Application of anti-PD-L1 / CD47 bispecific antibodies in combination therapy

Abstract

The use of anti-PD-L1 / CD47 bispecific antibodies in combination therapy includes administering effective amounts of anti-PD-L1 / CD47 bispecific antibodies and anti-CTLA-4 antibodies to patients in need.

Description

Application of anti-PD-L1 / CD47 bispecific antibodies in combination therapy Technical Field

[0001] The present invention belongs to the field of biomedicine, and specifically relates to the use of anti-PD-L1 / CD47 bispecific antibodies in combined medication. Background Art

[0002] Monoclonal antibody immunotherapy has brought lasting survival benefits to some patients, realizing their hope for a cure. With in-depth research into cancer and various other diseases, we have come to realize that the occurrence and progression of diseases are involved in multiple signaling pathways and mechanisms. However, the stimulation or inhibition of a single monoclonal antibody alone cannot completely kill the tumor, resulting in a limited response rate for single-agent therapy. Bispecific monoclonal antibodies (BsAbs) can simultaneously bind to two antigens or two different epitopes of a single antigen, blocking or activating different signaling pathways and more effectively mediating immune cell killing of tumor cells, thus potentially achieving better therapeutic effects. Summary of the Invention

[0003] The purpose of the present invention is to provide a method or use of an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment for combined treatment of tumors.

[0004] In one aspect, the present invention provides a pharmaceutical composition for treating tumors, comprising an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment.

[0005] In another aspect, the present invention provides a combination of an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment for use in treating tumors.

[0006] On the other hand, the present invention provides the use of an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment in the preparation of a combination with an anti-CTLA-4 antibody or antigen-binding fragment for treating tumors.

[0007] In another aspect, the present invention provides use of an anti-CTLA-4 antibody or antigen-binding fragment in the preparation of a medicament for treating tumors in combination with an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment.

[0008] On the other hand, the present invention provides use of an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment in preparing a pharmaceutical composition for treating tumors.

[0009] In another aspect, the present invention provides a method for treating tumors, comprising: administering an effective amount of an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment to a patient in need thereof.

[0010] On the other hand, the present invention also provides a kit comprising an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment (or preparation), an anti-CTLA-4 antibody or antigen-binding fragment (or preparation), and instructions for guiding administration to patients in need thereof.

[0011] In some embodiments, the anti-PD-L1 / CD47 bispecific antibody is selected from the antibodies Kh2NF-PC and Kh2NF-PC-NL in patent CN109970860A, the antibodies BsAb1, BsAb2, and BsAb3 in patent CN115397853A, and the antibodies 2MW1531-p and 2MW1531-m1 to m11 in patent CN114478770A.

[0012] In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment comprises a variable region a that specifically binds to PD-L1, and a variable region b that specifically binds to CD47; wherein

[0013] The variable region a comprises one or more amino acid sequences of (a)-(f):

[0014] (a) a VHa CDR1 comprising the amino acid sequence of SEQ ID NO: 1, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO: 1;

[0015] (b) a VHa CDR2 comprising the amino acid sequence shown in SEQ ID NO: 2, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO: 2;

[0016] (c) a VHa CDR3 comprising the amino acid sequence shown in SEQ ID NO: 3, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO: 3;

[0017] (d) VLa CDR1 comprising the amino acid sequence set forth in SEQ ID NO:4, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence set forth in SEQ ID NO:4;

[0018] (e) VLa CDR2 comprising the amino acid sequence shown in SEQ ID NO: 5, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO: 5;

[0019] (f) a VLa CDR3 comprising the amino acid sequence of SEQ ID NO: 6, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO: 6;

[0020] The variable region b comprises one or more amino acid sequences of (g)-(l):

[0021] (g) a VHb CDR1 comprising the amino acid sequence of SEQ ID NO: 7, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO: 7;

[0022] (h) a VHb CDR2 comprising the amino acid sequence of SEQ ID NO: 8, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO: 8;

[0023] (i) a VHb CDR3 comprising the amino acid sequence of SEQ ID NO: 9, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO: 9;

[0024] (j) a VLb CDR1 comprising the amino acid sequence of SEQ ID NO: 4, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO: 4;

[0025] (k) a VLb CDR2 comprising the amino acid sequence of SEQ ID NO: 5, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO: 5;

[0026] (1) VLb CDR3 comprising the amino acid sequence shown in SEQ ID NO: 6, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO: 6.

[0027] In some embodiments, the variable region a comprises VHa CDR1, VHa CDR2, and VHa CDR3; wherein, VHa CDR1 comprises the amino acid sequence of SEQ ID NO: 1, VHa CDR2 comprises the amino acid sequence of SEQ ID NO: 2, and VHa CDR3 comprises the amino acid sequence of SEQ ID NO: 3; the variable region b comprises VHb CDR1, VHb CDR2, and VHb CDR3; wherein, VHb CDR1 comprises the amino acid sequence of SEQ ID NO: 7, VHb CDR2 comprises the amino acid sequence of SEQ ID NO: 8, and VHb CDR3 comprises the amino acid sequence of SEQ ID NO: 9.

[0028] In some embodiments, the variable region a comprises VLa CDR1, VLa CDR2, and VLa CDR3; wherein VLa CDR1 comprises the amino acid sequence of SEQ ID NO:4, VLa CDR2 comprises the amino acid sequence of SEQ ID NO:5, and VLa CDR3 comprises the amino acid sequence of SEQ ID NO:6; the variable region b comprises VLb CDR1, VLb CDR2, and VLb CDR3; wherein VLb CDR1 comprises the amino acid sequence of SEQ ID NO:4, VLb CDR2 comprises the amino acid sequence of SEQ ID NO:5, and VLb CDR3 comprises the amino acid sequence of SEQ ID NO:6.

[0029] In some embodiments, the variable region a comprises VHa CDR1, VHa CDR2, VHa CDR3, VLa CDR1, VLa CDR2, and VLa CDR3; the variable region b comprises VHb CDR1, VHb CDR2, VHb CDR3, VLb CDR1, VLb CDR2, and VLb CDR3; wherein VHa CDR1 comprises the amino acid sequence of SEQ ID NO: 1, VHa CDR2 comprises the amino acid sequence of SEQ ID NO: 2, VHa CDR3 comprises the amino acid sequence of SEQ ID NO: 3, VHb CDR1 comprises the amino acid sequence of SEQ ID NO: 7, VHb CDR2 comprises the amino acid sequence of SEQ ID NO: 8, VHb CDR3 comprises the amino acid sequence of SEQ ID NO: 9, VLa CDR1 and VLb CDR1 both comprise the amino acid sequence of SEQ ID NO: 4, VLa CDR2 and VLb CDR2 both comprise the amino acid sequence of SEQ ID NO: 5, VLa CDR3 and VLb The CDR3s all comprise the amino acid sequence shown in SEQ ID NO:6.

[0030] In some embodiments, the variable region a comprises VHa and VLa; wherein, the VHa comprises the amino acid sequence shown in SEQ ID NO: 10, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO: 10, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO: 10; and / or the VLa comprises the amino acid sequence shown in SEQ ID NO: 12, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO: 12, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO: 12.

[0031] In some embodiments, the variable region b comprises VHb and VLb; wherein the VHb comprises the amino acid sequence shown in SEQ ID NO: 11, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO: 11, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO: 11; and / or the VLb comprises the amino acid sequence shown in SEQ ID NO: 12, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO: 12, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO: 12.

[0032] In some embodiments, the variable region a comprises VHa and VLa; the variable region b comprises VHb and VLb; the VHa comprises the amino acid sequence of SEQ ID NO: 10, or an amino acid sequence having at least 80% identity with the amino acid sequence of SEQ ID NO: 10, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO: 10; the VHb comprises the amino acid sequence of SEQ ID NO: 11, or an amino acid sequence having at least 80% identity with the amino acid sequence of SEQ ID NO: 11, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO: 11; the VLa and VLb both comprise the amino acid sequence of SEQ ID NO: 12, or an amino acid sequence having at least 80% identity with the amino acid sequence of SEQ ID NO: 12, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO: 12.

[0033] In some embodiments, the variable region a comprises VHa and VLa; the variable region b comprises VHb and VLb; the VHa comprises the amino acid sequence shown in SEQ ID NO: 10, the VHb comprises the amino acid sequence shown in SEQ ID NO: 11, and the VLa and VLb both comprise the amino acid sequence shown in SEQ ID NO: 12.

[0034] In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment further comprises a heavy chain constant region a (CHa), a light chain constant region a (CLa), a heavy chain constant region b (CHb), and a light chain constant region b (CLb).

[0035] In some embodiments, the VHa is connected to CHa to form heavy chain a, and the VHb is connected to CHb to form heavy chain b. In some embodiments, the VLa is connected to CLa to form light chain a, and the VLb is connected to CLb to form light chain b.

[0036] In some embodiments, the CHa is of IgG1 subtype. In some embodiments, the CHb is of IgG1 subtype.

[0037] In some embodiments, the CHa and / or CHb comprises one or more of the following amino acid mutations: Y349C, S354C, T366W, T366S, L368A, and Y407V, wherein the amino acid positions are numbered according to Eu.

[0038] In some embodiments, the CHa and / or CHb comprises the following amino acid mutation: N297A, wherein amino acid positions are numbered according to Eu.

[0039] In some embodiments, one of the heavy chain constant regions of CHa and CHa comprises one or more of the following amino acid mutations: N297A, Y349C, T366S, L368A, and Y407V, wherein the amino acid positions are numbered according to Eu. In some embodiments, the other of the heavy chain constant regions of CHa and CHa comprises one or more of the following amino acid mutations: N297A, S354C, and T366W, wherein the amino acid positions are numbered according to Eu.

[0040] In some embodiments, the CHa comprises the amino acid sequence set forth in SEQ ID NO: 13 or 15, or an amino acid sequence having at least 80% identity with the amino acid sequence set forth in SEQ ID NO: 13 or 15, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence set forth in SEQ ID NO: 13 or 15; the CHb comprises the amino acid sequence set forth in SEQ ID NO: 13 or 15, or an amino acid sequence having at least 80% identity with the amino acid sequence set forth in SEQ ID NO: 13 or 15, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence set forth in SEQ ID NO: 13 or 15.

[0041] In some embodiments, the CHa comprises the amino acid sequence set forth in SEQ ID NO: 13, and the CHb comprises the amino acid sequence set forth in SEQ ID NO: 15. In some embodiments, the CHa comprises the amino acid sequence set forth in SEQ ID NO: 15, and the CHb comprises the amino acid sequence set forth in SEQ ID NO: 13.

[0042] In some embodiments, the CLa or CLb comprises the amino acid sequence shown in SEQ ID NO: 14, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO: 14, or an amino acid sequence having one or more conservative amino acid substitutions with the amino acid sequence shown in SEQ ID NO: 14.

[0043] In some embodiments, both CLa and CLb comprise the amino acid sequence shown in SEQ ID NO:14.

[0044] In some embodiments, the anti-PD-L1 / CD47 bispecific antibody comprises heavy chain a, heavy chain b, light chain a, and light chain b.

[0045] In some embodiments, the heavy chain a comprises the amino acid sequence shown in SEQ ID NO: 16, or an amino acid sequence that is at least 80% identical to the amino acid sequence shown in SEQ ID NO: 16, or an amino acid sequence that has one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO: 16; the heavy chain b comprises the amino acid sequence shown in SEQ ID NO: 17, or an amino acid sequence that is at least 80% identical to the amino acid sequence shown in SEQ ID NO: 17, or an amino acid sequence that has one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO: 17.

[0046] In some embodiments, the light chain a and light chain b comprise the same amino acid sequence. In some embodiments, the light chain a or light chain b comprises the amino acid sequence set forth in SEQ ID NO: 18, or an amino acid sequence having at least 80% identity to the amino acid sequence set forth in SEQ ID NO: 18, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence set forth in SEQ ID NO: 18.

[0047] In some embodiments, the anti-PD-L1 / CD47 bispecific antibody comprises heavy chain a shown in SEQ ID NO: 16, heavy chain b shown in SEQ ID NO: 17, and light chain a and light chain b shown in SEQ ID NO: 18.

[0048] In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment provided herein has an affinity index KD ≤ 10 nM for PD-L1. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment provided herein has an affinity index KD ≤ 1 nM for PD-L1. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment provided herein has an affinity index KD ≤ 0.5 nM for PD-L1. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment provided herein has an affinity index KD ≤ 0.2 nM for PD-L1. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment provided herein has an affinity index KD ≤ 100 nM for CD47. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment provided herein has an affinity index KD ≤ 30 nM for CD47. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment provided herein has an affinity index KD ≤ 10 nM for CD47. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment provided herein has an affinity index KD ≤ 5 nM for CD47. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment provided herein has an affinity index KD ≤ 4 nM for CD47. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment provided herein binds to PD-L1 with greater affinity than to CD47.

[0049] In some embodiments, the affinity index KD of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment to PD-L1 is ≤1 nM, and the affinity index KD of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment to CD47 is ≤30 nM.

[0050] In some embodiments, the affinity index KD of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment to PD-L1 is ≤1 nM, and the affinity index KD of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment to CD47 is ≤10 nM.

[0051] In some embodiments, the anti-PD-L1 / CD47 bispecific antibody is antibody BsAb-71-N297A, comprising heavy chain a shown in SEQ ID NO: 16, heavy chain b shown in SEQ ID NO: 17, and light chain a and light chain b shown in SEQ ID NO: 18.

[0052] In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is expressed by CHO cells. The anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment can also be expressed by other cells (e.g., HEK cells (such as HEK293F cells), BHK cells, Cos1 cells, Cos7 cells, CV1 cells, or mouse L cells).

[0053] In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is selected from the bispecific antibodies or antigen-binding fragments disclosed in patent WO2022095970A.

[0054] In some embodiments, the anti-CTLA-4 antibody is selected from tremelimumab ( or its biosimilars), ipilimumab ( or their biosimilars), antibodies huC1D1-V1 to huC1D1-V12, huD1D5-V1 to huD1D5-V12 in patent CN115443291A, antibody 1, antibody 1-1, antibody 1-2, antibody 2, antibody 2-1 to 2-6 in patent CN113490687A, antibodies BA-087-05-19, BA-087-08-32 in patent CN112996813A, antibodies AH01672, AH01674, AH01679, AH01686, AH01695, AH01696, AH01704 in patent CN115850475A.

[0055] In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment comprises one or more amino acid sequences of (1)-(6):

[0056] (1) VH CDR1 comprising the amino acid sequence of SEQ ID NO: 19, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO: 19;

[0057] (2) VH CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 20, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence set forth in SEQ ID NO: 20;

[0058] (3) a VH CDR3 comprising the amino acid sequence set forth in SEQ ID NO: 21, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence set forth in SEQ ID NO: 21;

[0059] (4) a VL CDR1 comprising the amino acid sequence set forth in SEQ ID NO: 22, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence set forth in SEQ ID NO: 22;

[0060] (5) VL CDR2 comprising the amino acid sequence set forth in SEQ ID NO: 23, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence set forth in SEQ ID NO: 23;

[0061] (6) VL CDR3 comprising the amino acid sequence shown in SEQ ID NO: 24, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO: 24.

[0062] In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment comprises VH CDR1, VH CDR2, and VH CDR3; wherein VH CDR1 comprises the amino acid sequence of SEQ ID NO: 19, VH CDR2 comprises the amino acid sequence of SEQ ID NO: 20, and VH CDR3 comprises the amino acid sequence of SEQ ID NO: 21.

[0063] In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment comprises VL CDR1, VL CDR2, and VL CDR3; wherein, VL CDR1 comprises the amino acid sequence of SEQ ID NO:22, VL CDR2 comprises the amino acid sequence of SEQ ID NO:23, and VL CDR3 comprises the amino acid sequence of SEQ ID NO:24.

[0064] In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment comprises VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2, and VL CDR3; wherein VH CDR1 comprises the amino acid sequence of SEQ ID NO:19, VH CDR2 comprises the amino acid sequence of SEQ ID NO:20, VH CDR3 comprises the amino acid sequence of SEQ ID NO:21, VL CDR1 comprises the amino acid sequence of SEQ ID NO:22, VL CDR2 comprises the amino acid sequence of SEQ ID NO:23, and VL CDR3 comprises the amino acid sequence of SEQ ID NO:24.

[0065] In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment comprises a heavy chain variable region and a light chain variable region; wherein the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:25, or an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO:25, or an amino acid sequence that has one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO:25; and / or the light chain variable region comprises the amino acid sequence of SEQ ID NO:26, or an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO:26, or an amino acid sequence that has one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO:26.

[0066] In some embodiments, the heavy chain variable region of the anti-CTLA-4 antibody or antigen-binding fragment comprises the amino acid sequence of SEQ ID NO:25, and the light chain variable region of the anti-CTLA-4 antibody or antigen-binding fragment comprises the amino acid sequence of SEQ ID NO:26.

[0067] In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment further comprises a heavy chain constant region and a light chain constant region.

[0068] In some embodiments, the heavy chain constant region of the anti-CTLA-4 antibody or antigen-binding fragment comprises the amino acid sequence of SEQ ID NO:27, or an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO:27, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO:27; and / or the light chain constant region of the anti-CTLA-4 antibody or antigen-binding fragment comprises the amino acid sequence of SEQ ID NO:14, or an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO:14, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO:14.

[0069] In some embodiments, the heavy chain constant region of the anti-CTLA-4 antibody or antigen-binding fragment comprises the amino acid sequence of SEQ ID NO:27, and the light chain constant region of the anti-CTLA-4 antibody or antigen-binding fragment comprises the amino acid sequence of SEQ ID NO:14.

[0070] In some embodiments, the anti-CTLA-4 antibody comprises a heavy chain and a light chain.

[0071] In some embodiments, the heavy chain of the anti-CTLA-4 antibody comprises the amino acid sequence of SEQ ID NO:28, or an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO:28, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO:28.

[0072] In some embodiments, the light chain of the anti-CTLA-4 antibody comprises the amino acid sequence of SEQ ID NO:29, or an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO:29, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO:29.

[0073] In some embodiments, the heavy chain of the anti-CTLA-4 antibody comprises the amino acid sequence of SEQ ID NO:28, or an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO:28, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO:28; and the light chain of the anti-CTLA-4 antibody comprises the amino acid sequence of SEQ ID NO:29, or an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO:29, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO:29.

[0074] In some embodiments, the anti-CTLA-4 antibody is Antibody 1 or Antibody 2, whose heavy chain comprises the amino acid sequence of SEQ ID NO: 28, and whose light chain comprises the amino acid sequence of SEQ ID NO: 29. In some embodiments, the anti-CTLA-4 antibody contains two identical heavy chains and two identical light chains.

[0075] In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment is expressed by CHO cells. The anti-CTLA-4 antibody or antigen-binding fragment can also be expressed by other cells (e.g., HEK cells (such as HEK293F cells), BHK cells, Cos1 cells, Cos7 cells, CV1 cells or mouse L cells). In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment is expressed by cells in which the α-(1,6)-fucosyltransferase gene (Fut-8) is knocked out. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment is expressed by CHO cells in which the α-(1,6)-fucosyltransferase gene (Fut-8) is knocked out (for construction methods, see patent CN109096399A).

[0076] In some embodiments, the total amount of high mannose glycoforms in the anti-CTLA-4 antibody (e.g., Antibody 1 or 2) or antigen-binding fragment is <5% and / or the total amount of sialylated glycoforms is <3%. In some embodiments, the total amount of high mannose glycoforms in the anti-CTLA-4 antibody (e.g., Antibody 1 or 2) or antigen-binding fragment is about 0.1%, about 0.3%, about 0.9%, about 1.2%, about 1.7%, about 2.6%, about 3.3%, about 4.1%, about 4.9%, about 5%, or a range between any two values ​​(including the endpoints) or any value therein. In some embodiments, the total amount of sialylated glycoforms in the anti-CTLA-4 antibody (e.g., Antibody 1 or 2) or antigen-binding fragment is about 0.1%, about 0.2%, about 0.4%, about 0.8%, about 1.5%, about 2.2%, about 2.7%, about 2.9%, 3%, or a range between any two values ​​(including the endpoints) or any value therein. In some embodiments, the total amount of high mannose glycoforms in the anti-CTLA-4 antibody (e.g., Antibody 1 or 2) or antigen-binding fragment is <3% and / or the total amount of sialylated glycoforms is <2%. In some embodiments, the total amount of high mannose glycoforms in the anti-CTLA-4 antibody (e.g., Antibody 1 or 2) or antigen-binding fragment is <2% and / or the total amount of sialylated glycoforms is <1%.

[0077] In some embodiments, the anti-CTLA-4 antibody (e.g., Antibody 1) or antigen-binding fragment has a fucosylation level of ≤5%, such as about 0, about 0.1%, about 0.3%, about 0.4%, about 0.6%, about 1.3%, about 1.9%, about 2.2%, about 2.8%, about 3.3%, about 3.7%, about 4.1%, about 4.5%, about 5%, or a range (including endpoints) between any two of these values ​​or any value therein. In some embodiments, the anti-CTLA-4 antibody (e.g., Antibody 1) or antigen-binding fragment has a fucosylation level of about 0. In some embodiments, the anti-CTLA-4 antibody (e.g., Antibody 1) or antigen-binding fragment has an enhanced ADCC effect (antibody-dependent cell-mediated cytotoxicity).

[0078] In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment is an anti-CTLA-4 antibody or antigen-binding fragment disclosed in WO2021129775A1.

[0079] In some embodiments, the antibody or antigen-binding fragment can be purified by conventional methods, such as centrifuging the cell suspension and collecting the supernatant, followed by further centrifugation to further remove impurities. Protein A affinity columns and ion exchange columns can be used to purify antibodies.

[0080] In some embodiments, the pharmaceutical composition is a pharmaceutical composition suitable for injection, such as a push-in pharmaceutical composition or an infusion (drip) pharmaceutical composition. In some embodiments, the pharmaceutical composition contains at least 0.1% of an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment. In some embodiments, the pharmaceutical composition contains at least 0.1% of an anti-CTLA-4 antibody or antigen-binding fragment. The percentage of the antibody can vary and can be between about 2% and about 90% of the weight of a given dosage form. The antibody or antigen-binding fragment in such a therapeutically useful pharmaceutical composition can be an effective amount for administration.

[0081] In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is administered at a dose of about 0.01 mg / kg to 200 mg / kg, about 0.1 mg / kg to 100 mg / kg, about 1 mg / kg to 60 mg / kg, or a formulation containing such a dose of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is administered at a dose of about 0.01 mg / kg, about 0.05 mg / kg, about 0.1 mg / kg, about 1 mg / kg, about 5 mg / kg, about 10 mg / kg, about 20 mg / kg, about 30 mg / kg, about 35 mg / kg, about 40 mg / kg, about 45 mg / kg, about 50 mg / kg, about 60 mg / kg, about 80 mg / kg, about 100 mg / kg, about 120 mg / kg, about 200 mg / kg, or a range (including endpoints) between any two of these values ​​or any value therein, or a formulation containing such a dose of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody is antibody BsAb-71-N297A.

[0082] In some embodiments, the dose of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment administered per treatment cycle is about 0.01 mg / kg to 200 mg / kg, about 0.1 mg / kg to 100 mg / kg, about 1 mg / kg to 60 mg / kg, or a formulation containing such a dose of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment. In some embodiments, the dose of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment administered per treatment cycle is about 0.01 mg / kg, about 0.05 mg / kg, about 0.1 mg / kg, about 1 mg / kg, about 5 mg / kg, about 10 mg / kg, about 20 mg / kg, about 30 mg / kg, about 35 mg / kg, about 40 mg / kg, about 45 mg / kg, about 50 mg / kg, about 60 mg / kg, about 80 mg / kg, about 100 mg / kg, about 120 mg / kg, about 200 mg / kg, or a range between any two of these values ​​(including the endpoints) or any value therein, or a formulation containing such a dose of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment. In some embodiments, one treatment cycle is about 1 week, about 2 weeks, about 3 weeks, about 4 weeks, about 5 weeks, about 6 weeks, about 2 months, about 5 months, about 8 months, about 1 year, or a range (including endpoints) between any two of these values ​​or any value therein. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody is antibody BsAb-71-N297A.

[0083] In some embodiments, the dose of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment administered each time is about 0.01 mg / kg to 200 mg / kg, about 0.1 mg / kg to 100 mg / kg, about 1 mg / kg to 60 mg / kg, or a formulation containing such a dose of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment. In some embodiments, the dose of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment administered each time is about 0.01 mg / kg, about 0.05 mg / kg, about 0.1 mg / kg, about 1 mg / kg, about 5 mg / kg, about 10 mg / kg, about 20 mg / kg, about 30 mg / kg, about 35 mg / kg, about 40 mg / kg, about 45 mg / kg, about 50 mg / kg, about 60 mg / kg, about 80 mg / kg, about 100 mg / kg, about 120 mg / kg, about 200 mg / kg, or a range (including endpoints) between any two of these values ​​or any value therein, or a formulation containing such a dose of an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment. In some embodiments, the dose is administered once a day to once every seven weeks. In some embodiments, the dose is administered once every two days to once every six weeks. In some embodiments, the dose is administered once every two weeks to four weeks. In some embodiments, the drug is administered about twice a week, or once about a week, about two weeks, about three weeks, about four weeks, about five weeks, about six weeks, or about seven weeks. In some embodiments, the drug is administered once a day, once every two days, once every three days, once every four days, once every five days, twice a week, once a week, once every two weeks, once every three weeks, once every four weeks, once every five weeks, once every six weeks, or once every seven weeks. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody is antibody BsAb-71-N297A.

[0084] In some embodiments, about 0.01 mg / kg to 200 mg / kg, about 0.1 mg / kg to 100 mg / kg, about 1 mg / kg to 60 mg / kg of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment, or a formulation containing such a dose of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment, is administered once every three weeks. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is administered once every three weeks at about 0.01 mg / kg, about 0.05 mg / kg, about 0.1 mg / kg, about 1 mg / kg, about 5 mg / kg, about 10 mg / kg, about 20 mg / kg, about 30 mg / kg, about 35 mg / kg, about 40 mg / kg, about 45 mg / kg, about 50 mg / kg, about 60 mg / kg, about 80 mg / kg, about 100 mg / kg, about 120 mg / kg, about 200 mg / kg, or a range (including endpoints) between any two of these values ​​or any value therein, or a formulation containing such a dose of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody is antibody BsAb-71-N297A.

[0085] In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment is administered at a dose of about 0.01 mg / kg to 100 mg / kg, about 0.1 mg / kg to 50 mg / kg, about 0.1 mg / kg to 20 mg / kg, about 0.1 mg / kg to 10 mg / kg, about 0.5 mg / kg to 6 mg / kg, or a formulation containing such a dose of the anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment is administered at a dose of about 0.01 mg / kg, about 0.1 mg / kg, about 0.5 mg / kg, about 1 mg / kg, about 2 mg / kg, about 3 mg / kg, about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 10 mg / kg, about 20 mg / kg, about 50 mg / kg, about 100 mg / kg, or a range (including endpoints) between any two of these values ​​or any value therein, or a formulation containing such a dose of the anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, the anti-CTLA-4 antibody is Antibody 1 or Antibody 2.

[0086] In some embodiments, the dose of the anti-CTLA-4 antibody or antigen-binding fragment administered per treatment cycle is about 0.01 mg / kg to 100 mg / kg, about 0.1 mg / kg to 50 mg / kg, about 0.1 mg / kg to 20 mg / kg, about 0.1 mg / kg to 10 mg / kg, about 0.5 mg / kg to 6 mg / kg, or a formulation containing such a dose of the anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, the dose of the anti-CTLA-4 antibody or antigen-binding fragment administered per treatment cycle is about 0.01 mg / kg, about 0.1 mg / kg, about 0.5 mg / kg, about 1 mg / kg, about 2 mg / kg, about 3 mg / kg, about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 10 mg / kg, about 20 mg / kg, about 50 mg / kg, about 100 mg / kg, or a range (including endpoints) between any two of these values ​​or any value therein, or a formulation containing such a dose of the anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, a treatment cycle is about 1 week, about 2 weeks, about 3 weeks, about 4 weeks, about 5 weeks, about 6 weeks, about 2 months, about 5 months, about 8 months, about 1 year, or a range (including endpoints) between any two of these values ​​or any value therein. In some embodiments, the anti-CTLA-4 antibody is Antibody 1 or Antibody 2.

[0087] In some embodiments, the dose of the anti-CTLA-4 antibody or antigen-binding fragment administered per administration is about 0.01 mg / kg to 100 mg / kg, about 0.1 mg / kg to 50 mg / kg, about 0.1 mg / kg to 20 mg / kg, about 0.1 mg / kg to 10 mg / kg, about 0.5 mg / kg to 6 mg / kg, or a formulation containing such a dose of the anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, the dose of the anti-CTLA-4 antibody or antigen-binding fragment administered per administration is about 0.01 mg / kg, about 0.1 mg / kg, about 0.5 mg / kg, about 1 mg / kg, about 2 mg / kg, about 3 mg / kg, about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 10 mg / kg, about 20 mg / kg, about 50 mg / kg, about 100 mg / kg, or a range (including endpoints) between any two of these values ​​or any value therein, or a formulation containing such a dose of the anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, the dose is administered once a day to once every seven weeks. In some embodiments, the dose is administered once every two days to once every six weeks. In some embodiments, the dose is administered once every two weeks to once every four weeks. In some embodiments, the dose is administered about twice a week, or about once a week, about two weeks, about three weeks, about four weeks, about five weeks, about six weeks, or about seven weeks. In some embodiments, the administration is once a day, once every two days, once every three days, once every four days, once every five days, twice a week, once a week, once every two weeks, once every three weeks, once every four weeks, once every five weeks, once every six weeks, or once every seven weeks. In some embodiments, the anti-CTLA-4 antibody is Antibody 1 or Antibody 2.

[0088] In some embodiments, about 0.01 mg / kg to 100 mg / kg, about 0.1 mg / kg to 50 mg / kg, about 0.1 mg / kg to 20 mg / kg, about 0.1 mg / kg to 10 mg / kg, about 0.5 mg / kg to 6 mg / kg, or a formulation containing such a dose of the anti-CTLA-4 antibody or antigen-binding fragment is administered once every three weeks. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment is administered once every three weeks at about 0.01 mg / kg, about 0.1 mg / kg, about 0.5 mg / kg, about 1 mg / kg, about 2 mg / kg, about 3 mg / kg, about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 10 mg / kg, about 20 mg / kg, about 50 mg / kg, about 100 mg / kg, or a range (including endpoints) between any two of these values, or any value therein, or a formulation containing such a dose of the anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, the anti-CTLA-4 antibody is Antibody 1 or Antibody 2.

[0089] In some embodiments, the present invention provides a method for treating a tumor, comprising: administering to a patient in need thereof about 0.01 mg / kg to 200 mg / kg, about 0.1 mg / kg to 100 mg / kg, about 1 mg / kg to 60 mg / kg, such as about 0.01 mg / kg, about 0.05 mg / kg, about 0.1 mg / kg, about 1 mg / kg, about 5 mg / kg, about 10 mg / kg, about 20 mg / kg, about 30 mg / kg, about 35 mg / kg, about 40 mg / kg, about 45 mg / kg, about 50 mg / kg, about 60 mg / kg, about 80 mg / kg, about 100 mg / kg, about 120 mg / kg, about 200 mg / kg of an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment, wherein the dose of anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is about 0.01 mg / kg to 200 mg / kg, about 0.05 mg / kg, about 0.1 mg / kg, about 1 mg / kg, about 5 mg / kg, about 10 mg / kg, about 20 mg / kg, about 30 mg / kg, about 35 mg / kg, about 40 mg / kg, about 45 mg / kg, about 50 mg / kg, about 60 mg / kg, about 80 mg / kg, about 100 mg / kg, about 120 mg / kg, about 200 mg / kg. A formulation of a D-L1 / CD47 bispecific antibody or antigen-binding fragment thereof; and further administering to the patient about 0.01 mg / kg to 100 mg / kg, about 0.1 mg / kg to 50 mg / kg, about 0.1 mg / kg to 20 mg / kg, about 0.1 mg / kg to 10 mg / kg, about 0.5 mg / kg to 6 mg / kg, such as about 0.01 mg / kg, about 0.1 mg / kg, about 0.5 mg / kg, about 1 mg / kg, about 2 mg / kg, about 3 mg / kg, about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 10 mg / kg, about 20 mg / kg, about 50 mg / kg, about 100 mg / kg of an anti-CTLA-4 antibody or antigen-binding fragment thereof. In some embodiments, the dosage is administered from once daily to once every seven weeks. In some embodiments, the dosage is administered from once every two days to once every six weeks. In some embodiments, the drug is administered once every two weeks to four weeks. In some embodiments, the drug is administered twice a week, or once every one week, about two weeks, about three weeks, about four weeks, about five weeks, about six weeks, or about seven weeks. In some embodiments, the drug is administered once a day, once every two days, once every three days, once every four days, once every five days, twice a week, once a week, once every two weeks, once every three weeks, once every four weeks, once every five weeks, once every six weeks, or once every seven weeks. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody is antibody BsAb-71-N297A. In some embodiments, the anti-CTLA-4 antibody is antibody 1.

[0090] In some embodiments, the present invention provides a method for treating a tumor, comprising: administering to a patient in need thereof about 0.01 mg / kg to 200 mg / kg, about 0.1 mg / kg to 100 mg / kg, about 1 mg / kg to 60 mg / kg, such as about 0.01 mg / kg, about 0.05 mg / kg, about 0.1 mg / kg, about 1 mg / kg, about 5 mg / kg, about 10 mg / kg, about 20 mg / kg, about 30 mg / kg, about 35 mg / kg, about 40 mg / kg, about 45 mg / kg, about 50 mg / kg, about 60 mg / kg, about 80 mg / kg, about 100 mg / kg, about 120 mg / kg, about 200 mg / kg of an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment once every three weeks, wherein the dose of anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is about 0.01 mg / kg to 200 mg / kg, about 0.1 mg / kg to 100 mg / kg, about 1 mg / kg, about 60 mg / kg, about 80 mg / kg, about 100 mg / kg, about 120 mg / kg, about 200 mg / kg. formulations of D-L1 / CD47 bispecific antibodies or antigen-binding fragments; and also administering to the patient about 0.01 mg / kg to 100 mg / kg, about 0.1 mg / kg to 50 mg / kg, about 0.1 mg / kg to 20 mg / kg, about 0.1 mg / kg to 10 mg / kg, about 0.5 mg / kg to 6 mg / kg, such as about 0.01 mg / kg, about 0.1 mg / kg, about 0.5 mg / kg, about 1 mg / kg, about 2 mg / kg, about 3 mg / kg, about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 10 mg / kg, about 20 mg / kg, about 50 mg / kg, about 100 mg / kg of an anti-CTLA-4 antibody or antigen-binding fragment once every three weeks. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody is antibody BsAb-71-N297A. In some embodiments, the anti-CTLA-4 antibody is Antibody 1.

[0091] In some embodiments, the present invention provides a method for treating tumors, comprising: administering to a patient in need thereof about 40 mg / kg of an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment, or a formulation comprising such a dose, once every three weeks; and further administering to the patient about 0.5 mg / kg, about 1 mg / kg, about 2 mg / kg, about 3 mg / kg, about 4 mg / kg, about 5 mg / kg, or about 6 mg / kg of an anti-CTLA-4 antibody or antigen-binding fragment, or a formulation comprising such a dose, once every three weeks. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody is antibody BsAb-71-N297A. In some embodiments, the anti-CTLA-4 antibody is Antibody 1.

[0092] In some embodiments, the present invention provides a method for treating a tumor, comprising: administering about 40 mg / kg of antibody BsAb-71-N297A to a patient in need thereof once every three weeks; and further administering about 0.5 mg / kg of Antibody 1 to the patient once every three weeks.

[0093] In some embodiments, the present invention provides a method for treating a tumor, comprising: administering about 40 mg / kg of antibody BsAb-71-N297A to a patient in need thereof once every three weeks; and further administering about 1 mg / kg of Antibody 1 to the patient once every three weeks.

[0094] In some embodiments, the present invention provides a method for treating a tumor, comprising: administering about 40 mg / kg of antibody BsAb-71-N297A to a patient in need thereof once every three weeks; and further administering about 2 mg / kg of Antibody 1 to the patient once every three weeks.

[0095] In some embodiments, the present invention provides a method for treating a tumor, comprising: administering about 40 mg / kg of antibody BsAb-71-N297A to a patient in need thereof once every three weeks; and further administering about 3 mg / kg of Antibody 1 to the patient once every three weeks.

[0096] In some embodiments, the present invention provides a method for treating a tumor, comprising: administering about 40 mg / kg of antibody BsAb-71-N297A to a patient in need thereof once every three weeks; and further administering about 6 mg / kg of Antibody 1 to the patient once every three weeks.

[0097] In some embodiments, a patient receives a single dose of an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, after a single dose, the patient's symptoms are alleviated. In some embodiments, after a single dose, if the patient's symptoms are not alleviated as expected, the patient is administered an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, the patient receives treatment until the condition is alleviated and no longer requires treatment. In some embodiments, the patient is administered an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment once per treatment cycle. In some embodiments, the patient is administered an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment multiple times per treatment cycle, for example, 2, 3, 4, or 5 times. In some embodiments, the patient receives treatment for one treatment cycle. In some embodiments, the patient receives treatment for multiple cycles (eg, at least 2, 3, or 4).

[0098] In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and / or anti-CTLA-4 antibody or antigen-binding fragment is administered by injection. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and / or anti-CTLA-4 antibody or antigen-binding fragment is administered by subcutaneous (sc) injection, intraperitoneal (ip) injection, parenteral injection, intraarterial injection or intravenous (iv) injection or infusion. The anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment can be administered in the same or different ways. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and / or anti-CTLA-4 antibody or antigen-binding fragment is administered by intravenous injection, such as intravenous infusion.

[0099] In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are administered as separate dosing units and are administered in combination. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are administered at intervals in time. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment is administered before the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment is administered after the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is administered. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are administered simultaneously. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are formulated into pharmaceutical compositions, respectively, and administered to the patient in a form suitable for the selected route of administration, such as parenteral, intravenous (iv), intramuscular, topical or subcutaneous routes.

[0100] In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment simultaneously form a combined administration unit and are administered in combination. In some embodiments, the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are combined to form a pharmaceutical composition and administered to the patient in a form suitable for the selected route of administration, such as parenteral, intravenous (iv), intramuscular, topical or subcutaneous routes.

[0101] In some embodiments, anti-PD-L1 / CD47 bispecific antibodies or antigen-binding fragments (or preparations), anti-CTLA-4 antibodies or antigen-binding fragments (or preparations) can be used in combination with other treatment methods for treating tumors, such as chemotherapy, radiotherapy, and surgical treatment.

[0102] In some embodiments, tumors include but are not limited to benign tumors, cancers. In some embodiments, tumors include but are not limited to blood tumors, solid tumors. In some embodiments, blood tumors include but are not limited to leukemia, lymphoma and myeloma. In some embodiments, blood tumors include but are not limited to acute lymphocytic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), chronic myeloid leukemia (CML), myeloproliferative disease / tumor (MPDS), Hodgkin lymphoma, non-Hodgkin lymphoma, Burkitt lymphoma, follicular lymphoma (small cell and large cell), multiple myeloma (MM), giant cell myeloma, heavy chain myeloma, light chain or Bence-Jones myeloma. In some embodiments, solid tumors include, but are not limited to, breast cancer, pancreatic cancer, prostate cancer, melanoma, head and neck cancer, liver cancer (such as hepatocellular carcinoma (HCC)), kidney cancer, squamous cell carcinoma (such as esophageal squamous cell carcinoma), lung cancer (such as non-small cell lung cancer (NSCLC)), cervical cancer, endometrial cancer, ovarian cancer, colorectal cancer, urothelial cancer, bladder cancer, brain cancer, gastric cancer, thyroid cancer, glioma, salivary gland cancer, leiomyosarcoma, thymic cancer. In some embodiments, the tumor is non-small cell lung cancer. In some embodiments, the tumor is locally advanced or metastatic non-small cell lung cancer. In some embodiments, the tumor is advanced non-small cell lung cancer.

[0103] In some embodiments, the patient is a patient with an advanced malignant tumor confirmed by cytology or pathology who has failed standard treatment.

[0104] In some embodiments, the patient is a patient with an advanced malignant tumor confirmed by cytology or pathology and for whom no standard treatment is available.

[0105] In some embodiments, the patient is a patient with an advanced malignancy confirmed by cytology or pathology who is intolerant to standard treatment.

[0106] In some embodiments, the patient is a patient with a cytologically or pathologically confirmed advanced malignancy who has refused standard treatment.

[0107] In some embodiments, the patient is a patient with histologically or cytologically confirmed locally advanced or metastatic (referring to the AJCC eighth edition staging criteria) NSCLC.

[0108] In some embodiments, the patient has disease progression after frontline treatment with a PD-1 / PD-L1 antibody and platinum-containing doublet chemotherapy or has been unable to receive standard treatment.

[0109] In some embodiments, the patient has a driver gene mutation and has failed targeted therapy and chemotherapy or is unable to receive standard treatment.

[0110] In some embodiments, the patient is a patient with histologically or cytologically confirmed advanced or metastatic solid tumors who have failed standard treatment, have no standard treatment, or are intolerant to standard treatment.

[0111] In some embodiments, the patient is a patient with histologically or cytologically confirmed advanced or metastatic hepatocellular carcinoma (liver function Child-Pugh grade A or better B, i.e., ≤7 points), renal clear cell carcinoma, cervical cancer, colorectal cancer, esophageal cancer, gastric or esophageal junction cancer, who has failed standard treatment, has no standard treatment, or is intolerant to standard treatment.

[0112] In some embodiments, the patient has at least one measurable tumor lesion according to RECIST 1.1.

[0113] In some embodiments, the patient with advanced malignant tumor has not previously received treatment with anti-CTLA-4, CD47, SIRPα antibodies or drugs targeting the same.

[0114] In some embodiments, the patient is a patient with at least two of the above. DETAILED DESCRIPTION

[0115] Unless otherwise stated, each of the following terms shall have the meaning set forth below.

[0116] definition

[0117] It should be noted that the term "a" entity refers to one or more of that entity, e.g., "an antibody" should be understood as one or more antibodies, and thus, the terms "a" (or "an"), "one or more" and "at least one" can be used interchangeably herein.

[0118] As used herein, the terms "comprising" or "including" mean that the antibody, composition, or method, etc. includes the listed elements, such as components or steps, but does not exclude others. "Essentially consisting of" means that the antibody, composition, or method, etc. excludes other elements that have a fundamental effect on the characteristics of the combination, but does not exclude elements that do not substantially affect the antibody, composition, or method, etc. "Consisting of" means excluding elements not specifically listed.

[0119] "About" refers to the normal error range of the corresponding numerical value that is easily known to those skilled in the relevant art. In some embodiments, "about" mentioned herein refers to the described numerical value and its ±10%, ±5% or ±1% range.

[0120] "Treatment" refers to both therapeutic treatment and prophylactic or preventative measures, the purpose of which is to prevent, slow, ameliorate, and halt an undesirable physiological change or disorder, such as the progression of a disease, including but not limited to the following results, whether detectable or undetectable, relief of symptoms, reduction in the extent of the disease, stabilization of the disease state (i.e., no worsening), delay or slowing of disease progression, improvement or alleviation of the disease state, alleviation or elimination (whether partial or total), prolongation of life expectancy compared to that expected in the absence of treatment, etc. Patients in need of treatment include those already suffering from a condition or disorder, those susceptible to a condition or disorder, or those in need of prevention of the condition or disorder, and those who can or are expected to benefit from the administration of the antibodies or compositions disclosed herein for detection, diagnostic procedures, and / or treatment.

[0121] The term "cancer" refers to or is intended to describe a physiological state in mammals that is typically characterized by uncontrolled cell growth. Examples of cancer include, but are not limited to, carcinoma, lymphoma, blastoma, sarcoma, or leukemia. More specific examples of such cancers include, but are not limited to, colorectal cancer, lung cancer, ovarian cancer, uterine cancer, endometrial cancer, salivary gland cancer, peritoneal cancer, fallopian tube cancer, pancreatic cancer, thyroid cancer, head and neck squamous cell carcinoma, nasopharyngeal cancer, laryngeal cancer, lung adenocarcinoma, lung squamous cell carcinoma, liver cancer, hepatocellular carcinoma, gastrointestinal cancer, glioblastoma, breast cancer, brain cancer, kidney cancer, renal cell carcinoma, colon cancer, rectal cancer, prostate cancer, vulvar cancer, testicular cancer, squamous cell carcinoma, small cell lung cancer, cervical cancer, cervical cancer, tongue cancer, bladder cancer, retinoblastoma, glioblastoma, mesothelioma, oral epithelioid carcinoma, choriocarcinoma, and head and neck cancer.

[0122] The term "in need thereof" refers to a patient who has been identified as being in need of a particular approach or treatment. In some embodiments, identification can be performed by any diagnostic means.

[0123] The term "patient" refers to any mammal in need of diagnosis, prognosis, or treatment, including humans, dogs, cats, guinea pigs, rabbits, rats, mice, horses, cows, and the like.

[0124] As used herein, the term "administration" or "application" refers to the administration of a substance to achieve a therapeutic purpose (e.g., to treat a tumor). The administration route can be parenteral, enteral, and topical. Parenteral administration is typically by injection, including but not limited to intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcutaneous, intraarticular, subcapsular, subarachnoid, intraspinal, and intrasternal injection and infusion.

[0125] An "effective amount" is an amount of an active compound or agent that elicits a biological or medicinal response in a tissue, system, animal or human.

[0126] The present invention is further described below by way of specific examples, which do not limit the scope of protection of the present invention. Non-essential modifications and adjustments made by others based on the concept of the present invention still fall within the scope of protection of the present invention.

[0127] Unless otherwise specified, the materials and reagents used in the following examples can be obtained from commercial sources.

[0128] Example 1: Preparation of antibodies

[0129] The DNA sequences encoding the antibody heavy and light chains are cloned into expression vectors, then transferred into host cells, cultured and purified to obtain antibodies.

[0130] The sequences of antibodies BsAb-71-N297A, Antibody 1, and Antibody 2 are shown in Tables 1 and 2. The anti-PD-L1 heavy chain a of antibody BsAb-71-N297A is shown in SEQ ID NO:16, the anti-CD47 heavy chain b is shown in SEQ ID NO:17, and the light chains a and b are shown in SEQ ID NO:18. Antibodies 1 and 2 share identical amino acid sequences, with their heavy chains shown in SEQ ID NO:28 and their light chains shown in SEQ ID NO:29. Antibodies BsAb-71-N297A and Antibody 2 were expressed in CHO cells; Antibody 1 was expressed in CHO cells with the α-(1,6)-fucosyltransferase gene (Fut-8) knockout (see patent CN109096399A for the construction method). Compared to Antibody 2, Antibody 1 has a predominantly defucosylated glycoform, with a fucosylation level of approximately 0, while Antibody 2 has a fucosylation content exceeding 70%. Both Antibody 1 and Antibody 2 have low levels of total high mannose glycoforms (<2%) and total sialylated glycoforms (<2%).

[0131] Table 1 Amino acid sequence of antibody BsAb-71-N297A

[0132] Table 2 Amino acid sequences of Antibody 1 and Antibody 2

[0133] Example 2: Clinical study of antibody BsAb-71-N297A combined with antibody 1

[0134] This study is a multicenter, open-label Phase Ib / II clinical study to evaluate the safety, tolerability, pharmacokinetic characteristics and preliminary clinical efficacy of antibody BsAb-71-N297A injection combined with antibody 1 injection in patients with advanced malignant tumors.

[0135] Study Objectives

[0136] Main Purpose:

[0137] Phase Ib study:

[0138] To evaluate the safety and tolerability of antibody BsAb-71-N297A injection combined with antibody 1 injection in patients with advanced malignancies;

[0139] Explore the maximum tolerated dose (MTD) of combined therapy and provide a recommended dose (RP2D) and a reasonable dosing regimen for Phase II or subsequent clinical studies;

[0140] Phase II study:

[0141] Evaluate the safety and tolerability of BsAb-71-N297A injection combined with Antibody 1 injection in patients with advanced non-small cell lung cancer and other advanced solid tumors, and determine an optimized RP2D and dosing schedule for subsequent studies;

[0142] To evaluate the preliminary efficacy of antibody BsAb-71-N297A injection combined with antibody 1 injection in advanced non-small cell lung cancer and other advanced solid tumors.

[0143] Secondary objectives:

[0144] Phase Ib study:

[0145] To preliminarily evaluate the efficacy of antibody BsAb-71-N297A injection combined with antibody 1 injection in the treatment of patients with advanced malignant tumors;

[0146] To evaluate the pharmacokinetic (PK) characteristics of antibody BsAb-71-N297A injection combined with antibody 1 injection after single and repeated administration in patients with advanced malignancies;

[0147] Evaluate the immunogenicity of antibody BsAb-71-N297A injection combined with antibody 1 injection;

[0148] Preliminary evaluation of the pharmacodynamic properties of antibody BsAb-71-N297A injection combined with antibody 1 injection.

[0149] Phase II study:

[0150] Further evaluate the pharmacokinetic (PK) characteristics of single and multiple-dose administration of antibody BsAb-71-N297A injection combined with antibody 1 injection in advanced non-small cell lung cancer and other advanced solid tumors;

[0151] Further evaluate the immunogenicity of antibody BsAb-71-N297A injection combined with antibody 1 injection;

[0152] Evaluate the pharmacodynamic properties of antibody BsAb-71-N297A injection combined with antibody 1 injection.

[0153] Study endpoints

[0154] Primary endpoint:

[0155] Phase Ib study:

[0156] Tolerability and safety endpoints: occurrence of dose-limiting toxicity (DLT), vital signs and physical examinations, various adverse events (AEs), clinical laboratory tests, and clinical auxiliary examinations (such as electrocardiograms);

[0157] Phase II study:

[0158] Safety assessment endpoints: adverse events (AEs), vital signs and physical examinations, clinical laboratory tests, and clinical auxiliary examinations (such as electrocardiograms);

[0159] Efficacy evaluation endpoint: objective response rate (ORR).

[0160] Secondary endpoints:

[0161] Phase Ib study:

[0162] Efficacy evaluation endpoints: ORR, progression-free survival (PFS), duration of response (DoR), disease control rate (DCR), and overall survival (OS) for advanced malignancies;

[0163] Pharmacokinetic parameters under single and multiple doses, including:

[0164] Single dose: C max 、T max 、T 1 / 2 , CL, Vd, Ke, MRT, AUC (0-τ) , AUC (0-∞) ;

[0165] Multiple doses: C max,ss 、C avg,ss 、C min,ss , AUC (0-τ)ss , AUC (0-∞)ss 、T max,ss 、T 1 / 2,ss 、

[0166] CL, V ss , Ke, MRT, accumulation index (R ac ), volatility index DF.

[0167] Immunogenicity evaluation indicators: anti-drug antibodies (ADA) / neutralizing antibodies (NAb);

[0168] Pharmacodynamic indicators: The relationship between the expression levels of PD-L1 and (or) CD47 in tumor tissue before treatment and the therapeutic effect.

[0169] Phase II study:

[0170] To evaluate PFS, DoR, DCR, and OS in advanced non-small cell lung cancer and other advanced solid tumors;

[0171] Pharmacokinetic parameters under single and repeated doses (same as in Phase Ib study);

[0172] Immunogenicity evaluation indicators (same as Phase Ib study);

[0173] Pharmacodynamic indicators (same as Phase Ib study).

[0174] Inclusion criteria

[0175] Patients must meet the following criteria to be included in this study:

[0176] 1. Age ≥ 18 years old;

[0177] 2. For different phases or cohorts, subjects must meet the following requirements:

[0178] 2-1) Phase Ib: Patients with advanced malignant tumors confirmed by cytology or pathology who have failed standard treatment, have no standard treatment, are intolerant to standard treatment, or refuse standard treatment;

[0179] 2-2) Phase II Cohort 1: Patients with histologically or cytologically confirmed locally advanced or metastatic NSCLC (referring to the eighth edition of the AJCC staging criteria):

[0180] a. Patients whose disease progresses after frontline treatment with PD-1 / PD-L1 antibodies and platinum-containing doublet chemotherapy or who are unable to receive standard treatment;

[0181] b. Patients with driver gene mutations who have failed targeted therapy and chemotherapy or are unconditionally eligible for standard treatment;

[0182] 2-3) Phase II Cohort 2: Patients with histologically or cytologically confirmed advanced or metastatic solid tumors who have failed standard treatment, have no standard treatment, or are intolerant to standard treatment. Only patients with hepatocellular carcinoma (Child-Pugh liver function grade A or better B, i.e., ≤7 points), renal clear cell carcinoma, cervical cancer, colorectal cancer, esophageal cancer, and gastric or esophageal junction cancer are included (Note: For hepatocellular carcinoma, patients with clinically diagnosed advanced disease are accepted);

[0183] 3. The presence of at least one measurable tumor lesion according to RECIST 1.1 (a tumor lesion previously treated with radiotherapy cannot be considered a measurable lesion unless the lesion has shown clear disease progression after radiotherapy);

[0184] 4. Eastern Cooperative Oncology Group (ECOG) performance status score of 0 or 1;

[0185] 5. The expected survival time as assessed by the investigator is ≥12 weeks;

[0186] 6. Adequate organ and bone marrow function, as defined below:

[0187] 7. Agree to provide archived pathological tissue or fresh biopsy tumor tissue for detection of PD-L1 and CD47 expression levels (not a necessary inclusion criterion);

[0188] 8. For female subjects of childbearing potential, a serum pregnancy test should be conducted within seven days before the first dose of the trial drug (Cycle 1, Day 1), and the result should be negative. Subjects must agree to use effective contraception or undergo surgical sterilization during the trial and within 90 days after the last dose of the trial drug;

[0189] 9. Able to comply with research and follow-up procedures;

[0190] 10. Sign written informed consent.

[0191] Exclusion criteria

[0192] Patients will be excluded from the study if they have the following conditions:

[0193] 1. Cohort 1: Patients who had received lung radiotherapy > 30 Gy within 6 months before enrollment;

[0194] Cohort 2: Patients whose hepatocellular carcinoma pathology suggests fibrolamellar carcinoma, sarcomatoid carcinoma, or mixed bile duct carcinoma; patients currently suffering from hepatic encephalopathy;

[0195] 2. Patients have previously received treatment with anti-CTLA-4, CD47, SIRPα antibodies or drugs with the same targets;

[0196] 3. Patients who have received chemotherapy, radiotherapy, biological therapy, endocrine therapy, immunotherapy or other anti-tumor treatment within 4 weeks before the first use of study drugs, excluding the following: ① Nitrosourea or mitomycin C within 6 weeks before the first use of study drugs; ② Oral fluorouracil and small molecule targeted drugs within 2 weeks before the first use of study drugs or 5 half-lives of the drug (whichever is longer); ③ Systemic treatment with traditional Chinese medicine / Chinese patent medicine with clear anti-tumor effects, immunomodulatory drugs (including but not limited to thymosin, interferon, interleukin, etc.) within 2 weeks before the first use of study drugs;

[0197] 4. Participants who are currently participating in or have participated in an interventional clinical study of an experimental drug or medical device within 4 weeks before the first dose of this study are not eligible for inclusion. During the follow-up phase of the interventional study, participants may be included if the time between the first dose and the end of the previous study (the last dose) meets the third exclusion criterion above.

[0198] 5. Vaccinated with live / attenuated or mRNA vaccine within 4 weeks before screening or planned to be vaccinated during the study period;

[0199] 6. Pregnant or breastfeeding women;

[0200] 7. Patients with primary central nervous system tumors, meningeal metastases, or symptomatic central nervous system metastases must be excluded. Patients with asymptomatic central nervous system metastases that have achieved clinical control, or those with symptoms but are judged by the investigator to be stable, may be included, but must meet the following conditions at the same time: a. The disease has been stable for ≥4 weeks before the first dose; b. No evidence of central nervous system disease progression has been found on plain / enhanced cranial MRI within 4 weeks before the first dose; c. Antiepileptic drugs have been discontinued for ≥2 weeks before the first dose, and prednisone dosage is ≤10 mg / day or equivalent dose of hormones;

[0201] 8. Patients who have undergone major surgical surgery on major organs (excluding puncture biopsy) within 4 weeks before the first use of the study drug or have not yet recovered after the surgery, or who need to undergo elective surgery during the trial;

[0202] 9. Those with a history of tissue or organ transplant surgery;

[0203] 10. Subjects who have had a serious infection as determined by the investigator within 4 weeks before the first dose of the drug or subjects who have an active infection within 2 weeks before the first dose of the drug;

[0204] 11. Known history of human immunodeficiency virus (HIV) infection;

[0205] 12. Untreated active hepatitis B must be excluded; Note: HBV viral load <1000IU / ml or <10 3Copies / ml can not be excluded (for patients with hepatocellular carcinoma, HBV viral load <2000IU / ml or <10 4 Copies / ml may not be excluded), but researchers need to consider giving anti-HBV treatment during the study; for subjects with anti-HBc(+), HBsAg(-), anti-HBs(-) and HBV viral load(-), routine preventive anti-HBV treatment is not required, but viral reactivation needs to be closely monitored;

[0206] 13. Subjects with active HCV infection (defined as: HCV antibody positive and HCV-RNA level above the detection limit);

[0207] 14. Subjects with tuberculosis who have not been treated or are currently being treated, including but not limited to pulmonary tuberculosis; subjects who have received standard anti-tuberculosis treatment and have been confirmed to be cured by the researchers may be included;

[0208] 15. Subjects with a known history of severe allergies, or subjects known to have had a grade ≥ 3 allergic reaction to large molecule protein preparations / antibodies;

[0209] 16. Patients with a history of autoimmune diseases (excluding vitiligo, thyroid diseases that can be treated with hormone replacement therapy, and type 1 diabetes);

[0210] 17. Received systemic glucocorticoids (prednisone > 10 mg / day or equivalent doses of similar drugs) or other immunosuppressive treatment within 14 days before the first use of study drug; excluding the following situations: use of topical, ocular, intra-articular, intranasal and inhaled glucocorticoids, and short-term use of glucocorticoids for prophylaxis (e.g., prevention of contrast agent allergy);

[0211] 18. Patients who have previously experienced grade ≥3 irAEs or grade ≥2 irAEs requiring permanent discontinuation of immunotherapy as specified in the CSCO Guidelines for the Management of Toxicity of Immune Checkpoint Inhibitors (2023 edition or later);

[0212] 19. Severe cardiovascular disease: New York Heart Association (NYHA) class III or higher heart failure, left ventricular ejection fraction (LVEF) <50%, unstable angina, uncontrolled hypertension (defined in this protocol as systolic blood pressure >160 mmHg and / or diastolic blood pressure >100 mmHg despite optimal antihypertensive therapy), a history of myocardial infarction within the past 6 months, or severe arrhythmias requiring medication (patients with atrial fibrillation or supraventricular tachycardia were included at the investigator's discretion);

[0213] 20. Subjects with a history of non-infectious pneumonia requiring glucocorticoid treatment within one year prior to enrollment, or currently experiencing or not excluding the presence of interstitial pneumonitis;

[0214] 21. Patients with a history of psychotropic drug abuse or drug use that is considered to affect compliance with this study;

[0215] 22. In addition to the tumor suffered at the time of enrollment in the study, other active malignant tumors existed within 3 years before the first dose (locally cured tumors such as basal cell carcinoma of the skin, superficial bladder cancer, or breast carcinoma in situ are not excluded);

[0216] 23. Patients with effusion in the pericardial cavity, pleural cavity, abdominal cavity, pelvic cavity, or other serous cavities who have recently required puncture, drainage, or other treatment (patients whose effusion has stabilized after recent puncture and drainage can be included);

[0217] 24. Patients who are deemed by the researcher to be unsuitable for participation in this study due to other conditions.

[0218] Overall study design

[0219] This study is designed as a multicenter, open-label Phase Ib / II clinical trial, primarily to evaluate the safety, tolerability, PK characteristics, and efficacy of antibody BsAb-71-N297A combined with Antibody 1 in patients with advanced solid tumors, explore the maximum tolerated dose (MTD) of the combined therapy, and provide a recommended dose (RP2D) and a reasonable dosing regimen for subsequent clinical studies.

[0220] 1. Phase Ib dose-escalation study

[0221] The "3+3" dose escalation rule was used to explore the safety and tolerability of antibody BsAb-71-N297A injection combined with antibody 1 injection in advanced solid tumors.

[0222] Five dose groups were set up for antibody 1, namely escalating groups A (0.5 mg / kg), B (1 mg / kg), C (2 mg / kg), D (3 mg / kg), and E (6 mg / kg). The antibody BsAb-71-N297A was administered at a fixed dose of 40 mg / kg, as shown below:

[0223] 2. Phase II dose expansion study

[0224] In the Phase II study, based on the preliminary safety and efficacy results of the Phase Ib study, two doses of Antibody 1 will be selected for combination with Antibody BsAb-71-N297A 40 mg / kg at a dosing frequency of once every three weeks (Q3W) for an extended study in locally advanced or metastatic non-small cell lung cancer and other advanced solid tumors. This will further explore the safety and efficacy of Antibody BsAb-71-N297A injection combined with Antibody 1 injection, providing a basis for subsequent clinical studies.

[0225] Dosage regimen

[0226] Antibody 1 dosing regimen: intravenous infusion, with an infusion time of 60±10 minutes in the Phase Ib study and a recommended infusion time of 30-60 minutes in the Phase II study, administered once every three weeks (Q3W).

[0227] Antibody BsAb-71-N297A dosing regimen: Intravenous infusion begins 30 (± 10) minutes after the completion of antibody 1 infusion. In the Phase Ib study, the infusion time was 60 ± 10 minutes. In the Phase II study, the recommended infusion time was 30 to 60 minutes. The drug was administered once every three weeks (Q3W).

[0228] DLT Definition

[0229] Adverse events (AEs) will be assessed based on CTCAE v5.0. DLTs are defined as AEs occurring within 21 days after the first dose and considered at least possibly related to the study drug. The specific definitions of DLTs for this study are as follows:

[0230] Level 5 AE.

[0231] Non-hematologic toxicities:

[0232] ① Non-hematological toxicity of grade 3 or higher (AEs not detected by simple laboratory tests);

[0233] ② Grade 3 or 4 non-hematological toxicity discovered by clinical laboratory tests that meets any of the following conditions:

[0234] 1) requiring clinical intervention; 2) judged as a SAE; 3) any treatment-related toxicity that causes a delay of more than 3 weeks in the second cycle of administration.

[0235] Hematological toxicity:

[0236] ① Grade 4 anemia or above or grade 3 anemia requiring red blood cell transfusion intervention;

[0237] ② Grade 4 thrombocytopenia persists for ≥7 days, or grade 3 thrombocytopenia is accompanied by bleeding (≥grade 2 bleeding) or requires platelet transfusion;

[0238] ③ Grade 4 neutropenia persists for ≥7 days after treatment; Grade 3 or higher neutropenia is accompanied by fever (≥38.0℃ for 1 hour or ≥38.3℃);

[0239] However, the following situations were not considered DLT in this study:

[0240] ① Lymphocytopenia ≥ grade 3;

[0241] ②Grade 3 endocrine toxicity can be controlled by hormone replacement therapy;

[0242] ③ Grade 3 or higher rash;

[0243] ④ Nausea, vomiting, and diarrhea of ​​grade 3 or higher can be relieved to grade 2 or lower within 72 hours after clinical intervention;

[0244] ⑤ Grade 3 or higher infusion reaction (without preventive medication) can be relieved to grade 2 or lower within 6 hours after clinical intervention;

[0245] ⑥ Fatigue of grade 3 or higher lasting for ≤7 days;

[0246] ⑦ A simple electrolyte abnormality of ≥ grade 3 is relieved to ≤ grade 2 within 72 hours and is not associated with other clinically significant adverse events;

[0247] ⑧Cancer pain of grade 3 or higher.

[0248] Safety evaluation

[0249] Including vital signs and physical examination, laboratory tests (blood routine, blood biochemistry, thyroid function, coagulation routine, troponin, BNP, urine routine, stool routine, pregnancy test), ECOG score, electrocardiogram, adverse events (including immune-related adverse events), etc.

[0250] Clinical effectiveness evaluation

[0251] Objective response rate (ORR), duration of response (DOR), disease control rate (DCR), progression-free survival (PFS), and overall survival (OS) as defined by RECIST 1.1;

[0252] ORR, DOR, DCR, and PFS according to iRECIST evaluation.

[0253] Pharmacodynamics / biomarker exploratory

[0254] Pharmacodynamic evaluation will be based on detecting the expression levels of PD-L1 and / or CD47 in tumor tissue before treatment to evaluate their correlation with clinical efficacy.

[0255] Pharmacokinetic evaluation

[0256] In the Phase Ib study, blood samples were collected at defined time points during treatment (first 6 cycles) and plasma concentrations (C trough Blood samples will be collected at each time point to measure serum drug concentrations and study pharmacokinetic (PK) characteristics. In each Phase II dose group, at least 10 subjects (regardless of tumor type) will undergo intensive PK blood sampling, with the remaining subjects undergoing sparse blood sampling.

[0257] PK parameters were calculated using the actual administered dose, actual sampling time, and a non-compartmental model.

[0258] Immunogenicity evaluation

[0259] The immunogenicity evaluation indicators are as follows:

[0260] The sample and individual positive rates of anti-drug antibodies (ADA);

[0261] ADA-positive samples will continue to be tested for neutralizing antibodies (NAb);

[0262] The sample and individual positive rates of ADA-positive and NAb-positive samples.

Claims

1. A pharmaceutical composition for treating tumors, comprising an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment.

2. A combination of an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment for treating tumors.

3. Use of an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment in the preparation of a pharmaceutical composition for treating tumors.

4. A method for treating a tumor, comprising: Administering an effective amount of an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment to a patient in need.

5. The pharmaceutical composition, combination, use, and method according to any one of claims 1-4, wherein the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment comprises a variable region a specifically binding to PD-L1 and a variable region b specifically binding to CD47; the variable region a comprises VHa CDR1 shown in SEQ ID NO:1, VHa CDR2 shown in SEQ ID NO:2, VHa CDR3 shown in SEQ ID NO:3, VLa CDR1 shown in SEQ ID NO:4, VLa CDR2 shown in SEQ ID NO:5, and VLa CDR3 shown in SEQ ID NO:6; the variable region b comprises VHb CDR1 shown in SEQ ID NO:7, VHb CDR2 shown in SEQ ID NO:8, VHb CDR3 shown in SEQ ID NO:9, VLb CDR1 shown in SEQ ID NO:4, VLb CDR2 shown in SEQ ID NO:5, and VLb CDR3 shown in SEQ ID NO:

6.

6. The pharmaceutical composition, combination, use, and method according to claim 5, wherein the variable region a comprises a heavy chain variable region VHa and a light chain variable region VLa; the VHa comprises the amino acid sequence shown in SEQ ID NO:10, or an amino acid sequence having at least 80% identity compared to the amino acid sequence shown in SEQ ID NO:10, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:10; the VLa comprises the amino acid sequence shown in SEQ ID NO:12, or an amino acid sequence having at least 80% identity compared to the amino acid sequence shown in SEQ ID NO:12, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:

12.

7. The pharmaceutical composition, combination, use and method according to claim 5 or 6, wherein the variable region b comprises a heavy chain variable region VHb and a light chain variable region VLb; the VHb comprises the amino acid sequence shown in SEQ ID NO: 11, or an amino acid sequence having at least 80% identity compared to the amino acid sequence shown in SEQ ID NO: 11, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO: 11; the VLb comprises the amino acid sequence shown in SEQ ID NO: 12, or an amino acid sequence having at least 80% identity compared to the amino acid sequence shown in SEQ ID NO: 12, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:

12.

8. The pharmaceutical composition, combination, use and method according to any one of claims 1-7, wherein the anti-PD-L1 / CD47 bispecific antibody comprises heavy chain a, heavy chain b, light chain a and light chain b; the heavy chain a comprises the amino acid sequence shown in SEQ ID NO: 16, or an amino acid sequence having at least 80% identity compared to the amino acid sequence shown in SEQ ID NO: 16, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO: 16; the heavy chain b comprises the amino acid sequence shown in SEQ ID NO: 17, or an amino acid sequence having at least 80% identity compared to the amino acid sequence shown in SEQ ID NO: 17, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO: 17; the light chain a and light chain b comprise the same amino acid sequence; or, the light chain a and light chain b comprise the amino acid sequence shown in SEQ ID NO: 18, or an amino acid sequence having at least 80% identity compared to the amino acid sequence shown in SEQ ID NO: 18, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:

18.

9. The pharmaceutical composition, combination, use and method according to any one of claims 1-8, wherein the anti-PD-L1 / CD47 bispecific antibody comprises heavy chain a, heavy chain b, light chain a and light chain b; the heavy chain a comprises the amino acid sequence shown in SEQ ID NO: 16, the heavy chain b comprises the amino acid sequence shown in SEQ ID NO: 17, and the light chain a and light chain b comprise the amino acid sequence shown in SEQ ID NO:

18.

10. The pharmaceutical composition, combination, use and method according to any one of claims 1-9, wherein the anti-CTLA-4 antibody or antigen-binding fragment comprises a VH CDR1 shown in SEQ ID NO:19, a VH CDR2 shown in SEQ ID NO:20, a VH CDR3 shown in SEQ ID NO:21, a VL CDR1 shown in SEQ ID NO:22, a VL CDR2 shown in SEQ ID NO:23 and a VL CDR3 shown in SEQ ID NO:

24.

11. The pharmaceutical composition, combination, use and method according to any one of claims 1-10, wherein the anti-CTLA-4 antibody or antigen-binding fragment comprises a heavy chain variable region and a light chain variable region; the heavy chain variable region of the anti-CTLA-4 antibody or antigen-binding fragment comprises the amino acid sequence shown in SEQ ID NO:25, or an amino acid sequence having at least 80% identity compared to the amino acid sequence shown in SEQ ID NO:25, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:25; the light chain variable region of the anti-CTLA-4 antibody or antigen-binding fragment comprises the amino acid sequence shown in SEQ ID NO:26, or an amino acid sequence having at least 80% identity compared to the amino acid sequence shown in SEQ ID NO:26, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:

26.

12. The pharmaceutical composition, combination, use and method according to any one of claims 1-11, wherein the anti-CTLA-4 antibody comprises a heavy chain and a light chain; the heavy chain of the anti-CTLA-4 antibody comprises the amino acid sequence shown in SEQ ID NO:28, or an amino acid sequence having at least 80% identity compared to the amino acid sequence shown in SEQ ID NO:28, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:28; the light chain of the anti-CTLA-4 antibody comprises the amino acid sequence shown in SEQ ID NO:29, or an amino acid sequence having at least 80% identity compared to the amino acid sequence shown in SEQ ID NO:29, or an amino acid sequence having one or more conservative amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:

29.

13. The pharmaceutical composition, combination, use and method according to any one of claims 1-12, wherein the fucosylation level of the anti-CTLA-4 antibody or antigen-binding fragment is 0-5%.

14. The pharmaceutical composition, combination, use and method according to claim 13, wherein the anti-CTLA-4 antibody or antigen-binding fragment is expressed by cells with α-(1,6)-fucosyltransferase gene knockout.

15. The pharmaceutical composition, combination, use and method according to claim 13, wherein the anti-CTLA-4 antibody or antigen-binding fragment is expressed by CHO cells with knockout of the α-(1,6)-fucosyltransferase gene.

16. The pharmaceutical composition, combination, use and method according to any one of claims 1-15, wherein the dosage of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is 0.01-200 mg / kg.

17. The pharmaceutical composition, combination, use and method according to any one of claims 1-15, wherein the dosage of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is 0.1-100 mg / kg.

18. The pharmaceutical composition, combination, use and method according to any one of claims 1-15, wherein the dosage of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is 1-60 mg / kg.

19. The pharmaceutical composition, combination, use and method according to any one of claims 1-15, wherein the dosage of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is about 40 mg / kg.

20. The pharmaceutical composition, combination, use and method according to any one of claims 1-19, wherein the dosage of the anti-CTLA-4 antibody or antigen-binding fragment is 0.01-100 mg / kg.

21. The pharmaceutical composition, combination, use and method according to any one of claims 1-19, wherein the dosage of the anti-CTLA-4 antibody or antigen-binding fragment is 0.1-50 mg / kg.

22. The pharmaceutical composition, combination, use and method according to any one of claims 1-19, wherein the dosage of the anti-CTLA-4 antibody or antigen-binding fragment is 0.1-20 mg / kg.

23. The pharmaceutical composition, combination, use and method according to any one of claims 1-19, wherein the dosage of the anti-CTLA-4 antibody or antigen-binding fragment is 0.1-10 mg / kg.

24. The pharmaceutical composition, combination, use and method according to any one of claims 1-19, wherein the dosage of the anti-CTLA-4 antibody or antigen-binding fragment is 0.5-6 mg / kg.

25. The pharmaceutical composition, combination, use and method according to any one of claims 1-19, wherein the dosage of the anti-CTLA-4 antibody or antigen-binding fragment is about 0.5 mg / kg.

26. The pharmaceutical composition, combination, use and method according to any one of claims 1-19, wherein the dosage of the anti-CTLA-4 antibody or antigen-binding fragment is about 1 mg / kg.

27. The pharmaceutical composition, combination, use and method according to any one of claims 1-19, wherein the dosage of the anti-CTLA-4 antibody or antigen-binding fragment is about 2 mg / kg.

28. The pharmaceutical composition, combination, use and method according to any one of claims 1-19, wherein the administration dose of the anti-CTLA-4 antibody or antigen-binding fragment is about 3 mg / kg.

29. The pharmaceutical composition, combination, use and method according to any one of claims 1-19, wherein the administration dose of the anti-CTLA-4 antibody or antigen-binding fragment is about 6 mg / kg.

30. The pharmaceutical composition, combination, use and method according to any one of claims 1-29, wherein the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is administered once a week to once every seven weeks.

31. The pharmaceutical composition, combination, use and method according to any one of claims 1-29, wherein the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is administered once every two weeks to once every four weeks.

32. The pharmaceutical composition, combination, use and method according to any one of claims 1-29, wherein the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is administered once every three weeks.

33. The pharmaceutical composition, combination, use and method according to any one of claims 1-32, wherein the anti-CTLA-4 antibody or antigen-binding fragment is administered once a week to once every seven weeks.

34. The pharmaceutical composition, combination, use and method according to any one of claims 1-32, wherein the anti-CTLA-4 antibody or antigen-binding fragment is administered once every two weeks to once every four weeks.

35. The pharmaceutical composition, combination, use and method according to any one of claims 1-32, wherein the anti-CTLA-4 antibody or antigen-binding fragment is administered once every three weeks.

36. The pharmaceutical composition, combination, use and method according to any one of claims 1-35, wherein the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are administered simultaneously.

37. The pharmaceutical composition, combination, use and method according to any one of claims 1-35, wherein the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are administered at intervals in time.

38. The pharmaceutical composition, combination, use and method according to any one of claims 1-35, wherein the anti-CTLA-4 antibody or antigen-binding fragment is administered before the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment.

39. The pharmaceutical composition, combination, use and method according to any one of claims 1-38, wherein the administration mode of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and / or the anti-CTLA-4 antibody or antigen-binding fragment is injection.

40. The pharmaceutical composition, combination, use and method according to any one of claims 1-38, wherein the administration mode is intravenous injection or subcutaneous injection; or, the administration mode is intravenous infusion.

41. The pharmaceutical composition, combination, use and method according to any one of claims 1-40, wherein the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is administered by intravenous injection once every two to four weeks, and the dosage is about 40 mg / kg; the anti-CTLA-4 antibody or antigen-binding fragment is administered by intravenous injection once every two to four weeks, and the dosage is 0.5-6 mg / kg.

42. The pharmaceutical composition, combination, use and method according to any one of claims 1-40, wherein the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment is administered by intravenous injection once every three weeks, and the dosage is about 40 mg / kg; the anti-CTLA-4 antibody or antigen-binding fragment is administered by intravenous injection once every three weeks, and the dosage is about 0.5 mg / kg, about 1 mg / kg, about 2 mg / kg, about 3 mg / kg or about 6 mg / kg.

43. The pharmaceutical composition, combination, use and method according to any one of claims 1-42, wherein the tumor is a benign tumor or cancer.

44. The pharmaceutical composition, combination, use and method according to any one of claims 1-43, wherein the tumor is a hematological tumor or a solid tumor.

45. The pharmaceutical composition, combination, use and method according to any one of claims 1-44, wherein the tumor is selected from acute lymphoblastic leukemia, acute myeloid leukemia, chronic lymphocytic leukemia, chronic myeloid leukemia, myeloproliferative diseases / tumors, Hodgkin lymphoma, non-Hodgkin lymphoma, Burkitt lymphoma, follicular lymphoma, multiple myeloma, giant cell myeloma, heavy chain myeloma, light chain or Bence-Jones myeloma, breast cancer, pancreatic cancer, prostate cancer, melanoma, head and neck cancer, liver cancer (such as hepatocellular carcinoma), kidney cancer, squamous cell carcinoma (such as esophageal squamous cell carcinoma), lung cancer (such as non-small cell lung cancer), cervical cancer, endometrial cancer, ovarian cancer, colorectal cancer, urothelial cancer, bladder cancer, brain cancer, gastric cancer, thyroid cancer, glioma, salivary gland cancer, leiomyosarcoma and thymic cancer.

46. The pharmaceutical composition, combination, use and method according to any one of claims 1-45, wherein the tumor is non-small cell lung cancer.

47. The pharmaceutical composition, combination, use and method according to any one of claims 1-45, wherein the tumor is locally advanced or metastatic non-small cell lung cancer.

48. The pharmaceutical composition, combination, use and method according to any one of claims 1-45, wherein the tumor is advanced non-small cell lung cancer.

49. A kit, comprising an anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment, an anti-CTLA-4 antibody or antigen-binding fragment, and an instruction manual for guiding the administration of the anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment to a patient in need; The anti-PD-L1 / CD47 bispecific antibody or antigen-binding fragment thereof comprises a variable region a that specifically binds to PD-L1 and a variable region b that specifically binds to CD47; the variable region a comprises a VHa CDR1 shown in SEQ ID NO:1, a VHa CDR2 shown in SEQ ID NO:2, a VHa CDR3 shown in SEQ ID NO:3, a VLa CDR1 shown in SEQ ID NO:4, a VLa CDR2 shown in SEQ ID NO:5, and a VLa CDR3 shown in SEQ ID NO:6; the variable region b comprises a VHb CDR1 shown in SEQ ID NO:7, a VHb CDR2 shown in SEQ ID NO:8, a VHb CDR3 shown in SEQ ID NO:9, a VLb CDR1 shown in SEQ ID NO:4, a VLb CDR2 shown in SEQ ID NO:5, and a VLb CDR3 shown in SEQ ID NO:6; or the variable region a comprises a heavy chain variable region VHa and a light chain variable region VLa; the VHa comprises the amino acid sequence shown in SEQ ID NO:10, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO:10, or an amino acid sequence having one or more conservative amino acid substitutions compared with the amino acid sequence shown in SEQ ID NO:10; the VLa comprises the amino acid sequence shown in SEQ ID NO:12, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO:12, or an amino acid sequence having one or more conservative amino acid substitutions compared with the amino acid sequence shown in SEQ ID NO:12; or the variable region b comprises a heavy chain variable region VHb and a light chain variable region VLb; the VHb comprises the amino acid sequence shown in SEQ ID NO:11, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO:11, or an amino acid sequence having one or more conservative amino acid substitutions compared with the amino acid sequence shown in SEQ ID NO:11; the VLb comprises the amino acid sequence shown in SEQ ID NO:12, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO:12, or an amino acid sequence having one or more conservative amino acid substitutions compared with the amino acid sequence shown in SEQ ID NO:12; or The anti-PD-L1 / CD47 bispecific antibody comprises heavy chain a, heavy chain b, light chain a and light chain b; the heavy chain a comprises the amino acid sequence shown in SEQ ID NO:16, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO:16, or an amino acid sequence having one or more conservative amino acid substitutions compared with the amino acid sequence shown in SEQ ID NO:16; the heavy chain b comprises the amino acid sequence shown in SEQ ID NO:17, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO:17, or an amino acid sequence having one or more conservative amino acid substitutions compared with the amino acid sequence shown in SEQ ID NO:17; the light chain a and light chain b comprise the same amino acid sequence; or, the light chain a and light chain b comprise the amino acid sequence shown in SEQ ID NO:18, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO:18, or an amino acid sequence having one or more conservative amino acid substitutions compared with the amino acid sequence shown in SEQ ID NO:18; or The anti-PD-L1 / CD47 bispecific antibody comprises heavy chain a, heavy chain b, light chain a and light chain b; the heavy chain a comprises the amino acid sequence shown in SEQ ID NO:16, the heavy chain b comprises the amino acid sequence shown in SEQ ID NO:17, and the light chain a and light chain b comprise the amino acid sequence shown in SEQ ID NO:18; or The anti-CTLA-4 antibody or antigen-binding fragment comprises VH CDR1 shown in SEQ ID NO:19, VH CDR2 shown in SEQ ID NO:20, VH CDR3 shown in SEQ ID NO:21, VL CDR1 shown in SEQ ID NO:22, VL CDR2 shown in SEQ ID NO:23, and VL CDR3 shown in SEQ ID NO:24; or The anti-CTLA-4 antibody or antigen-binding fragment comprises a heavy chain variable region and a light chain variable region; the heavy chain variable region of the anti-CTLA-4 antibody or antigen-binding fragment comprises the amino acid sequence shown in SEQ ID NO:25, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO:25, or an amino acid sequence having one or more conservative amino acid substitutions compared with the amino acid sequence shown in SEQ ID NO:25; the light chain variable region of the anti-CTLA-4 antibody or antigen-binding fragment comprises the amino acid sequence shown in SEQ ID NO:26, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO:26, or an amino acid sequence having one or more conservative amino acid substitutions compared with the amino acid sequence shown in SEQ ID NO:26; or The anti-CTLA-4 antibody comprises a heavy chain and a light chain; the heavy chain of the anti-CTLA-4 antibody comprises the amino acid sequence shown in SEQ ID NO: 28, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO: 28, or an amino acid sequence having one or more conservative amino acid substitutions compared with the amino acid sequence shown in SEQ ID NO: 28; the light chain of the anti-CTLA-4 antibody comprises the amino acid sequence shown in SEQ ID NO: 29, or an amino acid sequence having at least 80% identity with the amino acid sequence shown in SEQ ID NO: 29, or an amino acid sequence having one or more conservative amino acid substitutions compared with the amino acid sequence shown in SEQ ID NO: 29; or the fucosylation level of the anti-CTLA-4 antibody or antigen-binding fragment is 0-5%; or, the anti-CTLA-4 antibody or antigen-binding fragment is expressed by cells with knockout of the α-(1,6)-fucosyltransferase gene; or, the anti-CTLA-4 antibody or antigen-binding fragment is expressed by CHO cells with knockout of the α-(1,6)-fucosyltransferase gene.