. 296 / 410 26 / 126 targeting HER3 (e.g., any of the HER3-targeted variable chains described herein), a heavy chain variable region targeting MUC1 (e.g., any of the MUC1-targeted variable chains described herein), and two identical common light chain variable regions.

[0095] The CDR sequences for the antigen-binding domain of 1B2 include CDRs of the heavy chain variable domain, SEQ ID NOs: 4 to 6, and CDRs of the light chain variable domain, SEQ ID NOs: 1 to 3, as defined by Kabat's definition. CDRs can also be defined by Chothia's definition. Under Chothia's definition, the heavy chain variable domain CDR sequences are presented in SEQ ID NOs: 16 to 18 and the light chain variable domain CDR sequences are presented in SEQ ID NOs: 13 to 15. The human light chain variable region and the human heavy chain variable region for 1B2 are presented in SEQ ID NO: 25 and SEQ ID NO: 26, respectively.

[0096] The CDR sequences for the antigen-binding domain of 3E1 include CDRs of the heavy chain variable domain, SEQ ID NOs: 7 to 9, and CDRs of the light chain variable domain, SEQ ID NOs: 1-3, as defined by the Kabat definition. Under the Chothia definition, the CDR sequences of the heavy chain variable domain are presented in SEQ ID NOs: 19 to 21 and the CDRs of the light chain variable domain are presented in SEQ ID NOs: 13 to 15. The human light chain variable region and the human heavy chain variable region for 3E1 are presented in SEQ ID NO: 25 and SEQ ID NO: 27, respectively.

[0097] The CDR sequences for the antigen-binding domain of 3G6 include CDRs of the heavy chain variable domain, SEQ ID NOs: 7, 42, 43 and CDRs of the light chain variable domain, SEQ ID NOs: 1-3, as defined by the Kabat definition. Under the Chothia definition, the Petition 870250084299, dated 09 / 18 / 2025, pp. 297 / 410 27 / 126 CDR sequences of the heavy chain variable domain are presented in SEQ ID NOs: 19, 20, 44, and the CDRs of the light chain variable domain are presented in SEQ ID NOs: 13 to 15. The human light chain variable region and the human heavy chain variable region for 3E1 are presented in SEQ ID NO: 25 and SEQ ID NO: 45, respectively.

[0098] The CDR sequences for the antigen-binding domain of 10D1 include CDRs of the heavy chain variable domain, SEQ ID NOs: 10 to 12, and CDRs of the light chain variable domain, SEQ ID NOs: 1-3, as defined by the Kabat definition. Under the Chothia definition, the CDR sequences of the heavy chain variable domain are presented in SEQ ID NOs: 22 to 24 and the CDRs of the light chain variable domain are presented in SEQ ID NOs: 13 to 15. The human light chain variable region and the human heavy chain variable region for 3E1 are presented in SEQ ID NO: 25 and SEQ ID NO: 28, respectively.

[0099] In some embodiments, the anti-HER3 / MUC1 antibodies described herein may contain one, two, or three variable region heavy chain CDRs selected from the group of SEQ ID NOs: 4-6, SEQ ID NOs: 7-9, SEQ ID NOs: 10-12, SEQ ID NOs: 7, 42, 43, SEQ ID NOs: 16-18, SEQ ID NOs: 19-21, SEQ ID NOs: 19, 20, 44, and SEQ ID NO: 22-24; and / or one, two, or three variable region light chain CDRs selected from the group of SEQ ID NOs: 1-3 and SEQ ID NOs: 13 to 15.

[0100] In some embodiments, anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof may have a variable heavy chain (VH) region comprising complementarity-determining regions (CDRs) 1, 2 and 3, wherein the CDR1 region comprises or consists of an amino acid sequence that is at least 80%, 85%, 90% or 95% identical to a selected VH CDR1 amino acid sequence, the CDR2 region comprises or consists of a sequence of Petition 870250084299, dated 09 / 18 / 2025, pp. 298 / 410 28 / 126 amino acids that are at least 80%, 85%, 90%, or 95% identical to a selected VH CDR2 amino acid sequence, and the CDR3 region comprises or consists of an amino acid sequence that is at least 80%, 85%, 90%, or 95% identical to a selected VH CDR3 amino acid sequence, and a variable light chain (VL) region comprising CDRs 1, 2, and 3, wherein the CDR1 region comprises or consists of an amino acid sequence that is at least 80%, 85%, 90%, or 95% identical to a selected VL CDR1 amino acid sequence, the CDR2 region comprises or consists of an amino acid sequence that is at least 80%, 85%, 90%, or 95% identical to a selected VL CDR2 amino acid sequence, and the region CDR3 comprises or consists of an amino acid sequence that is at least 80%, 85%, 90%, or 95% identical to a selected VL CDR3 amino acid sequence.The amino acid sequences of the selected VH CDRs 1, 2, and 3 and the amino acid sequences of the selected VL CDRs 1, 2, and 3 are shown in Tables 1 and 2.

[0101] In some embodiments, the anti-HER3 / MUC1 antibody or antigen-binding fragment described in this document may contain a variable heavy chain domain containing one, two, or three of the CDRs of SEQ ID NO: 4 with zero, one, or two amino acid insertions, deletions, or substitutions; of SEQ ID NO: 5 with zero, one, or two amino acid insertions, deletions, or substitutions; of SEQ ID NO: 6 with zero, one, or two amino acid insertions, deletions, or substitutions.

[0102] In some embodiments, the anti-HER3 / MUC1 antibody or antigen-binding fragment described in this document may contain a variable heavy chain domain containing one, two, or three of the CDRs of SEQ ID NO: 16 with zero, one, or two insertions, deletions, or amino acid substitutions; of SEQ ID NO: 17 with zero, one, or two insertions, deletions Petition 870250084299, dated 09 / 18 / 2025, pp. 299 / 410 29 / 126 or amino acid substitutions; of SEQ ID NO: 18 with zero, one or two insertions, deletions or amino acid substitutions.

[0103] In some embodiments, the anti-HER3 / MUC1 antibody or antigen-binding fragment described in this document may contain a variable heavy chain domain containing one, two, or three of the CDRs of SEQ ID NO: 7 with zero, one, or two amino acid insertions, deletions, or substitutions; of SEQ ID NO: 8 with zero, one, or two amino acid insertions, deletions, or substitutions; of SEQ ID NO: 9 with zero, one, or two amino acid insertions, deletions, or substitutions.

[0104] In some embodiments, the anti-HER3 / MUC1 antibody or antigen-binding fragment described in this document may contain a variable heavy chain domain containing one, two, or three of the CDRs of SEQ ID NO: 19 with zero, one, or two amino acid insertions, deletions, or substitutions; of SEQ ID NO: 20 with zero, one, or two amino acid insertions, deletions, or substitutions; of SEQ ID NO: 21 with zero, one, or two amino acid insertions, deletions, or substitutions.

[0105] In some embodiments, the anti-HER3 / MUC1 antibody or antigen-binding fragment described in this document may contain a variable heavy chain domain containing one, two, or three of the CDRs of SEQ ID NO: 7 with zero, one, or two amino acid insertions, deletions, or substitutions; of SEQ ID NO: 42 with zero, one, or two amino acid insertions, deletions, or substitutions; of SEQ ID NO: 43 with zero, one, or two amino acid insertions, deletions, or substitutions.

[0106] In some embodiments, the anti-HER3 / MUC1 antibody or antigen-binding fragment described in this document may contain a variable heavy chain domain containing one, two, or three of the CDRs of SEQ ID NO: 19 with zero, one, or two insertions, deletions, or amino acid substitutions; of SEQ ID NO: 20 with zero, one, or two insertions, deletions Petition 870250084299, dated 09 / 18 / 2025, pp. 300 / 410 30 / 126 or amino acid substitutions; of SEQ ID NO: 44 with zero, one or two insertions, deletions or amino acid substitutions.

[0107] In some embodiments, the anti-HER3 / MUC1 antibody or antigen-binding fragment described in this document may contain a variable heavy chain domain containing one, two, or three of the CDRs of SEQ ID NO: 10 with zero, one, or two amino acid insertions, deletions, or substitutions; of SEQ ID NO: 11 with zero, one, or two amino acid insertions, deletions, or substitutions; of SEQ ID NO: 12 with zero, one, or two amino acid insertions, deletions, or substitutions.

[0108] In some embodiments, the anti-HER3 / MUC1 antibody or antigen-binding fragment described in this document may contain a variable heavy chain domain containing one, two, or three of the CDRs of SEQ ID NO: 22 with zero, one, or two amino acid insertions, deletions, or substitutions; of SEQ ID NO: 23 with zero, one, or two amino acid insertions, deletions, or substitutions; of SEQ ID NO: 24 with zero, one, or two amino acid insertions, deletions, or substitutions.

[0109] In some embodiments, the anti-HER3 / MUC1 antibody or antigen-binding fragment described in this document may contain a variable light chain domain containing one, two, or three of the CDRs of SEQ ID NO: 1 with zero, one, or two amino acid insertions, deletions, or substitutions; of SEQ ID NO: 2 with zero, one, or two amino acid insertions, deletions, or substitutions; of SEQ ID NO: 3 with zero, one, or two amino acid insertions, deletions, or substitutions.

[0110] In some embodiments, the anti-HER3 / MUC1 antibody or antigen-binding fragment described in this document may contain a variable light chain domain containing one, two, or three of the CDRs of SEQ ID NO: 13 with zero, one, or two amino acid insertions, deletions, or substitutions; of SEQ ID NO: 14 with zero, one, or two insertions, deletions, or Petition 870250084299, dated 09 / 18 / 2025, pp. 301 / 410 31 / 126 amino acid substitutions; of SEQ ID NO: 15 with zero, one, or two insertions, deletions, or amino acid substitutions.

[0111] Insertions, deletions, and replacements can be within the CDR sequence or at one or both terminal ends of the CDR sequence.

[0112] In some embodiments, the anti-HER3 / MUC1 antibodies contain a variable heavy chain (VH) region comprising or consisting of an amino acid sequence that is at least 80%, 85%, 90%, or 95% identical to a selected VH sequence, and a variable light chain (VL) region comprising or consisting of an amino acid sequence that is at least 80%, 85%, 90%, or 95% identical to a selected VL sequence. In some embodiments, the selected VH sequence is SEQ ID NO: 26, 27, 45, or 28, and the selected VL sequence is SEQ ID NO: 25.

[0113] In some embodiments, the anti-HER3 / MUC1 antibody or antigen-binding fragment may have 3 HV CDRs that are identical to the CDRs of any HV sequences as described herein. In some embodiments, the anti-HER3 / MUC1 antibody or antigen-binding fragment may have 3 VL CDRs that are identical to the CDRs of any VL sequences as described herein.

[0114] The invention also provides nucleic acid comprising a polynucleotide encoding an anti-HER3 / MUC1 antibody. The immunoglobulin heavy chain or the immunoglobulin light chain in the anti-HER3 / MUC1 antibody comprises CDRs as shown in Table 1, Table 2 or Table 3 below.

[0115] Table 1 shows CDR sequences of the variable region of the antigen-binding domain anti-HER3 heavy chain (1B2, 3G6 and 3E1) and an antigen-binding domain anti-MUC1 (10D1) in antibodies against Petition 870250084299, dated 09 / 18 / 2025, page 302 / 410 32 / 126 HER3 / MUC1 as defined by Kabat's definition. Table 1 Ab CDR1 de VH SEQ ID NO: CDR2deVH SEQ ID NO: CDR3 de VH SEQID NO: CDR1 deVL SEQ ID NO: CDR2 deVL SEQ ID NO: CDR3 de VL SEQ ID NO: 1B2 NARMGVS 4 HIFSNDDKSYSP SLKS 5 MRYSGGPFDY 6 RASQSVSSYLA 1 DASNRAT 2 QQRSNWPP T 3 3E1 MRMGVS 7 HIFSNDDKSYSP SLKN 8 MKYSGGPFDY 9 RASQSVSSYLA 1 DASNRAT 2 QQRSNWPP T 3 10D1 SYGMH 10 VISYDGSNKYYA DSVKG 11 DRRGFYESGN YYNVPFDY 12 RASQSVSSYLA 1 DASNRAT 2 QQRSNWPP T 3

[0116] Table 2 shows CDR sequences of the antigen-binding domain heavy chain variable region (1B2, 3G6 and 3E1) and an anti-MUC1 antigen-binding domain (10D1) in antiHER3 / MUC1 antibodies as defined by Chothia's definition. Table 2 SEQ D NO: LA ΙΛ ui CDR3deVL QQRSNWPP T QQRSNWPP T QQRSNWPP T QQRSNWPP T SEQ ID NO: 3 1—1 t—1 i—l cm O. > Ο Ό DASNRAT DASNRAT DASNRAT DASNRAT SEQ ID NO: rn CDR1 deVL RASQSVSSYL A RASQSVSSYL A RASQSVSSYL A RASQSVSSYL A SEQ ID NO: oo Tl 3 CDR3deVH MRYSGGPFDY MKYSGGPFDY MRYSGGPFDY DRRGFYESGNYY NVPFDY â σ g WZ tf» oo CM X CC > O u U Ή FSNDD FSNDD FSNDD SYDGSN SEQ ID NO: 3 Ím CDR1 de VH TFSLSNARM GFSLSNIRM GFSLSNIRM GFTFSSY < 1B2 £ 3G6 10D1 Petition: 870250084299, on September 18, 2025, page. 303 / 410 33 / 126

[0117] There are 3 different types of amino acids extracted from the box. Tabela 3 SEQ ID NO ϋΊ CM 45 CO 29 30 31 32 Sequência de aminoácidos EIVLTQSPATLSLSPGERATLSCRASQSVS5YLAWYQQKPGQAPRLLIYDASNRATGIPARFSGSGSGTDFTLTISSLEPEDFAVYYCQ QRSNWPPTFGQGTKVEIK QVQLVQSGPVLVKPTETLILTCTVSTFSLSNARMGVSWIRQPPGKALEWLAHIFSNDDKSYSPSLKSRLTISKDTSKTQWLTMTN M DPVDIATYYCSRM RYSGGPFDYWGQGSLVTVSS EVQLVQSG PVLAKPTETLTLTCTVSGF5LSNIRMG V5WIRQPPGKALEWLAHIFSNDDK5YSPSLKN RLTISKDTSKSQWLTMnM DPM DTAIYYCTRM KYSGGPFDYWGQGTLVTVSS QVQLVQSGPVLVKPTETLTLTCTVSGFSLSNIRMGVSWIRQPPGKALEWLAHIFSNDDKSYSTSLKSRLTISKDTSKSQWLTMTNV DPVDTATYYCTRMRYSGGPFDYWGQGTLVTV5S QVTLKESGGGWQPG RSLRLSCAASGFTFSSYGM HWVRQAPG KGLEWVAVISYDGSNKYYADSVKG RFTI5RDN5KNTLYLQMN SLRAEDTAVYYCANDRRGFYESGNYYNVPFDYWGQGTLVTVSS RTVAAPSVFIFPPSDEQLKSGTASWCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYAC EVTHQGLSSPVTKSFNRGEC A5TKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNVNHK PSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVWDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPCIVYTLPPCREEMTKNQVSLWCLVKGFYPSDI AVEWESNGQPENNYKTTPPVLDSDGSFFLYSKL7VDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNVNHK PSNTKVDKKVEPKSCDKTHTCPPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVWDVSHEDPEVKFNWYVDGVEVHNAKTKP REEQYNSTYRWSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVCTLPPSREEMTKNQVSLSCAVKGFYPSDIA VEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEVGNSQAVCPGTLNGLSVTGDAENQYQTLYKLYERCEWMGNLEIVLTGHNADLSFLQWIREVTGYVLVAMNEFSTLPLPNLRV VRGTQVYDGKFAIFVMLNYNTNSSHALRQLRLTQLTEILSGGVYIEKNDKLCHMDTIDWRDIVRDRDAEIWKDNGRSCPPCHEVC Antibody Domains Common human light chain variable region Human heavy chain variable region 1B2 Human heavy chain variable region 3E1 Human heavy chain variable region 3G6 Human heavy chain variable region Human heavy chain 10D1 Constant region of the light chain lgG1 Constant region of the heavy chain lgG1 with a protrusionConstant region of the IgG 1 heavy chain with a human HER3 hole (hHER3) Petition 870250084299, dated 09 / 18 / 2025, pp. 304 / 410 34 / 126 33 34 KGRCWGPGSEDCQTLTKTICAPQCNGHCFGPNPNQCCHDECAGGCSGPQDTDCFACRHFNDSGACVPRCPQPLVYNKLTFQLE PNPHTKYQYGGVCVASCPHNFWDQTSCVRACPPDKMEVDKNGLKIVICEPCGGLCPKACEGTGSGSRFQTVDSSNIDGFVNCTK ILGNLDFLITGLNGDPWHKIPALDPEKLNVFRTVREITGYLNIQSWPPHMHNFSVFSNLTTIGGRSLYNRGFSLUMKNLNVTSLGFR SLKEISAGRIYISANRQLCYHHSLNWTKVLRGPTEERLDIKHNRPRRDCVAEGKVCDPLCSSGGCWGPGPGQCLSCRNYSRGGVCV THCNFLNGEPREFAHEAECFSCHPECQPMEGTATCNGSGSDTCAQCAHFRDGPHCVSSCPHGVLGAKGPIYKYPDVQNECRPCH ENCTQGCKG PELQDCLGQTLVLIGKTH LT SEVGNSQAVCPGTLNGLSVTGDAENQYQTLYKLYERCEWMGNLEIVLTGHNADLSFLQWIREVTGYVLVAMNEFSTLPLPNLRV VRGTQVYDGKFAIFVMLNYNTNSSHALRQLRLTQLTEILSGGVYIEKNDKLCHMDTJDWKDIVRDQDAEIWKDNGRSCPLCHEVC KGRCWGPGPEDCQTLTKTICAPQCNGHCFGPNPNQCCHDECAGGCSGPQPTDCFACRHFNDSGACVPRCPQPLVYNKLTFQLE PNPHTKYQYGGVCVASCPHNFWDQTSCVRACPPDKMEVDKNGLKMCEPCGGLCPKACEGTGSGSRFQTVDSSNIDGFVNCTK ILGNLDFLITGLNGDPWHKIPALDPEKLNVFRTVREITGYLNIQ5WPPHMYNFSVFSNLTTIGGRSLYNRGFSLLIMKNLNVTSLGFR SLKEISAGRIYISANRQLCYHHSLNWTKVLRGPTEERLDIKHNRPRRDCVAEGKVCDPLCSSGGCWGPGPGQCLSCRNYSRGGVCVTHCNFLNGEPREFAHEAECFSCHPECQPMEGTATCNGSGSDTCAQCAHFRDGPHCVSSCPHGVLGAKGPIYKYPDVQNECRPCH ENCTQGCKG PELQDCLGQTLVLIGKTH LT MTPGTQSPFFLLLLLTVLTWTGSGHASSTPGGEKETSATQRSSVPSSTEKNAVSMTSSVLSSHSPGSGSSTTQGQDVTLAPATEPA SGSAATWGQpVTSVPVTRPALGSTTPPAHDVTSAPDNKPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGST APPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPA HGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVT SAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVT5APDTRPAPGSTAPPAHGVT5APD TRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAP GSTAPPAHGVT5APDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTA PPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAH GVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVrSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTS APDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAPGSTAPPAHGVTSAPDTRPAP monkey HER3 (fasHER3) human MUC1 (hMUC1) Petition 870250084299, dated 09 / 18 / 2025, pages 305 / 410 35 / 126 LTJ 37 00 OΊ 40 41 GSTAPPAHGVTSAPDNRPALGSTAPPVHNVTSASGSASGSASTLVHNGTSARATTTPASKSTPFSIPSHHSDTPTTLASHSTKTDAS STHHSSVPPLTSSNHSTSPQLSTGVSFFFLSFHISNLQFNSSLEDPSTDYYQELQRDISEMFLQIYKQGGFLGLSNIKFRPGSVWQLTL AFREGTINVHDVETQFNQYKTEAASRYNLTISDVSVSDVPFPFSAQSGAGVPGWGIALLVLVCVLVALAIVYUALAVCQCRRKNYG QLDIFPARDTYHPMSEYPTYHTHGRYVPPS5TDRSPYEKVSAGNGGS5LSYTNPAVAATSANL ASDSASGSASTLVHSTTSARATTTPASKSTPFSIPSHHSDTPTTLASHSTKTDASSTHHSTVPPFTSSNHSTSPQLSLGVSFFFLSFHIS NLQFNSSLEDPSTNYYQQLQRDISELFLQIYKQGDFLGLSNIMFRPGSVWQSTLVFREGTTNVHDVETQFNQRKTEAASRYNLTIS DISVRDVPFPFSAQTGAGVPGWG QVQLQQWGAGLLKPSETLSLTCAVYGGSFSGYYWSWIRQPPGKGLEWIGEINHSGSTNYNPSLKSRVTISVETSKNQFSLKLSSVT AADTAVYYCARDKWTWYFDLWG RGTLVTVSS DIEMTQ5PDSLAVSLGERATINCRSSQSVLYSSSNRNYLAWYQQNPGQPPKLLIYWA5TRESGVPDRFSGSGSGTDFTLTISSLQAE DVAVYYCQQYYSTPRTFGQGTKVEIK QVQLQQWGAGLLKPSETLSLTCAVYGGSFSGYYWSWIRQPPGKGLEWIGEINHSGSTNYNPSLKSRVTISVETSKNQFSLKLSSVT AADTAVYYCARDKWTWYFDLWG RGTLVTVSS DIEMTQSPDSLAVSLGERATINCRSSQSVLYSSSNRNYLAWYQQNPGQPPKLLIYWASTRESGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCQQYYSTPRTFGQGTKVEI K EVQLQQSGAELAKPGASVKMSCKVSGYTFTSYWMHWVKQRPGQGLEWIGYINPGTGYIEYNQKFKDKATLTADKSSSTAYMQL SSLTSE DSAVYYCASSTAPFDYWGQGTTLTVSS DIVITQSPSSMYASLGERVTITCKASQDIKSYLSWYQQKPWKSPKTLIYYATRLADGVPSRFSGSGSGQDYSLTISSLESDDTATYYCL QYDESPYTFGGGTKLEIK Monkey MUC1 (fasMUCI) Gatipotuzumab heavy chain variable region Gatipotuzumab light chain variable region Patritumab heavy chain variable region Patritumab light chain variable region Patritumab heavy chain variable region 1H7 light chain variable region 1H7 Petition 870250084299, dated 09 / 18 / 2025, pp. 306 / 410 36 / 126

[0118] When polypeptides are paired with a corresponding polypeptide (for example, a corresponding heavy chain variable region or a corresponding light chain variable region), the paired polypeptides bind to MUC1 and / or HER3.

[0119] Anti-HER3 / MUC1 antibodies may also be variants of anti-HER3 / MUC1 antibodies (including derivatives and conjugates) or antibody fragments. Additional anti-HER3 / MUC1 antibodies provided herein are polyclonal, monoclonal, multispecific (multimeric, e.g., bispecific), human antibodies, chimeric antibodies (e.g., mouse-human chimera), single-chain antibodies, intracellularly produced (i.e., intrabody) antibodies, and antigen-binding fragments thereof. Anti-HER3 / MUC1 antibodies may be of any type (e.g., IgG, IgE, IgM, IgD, IgA, and IgY), class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or subclass. In some embodiments, the antiHER3 / MUC1 antibody or antigen-binding fragment is an IgG antibody (e.g., IgG1) or antigen-binding fragment of the same.

[0120] Anti-HER3 / MUC1 antibody fragments are suitable for use in the methods provided as long as they retain the desired affinity and specificity for both MUC1 and HER3. Thus, a fragment of an anti-HER3 / MUC1 antibody will retain the ability to bind to both MUC1 and HER3. Antibodies and antigen-binding fragments thereof

[0121] In some embodiments, the multispecific anti-HER3 / MUC1 antibody (e.g., bispecific antibody) includes an antigen-binding domain that is derived from an anti-HER3 antibody and an antigen-binding domain that is derived from an anti-MUC1 antibody. These anti-HER3 / MUC1 antibodies and their antigen-binding fragments may have several Petition 870250084299, dated 09 / 18 / 2025, pp. 307 / 410 37 / 126 forms.

[0122] In general, antibodies (also called immunoglobulins) can be composed of two classes of polypeptide chains, light chains and heavy chains. A non-limiting anti-HER3 / MUC1 antibody of the present invention may be an intact antibody with four immunoglobulin chains comprising two heavy chains and two light chains. The heavy chain of the anti-HER3 / MUC1 antibody may be of any isotype including IgM, IgG, IgE, IgA or IgD or subisotype including IgG1, IgG2, IgG2a, IgG2b, IgG3, IgG4, IgE1, IgE2, etc. The light chain may be a kappa light chain or a lambda light chain.

[0123] The hypervariable regions, known as complementarity-determining regions (CDRs), form loops that comprise the main antigen-binding surface of the antibody. The four scaffold regions largely adopt a beta-sheet conformation, and the CDRs form loops that connect and, in some cases, form part of the beta-sheet structure. The CDRs in each chain are held in close proximity by the scaffold regions and, with the CDRs of the other chain, contribute to the formation of the antigen-binding domain.

[0124] Methods for identifying the CDR regions of an antibody by analyzing the antibody's amino acid sequence are well known, and several definitions of CDRs are commonly used. The Kabat definition is based on sequence variability, and the Chothia definition is based on the location of structural loop regions. These methods and definitions are described in, for example, Martin, “Protein sequence and structure analysis of antibody variable domains”, Antibody engineering, Springer Berlin Heidelberg, 2001. 422 to 439; Abhinandan, et al. “Analysis and improvements to Kabat and structurally correct numbering of antibody variable domains”, Molecular immunology 45(14) (2008): 3832 to 3839; Wu, TT and Kabat, EA (1970) J. Exp. Petition 870250084299, dated 09 / 18 / 2025, pp. 308 / 410 38 / 126 Med. 132: 211 to 250; Martin et al., Enzymol. Methods 203:121-53 (1991); Morea et al., Biophys Chem. 68(1 to 3): 9 to 16 (October 1997); Morea et al., J Mol Biol. 275(2):269-94 (January 1998); Chothia et al., Nature 342(6252):877-83 (December 1989); Ponomarenko and Bourne, BMC Structural Biology 7:64 (2007); each of which is incorporated into this document by reference in its entirety.

[0125] CDRs are important for recognizing an antigen epitope. As used in this document, an “epitope” is the smallest portion of a target molecule capable of being specifically bound by the antigen-binding domain of an antibody. The minimum size of an epitope can be about three, four, five, six, or seven amino acids, but these amino acids do not need to be in a consecutive linear sequence of the antigen's primary structure, as the epitope can depend on the three-dimensional configuration of an antigen based on the antigen's secondary and tertiary structures.

[0126] In some embodiments, the anti-HER3 / MUC1 antibody is an intact immunoglobulin molecule (e.g., IgG1, IgG2a, IgG2b, IgG3, IgM, IgD, IgE, IgA). IgG subclasses (IgG1, IgG2, IgG3, and IgG4) are highly conserved, differing in their constant region, particularly in their hinges and upper CH2 domains. The sequences and differences of IgG subclasses are known in the art and are described, for example, in Vidarsson, et al., “IgG subclasses and allotypes: from structure to effector functions”. Frontiers in Immunology 5 (2014); Irani, et al. “Molecular properties of human IgG subclasses and their implications for the design of therapeutic monoclonal antibodies against infectious diseases.” Molecular Immunology 67.2 (2015): 171-182; Shakib, Farouk, ed. “The subclasses of human IgG: molecular analysis of structure, function and regulation”. Elsevier, 2016; each of which is incorporated into this document by reference in its entirety. Petition 870250084299, dated 09 / 18 / 2025, pp. 309 / 410 39 / 126

[0127] The anti-HER3 / MUC1 antibody may also be an immunoglobulin molecule that is derived from any species (e.g., human, rodent, mouse, rat, camelid). The antigen-binding domain or antigen-binding fragment is a portion of an antibody that retains specific binding activity of the intact antibody, that is, any portion of an antibody that is capable of specifically binding to an epitope on the target molecule of the intact antibody. It includes, for example, Fab, Fab', F(ab')2 and variants of these fragments.Thus, in some embodiments, an anti-HER3 / MUC1 antibody or antigen-binding fragment thereof may comprise, for example, an scFv, an Fv, an Fd, a dAb, a bispecific antibody, a bispecific scFv, a diabody, a linear antibody, a single-chain antibody molecule, a multispecific antibody formed from antibody fragments, and any polypeptide that includes a binding domain that is, or is homologous to, an antibody-binding domain. Non-limiting examples of antigen-binding domains include, for example, the heavy chain and / or light chain CDRs of an intact antibody, the variable regions of the heavy and / or light chain of an intact antibody, full-length heavy or light chains of an intact antibody, or an individual heavy chain or light chain CDR of an intact antibody.

[0128] In some embodiments, the scFv in an anti-HER3 / MUC1 antibody has two variable heavy chain domains and two variable light chain domains. In some embodiments, the anti-HER3 / MUC1 scFv has two antigen-binding regions (antigen-binding regions: A and B), and the two antigen-binding regions can bind to the respective target antigens with different affinities.

[0129] In some embodiments, anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof may comprise one, two, or three variable region heavy chain CDRs selected from Petition 870250084299, dated 09 / 18 / 2025, pages 310 / 410 40 / 126 Tables 1 and 2.

[0130] In some embodiments, the anti-HER3 / MUC1 antibodies described herein may be conjugated with a therapeutic agent. The anti-HER3 / MUC1 antibody-drug conjugate comprising the antibody or antigen-binding fragment thereof may bind covalently or non-covalently to a therapeutic agent. In some embodiments, the therapeutic agent is a cytotoxic or cytostatic agent (e.g., monomethylauristatin E, monomethylauristatin F, cytochalasin B, gramicidin D, ethidium bromide, emetine, mitomycin, etoposide, tenoposide, vincristine, vinblastine, colchicine, doxorubicin, daunorubicin, dihydroxyanthracine, maytansinoids such as DM-1 and DM-4, ​​dione, mitoxantrone, mithramycin, actinomycin D, 1-dehydrotestosterone, glucocorticoids, procaine, tetracaine, lidocaine, propranolol, puromycin, epirubicin, and cyclophosphamide and analogues). In some embodiments, the therapeutic agent is MMAE or MMAF.In some embodiments, the therapeutic agent is conjugated via a linker, for example, a VC linker. Details of the linkers used for ADCs can be found, for example, in Su, Z. et al. “Antibody-drug conjugates: Recent advances in linker chemistry”. Acta Pharmaceutica Sinica B (2021), which is incorporated herein by reference in its entirety.

[0131] In some embodiments, the anti-HER3 / MUC1 antibody is a bispecific antibody. Bispecific antibodies can be produced by manipulating the interface between a pair of antibody molecules to maximize the percentage of heterodimers that are recovered from recombinant cell culture. For example, the interface may contain at least a portion of the CH3 domain of an antibody constant domain. In this method, one or more small amino acid side chains of the interface of the first antibody molecule are replaced by larger side chains (by Petition 870250084299, dated 09 / 18 / 2025, pages 311 / 410 41 / 126 example, tyrosine or tryptophan). Compensatory “cavities” of identical or similar size to the large side chain(s) are created at the interface of the second antibody molecule by replacing large amino acid side chains with smaller chains (e.g., alanine or threonine). This provides a mechanism for increasing heterodimer yield relative to other undesirable end products, such as homodimers. This method is described, for example, in document no. WO 96 / 27011, which is incorporated by reference in its entirety.

[0132] Any of the anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof described herein may be conjugated with a stabilizing molecule (e.g., a molecule that increases the half-life of the antibody or antigen-binding fragment thereof in a subject or in solution). Non-limiting examples of stabilizing molecules include: a polymer (e.g., a polyethylene glycol) or a protein (e.g., serum albumin, such as human serum albumin). Conjugation with a stabilizing molecule may increase the half-life or prolong the biological activity of an anti-HER3 / MUC1 antibody or antigen-binding fragment in vitro (e.g., in tissue culture or when stored as a pharmaceutical composition) or in vivo (e.g., in a human).

[0133] Anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof can also have various forms. Many different formats of bispecific antibodies or antigen-binding fragments thereof are known in the art and are described, for example, in Suurs, et al. “A review of bispecific antibodies and antibody constructs in oncology and clinical challenges”, Pharmacology & therapeutics (2019), which is incorporated herein by reference in its entirety.

[0134] In some modalities, the anti-HER3 / MUC1 antibody is Petition 870250084299, dated 09 / 18 / 2025, pp. 312 / 410 42 / 126 a BiTe, a (scFv)2, a nanobody, a nanobody-HSA, a DART, a TandAb, a scDiabody, a scDiabody-CH3, a scFv-CH-CL-scFv, an HSAbody, a scDiabody-HAS or a tandem scFv.In some embodiments, the anti-HER3 / MUC1 antibody is a VHH-scAb, a VHH-Fab, a Dual scFab, an F(ab')2, a diabody, a crossMab, a DAF (two in one), a DAF (four in one), a DutaMab, a DT-IgG, a common light chain of bumps in holes, a set of bumps in holes, a cargo pair, a Fab arm exchange, a SEEDbody, a LUZ-Y, an Fcab, a κλ body, an orthogonal Fab, a DVD-IgG, an IgG(H)-scFv, a scFv-(H)IgG, IgG(L)-scFv, scFv-(L)IgG, IgG(L,H)Fv, IgG(H)-V, V(H)-IgG, IgG(L)-V, V(L)-IgG, KIH IgG-scFab, 2scFv-IgG, IgG2scFv, scFv4-Ig, Zybody, DVI-IgG, Diabody-CH3, a triple body, a miniantibody, a minibody, a TriBi minibody, scFv-CH3 KIH, Fab-scFv, an F(ab')2-scFv2, a scFv-KIH, a Fab-scFv-Fc, a tetravalent HCAb, a scDiabody-Fc, a Diabody-Fc, a tandem scFv-Fc, an intrabody, a Dock and Lock, an lmmTAC, an IgG-IgG conjugate, a Cov-X-Body, or a scFv1PEG-scFv2.

[0135] In some embodiments, the anti-HER3 / MUC1 antibody may be a TrioMab. In a TrioMab, the two heavy chains are of different species, where different sequences restrict the pairing of heavy and light chains.

[0136] In some embodiments, the anti-HER3 / MUC1 antibody has two different heavy chains and a common light chain. Heavy chain heterodimerization may be based on bump-in-holes or some other heavy chain pairing technique.

[0137] In some modalities, the CrossMAb technique can be used to produce bispecific anti-HER3 / MUC1 antibodies. The CrossMAb technique can be used to enhance the correct association of light chains. Petition 870250084299, dated 09 / 18 / 2025, pp. 313 / 410 43 / 126 in bispecific heterodimeric IgG antibodies, this technique allows the generation of various bispecific antibody formats, including bi-(1+1), tri-(2+1), and tetra-(2+2)valent bispecific antibodies, as well as antibodies based on non-Fc tandem antigen-binding fragment (Fab). These formats can be derived from any existing antibody pair using domain permutation, without the need for common light chain identification, post-translational processing / in vitro chemical assembly, or the introduction of a set of mutations that reinforce the correct light chain association. The method is described in Klein et al., “The use of CrossMAb technology for the generation of bi- and multispecific antibodies”. MAbs. Vol. 8. No. 6. Taylor & Francis, 2016, which is incorporated by reference in full. In some embodiments, the CH1 in the heavy chain and the CL domain in the light chain are exchanged.

[0138] The anti-HER3 / MUC1 antibody may be a duobody. The Fab exchange mechanism that occurs naturally in IgG4 antibodies is mimicked in a controlled manner in IgG1 antibodies, a mechanism called controlled Fab exchange. This format may ensure a specific pairing between the heavy and light chains.

[0139] In dual-variable-domain immunoglobulin (DVD-Ig), additional VH and variable light chain (VL) domains are added to each N-terminus for bispecific targeting. This format resembles IgG-scFv, but the added binding domains are individually linked to their respective N-terminus instead of one scFv to each N-terminus of the heavy chain.

[0140] In scFv-IgG, the two scFv molecules are connected to the C-terminal of the heavy chain (CH3). The scFv-IgG format has two different bivalent binding sites and is consequently also called tetravalent. There is no heavy chain-light chain pairing problem in scFv-IgG. Petition 870250084299, dated 09 / 18 / 2025, pp. 314 / 410 44 / 126

[0141] In some embodiments, the anti-HER3 / MUC1 antibody may have an IgG-IgG format. Two intact IgG antibodies are conjugated by chemically linking the C-terminus of the heavy chains.

[0142] The anti-HER3 / MUC1 antibody can also have a Fab-scFv-Fc format. In the Fab-scFv-Fc format, a light chain, a heavy chain, and a third chain containing the Fc and scFv regions are assembled. This can ensure efficient fabrication and purification.

[0143] In some embodiments, the anti-HER3 / MUC1 antibody may be a TF. Three Fab fragments are linked by disulfide bridges. Two fragments are directed to the tumor-associated antigen (TAA) and one fragment is directed to a hapten. The TF form does not have an Fc region.

[0144] ADAPTIR has two scFvs attached to each side of an Fc region. It abandons intact IgG as a base for its construction, but retains the Fc region to prolong half-life and facilitate purification.

[0145] Dual affinity retargeting (DART) has two peptide chains connecting opposite fragments, such as VLA with VHB and VLB with VHA, and a sulfur bond at their C-terminus fusing them together. In DART, the sulfur bond can improve stability compared to BiTEs.

[0146] In DART-Fc, an Fc region is attached to the DART structure. It can be generated by assembling three chains, two through a disulfide bond, as with DART. One chain contains half of the Fc region which dimerizes with the third chain, expressing only the Fc region. The addition of the Fc region increases the half-life leading to effective concentrations for longer, avoiding continuous IR.

[0147] In tetravalent DART, four peptide chains are assembled. Basically, two DART molecules are created with half of an Fc region and will undergo dimerization. This format has a bivalent bond to both. Petition 870250084299, dated 09 / 18 / 2025, pages 315 / 410 45 / 126 targets, so it is a tetravalent molecule.

[0148] Tandem diabody (TandAb) comprises two diabodies. Each diabody consists of a VHA and VLB fragment and a VHA and VLB fragment that are covalently associated. The two diabodies are linked by a peptide chain. It may improve stability compared to the diabody consisting of two scFvs. It has two bivalent binding sites.

[0149] The ScFv-scFv-toxin includes toxin and two scFv with a stabilizing linker. It can be used for targeted payload delivery.

[0150] In some embodiments, the anti-HER3 / MUC1 antibody is a bispecific antibody. In some embodiments, the bispecific antibody in the present invention is designed to be 1+1 (monovalent for each target) and has an IgG1 subtype structure. This can reduce avidity for cells with low levels of HER3 and MUC1 expression, and increase avidity for cells that co-express HER3 and MUC1, to achieve enhanced targeting function.

[0151] In some embodiments, anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof have a constant light chain region that is at least 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 29, and a constant heavy chain region that is at least 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to any of the SEQ ID Nos: 30 and 31.

[0152] In some embodiments, anti-HER3 / MUC1 antibodies include KIH mutations. In some embodiments, the anti-HER3 / MUC1 antibody includes a first antigen-binding domain that binds specifically to HER3 and a second antigen-binding domain that binds specifically to MUC1. In some embodiments, the first antigen-binding domain Petition 870250084299, dated 09 / 18 / 2025, pp. 316 / 410 46 / 126 to the antigen includes a heavy chain that includes one or more protrusion mutations (a protrusion heavy chain), and the second antigen-binding domain includes a heavy chain that includes one or more hole mutations (a hole heavy chain). In some embodiments, the first antigen-binding domain includes a heavy chain that includes one or more hole mutations (a hole heavy chain), and the second antigen-binding domain includes a heavy chain that includes one or more protrusion mutations (a protrusion heavy chain). In some embodiments, the anti-HER3 / MUC1 antibody includes a protrusion heavy chain comprising a constant region that is at least 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 30.In some embodiments, the anti-HER3 / MUC1 antibody includes a hole heavy chain comprising a constant region that is at least 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 31. Characteristics of Antibodies

[0153] Anti-HER3 / MUC1 antibodies may include an anti-HER3 antigen-binding domain and any anti-MUC1 antigen-binding domain as described in this document.

[0154] The invention provides anti-HER3 / MUC1 antibodies and antigen-binding fragments thereof that can specifically bind to HER3. These anti-HER3 / MUC1 antibodies can be agonists or antagonists. The anti-HER3 / MUC1 antibodies, or antigen-binding fragments thereof described herein, can bind to HER3 and block the binding between HER3 and its ligands. By blocking the binding between HER3 and its ligands, the anti-HER3 / MUC1 antibodies can inhibit the HER3-associated signaling pathway and thus treat cancer. In some Petition 870250084299, dated 09 / 18 / 2025, pp. 317 / 410 47 / 126 modes, anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof can initiate CMC or ADCC.

[0155] General techniques that can be used to measure the affinity of an antibody for an antigen include, for example, ELISA, RIA, and surface plasmon resonance (SPR). Affinities can be deduced from the quotient of the kinetic rate constants (KD = koff / kon). In some implementations, anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof can bind to HER3 (e.g., human HER3, monkey HER3, mouse HER3, and / or chimeric HER3) with a dissociation rate (koff) less than 0.1 s-1, less than 0.01 s-1, less than 0.001 s-1, less than 0.0001 s-1, or less than 0.00001 s-1. In some modalities, the dissociation rate (koff) is greater than 0.01 s-1, greater than 0.001 s-1, greater than 0.0001 s-1, greater than 0.00001 s-1, or greater than 0.000001 s-1.

[0156] In some modalities, the kinetic association rates (kon) are greater than 1 x 102 / Ms, greater than 1 x 103 / Ms, greater than 1 x 104 / Ms, greater than 1 x 105 / Ms or greater than 1 x 106 / Ms. In some modalities, the kinetic association rates (kon) are less than 1 x 105 / Ms, less than 1 x 106 / Ms or less than 1 x 107 / Ms.

[0157] In some embodiments, anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof can bind to HER3 (e.g., human HER3, monkey HER3, mouse HER3, and / or chimeric HER3) with a KD less than 1 x 10-6M, less than 1 x 10-7M, less than 1 x 10-8M, less than 1 x 10-9M, or less than 1 χ 10-10M. In some embodiments, the KD is less than 50 nM, 40 nM, 30 nM, 20 nM, 10 nM, 9 nM, 8 nM, 7 nM, 6 nM, 5 nM, 4 nM, 3 nM, 2 nM, or 1 nM. In some disciplines, the KD is greater than 1 x 10-7M, greater than 1 x 10-8M, greater than 1 x 10-9M, or greater than 1 x 10-10M.

[0158] Anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof may also include a binding domain to Petition 870250084299, dated 09 / 18 / 2025, pages 318 / 410 48 / 126 antigen that can specifically bind to MUC1. Anti-HER3 / MUC1 antibodies, or antigen-binding fragments thereof described herein, can block the binding between MUC1 and its ligands. In some embodiments, by binding to MUC1, the anti-HER3 / MUC1 antibody can also inhibit signaling pathways associated with MUC1, thereby inhibiting cell proliferation, differentiation, and / or metastasis. Thus, in some embodiments, the anti-HER3 / MUC1 antibodies as described herein are MUC1 agonists. In some embodiments, the anti-HER3 / MUC1 antibodies are MUC1 antagonists.

[0159] In some implementations, antiHER3 / MUC1 antibodies or antigen-binding fragments thereof can bind to MUC1 (e.g., human MUC1, monkey MUC1, mouse MUC1, and / or chimeric MUC1) with a dissociation rate (koff) less than 0.1 s-1, less than 0.01 s-1, less than 0.001 s-1, less than 0.0001 s-1, or less than 0.00001 s-1. In some embodiments, the dissociation rate (koff) is greater than 0.01 s-1, greater than 0.001 s-1, greater than 0.00001 s-1, greater than 0.000001 s-1, or greater than 0.000001 s-1.

[0160] In some modalities, the kinetic association rates (kon) are greater than 1 x 102 / Ms, greater than 1 x 103 / Ms, greater than 1 x 104 / Ms, greater than 1 x 105 / Ms or greater than 1 x 106 / Ms. In some modalities, the kinetic association rates (kon) are less than 1 x 105 / Ms, less than 1 x 106 / Ms or less than 1 x 107 / Ms.

[0161] Affinities can be deduced from the quotient of the kinetic rate constants (KD = koff / kon). In some modalities, KD is less than 1 x 10⁻⁶ M, less than 1 x 10⁻⁷ M, less than 1 x 10⁻⁸ M, less than 1 x 10⁹ M, or less than 1 x 10⁻¹⁰ M. In some modalities, KD is less than 50 nM, 40 nM, 30 nM, 20 nM, 15 nM, 10 nM, 9 nM, 8 nM, 7 nM, 6 nM, 5 nM, 4 nM, 3 nM, 2 nM, or 1 nM. In some modalities, KD is greater than 1 x 10⁻⁷ M, greater Petition 870250084299, dated 09 / 18 / 2025, pp. 319 / 410 49 / 126 at 1 x 10-8M, greater than 1 x 10-9M or greater than 1 x 10-10M.

[0162] Since the anti-HER3 / MUC1 antibody (e.g., bispecific antibody) binds to both MUC1 and HER3, for cells expressing both MUC1 and HER3, the antibody has a higher binding affinity to these cells. Avidity can be used to measure the binding affinity of an antibody to these cells. Avidity is the cumulative strength of multiple affinities of individual non-covalent binding interactions.

[0163] Thermal stabilities can also be determined. Anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof, as described in this document, may have Tm values ​​greater than 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, or 95 °C. Since IgG can be described as a multidomain protein, the melting curve sometimes shows two transitions, with a first denaturation temperature, Tm D1, and a second denaturation temperature, Tm D2. The presence of these two peaks often indicates the denaturation of the Fc (Tm D1) and Fab (Tm D2) domains, respectively. When there are two peaks, Tm usually refers to Tm D2.Thus, in some embodiments, anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof, as described in this document, have a Tm D1 greater than 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, or 95 °C. In some embodiments, the anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof, as described in this document, have a Tm D2 greater than 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, or 95 °C. In some modalities, Tm, Tm D1, Tm D2 are lower than 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, or 95 °C. Petition 870250084299, dated 09 / 18 / 2025, pp. 320 / 410 50 / 126

[0164] In some embodiments, anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof can bind to human HER3 or monkey HER3. In some embodiments, anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof cannot bind to human HER3 or monkey HER3. In some embodiments, anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof can bind to human MUC1 or monkey MUC1. In some embodiments, anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof cannot bind to human MUC1 or monkey MUC1.

[0165] In some embodiments, the anti-HER3 / MUC1 antibody, antigen-binding fragment or ADC has a purity that is greater than 30%, 40%, 50%, 60%, 70%, 72.5%, 75%, 77.5%, 80%, 82.5%, 85%, 87.5%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%, for example, as measured by HPLC. In some forms, the purity is less than 30%, 40%, 50%, 60%, 70%, 72.5%, 75%, 77.5%, 80%, 82.5%, 85%, 87.5%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%, for example, as measured by HPLC.

[0166] In some embodiments, the anti-HER3 / MUC1 antibody, antigen-binding fragment, or ADC has a purity above 90%, above 91%, above 92%, above 93%, above 94%, above 95%, above 96%, above 97%, or above 98%, as determined by size exclusion chromatography (SEC). In some embodiments, the anti-HER3 / MUC1 antibody, antigen-binding fragment, or ADC has a hydrophobic interaction chromatography (HIC) retention time greater than 2 minutes, greater than 3 minutes, greater than 4 minutes, or greater than 5 minutes. In some embodiments, the HIC retention time is less than 2 minutes, less than 3 minutes, less than 4 minutes, less than 5 minutes, or less than Petition 870250084299, dated 09 / 18 / 2025, pp. 321 / 410 51 / 126 minutes.

[0167] In some embodiments, the anti-HER3 / MUC1 antibody, antigen-binding fragment or ADC has a purity above 85%, above 86%, above 87%, above 88%, above 89%, above 90%, above 91%, above 92%, above 93%, above 94%, above 95%, above 96%, above 97%, above 98% or above 99%, as determined by capillary electrophoresis - sodium dodecyl sulfate (CE-SDS).

[0168] In some embodiments, the anti-HER3 / MUC1 antibody, antigen-binding fragment or ADC has a main peak that constitutes more than 40%, more than 50%, more than 55%, more than 60%, more than 65%, more than 70%, more than 75%, more than 80%, more than 85% or more than 90% of the total sample, as determined by capillary isoelectric focusing (cIEF). In some embodiments, the anti-HER3 / MUC1 antibody, antigen-binding fragment, or ADC has an acid peak that constitutes less than 10%, less than 15%, less than 20%, less than 25%, less than 30%, less than 35%, less than 40%, less than 45%, less than 50%, less than 55%, less than 60%, less than 65%, less than 70%, or less than 75% of the total sample, as determined by capillary isoelectric focusing (cIEF).

[0169] In some embodiments, the anti-HER3 / MUC1 antibody, antigen-binding fragment or ADC has a rate or percentage of tumor growth inhibition (% ICT) that is greater than 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 110%, 120%, 130%, 140%, 150%, 160%, 170%, 180%, 190%, or 200%. In some modalities, the antiHER3 / MUC1 antibody, antigen-binding fragment, or ADC has a percentage of tumor growth inhibition that is less than 60%, 70%, 80%, 90%, 100%, 110%, 120%, 130%, 140%, or 150%. The ICT (%) can be determined, for example, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, or 41 days after Petition 870250084299, dated 09 / 18 / 2025, pp. 322 / 410 52 / 126 the start of treatment. As used in this document, the rate or percentage of tumor growth inhibition (% ICT) is calculated using the following formula: ICT (%) = [1-(Ti-T0) / (Vi-V0)]x100 %

[0170] Ti is the mean tumor volume in the treatment group on day i. T0 is the mean tumor volume in the treatment group on day zero. Vi is the mean tumor volume in the control group on day i. V0 is the mean tumor volume in the control group on day zero.

[0171] In some embodiments, the anti-HER3 / MUC1 antibody, antigen-binding fragment, or ADC has a functional Fc region. In some embodiments, the effector function of a functional Fc region is antibody-dependent cell-mediated cytotoxicity (ADCC). In some embodiments, the effector function of a functional Fc region is phagocytosis. In some embodiments, the effector function of a functional Fc region is ADCC and phagocytosis. In some embodiments, the Fc region is human IgG1, human IgG2, human IgG3, or human IgG4.

[0172] In some embodiments, the anti-HER3 / MUC1 antibody, antigen-binding fragment, or ADC does not have a functional Fc region. For example, anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof are Fab, Fab', F(ab')2, and Fv fragments. In some embodiments, the anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof as described herein have an Fc region without effector function. In some embodiments, the Fc is a human IgG4 Fc. In some embodiments, the Fc does not have a functional Fc region. For example, the Fc region has LALA mutations (L234A and L235A mutations in EU numbering) or LALA-PG mutations (L234A, L235A, P329G mutations in EU numbering).

[0173] Some other modifications can be made to the Fc region. For example, a cysteine ​​residue(s) can be introduced into Petition 870250084299, dated 09 / 18 / 2025, pp. 323 / 410 53 / 126 Fc region, thus allowing the formation of an interchain disulfide bond in this region. The homodimeric fusion protein thus generated can have any half-life increased in vitro and / or in vivo.

[0174] In some forms, IgG4 has the S228P mutation (EU numbering). The S228P mutation prevents Fab arm exchange of IgG4 in vivo and in vitro.

[0175] In some embodiments, the Fc regions are provided with a carbohydrate structure that lacks fucose attached (directly or indirectly) to an Fc region. For example, the amount of fucose in such an Fc region composition may be 1% to 80%, 1% to 65%, 5% to 65%, or 20% to 40%. The amount of fucose is determined by calculating the average amount of fucose in the sugar chain at Asn297, relative to the sum of all glycostructures attached to Asn297 (e.g., complex, hybrid, and high-mannose structures) as measured by MALDI-TOF mass spectrometry, as described in document no. WO 2008 / 077546, for example.Asn297 refers to the asparagine residue located at approximately position 297 in the Fc region (EU numbering of Fc region residues; or position 314 in Kabat numbering); however, Asn297 may also be located approximately ± 3 amino acids upstream or downstream of position 297, i.e., between positions 294 and 300, due to small sequence variations in Fc region sequences. Such fucosylation variants may have enhanced ADCC function. In some embodiments, to reduce glycan heterogeneity, the Fc region may be further manipulated to replace asparagine at position 297 with alanine (N297A).

[0176] In some embodiments, the main HPLC-SEC peak represents at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, 99.5%, 100% of the protein complex described in this document after purification by protein A-based affinity chromatography and / or Petition 870250084299, dated 09 / 18 / 2025, pp. 324 / 410 54 / 126 size exclusion chromatography.

[0177] In some embodiments, the anti-HER3 / MUC1 ADC described herein has an IC50 for in vitro cancer cell killing of less than 5 μg / ml, less than 4.5 μg / ml, less than 4 μg / ml, less than 3.5 μg / ml, less than 3 μg / ml, less than 2.5 μg / ml, less than 2 μg / ml, less than 1.5 μg / ml, less than 1 μg / ml, less than 0.9 μg / ml, less than 0.8 μg / ml, less than 0.7 μg / ml, less than 0.6 μg / ml, less than 0.5 μg / ml, less than 0.4 μg / ml, less than 0.3 μg / ml, less than 0.2 μg / ml, less than 0.1 μg / ml, less than 0.05 μg / ml, less than 0.025 μg / ml, less than 0.0125 μg / ml, less than 0.005 μg / ml, or less than 0.0025 μg / ml.

[0178] In some embodiments, the bispecific anti-HER3 / MUC1 antibody described herein has a higher endocytosis rate than the corresponding monoclonal antibodies and / or bispecific control antibodies described herein. In some embodiments, the anti-HER3 / MUC1 antibody described herein has a higher endocytosis rate than the Patritumab analog and / or Gatipotuzumab analog. Antibody-drug conjugates (ADGs)

[0179] Anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof described in this document may be conjugated with a therapeutic agent (a drug). The therapeutic agent may be covalently or non-covalently linked to the anti-HER3 / MUC1 antibody. In some embodiments, the anti-HER3 / MUC1 antibody is a bispecific anti-HER3 / MUC1 antibody. In some embodiments, the bispecific antibody has a common light chain.

[0180] In some embodiments, the therapeutic agent is a cytotoxic or cytostatic agent (e.g., monomethylauristatin E, monomethylauristatin F, cytochalasin B, gramicidin D, ethidium bromide, emetine, mitomycin, etoposide, tenoposide, vincristine, vinblastine, Petition 870250084299, dated 09 / 18 / 2025, pp. 325 / 410 55 / 126 colchicine, doxorubicin, daunorubicin, dihydroxyanthracine, maytansinoids such as DM-1 and DM-4, ​​dione, mitoxantrone, mithramycin, actinomycin D, 1-dehydrotestosterone, glucocorticoids, procaine, tetracaine, lidocaine, propranolol, puromycin, epirubicin and cyclophosphamide and analogues). Useful classes of cytotoxic, cytostatic or immunomodulatory agents include, for example, anti-tubulin agents, minor groove DNA ligands, DNA replication inhibitors and alkylating agents.

[0181] In some embodiments, the therapeutic agent may include, but is not limited to, cytotoxic reagents, such as chemotherapeutic agents, immunotherapeutic agents and the like, antiviral agents or antimicrobial agents. In some embodiments, the therapeutic agent to be conjugated may be selected from, but is not limited to, MMAE (monomethylauristatin E), MMAD (monomethylauristatin D) or MMAF (monomethylauristatin F).

[0182] Definitions of specific functional groups and chemical terms are described in more detail below. For the purposes of the present invention, chemical elements are identified according to the “Periodic Table of the Elements”, CAS version, Handbook of Chemistry and Physics, 75th edition, inside cover, and specific functional groups are generally defined as described therein. Additionally, general principles of organic chemistry, as well as specific functional moieties and reactivity, are described in Organic Chemistry, Thomas Sorrell, University Science Books, Sausalito, 1999; Smith and March, March's Advanced Organic Chemistry, 5th edition, John Wiley & Sons, Inc., New York, 2001; Larock, Comprehensive Organic Transformations, VCH Publishers, Inc., New York, 1989; Carruthers, Some Modern Methods of Organic Synthesis, 3rd edition, Cambridge University Press, Cambridge, 1987.

[0183] All ranges mentioned in this document are inclusive, unless expressly stated otherwise. When a Petition 870250084299, dated 09 / 18 / 2025, pp. 326 / 410 The 56 / 126 range of values ​​is mentioned, it is intended to cover each value and sub-range within the range. For example, “C1-6” is intended to cover C1, C2, C3, C4, C5, C6, C1-6, C1-5, C1-4, C1-3, C1-2, C2-6, C2-5, C2-4, C2-3, C3-6, C3-5, C3-4, C4-6, C4-5 and C5-6.

[0184] The compounds or any formula representing and describing the compounds of the present invention may have one or more chiral (asymmetric) centers. The present invention encompasses all stereoisomeric forms of the compounds or any formula representing and describing the compounds of the present invention. The asymmetry centers present in the compounds or any formula representing and describing the compounds of the present invention may all independently of each other have either (R) or (S) configuration. When the bonds to a chiral carbon are represented as straight lines in the structural formulas, or when the name of a compound is recited without a chiral (R) or (S) designation for a chiral carbon, it is understood that both (R) and (S) configurations of each such chiral carbon, and therefore each enantiomer or diastereomer and mixtures thereof, are encompassed in the formula or by the name.

[0185] The invention includes all possible enantiomers and diastereomers and mixtures of two or more stereoisomers, for example, mixtures of enantiomers and / or diastereomers, in all ratios. Thus, enantiomers are an object of the invention in enantiomerically pure form, both as levorotatory and dextrorotatory antipodes, in the form of racemates and in the form of mixtures of the two enantiomers in all ratios. In the case of cis / trans isomerism, the invention includes both the cis and trans forms, as well as mixtures of these forms in all ratios. The preparation of individual stereoisomers can be carried out, if desired, by separating a mixture by conventional methods, for example by chromatography or crystallization, by using stereochemically uniform starting materials for the synthesis or by stereoselective synthesis. Petition 870250084299, dated 09 / 18 / 2025, pp. 327 / 410 57 / 126 Optionally, derivatization can be performed prior to stereoisomer separation. Separation of a mixture of stereoisomers can be performed at an intermediate step during the synthesis of a compound or can be done on a final racemic product. Absolute stereochemistry can be determined by X-ray crystallography of crystalline products or crystalline intermediates that are derivatized, if necessary, with a reagent containing a stereogenic center of known configuration. Alternatively, absolute stereochemistry can be determined by vibrational circular dichroism (VCD) spectroscopy analysis.

[0186] Unless otherwise indicated, the structures represented herein are also intended to include compounds that differ only in the presence of one or more isotopically enriched atoms, in other words, compounds in which one or more atoms are replaced by atoms having the same atomic number but an atomic mass or mass number different from the atomic mass or mass number that predominates in nature. Such compounds are called an “isotopic variant”. The present invention is intended to include all pharmaceutically acceptable isotopic variants of the compounds or any formula representing and describing the compounds of the present invention.Examples of isotopes suitable for inclusion in the compounds of the present invention include, but are not limited to, isotopes of hydrogen, such as 2H (i.e., D) and 3H; carbon, such as 11C, 13C and 14C; chlorine, such as 36Cl; fluorine, such as 18F; iodine, such as 123I and 125I; nitrogen, such as 13N and 15N; oxygen, such as 15O, 17O and 18O; phosphorus, such as 32P; and sulfur, such as 35S. Certain isotopic variants of the compounds or of any formula representing and describing the compounds of the present invention, for example, those incorporating a radioactive isotope, may be useful in studies of tissue distribution of drugs and / or substrates. In particular, the compounds... Petition 870250084299, dated 09 / 18 / 2025, pp. 328 / 410 58 / 126 with the represented structures, differing only in the substitution by heavier isotopes, such as the substitution of hydrogen by deuterium (2H or D), may provide certain therapeutic advantages, for example, resulting from greater metabolic stability, increased in vivo half-life or reduced dosage requirements, and therefore may be used in some particular circumstances. Isotopic variants of compounds or any formula representing and describing the compounds of the present invention can generally be prepared by techniques known to those skilled in the art or by processes analogous to those described in the accompanying examples and synthesis using an appropriate isotopically labeled reagent in place of the unlabeled reagent previously employed.

[0187] The compounds as provided in this document are described with reference to both generic formulas and specific compounds. Furthermore, the compounds of the present invention may exist in various different forms or derivatives, all within the scope of the invention. These include, for example, pharmaceutically acceptable salts, tautomers, stereoisomers, racemic mixtures, regioisomers, prodrugs, solvated forms, different crystalline forms or polymorphs, and active metabolites, etc.

[0188] As used in this document, the term “pharmaceutically acceptable salt,” unless otherwise indicated, includes salts that retain the biological efficacy of the free acid / base form of the specified compound and that are not biologically or otherwise undesirable. Pharmaceutically acceptable salts may include salts formed with inorganic bases or acids and organic bases or acids. In cases where the compounds of the present invention contain one or more acidic or basic groups, the invention also includes their corresponding pharmaceutically acceptable salts. Thus, the compounds of the present invention containing acidic groups, such as carboxyl groups, may be present in salt form and may be used Petition 870250084299, dated 09 / 18 / 2025, pp. 329 / 410 59 / 126 according to the invention, for example, as alkali metal salts, alkaline earth metal salts, aluminum salts or as ammonium salts. Further non-limiting examples of such salts include lithium salts, sodium salts, potassium salts, calcium salts, magnesium salts, barium salts or salts with ammonia or organic amines, such as ethylamine, ethanolamine, diethanolamine, triethanolamine, piperidine, n-methylglutamine or amino acids. These salts are readily available, for example, by reacting the compound with an acid group with a suitable base, for example, lithium hydroxide, sodium hydroxide, sodium propoxide, potassium hydroxide, potassium ethoxide, magnesium hydroxide, calcium hydroxide or barium hydroxide. Other base salts of compounds of the present invention include, but are not limited to, copper (I), copper (II), iron (II), iron (III), manganese (II) and zinc salts.The compounds of the present invention containing one or more basic groups, for example, groups that can be protonated, may be present in salt form and may be used according to the invention in the form of their addition salts with inorganic or organic acids.Examples of suitable acids include hydrogen chloride, hydrogen bromide, hydrogen iodide, phosphoric acid, sulfuric acid, nitric acid, methanesulfonic acid, p-toluenesulfonic acid, naphthalenedisulfonic acid, sulfoacetic acid, trifluoroacetic acid, oxalic acid, acetic acid, tartaric acid, lactic acid, salicylic acid, benzoic acid, carbonic acid, formic acid, propionic acid, pivalic acid, diethylacetic acid, malonic acid, succinic acid, pimelic acid, fumaric acid, malonic acid, maleic acid, malic acid, embonic acid, mandelic acid, sulfaminic acid, phenylpropionic acid, gluconic acid, ascorbic acid, isonicotinic acid, citric acid, adipic acid, taurocholic acid, glutaric acid, stearic acid, glutamic acid or aspartic acid, and other acids known to people versed in the technique.The salts that are formed include, among others, hydrochlorides, chlorides, hydrobromides, bromides, iodides, sulfates, and phosphates. Petition 870250084299, dated 09 / 18 / 2025, pages 330 / 410 60 / 126 methanesulfonates (mesylates), tosylates, carbonates, bicarbonates, formates, acetates, sulfoacetates, triflates, oxalates, malonates, maleates, succinates, tartrates, malates, embonates, mandelates, fumarates, lactates, citrates, glutarates, stearates, aspartates and glutamates. The stoichiometry of the salts formed from the compounds of the invention may, moreover, be an integral or non-integral multiple of one.

[0189] The compounds of the present invention containing basic nitrogen-containing groups can be quaternized using agents such as C1-4 alkyl halides, for example, methyl, ethyl, isopropyl and tert-butyl chloride, bromide and iodide; diC1-4 alkyl sulfates, for example, dimethyl, diethyl and diamyl sulfate; C10-18 alkyl halides, for example, decyl, dodecyl, lauryl, myristyl and stearyl chloride, bromide and iodide; and (C1-4) arylalkyl halides, for example, benzyl chloride and phenethyl bromide.

[0190] If the compounds of the present invention contain both acidic and basic groups in the molecule, the invention also includes, in addition to the salt forms mentioned, internal salts or betaines (zwitterions). The respective salts can be obtained by usual methods known to those skilled in the art, for example, by contacting them with an organic or inorganic acid or base in a solvent or dispersant, or by anionic exchange or cationic exchange with other salts. The present invention also includes all salts of the compounds of the present invention which, due to low physiological compatibility, are not directly suitable for use in pharmaceutical products, but which can be used, for example, as intermediates for chemical reactions or for the preparation of pharmaceutically acceptable salts. For a review of more suitable salts, see Stahl and Wermuth, Handbook of Pharmaceutical Salts: Properties, Selection, and Use (Wiley-VCH, 2002).

[0191] The compound or any formula that represents and describes Petition 870250084299, dated 09 / 18 / 2025, pp. 331 / 410 61 / 126 The compounds of the present invention and pharmaceutically acceptable salts thereof may exist in non-solvated and solvated forms. As used herein, the term “solvate” refers to a molecular complex comprising the compound of Formula (I), or a pharmaceutically acceptable salt thereof, and one or more molecules of pharmaceutically acceptable solvent. For example, the term “hydrate” is used when the solvent is water.

[0192] Pharmaceutically acceptable solvates according to the present invention may include those in which the crystallization solvent can be isotopically replaced, for example, D2O, d6-acetone, d6-DMSO. Binder (binding agent compound)

[0193] In some embodiments, the therapeutic agent is conjugated via a ligand (or a linking agent compound). As used herein, the term “ligand” or “linking agent compound” refers to a compound that can connect a ligand (e.g., antibodies or antigen-binding fragments thereof described herein) to a therapeutic agent (e.g., any of the therapeutic agents described herein) to form a ligand-drug conjugate by reaction with a group of the ligand compound and the therapeutic agent compound, respectively, by, for example, a coupling reaction.

[0194] In some embodiments, the ligand described in this document is a compound that has the following formula: QL Formula (I), or a pharmaceutically acceptable salt, solvate, stereoisomer or isotopic variant thereof, wherein Q denotes a chemical junction moiety capable of being coupled to a ligand via a linkage selected from the group consisting of carbonyl, thioether, amide, disulfide and hydrazone linkages; Petition 870250084299, dated 09 / 18 / 2025, pp. 332 / 410 62 / 126 L denotes a chemical portion of a ligand capable of connecting Q to a therapeutic agent.

[0195] In some embodiments, the chemical junction portion (Q in Formula (I)) has the following structure: the

[0196] In some embodiments, the chemical portion of the ligand (L in Formula (I)) has the following formula: φ-L-COOH L2 where Li is a polypeptide residue consisting of three to eight amino acid residues comprising at least one amino acid residue with a side-chain carboxyl group, for example, a glutamic acid residue or an aspartic acid residue, wherein “-COOH” denotes the carboxyl group of an amino acid residue at the C-terminus of the polypeptide residue; l_2 is absent or is a hydrophilic monodentate, bidentate, or tridentate group attached to the side-chain carboxyl group on the amino acid residue of the polypeptide residue Li, and l_2 has a structure of NHC(RL2a)(RL2b)(RL2c), wherein RL2a, RL2b, and RL2c are each independently selected from the group consisting of H, (CH2O)(CH2CH2O)m(CH2)pC(O)OH, and -(CH2O)(CH2CH2O)m(CH2)PC(O)NH; RL2d, RL2d, and RL2d are H or C1-6 alkyl optionally substituted with 1 to 6 hydroxyl groups, each m is independently an integer from 0 to 10, preferably 0 to 4, for example 0, 1, 2, 3, or 4, especially preferably m is 0, and each p is independently an integer from 1 to 4, for example For example, 1, 2, 3 or 4; and <► denotes the N-terminal side of the polypeptide residue covalently linked to the Q-junction chemical portion. Petition 870250084299, dated 09 / 18 / 2025, pp. 333 / 410 63 / 126

[0197] In some embodiments, the polypeptide residue Li is NHGlu-Val-Ala-COOH. In some embodiments, the hydrophilic group L2 has the following structure: OH OH where “*” denotes the site covalently linked to the polypeptide residue Li, for example, the side chain of the Glu residue in NH-Glu-Val-AlaCOOH

[0198] In some embodiments, the ligand described in this document is a compound that has the following structure: CPT-L

[0199] In some embodiments, the linker is a VC linker. Details of the linkers used for ADCs can be found, for example, in Su, Z. et al. “Antibody-drug conjugates: Recent advances in linker chemistry”. Acta Pharmaceutica Sinica B (2021), which is incorporated herein by reference in its entirety. Petition 870250084299, dated 09 / 18 / 2025, pp. 334 / 410 64 / 126 Therapeutic agent

[0200] In some embodiments, the therapeutic agent that is conjugated with the antibodies or antigen-binding fragments thereof described in this document is discussed as follows.

[0201] In some embodiments, the therapeutic agent described in this document is a cytotoxic agent. In some embodiments, the cytotoxic agent is a camptothecin compound, an analogue, or a derivative thereof. In some preferred embodiments, the camptothecin compound is a compound that has the following structure: where X is selected from the group consisting of -CH2-, O, and S; Y is selected from the group consisting of H, D, and F.

[0202] In some embodiments, the therapeutic agent is (S)-4-amino-9-ethyl-9-hydroxy-1,9,12,15-tetrahydro-13H-pyrano[3',4':6,7]indolizino[1,2b]thiopyrano[4,3,2-de]quinoline-10,13(2H)-dione) (CPT-1). The structure of CPT-1 is shown below: CPT-1

[0203] In some embodiments, the therapeutic agent is (S)-4-amino-9-ethyl-9-hydroxy-1,9,12,15-tetrahydro-13H-pyrano[4,3,2de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13(2H)-dione (CPT-2). The structure of CPT-2 is shown below: Petition 870250084299, dated 09 / 18 / 2025, pp. 335 / 410 65 / 126 CPT-2

[0204] In some modalities, the therapeutic agent is CPT3. The structure of CPT-3 is shown below:

[0205] In some embodiments, the therapeutic agent is (S)-4-amino-9-ethyl-5-fluoro-9-hydroxy-1,9,12,15-tetrahydro-13H-pyrano[4,3,2de]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13(2H)-dione (CPT-4). The structure of CPT-4 is shown below: CPT-4

[0206] In some embodiments, the therapeutic agent is an auristatin, such as auristatin E (also known in the art as a dolastatin-10 derivative) or a derivative thereof. The auristatin may be, for example, an ester formed between auristatin E and a keto acid. For example, auristatin E may be reacted with para-acetylbenzoic acid or benzoylvaleric acid to produce AEB and AEVB, respectively. Other typical auristatins include AFP, MMAF, and MMAE. The synthesis and structure of auristatins Petition 870250084299, dated 09 / 18 / 2025, pp. 336 / 410 Examples of patents 66 / 126 are described in Patent Application Publication No. US20030083263; International Patent Publication No. WO 04 / 010957, International Patent Publication No. WO 02 / 088172 and in US Patents Nos. 7,498,298, 6,884,869, 6,323,315; 6,239,104; 6,034,065; 5,780,588; 5,665,860; 5,663,149; 5,635,483; 5,599,902; 5,554,725; 5,530,097; 5,521,284; 5,504,191; 5,410,024; 5,138,036; 5,076,973; 4,986,988; 4,978,744; 4,879,278; 4,816,444 and 4,486,414, each of which is incorporated by reference into this document in its entirety and for all purposes.

[0207] Auristatins have been shown to interfere with microtubule dynamics and nuclear and cell division and have anticancer activity. Auristatins bind to tubulin and can exert a cytotoxic or cytostatic effect on the cancer cell. There are a number of different assays, known in the art, that can be used to determine whether an auristatin or resulting antibody-drug conjugate exerts a cytostatic or cytotoxic effect on a desired cell.

[0208] In some embodiments, the therapeutic agent is a chemotherapeutic agent. Examples of chemotherapeutic agents include alkylating agents, such as thiotepa and cyclophosphamide (CYTOXAN™); alkyl sulfonates, such as busulfan, improsulfan and piposulfan; aziridines, such as benzodopa, carboquone, meturedopa and uredopa; ethyleneimines and methylamylamines, including altretamine, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphoramide and trimethylolomelamine; nitrogen mustards, such as chlorambucil, chlornaphazine, colofosfamide, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, melphalan, novembichin, fenesterin, prednimustine, trofosfamide, uracil mustard; nitrosureas, such as carmustine, chlorozotocin, fotemustine, lomustine, nimustine, ranimustine; antibiotics, such as aclacinomysins, actinomycin, authramycin, azaserine, bleomycins, cactinomycin, calicheamicin, carabicin, Petition 870250084299, dated 09 / 18 / 2025, pp. 337 / 410 67 / 126 carminomycin, carzinophylline, chromomycins, dactinomycin, daunorubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin, epirubicin, esorrubicin, idarubicin, marcelomycin, mitomycins, mycophenolic acid, nogalamycin, olivomycins, peplomycin, potfiromycin, puromycin, chelamycin, rhodorubicin, streptonigrin, streptozocin, tubecidin, ubenimex, zinostatin, zorubicin; antimetabolites such as methotrexate and 5-fluorouracil (5-FU); folic acid analogues, such as denopterin, methotrexate, pteropterin, trimetrexate; Purine analogs, such as fludarabine, 6-mercaptopurine, tiamiprine, thioguanine; pyrimidine analogs, such as ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxyfluridine, enocitabine, floxuridine, 5-FU; androgens, such as calusterone, dromostanolone propionate, epitiostanol, mepitiostane, testolactone; antiadrenals, such as aminoglutethimide, mitotane, trilostane;folic acid supplementation, such as frolinic acid; aceglathone; aldophosphamide glycoside; aminolevulinic acid; amsacrine; bestrabucil; bisanthrene; edatraxate; defofamine; demecolcine; diaziquone; elfornitrine; elliptinium acetate; etoglucid; gallium nitrate; hydroxyurea; lentinan; lonidamine; mitoguazone; mitoxantrone; mopidamol; nitracrine; pentostatin; fenamet; pirarubicin; podophyllinic acid; 2-ethylhydrazide; procarbazine; PSK7; razoxane; sizofiran; spirogermanium; tenuazonic acid; triaziquone; 2',2',2'-trichlorotriethylamine; urethane; vindesine; dacarbazine; manomustine; mitobronitol; mitolactol; pipobroman; gacytosine; arabinoside (“Ara-C”); cyclophosphamide; taxanes, for example, paclitaxel (TAXOL®, Bristol-Myers Squibb Oncology, Princeton, NJ) and doxetaxel (TAXOTERE®, Rhone-Poulenc Rorer, Antony, France); chlorambucil; gemcitabine; 6-thioguanine; platinum analogs, such as cisplatin and carboplatin; vinblastine; platinum; etoposide (VP-16); ifosfamide;mitomycin C; mitoxantrone; vincristine; vinorelbine; navelbine; novantron; teniposide; daunomycin; aminopterin; xeloda; ibandronate; CPT-11; inhibitor of; Petition 870250084299, dated 09 / 18 / 2025, pp. 338 / 410 68 / 126 topoisomerase RFS 2000; difluoromethylornithine (DMFO); retinoic acid; esperamycins; capecitabine and pharmaceutically acceptable salts, acids or derivatives of any of the above. Also included in this definition are anti-hormonal agents that act to regulate or inhibit hormonal action in tumors, such as antiestrogens including, for example, tamoxifen, raloxifene, 4(5)-imidazole aromatase inhibitors, 4-hydroxytamoxifen, trioxifene, ceoxifene, LY117018, onapristone and toremifene (Fareston); and antiandrogens, such as flutamide, nilutamide, bicalutamide, leuprolide and goserelin; and pharmaceutically acceptable salts, acids or derivatives of any of the above. A detailed description of chemotherapeutic agents can be found in, for example, US20180193477A1, which is incorporated by reference in its entirety. Binding compound - therapeutic agent

[0209] In some embodiments, a ligand (for example, any of the ligands described in this document) and a therapeutic agent (for example, any of the therapeutic agents described in this document) can be linked to form a “ligand-therapeutic agent” compound.

[0210] In some embodiments, the therapeutic agent-binding compound has the following structure:

[0211] In some embodiments, the binding agent compound Petition 870250084299, dated 09 / 18 / 2025, pp. 339 / 410 The therapeutic 69 / 126 has the following structure:

[0212] In some embodiments, an antibody (“Ab”), for example, any of the antibodies or antigen-binding fragments thereof described in this document, may be linked to a therapeutic agent-linker compound (for example, any of the therapeutic agent-linker compounds described in this document) to generate an antibody-drug conjugate. In some embodiments, the antibody-drug conjugate has the following structure: where n = 1 to 8. In some embodiments, n = 1 to 8. In some embodiments, n is approximately 1, approximately 2, approximately 3, approximately 4, approximately 5, approximately 6, approximately 7, approximately 8. In some embodiments, n is approximately 1-8, 1-7, 1-6, 1-5, 1-4, 1-3, 1-2, 2-8, 2-7, 2-6, 2-5, 2-4, 2-3, 3-8, 3-7, 3-6, 3-5, 3-4, 4-8, 4-7, 4-6, 4-5, 5-8, 5-7, 5-6, 6-8, 6-7, or 7-8. In some embodiments, n is an integral or non-integral multiple of one. Petition 870250084299, dated 09 / 18 / 2025, pp. 340 / 410 70 / 126

[0213] In some embodiments, the anti-HER3 / MUC1 antibody is coupled to the drug via a cleavable ligand, for example, an SPBD ligand or a maleimidocaproyl-valine-citrulline-p-aminobenzyloxycarbonyl (VC) ligand. In some embodiments, the anti-HER3 / MUC1 antibody is coupled to the drug via a non-cleavable ligand, for example, an MCC ligand formed using SMCC or sulfo-SMCC. The selection of an appropriate ligand for a given ADC can be easily made by a skilled person knowledgeable in the art, taking into account relevant factors such as the binding site of the anti-HER3 / MUC1 antibody, any structural constraints of the drug, and the hydrophobicity of the drug (see, for example, the review in Nolting, Chapter 5, “Antibody-Drug Conjugates: Methods in Molecular Biology”, 2013, Ducry (Ed.), Springer).Several specific ligand-toxin combinations have been described and can be used with the anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof described herein to prepare ADCs in certain embodiments. Examples include, but are not limited to, peptide-based ligands clivable with auristatins such as MMAE and MMAF, camptothecins such as SN-38, duocarmycins and PBD dimers; MC-based ligands not clivable with auristatins MMAF and MMAE; acid-labile hydrazone-based ligands with calicheamicins and doxorubicin; disulfide-based ligands with maytansinoids such as DM1 and DM4, and bis-maleimidotrioxyethylene glycol (BMPEO)-based ligands with maytansinoid DM1. Some of these therapeutic agents and ligands are described, for example, in Peters & Brown, (2015) Biosci. Rep. e00225; Dosio et al., (2014) Recent Patents on Anti-Cancer Drug Discovery 9: 35 to 65; US Patent Publication No.US 2015 / 0374847 and US20180193477A1; which are incorporated herein by reference in their entirety.

[0214] Depending on the desired drug and the selected ligand, Petition 870250084299, dated 09 / 18 / 2025, pp. 341 / 410 71 / 126 Those skilled in the art may select a suitable method for coupling them together. For example, some conventional coupling methods, such as amine coupling methods, can be used to form the desired drug-ligand complex that still contains reactive groups for conjugation with anti-HER3 / MUC1 antibodies or antigen-binding fragments thereof via covalent linkage. In some embodiments, a drug-maleimide complex (i.e., maleimide drug-ligand) can be used for the payload-bearing reactive group in the present invention. The most common reactive group capable of binding to the thiol group in the preparation of ADCs is maleimide. In addition, organic iodides and bromides are also frequently used.

[0215] The anti-HER3 / MUC1 ADC can be prepared by one of several routes known in the art, employing organic chemical reactions, conditions and reagents known to those skilled in the art (see, for example, “Bioconjugate Techniques” (GT Hermanson, 2013, Academic Press). For example, conjugation can be achieved by (1) reacting a nucleophilic group or an electrophilic group of an antibody with a bivalent ligand reagent to form the antibody-ligand intermediate Ab-L via a covalent bond, followed by reaction with an activated drug moiety D; or (2) reacting a nucleophilic group or an electrophilic group of a drug moiety with a ligand reagent to form the drug-ligand intermediate DL via a covalent bond, followed by reaction with the nucleophilic group or an electrophilic group of an antibody.Conjugation methods (1) and (2) can be employed with a variety of antibodies, drug fractions and ligands to prepare the anti-HER3 / MUC1 ADCs described in this document. Several ligands, ligand components and prepared toxins are commercially available or can be prepared using standard synthetic organic chemistry techniques. These methods are described, by. Petition 870250084299, dated 09 / 18 / 2025, pp. 342 / 410 72 / 126 example, in “March's Advanced Organic Chemistry” (Smith and March, 2006, Sixth Ed., Wiley); Toki et al., (2002) J. Org. Chem. 67: 1866 to 1872; Frisch et al., (1997) Bioconj. Chem. 7: 180 to 186; “Bioconjugate Techniques” (GT Hermanson, 2013, Academic Press); US20210379193A1 and US20180193477A1, which are incorporated herein by reference in their entirety. In addition, several pre-formed drug-ligands suitable for reaction with an anti-HER3 / MUC1 antibody or selected antigen-binding fragment are also commercially available, for example, toxin-ligands comprising DM1, DM4, MMAE, MMAF or duocarmycin SA are available from Creative BioLabs (Shirley, NY).

[0216] Several specific examples of methods for preparing anti-HER3 / MUC1 ADCs are known in the art and are described in U.S. Patent No. 8,624,003 (pot method), U.S. Patent No. 8,163,888 (one-step method), and U.S. Patent No. 5,208,020 (two-step method), and U.S. 20180193477A1, which are incorporated herein by reference in their entirety. Other methods are known in the art and include those described in “Antibody-Drug Conjugates: Methods in Molecular Biology”, 2013, Ducry (Ed.), Springer.

[0217] Drug loading is represented by the number of drug moieties per antibody in an ADC molecule. For some antibody-drug conjugates, drug loading may be limited by the number of binding sites on the antibody. For example, when the binding site is a cysteine ​​thiol, as in certain exemplary embodiments described in this document, the drug loading may range from 0 to 8 drug moieties per antibody. In certain embodiments, higher drug loading, e.g., p > 5, may cause aggregation, insolubility, toxicity, or loss of cell permeability of certain antibody-drug conjugates. In certain embodiments, the average drug loading for an anti-HER3 / MUC1 antibody-drug conjugate in Petition 870250084299, dated 09 / 18 / 2025, pp. 343 / 410 73 / 126 range from 1 to about 8; from about 2 to about 6; or from about 3 to about 5. In fact, it has been shown that for certain antibody-drug conjugates, the optimal ratio of drug fractions to antibody can be around 4. In some embodiments, the DAR for an anti-HER3 / MUC1 ADC composition is about or at least 1, 2, 3, 4, 5, 6, 7, or 8. In some embodiments, the average DAR in the anti-HER3 / MUC1 ADC composition is about 1 to about 2, about 2 to about 3, about 3 to about 4, about 3 to about 5, about 4 to about 5, about 5 to about 6, about 6 to about 7, or about 7 to about 8.

[0218] In some embodiments, antiHER3 / MUC1 antibody variants are provided with a carbohydrate structure that lacks fucose attached (directly or indirectly) to an Fc region. For example, the amount of fucose in such an antibody may be 1% to 80%, 1% to 65%, 5% to 65%, or 20% to 40%. The amount of fucose is determined by calculating the average amount of fucose in the sugar chain at Asn297, relative to the sum of all glycostructures attached to Asn297 (e.g., complex, hybrid, and high-mannose structures), as measured by MALDI-TOF mass spectrometry, as described in document no. WO 2008 / 077546, for example.Asn297 refers to the asparagine residue located at approximately position 297 in the Fc region (EU numbering of Fc region residues; or position 314 in Kabat numbering); however, Asn297 may also be located approximately ± 3 amino acids upstream or downstream of position 297, i.e., between positions 294 and 300, due to small sequence variations in antibodies. Such fucosylation variants may have enhanced ADCC function. In some embodiments, to reduce glycan heterogeneity, the Fc region of the anti-HER3 / MUC1 antibody may be further manipulated to replace asparagine at position 297 with alanine (N297A).

[0219] In some modalities, to facilitate efficiency of Petition 870250084299, dated 09 / 18 / 2025, pp. 344 / 410 74 / 126 production preventing Fab arm exchange, the Fc region of antiHER3 / MUC1 antibodies or antigen-binding fragments thereof was further manipulated to replace serine at position 228 (EU numbering) of IgG4 with proline (S228P). A detailed description of the S228 mutation is described, for example, in Silva et al. “The S228P mutation prevents in vivo and in vitro IgG4 Fab-arm exchange as demonstrated using a combination of novel quantitative immunoassays and physiological matrix preparation”. Journal of Biological Chemistry 290(9) (2015): 5462-5469, which is incorporated by reference in its entirety.

[0220] In some embodiments, the methods described herein are designed to produce a bispecific anti-HER3 / MUC1 antibody. Bispecific anti-HER3 / MUC1 antibodies can be produced by genetically modifying the interface between a pair of antibody molecules to maximize the percentage of heterodimers that are recovered from recombinant cell culture. For example, the interface may contain at least a portion of the CH3 domain of an antibody constant domain. In this method, one or more small amino acid side chains of the interface of the first antibody molecule are replaced with larger side chains (e.g., tyrosine or tryptophan). Compensatory “cavities” of identical or similar size to the large side chain(s) are created at the interface of the second antibody molecule by replacing large amino acid side chains with smaller chains (e.g., alanine or threonine).This provides a mechanism for increasing the yield of heterodimer relative to other undesirable end products, such as homodimers. This method is described, for example, in document no. WO 96 / 27011, which is incorporated by reference in its entirety.

[0221] In some embodiments, bulge-in-hole (KIH) technology can be used, which involves the production of CH3 domains. Petition 870250084299, dated 09 / 18 / 2025, pages 345 / 410 75 / 126 to create a “bulge” or a “hole” in each heavy chain to promote heterodimerization. The KIH technique is described, for example, in Xu, Yiren, et al. “Production of bispecific antibodies in ‘knobs-into-holes’ using a cell-free expression system”. MAbs. Vol. 7. No. 1. Taylor & Francis, 2015, which is incorporated by reference in its entirety. In some embodiments, one heavy chain has a T366W and / or S354C substitution (bulge) (EU numbering) and the other heavy chain has a Y349C, T366S, L368A and / or Y407V substitution (hole) (EU numbering). In some embodiments, one heavy chain has one or more of the following Y349C and T366W substitutions (EU numbering). The other heavy chain may have one or more of the following substitutions: E356C, T366S, L368A, and Y407V (EU numbering). Additionally, a substitution (-ppcpScp-->-ppcpPcp-) may also be introduced into the hinge regions of both substituted IgGs. Recombinant vectors

[0222] The present invention also provides recombinant vectors (e.g., expression vectors) that include an isolated polynucleotide disclosed herein (e.g., a polynucleotide encoding a polypeptide disclosed herein), host cells into which the recombinant vectors are introduced (i.e., so that the host cells contain the polynucleotide and / or a vector comprising the polynucleotide), and the production of anti-HER3 / MUC1 antibody polypeptides or fragments thereof by recombinant techniques.

[0223] As used in this document, a “vector” is any construct capable of delivering one or more polynucleotides of interest to a host cell when the vector is introduced into the host cell. An “expression vector” is capable of delivering and expressing one or more polynucleotides of interest as an encoded polypeptide in a host cell in which Petition 870250084299, dated 09 / 18 / 2025, pp. 346 / 410 76 / 126 the expression vector was introduced. Thus, in an expression vector, the polynucleotide of interest is positioned for expression in the vector by being operationally linked to regulatory elements, such as a promoter, enhancer and / or a poly-A tail, within the vector or in the host cell genome at or near or flanking the integration site of the polynucleotide of interest, so that the polynucleotide of interest is translated in the host cell into which the expression vector was introduced.

[0224] A vector can be introduced into the host cell by methods known in the art, for example, electroporation, chemical transfection (e.g., DEAE-dextran), transformation, transfection and infection and / or transduction (e.g., with recombinant virus). Thus, non-limiting examples of vectors include viral vectors (which can be used to generate recombinant virus), naked DNA or RNA, plasmids, cosmids, phage vectors, and DNA or RNA expression vectors associated with cationic condensation agents.

[0225] In some implementations, a polynucleotide disclosed in this document (e.g., a polynucleotide encoding a polypeptide disclosed in this document) is introduced using a viral expression system (e.g., vaccinia virus or other poxvirus, retrovirus, or adenovirus), which may involve the use of a non-pathogenic (deficient) virus competent for replication or may use a virus deficient for replication. In the latter case, viral propagation will generally occur only in complementation virus packaging cells. Suitable systems are disclosed, for example, in Fisher-Hoch et al., 1989, Proc. Natl. Acad. Sci. USA 86: 317 to 321; Flexner et al., 1989, Ann. NY Acad Sci. 569: 86 to 103; Flexner et al., 1990, Vaccine, 8: 17 to 21; US Patents Nos. 4,603,112, 4,769,330 and 5,017,487; WO 89 / 01973; US Patent No. 4,777,127; GB 2,200,651; EP 0,345,242; WO 91 / 02805; Berkner Biotechniques, 6: 616 to 627, 1988; Rosenfeld Petition 870250084299, dated 09 / 18 / 2025, pp. 347 / 410 77 / 126 et al., 1991, Science, 252: 431-434; Kolls et al., 1994, Proc. Natl. Acad. Sci. USA, 91: 215-219; Kass-Eisler et al., 1993, Proc. Natl. Acad. Sci. USA, 90: 11498-11502; Guzman et al., 1993, Circulation, 88: 2838-2848; and Guzman et al., 1993, Cir. Res., 73: 1202-1207. Techniques for incorporating DNA into such expression systems are well known to people of ordinary technical skill. DNA can also be “naked,” as described, for example, in Ulmer et al., 1993, Science, 259: 1745–1749, and Cohen, 1993, Science, 259: 1691–1692. The uptake of naked DNA can be increased by coating biodegradable microspheres with DNA that are efficiently transported into cells.

[0226] For expression, the DNA insert comprising a polynucleotide encoding polypeptide disclosed in this document can be operationally ligated to an appropriate promoter (e.g., a heterologous promoter), such as the lambda phage PL promoter, the E. coli lac, trp, and tac promoters, the SV40 early and late promoters, and retroviral LTR promoters, to name a few. Other suitable promoters are known to those skilled in the art. The expression constructs may additionally contain transcription initiation and termination sites and, in the transcript region, a ribosome binding site for translation. The coding portion of the mature transcripts expressed by the constructs may include a translation initiation codon at the beginning and a termination codon (UAA, UGA, or UAG) appropriately positioned at the end of the polypeptide to be translated.

[0227] As indicated, expression vectors may include at least one selection marker. Such markers include dihydrofolate reductase or neomycin resistance for eukaryotic cell culture and tetracycline or ampicillin resistance genes for culture in E. coli and other bacteria. Representative examples of suitable hosts include, but are not limited to, bacterial cells such as E. coli, Streptomyces and Petition 870250084299, dated 09 / 18 / 2025, pages 348 / 410 78 / 126 Salmonella typhimurium; fungal cells, such as yeast cells; insect cells, such as Drosophila S2 and Spodoptera Sf9 cells; animal cells, such as CHO, COS, Bowes melanoma, and HK 293 cells; and plant cells. Appropriate culture media and conditions for the host cells described herein are known in the art.

[0228] Non-limiting vectors for use in bacteria include pQE70, pQE60, and pQE-9, available from Qiagen; pBS vectors, Phagescript vectors, Bluescript vectors, pNH8A, pNH16a, pNH18A, pNH46A, available from Stratagene; and ptrc99a, pKK223-3, pKK233-3, pDR540, pRIT5 available from Pharmacia. Non-limiting eukaryotic vectors include pWLNEO, pSV2CAT, pOG44, pXT1, and pSG available from Stratagene; and pSVK3, pBPV, pMSG, and pSVL available from Pharmacia. Other suitable vectors will be readily apparent to the knowledgeable person.

[0229] Suitable non-limiting bacterial promoters for use include the E. coli lacI and lacZ promoters, the T3 and T7 promoters, the gpt promoter, the lambda PR and PL promoters, and the trp promoter. Suitable eukaryotic promoters include the immediate early promoter of CMV, the thymidine kinase promoter of HSV, the early and late promoters of SV40, the promoters of retroviral LTRs, such as those of Rous sarcoma virus (RSV), and metallothionein promoters, such as the mouse metallothionein-I promoter.

[0230] In the yeast Saccharomyces cerevisiae, several vectors containing constitutive or inducible promoters, such as factor alpha, alcohol oxidase, and PGH, can be used. For reviews, see Ausubel et al. (1989) “Current Protocols in Molecular Biology”, John Wiley & Sons, New York, N.Y., and Grant et al., Methods Enzymol., 153: 516 to 544 (1997).

[0231] The introduction of the construct into the host cell can be carried out by transfection with calcium phosphate, DEAE-mediated transfection Petition 870250084299, dated 09 / 18 / 2025, pp. 349 / 410 79 / 126 dextran, cationic lipid-mediated transfection, electroporation, transduction, infection, or other methods. Such methods are described in various standard laboratory manuals, such as Davis et al., “Basic Methods in Molecular Biology” (1986), which is incorporated herein by reference in its entirety.

[0232] The transcription of DNA encoding an anti-HER3 / MUC1 antibody of the present invention by higher eukaryotes can be enhanced by inserting an enhancer sequence into the vector. Enhancers are cis-acting DNA elements, generally of about 10 to 300 bp, that act to increase the transcriptional activity of a promoter in a given type of host cell. Examples of enhancers include the SV40 enhancer, which is located on the late side of the origin of replication at base pairs 100 to 270, the cytomegalovirus early promoter enhancer, the polyoma late-side enhancer, and adenovirus enhancers.

[0233] For secretion of the translated protein into the endoplasmic reticulum lumen, the periplasmic space, or the extracellular environment, appropriate secretion signals can be incorporated into the expressed polypeptide. The signals can be endogenous to the polypeptide or they can be heterologous signals.

[0234] The polypeptide (e.g., an antiHER3 / MUC1 antibody) can be expressed in a modified form, such as a fusion protein (e.g., a fusion with GST) or with a histidine tail, and may include not only secretory signals but also additional heterologous functional regions. For example, a region of additional amino acids, particularly charged amino acids, can be added to the N-terminus of the polypeptide to improve stability and persistence in the host cell, during purification, or during subsequent handling and storage. Furthermore, peptide moieties can be added to Petition 870250084299, dated 09 / 18 / 2025, pages 350 / 410 80 / 126 polypeptide to facilitate purification. Such regions can be removed before the final preparation of the polypeptide. The addition of peptide moieties to polypeptides to induce secretion or excretion, to improve stability and facilitate purification, among other things, are familiar and routine techniques in the art.

[0235] The invention also provides a nucleic acid sequence that is at least 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% identical to any nucleotide sequence as described in this document and an amino acid sequence that is at least 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% identical to any amino acid sequence as described in this document.

[0236] The invention also provides a nucleic acid sequence having a homology of at least 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% with any nucleotide sequence as described herein and an amino acid sequence having a homology of at least 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% with any amino acid sequence as described in this document.

[0237] In some embodiments, the invention relates to nucleotide sequences encoding any peptides that are described herein or any amino acid sequences that are encoded by any nucleotide sequences as described herein. Petition 870250084299, dated 09 / 18 / 2025, pp. 351 / 410 81 / 126 document. In some forms, the nucleic acid sequence is less than 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 150, 200, 250, 300, 350, 400, 500, or 600 nucleotides. In some forms, the amino acid sequence is less than 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 250, 300, 350, or 400 amino acid residues.

[0238] In some embodiments, the amino acid sequence (i) comprises an amino acid sequence; or (ii) consists of an amino acid sequence, wherein the amino acid sequence is any of the sequences as described herein.

[0239] In some embodiments, the nucleic acid sequence (i) comprises a nucleic acid sequence; or (ii) consists of a nucleic acid sequence, wherein the nucleic acid sequence is any of the sequences as described herein.

[0240] To determine the percentage of identity of two amino acid sequences, or two nucleic acid sequences, the sequences are aligned for optimal comparison purposes (e.g., gaps may be introduced in one or both of a first and second amino acid or nucleic acid sequence for optimal alignment, and non-homologous sequences may be disregarded for comparison purposes). The amino acid or nucleotide residues at corresponding amino acid or nucleotide positions are then compared. When a position in the first sequence is occupied by the same amino acid or nucleotide residue as the corresponding position in the second sequence, then the molecules are identical at that position (as used in this document, the “identity” of amino acids or nucleic acids is equivalent to the “homology” of amino acids or nucleic acids). The percentage of identity between the two sequences is a function of the number Petition 870250084299, dated 09 / 18 / 2025, pp. 352 / 410 82 / 126 of identical positions are shared by the sequences, taking into account the number of gaps and the length of each gap, which need to be introduced for ideal alignment of the two sequences. For example, sequence comparison and determination of the percentage of identity between two sequences can be performed using a Blossum 62 scoring matrix with a gap penalty of 12, a gap extension penalty of 4, and a reading phase change gap penalty of 5.

[0241] The percentage of sequence homology (e.g., amino acid sequence homology or nucleic acid homology) can also be determined. How to determine the percentage of sequence homology is known in the art. In some embodiments, conserved amino acid residues with similar physicochemical properties (percentage of homology), for example, leucine and isoleucine, can be used to measure sequence similarity. Families of amino acid residues that have similar physicochemical properties have been defined in the art.These families include, for example, amino acids with basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), branched beta side chains (e.g., threonine, valine, isoleucine), and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). The percentage of homology, in many cases, is greater than the percentage of identity.

[0242] The invention provides one or more nucleic acids encoding any of the polypeptides as described herein. In some embodiments, the nucleic acid (e.g., cDNA) Petition 870250084299, dated 09 / 18 / 2025, pp. 353 / 410 83 / 126 includes a polynucleotide encoding a single-chain heavy polypeptide as described herein. In some embodiments, the nucleic acid includes a polynucleotide encoding a single-chain light polypeptide as described herein. In some embodiments, the nucleic acid includes a polynucleotide encoding an scFv polypeptide as described herein.

[0243] In some embodiments, the vector may have two of the nucleic acids as described herein, wherein the vector encodes the VL region and the VH region which together bind to HER3. In some embodiments, a pair of vectors is provided, wherein each vector comprises one of the nucleic acids as described herein, wherein together the pair of vectors encodes the VL region and the VH region which together bind to HER3.

[0244] In some embodiments, the vector includes two of the nucleic acids as described herein, wherein the vector encodes the VL region and the VH region which together bind to MUC1. In some embodiments, a pair of vectors is provided, wherein each vector comprises one of the nucleic acids as described herein, wherein together the pair of vectors encodes the VL region and the VH region which together bind to MUC1. Treatment methods

[0245] The methods described in this document include methods for the treatment of cancer-associated disorders. Generally, the methods involve administering a therapeutically effective amount of anti-HER3 / MUC1 antibodies or anti-HER3 / MUC1 antibody-drug conjugates, as described in this document, to a subject in need of, or who has been determined to be in need of, such treatment.

[0246] As used in this context, “treat” means to improve at least one symptom of the disorder associated with cancer. Cancer often results in death; thus, treatment may result in an expectation of Petition 870250084299, dated 09 / 18 / 2025, pp. 354 / 410 84 / 126 increased life expectancy (e.g., by at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months or at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 years). Administration of a therapeutically effective amount of an agent described herein for the treatment of a cancer-related condition will result in a reduced number of cancer cells and / or relieved symptoms.

[0247] As used in this document, the term “cancer” refers to cells that have the capacity for autonomous growth, that is, an abnormal state or condition characterized by rapidly proliferating cell growth. The term is intended to include all types of cancerous growths or oncogenic processes, metastatic tissues or cells, malignantly transformed tissues or organs, regardless of histopathological type or stage of invasiveness. The term “tumor,” as used in this document, refers to cancerous cells, for example, a mass of cancerous cells.Cancers that can be treated or diagnosed using the methods described herein include malignancies of various organ systems, such as those affecting the lung, breast, thyroid, lymphatic system, gastrointestinal tract, and genitourinary tract, as well as adenocarcinomas, which include malignancies such as most colon cancers, renal cell carcinoma, prostate cancer and / or testicular tumors, non-small cell lung carcinoma, small bowel cancer, and esophageal cancer. In some embodiments, the agents described herein are designed for the treatment or diagnosis of a carcinoma in a subject.The term "carcinoma" is recognized in the technique and refers to malignancies of epithelial or endocrine tissues, including carcinomas of the respiratory system, carcinomas of the gastrointestinal system, carcinomas of the genitourinary system, testicular carcinomas, breast carcinomas, prostate carcinomas, carcinomas of the endocrine system, and melanomas. In some modalities, the cancer is renal carcinoma or melanoma. Carcinomas. Petition 870250084299, dated 09 / 18 / 2025, pages 355 / 410 Examples of 85 / 126 include those that form from tissue of the cervix, lung, prostate, breast, head and neck, colon, and ovary. The term also includes carcinosarcomas, for example, which include malignant tumors composed of carcinomatous and sarcomatous tissues. An “adenocarcinoma” refers to a carcinoma derived from glandular tissue or in which the tumor cells form recognizable glandular structures. The term “sarcoma” is recognized in the art and refers to malignant tumors of mesenchymal origin.

[0248] In some forms, the cancer is chemotherapy-resistant.

[0249] In one aspect, the invention also provides methods for treating cancer in a subject, methods for reducing the rate of increase in tumor volume in a subject over time, methods for reducing the risk of developing metastasis, or methods for reducing the risk of developing further metastasis in a subject. In some embodiments, the treatment may stop, slow, delay, or inhibit the progression of cancer. In some embodiments, the treatment may result in a reduction in the number, severity, and / or duration of one or more cancer symptoms in a subject.

[0250] In one aspect, the invention provides methods that include administering a therapeutically effective amount of anti-HER3 / MUC1 antibodies or anti-HER3 / MUC1 antibody-drug conjugates disclosed herein to a subject in need thereof, for example, a subject who has, or has been identified or diagnosed as having, cancer, for example, solid tumor, lung cancer (e.g., non-small cell lung cancer, lung adenocarcinoma or lung carcinoma), gastric cancer (e.g., gastric carcinoma), skin cancer (e.g., skin carcinoma), colorectal cancer, breast cancer, cancer of Petition 870250084299, dated 09 / 18 / 2025, pp. 356 / 410 86 / 126 head and neck cancer, ovarian cancer, prostate cancer, thyroid cancer, pancreatic cancer, CNS cancer, liver cancer, nasopharyngeal cancer, brain cancer, colon cancer, bladder cancer, oral squamous cell carcinoma, cervical cancer, or esophageal cancer. In some modalities, the cancer is esophageal cancer, colorectal cancer, gastric cancer, breast cancer, endometrial carcinoma, lung cancer, melanoma, ovarian cancer, bladder cancer, stomach cancer, non-Hodgkin lymphoma, head and neck cancer, pancreatic cancer, and cervical cancer.

[0251] As used in this document, the terms “subject” and “patient” are used interchangeably throughout the descriptive report and describe an animal, human or non-human, that is provided with treatment according to the methods of the present invention. Veterinary and non-veterinary applications are contemplated by the present invention. Human patients may be adult humans or juvenile humans (e.g., humans under the age of 18 years). In addition to humans, patients include, but are not limited to, mice, rats, hamsters, guinea pigs, rabbits, ferrets, cats, dogs, and primates. These include, for example, non-human primates (e.g., monkeys, chimpanzees, gorillas and the like), rodents (e.g., rats, mice, gerbils, hamsters, ferrets, rabbits), lagomorphs, swine (e.g., pig, miniature pig), equines, canines, felines, bovines and other domestic animals, farm animals and zoo animals.

[0252] In some embodiments, the compositions and methods disclosed herein may be used for the treatment of patients at risk of cancer. Patients with cancer may be identified by various methods known in the art.

[0253] As used in this document, an “effective amount” means an amount or dosage sufficient to effect Petition 870250084299, dated 09 / 18 / 2025, pp. 357 / 410 87 / 126 beneficial or desired results, including stopping, slowing, delaying or inhibiting the progression of a disease, for example, cancer. An effective amount will vary depending, for example, on the age and body weight of a subject to whom the anti-HER3 / MUC1 antibody, anti-HER3 / MUC1 antigen-binding fragment, anti-HER3 / MUC1 antibody-drug conjugates, polynucleotide encoding anti-HER3 / MUC1 antibody, vector comprising the polynucleotide and / or compositions thereof are to be administered, the severity of the symptoms and the route of administration, and thus administration can be determined individually.

[0254] An effective amount may be administered in one or more administrations. For example, an effective amount of an anti-HER3 / MUC1 antibody, an anti-HER3 / MUC1 antigen-binding fragment, or an anti-HER3 / MUC1 antibody-drug conjugate is an amount sufficient to improve, stop, stabilize, reverse, inhibit, delay, and / or retard the progression of an autoimmune disease or cancer in a patient, or is an amount sufficient to improve, stop, stabilize, reverse, retard, and / or retard the proliferation of a cell (e.g., a biopsied cell, any of the cancer cells described herein, or a cell line (e.g., a cancer cell line) in vitro.As understood in the art, an effective amount of an anti-HER3 / MUC1 antibody, anti-HER3 / MUC1 antigen-binding fragment, or anti-HER3 / MUC1 antibody-drug conjugate may vary depending, among other things, on the patient's history, as well as other factors such as the type (and / or dosage) of the agent used.

[0255] Effective amounts and regimens for administration of anti-HER3 / MUC1 antibodies, anti-HER3 / MUC1 antigen-binding fragments thereof, polynucleotides encoding anti-HER3 / MUC1 antibodies, anti-HER3 / MUC1 antibody-drug conjugates and / or Petition 870250084299, dated 09 / 18 / 2025, pp. 358 / 410 88 / 126 compositions disclosed herein can be determined empirically, and performing such determinations is within the skill of the art. Those skilled in the art will understand that the dosage to be administered will vary depending, for example, on the mammal receiving the anti-HER3 / MUC1 antibodies, anti-HER3 / MUC1 antigen-binding fragments thereof, anti-HER3 / MUC1 antibody-encoding polynucleotides, anti-HER3 / MUC1 antibody-drug conjugates and / or compositions disclosed herein, the route of administration, the particular type of agent or compositions disclosed herein used, and other drugs administered to the mammal.

[0256] A typical daily dosage of an effective amount of an anti-HER3 / MUC1 antibody or anti-HER3 / MUC1 ADC is 0.01 mg / kg to 100 mg / kg. In some embodiments, the dosage may be less than 100 mg / kg, 30 mg / kg, 20 mg / kg, 10 mg / kg, 9 mg / kg, 8 mg / kg, 7 mg / kg, 6 mg / kg, 5 mg / kg, 4 mg / kg, 3 mg / kg, 2 mg / kg, 1 mg / kg, 0.5 mg / kg or 0.1 mg / kg. In some formulations, the dosage may exceed 10 mg / kg, 9 mg / kg, 8 mg / kg, 7 mg / kg, 6 mg / kg, 5 mg / kg, 4 mg / kg, 3 mg / kg, 2 mg / kg, 1 mg / kg, 0.5 mg / kg, 0.1 mg / kg, 0.05 mg / kg, or 0.01 mg / kg. In some forms, the dosage is approximately or at least 10 mg / kg, 9 mg / kg, 8 mg / kg, 7 mg / kg, 6 mg / kg, 5 mg / kg, 4 mg / kg, 3 mg / kg, 2 mg / kg, 1 mg / kg, 0.9 mg / kg, 0.8 mg / kg, 0.7 mg / kg, 0.6 mg / kg, 0.5 mg / kg, 0.4 mg / kg, 0.3 mg / kg, 0.2 mg / kg, or 0.1 mg / kg.

[0257] In any of the methods described in this document, at least one anti-HER3 / MUC1 antibody, the anti-HER3 / MUC1 antigen-binding fragment thereof, anti-HER3 / MUC1 antibody-drug conjugates or pharmaceutical composition (for example, comprising any of the anti-HER3 / MUC1 antibodies, anti-HER3 / MUC1 antigen-binding antibody fragments or anti-HER3 / MUC1 ADCs) and, optionally, at least one additional therapeutic agent may be Petition 870250084299, dated 09 / 18 / 2025, pp. 359 / 410 89 / 126 administered to the subject (e.g., once a week, twice a week, three times a week, four times a week, once a day, twice a day, or three times a day).

[0258] In some embodiments, one or more additional therapeutic agents may be administered to the subject before or after the administration of at least one anti-HER3 / MUC1 antibody, anti-HER3 / MUC1 antigen-binding antibody fragment, anti-HER3 / MUC1 antibody-drug conjugate or pharmaceutical composition (for example, comprising any of the anti-HER3 / MUC1 antibodies, anti-HER3 / MUC1 antigen-binding antibody fragments or anti-HER3 / MUC1 ADCs). In some embodiments, one or more additional therapeutic agents and at least one anti-HER3 / MUC1 antibody, anti-HER3 / MUC1 antigen-binding antibody fragment, or anti-HER3 / MUC1 antibody-drug conjugate are administered to the subject so that there is an overlap in the bioactive period of one or more additional therapeutic agents and at least one anti-HER3 / MUC1 antibody, anti-HER3 / MUC1 antigen-binding fragment, or anti-HER3 / MUC1 ADC in the subject.

[0259] In some embodiments, the subject may be administered at least one anti-HER3 / MUC1 antibody, anti-HER3 / MUC1 antigen-binding antibody fragment, anti-HER3 / MUC1 antibody-drug conjugate or pharmaceutical composition (e.g., comprising any of the anti-HER3 / MUC1 antibodies, anti-HER3 / MUC1 antigen-binding antibody fragments or anti-HER3 / MUC1 ADCs) over a prolonged period of time (e.g., over a period of at least 1 week, 2 weeks, 3 weeks, 1 month, 2 months, 3 months, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months, 12 months, 1 year, 2 years, 3 years, 4 years or 5 years). A specialist medical professional may determine the length of the treatment period using any of the methods described. Petition 870250084299, dated 09 / 18 / 2025, pages 360 / 410 90 / 126 in this document for the diagnosis or monitoring of treatment efficacy (e.g., observation of at least one symptom of cancer). As described in this document, a qualified medical professional may also alter the identity and number (e.g., increase or decrease) of anti-HER3 / MUC1 antibodies or anti-HER3 / MUC1 antigen-binding antibody fragments, anti-HER3 / MUC1 antibody-drug conjugates (and / or one or more additional therapeutic agents) administered to the subject and may also adjust (e.g., increase or decrease) the dosage or frequency of administration of at least one anti-HER3 / MUC1 antibody, anti-HER3 / MUC1 antigen-binding antibody fragment or anti-HER3 / MUC1 ADC (and / or one or more additional therapeutic agents) to the subject based on an assessment of treatment efficacy (e.g., using any of the methods described in this document and known in the art).

[0260] In some embodiments, one or more additional therapeutic agents may be administered to the subject. The additional therapeutic agent may comprise one or more inhibitors selected from the group consisting of a B-Raf inhibitor, a HER3 inhibitor, a MEK inhibitor, an ERK inhibitor, a K-Ras inhibitor, a c-Met inhibitor, a MUC1 inhibitor, an anaplastic lymphoma kinase (ALK) inhibitor, a phosphatidylinositol 3-kinase (PI3K) inhibitor, an Akt inhibitor, an mTOR inhibitor, a dual PI3K / mTOR inhibitor, a Bruton's tyrosine kinase (BTK) inhibitor, and an isocitrate dehydrogenase 1 (IDH1) and / or isocitrate dehydrogenase 2 (IDH2) inhibitor. In some modalities, the additional therapeutic agent is an indoleamine 2,3-dioxygenase-1 (IDO1) inhibitor (e.g., epacadostate).

[0261] In some embodiments, the additional therapeutic agent may comprise one or more inhibitors selected from the group consisting of a HER3 inhibitor, an LSD1 inhibitor, an MDM2 inhibitor, Petition 870250084299, dated 09 / 18 / 2025, pp. 361 / 410 91 / 126 a BCL2 inhibitor, a CHK1 inhibitor, an inhibitor of the hedgehog-activated signaling pathway, and an agent that selectively degrades the estrogen receptor.

[0262] In some embodiments, the additional therapeutic agent may comprise one or more therapeutic agents selected from the group consisting of trabectedin, nab-paclitaxel, trebananib, pazopanib, cediranib, palbociclib, everolimus, fluoropyrimidine, IFL, regorafenib, reolisin, alimta, Zykadia, Sutent, temsirolimus, axitinib, everolimus, sorafenib, Votrient, pazopanib, IMA-901, AGS-003, cabozantinib, vinflunine, an Hsp90 inhibitor, Ad-GM-CSF, temazolomide, IL-2, IFNα, vinblastine, Thalomid, dacarbazine, cyclophosphamide, lenalidomide, azacitidine, lenalidomide, bortezomide, amrubicin, carfilzomib, pralatrexate and enzastaurine.

[0263] In some embodiments, the additional therapeutic agent may comprise one or more therapeutic agents selected from the group consisting of an adjuvant, a TLR agonist, tumor necrosis factor (TNF) alpha, IL-1, HMGB1, an IL-10 antagonist, an IL-4 antagonist, an IL-13 antagonist, an IL-17 antagonist, an HVEM antagonist, an ICOS agonist, a CX3CL1 targeted treatment, a CXCL9 targeted treatment, a CXCL10 targeted treatment, a CCL5 targeted treatment, an LFA-1 agonist, an ICAM1 agonist, and a selectin agonist.

[0264] In some modalities, carboplatin, nab-paclitaxel, paclitaxel, cisplatin, pemetrexed, gemcitabine, FOLFOX or FOLFIRI are administered to the subject.

[0265] In some embodiments, the additional therapeutic agent is an anti-PD-1 antibody, an anti-PD-L1 antibody, an anti-PD-L2 antibody, an anti-LAG-3 antibody, an anti-TIGIT antibody, an anti-BTLA antibody, an anti-CTLA4 antibody, an anti-CD40 antibody, an anti-OX40 antibody, a Petition 870250084299, dated 09 / 18 / 2025, pp. 362 / 410 92 / 126 anti-4-1BB antibody, an anti-TIM3 antibody, or an anti-GITR antibody. Pharmaceutical compositions and routes of administration

[0266] Pharmaceutical compositions containing at least one (e.g., one, two, three or four) of the anti-HER3 / MUC1 antibodies (e.g., bispecific antibodies), anti-HER3 / MUC1 antigen-binding fragments or anti-HER3 / MUC1 antibody-drug conjugates described herein are also provided herein. The pharmaceutical compositions may be formulated in any manner known in the art.

[0267] Pharmaceutical compositions are formulated to be compatible with their intended route of administration (e.g., intravenous, intra-arterial, intramuscular, intradermal, subcutaneous, or intraperitoneal). The compositions may include a sterile diluent (e.g., sterile water or saline solution), a fixed oil, polyethylene glycol, glycerin, propylene glycol or other synthetic solvents, antibacterial or antifungal agents such as benzyl alcohol or methylparabens, chlorobutanol, phenol, ascorbic acid, thimerosal and the like, antioxidants such as ascorbic acid or sodium bisulfite, chelating agents such as ethylenediaminetetraacetic acid, buffers such as acetates, citrates or phosphates, and isotonic agents such as sugars (e.g., dextrose), polyalcohols (e.g., mannitol or sorbitol) or salts (e.g., sodium chloride), or any combination thereof.Liposomal suspensions can also be used as pharmaceutically acceptable carriers (see, for example, US Patent No. 4,522,811). The compositions can be formulated and packaged in ampoules, disposable syringes, or multi-dose vials. When necessary (as in, for example, injectable formulations), adequate fluidity can be maintained, for example, by the use of a coating, such as lecithin, or a surfactant. The absorption of the anti-HER3 / MUC1 antibody, binding fragment to... Petition 870250084299, dated 09 / 18 / 2025, pp. 363 / 410 93 / 126 anti-HER3 / MUC1 antigen or anti-HER3 / MUC1 ADC may be prolonged by including an agent that retards absorption (e.g., aluminum monostearate and gelatin). Alternatively, controlled release may be achieved by microencapsulated implants and delivery systems, which may include biodegradable biocompatible polymers (e.g., ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid; Alza Corporation and Nova Pharmaceutical, Inc.).

[0268] Compositions containing one or more of any of the anti-HER3 / MUC1 antibodies, anti-HER3 / MUC1 antigen-binding fragments, anti-HER3 / MUC1 antibody-drug conjugates described herein may be formulated for parenteral administration (e.g., intravenous, intra-arterial, intramuscular, intradermal, subcutaneous or intraperitoneal) in unit dose form (i.e., physically discrete units containing a predetermined amount of active compound for ease of administration and uniformity of dosage).

[0269] The toxicity and therapeutic efficacy of the compositions can be determined by standard pharmaceutical procedures in cell cultures or experimental animals (e.g., monkeys). The LD50 (the lethal dose for 50% of the population) and the ED50 (the therapeutically effective dose in 50% of the population) can be determined, the therapeutic index being the ratio of LD50:ED50. Agents exhibiting high therapeutic indices are preferred. When an agent exhibits an undesirable side effect, care should be taken to minimize potential harm (i.e., reduce unwanted side effects). Toxicity and therapeutic efficacy can be determined by other standard pharmaceutical procedures.

[0270] Data obtained from cell culture assays and animal studies can be used in formulating an appropriate dosage of Petition 870250084299, dated 09 / 18 / 2025, pp. 364 / 410 94 / 126 any given agent for use in a subject (e.g., a human being). A therapeutically effective amount of anti-HER3 / MUC1 antibodies, an anti-HER3 / MUC1 antigen-binding fragment thereof, or an anti-HER3 / MUC1 ADC will be an amount that treats the disease in a subject (e.g., kills cancer cells) in a subject (e.g., a human subject identified as having cancer), or a subject identified as being at risk of developing the disease (e.g., a subject who previously developed cancer but has now been cured), decreases the severity, frequency, and / or duration of one or more symptoms of a disease in a subject (e.g., a human being).The efficacy and dosage of any of the anti-HER3 / MUC1 antibodies, the anti-HER3 / MUC1 antigen-binding fragment thereof, or the anti-HER3 / MUC1 ADC described herein may be determined by a healthcare professional or veterinary professional using methods known in the art, as well as by observing one or more disease symptoms in a subject (e.g., a human). Certain factors may influence the dosage and time required to effectively treat a subject (e.g., the severity of the disease or disorder, previous treatments, the subject's general health and / or age, and the presence of other diseases).

[0271] Exemplary doses include milligram or microgram amounts of any of the anti-HER3 / MUC1 antibodies, the anti-HER3 / MUC1 antigen-binding fragments thereof, or the anti-HER3 / MUC1 ADCs described in this document per kilogram of subject weight (e.g., about 1 μg / kg to about 500 mg / kg; about 100 μg / kg to about 500 mg / kg; about 100 μg / kg to about 50 mg / kg; about 10 μg / kg to about 5 mg / kg; Although these doses cover a wide range, a person of ordinary skill in the art will understand that agents Petition 870250084299, dated 09 / 18 / 2025, pp. 365 / 410 95 / 126 Therapeutic agents vary in their potency, and effective amounts can be determined by methods known in the art. Typically, relatively low doses are administered first, and the attending healthcare professional or veterinarian (in the case of therapeutic application) or an investigator (when still working in the development phase) may subsequently and gradually increase the dose until an adequate response is obtained. Furthermore, it is understood that the specific dose level for any particular subject will depend on a variety of factors, including the activity of the specific compound employed, the subject's age, body weight, general health, sex and diet, time of administration, route of administration, excretion rate, and half-life of the therapeutic agent in vivo.

[0272] Pharmaceutical compositions may be included in a container, packaging or dispenser together with instructions for administration. The invention also provides methods for manufacturing anti-HER3 / MUC1 antibodies, the anti-HER3 / MUC1 antigen-binding fragment thereof or the anti-HER3 / MUC1 ADC for various uses as described herein. Examples

[0273] The invention is further described in the following examples, which do not limit the scope of the invention described in the claims. Example 1. Preparation of bispecific anti-HER3 / MUC1 antibodies

[0274] Bispecific antigen-binding molecules targeting HER3 and MUC1 are provided in this document. These antigen-binding molecules are referred to as the anti-HER3 / MUC1 bispecific antibody below.

[0275] Anti-HER3 antibodies (1B2, VH SEQ ID NO: 26, VL SEQ ID NO: 25; 3E1, VH SEQ ID NO: 27, VL SEQ ID NO: 25; and 3G6, VH SEQ ID NO: 45, VL SEQ ID NO: 25) and anti-MUC1 antibody (10D1, VH SEQ ID NO: 28, VL Petition 870250084299, dated 09 / 18 / 2025, pp. 366 / 410 96 / 126 (SEQ ID NO: 25) can be paired to form bispecific antibodies. Vectors encoding both the light and heavy chains of the antibodies were constructed. CHO-S cells were co-transfected with three vectors, including a first vector encoding the heavy chain of an anti-HER3 antibody, a second vector encoding the heavy chain of an anti-MUC1 antibody, and a third vector encoding the common light chain. After 14 days of culture, the cell supernatant was collected and purified by protein A affinity chromatography. Exemplary bispecific antibodies obtained include 3E1-10D1, 3G6-10D1, and 1B2-10D1.

[0276] To verify the binding affinity of bispecific antibodies, bispecific control anti-HER3 antibody and bispecific control anti-MUC1 antibody were also generated, wherein one arm of the bispecific control antibody recognizes HER3 or MUC1 and the other arm recognizes CD28. Similar methods were used to generate these bispecific control antibodies, for example, obtaining VH sequences by immunizing RenLite™ mice. Exemplary bispecific control antibodies are named 3E1-CD28, 3G6-CD28, 1B2-CD28 and 10D1CD28.

[0277] Several methods can be used to reduce the chance of mismatch between the two heavy chains. For example, bulge-hole mutations have been introduced into the Fc regions of the anti-HER3 arm heavy chain, the anti-MUC1 arm heavy chain, and the anti-CD28 arm heavy chain. For example, in 3E1-10D1, the 3E1 heavy chain constant region includes bulge mutations and the 10D1 heavy chain constant region includes hole mutations. In 3E1-CD28, the 3E1 heavy chain constant region includes bulge mutations and the CD28 heavy chain constant region includes hole mutations.

[0278] The sequences of the constant light chain region, of the region Petition 870250084299, dated 09 / 18 / 2025, pp. 367 / 410 97 / 126 heavy chain constant with bulge mutations and the heavy chain constant region with hole mutations are shown in SEQ ID NO: 29, SEQ ID NO: 30 and SEQ ID NO: 31, respectively. Example 2. Binding activity between bispecific anti-HER3 / MUC1 antibody species

[0279] CHO-hHER3 cells, CHO-fasHER3 cells, H293F-hMUC1 cells, CHO-fasMUC1 cells, NUGC-4 cells (Cobioer, Cat. No.: CBP60493) or HCC827 cells (ATCC, Cat. No.: CRL-2868) were transferred to a 96-well plate at a density of 5 χ 104 cells / well, respectively. The bispecific anti-HER3 / MUC1 antibody was added to the 96-well plate and incubated at 4 °C for 30 min. Next, the cells were incubated with the secondary antibody anti-hIgG-Fc-Alex Flour 647 (RL1-H) (Jackson ImmunoResearch Laboratories, Inc., Cat. No.: 109-606-170) at 4 °C in the dark for 15 minutes before flow cytometry analysis.

[0280] CHO-hHER3 cells, CHO-fasHER3 cells, and CHO-fasMUC1 cells were obtained by transfecting CHO-S cells with vectors expressing human HER3 (hHER3, SEQ ID NO: 32), macaque (Macaca fascicularis) HER3 (fasHER3, SEQ ID NO: 33), and macaque (Macaca fascicularis) MUC1 (fasMUC1, SEQ ID NO: 35), respectively. H293F-hMUC1 cells were obtained by transfecting H293F cells with a vector expressing human MUC1 (hMUC1, SEQ ID NO: 34, positions 961-1152). RNA sequencing analyses showed that HER3 and MUC1 expression levels in NUGC-4 cells were 155 and 79, respectively, and expression levels in HCC827 cells were 27 and 41, respectively.

[0281] Gatipotuzumab, a glycogen-modified humanized monoclonal antibody that recognizes the tumor-associated epitope of MUC1, is in phase II clinical development at Glycotope for the treatment of recurrent epithelial ovarian, fallopian tube or Petition 870250084299, dated 09 / 18 / 2025, pp. 368 / 410 98 / 126 of primary peritoneal cancer. The variable heavy chain region and the variable light chain region of Gatipotuzumab are shown in SEQ ID NO: 36 and SEQ ID NO: 37, respectively.

[0282] Test results are shown in Table 4 below. 3E1-10D1, 3G6-10D1 and 1B2-10D1 can bind to human HER3, monkey HER3, human MUC1 and monkey MUC1. Table 4 Antibody Protein Percentage of Positive Cells NC Assessment hHER3 0.24% No Binding fasHER3 0.46% No Binding hMUC1 1.42% No Binding fasMUC1 0.15% No Binding HCC827 0.31% No Binding NUGC-4 0.43% No Binding NC+ secondary antibody hHER3 0.31% No Binding fasHER3 0.39% No Binding hMUC1 0.96% No Binding Petition 870250084299, dated 09 / 18 / 2025, pp. 369 / 410 99 / 126 Antibody Protein Percentage of Positive Cells Evaluation fasMUC1 0.20% No Binding HCC827 0.49% No Binding NUGC-4 1.15% No Binding Gatipotuzumab analog hHER3 1.63% No Binding fasHER3 1.94% No Binding hMUC1 22.0% Binding fasMUC1 0.65% No Binding HCC827 96.9% Binding NUGC-4 95.3% Binding 3E1-10D1 hHER3 28.5% Binding fasHER3 39.7% Binding hMUC1 68.2% Binding fasMUC1 36.9% Binding HCC827 98.7% Binding NUGC-4 94.4% Binding 1B2-10D1 hHER3 27.3% Binding fasHER3 34.3% Linkage hMUC1 69.4% Linkage fasMUC1 40.4% Linkage Petition 870250084299, dated 09 / 18 / 2025, pp. 370 / 410 100 / 126 Antibody Protein Percentage of Positive Cells Evaluation HCC827 99.0% NUGC-4 Binding 91.5% 3G6-10D1 Binding hHER3 16.5% fasHER3 Binding 23.0% hMUC1 Binding 72.0% fasMUC1 Binding 40.5% HCC827 Binding 98.6% NUGC-4 Binding 93.0%

[0283] In another experiment, the binding activity of bispecific anti-HER3 / MUC1 antibody to NUGC-4 or NCI-H226 tumor cells was measured by flow cytometry. The secondary antibody used in the experiment was: Alexa Fluor® 647 anti-human IgG Fcy (Jackson ImmunoResearch Laboratories, Inc., Cat. No.: 109-606-170). Human IgG1 was used as an isotype control (ISO). The MFI was determined using serially diluted sample antibodies (maximum concentration: 30 μg / ml, 3-fold dilutions). A fitting curve was obtained using concentration (μg / ml) as the X-axis and MFI as the Y-axis. The results are shown in Figures 14A-14B. Example 3. Internalization of bispecific anti-HER3 / MUC1 antibodies

[0284] Anti-HER3 antibodies, anti-MUC1 antibodies, bispecific anti-HER3 / MUC1 antibodies, bispecific anti-HER3 / CD28 antibodies, or bispecific anti-MUC1 / CD28 antibodies, together with the secondary goat anti-human IgG antibody pHAb-AffiniPure Fab, were added to HCC70 cells (ATCC, Cat. No.: CRL-2315) and incubated for 6 hours. The cells were centrifuged and washed with FACS buffer. IMF was measured using a flow cytometer. Antibody endocytosis rates Petition 870250084299, dated 09 / 18 / 2025, pp. 371 / 410 101 / 126 were calculated. Human IgG1 protein was used for isotype control (ISO). The results are shown in Table 5 below.

[0285] RNA sequencing analyses showed that the expression levels of HER3 and MUC1 in HCC70 cells were 19 and 110, respectively.

[0286] Patritumab is a fully human anti-HER3 monoclonal antibody that was developed at Daiichi Sankyo, and its variable heavy chain region and variable light chain region are shown in SEQ ID NO: 38 and SEQ ID NO: 39, respectively. Table 5 HCC70 IMF Antibodies Population Positive ISO 11727 4.43% NC 10083 2.29% NC+ secondary antibody 11259 4.14% Patritumab Analog 15915 9.21% Gatipotuzumab Analog 94218 74.9% 3E1 13784 6.34% 1B2 14178 6.38% 3G6 14697 8.24% 10D1 54095 96.0% 3E1-CD28 11894 3.76% 1B2-CD28 11869 4.32% 3G6-CD28 11878 4.15% 10D1-CD28 16411 13.6% 3E1-10D1 46923 96.4% 1B2-10D1 45503 95.2% Petition 870250084299, dated 09 / 18 / 2025, pp. 372 / 410 102 / 126 HCC70 IMF Antibodies Positive Population 3G6-10D1 37576 91.6%

[0287] The results showed that the endocytosis rates of the bispecific antibodies 3E1-10D1, 3G6-10D1 and 1B2-10D1 were equal to or greater than those of the corresponding monoclonal antibody 3E1, 3G6, 1B2 or 10D1 in HCC70 cells. Example 4. Binding activity of bispecific antiHER3 / MUC1 antibodies

[0288] The binding activity of bispecific antiHER3 / MUC1 antibodies against human HER3, human MUC1, monkey HER3 and monkey MUC1 was verified by surface plasmon resonance (SPR) using an 8K Biacore™ biosensor (Biacore, Inc., Piscataway NJ) equipped with pre-immobilized protein A sensor chips.

[0289] Specifically, hHER3-His (ACROBiosystems Inc., catalog no.: ER3-H5223), hMUC1(24-1158)-His (SEQ ID NO: 34, position 14-1158), hMUC1(961-1152)-His (SEQ ID NO: 34, position 961-1152), fasHER3-His (Sino Biological, Inc., Cat. no.: 90043-K08H), and fasMUC1-His (SEQ ID NO: 35) were diluted to 200 nM with 1x HBS-EP+ buffer (pH 7.4) and then injected into the 8K Biacore™ biosensor at 10 μm / min for approximately 50 seconds to achieve the desired protein density (e.g., approximately 200 response units (RU)). Purified antibodies at concentrations of 2 μg / ml in 1x HBS-EP+ buffer (pH 7.4) were then injected at 10 μg / min for 50 seconds. Dissociation was monitored for 400 seconds. The chip was regenerated after the last injection of each titration with a glycine solution (pH 1.5) at 30 ml / min for 30 seconds.

[0290] The kinetic association rates (kon) and dissociation rates (koff) were obtained simultaneously by fitting the data Petition 870250084299, dated 09 / 18 / 2025, pp. 373 / 410 103 / 126 globally to a 1:1 Langmuir linkage model (Karlsson, R. Roos, H. Fagerstam, L. Petersson, B., 1994. Methods Enzymology 6. 99 to 110) using the Biacore™ 8K 3.0 evaluation software. Affinities were deduced from the quotient of the kinetic rate constants (KD = koff / kon).

[0291] As a person of ordinary skill in the technique would understand, the same method with appropriate adjustments for parameters (e.g., antibody concentration) was performed for each antibody tested.

[0292] The results of the tested antibodies are summarized in Table 6, which showed that the bispecific anti-HER3 / MUC1 antibodies 3E110D1, 3G6-10D1 and 1B2-10D1 showed good binding affinity to human HER3, monkey HER3, human MUC1 and monkey MUC1. Table 6 Antibody Analyte solution 1 kon (1 / Ms) koff (1 / s) KD (M) 3E1-10D1 fasMUC1-His 1.99E+05 7.76E-04 3.89E-09 hMUC1(24-1158)-His 5.20E+04 1.03E-03 1.98E-08 hMUC1(961-1152)-His 2.11E+05 1.30E-03 6.14E-09 hHER3-His 1.98E+05 3.14E-03 1.58E-08 fasHER3-His 1.15E+05 3.18E-03 2.75E-08 1B2-10D1 fasMUC1-His 1.88E+05 7.26E-04 3.85E-09 hMUC1(24-1158)- His 5.01E+04 9.56E-04 1.91E-08 hMUC1(961-1152)- His 1.97E+05 1.19E-03 6.06E-09 hHER3-His 1.30E+05 2.41E-03 1.85E-08 Petition 870250084299, dated 09 / 18 / 2025, pp. 374 / 410 104 / 126 Antibody Analyte Solution 1 kon (1 / Ms) koff (1 / s) KD (M) fasHER3-His 7.82E+04 2.47E-03 3.16E-08 3G6-10D1 fasMUC1-His 1.94E+05 8.08E-04 4.17E-09 hMUC1(24-1158)- His 4.65E+04 9.94E-04 2.14E-08 hMUC1(961-1152)- His 2.02E+05 1.33E-03 6.60E-09 hHER3-His 1.84E+05 9.33E-03 5.08E-08 fasHER3-His 1.73E+05 1.19E-02 6.90E-08 Example 5. Stability of bispecific anti-HER3 / MUC1 antibodies

[0293] The bispecific anti-HER3 / MUC1 antibodies 3E1-10D1 and 1B2-10D1 were exchanged for buffer at pH 6.0 (3 mg / ml histidine, 80 mg / ml sucrose, and 0.2 mg / ml Tween 80). The antibodies were kept in sealed Eppendorf tubes at 40 ± 2 °C, 60 % ± 5 % RH, and 4 ± 3 °C for 7 days, and their thermal stability was evaluated. Alternatively, the bispecific antibodies were loaded onto a protein A column and eluted with a buffer (0.1 mol / l HAc) at pH 3.5. Half of the antibodies received 2M Tris buffer to immediately bring the pH to 7.5. The remaining half was kept at pH 3.5 for 6 hours, and then the pH was adjusted to 7.5. The diluted antibodies were kept in sealed Eppendorf tubes at pH 3.5 ± 0.1, 25 ± 2 °C (hereinafter referred to as pH 3.5) for 6 hours or 24 hours to test stability at low pH.

[0294] After the above treatments, the following tests were performed: (1) Antibody purity was measured by Size Exclusion High Performance Liquid Chromatography (SEC-HPLC) (indicated as the percentage of the area of ​​the main peak relative to the sum of all peak areas) Petition 870250084299, dated 09 / 18 / 2025, pp. 375 / 410 105 / 126 peak (purity, %)); (2) Antibody hydrophobicity measured by Hydrophobic Interaction Chromatography-High Performance Liquid Chromatography (HIC-HPLC) method (indicated as the retention time of the main peak (HIC, min)); (3) Antibody pI (isoelectric point) and charge variants were measured by Capillary Isoelectric Focusing (cIEF) method (indicated as percentages of the main component, acid component, and alkaline component); (4) Changes in antibody purity by capillary electrophoresis-sodium dodecyl sulfate (CE-SDS) under non-reducing conditions (CE-SDS(NR)) (indicated as the percentage of the main peak area relative to the sum of all peak areas (Purity, %)); (5) Appearance and presence of visible non-soluble particles.

[0295] In SEC-HPLC experiments, antibody samples were diluted to 1 mg / ml with purified water and an Agilent 1290 chromatography system (connected with XBridge Protein BEH SEC column (200 A, Waters Corporation)) was used. The following parameters were used: mobile phase: 0.1 M phosphate buffer (PB) + 10% ACN, pH 7.4; flow rate: 1.8 ml / min; column temperature: 25 °C; detection wavelength: 280 nm, 220 nm; injection volume: 10 μL; sample tray temperature: approximately 8 °C; and run time: 7 minutes.

[0296] In the HIC-HPLC experiments, an Agilent 1260 chromatography system (connected with a ProPac HIC-10 column (4.6 x 250 mm, Thermo Scientific)) was used, and the samples were 10 times diluted using mobile phase A. The following parameters were used: mobile phase A: 0.9 M ammonium sulfate, 0.1 M phosphate buffer (PB), 10% acetonitrile pH 6.5; mobile phase B: 0.1 M phosphate buffer (PB), 10% acetonitrile pH 6.5; flow rate: 0.8 ml / min; gradient: 0 min 100% of A, 2 min 100% of A, 32 min 100% of B, 34 min 100% of B, 35 min 100% of A and 45 min 100% of A; column temperature: 30°C; detection wavelength: 280 nm, 220 nm; volume of Petition 870250084299, dated 09 / 18 / 2025, pp. 376 / 410 106 / 126 injection: 10 μg; sample tray temperature: approximately 10 °C; and run time: 50 minutes.

[0297] In the cIEF experiments, a Maurice cIEF Method Development Kit (Protein Simple, catalog number: PS-MDK01-C) was used for sample preparation. Specifically, 8 μL (30 μg) of the protein sample were mixed with the following reagents in the kit: 1 μL of Maurice cIEF pI Marker 7.05, 1 μL of Maurice cIEF pI Marker 10.10, 35 μL of 1% Methylcellulose Solution, 2 μL of Maurice cIEF 500 mM Arginine, 1.33 μL of Ampholytes (Pharmalyte pH ranges 3-10), 6.66 μL of Ampholytes (Pharmalyte pH ranges 8-10.5), and water (added to make a final volume of 100 μL). In the Maurice analyzer (Protein Simple, Santa Clara, CA), Maurice cIEF Cartridges (PSMC02-C) were used to generate capillary isoelectric focusing spectra of The sample was focused for a total of 10 minutes. The analysis software installed on the instrument was used to integrate the absorbance of the protein focused at 280 nm.

[0298] In CE-SDS(NR) experiments, Maurice (Protein The Simple, Maurice™ microcentrifuge and the Maurice CE-SDS Size Application Kit (Protein Simple, catalog number: PS-MAK02-S) were used. In the CE-SDS(NR), 30 μl of sample buffer, 30 μl of 30 µg antibody sample, 1.5 μl of 25x internal standard, and 3 μl of 250 nM iodoacetamide (SIGMA, catalog number: 16125) were added to a microcentrifuge tube, followed by centrifugation at 3000 rpm for 1 min and heating in a water bath at 70 °C for 10 min. The samples were then cooled to room temperature, followed by centrifugation at 10000 rpm for 3 minutes. The supernatant sample preparations were then transferred to a 96-well plate and tested on the Maurice. The following parameters were used: injection voltage 4.6 kV, injection time 20 s, separation voltage 5.75 kV, and separation time 40 min.

[0299] Detailed results of bispecific anti antibodies Petition 870250084299, dated 09 / 18 / 2025, pp. 377 / 410 107 / 126 HER3 / MUC1 are shown in Table 7. The results showed that 3E110D1 and 1B2-10D1 had good stability, as well as good physical and chemical properties. Table 7 Antibody Treatment Purity of SEC (%) hic (min) CE-SDS (NR) cIEF Purity a (%) LMW (%) HMW (%) Acid Peak (%) Principal Peak (%) Alkaline Peak (%) 3E1-10D1 0d 96.03 3.88 92.8 7.2 0 22.1 72.2 5.7 40 °C 7 d 95.13 3.89 92.3 7.7 0 30.5 59.5 10.0 pH3.5 0h 94.29 3.90 94.0 5.9 0 22.6 70.7 6.7 pH3.5 6h 92.37 3.91 94.5 5.5 0 16.7 76.9 6.4 pH3.5 24h 92.49 3.91 93.4 6.6 0 17.6 76.4 6.0 1B2-10D1 0d 85.63 5.12 85.5 14.5 0 14.9 78.6 6.4 40 °C 7 d 86.48 5.05 85, 14.8 0 36.8 58.2 5.0 pH3.5 0h 88.93 5.16 88.8 11.2 0 25.9 69.5 4.5 pH3.5 24h 88.75 5.16 88.7 11.2 0 26.8 68.4 4.9 Example 6. Preparation of Antibody-Drug Conjugates antiHER3 / MUC1

[0300] Each purified antibody (3E1, 3G6, 1B2, 10D1, 3E1-10D1, 3G6-10D1 or 1B2-10D1) was coupled to MMAE (monomethylauristatin E) or MMAF (monomethylauristatin F) via a maleimidocaproyl-valinecitrulline-p-aminobenzyloxycarbonyl (VC) linker.

[0301] For antibody-drug conjugate names, “ADC” is added directly after the antibody name. For example, if 3E1-10D1 with a constant region of IgG1 is coupled to MMAE, it is named 3E1-10D1-ADC. Antibody-drug conjugates produced by similar methods also included Patritumab-ADC, Gatipotuzumab-ADC, and 1H7ADC. For isotype control, human IgG1 was coupled to MMAE to form ISO-ADC. Petition 870250084299, dated 09 / 18 / 2025, pp. 378 / 410 108 / 126

[0302] 1H7 was a murine monoclonal IgG antibody targeting the extracellular region of the C-terminal subunit of human MUC1, developed by Chungbuk National University and Peptron Co. Ltd. The variable region of the heavy chain and the variable region of the light chain of 1H7 are shown in SEQ ID NO: 40 and SEQ ID NO: 41, respectively.

[0303] HIC-HPLC were used to detect antibody coupling with drug molecules. In the HIC-HPLC experiments, an Agilent 1260 chromatography system (connected with a ProPac™ HIC-10 column (4.6 x 250 mm, Thermo Scientific)) was used, and samples were diluted using mobile phase A at 0.5 mg / ml. The following parameters were used: mobile phase A: 0.9 M ammonium sulfate, 0.1 M phosphate buffer (PB), 10% acetonitrile pH 6.5; mobile phase B: 0.1 M PB, 10% acetonitrile pH 6.5; flow rate: 0.8 ml / min; Gradient: 0 min 100% of A, 2 min 100% of A, 32 min 100% of B, 34 min 100% of B, 35 min 100% of A, and 45 min 100% of A; column temperature: 30 °C; detection wavelength: 280 nm; injection volume: 10 μL; sample tray temperature: approximately 6 °C; and run time: 45 minutes.

[0304] HIC-HPLC detection results show that the drug-to-antibody ratio (DAR) of each ADC is approximately 4. Example 7. Antitumor activity in a NUGC-4 xenograft model

[0305] ADCs were tested for their effect on tumor growth in vivo in a NUGC-4 xenograft model of gastric cancer. HER3 and MUC1 expression levels in NUGC-4 cells were 154.99 and 79.44 respectively, as determined by RNAseq. Specifically, approximately 5 x 106 NUGC-4 cells were injected subcutaneously into B-NDG mice (Biocytogen Pharmaceuticals (Beijing) Co., Ltd., catalog number: B-CM-002). When the tumors in the mice reached a volume of approximately 200 mm3, the mice Petition 870250084299, dated 09 / 18 / 2025, pp. 379 / 410 109 / 126 mice were randomly assigned to different groups based on tumor volumes. Mice were then injected with phosphate-buffered saline (PBS), Patritumab-ADC, 1H7-ADC, 3E1-10D1-ADC, 1B2-10D1-ADC, 3E1-ADC, 1B2-ADC, or 10D1-ADC by intravenous (iv) administration. The frequency of administration was once a week (2 administrations in total). Details are shown in Table 8. Table 8 Group No. of mice Antibodies Dosage Route Frequency Total no. of administrations G1 5 PBS - iv QW 2 G2 5 Patritumab-ADC 3 mg / kg iv QW 2 G3 5 1H7-ADC 3 mg / kg iv QW 2 G4 5 3E1-10D1-ADC 3 mg / kg iv QW 2 G5 5 1B2-10D1-ADC 3 mg / kg iv QW 2 G6 5 3E1-ADC 3 mg / kg iv QW 2 G7 5 1B2-ADC 3 mg / kg iv QW 2 G8 5 10D1-ADC 3 mg / kg iv QW 2

[0306] Tumor volumes were measured twice a week and the body weights of the mice were also recorded. Euthanasia was performed when a mouse's tumor volume reached 3000 mm3.

[0307] The long geometric axis and short geometric axis lengths of the tumor were measured, and the tumor volume was calculated as 0.5 x (long axis) χ (short geometric axis)2. Tumor growth inhibition (TGI) was calculated using the following formula: TGI (%) = [1-(Ti-T0) / (ViV0)]x100. Ti is the mean tumor volume in the treatment group on day i. T0 is the mean tumor volume in the treatment group on day zero. Vi is the mean tumor volume in the control group on day i. V0 is the mean tumor volume in the control group on day zero. The t-test was performed for statistical analysis. P < 0.05 is a threshold to indicate a significant difference. Petition 870250084299, dated 09 / 18 / 2025, pages 380 / 410 110 / 126

[0308] During the experimental period, no significant difference in body weight was observed between each group.

[0309] Table 9 summarizes the results for this experiment, including tumor volumes on the day of pooling (Day 0), 11 days after pooling (Day 11), 21 days after pooling (Day 21), and 32 days after pooling (Day 32); the ICT (%); and the statistical differences (p-value) of body weights and tumor volumes between the treatment and control groups on Day 21. Table 9 Group Tumor volume (mm3) ICT (%) (Day 21) p-value (Day 21) Day 0 Day 11 Day 21 Day 32 Body weight Tumor volume G1 186±10 445±47 902±89 1649±127 NA NA NA G2 185±14 332±50 706±79 1654±112 27.4 1.000 0.135 G3 185±12 314±33 645±146 1376±194 35.9 0.296 0.158 G4 185±15 99±12 160±58 493±105 103.5 0.273 1.10E-04 G5 185±11 66±7 83±34 366±166 114.2 0.120 2.53E-05 G6 186±14 146±20 298±52 1029±179 84.3 0.025 3.68E-04 G7 185±17 93±11 183±38 712±108 100.3 0.412 7.24E-05 G8 185±13 119±22 315±95 779±185 81.9 0.201 0.002

[0310] Tumor volumes in groups treated with ADCs are shown in Figure 1. The treatment groups exhibited different tumor-inhibiting effects. The bispecific anti-HER3 / MUC1 antibody ADCs 3E1-10D1-ADC (G4) and 1B2-10D1-ADC (G5) showed better antitumor activities than the monoclonal ADCs (G6, G7 and G8) and the positive controls (G2 and G3) in the gastric cancer model. Example 8. Antitumor Activity in a xenograft model derived from a patient with gastric cancer.

[0311] ADCs were tested for their effects on Petition 870250084299, dated 09 / 18 / 2025, pp. 381 / 410 111 / 126 In vivo tumor growth in a patient-derived xenograft model of gastric cancer. Immunofluorescence labeling of patient-derived gastric tumor fragments was performed, and images were analyzed using HALO version 3.2. Results showed that HER3-positive cells constituted 60.23% of the total cells and MUC1-positive cells constituted 91.30% of the total cells in human gastric tumor tissues. Specifically, patient-derived gastric tumor fragments (2 mm*2 mmx2 mm) were grafted into the right flank of B-NDG mice. When the tumors in the mice reached a volume of approximately 250–300 mm3, the mice were randomly placed into different groups based on tumor volumes. The mice were then injected with PBS, Patritumab-ADC, 1H7-ADC, Gatipotuzumab-ADC, 3E1-10D1-ADC, or 1B210D1-ADC. Details are shown in Table 10. Table 10 Group No. of mice Antibodies Dosage Route Frequency Total No. of administrations G1 6 PBS - iv QW 2 G2 6 Patritumab-ADC 3 mg / kg iv QW 2 G3 6 1H7-ADC 3 mg / kg iv QW 2 G4 6 Gatipotuzumab-ADC 3 mg / kg iv QW 2 G5 6 3E1-10D1-ADC 3 mg / kg iv QW 2 G6 6 1B2-10D1-ADC 3 mg / kg iv QW 2

[0312] Body weights were measured twice a week. During the experiment, there was no significant difference in body weights between the groups, indicating that the tested ADCs were well tolerated and were not obviously toxic to the mice.

[0313] Tumor volumes of mice in different groups are shown in Figure 2. The treatment groups (G2-G6) showed Petition 870250084299, dated 09 / 18 / 2025, pp. 382 / 410 112 / 126 greater tumor inhibition compared to the PBS group (G1). Furthermore, both 1B2-10D1-ADC (G6) and 3E1-10D1-ADC (G5) showed a higher % ICT on day 20 (50.1% and 69.1%) than the positive controls Patritumab-ADC (G2, 18.9%), 1H7-ADC (G3, 24.8%), and Gatipotuzumab-ADC (G4, 19.2%). Mouse tumor volume and survival rate after day 20 were also continuously monitored. Euthanasia was performed when the mouse tumor volume reached 3000 mm3. At the end of the experiment (Day 34), 1B2-10D1-ADC (G6, % ICT=52.7%) and 3E1-10D1-ADC (G5, % of ICT=37.8%) maintained better antitumor effects compared to the positive controls Patritumab-ADC (G2, % ICT=10.4%), 1H7-ADC (G3, % ICT=28.4%) and Gatipotuzumab-ADC (G4, % ICT=24.9%). Example 9. Antitumor activity in an HCC70 xenograft model

[0314] ADCs were tested for their effects on tumor growth in vivo in an HCC70 cell xenograft model of ductal carcinoma of the breast. Specifically, approximately 1 χ¹⁰⁷ HCC70 cells were injected subcutaneously into B-NDG mice. When the tumors in the mice reached a volume of approximately 200 mm³, the mice were randomly placed into different groups based on tumor volumes. The mice were then injected with PBS, Patritumab-ADC, 1H7-ADC, 3E1-10D1-ADC, 3G6-10D1-ADC, or 1B2-10D1ADC by intravenous (iv) administration. The frequency of administration was once weekly (2 administrations in total). Details are shown in Table 11. Table 11 Group Number of mice Antibodies Dosage Route Frequency Total number of administrations G1 5 PBS - iv QW 2 G2 5 Patritumab-ADC 3 mg / kg iv QW 2 Petition 870250084299, dated 09 / 18 / 2025, pp. 383 / 410 113 / 126 Group Number of mice Antibodies Dosage Route Frequency Total number of administrations G3 5 1H7-ADC 3 mg / kg iv QW 2 G4 5 Gatipotuzumab-ADC 3 mg / kg iv QW 2 G5 5 3E1-10D1-ADC 3 mg / kg iv QW 2 G6 5 3G6-10D1-ADC 3 mg / kg iv QW 2 G7 5 1B2-10D1-ADC 3 mg / kg iv QW 2

[0315] Tumor volumes were measured twice a week and the body weights of the mice were also recorded. Euthanasia was performed when a mouse's tumor volume reached 3000 mm3.

[0316] The weight of mice in different groups increased. On Day 0, the average weight of each group was in the range of 21.6 g-22.0 g. At the end of the experiment (Day 39), the average weight of each group was in the range of 24.3 g-26.6 g. Thus, the average weight of each group was in the range of 112.5%-121.2%. The results showed that the tested ADCs were well tolerated and were not obviously toxic to the mice.

[0317] Table 12 summarizes the results for this experiment, including tumor volumes on the day of clustering (Day 0), 18 days after clustering (Day 18), 28 days after clustering (Day 28), and 39 days after clustering (Day 39); the survival rate of the mice; the ICT (%); and the statistical differences (p-value) of body weights and tumor volumes between the treatment and control groups. Table 12 Group Tumor volume (mm3) ICT (%) (Day 39) p-value (Day 39) Day 0 Day 18 Day 28 Day 39 Body weight Tumor volume G1 204±11 735±79 980±106 1565±131 NA NA NA G2 204±14 657±103 928±173 1409±301 11.5 0.737 0.647 Petition 870250084299, dated 09 / 18 / 2025, pp. 384 / 410 114 / 126 Group Tumor volume (mm3) ICT (%) (Day 39) p-value (Day 39) Day 0 Day 18 Day 28 Day 39 Body weight Tumor volume G3 204±13 164±62 240±116 373±131 87.6 0.001 2.04E-04 G4 204±14 262±62 396±83 646±72 67.5 0.227 2.77E-04 G5 204±10 42±8 36±6 65±11 110.2 0.086 3.18E-06 G6 204±18 177±40 218±72 359±123 88.6 0.122 1.54E-04 G7 204±15 93±37 99±58 199±124 100.4 0.021 6.44E-05

[0318] Tumor sizes in groups treated with ADCs are shown in Figure 3. 3E1-10D1-ADC (G5), 3G6-10D1-ADC (G6) and 1B210D1-ADC (G7) showed better antitumor activities compared to the positive controls Patritumab-ADC (G2), 1H7-ADC (G3) and Gatipotuzumab-ADC (G4) in the ductal carcinoma model of the breast. Example 10. Antitumor Activity in a xenograft model derived from a patient with pancreatic cancer.

[0319] ADCs were tested for their effects on tumor growth in vivo in a patient-derived xenograft model of pancreatic cancer. Immunofluorescence labeling of patient-derived pancreatic tumor fragments was performed, and images were analyzed by HALO version 3.2. Results showed that HER3-positive cells constituted 26.78% of the total cells and MUC1-positive cells constituted 64.77% of the total cells in human pancreatic tumor tissues. Specifically, patient-derived pancreatic tumor fragments (2 mm*2 mm*2 mm) were grafted into the right flank of B-NDG mice. When the tumors in the mice reached a volume of approximately 250–300 mm3, the mice were randomly placed into different groups based on tumor volumes (5 mice per group). The mice were then injected with PBS (G1), Patritumab-ADC (G2), 1H7-ADC (G3), Gatipotuzumab-ADC (G4), 3E1 Petition 870250084299, dated 09 / 18 / 2025, pages 385 / 410 115 / 126 10D1-ADC (G5) or 1B2-10D1-ADC (G6) at 3 mg / kg by intravenous (IV) administration. The frequency of administration was once a week (2 administrations in total).

[0320] Tumor volumes of mice in different groups are shown in Table 13 and Figure 4. Treatment groups (G2-G6) showed significant tumor inhibition compared to PBS groups (G1). In addition, 3E1-10D1-ADC (G5) and 1B2-10D1-ADC (G6) showed a higher % ICT on day 56 compared to positive controls Patritumab-ADC (G2), 1H7-ADC (G3), and Gatipotuzumab-ADC (G4). Bispecific anti-HER3 / MUC1 ADCs showed robust and sustained antitumor effects in the pancreatic cancer model. Table 13 Group Tumor volume (mm3) ICT (%) (Day 56) P value (Day 56) Day 0 Day 21 Day 42 Day 56 G1 255±17 1133±48 2103±112 3119±147 NA NA G2 255±17 635±105 1201±214 1988±280 39.5 0.007 G3 255±31 392±75 973±188 1880±331 43.3 0.009 G4 255±18 547±78 968±194 1489±305 56.9 0.001 G5 255±20 194±14 325±38 704±69 84.3 2.64E-06 G6 255±28 278±29 698±100 1098±167 70.6 1.71E-05 Example 11. Antitumor Activity in a xenograft model derived from PATIENT WITH LUNG CANCER

[0321] ADCs were tested for their effects on tumor growth in vivo in a patient-derived xenograft model of lung cancer. Immunofluorescence labeling of patient-derived lung tumor fragments was performed and the images were analyzed by HALO version 3.2. The results showed that HER3-positive cells constituted 22.84% of the total cells and MUC1-positive cells Petition 870250084299, dated 09 / 18 / 2025, pp. 386 / 410 116 / 126 constituted 82.88% of the total cells in human lung tumor tissues. Specifically, patient-derived lung tumor fragments (2 mm x 2 mm x 2 mm) were grafted onto the right flank of B-NDG mice. When the tumors in the mice reached a volume of approximately 250 to 300 mm3, the mice were randomly placed into different groups based on tumor volumes (5 mice per group). The mice were then injected with PBS, Patritumab-ADC, 1H7-ADC, Gatipotuzumab-ADC, 3E1-10D1-ADC, 1B2-10D1-ADC, 3E1-ADC, 1B2-ADC, or 10D1-ADC by intravenous (iv) administration. Details are shown in Table 14. Table 14 Group No. of mice Antibodies Dosage Route Frequency Total no. of administrations G1 5 PBS - iv QW 2 G2 5 Patritumab-ADC 3 mg / kg iv QW 2 G3 5 1H7-ADC 3 mg / kg iv QW 2 G4 5 Gatipotuzumab-ADC 3 mg / kg iv QW 2 G5 5 3E1-10D1-ADC 3 mg / kg iv QW 2 G6 5 1B2-10D1-ADC 3 mg / kg iv QW 2 G7 5 3E1-10D1-ADC 6 mg / kg iv QW 2 G8 5 1B2-10D1-ADC 6 mg / kg iv QW 2 G9 5 3E1-ADC 3 mg / kg iv QW 2 G10 5 1B2-ADC 3 mg / kg iv QW 2 G11 5 10D1-ADC 3 mg / kg iv QW 2

[0322] Tumor volumes of mice in different groups are shown in Figure 5. As all mice in groups G1, G2, G3 and G4 reached the euthanasia standard before day 39 after pooling, tumor size on Day 39 was not available.

[0323] 3E1-10D1-ADC and 1B2-10D1-ADC displayed the best Petition 870250084299, dated 09 / 18 / 2025, pp. 387 / 410 117 / 126 tumor growth inhibition effects at both 3 mg / kg and 6 mg / kg (G5-G8). Furthermore, 3E1-ADC, 1B2-ADC, and 10D1-ADC (G9-G11) showed better efficacy than the positive controls Patritumab-ADC, 1H7-ADC, and Gatipotuzumab-ADC (G2-G4). Example 12. Internalization of bispecific anti-HER3 / MUC1 ADCs

[0324] Anti-HER3 antibodies, anti-MUC1 antibodies, or bispecific anti-HER3 / MUC1 antibodies and ADCs were used to treat NUGC4 cells cultured in a cell culture plate, and internalization activity was detected after 15-24 hours of incubation with IncuCyte (Sartorius AG, IncuCyte® S3). The results are shown in Figures 6A-6C.

[0325] The data showed that the endocytosis activities of 3E1-10D1 and 1B2-10D1 were not altered after MMAE conjugation. Furthermore, 3E1-10D1 and 1B2-10D1 exhibited better internalization effects than the corresponding parental monoclonal antibodies (3E1, 1B2 and 10D1) and the positive controls (Patritumab analog, 1H7 analog and Gatipotuzumab analog). Example 13. Antibody-Drug Conjugates

[0326] The purified antibodies (analog 1B2, 3E1, 10D1, 1B210D1, 3E1-10D1, 1H7 and analog Gatipotuzumab) were coupled with CPT-1, CPT-2, CPT-3 or CPT-4 via a CPT-L ligand. For the antibody-drug conjugate names, CPTx (x = 1, 2, 3 or 4) is added directly after the antibody name. For example, when 1B2 is coupled to CPT-1, it is named 1B2-CPT1. As another example, when 1B2-10D1 is coupled to CPT-2, it is named 1B2-10D1-CPT2. Exemplary ADCs obtained by this method included: 1B2-10D1-CPT2, 3E1-10D1-CPT2, 1H7-CPT2 and Gatipotuzumab-CPT2.

[0327] The Patritumab analog and the Gatipotuzumab analog were also coupled to Dxd via the GGFG ligand, respectively, Petition 870250084299, dated 09 / 18 / 2025, pp. 388 / 410 118 / 126 for comparative purposes, and the resulting ADCs were named PatritumabDxd and Gatipotuzumab-Dxd.

[0328] An IgG1 monoclonal antibody targeting an irrelevant target was coupled to CPT-2 via the CPT-L ligand and to Dxd via the GGFG ligand, respectively. This formed the CPT2 isotype (ISO-CPT2) and the Dxd isotype (ISO-Dxd), which were used as isotype controls.

[0329] MS (mass spectrometry) was used to detect antibody coupling with drug molecules. MS detection results showed that the drug-to-antibody ratio (DAR) of the ADCs was approximately 4 or 8. All DARs of the controls (including 1H7CPT2, Gatipotuzumab-CPT2, ISO-CPT2, Gatipotuzumab-Dxd, Patritumab-Dxd, and ISO-Dxd) were 8. Regarding the names of 1B2-10D1-CPT2 and 3E1-10D1CPT2, if the DAR is approximately 4, the ADCs will be named 1B2-10D1CPT2(DAR4) and 3E1-10D1-CPT2(DAR4). If the DAR is around 8, the ADCs will be named 1B2-10D1-CPT2(DAR8) and 3E1-10D1-CPT2(DAR8). Example 14. Antitumor Activity in a PDX Model with Lung Cancer

[0330] The antitumor activities of 1B2-10D1-CPT2 and 3E1-10D1CPT2 were tested in vivo in a PDX model of lung cancer with low HER3 expression / high MUC1 expression. Specifically, patient-derived lung tumor fragments (2 mm*2 mm*2 mm) were grafted into the right flank of B-NDG mice. When the tumors in the mice reached a volume of approximately 200-300 mm3, the mice were randomly placed into different groups based on tumor volumes (5 mice per group) and then injected with PBS, 1B2-10D1-CPT2, or 3E1-10D1-CPT2 by intravenous administration. Tumor volumes were measured twice weekly.

[0331] As shown in Figure 7, tumor volumes in Petition 870250084299, dated 09 / 18 / 2025, pp. 389 / 410 119 / 126 all treatment groups (G2-G13) were inferior to those of the control group (G1). The results also showed that 1B2-10D1-CPT2 and 3E110D1-CPT2, both with DAR4 and DAR8, had different inhibitory effects on tumors, which were dose-dependent. 1B2-10D1-CPT2 and 3E110D1-CPT2 can significantly inhibit tumor growth with a % ICT (e.g., on day 24) greater than 100% when administered at a dose of 3 mg / kg or higher (G7, G9, G10, G11, G12, and G13).

[0332] Mouse tumor volume and survival after day 24 continued to be monitored.

[0333] On day 28, all mice in group G1 died, while groups G2 to G13 had better survival rates. At the end of the experiment on day 49, all mice in groups G1-G5 died; only one mouse died in G6, G8, G9 and G11; all mice in groups G7, G10, G12 and G13 survived, indicating that 1B2-10D1-CPT2 and 3E1-10D1-CPT2 have an excellent tumor inhibitory effect and good safety. Example 15. Antitumor activity in a NUGC-4 xenograft model

[0334] ADCs were tested for their effect on tumor growth in vivo in a gastric cancer xenograft NUGC4 model. Specifically, approximately 5 χ¹⁰⁵ NUGC-4 cells were injected subcutaneously into B-NDG mice. When the tumors in the mice reached a volume of approximately 200 mm³, the mice were randomly placed into different groups based on tumor volumes and then injected with PBS, Patritumab-Dxd, Gatipotuzumab-Dxd, 1B2-10D1-CPT2, or 3E1-10D1-CPT2 by intravenous administration. The frequency of administration was once weekly (2 administrations in total). Tumor volumes were measured twice weekly, and the results are shown in Figure 8. Petition 870250084299, dated 09 / 18 / 2025, pages 390 / 410 120 / 126

[0335] The results showed that 1B2-10D1-CPT2 and 3E1-10D1-CPT2, both with DAR4 and DAR8, inhibited tumor growth with a higher % ICT (e.g., on day 35) than the positive controls (G10 and G11). For example, when administered at a dose of 6 mg / kg, the % ICT in G6-G9 was greater than 90%, while it was 50.7% in G10 and 40.4% in G11, respectively. 1B2-10D1-CPT2 and 3E1-10D1-CPT2 have a good tumor inhibitory effect in a gastric cancer model. Example 16. Antitumor activity in an HCC70 xenograft model

[0336] ADCs were tested for their effect on tumor growth in vivo in an HCC70 breast ductal carcinoma xenograft model. Specifically, approximately 1 χ¹⁰⁷ HCC70 cells were injected subcutaneously into B-NDG mice. When the tumors in the mice reached a volume of approximately 200 mm³, the mice were randomly placed into different groups based on tumor volumes and then injected with PBS, Patritumab-Dxd, Gatipotuzumab-Dxd, ISO-Dxd, 1B2-10D1-CPT2, or 3E1-10D1-CPT2 by intravenous administration. Tumor volumes were measured twice weekly, and the body weights of the mice were also recorded. The results are shown in Figure 9A and Figure 9B.

[0337] Figure 9A presents the results of tumor volume measurements, in which 1B2-10D1-CPT2 and 3E1-10D1-CPT2, both with DAR4 and DAR8, exhibited different tumor inhibitory effects, which were dose-dependent. At a dose of 3 mg / kg, the % of ICT (e.g., on day 38) in groups G6-G9 was higher than in the control groups G10-G12. Specifically, on Day 38 the % of ICT was 89.1%, 106.1%, 100.0%, and 106.3% in groups G6 to G9, respectively; while it was lower with values ​​of 68.5% in group G10, 52.1% in group G11 and 19.7% in group G12, indicating that 1B2-10D1-CPT2 and 3E1-10D1-CPT2 have a good tumor inhibitory effect in Petition 870250084299, dated 09 / 18 / 2025, pp. 391 / 410 121 / 126 breast cancer model.

[0338] The weight of mice in different groups increased (as shown in Figure 9B), indicating that all antiHER3 / MUC1 ADCs were well tolerated and non-toxic to the mice. Example 17. Antitumor Activity in a PDX Cancer Model COLORECTAL

[0339] The antitumor activities of 1B2-10D1-CPT2 and 3E1-10D1CPT2 were tested in vivo in a PDX model of colorectal cancer with high HER3 expression / low MUC1 expression. Specifically, patient-derived colorectal tumor fragments (2 mm*2 mm*2 mm) were grafted into the right flank of B-NDG mice. When the tumors in the mice reached a volume of approximately 200-300 mm3, the mice were randomly placed into different groups based on tumor volumes (5 mice per group) and then injected with PBS, 1B2-10D1-CPT2, 3E1-10D1-CPT2, 1H7-CPT2, or Patritumab-Dxd, by intravenous administration. Tumor volumes were measured twice weekly.

[0340] As shown in Figure 10, 1B2-10D1-CPT2 and 3E110D1-CPT2, both with DAR4 and DAR8, inhibited tumor growth with a higher % ICT (e.g., on day 31) than the positive controls 1H7CPT2 and Patritumab-Dxd.

[0341] In a similar experiment, PDX model mice of colorectal cancer were injected with PBS, ISO-CPT2, 1B2-10D1CPT2(DAR8), 3E1-10D1-CPT2(DAR8), 1H7-CPT2 or Patritumab-Dxd by intravenous administration. Tumor volumes were measured twice weekly. The results are shown in Figure 11, 1B2-10D1-CPT2(DAR8) and 3E1-10D1-CPT2(DAR8) exhibited better antitumor activities than the positive controls 1H7-CPT2 and Patritumab-Dxd. Petition 870250084299, dated 09 / 18 / 2025, pp. 392 / 410 122 / 126

[0342] In another similar experiment, PDX model mice of colorectal cancer were injected with PBS (G1), ISO-CPT2 (G2), 3E1-10D1-CPT2(DAR8) (G3 and G4), a combination of 3E1-CPT2(DAR8) and 10D1-CPT2(DAR8) (G5), ISO-Dxd (G6), Patritumab-Dxd (G7), Gatipotuzumab-Dxd (G8) or 3E1-10D1-Dxd(DAR8) (G9) by intravenous administration. The frequency of administration was once a week (2 administrations in total). Tumor volumes were measured twice weekly, and the results are shown in Figure 16. 3E1-10D1-CPT2(DAR8) (G3 and G4) exhibited the best tumor inhibition effect compared to positive controls (G7 and G8) and combination therapy (G5), in a dose-dependent manner. Furthermore, 3E110D1-Dxd (G9) also inhibited tumor growth with superior efficacy compared to positive controls (G7 and G8). Example 18. Antitumor activity in a PDX model of pancreatic cancer.

[0343] The antitumor activities of 1B2-10D1-CPT2 and 3E1-10D1CPT2 were tested in vivo using PDX models of pancreatic cancer with HER3 / MUC1 co-overexpression. Patient-derived pancreatic tumor fragments (2 mm*2 mm*2 mm) were grafted into the right flank of B-NDG mice. When the tumors in the mice reached a volume of approximately 200-300 mm3, the mice were randomly placed into different groups based on tumor volumes (5 mice per group) and then injected with PBS, 1B2-10D1CPT2(DAR8), 3E1-10D1-CPT2(DAR8), 1H7-CPT2, Gatipotuzumab-CPT2 or Patritumab-Dxd was administered intravenously. Tumor volumes were measured twice weekly, and the body weights of the mice were also recorded.

[0344] As shown in Figure 12A, 1B2-10D1CPT2(DAR8), 3E1-10D1-CPT2(DAR8) induced significant inhibition of Petition 870250084299, dated 09 / 18 / 2025, pp. 393 / 410 123 / 126 tumor growth at 3 mg / kg with a higher % ICT (e.g., on Day 31) than that of positive controls 1H7-CPT2, Gatipotuzumab-CPT2, or Patritumab-Dxd.

[0345] As shown in Figure 12B, the weight of mice in the 1B2-10D1-CPT2(DAR8) and 3E1-10D1CPT2(DAR8) treatment groups (G3 and G4) increased. On the day of pooling, the mean weights of groups G3 and G4 were 21.8 g and 21.6 g, respectively. 31 days after pooling, the mean weight of groups G3 and G4 was 24.8 g and 24.4 g, respectively, resulting in weight changes of 114.4% and 113.1%. However, the body weight of the 1H7-CPT2, Gatipotuzumab-CPT2, and Patritumab-Dxd treatment groups remained constant or decreased slightly. The results indicate that both 1B2-10D1-CPT2 and 3E110D1-CPT2 were well tolerated by mice without causing any toxicity. Example 19. Antitumor activity in a PDX model of breast cancer.

[0346] The antitumor activities of 1B2-10D1-CPT2 and 3E1-10D1CPT2 were tested in vivo using PDX models of breast cancer with low HER3 expression / high MUC1 expression. Patient-derived breast tumor fragments (2 mm*2 mm*2 mm) were grafted onto the right flank of B-NDG mice. Tumor volumes were measured twice weekly and the results are shown in Figure 13.

[0347] 1B2-10D1-CPT2 and 3E1-10D1-CPT2, both with DAR4 and DAR8, significantly inhibited tumor growth with a higher % ICT (e.g., on day 16) than the positive controls GatipotuzumabCPT2 and Patritumab-Dxd. This indicates that anti-HER3 / MUC1 ADCs have demonstrated potential in the treatment of breast cancer. Example 20. Antitumor activity in a PDX model of gastric cancer.

[0348] ADCs were tested for their effects on Petition 870250084299, dated 09 / 18 / 2025, pp. 394 / 410 124 / 126 In vivo tumor growth in a patient-derived xenograft model of gastric cancer. Specifically, patient-derived gastric tumor fragments (2 mm*2 mm*2 mm) were grafted into the right flank of B-NDG mice. When tumor volumes reached approximately 250 mm3, mice were randomly placed into different groups based on tumor volumes. Mice were then injected with PBS (G1), ISO-CPT2 (G2), 3E1-10D1-CPT2 (DAR8) (G3 and G4), a combination of 3E1-CPT2 (DAR8) and 10D1-CPT2 (DAR8) (G5), Patritumab-Dxd (G6), Gatipotuzumab-Dxd (G7), ISO-ADC (G8), or 3E1-10D1-ADC (G9) via intravenous (iv) administration. The administration frequency was once a week (2 administrations in total).

[0349] Tumor volumes were measured twice weekly and the results are shown in Figure 15. The treatment groups (G2-G9) showed different tumor inhibition compared to the PBS group (G1). 3E1-10D1-CPT2(DAR8) (G3 and G4) exhibited a better tumor inhibition effect compared to the positive controls (G6 and G7) and the combination therapy (G5), in a dose-dependent manner. Furthermore, 3E1-10D1-ADC (G9) also inhibited tumor growth with superior efficacy compared to the positive controls (G6 and G7). Example 21. Antitumor Activity in a PDX Model with Lung Cancer

[0350] 3E1-10D1-CPT2(DAR8) was tested for its effect on tumor growth in vivo in a patient-derived xenograft model of lung cancer. Immunohistochemical (IHC) staining of patient-derived tumor tissues was performed, and the results showed that the histochemical score (H-score) of HER3 and MUC1 expression levels in patient-derived tumor tissues were 104.68 and 89.9, respectively. Patient-derived lung tumor fragments (2 Petition 870250084299, dated 09 / 18 / 2025, pages 395 / 410 Tumors (125 / 126 mm*2 mmx2 mm) were grafted onto the right flank of naked BALB / c mice. When tumor volumes reached approximately 200 mm3, mice were randomly placed into different groups based on tumor volumes. Mice were then injected with 5% glucose (G1) or 3 mg / kg of 3E1-10D1-CPT2(DAR8) (G2) by intravenous (iv) administration.

[0351] Tumor volumes were measured twice a week, and the results are shown in Figure 17, in which 3E1-10D1ADC(DAR8) exhibited a good tumor growth inhibition effect in the lung cancer model. Example 22. Antitumor activity in a PDX model of ovarian cancer.

[0352] 3E1-10D1-CPT2(DAR8) tests its effect on tumor growth in a patient-derived xenograft model of ovarian cancer. IHC staining of patient-derived tumor tissues was performed, and the results showed that the Hf of HER3 and MUC1 expression levels in patient-derived tumor tissues were 245.87 and 112.00, respectively. When tumor volumes reached approximately 200 mm3, mice were randomly placed into different groups based on tumor volumes. Mice were then injected with 5% glucose (G1) or 6 mg / kg of 3E1-10D1-CPT2(DAR8) (G2) via intravenous administration.

[0353] Tumor volumes are measured twice a week, and periodic results are shown in Figure 18, in which 3E1-10D1ADC(DAR8) also exhibits a good tumor growth inhibition effect in the ovarian cancer model. Example 23. Toxicological evaluation

[0354] In a preliminary experiment to investigate the safety and toxicokinetic (TC) profile, 1B2-10D1-CPT2(DAR4), 1B2-10D1CPT2(DAR8), 3E1-10D1-CPT2(DAR4) or 3E1-10D1-CPT2(DAR8) was administered by IV injection to cynomolgus monkeys, respectively, three times Petition 870250084299, dated 09 / 18 / 2025, pages 396 / 410 126 / 126 with a 3-week interval (on Day 1, Day 22, and Day 43). The dose formulation is shown in Table 15. Animals were then sacrificed on Day 50 for macroscopic and histopathological examination. Mortality / moribundity, general observations, body weights, feed intake, clinical pathology (hematology, coagulation, serum chemistry, and urinalysis), and macroscopic lesions were evaluated. Blood samples were also collected for CT analysis, and primary CT parameters, e.g., Tmax, Cmax, and AUC(0-t) for payload, total antibody, and ADC, were calculated. As a result, it was found that T1B2-10D1-CPT2(DAR4), 1B2-10D1-CPT2(DAR8), 3E1-10D1-CPT2(DAR4) and 3E1-10D1-CPT2(DAR8) have favorable safety profiles. Table 15 ADC Group Dose (mg / kg) Concentration (mg / ml) 1B2-10D1-CPT2(DAR4) G1 30 mg / kg 3 mg / ml G2 40 mg / kg 4 mg / ml 1B2-10D1-CPT2(DAR8) G3 15 mg / kg 1.5 mg / ml G4 20 mg / kg 2 mg / ml 3E1-10D1-CPT2(DAR4) G5 30 mg / kg 3 mg / ml G6 40 mg / kg 4 mg / ml 3E1-10D1-CPT2(DAR8) G7 15 mg / kg 1.5 mg / ml G8 20 mg / kg 2 mg / ml Other Modalities

[0355] It should be understood that, although the invention has been described in conjunction with the detailed description thereof, the preceding description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages and modifications are within the scope of the following claims. Petition 870250084299, dated 09 / 18 / 2025, pp. 397 / 410

Claims

1 / 11 Claims 1. ANTI-HER3 / MUC1 ANTIBODY OR ANTIGEN-BINDING FRAGMENT THEREOF, characterized by comprising a first antigen-binding domain that specifically binds to HER3; and a second antigen-binding domain that specifically binds to MUC1.

2. ANTIBODY, according to claim 1, characterized in that the first antigen-binding domain comprises a first variable heavy chain region (VH1) and a first variable light chain region (VL1); and the second antigen-binding domain comprises a second variable heavy chain region (VH2) and a second variable light chain region (VL2); and wherein (i) the first variable heavy chain region (VH1) comprises complementarity-determining regions (CDRs) 1, 2 and 3, wherein the CDR1 region of VH1 comprises an amino acid sequence that is at least 80% identical to a selected CDR1 amino acid sequence of VH1,The CDR2 region of VH1 comprises an amino acid sequence that is at least 80% identical to a selected CDR2 amino acid sequence of VH1, and the CDR3 region of VH1 comprises an amino acid sequence that is at least 80% identical to a selected CDR3 amino acid sequence of VH1; and the first variable light chain region (VL1) comprises CDRs 1, 2, and 3, wherein the CDR1 region of VL1 comprises an amino acid sequence that is at least 80% identical to a selected CDR1 amino acid sequence of VL1, the CDR2 region of VL1 comprises an amino acid sequence that is at least 80% identical to a selected CDR2 amino acid sequence of VL1, and the CDR3 region of VL1 comprises an amino acid sequence that is at least 80% identical to Petition 870250084299, dated 09 / 18 / 2025, p. 398 / 410 2 / 11 a selected amino acid sequence of CDR3 from VL1, wherein the amino acid sequences of selected CDRs 1, 2 and 3 from VH1,The amino acid sequences of selected VL1 CDRs 1, 2, and 3 are one of the following: (1) the amino acid sequences of selected VH1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 7 to 9, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2, and 3 are presented in SEQ ID NOs: 1 to 3, respectively; (2) the amino acid sequences of selected VH1 CDRs 1, 2 and 3 are presented in SEQ ID NOs: 19 to 21, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2 and 3 are presented in SEQ ID NOs: 13 to 15, respectively; (3) the amino acid sequences of selected VH1 CDRs 1, 2 and 3 are presented in SEQ ID NOs: 4 to 6, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2 and 3 are presented in SEQ ID NOs: 1 to 3, respectively; and (4) the amino acid sequences of selected VH1 CDRs 1, 2 and 3 are presented in SEQ ID NOs: 16 to 18, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2 and 3 are presented in SEQ ID NOs: 13 to 15, respectively; and / or (ii) the second variable heavy chain region (VH2) comprises CDRs 1, 2 and 3, wherein the CDR1 region of VH2 comprises an amino acid sequence that is at least 80% identical to a selected CDR1 amino acid sequence of VH2, the CDR2 region of VH2 comprises an amino acid sequence that is at least 80% identical to a selected CDR2 amino acid sequence of VH2,and the CDR3 region of VH2 comprises an amino acid sequence that is at least 80% identical to a selected CDR3 amino acid sequence from Petition 870250084299, dated 09 / 18 / 2025, page 399 / 410 3 / 11; and the second variable light chain region (VL2) comprises CDRs 1, 2 and 3, wherein the CDR1 region of VL2 comprises an amino acid sequence that is at least 80% identical to a selected CDR1 amino acid sequence of VL2, the CDR2 region of VL2 comprises an amino acid sequence that is at least 80% identical to a selected CDR2 amino acid sequence of VL2, and the CDR3 region of VL2 comprises an amino acid sequence that is at least 80% identical to a selected CDR3 amino acid sequence of VL2, wherein the amino acid sequences of selected CDRs 1, 2 and 3 of VL2 and the amino acid sequences of selected CDRs 1, 2 and 3 of VL2 are one of the following: (1) the amino acid sequences of CDRs 1,(1) The amino acid sequences of selected VH2 CDRs 2 and 3 are shown in SEQ ID NOs: 10 to 12, respectively, and the amino acid sequences of selected VL2 CDRs 1, 2, and 3 are shown in SEQ ID NOs: 1 to 3, respectively; and (2) The amino acid sequences of selected VH2 CDRs 1, 2, and 3 are shown in SEQ ID NOs: 22 to 24, respectively, and the amino acid sequences of selected VL2 CDRs 1, 2, and 3 are shown in SEQ ID NOs: 13 to 15, respectively.

3. ANTIBODY, according to claim 2, characterized by: (1) the amino acid sequences of selected VH1 CDRs 1, 2 and 3 being presented in SEQ ID NOs: 7 to 9, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2 and 3 being presented in SEQ ID NOs: 1 to 3, respectively, and the amino acid sequences of selected VH2 CDRs 1, 2 and 3 being presented in SEQ ID NOs: 10 to 12, respectively, and the amino acid sequences of Petition 870250084299, dated 09 / 18 / 2025,pg. 400 / 410 4 / 11 CDRs 1, 2 and 3 of VL2 selected to be displayed in SEQ ID NOs: 1 to 3, respectively; (2) the amino acid sequences of CDRs 1, 2 and 3 of VH1 selected being presented in SEQ ID NOs: 19 to 21, respectively, and the amino acid sequences of CDRs 1, 2 and 3 of VL1 selected being presented in SEQ ID NOs: 13 to 15, respectively, and the amino acid sequences of CDRs 1, 2 and 3 of VH2 selected being presented in SEQ ID NOs: 22 to 24, respectively, and the amino acid sequences of CDRs 1, 2 and 3 of VL2 selected being shown in SEQ ID NOs: 13 to 15, respectively; (3) the amino acid sequences of selected VH1 CDRs 1, 2 and 3 being presented in SEQ ID NOs: 4 to 6, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2 and 3 being presented in SEQ ID NOs: 1 to 3, respectively, and the amino acid sequences of CDRs 1,2 and 3 of selected VH2 being displayed in SEQ ID NOs: 10 to 12, respectively, and the amino acid sequences of CDRs 1, 2 and 3 of selected VL2 being displayed in SEQ ID NOs: 1 to 3, respectively; or (4) the amino acid sequences of selected VH1 CDRs 1, 2 and 3 being displayed in SEQ ID NOs: 16 to 18, respectively, and the amino acid sequences of selected VL1 CDRs 1, 2 and 3 being displayed in SEQ ID NOs: 13 to 15, respectively, and the amino acid sequences of selected VH2 CDRs 1, 2 and 3 being displayed in SEQ ID NOs: 22 to 24, respectively, and the amino acid sequences of selected VL2 CDRs 1, 2 and 3 being displayed in SEQ ID NOs: 13 to 15, respectively.

4. ANTIBODY, according to any one of claims 2 to 3, characterized by: Petition 870250084299, dated 09 / 18 / 2025, p. 401 / 410 5 / 11 (i) a VH1 comprise a sequence that is at least 80%, 85%, 90%, 95%, 99% or 100% identical to SEQ ID NO: 27,VL1 comprises a sequence that is at least 80%, 85%, 90%, 95%, 99%, or 100% identical to SEQ ID NO: 25, VH2 comprises a sequence that is at least 80%, 85%, 90%, 95%, 99%, or 100% identical to SEQ ID NO: 28, and VL2 comprises a sequence that is at least 80%, 85%, 90%, 95%, 99%, or 100% identical to SEQ ID NO: 25; or (ii) VH1 comprises a sequence that is at least 80%, 85%, 90%, 95%, 99% or 100% identical to SEQ ID NO: 26, VL1 comprises a sequence that is at least 80%, 85%, 90%, 95%, 99% or 100% identical to SEQ ID NO: 25, VH2 comprises a sequence that is at least 80%, 85%, 90%, 95%, 99% or 100% identical to SEQ ID NO: 28, and VL2 comprises a sequence that is at least 80%, 85%, 90%, 95%, 99% or 100% identical to SEQ ID NO: 25; optionally wherein: (i) (a) VH1 comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 27,and VL1 comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 25, or (b) VH1 comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 26, and VL1 comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 25; and / or (ii) VH2 comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 28, and VL2 comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 25; optionally where: (i) (a) VH1 comprises the sequence of SEQ ID NO: 27 and VL1 comprises the sequence of SEQ ID NO: 25; or (b) VH1 comprises the sequence with SEQ ID NO: 26 and VL1 comprises the sequence with SEQ ID NO: 25; and / or, (ii) VH2 comprises the sequence with SEQ ID NO: 28 and VL2 comprises the sequence with SEQ ID NO:

25.

5. ANTIBODY, according to any one of claims 2 to 4,characterized by: (i) VH1 comprising VH1 CDR1, VH1 CDR2 and VH1 CDR3 that are identical to VH CDR1, VH CDR2 and VH CDR3 of a selected VH sequence; and VL1 comprising VL1 CDR1, VL1 CDR2 and VL1 CDR3 that are identical to VL CDR1, VL CDR2 and VL CDR3 of a selected VL sequence, wherein the selected VH sequence and the selected VL sequence are one of the following: (1) the selected VH sequence is SEQ ID NO: 26 and the selected VL sequence is SEQ ID NO: 25; and (2) the selected VH sequence is SEQ ID NO: 27 and the selected VL sequence is SEQ ID NO: 25; and / or (ii) VH2 comprises VH2 CDR1, VH2 CDR2, and VH2 CDR3 that are identical to VH CDR1, VH CDR2, and VH CDR3 of a selected VH sequence; and VL2 comprises VL2 CDR1, VL2 CDR2, and VL2 CDR3 that are identical to VL CDR1, VL CDR2, and VL CDR3 of a selected VL sequence,wherein the selected VH sequence is SEQ ID NO: 28 and the selected VL sequence is SEQ ID NO:

25.

6. ANTIBODY, according to any one of claims 1 to 5, characterized by one or more of the following: (i) the first antigen-binding domain binds specifically to human or monkey HER3; and / or the second antigen-binding domain binds specifically to human or monkey MUC1; Petition 870250084299, dated 09 / 18 / 2025, pp. 403 / 410 7 / 11 (ii) the first antigen-binding domain is human or humanized; and / or the second antigen-binding domain is human or humanized; (iii) the antibody is a multispecific antibody (e.g., a bispecific antibody); (iv) the first antigen-binding domain is a single-strand variable fragment (scFv); and / or the second antigen-binding domain is an scFv; (v) the first variable region of the light chain and the second variable region of the light chain are identical.

7. NUCLEIC ACID,characterized by comprising a polynucleotide encoding the anti-HER3 / MUC1 antibody or antigen-binding fragment thereof, as defined in any one of claims 1 to 6.

8. VECTOR, characterized by comprising the nucleic acid, as defined in claim 7.

9. CELL, characterized by comprising the nucleic acid, as defined in claim 7, or a vector comprising the nucleic acid.

10. METHOD FOR PRODUCING AN ANTI-HER3 / MUC1 ANTIBODY OR AN ANTIGEN-BINDING FRAGMENT THEREOF, characterized by comprising: (a) culturing the cell, as defined in claim 9, under conditions sufficient for the cell to produce the anti-HER3 / MUC1 antibody or antigen-binding fragment thereof; and (b) collecting the anti-HER3 / MUC1 antibody or antigen-binding fragment thereof produced by the cell.

11. ANTI-HER3 / MUC1 ANTIBODY CONJUGATE - Petition 870250084299, dated 09 / 18 / 2025, pp. 404 / 410 8 / 11 DRUG (ADC),characterized by comprising a therapeutic agent covalently linked to an anti-HER3 / MUC1 antibody or antigen-binding fragment thereof, as defined in any one of claims 1 to 6.

12. CONJUGATE, according to claim 11, characterized in that the therapeutic agent is a cytotoxic or cytostatic agent; optionally, the therapeutic agent is selected from MMAE, MMAF or one of the following: claims 11 to 12, characterized in that the therapeutic agent is linked to the antibody or antigen-binding fragment thereof, or to the antigen-binding protein construct by means of a ligand, and the ligand has a structure of: OH OH 14. CONJUGATE, according to any one of claims 11 to 13, characterized in that it has a structure of: Petition 870250084299, dated 09 / 18 / 2025, p. 405 / 410 9 / 11 where n = 1 to 8; where “Ab” represents the antibody or antigen-binding fragment thereof,or the construction of an antigen-binding protein.

15. USE OF ANTI-HER3 / MUC1 ANTIBODY OR ANTIGEN-BINDING FRAGMENT THEREOF, as defined in any one of claims 1 to 6, or of the anti-HER3 / MUC1 antibody-drug conjugate, as defined in any one of claims 11 to 14, characterized in being in the manufacture of a medicament for the treatment of cancer in a subject.

16. USE OF ANTI-HER3 / MUC1 ANTIBODY OR ANTIGEN-BINDING FRAGMENT THEREOF, as defined in any one of claims 1 to 6, or of the anti-HER3 / MUC1 antibody-drug conjugate, as defined in any one of claims 11 to 14, and of an anti-PD1 antibody, characterized in being in the manufacture of a medicament or combination for the treatment of cancer in a subject. Petition 870250084299, dated 09 / 18 / 2025, pp. 406 / 410 10 / 11 17. USE, in accordance with any of claims 15 to 16,characterized by one or more of the following: (i) the cancer being a cancer that expresses HER3 and / or MUC1 (e.g., both HER3 and MUC1); (ii) the cancer being esophageal cancer, colorectal cancer, gastric cancer, breast cancer, endometrial carcinoma, lung cancer, melanoma, ovarian cancer, bladder cancer, stomach cancer, non-Hodgkin lymphoma, head and neck cancer, pancreatic cancer, lung adenocarcinoma, and cervical cancer; (iii) the subject being a human being; (iv) an anti-PD1 antibody being administered; (v) chemotherapy being administered.

18. METHOD FOR REDUCING THE TUMOR GROWTH RATE or killing a tumor cell, characterized by comprising placing a tumor cell in contact with an effective amount of a composition comprising the anti-HER3 / MUC1 antibody or antigen-binding fragment thereof, as defined in any of claims 1 to 6, or an anti-HER3 / MUC1 antibody-drug conjugate,as defined in any one of claims 11 to 14.

19. PHARMACEUTICAL COMPOSITION, characterized by comprising a pharmaceutically acceptable carrier and (a) the anti-HER3 / MUC1 antibody or antigen-binding fragment thereof, as defined in any one of claims 1 to 6, and / or (b) the anti-HER3 / MUC1 antibody-drug conjugate, as defined in any one of claims 11 to 14.

20. ANTI-HER3 / MUC1 ANTIBODY-DRUG CONJUGATE (ADC), characterized by comprising a therapeutic agent covalently linked to a bispecific antibody or antigen-binding fragment thereof comprising: a first antigen-binding domain that specifically binds to HER3; and a second antigen-binding domain that binds specifically to MUC1.

21. CONJUGATED, according to any one of claims 11 to 14 or 20,characterized by the drug-to-antibody ratio (DAR) being approximately 4 or 8. Petition 870250084299, dated 09 / 18 / 2025, pp. 408 / 410.

Images