CN117618555A

CN117618555A - Use of anti-TIGIT antibodies in combination with anti-CTLA-4 antibodies for the treatment of tumors

Info

Publication number: CN117618555A
Application number: CN202311112053.8A
Authority: CN
Inventors: 黄俊杰; 吴精博; 张少榆; 黄贤明; 李胜峰
Original assignee: Bio Thera Solutions Ltd
Current assignee: Bio Thera Solutions Ltd
Priority date: 2022-08-31
Filing date: 2023-08-30
Publication date: 2024-03-01
Also published as: WO2024046389A1

Abstract

The present invention relates to a combination comprising an anti-TIGIT antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment. The invention also relates to the use of an anti-TIGIT antibody in combination with an anti-CTLA-4 antibody for the treatment of a tumor.

Description

Use of anti-TIGIT antibodies in combination with anti-CTLA-4 antibodies for the treatment of tumors

Technical Field

The invention belongs to the field of biological medicine, and in particular relates to application of an anti-TIGIT antibody and an anti-CTLA-4 antibody in combination for treating tumors.

Background

TIGIT (T cell immunoreceptor with Ig and ITIM domains) is a T cell immunoreceptor with immunoglobulin (Ig) and tyrosine inhibitor motif (ITIM) domains, expressed predominantly on activated T cells and NK cells. TIGIT structures are shown to contain one extracellular immunoglobulin domain, one type I transmembrane region, and two ITIM motifs. TIGIT is part of a co-stimulatory network consisting essentially of the activating receptor CD226 and the inhibitory receptor TIGIT on T cells, as well as the ligand CD155 (also known as PVR, a polio virus receptor protein encoded by the PVR gene in humans) and CD112 expressed on APC, tumor cells, infected cell surfaces. TIGIT binds to PVR or CD112, which causes the intracytoplasmic Tyr225 of TIGIT to be phosphorylated, and TIGIT binds to the cell-adaptive growth factor receptor binding protein 2 (GRB 2). GRB2 can recruit SHIP1 to inhibit phosphatidylinositol triskinase (PI 3K) and Mitogen Activated Protein Kinase (MAPK) signals. In addition, phosphorylated TIGIT recruits SHIP1 via Beta-arrestin 2 and disrupts nuclear factor KB (NF-KB) activation by blocking self-ubiquitination of TNF receptor-related factor 6 (TRAF 6), a series of signaling ultimately resulting in inhibition of T cell or NK cell function and inhibition of cytokine production. PVR is a ligand for both TIGIT and CD226 molecules. After binding to CD112 or CD155, ser329 and Tyr322 of the intracellular domain of CD226 are phosphorylated; ser329 phosphorylation promotes activation of Protein Kinase (PKC) and binding of CD226 to lymphocyte-associated antigen 1 (LFA 1) to each other. LFA1 was then used for TYN-mediated Tyr322 phosphorylation and CD 226-mediated downstream signaling. A series of signalling ultimately results in activation of the function of T cells or NK cells, promoting cytokine production. Interaction also occurs between TIGIT molecules present on the surface of T cells or NK cells and CD226 molecules, which are shown to directly disrupt the normal dimer formation of CD226, thereby disrupting the normal physiological function of CD 226. TIGIT and CD226 act like a balance at both ends, through PVR, which regulates the immune function of the organism smartly through co-stimulation and co-suppression signaling.

Cytotoxic T lymphocyte antigen-4 (CTLA-4) is a key inhibitory receptor affecting T cell function, playing a key role in the initiation of the immune response (Scaalapino KJ1, daikh DI. (2008), immunol Rev. 223:143-155). When antigen is transported by MHC class I or class II antigen presenting cells to a T Cell Receptor (TCR), the signal on the TCR is amplified and counteracted by a co-stimulatory molecule. After binding of CD28 on T cells to B7-1 (CD 80) and B7-2 (CD 86) on antigen presenting cells, a signal is emitted that total activation of the T cells is required. This binding of CD28/B7 results in increased production of interleukin 2 and other stimulatory cytokines, increased metabolism, promotion of cell cycle progression, up-regulation of cell survival genes, and ultimately proliferation and differentiation of T cells. When CD28 binds and causes T-cell proliferation, CTLA-4 is transported and expressed on the surface of T-cells (Linsley PS, et al (1996), immunity.4 (6): 535-543). The stronger the activation signal through the TCR, the more CTLA-4 is transported and expressed. When on the cell surface, the CTLA-4 inhibition signal is transmitted (Egen JG, allison JP. (2002), immunity.16 (1): 23-25). CTLA-4 has a higher affinity for B7 than CD28 and can block further coactivation (Krummel MF, allison JP. (1995), J Exp Med.182 (2): 458-465). Furthermore, CTLA-4 expressing cells were able to capture and degrade B7-1 and B7-2 by endocytosis (Qureshi OS, et al (2011), science.332 (6029): 600-603).

In view of the importance of immune checkpoints, there is a need to develop new combination therapies.

Disclosure of Invention

The invention aims to provide a combined drug and application thereof in treating tumors.

In some embodiments, the combination comprises an anti-TIGIT antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, the combination comprises a therapeutically effective amount of an anti-TIGIT antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment.

In another aspect, the invention provides a pharmaceutical combination for treating a tumor comprising an anti-TIGIT antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment.

In another aspect, the invention provides a combination of an anti-TIGIT antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment for treating a tumor. In some embodiments, the invention provides a method of treating a tumor comprising administering to a patient in need thereof an effective amount of an anti-TIGIT antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment.

In another aspect, the invention provides the use OF an anti-TIGIT antibody (e.g., antibody h10D8OF or h10D8 OFKF) or antigen-binding fragment in the manufacture OF a medicament for use in combination with an anti-CTLA-4 antibody or antigen-binding fragment in the treatment OF a tumor.

In another aspect, the invention provides the use OF an anti-TIGIT antibody (e.g., antibody h10D8OF or h10D8 OFKF) or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment in the manufacture OF a medicament for treating a tumor.

In another aspect, the invention provides the use OF an anti-CTLA-4 antibody or antigen-binding fragment in the manufacture OF a medicament for use in combination with an anti-TIGIT antibody (e.g., antibody h10D8OF or h10D8 OFKF) or antigen-binding fragment in the treatment OF a tumor.

In another aspect, the invention provides the use OF an anti-CTLA-4 antibody or antigen-binding fragment and an anti-TIGIT antibody (e.g., antibody h10D8OF or h10D8 OFKF) or antigen-binding fragment in the manufacture OF a medicament for treating a tumor.

In another aspect, the invention provides the use OF an anti-TIGIT antibody (e.g., antibody h10D8OF or h10D8 OFKF) or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment in the preparation OF a pharmaceutical composition for treating a tumor.

In another aspect, the invention provides a kit comprising an anti-TIGIT antibody and a package insert comprising instructions for using the anti-TIGIT antibody in combination with an anti-CTLA-4 antibody to treat a tumor.

In another aspect, the invention provides a kit comprising an anti-CTLA-4 antibody and a package insert comprising instructions for using the anti-CTLA-4 antibody in combination with an anti-TIGIT antibody to treat a tumor.

In another aspect, the invention provides a kit comprising an anti-TIGIT antibody and an anti-CTLA-4 antibody agent, and a package insert comprising instructions for using the anti-TIGIT antibody and anti-CTLA-4 antibody agent to treat a tumor.

In another aspect, the invention provides a kit comprising an agent that is an anti-TIGIT antibody and a package insert comprising instructions for using the agent that is the anti-TIGIT antibody in combination with an anti-CTLA-4 antibody to treat a tumor.

In another aspect, the invention provides a kit comprising an agent that is an anti-CTLA-4 antibody and a package insert comprising instructions for using the agent that is an anti-CTLA-4 antibody in combination with an anti-TIGIT antibody to treat a tumor.

In another aspect, the invention provides a kit comprising an anti-TIGIT antibody or antigen-binding fragment (or formulation), an anti-CTLA-4 antibody or antigen-binding fragment (or formulation), and instructions for administering the anti-TIGIT antibody or antigen-binding fragment (or formulation) and the anti-CTLA-4 antibody or antigen-binding fragment (or formulation) to a patient in need thereof.

In some embodiments, the anti-TIGIT antibody or antigen-binding fragment comprises at least one or more of HCDR1 shown in SEQ ID No. 1, HCDR2 shown in SEQ ID No. 2, HCDR3 shown in SEQ ID No. 3, LCDR1 shown in SEQ ID No. 4, LCDR2 shown in SEQ ID No. 5, LCDR3 shown in SEQ ID No. 6.

In some embodiments, the anti-TIGIT antibody or antigen-binding fragment comprises HCDR1 shown in SEQ ID No. 1, HCDR2 shown in SEQ ID No. 2, HCDR3 shown in SEQ ID No. 3, LCDR1 shown in SEQ ID No. 4, LCDR2 shown in SEQ ID No. 5, and LCDR3 shown in SEQ ID No. 6.

In some embodiments, the anti-TIGIT antibody or antigen-binding fragment comprises a heavy chain variable region and a light chain variable region.

In some embodiments, the heavy chain variable region of the anti-TIGIT antibody or antigen-binding fragment comprises the amino acid sequence shown in SEQ ID No. 7, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID No. 7, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID No. 7.

In some embodiments, the light chain variable region of the anti-TIGIT antibody or antigen-binding fragment comprises the amino acid sequence shown in SEQ ID No. 8, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID No. 8, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID No. 8.

In some embodiments, the heavy chain variable region of the anti-TIGIT antibody or antigen-binding fragment comprises the amino acid sequence shown in SEQ ID No. 7, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID No. 7, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID No. 7; the light chain variable region of the anti-TIGIT antibody or antigen-binding fragment comprises the amino acid sequence shown in SEQ ID No. 8, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID No. 8, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID No. 8.

In some embodiments, the heavy chain variable region of the anti-TIGIT antibody or antigen-binding fragment comprises the amino acid sequence shown in SEQ ID No. 7 and the light chain variable region of the anti-TIGIT antibody or antigen-binding fragment comprises the amino acid sequence shown in SEQ ID No. 8.

In some embodiments, the heavy chain of the anti-TIGIT antibody comprises the amino acid sequence shown in SEQ ID No. 9, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID No. 9, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID No. 9.

In some embodiments, the light chain of the anti-TIGIT antibody comprises the amino acid sequence shown in SEQ ID No. 10, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID No. 10, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID No. 10.

In some embodiments, the heavy chain of the anti-TIGIT antibody comprises the amino acid sequence shown in SEQ ID No. 9, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID No. 9, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID No. 9; the light chain of the anti-TIGIT antibody comprises the amino acid sequence shown in SEQ ID No. 10, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID No. 10, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID No. 10.

In some embodiments, the anti-TIGIT antibody is antibody h10D8OF or h10D8OFKF, the heavy chain OF which comprises the amino acid sequence shown in SEQ ID No. 9, and the light chain OF which comprises the amino acid sequence shown in SEQ ID No. 10. In some embodiments, the antibody h10D8OF or h10D8OFKF contains two heavy chains with identical sequences and two light chains with identical sequences.

In some embodiments, the anti-TIGIT antibody (e.g., antibody h10D8 OFKF) or antigen binding fragment has a fucosylation level of 0-10%. In some embodiments, the anti-TIGIT antibody (e.g., antibody h10D8 OFKF) or antigen binding fragment has a fucosylation level of 0-5%. In some embodiments, the anti-TIGIT antibody (e.g., antibody h10D8 OFKF) or antigen-binding fragment has a fucosylation level of about 0, about 0.1%, about 0.5%, about 0.8%, about 1%, about 1.3%, about 1.6%, about 2.1%, 2.9%, about 3%, about 3.3%, 3.8%, about 4%, about 4.2%, 4.3%, about 4.6%, about 5%, or a range between any two of these values (inclusive), or any value therein. In some embodiments, the anti-TIGIT antibody (e.g., antibody h10D8 OFKF) or antigen binding fragment does not bind to fucose. In some embodiments, the anti-TIGIT antibody (e.g., antibody h10D8 OFKF) or antigen binding fragment has enhanced ADCC effect (anti-independent cell-mediated cytotoxicity).

In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment comprises at least one or more of HCDR1 shown in SEQ ID NO. 11, HCDR2 shown in SEQ ID NO. 12, HCDR3 shown in SEQ ID NO. 13, LCDR1 shown in SEQ ID NO. 14, LCDR2 shown in SEQ ID NO. 15, LCDR3 shown in SEQ ID NO. 16.

In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment comprises HCDR1 shown in SEQ ID NO. 11, HCDR2 shown in SEQ ID NO. 12, HCDR3 shown in SEQ ID NO. 13, LCDR1 shown in SEQ ID NO. 14, LCDR2 shown in SEQ ID NO. 15, and LCDR3 shown in SEQ ID NO. 16.

In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment comprises a heavy chain variable region and a light chain variable region.

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. 17, or an amino acid sequence having at least 80% identity to the sequence of SEQ ID No. 17, or an amino acid sequence having one or more conservative amino acid substitutions as compared to the sequence of SEQ ID No. 17.

In some embodiments, the light chain variable region of the anti-CTLA-4 antibody or antigen-binding fragment comprises the amino acid sequence of SEQ ID No. 18, or an amino acid sequence having at least 80% identity to the sequence of SEQ ID No. 18, or an amino acid sequence having one or more conservative amino acid substitutions as compared to the sequence of SEQ ID No. 18.

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. 17, or an amino acid sequence having at least 80% identity to the sequence of SEQ ID No. 17, or an amino acid sequence having one or more conservative amino acid substitutions as compared to the sequence of SEQ ID No. 17; the light chain variable region of the anti-CTLA-4 antibody or antigen-binding fragment comprises the amino acid sequence shown as SEQ ID NO. 18, or an amino acid sequence having at least 80% identity to the sequence shown as SEQ ID NO. 18, or an amino acid sequence having one or more conservative amino acid substitutions as compared to the sequence shown as SEQ ID NO. 18.

In some embodiments, 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. 17 and 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. 18.

In some embodiments, the heavy chain of the anti-CTLA-4 antibody comprises the amino acid sequence of SEQ ID NO. 19, or an amino acid sequence having at least 80% identity to the sequence of SEQ ID NO. 19, or an amino acid sequence having one or more conservative amino acid substitutions as compared to the sequence of SEQ ID NO. 19.

In some embodiments, the light chain of the anti-CTLA-4 antibody comprises the amino acid sequence of SEQ ID NO. 20, or an amino acid sequence having at least 80% identity to the sequence of SEQ ID NO. 20, or an amino acid sequence having one or more conservative amino acid substitutions as compared to the sequence of SEQ ID NO. 20.

In some embodiments, the heavy chain of the anti-CTLA-4 antibody comprises the amino acid sequence of SEQ ID No. 19, or an amino acid sequence having at least 80% identity to the sequence of SEQ ID No. 19, or an amino acid sequence having one or more conservative amino acid substitutions as compared to the sequence of SEQ ID No. 19; the light chain of the anti-CTLA-4 antibody comprises an amino acid sequence shown as SEQ ID NO. 20, or an amino acid sequence which has at least 80% identity with the sequence shown as SEQ ID NO. 20, or an amino acid sequence which has one or more conservative amino acid substitutions with the sequence shown as SEQ ID NO. 20.

In some embodiments, the heavy chain of the anti-CTLA-4 antibody comprises the amino acid sequence shown in SEQ ID NO. 19 and the light chain of the anti-CTLA-4 antibody comprises the amino acid sequence shown in SEQ ID NO. 20. In some embodiments, an anti-CTLA-4 antibody contains two heavy chains that are identical in sequence and two light chains that are identical in sequence.

In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment has a total amount of high mannose glycoforms of <5% and/or a total amount of sialylated glycoforms of <3%. In some embodiments, the total amount of high mannose glycoforms of the anti-CTLA-4 antibody or antigen-binding fragment is about 0.1%, about 0.3%, about 0.9%, about 1.18%, about 1.7%, about 2.6%, about 3.3%, about 4.1%, about 4.9%, about 4.99%, or a range between any two of these values (inclusive) or any value therein. In some embodiments, the total amount of sialylated glycoforms of the anti-CTLA-4 antibody or antigen-binding fragment is about 0.1%, 0.2%, about 0.36%, about 0.8%, about 1.5%, about 2.2%, about 2.7%, about 2.9%, 2.99%, or a range between any two of these values (inclusive) or any value therein.

In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment has a total amount of high mannose glycoforms of <2% and/or a total amount of sialylated glycoforms of <1%.

In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment has a fucosylation level of 0-10%. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment has a fucosylation level of 0-5%. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment has a fucosylation level of about 0, about 0.1%, about 0.5%, about 0.8%, about 1%, about 1.3%, about 1.6%, about 2.1%, 2.9%, about 3%, about 3.3%, 3.8%, about 4%, about 4.2%, 4.3%, about 4.6%, about 5%, or a range between any two of these values (inclusive) or any value therein. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment does not bind fucose. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment has enhanced ADCC effect (anti-independent cell-mediated cytotoxicity).

In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment has a fucosylation level of 0-10%, a total amount of high mannose glycoforms of <5% and/or a total amount of sialylated glycoforms of <3%. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment has a fucosylation level of 0-5%, a total amount of high mannose glycoforms of <5% and/or a total amount of sialylated glycoforms of <3%. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment has a fucosylation level of about 0%, a total amount of high mannose glycoforms of <5% and/or a total amount of sialylated glycoforms of <3%. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment has a fucosylation level of 0-10%, a total amount of high mannose glycoforms of <3% and/or a total amount of sialylated glycoforms of <2%. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment has a fucosylation level of 0-5%, a total amount of high mannose glycoforms of <3% and/or a total amount of sialylated glycoforms of <2%. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment has a fucosylation level of about 0%, a total amount of high mannose glycoforms of <3% and/or a total amount of sialylated glycoforms of <2%. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment has a fucosylation level of 0-10%, a total amount of high mannose glycoforms of <2% and/or a total amount of sialylated glycoforms of <1%. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment has a fucosylation level of 0-5%, a total amount of high mannose glycoforms of <2% and/or a total amount of sialylated glycoforms of <1%. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment has a fucosylation level of about 0%, a total amount of high mannose glycoforms of <2% and/or a total amount of sialylated glycoforms of <1%.

In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment is ipilimumab (Yervoy ^TM Or a biological analogue thereof).

Antibodies can be expressed in host cells by genetic engineering and obtained by purification; purification can be carried out by conventional methods, for example, centrifugation of the cell suspension followed by collection of the supernatant, and centrifugation again to further remove impurities. ProteinA affinity columns and ion exchange columns can be used to purify antibodies.

In some embodiments, the hypofucosylated or afucosylated anti-TIGIT antibody (e.g., antibody h10D8 OFKF) or antigen-binding fragment or anti-CTLA-4 antibody or antigen-binding fragment is expressed by an a- (1, 6) -fucosyltransferase knockout cell line, e.g., by a CHO-K1 cell with an a- (1, 6) -fucosyltransferase knockout.

In some embodiments, the invention also provides pharmaceutical compositions comprising an anti-TIGIT antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, the pharmaceutical composition is a pharmaceutical composition suitable for injection, such as a bolus type pharmaceutical composition or an infusion (instillation) type pharmaceutical composition. In some embodiments, the pharmaceutical composition comprises at least 0.1% of an anti-TIGIT antibody or antigen-binding fragment and 0.1% of an anti-CTLA-4 antibody or antigen-binding fragment. The percentage of antibody may vary and may be between about 2% and about 90% of the weight of a given dosage form. The anti-TIGIT antibodies or antigen-binding fragments and anti-CTLA-4 antibodies or antigen-binding fragments in such therapeutically useful pharmaceutical compositions can be administered in effective amounts.

On the other hand, the invention also provides a preparation method of the pharmaceutical composition, which comprises the following steps: the anti-TIGIT antibodies or antigen-binding fragments and anti-CTLA-4 antibodies or antigen-binding fragments, respectively, described herein are admixed with a pharmaceutically acceptable carrier suitable for injection (e.g., water for injection, physiological saline, etc.). In some embodiments, the method of preparing the above pharmaceutical composition comprises: the anti-TIGIT antibodies or antigen-binding fragments and anti-CTLA-4 antibodies or antigen-binding fragments described herein are mixed with a pharmaceutically acceptable carrier suitable for injection (e.g., water for injection, physiological saline, etc.). Methods of mixing the above antibodies or antigen binding fragments with a carrier are generally known in the art.

In some embodiments, the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment is 1-50:1. In some embodiments, the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment is 5-25:1. In some embodiments, the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment is 1-15:1. In some embodiments, the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment is 1-10:1. In some embodiments, the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment is about 5:1. In some embodiments, the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment is 15-30:1. In some embodiments, the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment is 20-30:1. In some embodiments, the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment is about 25:1.

In some embodiments, the dose of anti-TIGIT antibody or antigen-binding fragment administered per administration is about 0.05mg to 1200mg, about 0.5mg to 1000mg, about 5mg to 500mg, about 5mg to 100mg, about 10mg to 50mg, or a formulation comprising such a dose of anti-TIGIT antibody or antigen-binding fragment. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered at a dose of about 0.05mg, about 0.1mg, about 0.5mg, about 1mg, about 2mg, about 3mg, about 4mg, about 5mg, about 6mg, about 7mg, about 8mg, about 9mg, about 10mg, about 12mg, about 20mg, about 30mg, about 40mg, about 50mg, about 60mg, about 80mg, about 100mg, about 120mg, about 200mg, about 250mg, about 290mg, about 300mg, about 330mg, about 380mg, about 400mg, about 480mg, about 500mg, about 580mg, about 600mg, about 800mg, about 900mg, about 1000mg, about 1200mg, or a range between any two of these values (inclusive) or any value therein, or a formulation containing the dose of the anti-TIGIT antibody or antigen-binding fragment. In some embodiments, the anti-TIGIT antibody is antibody h10D8OF or h10D8OFKF.

In some embodiments, the dose of anti-TIGIT antibody or antigen-binding fragment administered per administration is about 0.01mg/kg to 20mg/kg, about 0.01mg/kg to 2mg/kg, about 0.01mg/kg to 1mg/kg, about 0.05mg/kg to 1mg/kg, about 0.1mg/kg to 1mg/kg, or a formulation comprising such a dose of anti-TIGIT antibody or antigen-binding fragment. In some embodiments, the dose of anti-TIGIT antibody or antigen-binding fragment administered per administration is about 0.01mg/kg, about 0.05mg/kg, about 0.1mg/kg, about 0.3mg/kg, about 0.5mg/kg, about 0.8mg/kg, about 0.9mg/kg, about 1mg/kg, about 2mg/kg, about 3mg/kg, about 4mg/kg, about 5mg/kg, about 6mg/kg, about 7mg/kg, about 8mg/kg, about 9mg/kg, about 10mg/kg, about 12mg/kg, about 14mg/kg, about 15mg/kg, about 18mg/kg, about 20mg/kg, or a range between any two of these values (inclusive), or a formulation comprising such a dose of anti-TIGIT antibody or antigen-binding fragment. In some embodiments, the anti-TIGIT antibody is antibody h10D8OF or h10D8OFKF.

In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered at a dose of about 0.05mg to 1200mg, about 0.5mg to 1000mg, about 5mg to 500mg, about 5mg to 100mg, about 10mg to 50mg, or a formulation comprising such a dose of the anti-TIGIT antibody or antigen-binding fragment per treatment cycle. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered at a dose of about 0.05mg, about 0.1mg, about 0.5mg, about 1mg, about 2mg, about 3mg, about 4mg, about 5mg, about 6mg, about 7mg, about 8mg, about 9mg, about 10mg, about 12mg, about 20mg, about 30mg, about 40mg, about 50mg, about 60mg, about 80mg, about 100mg, about 120mg, about 200mg, about 250mg, about 290mg, about 300mg, about 330mg, about 380mg, about 400mg, about 480mg, about 500mg, about 580mg, about 600mg, about 800mg, about 900mg, about 1000mg, about 1200mg, or a range between any two of these values (inclusive), or any value therein, or formulation containing such dose of anti-TIGIT antibody or antigen-binding fragment per treatment cycle. In some embodiments, the anti-TIGIT antibody is antibody h10D8OF or h10D8OFKF.

In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered at a dose of about 0.01mg/kg to 20mg/kg, about 0.01mg/kg to 2mg/kg, about 0.01mg/kg to 1mg/kg, about 0.05mg/kg to 1mg/kg, about 0.1mg/kg to 1mg/kg per treatment cycle, or a formulation comprising such dose of the anti-TIGIT antibody or antigen-binding fragment. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered at a dose of about 0.01mg/kg, about 0.05mg/kg, about 0.1mg/kg, about 0.3mg/kg, about 0.5mg/kg, about 0.8mg/kg, about 0.9mg/kg, about 1mg/kg, about 2mg/kg, about 3mg/kg, about 4mg/kg, about 5mg/kg, about 6mg/kg, about 7mg/kg, about 8mg/kg, about 9mg/kg, about 10mg/kg, about 12mg/kg, about 14mg/kg, about 15mg/kg, about 18mg/kg, about 20mg/kg, or a range between any two of these values (inclusive), or a formulation containing the dose of the anti-TIGIT antibody or antigen-binding fragment. In some embodiments, the anti-TIGIT antibody is antibody h10D8OF or h10D8OFKF.

In some embodiments, the dose of anti-CTLA-4 antibody or antigen-binding fragment per administration is about 0.05mg to 600mg, about 0.5mg to 600mg, about 1mg to 100mg, about 1mg to 50mg, about 1mg to 10mg, or a formulation containing such dose of 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 0.05mg, about 0.1mg, about 0.2mg, about 0.3mg, about 0.5mg, about 0.8mg, about 1mg, about 2mg, about 3mg, about 4mg, about 5mg, about 6mg, about 7mg, about 8mg, about 9mg, about 10mg, about 12mg, about 20mg, about 30mg, about 50mg, about 60mg, about 80mg, about 100mg, about 120mg, about 200mg, about 250mg, about 290mg, about 300mg, about 330mg, about 380mg, about 400mg, about 434mg, about 480mg, about 500mg, about 567mg, about 580mg, about 600mg, or a range between any two of these values (inclusive), or any value therein, or formulation containing such dose of anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, the anti-CTLA-4 antibody is antibody 1.

In some embodiments, the dose of anti-CTLA-4 antibody or antigen-binding fragment per administration is about 0.01mg/kg to 10mg/kg, about 0.01mg/kg to 2mg/kg, about 0.01mg/kg to 1mg/kg, about 0.02mg/kg to 0.5mg/kg, about 0.02mg/kg to 0.2mg/kg, or a formulation comprising such dose of 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 0.01mg/kg, about 0.02mg/kg, about 0.03mg/kg, about 0.04mg/kg, about 0.05mg/kg, about 0.06mg/kg, about 0.07mg/kg, about 0.08mg/kg, about 0.1mg/kg, about 0.3mg/kg, about 0.5mg/kg, about 0.8mg/kg, about 0.9mg/kg, about 1mg/kg, about 2mg/kg, about 3mg/kg, about 4mg/kg, about 5mg/kg, about 6mg/kg, about 7mg/kg, about 8mg/kg, about 9mg/kg, about 10mg/kg, or a range between any two of these values (inclusive), or a formulation containing such dose of anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, the anti-CTLA-4 antibody is antibody 1.

In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment is administered at a dose of about 0.05mg to 600mg, about 0.5mg to 600mg, about 1mg to 100mg, about 1mg to 50mg, about 1mg to 10mg, or a formulation containing such dose of anti-CTLA-4 antibody or antigen-binding fragment per treatment cycle. In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment is administered at a dose of about 0.05mg, about 0.1mg, about 0.2mg, about 0.3mg, about 0.5mg, about 0.8mg, about 1mg, about 2mg, about 3mg, about 4mg, about 5mg, about 6mg, about 7mg, about 8mg, about 9mg, about 10mg, about 12mg, about 20mg, about 30mg, about 50mg, about 60mg, about 80mg, about 100mg, about 120mg, about 200mg, about 250mg, about 290mg, about 300mg, about 330mg, about 380mg, about 400mg, about 434mg, about 480mg, about 500mg, about 567mg, about 580mg, about 600mg, or a range between any two of these values (inclusive), or any value therein, or formulation containing such dose of anti-CTLA-4 antibody or antigen-binding fragment per treatment cycle. In some embodiments, the anti-CTLA-4 antibody is antibody 1.

In some embodiments, the anti-CTLA-4 antibody or antigen-binding fragment is administered at a dose of about 0.01mg/kg to 10mg/kg, about 0.01mg/kg to 2mg/kg, about 0.01mg/kg to 1mg/kg, about 0.02mg/kg to 0.5mg/kg, about 0.02mg/kg to 0.2mg/kg per treatment cycle, or a formulation comprising such dose of 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 0.01mg/kg, about 0.02mg/kg, about 0.03mg/kg, about 0.04mg/kg, about 0.05mg/kg, about 0.06mg/kg, about 0.07mg/kg, about 0.08mg/kg, about 0.1mg/kg, about 0.3mg/kg, about 0.5mg/kg, about 0.8mg/kg, about 0.9mg/kg, about 1mg/kg, about 2mg/kg, about 3mg/kg, about 4mg/kg, about 5mg/kg, about 6mg/kg, about 7mg/kg, about 8mg/kg, about 9mg/kg, about 10mg/kg, or a range between any two of these values (inclusive), or a formulation comprising such dose of anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, the anti-CTLA-4 antibody is antibody 1.

In some embodiments, the invention provides a method of treating a tumor, the method comprising: administering to a patient in need thereof about 0.05mg to 1200mg, about 0.5mg to 1000mg, about 5mg to 500mg, about 5mg to 100mg, about 10mg to 50mg, such as about 0.05mg, about 0.08mg, about 0.1mg, about 0.2mg, about 0.3mg, about 0.4mg, about 0.5mg, about 0.6mg, about 0.7mg, about 0.8mg, about 0.9mg, about 1mg, about 2mg, about 3mg, about 4mg, about 5mg, about 6mg, about 7mg, about 8mg, about 9mg, about 10mg, about 12mg, about 20mg, about 30mg, about 50mg, about 60mg, about 80mg, about 100mg, about 120mg, about 200mg, about 250mg, about 290mg, about 300mg, about 330mg, about 380mg, about 400mg, about 434mg, about 480mg, about 500mg, about 567mg, about 580mg, about 600mg, about 700mg, about 800mg, about 900mg, about 1000mg, about 1100mg, about 1200mg OF an anti-TIT antibody (e.g., antibody h10D8OF or OfD 8 OF) or a fragment thereof, or an antigen-binding antigen or a GIT-binding fragment thereof; an anti-CTLA antigen or antigen-binding fragment thereof is also administered to a patient in need thereof in a dose of about 0.05mg to 600mg, about 0.5mg to 600mg, about 1mg to 100mg, about 1mg to 50mg, about 1mg to 10mg, such as about 0.05mg, about 0.08mg, about 0.1mg, about 0.2mg, about 0.3mg, about 0.4mg, about 0.5mg, about 0.6mg, about 0.7mg, about 0.9mg, about 1mg, about 2mg, about 3mg, about 4mg, about 5mg, about 6mg, about 7mg, about 8mg, about 9mg, about 10mg, about 12mg, about 20mg, about 30mg, about 50mg, about 60mg, about 80mg, about 100mg, about 120mg, about 200mg, about 250mg, about 290mg, about 300mg, about 330mg, about 380mg, about 400mg, about 434mg, about 480mg, about 500mg, about 567mg, about 580mg, about 600mg, or an anti-CTLA-binding antibody or antigen-binding fragment thereof.

In some embodiments, the anti-TIGIT antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are administered in a mass ratio of 1-50:1. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are administered in a mass ratio of 5-25:1. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are administered at a mass ratio of 1-15:1. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are administered at a mass ratio of 1-10:1. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are administered at a mass ratio of about 5:1. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are administered at a mass ratio of 15-30:1. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are administered at a mass ratio of 20-30:1. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are administered at a mass ratio of about 25:1.

In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered at a dose of about 10mg, 30mg, 100mg, 300mg, 600mg, or 900mg per treatment cycle and the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment administered is 1-50:1. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered at a dose of about 10mg, 30mg, 100mg, 300mg, 600mg, or 900mg per treatment cycle and the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment administered is 5-25:1. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered at a dose of about 10mg, 30mg, 100mg, 300mg, 600mg, or 900mg per treatment cycle and the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment administered is 1-15:1. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered at a dose of about 10mg, 30mg, 100mg, 300mg, 600mg, or 900mg per treatment cycle and the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment administered is 1-10:1. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered at a dose of about 10mg, 30mg, 100mg, 300mg, 600mg, or 900mg per treatment cycle and the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment administered is about 5:1. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered at a dose of about 10mg, 30mg, 100mg, 300mg, 600mg, or 900mg per treatment cycle and the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment administered is 15-30:1. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered at a dose of about 10mg, 30mg, 100mg, 300mg, 600mg, or 900mg per treatment cycle and the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment administered is 20-30:1. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered at a dose of about 10mg, 30mg, 100mg, 300mg, 600mg, or 900mg per treatment cycle and the mass ratio of anti-TIGIT antibody or antigen-binding fragment to anti-CTLA-4 antibody or antigen-binding fragment administered is about 25:1.

In some embodiments, the anti-TIGIT antibody is antibody h10D8 kf and the anti-CTLA-4 antibody is antibody 1.

In some embodiments, the anti-TIGIT antibody is antibody h10D8OF and the anti-CTLA-4 antibody is antibody 1.

The formulation may be a formulation suitable for injectable use including sterile aqueous solutions or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. Suitable carriers include physiological saline, bacteriostatic or Phosphate Buffered Saline (PBS), ethanol, solvents or dispersion media for polyols (e.g., glycerol, propylene glycol, and liquid polyethylene glycols, and the like), and suitable mixtures thereof. In some embodiments, the formulation comprises at least 0.1% antibody or antigen binding fragment.

In some embodiments, one treatment cycle is 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 2 months, 5 months, 6 months, one year, or a range between any two of these values (inclusive) or any value therein. In some embodiments, the dosing frequency is from once daily to once every 7 weeks. In some embodiments, the dosing frequency is once daily, three times per week, twice per week, once every 2 weeks, once every 3 weeks, once every 4 weeks, once every 5 weeks, once every 6 weeks, or once every 7 weeks. The frequency of administration of the anti-TIGIT antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment may be the same or different.

In some embodiments, the patient is treated with a single anti-TIGIT antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, the symptoms of the patient are alleviated after a single administration. In some embodiments, after a single administration, the symptoms after the patient are not expected to be alleviated, and the patient is administered an anti-TIGIT 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 without further treatment. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment, respectively, are administered once per treatment cycle to the patient. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are administered multiple times, e.g., 2, 3, 4, or 5 times, per treatment cycle to the patient. In some embodiments, the patient receives one treatment cycle. In some embodiments, the patient receives multiple treatment cycles (e.g., 2, 3, or 4) of treatment.

In some embodiments, the anti-TIGIT antibody or antigen-binding fragment, anti-CTLA-4 antibody or antigen-binding fragment is administered by subcutaneous (s.c.) injection, intraperitoneal (i.p.) injection, parenteral injection, intraarterial injection, or intravenous (i.v.) injection or infusion. The anti-TIGIT 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-TIGIT antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are administered by intraperitoneal injection.

In some embodiments, the anti-TIGIT antibody (e.g., antibody h10D8OF or h10D8 OFKF) or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment, respectively, are separate dosing units, in combination. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment can be administered at different times. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered prior to administration of the anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered after administration of the anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered concurrently with the anti-CTLA-4 antibody or antigen-binding fragment. In some embodiments, an anti-TIGIT antibody (e.g., antibody h10D8OF or h10D8 OFKF) or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment, respectively, are formulated into a pharmaceutical composition and administered to a patient in a form suitable for the selected route OF administration, e.g., by parenteral, intravenous (iv), intramuscular, topical, or subcutaneous route.

In some embodiments, an anti-TIGIT antibody (e.g., antibody h10D8OF or h10D8 OFKF) or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment are administered in combination together forming a combined dosing unit. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment and the anti-CTLA-4 antibody or antigen-binding fragment are combined to formulate a pharmaceutical composition and administered to a patient in a form suitable for the selected route of administration, e.g., by parenteral, intravenous (iv), intramuscular, topical, or subcutaneous route.

In some embodiments, the anti-TIGIT antibodies or antigen-binding fragments (or formulations), anti-CTLA-4 antibodies or antigen-binding fragments (or formulations) can be used in combination with other therapeutic methods for treating tumors, such as chemotherapy, radiation therapy, and surgical therapies, among others.

In some embodiments, the patient has a tumor. In some embodiments, the tumor includes, but is not limited to, benign tumor, cancer. In some embodiments, the tumor includes, but is not limited to, hematological cancer, solid tumors. In some embodiments, hematological cancers include, but are not limited to, leukemia, lymphoma, and myeloma. In some embodiments, leukemia includes Acute Lymphoblastic Leukemia (ALL), acute Myelogenous Leukemia (AML), chronic Lymphocytic Leukemia (CLL), chronic Myelogenous Leukemia (CML), and myeloproliferative diseases/tumors (MPDS). In some embodiments, lymphomas include hodgkin's lymphoma, non-hodgkin's lymphoma, burkitt's lymphoma, and follicular lymphoma (small and large cells). In some embodiments, the myeloma includes Multiple Myeloma (MM), giant cell myeloma, heavy chain myeloma, and light chain or bens-jones myeloma. In some embodiments, the solid tumor comprises breast cancer, pancreatic cancer, prostate cancer, melanoma, head and neck cancer, liver cancer, kidney cancer, squamous cell carcinoma, esophageal squamous cell carcinoma, lung cancer, non-small cell lung cancer, cervical cancer, esophageal cancer, endometrial cancer, ovarian cancer, colon cancer, colorectal cancer, urothelial cancer, bladder cancer, brain cancer. In some embodiments, the tumor is a pathologically diagnosed locally advanced or metastatic malignant solid tumor for which no effective treatment has yet been established.

In some embodiments, the combination of an anti-TIGIT antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment can increase tumor suppression and/or increase survival as compared to treatment with the anti-TIGIT antibody or antigen-binding fragment and anti-CTLA-4 antibody or antigen-binding fragment alone.

Drawings

FIG. 1 is a graph of the tumor growth curve of the mice of example 2.

FIG. 2 is a survival curve of the mice of example 2.

FIG. 3 is a graph of the tumor growth curve of the mice of example 3.

FIG. 4 is a survival curve of the mice of example 3.

Terminology

Unless otherwise indicated, each term below shall have the meaning described below.

Definition of the definition

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

The terms "comprising" or "including" as used herein mean that the compositions and methods, etc., include the recited elements, e.g., components or steps, but do not exclude the others.

The term "polypeptide" is intended to encompass both the singular and the plural of "polypeptides" and refers to molecules composed of amino acid monomers that are linearly linked by amide bonds (also referred to as peptide bonds). The term "polypeptide" refers to any single chain or multiple chains of two or more amino acids, and does not refer to a particular length of product. Thus, the definition of "polypeptide" includes peptides, dipeptides, tripeptides, oligopeptides, "proteins", "amino acid chains" or any other term used to refer to two or more amino acid chains, and the term "polypeptide" may be used in place of, or in addition to, any of the terms described above. The term "polypeptide" is also intended to refer to products of modification of the polypeptide after expression, including but not limited to glycosylation, acetylation, phosphorylation, amidation, derivatization by known protecting/blocking groups, proteolytic cleavage, or non-naturally occurring amino acid modification. The polypeptide may be derived from a natural biological source or produced by recombinant techniques, but it need not be translated from the specified nucleic acid sequence, and it may be produced in any manner including chemical synthesis.

"amino acid" refers to an organic compound containing both amino and carboxyl groups, such as an alpha-amino acid, which may be encoded by a nucleic acid directly or in precursor form. A single amino acid is encoded by a nucleic acid consisting of three nucleotides, a so-called codon or base triplet. Each amino acid is encoded by at least one codon. The same amino acid is encoded by different codons called "degeneracy of the genetic code". Amino acids include natural amino acids and unnatural amino acids. Natural amino acids include alanine (three-letter code: ala, one-letter code: a), arginine (arg, R), asparagine (asn, N), aspartic acid (asp, D), cysteine (cys, C), glutamine (gln, Q), glutamic acid (glu, E), glycine (gly, G), histidine (his, H), isoleucine (ile, I), leucine (leu, L), lysine (lys, K), methionine (met, M), phenylalanine (phe, F), proline (pro, P), serine (ser, S), threonine (thr, T), tryptophan (trp, W), tyrosine (tyr, Y), and valine (val, V).

"conservative amino acid substitution" refers to the substitution of one amino acid residue with another amino acid residue that contains a side chain (R group) that is similar in chemical properties (e.g., charge or hydrophobicity). In general, conservative amino acid substitutions will not substantially alter the functional properties of the protein. Examples of classes of amino acids containing chemically similar side chains include: 1) Aliphatic side chain: glycine, alanine, valine, leucine and isoleucine; 2) Aliphatic hydroxyl side chains: serine and threonine; 3) Amide-containing side chains: asparagine and glutamine; 4) Aromatic side chain: phenylalanine, tyrosine, and tryptophan; 5) Basic side chain: lysine, arginine, and histidine; 6) Acidic side chain: aspartic acid and glutamic acid.

The number of amino acids of a "conservative amino acid substitution of VL, VH" may be about 1, about 2, about 3, about 4, about 5, about 6, about 8, about 9, about 10, about 11, about 13, about 14, about 15 conservative amino acid substitutions, or a range between any two of these values (inclusive) or any value therein. The number of amino acid substitutions of "conservative amino acid substitutions of a heavy or light chain" may be about 1, about 2, about 3, about 4, about 5, about 6, about 8, about 9, about 10, about 11, about 13, about 14, about 15, about 18, about 19, about 22, about 24, about 25, about 29, about 31, about 35, about 38, about 41, about 45 conservative amino acid substitutions, or a range between any two of these values (inclusive) or any value therein.

The term "encoding" when applied to a polynucleotide refers to a polynucleotide referred to as "encoding" a polypeptide, which polypeptide and/or fragment thereof may be produced by transcription and/or translation in its natural state or when manipulated by methods well known to those skilled in the art.

Antibodies, antigen binding fragments or derivatives of the disclosure include, but are not limited to, polyclonal, monoclonal, multispecific, fully human, humanized, primatized, chimeric antibodies, single chain antibodies, antigen binding fragments (e.g., fab ', F (ab') ₂ 、scFv)。

The term "recombinant" refers to a polypeptide or polynucleotide, meaning a form of the polypeptide or polynucleotide that does not exist in nature, and non-limiting examples can be combined to produce a polynucleotide or polypeptide that does not normally exist.

"homology" or "identity" or "similarity" refers to sequence similarity between two peptides or between two nucleic acid molecules. Homology can be determined by comparing the positions in each sequence that can be aligned. When a position in the compared sequences is occupied by the same base or amino acid, then the molecules are homologous at that position. The degree of homology between sequences is a function of the number of matched or homologous positions shared by the sequences.

"at least 80% identical" is about 80% identical, about 81% identical, about 82% identical, about 83% identical, about 85% identical, about 86% identical, about 87% identical, about 88% identical, about 90% identical, about 91% identical, about 92% identical, about 94% identical, about 95% identical, about 98% identical, about 99% identical, or a range between any two of these values (inclusive) or any value therein.

A nucleic acid or polynucleotide sequence (or polypeptide or antibody sequence) has a certain percentage (e.g., 90%, 95%, 98%, or 99%) of "identity" or "sequence identity" with another sequence, meaning that when the sequences are aligned, the percentage of bases (or amino acids) in the two sequences that are compared are identical. The percentage of alignment identity or sequence identity may be determined using visual inspection or software programs known in the art, such as the software program described in Ausubel et al eds. (2007) in Current Protocols in Molecular Biology. Preferably, the alignment is performed using default parameters. One such alignment program is BLAST using default parameters, such as BLASTN and BLASTP, both of which use the following default parameters: genetics code = standard; filter = none; strand = both; cutoff = 60; expect=10; matrix = BLOSUM62; descriptive = 50sequences; sortby=highscore; databases = non-redundants; genbank+embl+ddbj+pdb+genbank cdstransplations+swi ssprotein+spldate+pir. Biologically equivalent polynucleotides are those that have the indicated percent identity and encode polypeptides having the same or similar biological activity.

An "antibody" or "antigen-binding fragment" refers to a polypeptide or complex of polypeptides that specifically recognizes and binds an antigen. The antibody may be an intact antibody, any antigen-binding fragment thereof, or a single chain thereof. The term "antibody" thus includes any protein or peptide comprising at least a portion of an immunoglobulin molecule having biological activity for binding to an antigen in a molecule. Antibodies and antigen binding fragments include, but are not limited to, complementarity Determining Regions (CDRs), heavy chain variable regions (VH), light chain variable regions (VL), heavy chain constant regions (CH), light chain constant regions (CL), framework Regions (FR) or any portion thereof, or at least a portion of a binding protein, of a heavy chain or light chain or ligand binding portion thereof. The CDR regions include the CDR regions of the light chain (L CDR 1-3) and the heavy chain (HCDR 1-3).

The term "antibody" includes a wide variety of polypeptides that can be biochemically distinguished. Those skilled in the art will appreciate that the heavy chain classes include gamma, mu, alpha, delta or epsilon (γ, μ, α, δ, ε), some of which are also subclasses (e.g., γ1- γ4). The nature of this chain determines the "class" of antibody as IgG, igM, igA, igG or IgE, respectively. Immunoglobulin subclasses (isotypes), e.g., igG1, igG2, igG3, igG4, igG5, etc., have been well characterized and the functional specificities conferred are also known. All immunoglobulin classes are within the scope of the present disclosure. In some embodiments, the immunoglobulin molecule is an IgG class.

Light chains can be classified as kappa (kappa) or lambda (lambda). Each heavy chain may be associated with a kappa or lambda light chain. In general, when immunoglobulins are produced by hybridomas, B cells or genetically engineered host cells, the light and heavy chains thereof are bound by covalent bonds, and the "tail" portions of the two heavy chains are bound by covalent disulfide bonds or non-covalent bonds. In the heavy chain, the amino acid sequence extends from the N-terminus of the forked end of the Y-configuration to the C-terminus of the bottom of each chain. Immunoglobulin kappa light chain variable region vkappa; immunoglobulin lambda light chain variable region V _λ 。

The light chain variable region (VL) and the heavy chain variable region (VH) determine antigen recognition and specificity. The light chain constant region (CL) and the heavy chain constant region (CH) confer important biological properties such as secretion, transplacental movement, fc receptor binding, complement fixation, and the like. Conventionally, the numbering of constant regions increases as they become farther from the antigen binding site or amino terminus of an antibody. The N-terminal portion is a variable region and the C-terminal portion is a constant region; for example, the CH3 and CL domains of IgG1 actually comprise the carboxy-terminus of the heavy and light chains, respectively.

Where there are two or more definitions of terms used and/or accepted in the art, the definitions of terms used herein include all such meanings unless explicitly stated to the contrary. One specific example is the use of the term "complementarity determining regions" ("CDRs") to describe non-contiguous antigen binding sites found within the variable regions of heavy and light chain polypeptides. This particular region is described in Kabat et al, U.S. Dept. Of Health and Human Services, sequences of Proteins of Immunological Interest (1983) and Chothia et al, J.mol. Biol.196:901-917 (1987), which are incorporated herein by reference in their entirety.

CDRs defined according to Kabat and Chothia include overlapping or subsets of amino acid residues when compared to each other. Nevertheless, it is within the scope of the invention to apply either definition to refer to the CDRs of an antibody or variant thereof. The exact residue number comprising a particular CDR will vary depending on the sequence and size of the CDR. One skilled in the art can generally determine which specific residues a CDR comprises based on the variable region amino acid sequence of an antibody.

Kabat et al also define a numbering system for variable region sequences suitable for use with any antibody. The "Kabat numbering" system can be applied to any variable region sequence by one of ordinary skill in the art independent of other experimental data than the sequence itself. "Kabat numbering" refers to the numbering system set forth by Kabat et al, U.S. Dept. Ofhealth and Human Services at "Sequence ofProteins ofImmunological Interest" (1983). Antibodies may also use EU or Chothia numbering systems.

"treatment" refers to therapeutic treatment and prophylactic or preventative measures, with the object of preventing, slowing, ameliorating, and halting an undesirable physiological change or disorder, such as the progression of a disease, including but not limited to, a decrease in symptoms, a decrease in the degree of a disease, stabilization (i.e., not worsening) of a disease state, a delay or slowing of disease progression, an amelioration or palliation of a disease state, a decrease or disappearance (whether partial or total), an extension and an expected lifetime without treatment, and the like, whether detectable or undetectable. Patients in need of treatment include those already with the condition or disorder, those prone to the condition or disorder, or those in need of prophylaxis of the condition or disorder, for whom the administration of the disclosed antibodies or compositions for detection, diagnostic procedures, and/or treatment would be expected to benefit.

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

"about" refers to a conventional error range of corresponding numerical values as readily known to one of ordinary skill in the relevant art. In some embodiments, references herein to "about" refer to the values described and ranges thereof of ± 10%, ± 5% or ± 1%.

An "effective amount" refers to an amount of an active compound or agent that is capable of eliciting a biological or medical response in a tissue, system, animal, individual or human.

As used herein, "in need of" means that the patient has been identified as in need of a particular method or treatment. In some embodiments, the identification may be performed by any diagnostic means.

Preparation of antibodies

The DNA encoding the antibody may be synthesized according to the amino acid sequence design of the antibody described herein by conventional methods, placed into an expression vector, and then transfected into a host cell, and the transfected host cell is cultured in a medium to produce monoclonal antibodies. In some embodiments, the expression antibody vector comprises at least one promoter element, an antibody coding sequence, a transcription termination signal, and a polyA tail. Other elements include enhancers, kozak sequences, and donor and acceptor sites for RNA splicing flanking the insertion. Efficient transcription can be obtained by the early and late promoters of SV40, the long terminal repeats from retroviruses such as the early promoters of RSV, HTLV1, HIVI and cytomegalovirus, and other cellular promoters such as actin promoters may be used. Suitable expression vectors may include pIRES1neo, pRetro-Off, pRetro-On, PLXSN, or pLNCX, pcDNA3.1 (+/-), pcDNA/Zeo (+/-), pcDNA3.1/Hygro (+/-), PSVL, PMSG, pRSVcat, pSV dhfr, pBC12MI, pCS2, and the like. Commonly used host cells include HEK293 cells, cos1 cells, cos7 cells, CV1 cells, murine L cells, CHO cells, and the like.

All publications, patents, and patent applications cited herein are incorporated by reference in their entirety for all purposes.

Detailed Description

The technical solution of the present invention is further illustrated by the following specific examples, which do not represent limitations on the scope of the present invention. Some insubstantial modifications and adaptations of the invention based on the inventive concept by others remain within the scope of the invention.

Materials, reagents and the like used in the examples described below are commercially available unless otherwise specified.

Example 1: preparation of antibodies

Cloning the heavy chain and light chain DNA sequences of the antibody onto an expression vector, transferring the antibody into a host cell, culturing and purifying to obtain the antibody.

The amino acid sequences OF antibody h10D8OF, antibody h10D8OFKF, antibody 1, antibody Anti-mCTLA4, antibody Anti-mTIGIT and control antibody IgG1 are shown in tables 1-5, wherein the amino acid sequences OF antibody h10D8OF and h10D8OFKF are identical. Wherein the host cells expressing antibody h10D8OFKF and antibody 1 are CHO-K1 cells with the α - (1, 6) -fucosyltransferase gene (Fut-8) knocked out, the resulting antibody h10D8OFKF has a fucosylation level of about 0%, antibody 1 has a fucosylation level of about 0%, a total high mannose glycoform of <2%, and a total sialylation glycoform of <1%. The host cell expressing antibody h10D8OF was a CHO-K1 cell. The host cells expressing antibody Anti-mCTLA4, antibody Anti-mTIGAT and control antibody IgG1 were HEK293F cells.

TABLE 1 amino acid sequences OF antibodies h10D8OF and h10D8OFKF

TABLE 2 amino acid sequence of antibody 1

TABLE 3 amino acid sequence of antibody Anti-mCTLA4

TABLE 4 amino acid sequence of antibodies Anti-mTIGAT

TABLE 5 amino acid sequence of control antibody IgG1

Example 2: anti-mTIGAT and Anti-mCTLA4 combined drug efficacy test

The test evaluates the therapeutic effect of the test substance on Balb/C mice subcutaneous tumor-bearing CT26 cells. CT26 colon cancer cells (1X 10 in number) ⁶ /v) when the average tumor volume of the mice reached about 221mm ³ The time of the group and administration was carried out, and the group was recorded as day 0, i.e., the first administration was started, the administration modes (administration frequency, period, route) and the specific group administration schedule are shown in Table 6. Mice body weight and tumor volume were recorded during the course of the experiment, 2 times per week, and tumor inhibition TGI% and p-values were calculated, with results expressed as mean ± standard error. When the tumor volume of the mice is more than 3000mm ³ Euthanasia was performed above.

Table 6 modes of administration and group dosing regimens

Note that: BIW: 2 times per week; i.p.: intraperitoneal injection

The tumor growth curve of the mice is shown in FIG. 1, and the survival curve of the mice is shown in FIG. 2. Both the single (groups 2 and 3) and the combination (group 4) had some degree of tumor growth inhibition, and the groups did not show significant differences at 14 days post-dose. However, with prolonged treatment time, mice with tumor volume increase reaching the euthanasia standard were euthanized, and the survival curves of the mice revealed that the survival time of mice in the combination administration group (group 4) was significantly higher than that of the negative control group (group 1) and the single administration group (group 2 and group 3).

Example 3: drug efficacy test of antibody h10D8OFKF and antibody 1 combined administration

The test evaluates the anti-tumor effect of the test object in CT26 colon cancer cell subcutaneously transplanted BALB/c/hTIGIT/hCTLA4 mice (JieXtensikang, strain number: T051156), and explores the combined administration effect of the antibody h10D8OFKF and the antibody 1.

CT26 cells were inoculated subcutaneously in 1X 10 mice ⁶ When the average tumor volume of the mice was 88.33mm ³ The time of administration and the administration were grouped and the day of the grouping was recorded as day 0, i.e., the first administration was started, and the administration modes (administration frequency, period, route) and specific grouping administration schemes are shown in Table 7. Mice body weight and tumor volume were recorded during the course of the experiment, 2 times per week, and tumor inhibition TGI% and p-values were calculated, with results expressed as mean ± standard error. When the tumor volume of the mice is more than 3000mm ³ Euthanasia was performed above.

Table 7 modes of administration and group dosing regimens

Note that: BIW: 2 times per week; i.p.: intraperitoneal injection

The tumor growth curve of the mice is shown in FIG. 3, and the survival curve of the mice is shown in FIG. 4. The study results showed that the test antibody 1, antibody h10D8OFKF, antibody 1 and antibody h10D8OFKF all showed significant tumor inhibition (see fig. 3) in combination, and significantly prolonged the survival of mice (see fig. 4) compared to the control antibody IgG 1. The combination of antibody 1 with antibody h10D8OFKF (group 4) showed significant tumor inhibition (×p < 0.05) and significantly prolonged survival of mice (×p < 0.01) compared to the alone (group 3) of antibody h10D8 OFKF.

Claims

1. A combination comprising an anti-TIGIT antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment; the anti-TIGIT antibody or antigen-binding fragment comprises HCDR1 shown in SEQ ID No. 1, HCDR2 shown in SEQ ID No. 2, HCDR3 shown in SEQ ID No. 3, LCDR1 shown in SEQ ID No. 4, LCDR2 shown in SEQ ID No. 5 and LCDR3 shown in SEQ ID No. 6.

2. The combination of claim 1, wherein the anti-TIGIT 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 shown in SEQ ID NO. 7, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID NO. 7, or an amino acid sequence having one or more conservative amino acid substitutions to the sequence shown in SEQ ID NO. 7; and/or

The light chain variable region comprises the amino acid sequence shown in SEQ ID NO. 8, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID NO. 8, or an amino acid sequence having one or more conservative amino acid substitutions as compared to the sequence shown in SEQ ID NO. 8.

3. The combination of claim 1 or 2, wherein the heavy chain of the anti-TIGIT antibody comprises the amino acid sequence shown in SEQ ID No. 9, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID No. 9, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID No. 9; and/or

The light chain of the anti-TIGIT antibody comprises the amino acid sequence shown in SEQ ID No. 10, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID No. 10, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID No. 10.

4. The combination according to any one of claims 1 to 3, wherein the anti-CTLA-4 antibody or antigen-binding fragment comprises HCDR1 shown in SEQ ID No. 11, HCDR2 shown in SEQ ID No. 12, HCDR3 shown in SEQ ID No. 13, LCDR1 shown in SEQ ID No. 14, LCDR2 shown in SEQ ID No. 15 and LCDR3 shown in SEQ ID No. 16.

5. The combination of any one of claims 1-4, wherein 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 shown in SEQ ID NO. 17, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID NO. 17, or an amino acid sequence having one or more conservative amino acid substitutions as compared to the sequence shown in SEQ ID NO. 17; and/or

The light chain variable region comprises the amino acid sequence shown in SEQ ID NO. 18, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID NO. 18, or an amino acid sequence having one or more conservative amino acid substitutions as compared to the sequence shown in SEQ ID NO. 18.

6. The combination according to any one of claims 1 to 5, wherein the heavy chain of the anti-CTLA-4 antibody comprises the amino acid sequence of SEQ ID No. 19, or an amino acid sequence having at least 80% identity to the sequence of SEQ ID No. 19, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence of SEQ ID No. 19; and/or

The light chain of the anti-CTLA-4 antibody comprises an amino acid sequence shown as SEQ ID NO. 20, or an amino acid sequence which has at least 80% identity with the sequence shown as SEQ ID NO. 20, or an amino acid sequence which has one or more conservative amino acid substitutions with the sequence shown as SEQ ID NO. 20.

7. The combination according to any one of claims 1-6, wherein the mass ratio of the anti-TIGIT antibody or antigen-binding fragment to the anti-CTLA-4 antibody or antigen-binding fragment is 1-50:1; or 5-25:1; or about 5:1; or about 25:1.

8. A method of treating a tumor, comprising: administering to a patient in need thereof an effective amount of an anti-TIGIT antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment; the anti-TIGIT antibody or antigen-binding fragment comprises HCDR1 shown in SEQ ID No. 1, HCDR2 shown in SEQ ID No. 2, HCDR3 shown in SEQ ID No. 3, LCDR1 shown in SEQ ID No. 4, LCDR2 shown in SEQ ID No. 5 and LCDR3 shown in SEQ ID No. 6.

9. The method of claim 8, wherein the anti-TIGIT antibody or antigen-binding fragment comprises a heavy chain variable region and a light chain variable region, wherein:

10. The method of claim 8 or 9, wherein the heavy chain of the anti-TIGIT antibody comprises the amino acid sequence shown in SEQ ID No. 9, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID No. 9, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID No. 9; and/or

11. The method of any one of claims 8-10, wherein the anti-CTLA-4 antibody or antigen-binding fragment comprises HCDR1 as set forth in SEQ ID No. 11, HCDR2 as set forth in SEQ ID No. 12, HCDR3 as set forth in SEQ ID No. 13, LCDR1 as set forth in SEQ ID No. 14, LCDR2 as set forth in SEQ ID No. 15, and LCDR3 as set forth in SEQ ID No. 16.

12. The method of any one of claims 8-11, wherein the anti-CTLA-4 antibody or antigen-binding fragment comprises a heavy chain variable region and a light chain variable region, wherein:

13. The method of any one of claims 8-12, wherein the heavy chain of the anti-CTLA-4 antibody comprises the amino acid sequence of SEQ ID No. 19, or an amino acid sequence having at least 80% identity to the sequence of SEQ ID No. 19, or an amino acid sequence having one or more conservative amino acid substitutions as compared to the sequence of SEQ ID No. 19; and/or

14. The method of any one of claims 8-13, wherein the mass ratio of the anti-TIGIT antibody or antigen-binding fragment to the anti-CTLA-4 antibody or antigen-binding fragment is 1-50:1; or 5-25:1; or about 5:1; or about 25:1.

15. The method of any one of claims 8-14, wherein the anti-TIGIT antibody or antigen-binding fragment is administered at a dose of 0.01mg/kg to 20mg/kg or 0.05mg to 1200mg and the anti-CTLA-4 antibody or antigen-binding fragment is administered at a dose of 0.01mg/kg to 10mg/kg or 0.05mg to 600mg.

16. The method of any one of claims 8-15, wherein the anti-TIGIT antibody or antigen-binding fragment and/or anti-CTLA-4 antibody or antigen-binding fragment is administered once daily to once every 7 weeks; alternatively, the dosing frequency is once daily, three times weekly, twice weekly, once every 2 weeks, once every 3 weeks, once every 4 weeks, once every 5 weeks, once every 6 weeks, or once every 7 weeks.

17. The method of any one of claims 8-16, wherein the tumor is a benign tumor or cancer; alternatively, the tumor is a hematological cancer or a solid tumor; alternatively, the tumor is selected from acute lymphoblastic leukemia, acute myelogenous leukemia, chronic lymphocytic leukemia, chronic myelogenous leukemia, myeloproliferative diseases/tumors, hodgkin's lymphoma, non-hodgkin's lymphoma, burkitt's lymphoma, follicular lymphoma, multiple myeloma, giant cell myeloma, heavy chain myeloma, light chain or present-jones myeloma, breast cancer, pancreatic cancer, prostate cancer, melanoma, head and neck cancer, liver cancer, kidney cancer, squamous cell cancer, esophageal squamous cell cancer, lung cancer, non-small cell lung cancer, cervical cancer, esophageal cancer, endometrial cancer, ovarian cancer, multiple myeloma, colon cancer, colorectal cancer, urothelial cancer, bladder cancer, brain cancer.

18. A pharmaceutical combination for treating a tumor comprising an anti-TIGIT antibody or antigen-binding fragment and an anti-CTLA-4 antibody or antigen-binding fragment; the anti-TIGIT antibody or antigen-binding fragment comprises HCDR1 shown in SEQ ID No. 1, HCDR2 shown in SEQ ID No. 2, HCDR3 shown in SEQ ID No. 3, LCDR1 shown in SEQ ID No. 4, LCDR2 shown in SEQ ID No. 5 and LCDR3 shown in SEQ ID No. 6.

19. The pharmaceutical combination of claim 18, wherein the anti-TIGIT antibody or antigen-binding fragment comprises a heavy chain variable region and a light chain variable region, wherein:

20. The pharmaceutical combination of claim 18 or 19, wherein the heavy chain of the anti-TIGIT antibody comprises the amino acid sequence shown in SEQ ID No. 9, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID No. 9, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID No. 9; and/or

21. The pharmaceutical combination of any one of claims 18-20, wherein the anti-CTLA-4 antibody or antigen-binding fragment comprises HCDR1 as set forth in SEQ ID No. 11, HCDR2 as set forth in SEQ ID No. 12, HCDR3 as set forth in SEQ ID No. 13, LCDR1 as set forth in SEQ ID No. 14, LCDR2 as set forth in SEQ ID No. 15, and LCDR3 as set forth in SEQ ID No. 16.

22. The pharmaceutical combination of any one of claims 18-21, wherein the anti-CTLA-4 antibody or antigen-binding fragment comprises a heavy chain variable region and a light chain variable region, wherein:

23. The pharmaceutical combination of any one of claims 18-22, wherein the heavy chain of the anti-CTLA-4 antibody comprises the amino acid sequence of SEQ ID No. 19, or an amino acid sequence having at least 80% identity to the sequence of SEQ ID No. 19, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence of SEQ ID No. 19; and/or

24. The pharmaceutical combination of any one of claims 18-23, wherein the mass ratio of the anti-TIGIT antibody or antigen-binding fragment to the anti-CTLA-4 antibody or antigen-binding fragment is 1-50:1; or 5-25:1; or about 5:1; or about 25:1.

25. Use of a combination according to any one of claims 1 to 7 for the manufacture of a medicament for the treatment of a tumour.

26. Use of a combination according to any one of claims 1 to 7 or a pharmaceutical combination according to any one of claims 18 to 24 for the treatment of a tumor.

27. The use of claim 25 or 26, wherein the tumor is a benign tumor or cancer; alternatively, the tumor is a hematological cancer or a solid tumor; alternatively, the tumor is selected from acute lymphoblastic leukemia, acute myelogenous leukemia, chronic lymphocytic leukemia, chronic myelogenous leukemia, myeloproliferative diseases/tumors, hodgkin's lymphoma, non-hodgkin's lymphoma, burkitt's lymphoma, follicular lymphoma, multiple myeloma, giant cell myeloma, heavy chain myeloma, light chain or present-jones myeloma, breast cancer, pancreatic cancer, prostate cancer, melanoma, head and neck cancer, liver cancer, kidney cancer, squamous cell cancer, esophageal squamous cell cancer, lung cancer, non-small cell lung cancer, cervical cancer, esophageal cancer, endometrial cancer, ovarian cancer, multiple myeloma, colon cancer, colorectal cancer, urothelial cancer, bladder cancer, brain cancer.

28. A kit comprising an anti-TIGIT antibody and a package insert comprising instructions for using the anti-TIGIT antibody in combination with an anti-CTLA-4 antibody to treat a tumor.

29. A kit comprising an anti-CTLA-4 antibody and a package insert comprising instructions for using the anti-CTLA-4 antibody in combination with an anti-TIGIT antibody to treat a tumor.

30. A kit comprising an anti-TIGIT antibody and an anti-CTLA-4 antibody agent, and a package insert comprising instructions for using the anti-TIGIT antibody and anti-CTLA-4 antibody agent to treat a tumor.

31. A kit comprising an anti-TIGIT antibody agent and a package insert comprising instructions for using the anti-TIGIT antibody agent in combination with an anti-CTLA-4 antibody to treat a tumor.

32. A kit comprising an anti-CTLA-4 antibody agent and a package insert comprising instructions for using the anti-CTLA-4 antibody agent in combination with an anti-TIGIT antibody to treat a tumor.

33. The kit of any one of claims 28-32, wherein the anti-TIGIT antibody or antigen-binding fragment comprises HCDR1 shown in SEQ ID No. 1, HCDR2 shown in SEQ ID No. 2, HCDR3 shown in SEQ ID No. 3, LCDR1 shown in SEQ ID No. 4, LCDR2 shown in SEQ ID No. 5, and LCDR3 shown in SEQ ID No. 6; preferably, the anti-TIGIT antibody or antigen-binding fragment comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises an amino acid sequence set forth in SEQ ID No. 7, or an amino acid sequence having at least 80% identity to the sequence set forth in SEQ ID No. 7, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence set forth in SEQ ID No. 7; and/or the light chain variable region comprises the amino acid sequence shown in SEQ ID NO. 8, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID NO. 8, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID NO. 8; preferably, the heavy chain of the anti-TIGIT antibody comprises the amino acid sequence shown in SEQ ID No. 9, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID No. 9, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID No. 9; and/or the light chain of the anti-TIGIT antibody comprises the amino acid sequence shown in SEQ ID No. 10, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID No. 10, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID No. 10.

34. The kit of any one of claims 28-33, wherein the anti-CTLA-4 antibody or antigen-binding fragment comprises HCDR1 shown in SEQ ID No. 11, HCDR2 shown in SEQ ID No. 12, HCDR3 shown in SEQ ID No. 13, LCDR1 shown in SEQ ID No. 14, LCDR2 shown in SEQ ID No. 15, and LCDR3 shown in SEQ ID No. 16; preferably, 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 set forth in SEQ ID No. 17, or an amino acid sequence having at least 80% identity to the sequence set forth in SEQ ID No. 17, or an amino acid sequence having one or more conservative amino acid substitutions as compared to the sequence set forth in SEQ ID No. 17; and/or the light chain variable region comprises the amino acid sequence shown in SEQ ID NO. 18, or an amino acid sequence having at least 80% identity to the sequence shown in SEQ ID NO. 18, or an amino acid sequence having one or more conservative amino acid substitutions compared to the sequence shown in SEQ ID NO. 18; preferably, the heavy chain of the anti-CTLA-4 antibody comprises the amino acid sequence of SEQ ID NO. 19, or an amino acid sequence having at least 80% identity to the sequence of SEQ ID NO. 19, or an amino acid sequence having one or more conservative amino acid substitutions as compared to the sequence of SEQ ID NO. 19; and/or the light chain of the anti-CTLA-4 antibody comprises an amino acid sequence as set forth in SEQ ID No. 20, or an amino acid sequence having at least 80% identity to the sequence set forth in SEQ ID No. 20, or an amino acid sequence having one or more conservative amino acid substitutions as compared to the sequence set forth in SEQ ID No. 20.