WO2022184068A1 - 抗tigit抗体在治疗肿瘤或癌症中的应用 - Google Patents

抗tigit抗体在治疗肿瘤或癌症中的应用 Download PDF

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WO2022184068A1
WO2022184068A1 PCT/CN2022/078683 CN2022078683W WO2022184068A1 WO 2022184068 A1 WO2022184068 A1 WO 2022184068A1 CN 2022078683 W CN2022078683 W CN 2022078683W WO 2022184068 A1 WO2022184068 A1 WO 2022184068A1
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seq
antigen
amino acid
antibody
binding fragment
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WO2022184068A9 (zh
WO2022184068A8 (zh
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黄俊杰
徐振前
黄贤明
俞金泉
李胜峰
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百奥泰生物制药股份有限公司
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/28—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
    • C07K16/2803—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against the immunoglobulin superfamily
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/395—Antibodies; Immunoglobulins; Immune serum, e.g. antilymphocytic serum
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/02—Antineoplastic agents specific for leukemia
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/28—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/505—Medicinal preparations containing antigens or antibodies comprising antibodies

Definitions

  • the invention belongs to the field of biomedicine, and particularly relates to the application of anti-TIGIT antibodies or antigen-binding fragments in the treatment of tumors or cancers.
  • TIGIT T cell immunoreceptor with Ig and ITIM domains
  • Ig immunoglobulin
  • ITIM tyrosine inhibitor motif
  • TIGIT is part of a co-stimulatory network consisting mainly of the activating receptor CD226 and the inhibitory receptor TIGIT on T cells, and the ligand CD155 (also known as the ligand CD155 expressed on the surface of APCs, tumor cells, infected cells) PVR, a poliovirus receptor protein encoded in humans by the PVR gene) and CD112. Binding of TIGIT to PVR or CD112 will lead to the phosphorylation of Tyr225 in the cytoplasm of TIGIT, and the binding of TIGIT to cell-adaptive growth factor receptor binding protein 2 (GRB2).
  • ligand CD155 also known as the ligand CD155 expressed on the surface of APCs, tumor cells, infected cells
  • PVR a poliovirus receptor protein encoded in humans by the PVR gene
  • GRB2 can recruit SHIP1 to inhibit phosphatidylinositol tri-kinase (PI3K) and mitogen-activated protein kinase (MAPK) signaling.
  • PI3K phosphatidylinositol tri-kinase
  • MAPK mitogen-activated protein kinase
  • phosphorylated TIGIT recruits SHIP1 via beta arrestin 2 ( ⁇ -arrestin2) and by blocking autoubiquitination of TNF receptor-associated factor 6 (TRAF6) and disrupts nuclear factor KB (NF-KB) activation , a series of signal transduction eventually lead to the inhibition of T cell or NK cell function and the inhibition of cytokine production.
  • PVR is both a ligand of TIGIT and a ligand of CD226 molecule.
  • Ser329 and Tyr322 of the intracellular domain of CD226 are phosphorylated; Ser329 phosphorylation promotes the activation of protein kinase (PKC) and the mutual binding of CD226 to lymphocyte-associated antigen 1 (LFA1). LFA1 is then used for TYN-mediated phosphorylation of Tyr322 and CD226-mediated downstream signaling. A series of signal transduction finally leads to the activation of T cell or NK cell function, which promotes the production of cytokines.
  • PDC protein kinase
  • LFA1 lymphocyte-associated antigen 1
  • TIGIT molecules can directly disrupt the formation of normal dimers of CD226, thereby destroying the normal physiological function of CD226.
  • TIGIT and CD226 are like two ends of the balance, through the pivot point of PVR, they skillfully regulate the immune function of the body through the transduction of co-stimulatory and co-inhibitory signals.
  • TIGIT is a very potential inhibitory target in tumor immunotherapy, which can provide a new means for tumor immunotherapy.
  • the present invention discloses methods or uses of anti-TIGIT antibodies or antigen-binding fragments for treating tumors or cancers.
  • the anti-TIGIT antibody or antigen-binding fragment agent is used to treat tumors or cancer.
  • the anti-TIGIT antibody or antigen-binding fragment comprises at least HCDR1 set forth in SEQ ID NO:1, HCDR2 set forth in SEQ ID NO:2, HCDR3 set forth in SEQ ID NO:3, : one or more of LCDR1 shown in SEQ ID NO: 5, LCDR2 shown in SEQ ID NO: 6, and LCDR3 shown in SEQ ID NO: 6.
  • the anti-TIGIT antibody or antigen-binding fragment comprises HCDR1 set forth in SEQ ID NO:1, HCDR2 set forth in SEQ ID NO:2, HCDR3 set forth in SEQ ID NO:3, SEQ ID NO:3 LCDR1 shown in 4, LCDR2 shown in SEQ ID NO:5 and LCDR3 shown in SEQ ID NO:6.
  • the heavy chain variable region of the anti-TIGIT antibody or antigen-binding fragment comprises the amino acid sequence set forth in SEQ ID NO:7, or is at least 80% identical to the sequence set forth in SEQ ID NO:7
  • the amino acid sequence of SEQ ID NO: 7 has 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 set forth in SEQ ID NO:8, or is at least 80% identical to the sequence set forth in SEQ ID NO:8
  • the amino acid sequence of SEQ ID NO: 8 has one or more conservative amino acid substitutions.
  • the heavy chain variable region of the anti-TIGIT antibody or antigen-binding fragment comprises the amino acid sequence set forth in SEQ ID NO:7, or is at least 80% identical to the sequence set forth in SEQ ID NO:7
  • the light chain variable region of the anti-TIGIT antibody or antigen-binding fragment comprises SEQ ID NO: 8
  • the heavy chain variable region of the anti-TIGIT antibody or antigen-binding fragment comprises the amino acid sequence set forth in SEQ ID NO:7
  • the light chain variable region of the anti-TIGIT antibody or antigen-binding fragment comprises SEQ ID NO: 7 The amino acid sequence shown in ID NO:8.
  • the heavy chain of the anti-TIGIT antibody comprises the amino acid sequence set forth in SEQ ID NO:9, or an amino acid sequence that is at least 80% identical to the sequence set forth in SEQ ID NO:9, or The sequence shown in SEQ ID NO: 9 is compared to an amino acid sequence with one or more conservative amino acid substitutions.
  • the light chain of the anti-TIGIT antibody comprises the amino acid sequence set forth in SEQ ID NO: 10, or an amino acid sequence that is at least 80% identical to the sequence set forth in SEQ ID NO: 10, or is The sequence shown in SEQ ID NO: 10 is compared to the amino acid sequence with one or more conservative amino acid substitutions.
  • the heavy chain of the anti-TIGIT antibody comprises the amino acid sequence set forth in SEQ ID NO:9, or an amino acid sequence that is at least 80% identical to the sequence set forth in SEQ ID NO:9, or The amino acid sequence with 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 the amino acid sequence shown in SEQ ID NO:10.
  • the anti-TIGIT antibody is antibody h10D8OF or h10D8OFKF
  • the heavy chains of antibodies h10D8OF and h10D8OFKF comprise the amino acid sequence set forth in SEQ ID NO:9
  • the light chains of antibodies h10D8OF and h10D8OFKF comprise as shown in SEQ ID NO:
  • the amino acid sequences shown in 10; the antibodies h10D8OF and h10D8OFKF contain two heavy chains with identical sequences and two light chains with identical sequences, respectively.
  • the anti-TIGIT antibody eg, antibody h10D8OFKF
  • antigen-binding fragment has a fucosylation level of 0-10%. In some embodiments, the anti-TIGIT antibody (eg, antibody h10D8OFKF) or antigen-binding fragment has a fucosylation level of 0-5%.
  • the anti-TIGIT antibody (eg, antibody h10D8OFKF) 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 any two of these values range between (including the endpoints) or any value therein.
  • the anti-TIGIT antibody (eg, antibody h10D8OFKF) or antigen-binding fragment does not bind fucose.
  • the anti-TIGIT antibody (eg, antibody h10D8OFKF) or antigen-binding fragment has an enhanced ADCC effect (antibody-dependent cell-mediated cytotoxicity).
  • Anti-TIGIT antibodies or antigen-binding fragments can be expressed in CHO cells or HEK293 cells by genetic engineering and obtained by purification; purification can be carried out by conventional methods, such as centrifuging the cell suspension and collecting the supernatant, and centrifuging again to further remove impurities. Methods such as ProteinA affinity columns and ion exchange columns can be used to purify antibody proteins.
  • the hypofucosylated or afucosylated anti-TIGIT antibody or antigen-binding fragment is expressed by an alpha-(1,6)-fucosyltransferase knockout cell line.
  • the antibody h10D8OFKF is expressed by an ⁇ -(1,6)-fucosyltransferase knockout cell line, eg, an ⁇ -(1,6)-fucosyltransferase knockout CHO cells.
  • the method or use comprises administering to a patient in need thereof an effective amount of an anti-TIGIT antibody or antigen-binding fragment.
  • the anti-TIGIT antibody is the antibody h10D8OF or h10D8OFKF.
  • the anti-TIGIT antibody or antigen-binding fragment is administered in an effective dose of about 9 mg to 1200 mg per dose.
  • the patient has a tumor or cancer.
  • tumors and cancers include, but are not limited to, hematological cancers, solid tumors.
  • hematological cancers include, but are not limited to, leukemia, lymphoma, and myeloma.
  • the leukemia includes acute lymphocytic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), chronic myelogenous leukemia (CML), and myeloproliferative disorders/neoplastics (MPDS) ).
  • ALL acute lymphocytic leukemia
  • AML acute myeloid leukemia
  • CLL chronic lymphocytic leukemia
  • CML chronic myelogenous leukemia
  • MPDS myeloproliferative disorders/neoplastics
  • lymphomas include Hodgkin's lymphoma, indolent and aggressive non-Hodgkin's lymphoma, Burkitt's lymphoma, and follicular lymphoma (small cell and large cell).
  • the myeloma includes multiple myeloma (MM), giant cell myeloma, heavy chain myeloma, and light chain or Bence-Jones myeloma.
  • solid tumors include breast cancer, ovarian cancer, pancreatic cancer, prostate cancer, melanoma, colorectal cancer, colon cancer, lung cancer, head and neck cancer, bladder cancer, esophageal cancer, liver cancer, and kidney cancer.
  • the tumor and cancer are pathologically confirmed locally advanced or metastatic malignant solid tumors for which there is no effective treatment.
  • the present invention discloses a method for treating a tumor or cancer in a patient in need thereof, comprising administering an effective amount of an anti-TIGIT antibody or antigen-binding fragment, wherein the administration of the anti-TIGIT antibody or antigen-binding fragment is effective
  • the amount is about 9 mg to 1200 mg per treatment cycle.
  • a treatment cycle is 1 week, 2 weeks, 3 weeks, 4 weeks, 1 month, 5 weeks, 6 weeks, 7 weeks, or a range between any two of these values (including endpoint) or any of its values.
  • the anti-TIGIT antibody is the antibody h10D8OF or h10D8OFKF.
  • anti-TIGIT antibodies can be formulated into pharmaceutical compositions and administered to patients in a variety of forms suitable for the chosen route of administration, eg, parenteral, intravenous (iv), intramuscular , topical or subcutaneous.
  • the anti-TIGIT antibody can be infused intravenously.
  • the amount of anti-TIGIT antibody administered will depend on the nature of the drug, the extent to which internalization, transport, and release of the drug is triggered on the cell surface, as well as the disease being treated and the condition of the patient (eg, age, sex, weight, etc.).
  • the anti-TIGIT antibody eg, antibody h10D8OF or h10D8OFKF
  • antigen-binding fragment per administration is 0.01 mg/kg to 26 mg/kg or a formulation containing such doses of anti-TIGIT antibody or antigen-binding fragment.
  • the anti-TIGIT antibody or antigen-binding fragment is about 0.01 mg/kg, about 0.02 mg/kg, about 0.03 mg/kg, about 0.06 mg/kg, about 0.08 mg/kg, about 0.1 mg/kg per administration mg/kg, about 0.2 mg/kg, about 0.3 mg/kg, about 0.5 mg/kg, about 0.9 mg/kg, about 1 mg/kg, about 1.6 mg/kg, about 2 mg/kg, about 2.5 mg/kg, About 3 mg/kg, about 4 mg/kg, about 5 mg/kg, about 6 mg/kg, about 7 mg/kg, about 8 mg/kg, about 9 mg/kg, about 10 mg/kg, about 11 mg/kg, about 12 mg/kg, About 13mg/kg, about 14mg/kg, about 15mg/kg, about 16mg/kg, about 17mg/kg, about 18mg/kg, about 19mg/kg, about 20mg/kg, about 21mg/kg, about 22mg/kg, about 23 mg/kg
  • the present invention discloses a method for treating a tumor or cancer, the method comprising: administering to a patient in need thereof about 0.01 mg/kg, about 0.02 mg/kg, about 0.03 mg every 3 weeks /kg, about 0.06 mg/kg, about 0.08 mg/kg, about 0.1 mg/kg, about 0.2 mg/kg, about 0.3 mg/kg, about 0.5 mg/kg, about 0.9 mg/kg, about 1 mg/kg, About 1.6 mg/kg, about 2 mg/kg, about 2.5 mg/kg, about 3 mg/kg, about 4 mg/kg, about 5 mg/kg, about 6 mg/kg, about 7 mg/kg, about 8 mg/kg, about 9 mg/kg kg, about 10 mg/kg, about 11 mg/kg, about 12 mg/kg, about 13 mg/kg, about 14 mg/kg, about 15 mg/kg, about 16 mg/kg, about 17 mg/kg, about 18 mg/kg, about 19 mg/kg kg, about 20 mg/kg, about 21 mg/kg, about 22 mg/kg
  • the present invention discloses a method for treating a tumor or cancer, the method comprising: administering to a patient in need thereof about 0.01 mg/kg, about 0.02 mg/kg, about 0.03 mg every 3 weeks /kg, about 0.06 mg/kg, about 0.08 mg/kg, about 0.1 mg/kg, about 0.2 mg/kg, about 0.3 mg/kg, about 0.5 mg/kg, about 0.9 mg/kg, about 1 mg/kg, About 1.6 mg/kg, about 2 mg/kg, about 2.5 mg/kg, about 3 mg/kg, about 4 mg/kg, about 5 mg/kg, about 6 mg/kg, about 7 mg/kg, about 8 mg/kg, about 9 mg/kg kg, about 10 mg/kg, about 11 mg/kg, about 12 mg/kg, about 13 mg/kg, about 14 mg/kg, about 15 mg/kg, about 16 mg/kg, about 17 mg/kg, about 18 mg/kg, about 19 mg/kg kg, about 20 mg/kg, about 21 mg/kg, about 22 mg/kg
  • the present invention discloses a method for treating a tumor or cancer, the method comprising: administering to a patient in need thereof about 0.01 mg/kg, about 0.02 mg/kg, about 0.03 mg every 3 weeks /kg, about 0.06 mg/kg, about 0.08 mg/kg, about 0.1 mg/kg, about 0.2 mg/kg, about 0.3 mg/kg, about 0.5 mg/kg, about 0.9 mg/kg, about 1 mg/kg, About 1.6 mg/kg, about 2 mg/kg, about 2.5 mg/kg, about 3 mg/kg, about 4 mg/kg, about 5 mg/kg, about 6 mg/kg, about 7 mg/kg, about 8 mg/kg, about 9 mg/kg kg, about 10 mg/kg, about 11 mg/kg, about 12 mg/kg, about 13 mg/kg, about 14 mg/kg, about 15 mg/kg, about 16 mg/kg, about 17 mg/kg, about 18 mg/kg, about 19 mg/kg kg, about 20 mg/kg, about 21 mg/kg, about 22 mg/kg
  • the present invention discloses a method of treating a tumor or cancer, comprising administering to a patient in need thereof an effective amount of an anti-TIGIT antibody or antigen-binding fragment (or formulation); wherein the anti-TIGIT antibody or antigen binds
  • An effective amount of the fragment is 9 mg to 1200 mg in a single administration (or a formulation containing this dose of the anti-TIGIT antibody or antigen-binding fragment).
  • Dosage schedule and mode of administration depend on the benefit-risk assessment of the anti-TIGIT antibody or antigen-binding fragment (or preparation) in certain patient populations and general clinical practice guidelines.
  • the effective amount of anti-TIGIT antibody or antigen-binding fragment administered to a patient per treatment cycle is 9 mg to 1200 mg (or a formulation containing such dose of anti-TIGIT antibody or antigen-binding fragment).
  • the patient is administered an effective amount of an anti-TIGIT antibody (eg, antibody h10D8OF or h10D8OFKF) or antigen-binding fragment per treatment cycle of about 9 mg, about 10 mg, about 18 mg, about 26 mg, about 30 mg, about 38 mg, about 45mg, about 60mg, about 70mg, about 80mg, about 100mg, about 120mg, about 180mg, about 200mg, about 250mg, about 290mg, about 300mg, about 330mg, about 380mg, about 400mg, about 434mg, about 480mg, about 500mg, about 567 mg, about 580 mg, about 600 mg, about 700 mg, about 800 mg, about 900 mg, or a range between any two of these values (including endpoints) or any value therein, or containing such a dose of an anti-TIGIT antibody or antigen-binding fragment preparation.
  • an anti-TIGIT antibody eg, antibody h10D
  • one treatment cycle is administered once every 1 week to 7 weeks.
  • the effective amount of anti-TIGIT antibody or antigen-binding fragment administered per treatment cycle is about 10 mg to 900 mg, or a formulation containing such dose of anti-TIGIT antibody or antigen-binding fragment; wherein one treatment cycle is about 1 Week, about 2 weeks, about 3 weeks, about 4 weeks, about 5 weeks, about 6 weeks, about 7 weeks, or a range (including endpoints) between any two of these values or any value therein.
  • one treatment cycle is about 1 week, about 2 weeks, about 3 weeks, or about 4 weeks.
  • the patient is administered an effective amount of the anti-TIGIT antibody or antigen-binding fragment per treatment cycle of about 10 mg to about 300 mg, or a formulation containing such dose of the anti-TIGIT antibody or antigen-binding fragment; wherein one treatment cycle is About 1 week, about 2 weeks, about 3 weeks, or about 4 weeks.
  • the effective amount of the anti-TIGIT antibody or antigen-binding fragment administered to the patient per treatment cycle is about 10 mg, about 30 mg, about 40 mg, about 50 mg, about 78 mg, about 82 mg, about 96 mg, about 100 mg, about 124 mg , about 143 mg, about 157 mg, about 180 mg, about 196 mg, about 200 mg, about 226 mg, about 240 mg, about 260 mg, about 286 mg, about 300 mg, or a range (including endpoints) between any two of these values or in which any value, or formulation comprising such dose of anti-TIGIT antibody or antigen-binding fragment; wherein one treatment cycle is about 1 week, about 2 weeks, about 3 weeks, or about 4 weeks.
  • the effective amount of the anti-TIGIT antibody or antigen-binding fragment administered to the patient per treatment cycle is about 300 mg to 600 mg, or a formulation containing such dose of the anti-TIGIT antibody or antigen-binding fragment; wherein one treatment cycle is about 1 week, about 2 weeks, about 3 weeks, or about 4 weeks.
  • the effective amount of the anti-TIGIT antibody or antigen-binding fragment administered to the patient per treatment cycle is about 300 mg, about 355 mg, about 360 mg, about 380 mg, about 390 mg, about 400 mg, about 420 mg, about 460 mg, about 480 mg , about 500 mg, about 520 mg, about 540 mg, about 580 mg, about 600 mg, or a range between any two of these values (including endpoints), or any value therein, or an anti-TIGIT antibody or antigen-binding fragment containing such a dose formulations; wherein one treatment cycle is about 1 week, about 2 weeks, about 3 weeks, or about 4 weeks.
  • the effective amount of the anti-TIGIT antibody or antigen-binding fragment administered to the patient per treatment cycle is about 600 mg to 900 mg, or a formulation containing such dose of the anti-TIGIT antibody or antigen-binding fragment; wherein one treatment cycle is about 1 week, about 2 weeks, about 3 weeks, or about 4 weeks.
  • the effective amount of the anti-TIGIT antibody or antigen-binding fragment administered to the patient per treatment cycle is about 600 mg, about 630 mg, about 642 mg, about 660 mg, about 680 mg, about 700 mg, about 734 mg, about 750 mg, about 780 mg , about 790 mg, about 800 mg, about 820 mg, about 844 mg, about 878 mg, about 900 mg, or a range between any two of these values (including endpoints) or any value therein, or containing such a dose of anti-TIGIT antibody or antigen Formulations of binding fragments; wherein one treatment cycle is about 1 week, about 2 weeks, about 3 weeks, or about 4 weeks.
  • the patient is administered an effective amount of an anti-TIGIT antibody (e.g., antibody h10D8OF or h10D8OFKF) or antigen-binding fragment per treatment cycle of about 9 mg to 13 mg, or a formulation containing such an amount of anti-TIGIT antibody or antigen-binding fragment; For example, about 10 mg is administered once.
  • the effective amount of the anti-TIGIT antibody or antigen-binding fragment administered to the patient is about 10 mg per treatment cycle, or a formulation containing such dose of the anti-TIGIT antibody or antigen-binding fragment; wherein one treatment cycle is about 1 week , about 2 weeks, about 3 weeks, or about 4 weeks.
  • the patient is administered an effective amount of an anti-TIGIT antibody (eg, antibody h10D8OF or h10D8OFKF) or antigen-binding fragment per treatment cycle of about 25 mg to 33 mg, or a formulation containing such an amount of anti-TIGIT antibody or antigen-binding fragment; For example, about 30 mg is administered once.
  • the effective amount of the anti-TIGIT antibody or antigen-binding fragment administered to the patient per treatment cycle is about 30 mg, or a formulation containing this dose of the anti-TIGIT antibody or antigen-binding fragment; wherein one treatment cycle is about 1 week , about 2 weeks, about 3 weeks, or about 4 weeks.
  • the patient is administered an effective amount of an anti-TIGIT antibody (eg, antibody h10D8OF or h10D8OFKF) or antigen-binding fragment per treatment cycle, or a formulation containing such an amount of anti-TIGIT antibody or antigen-binding fragment; For example, about 100 mg is administered once.
  • the effective amount of the anti-TIGIT antibody or antigen-binding fragment administered to the patient is about 100 mg per treatment cycle, or a formulation containing such dose of the anti-TIGIT antibody or antigen-binding fragment; wherein one treatment cycle is about 1 week , about 2 weeks, about 3 weeks, or about 4 weeks.
  • the patient is administered an effective amount of an anti-TIGIT antibody (eg, antibody h10D8OF or h10D8OFKF) or antigen-binding fragment per treatment cycle, or a formulation containing such an amount of anti-TIGIT antibody or antigen-binding fragment; For example, about 300 mg is administered once.
  • the effective amount of the anti-TIGIT antibody or antigen-binding fragment administered to the patient per treatment cycle is about 300 mg, or a formulation containing this dose of the anti-TIGIT antibody or antigen-binding fragment; wherein one treatment cycle is about 1 week , about 2 weeks, about 3 weeks, or about 4 weeks.
  • the patient is administered an effective amount of an anti-TIGIT antibody (eg, antibody h10D8OF or h10D8OFKF) or antigen-binding fragment per treatment cycle, or a formulation containing such an amount of anti-TIGIT antibody or antigen-binding fragment; For example, about 400 mg is administered once.
  • the effective amount of the anti-TIGIT antibody or antigen-binding fragment administered to the patient is about 400 mg per treatment cycle, or a formulation containing this dose of the anti-TIGIT antibody or antigen-binding fragment; wherein one treatment cycle is about 1 week , about 2 weeks, about 3 weeks, or about 4 weeks.
  • the patient is administered an effective amount of an anti-TIGIT antibody (eg, antibody h10D8OF or h10D8OFKF) or antigen-binding fragment, or a formulation containing such an amount of anti-TIGIT antibody or antigen-binding fragment, per treatment cycle; For example, about 600 mg is administered once. In some embodiments, the effective amount of the anti-TIGIT antibody or antigen-binding fragment administered to the patient per treatment cycle is about 600 mg, or a formulation containing this dose of the anti-TIGIT antibody or antigen-binding fragment; wherein one treatment cycle is about 1 week , about 2 weeks, about 3 weeks, or about 4 weeks.
  • an anti-TIGIT antibody eg, antibody h10D8OF or h10D8OFKF
  • the effective amount of the anti-TIGIT antibody or antigen-binding fragment administered to the patient per treatment cycle is about 600 mg, or a formulation containing this dose of the anti-TIGIT antibody or antigen-binding fragment;
  • the patient is administered an effective amount of an anti-TIGIT antibody (eg, antibody h10D8OF or h10D8OFKF) or antigen-binding fragment per treatment cycle, or a formulation containing such an amount of anti-TIGIT antibody or antigen-binding fragment; For example, about 800 mg is administered once. In some embodiments, the effective amount of the anti-TIGIT antibody or antigen-binding fragment administered to the patient is about 800 mg per treatment cycle, or a formulation containing such dose of the anti-TIGIT antibody or antigen-binding fragment; wherein one treatment cycle is about 1 week , about 2 weeks, about 3 weeks, or about 4 weeks.
  • an anti-TIGIT antibody eg, antibody h10D8OF or h10D8OFKF
  • the patient is administered an effective amount of an anti-TIGIT antibody (eg, antibody h10D8OF or h10D8OFKF) or antigen-binding fragment per treatment cycle, or a formulation containing such an amount of anti-TIGIT antibody or antigen-binding fragment; For example, about 900 mg is administered once. In some embodiments, the patient is administered an effective amount of an anti-TIGIT antibody or antigen-binding fragment of about 900 mg per treatment cycle, or a formulation containing this dose of an anti-TIGIT antibody or antigen-binding fragment; wherein one treatment cycle is about 1 week , about 2 weeks, about 3 weeks, or about 4 weeks.
  • an anti-TIGIT antibody eg, antibody h10D8OF or h10D8OFKF
  • the effective amount of anti-TIGIT antibody (eg, antibody h10D8OF or h10D8OFKF) or antigen-binding fragment administered is 9 mg to 600 mg once every 3 weeks.
  • the anti-TIGIT antibody is administered in an effective amount of about 9 mg, about 10 mg, about 16 mg, about 18 mg, about 30 mg, about 60 mg, about 100 mg, about 180 mg, about 200 mg, about 300 mg, about 400 mg, about 500 mg, or About 600mg every 3 weeks.
  • the anti-TIGIT antibody or antigen-binding fragment is administered in an effective amount of about 10 mg, about 30 mg, about 100 mg, about 300 mg, or about 600 mg once every 3 weeks.
  • the anti-TIGIT antibody or antigen-binding fragment is administered in an effective amount of about 600 mg to 1200 mg once every 3 weeks. In some embodiments, the anti-TIGIT antibody or antigen-binding fragment is administered in an effective amount of about 700 mg, about 800 mg, 900 mg, or about 1200 once every 3 weeks.
  • the patient is administered an anti-TIGIT antibody or antigen-binding fragment (or formulation) once per treatment cycle.
  • the anti-TIGIT antibody or antigen-binding fragment (or formulation) is administered multiple times per treatment cycle, eg, 2, 3, 4, or 5 times.
  • the patient is only dosed 1 or 4 times per treatment cycle.
  • the patient is treated with one treatment cycle. In some embodiments, the patient is treated with multiple (eg, 2, 3, or 4) treatment cycles. In some embodiments, the patient receives treatment until the condition resolves and no longer requires treatment.
  • the present invention discloses a method for treating a tumor or cancer, the method comprising: administering to a patient in need thereof about 9 mg to 30 mg, about 30 mg to 100 mg, about 100 mg to 300 mg, about 300 mg to 600 mg, about 600 mg to 900 mg, or about 900 mg to 1200 mg, such as about 9 mg, about 10 mg, about 18 mg, about 20 mg, about 30 mg, about 60 mg, 100 mg, about 180 mg, about 200 mg, about 300 mg, about 400 mg, about 500 mg , about 600 mg, about 900 mg, or about 1200 mg (or a range between any two of these values (including endpoints) or any value therein) of an anti-TIGIT antibody or antigen-binding fragment, or containing such a dose of an anti-TIGIT antibody or antigen Preparation of binding fragments.
  • the anti-TIGIT antibody is antibody h10D8OF.
  • the anti-TIGIT antibody is antibody h10D8OFKF
  • about 9 mg of the anti-TIGIT antibody or antigen-binding fragment is administered once every 3 weeks. In some embodiments, about 10 mg of the anti-TIGIT antibody or antigen-binding fragment is administered once every 3 weeks. In some embodiments, about 18 mg of the anti-TIGIT antibody or antigen-binding fragment is administered once every 3 weeks. In some embodiments, about 20 mg of the anti-TIGIT antibody or antigen-binding fragment is administered once every 3 weeks. In some embodiments, about 30 mg of the anti-TIGIT antibody or antigen-binding fragment is administered once every 3 weeks. In some embodiments, about 60 mg of the anti-TIGIT antibody or antigen-binding fragment is administered once every 3 weeks.
  • about 100 mg of the anti-TIGIT antibody or antigen-binding fragment is administered once every 3 weeks. In some embodiments, about 300 mg of the anti-TIGIT antibody or antigen-binding fragment is administered once every 3 weeks. In some embodiments, about 600 mg of the anti-TIGIT antibody or antigen-binding fragment is administered once every 3 weeks. In some embodiments, about 900 mg of the anti-TIGIT antibody or antigen-binding fragment is administered once every 3 weeks. In some embodiments, about 1200 mg of the anti-TIGIT antibody or antigen-binding fragment is administered once every 3 weeks.
  • the patient's symptoms are relieved after a single dose is administered. In some embodiments, after a single dose is administered and the patient's symptoms are not relieved as expected, the patient is administered about 9 mg to 1200 mg or about 10 mg to 900 mg of the anti-TIGIT antibody or antigen-binding fragment until the patient's symptoms are relieved.
  • the anti-TIGIT antibody or antigen-binding fragment (or formulation) is administered by subcutaneous (s.c.) injection, intraperitoneal (i.p.) injection, parenteral injection, intraarterial injection, or intravenous (i.v.) injection, etc. medicine.
  • the anti-TIGIT antibody or antigen-binding fragment (or formulation) is administered by infusion.
  • the anti-TIGIT antibody or antigen-binding fragment (or formulation) is administered as a bolus injection.
  • the anti-TIGIT antibody or antigen-binding fragment (or formulation) is administered by intravenous (i.v.) infusion (ie, intravenous infusion).
  • intravenous infusion ie, intravenous infusion
  • the duration of the intravenous infusion is about 50 minutes, about 55 minutes, about 60 minutes, about 65 minutes, about 70 minutes, about 75 minutes, about 81 minutes, about 87 minutes, about 90 minutes, about 95 minutes minutes, or the range (including the endpoints) between any two of these values, or any value therein. In some embodiments, the duration of the intravenous infusion is > 60 minutes.
  • the anti-TIGIT antibody or antigen-binding fragment (or formulation) is used in combination with other therapeutic methods to treat tumors or cancers, such as chemotherapy, radiation therapy, immunotherapy, hormone therapy, targeted therapy, biological therapy, and surgery, etc. .
  • the anti-TIGIT antibody or antigen-binding fragment (or formulation) is combined with other tumor or cancer therapeutic agents to treat tumors or cancers, such as hormones, antibodies to treat tumors or cancer, and the like.
  • the present invention discloses the use of anti-TIGIT antibodies or antigen-binding fragments in the preparation of medicaments for treating tumors or cancers.
  • the drug for treating a tumor or cancer includes an anti-TIGIT antibody or antigen-binding fragment.
  • the anti-TIGIT antibody is antibody h10D8OF.
  • the anti-TIGIT antibody is antibody h10D8OFKF.
  • the antibody h10D8OFKF has a fucosylation level of 0-10%.
  • the present invention also discloses a kit comprising an anti-TIGIT antibody or antigen-binding fragment (or preparation) and instructions for instructing a patient in need to administer the anti-TIGIT antibody or antigen-binding fragment (or preparation) .
  • the anti-TIGIT antibody is antibody h10D8OF.
  • the anti-TIGIT antibody is antibody h10D8OFKF.
  • the antibody h10D8OFKF has a fucosylation level of 0-10%.
  • the present invention also discloses a pharmaceutical composition suitable for injection, such as a bolus injection type pharmaceutical composition or an infusion (drip) type pharmaceutical composition, comprising the anti-TIGIT antibody or the antigen-binding fragment.
  • a pharmaceutical composition suitable for injectable use include sterile aqueous solutions (herein water-soluble) 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 of polyols (eg, glycerol, propylene glycol, and liquid polyethylene glycol, and the like), and the like, and suitable mixture.
  • the pharmaceutical composition further includes a pharmaceutically acceptable carrier.
  • a pharmaceutically acceptable carrier may be implemented comprising antibacterial and/or antifungal agents, such as parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like.
  • the pharmaceutically acceptable carrier may contain isotonic agents such as sugars, polyols (such as mannitol, sorbitol), sodium chloride.
  • the pharmaceutical composition comprises at least 0.1% anti-TIGIT antibody or antigen-binding fragment. The percentage of antibody can vary and is between about 2% and 90% by weight of a given dosage form. The amount of anti-TIGIT antibody or antigen-binding fragment in such a therapeutically useful pharmaceutical composition can be an effective amount for administration.
  • the anti-TIGIT antibody is antibody h10D8OF. In some embodiments, the anti-TIGIT antibody is antibody h10D8OFKF.
  • the antibody h10D8OFKF is expressed by an alpha-(1,6)-fucosyltransferase knockout cell line. In some embodiments, the antibody h10D8OFKF is expressed by a-(1,6)-fucosyltransferase knockout CHO cells.
  • the present invention also discloses the preparation method of the above pharmaceutical composition: respectively combining the anti-TIGIT antibody or antigen-binding fragment described herein with a pharmaceutically acceptable carrier suitable for injection (such as water for injection, physiological saline, etc.) mix.
  • a pharmaceutically acceptable carrier suitable for injection such as water for injection, physiological saline, etc.
  • anti-TIGIT antibodies or antigen-binding fragments (or preparations) of the present invention can be used for the treatment of tumors or cancers.
  • Figures 1A and 1B show that anti-TIGIT antibody inhibits the proliferation of tumor cells; wherein, Isotype IgG 30mg/kg indicates G1 group, h10D8OF 30mg/kg indicates G2 group, h10D8OFKF 10mg/kg indicates G3 group, h10D8OFKF 30mg/kg indicates G4 group, Tiragolumab 30 mg/kg represents the G5 group.
  • an entity refers to one or more of such entities, eg "an antibody” should be understood to mean one or more antibodies, thus the term “an” (or “an” ), “one or more” and “at least one” are used interchangeably herein.
  • compositions, methods and the like include the recited elements, such as components or steps, but do not exclude others.
  • Consisting essentially of means that the compositions and methods exclude other elements that have an essential effect on the characteristics of the combination, but do not exclude elements that have no essential effect on the compositions or methods.
  • Consisting of means excluding elements not specifically recited.
  • polypeptide is intended to encompass the singular “polypeptide” as well as the plural “polypeptide”, and refers to a molecule composed of amino acid monomers linked linearly by amide bonds (also known as peptide bonds).
  • polypeptide refers to any single chain or chains of two or more amino acids, and does not refer to a particular length of the product.
  • the definition of “polypeptide” includes a peptide, dipeptide, tripeptide, oligopeptide, "protein”, “amino acid chain” or any other term used to refer to two or more amino acid chains, and the term “polypeptide” may Used in place of, or used interchangeably with, any of the above terms.
  • polypeptide is also intended to refer to the product of post-expression modifications of the polypeptide, including but not limited to glycosylation, acetylation, phosphorylation, amidation, derivatization by known protecting/blocking groups, proteolytic cleavage or non-native Amino acid modifications that occur.
  • a polypeptide may be derived from a natural biological source or produced by recombinant techniques, but it need not be translated from a given nucleic acid sequence, and it may be produced by any means including chemical synthesis.
  • Amino acid refers to an organic compound containing both an amino group and a carboxyl group, such as an alpha-amino acid, which can be encoded by a nucleic acid directly or in a precursor form.
  • a single amino acid is encoded by a nucleic acid consisting of three nucleotides, so-called codons or base triplets. 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 Amino acid (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).
  • alanine three-letter code: ala, one-letter code: A
  • arginine arg, R
  • asparagine asparag
  • Constant amino acid substitution refers to the replacement of one amino acid residue by another amino acid residue containing a side chain (R group) of similar chemical properties (eg, charge or hydrophobicity). In general, conservative amino acid substitutions will not substantially alter the functional properties of the protein.
  • amino acid classes containing chemically similar side chains include: 1) aliphatic side chains: 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 chains: phenylalanine, tyrosine and tryptophan; 5) Basic side chains: lysine, Arginine and histidine; 6) Acidic side chains: aspartic acid and glutamic acid.
  • the number of amino acids for "conservative amino acid substitutions of VL and VH" can 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 (including endpoints) between any two of these values, or any value therein.
  • the number of amino acids for a "conservative amino acid substitution of a heavy or light chain” can 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 (including endpoints) between any two of these values, or any value therein.
  • encoding when applied to a polynucleotide refers to a polynucleotide referred to as “encoding” a polypeptide, transcribed and/or in its native state or when manipulated by methods well known to those skilled in the art Or translation can yield the polypeptide and/or fragments thereof.
  • Antibodies, antigen-binding fragments or derivatives disclosed herein include, but are not limited to, polyclonal, monoclonal, multispecific, fully human, humanized, primatized, chimeric antibodies, single chain antibodies, epitope binding Fragments (eg, Fab-like, Fab'-like, and F(ab') 2 ), single-chain-like Fvs (scFv).
  • recombinant refers to a polypeptide or polynucleotide and means a form of the polypeptide or polynucleotide that does not occur in nature, non-limiting examples may be combined to produce polynucleotides that do not normally exist or peptide.
  • Identity refers to the sequence similarity between two peptides or between two nucleic acid molecules. Homology can be determined by comparing the positions within each sequence that can be aligned. A molecule is homologous when a position in the sequences being compared is occupied by the same base or amino acid. The degree of homology between sequences is a function of the number of matches or homologous positions shared by the sequences.
  • At least 80% identity is about 80% identity, about 81% identity, about 82% identity, about 83% identity, about 85% identity, about 86% identity, about 87% identity, about 88% identity, about 90% identity, about 91% identity, about 92% identity, about 94% identity, about 95% identity, about 98% identity, about 99% identity, or these A range (including endpoints) between any two values in a numerical value or any value therein.
  • a polynucleotide or polynucleotide sequence has a certain percentage (eg, 90%, 95%, 98% or 99%) "identity" or “sequence identity” to another sequence "Sex” means that when the sequences are aligned, the percentage of bases (or amino acids) in the two sequences being compared are identical.
  • the percent alignment or sequence identity can be determined using visual inspection or software programs known in the art, such as those described by Ausubel et al. eds. (2007) in Current Protocols in Molecular Biology. Alignments are preferably performed using default parameters.
  • Biologically equivalent polynucleotides are polynucleotides that have the above-specified percentages of identity and encode polypeptides having the same or similar biological activity.
  • Antibody refers to a polypeptide or polypeptide complex that specifically recognizes and binds an antigen.
  • Antibodies can be whole antibodies and any antigen-binding fragments thereof or single chains thereof.
  • the term “antibody” thus includes any protein or peptide in the molecule that contains at least a portion of an immunoglobulin molecule that has the biological activity of binding to an antigen.
  • Antibodies and antigen-binding fragments include, but are not limited to, the complementarity determining regions (CDRs), heavy chain variable regions (VH), light chain variable regions (VL), heavy chain constant regions of heavy or light chains or ligand binding portions thereof (CH), a light chain constant region (CL), a framework region (FR), or any portion thereof, or at least a portion of a binding protein.
  • the CDR regions include the CDR regions of the light chain (LCDR1-3) and the CDR regions of the heavy chain (HCDR1-3).
  • antibody includes a wide variety of biochemically distinguishable polypeptides. Those of skill in the art will appreciate that classes of heavy chains include gamma, mu, alpha, delta, or epsilon (gamma, mu, alpha, delta, epsilon), with some subclasses (eg, gamma1-gamma4). The nature of this chain determines the "class” of the antibody as IgG, IgM, IgA, IgG or IgE, respectively. Immunoglobulin subclasses (isotypes), eg, IgGl, IgG2, IgG3, IgG4, IgG5, etc., are well characterized and the functional specificities conferred are known. All immunoglobulin species are within the scope of the present disclosure. In some embodiments, the immunoglobulin molecule is of the IgG class.
  • Light chains can be classified as kappa ( ⁇ ) or lambda ( ⁇ ). Each heavy chain can bind to a kappa or lambda light chain.
  • ⁇ kappa
  • ⁇ lambda
  • Each heavy chain can bind to a kappa or lambda light chain.
  • immunoglobulins are produced by hybridomas, B cells or genetically engineered host cells, their light and heavy chains are joined by covalent bonds, and the "tail" portion of the two heavy chains is joined by a covalent disulfide bond or non-covalent bond.
  • the amino acid sequence extends from the N-terminus of the forked terminus in the Y configuration to the C-terminus at the bottom of each chain.
  • the variable region of immunoglobulin kappa light chain is V ⁇ ; the variable region of immunoglobulin lambda light chain is V ⁇ .
  • Both light and heavy chains are divided into regions of structural and functional homology.
  • the terms "constant” and “variable” are used according to function.
  • the light chain variable region (VL) and heavy chain variable region (VH) determine antigen recognition and specificity.
  • the light chain constant region (CL) and heavy chain constant region (CH) confer important biological properties such as secretion, transplacental movement, Fc receptor binding, complement fixation, and the like. By convention, the numbering of constant regions increases as they become further from the antigen binding site or amino terminus of the antibody.
  • the N-terminal portion is the variable region and the C-terminal portion is the constant region; the CH3 and CL domains actually comprise the carboxy-terminus of the heavy and light chains, respectively.
  • CDR complementarity determining region
  • CDRs as defined by Kabat and Chothia include overlaps or subsets of amino acid residues when compared to each other. Nonetheless, 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 numbers encompassing a particular CDR will vary depending on the sequence and size of the CDR. Those skilled in the art can usually determine which specific residues the CDRs contain based on the amino acid sequence of the variable region of the antibody.
  • Kabat et al. also define a numbering system applicable to variable region sequences of any antibody.
  • One of ordinary skill in the art can apply this "Kabat numbering" system to any variable region sequence independent of experimental data other than the sequence itself.
  • Kabat Numbering means the numbering system proposed by Kabat et al., U.S. Dept. of Health and Human Services in "Sequence of Proteins of Immunological Interest” (1983).
  • Antibodies may also use the EU or Chothia numbering system.
  • Treatment means therapeutic treatment and prophylactic or prophylactic measures, the purpose of which is to prevent, slow, ameliorate and stop adverse physiological changes or disorders, such as the progression of disease, including but not limited to the following whether detectable or undetectable As a result, alleviation of symptoms, reduction of disease severity, stabilization of disease state (ie, no worsening), delay or slowdown of disease progression, improvement or alleviation of disease state, alleviation or disappearance (whether in part or in whole), prolongation and Expected duration of survival when not receiving treatment, etc.
  • a patient in need of treatment includes a patient already suffering from a condition or disorder, a patient susceptible to a condition or disorder, or a patient in need of prevention of such a condition or disorder, may or may be expected from administration of the antibodies or compositions disclosed herein for detection, Patients who benefit from the diagnostic process and/or treatment.
  • 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. In some embodiments, the patient is a human.
  • an “effective amount” refers to the amount of active compound or agent that elicits a biological or medical response in a tissue, system, animal, individual, or human; an effective amount is sought by the researcher, veterinarian, physician, or other clinician.
  • IC50 means the 50% inhibitory concentration, that is, the concentration of drug or inhibitor required to inhibit half the indicated biological process.
  • the term "in need” means that a patient has been identified as in need of a particular method or treatment. In some embodiments, identification can be made by any diagnostic means. In any of the methods and treatments described herein, the patient may need.
  • Antibody-encoding DNA can be designed and synthesized according to the antibody amino acid sequences described herein by conventional methods, inserted into an expression vector, and then transfected into host cells, and the transfected host cells are cultured in culture to produce monoclonal antibodies.
  • an antibody expression vector includes 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 inserted sequence.
  • Efficient transcription can be obtained by the early and late promoters of SV40, long terminal repeats from retroviruses such as RSV, HTLV1, HIVI, and the early promoter of cytomegalovirus, and other cellular promoters such as muscle can 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, pSV2dhfr, pBC12MI and pCS2 etc.
  • Commonly used mammalian cells include HEK293 cells, Cos1 cells, Cos7 cells, CV1 cells, mouse L cells and CHO cells.
  • the heavy and light chain DNA sequences of the antibodies were optimized according to CHO codon preference. Modify the 5' end of the DNA sequence with PCR primers, add the kozak sequence and the signal peptide DNA sequence to the 5' end of the light chain and heavy chain DNA sequences, then clone into the existing expression vector, and verify the correctness of the recombinant plasmid by sequencing analysis Construct.
  • the above recombinant plasmid was transfected into expression cells for expression, and the supernatant was collected and purified to obtain antibody protein samples, which were used in the following various examples.
  • the expression vector used in the preparation process of the antibody h10D8OF, is pCDNA3.1 TM (+) (Invitrogen company, the product number is V79020), and the expression cells are CHO cells; 2) in the preparation process of the antibody h10D8OFKF, the expression vector used It is pCDNA3.1 TM (+), the expression cells are CHO cells with ⁇ -(1,6)-fucosyltransferase gene knockout, and the tested fucosylation level is about 0; 3) Reference antibody Tiragolumab In the preparation process, the expression vector used was pCDNA3.1 TM (+), and the expression cells were CHO cells.
  • the amino acid sequences of the antibodies h10D8OF and h10D8OFKF are shown in Table 1 (the sequences of the antibodies h10D8OF and h10D8OFKF are the same), the amino acid sequences of the reference antibody Tiragolumab are shown in Table 2, and the DNA sequences of the antibodies h10D8OF and h10D8OFKF are shown in Table 3 (the sequences of the antibodies h10D8OF and h10D8OFKF are identical).
  • the DNA sequences added to the 5' ends of the light chain DNA sequences of antibodies h10D8OF and h10D8OFKF are gccgc caccatgg actttcaggtgcagatcatctccttcctgctgatcagcgcctccgtgatcatgtccaggggc, as shown in SEQ ID NO: 19, the kozak sequence is shown in underline, and the signal peptide is shown in italics;
  • the DNA sequence added at the 5' end is gccgccaccatgg gctggagcctgatcctgctgttcctggtggccgtggccaccagagtgctgtccc, as shown in SEQ ID NO: 20, the kozak sequence is underlined, and the signal peptide is shown in italics.
  • the DNA sequence added to the 5' end of the light chain DNA sequence of the reference antibody Tira golumab is gccgccaccatgg acatgagggtgctggccc agctgctgggactgctgctgctgtgcttcccaggcgccagatgc, as shown in SEQ ID NO: 21, the kozak sequence is underlined, and the signal peptide is shown in italics;
  • the DNA sequence added at the 5' end is gccgccac catgg agtttgggctgagctgggtttttccttgttgctatattaaaaggtgtccagt, as shown in SEQ ID NO: 22, the ko zak sequence is underlined, and the signal peptide is shown in italics.
  • TIGIT-Jurkat cells The binding of free PVR-Fc to TIGIT on the surface of TIGIT-Jurkat cells was detected by flow cytometry.
  • the test steps are as follows: take TIGIT-Jurkat cells with good viability (cell viability greater than 90%), resuspend them with PBS to a density of 10 million/ml after centrifugation, and add 50 ⁇ l per well to a 96-well tip bottom plate, which is the number of cells per well.
  • PVR-Fc-bio biotinylated PVR-Fc
  • PBS phosphate buffered saline
  • PVR-Fc-bio dilution a final concentration of 12.5nM
  • Appropriate amount of anti-TIGIT antibody or reference antibody Tiragolumab was diluted with PVR-Fc-bio diluent.
  • the initial concentration of the antibody was 200nM, 2-fold gradient dilution, a total of 10 concentration gradients, and 3 duplicate wells were set for each concentration point; 2- Incubate at 8°C for 1 hour, then wash twice with PBS, add 1:1000 diluted fluorescent secondary antibody Streptavidin-PE (eBioscience, CAT#12-4317-87) dilution, 100 ⁇ l per well, 2-8°C Incubate for 30 min; then wash with PBS twice, and use a flow analyzer to detect the fluorescence intensity (Mean PE-A).
  • the preparation method of PVR-Fc-bio is as follows: the nucleic acid sequence of the extracellular region of human PVR is added with enzyme cleavage sites (HindIII and EcoRI), and is fused with the nucleic acid sequence of the constant region of the human IgG1 heavy chain through a linker; Inserted into pCDNA3.1 TM (+) vector, and then transiently transfected into HEK293F cells; the cultured cell supernatant was purified by ProteinA affinity chromatography, and the purified fusion protein was named PVR-Fc; take an appropriate amount of PVR-Fc protein , using a biotin labeling kit (EZ-Link HSulfo-NHS-LC-Biotinylation Kit, product number: 21435), biotinylated PVR-Fc according to the operation steps in the manual, and the labeled protein was named PVR-Fc-bio.
  • EZ-Link HSulfo-NHS-LC-Biotinylation Kit product
  • amino sequence of the extracellular region of human PVR is shown in SEQ ID NO: 13
  • nucleotide sequence of the linker is shown in SEQ ID NO: 14
  • amino acid sequence of the linker is shown in SEQ ID NO: 23
  • human The amino acid sequence of the constant region of the IgG1 heavy chain is shown in SEQ ID NO: 15
  • amino acid sequence of the PVR-Fc is shown in SEQ ID NO: 24 (see Table 4).
  • the preparation method of TIGIT-Jurkat cells is as follows: replace the target gene on pCMV2-CFD-Flag (Yiqiao Shenzhou, item number: HG10160-MF) with the human full-length TIGIT gene to obtain a recombinant plasmid, and use the restriction endonuclease ClaI (Bsu15I) to The recombinant plasmid was linearized and transfected into Jurkat cell line (ATCC, Clone E6-1, TIB-152 TM ) by electroporation.
  • the screening pressure is hygromycin, and the positive cell line is obtained and then subcloned to obtain a cell line that can stably express human TIGIT, namely: TIGIT-Jurkat cells.
  • TIGIT-Jurkat cells a cell line that can stably express human TIGIT, namely: TIGIT-Jurkat cells.
  • sequence of the human full-length TIGIT gene is shown in SEQ ID NO: 16 (see Table 4).
  • both the antibody h10D8OF and the antibody Tiragolumab can effectively block the binding of TIGIT to PVR-Fc, and their IC 50 values are 0.4409nM and 2.820nM respectively; the blocking ability of the antibody h10D8OF is better than that of the antibody Tiragolumab.
  • both the antibody h10D8OFKF and the antibody Tiragolumab can effectively block the binding of TIGIT to PVR-Fc, and their IC 50 values are 0.742nM and 2.820nM respectively; the blocking ability of the antibody h10D8OFKF is better than that of the antibody Tiragolumab.
  • mice BALB/c-hPD1/hTIGIT (Jiangsu JiCui Yaokang Biotechnology Co., Ltd.) were subcutaneously inoculated with CT26 colon cancer tumor cells; after inoculation with tumor cells, when the average tumor volume of the mice was 79.65 mm3 , the Group, 10 in each group.
  • the day of grouping was defined as D0 day, and was administered by intraperitoneal injection (IP) on D0 day, D4 day, D7 day, D11 day, D14 day and D18 day, and the dose was 10 mg/kg or 30 mg/kg.
  • IP intraperitoneal injection
  • TGItw inhibition rate of tumor weight
  • TGItw (1-(mean TW administration group)/(mean TW control group )) ⁇ 100%;
  • Mean TW control group control The mean value of tumor weight at the end point treatment of mice in the group.
  • both antibody h10D8OF and antibody Tiragolumab can inhibit the growth of CT26 colon cancer, and the effect of antibody h10D8OF in inhibiting tumor growth is better than that of antibody Tiragolumab; at the dose of 30 mg/kg, the relative tumor at the end of the trial (D20 day) The inhibition rate TGitw (%) was 92.89% for the former and 71.07% for the latter.
  • both antibody h10D8OFKF and antibody Tiragolumab can inhibit the growth of CT26 colon cancer, and the effect of antibody h10D8OFKF in inhibiting tumor growth is better than that of antibody Tiragolumab.
  • the maximum tolerated dose (MTD) of single intravenous administration of antibody injection is greater than 400 mg/kg.
  • Cynomolgus monkeys were given antibody (antibody h10D8OF or h10D8OFKF) injection by intravenous infusion once a week, and the toxicity and toxicokinetics were evaluated after five repetitions; after four weeks of recovery, the reversibility of damage or possible delayed onset were observed. Toxic reaction. After the administration, 3 animals per gender in each group were dissected, and the remaining 2 animals per gender in each group were dissected at the end of the recovery period.
  • the immunogenicity results showed that 1) after cynomolgus monkeys were given 10, 30, and 100 mg/kg of the antibody h10D8OF in the repeated administration toxicity test, the positive rates of anti-drug antibodies (ADA) in the 10 and 30 mg/kg dose groups were 50, respectively. % and 25%, no ADA was detected in the high-dose group; 2) After cynomolgus monkeys were given 10, 30, and 100 mg/kg of the antibody h10D8OFKF in the repeated-dose toxicity test, the anti-drug antibodies in the 10, 30, and 100 mg/kg dose groups The positive rates were 70%, 33%, and 50%, respectively.
  • This study is two multicenter, open-label, dose-escalation studies evaluating the safety, tolerability, pharmacokinetic (PK) characteristics, and preliminary clinical efficacy of the antibody h10D8OF and h10D8OFKF injection in patients with advanced malignant solid tumors, respectively.
  • Phase I clinical trial exploring maximum tolerated dose (MTD) or maximum administered dose (MAD).
  • MTD maximum tolerated dose
  • MAD maximum administered dose
  • the clinical study of antibody h10D8OF is divided into two stages.
  • the first stage is based on the accelerated titration method and the "3+3" dose escalation rule to explore the safe dose range. It is mainly divided into: 30mg (starting dose) group and 100mg group using accelerated titration.
  • Dose escalation study was conducted by titration method; 300mg group, 600mg group and 900mg group were conducted dose escalation study according to the standard "3+3" rule; the second stage, dose expansion study, further studied the safety and clinical efficacy of antibody injection.
  • the clinical study of the antibody h10D8OFKF is divided into two stages.
  • the first stage is based on the accelerated titration method and the "3+3" dose escalation rule to explore the safe dose range. It is mainly divided into: 10mg (starting dose) group, 30mg group, 100mg The 300mg group, 600mg group and 900mg group were carried out dose escalation study according to the standard "3+3" rule; the second stage, dose expansion study, further study the safety and clinical efficacy of antibody injection sex.
  • 1 subject is included. If no DLT event is observed during the DLT evaluation period, it can directly enter the next group for dose escalation.
  • the accelerated escalation to the (300mg) dose Group.
  • DLT evaluation period the first treatment cycle (from the first dose to 21 days after the dose).
  • Tolerability evaluation indicators involved dose-limiting toxicity (DLT) events and their incidence; safety evaluation indicators involved: vital signs and physical examination, laboratory tests (blood routine, blood biochemistry, thyroid function, coagulation routine, urine routine , stool routine, pregnancy test), ECOG score, electrocardiogram, adverse events (including immune-related adverse events), etc.
  • DLT dose-limiting toxicity
  • Pharmacodynamics receptor occupancy (RO) study The receptor occupancy study of antibody injection was achieved by detecting TIGIT receptor binding on the surface of T cells in peripheral blood. Subjects in all dose groups were required to have blood samples collected at specific time points during the treatment period. Pharmacodynamic receptor occupancy studies were conducted only in dose-escalation subjects. A 2 mL blood sample was planned to be collected at each time point, and intensive sampling was performed before the administration of the first cycle, at the end of the administration, 168h, and 336h, and before the second to sixth cycles.
  • the immunogenicity evaluation index involves: anti-drug antibody (ADA) sample positive rate and individual positive rate, ADA-positive sample titer, and ADA-positive samples will continue to be tested for neutralizing antibody (nab).
  • ADA anti-drug antibody
  • Plasma concentrations Ctrough
  • serum drug concentration level was determined to study the pharmacokinetic (PK) characteristics.
  • parameters involved in PK are: for a single dose, parameters include C max , T max , T 1/2 , CL, Vd, Ke, MRT, AUC (0- ⁇ ) , AUC (0- ⁇ ) ; multiple doses When the drug is used, the parameters include C max,ss , C avg,ss , C min,ss , AUC (0- ⁇ ) ,ss, AUC( 0- ⁇ ) ,ss, Tmax,ss, T 1/2 ,ss, CL, Vss, Ke, MRT, accumulation index (Rac), volatility index DF.
  • ORR objective response rate
  • DOR duration of response
  • DCR disease control rate
  • PFS progression-free survival
  • OS overall survival
  • Objective Response Rate (ORR): the proportion of subjects with complete response (CR) and partial response (PR); Duration of Response (DOR): the first assessment of the tumor as an objective response (Objective Response, OR) to the first assessment of PD (Progressive Disease) or the time of death from any cause before PD, reflecting the duration of ORR; Progression-Free Survival (PFS): from the first dose Time to objective tumor progression or all-cause death (whichever occurs first).
  • ORR Objective Response Rate
  • DOR Duration of Response
  • PD Objective Response
  • PFS Progression-Free Survival
  • MTD is the highest dose level of DLT explored in a dose group in which ⁇ 1/6 of the subjects observed during the DLT evaluation period.
  • DLT Dose-Limiting Toxicity
  • AE adverse event
  • Ophthalmic toxicity requiring systemic therapy (Note: any patient with visual changes or ocular toxicity of grade 2 or higher will be evaluated by an ophthalmologist);

Abstract

本发明提供抗TIGIT抗体或抗原结合片段在制备治疗肿瘤或癌症的药物中的用途。本发明还提供一种试剂盒，包含抗TIGIT抗体或抗原结合片段和用于指导有需要患者给药抗TIGIT抗体或抗原结合片段的说明书。

Description

[根据细则26改正23.03.2022]　抗TIGIT抗体在治疗肿瘤或癌症中的应用 技术领域
本发明属于生物医药领域，尤其涉及抗TIGIT抗体或抗原结合片段在治疗肿瘤或癌症中的应用。
背景技术
TIGIT(T cell immunoreceptor with Ig and ITIM domains)是一种具有免疫球蛋白(Ig)和酪氨酸抑制剂基序(ITIM)结构域的T细胞免疫受体，主要表达于激活的T细胞和NK细胞上(Yu,X.,etal.(2009)."The surface protein TIGIT suppresses T cell activation by promoting the generation of mature immunoregulatory dendritic cells."Nature immunology 10(1):48-57.)。TIGIT结构显示包含一个细胞外免疫球蛋白结构域，一个I型跨膜区和两个ITIM基序。TIGIT是共同刺激网络的一部分，这个共刺激网络主要由T细胞上的激活性受体CD226和抑制性受体TIGIT，以及在APC、肿瘤细胞、感染的细胞表面表达的配体CD155(也称为PVR，一种在人类中被PVR基因编码的脊髓灰质炎病毒受体蛋白质)和CD112组成。TIGIT与PVR或CD112结合后会引起TIGIT胞质内Tyr225被磷酸化，TIGIT和细胞自适应生长因子受体结合蛋白2(GRB2)进行结合。GRB2可以招募SHIP1抑制磷脂酰肌醇三激酶(PI3K)和促分裂原活化蛋白激酶(MAPK)信号。除此之外，磷酸化的TIGIT通过Beta抑制蛋白2(β-arrestin2)招募SHIP1和通过阻断TNF受体相关因子6(TRAF6)的自身泛素化并破坏核因子KB(NF-KB)激活，一系列的信号传导最终导致T细胞或NK细胞的功能受到抑制，细胞因子的产生受到抑制。PVR既是TIGIT的配体，又是CD226分子的配体。在和CD112或CD155结合之后，CD226的胞内结构域的Ser329和Tyr322被磷酸化；Ser329磷酸化促进蛋白激酶(PKC)的激活和CD226与淋巴细胞关联抗原1(LFA1)的相互结合。LFA1然后被用于TYN介导的Tyr322磷酸化和CD226介导的下游信号传导。一系列的信号传导最终导致T细胞或NK细胞的功能受到激活，促进细胞因子的产生。存在于T细胞或NK细胞表面的TIGIT分子与CD226分子之间也发生着相互作用，表现在TIGIT分子可以直接扰乱CD226形成正常的二聚体，从而破坏CD226的正常生理功能。TIGIT和CD226如同天平的两端，通过PVR这个支点，通过共刺激和共抑制信号的传导巧妙地调节着机体的免疫功能。
TIGIT是肿瘤免疫疗法中的一个非常有潜力的抑制性性靶点，能为肿瘤免疫治疗提供新的手段。
发明内容
本发明公开了抗TIGIT抗体或抗原结合片段用于治疗肿瘤或癌症的方法或用途。在一些实施方案中，抗TIGIT抗体或抗原结合片段剂用于治疗肿瘤或癌症。
在一些实施方案中，所述抗TIGIT抗体或抗原结合片段至少包含SEQ ID NO:1所示的HCDR1、SEQ ID NO:2所示的HCDR2、SEQ ID NO:3所示的HCDR3、SEQ ID NO:4所示的LCDR1、SEQ ID NO:5所示的LCDR2、SEQ ID NO:6所示的LCDR3中一个或多个。
在一些实施方案中，所述抗TIGIT抗体或抗原结合片段包含SEQ ID NO:1所示的HCDR1、SEQ ID NO:2所示的HCDR2、SEQ ID NO:3所示的HCDR3、SEQ ID NO:4所示的LCDR1、SEQ ID NO:5所示的LCDR2和SEQ ID NO:6所示的LCDR3。
在一些实施方案中，所述抗TIGIT抗体或抗原结合片段的重链可变区包含SEQ ID NO:7所示的氨基酸序列，或与SEQ ID NO:7所示序列相比具有至少80％同一性的氨基酸序列，或与SEQ ID NO:7所示序列相比具有一个或多个保守氨基酸取代的氨基酸序列。
在一些实施方案中，所述抗TIGIT抗体或抗原结合片段的轻链可变区包含SEQ ID NO:8所示的氨基酸序列，或与SEQ ID NO:8所示序列相比具有至少80％同一性的氨基酸序列，或与SEQ ID NO:8所示序列相比具有一个或多个保守氨基酸取代的氨基酸序列。
在一些实施方案中，所述抗TIGIT抗体或抗原结合片段的重链可变区包含SEQ ID NO:7所示的氨基酸序列，或与SEQ ID NO:7所示序列相比具有至少80％同一性的氨基酸序列，或与SEQ ID NO:7所示序列相比具有一个或多个保守氨基酸取代的氨基酸序列；所述抗TIGIT抗体或抗原结合片段的轻链可变区包含SEQ ID NO:8所示的氨基酸序列，或与SEQ ID NO:8所示序列相比具有至少80％同一性的氨基酸序列，或与SEQ ID NO:8所示序列相比具有一个或多个保守氨基酸取代的氨基酸序列。
在一些实施方案中，所述抗TIGIT抗体或抗原结合片段的重链可变区包含SEQ ID NO:7所示的氨基酸序列，所述抗TIGIT抗体或抗原结合片段的轻链可变区包含SEQ ID NO:8所示的氨基酸序列。
在一些实施方案中，所述抗TIGIT抗体的重链包含SEQ ID NO:9所示的氨基酸序列，或与SEQ ID NO:9所示序列相比具有至少80％同一性的氨基酸序列，或与SEQ ID NO:9所示序列相比具有一个或多个保守氨基酸取代的氨基酸序列。
在一些实施方案中，所述抗TIGIT抗体的轻链包含SEQ ID NO:10所示的氨基酸序列，或与SEQ ID NO:10所示序列相比具有至少80％同一性的氨基酸序列，或与SEQ  ID NO:10所示序列相比具有一个或多个保守氨基酸取代的氨基酸序列。
在一些实施方案中，所述抗TIGIT抗体的重链包含SEQ ID NO:9所示的氨基酸序列，或与SEQ ID NO:9所示序列相比具有至少80％同一性的氨基酸序列，或与SEQ ID NO:9所示序列相比具有一个或多个保守氨基酸取代的氨基酸序列；所述抗TIGIT抗体的轻链包含SEQ ID NO:10所示的氨基酸序列，或与SEQ ID NO:10所示序列相比具有至少80％同一性的氨基酸序列，或与SEQ ID NO:10所示序列相比具有一个或多个保守氨基酸取代的氨基酸序列。
在一些实施方案中，所述抗TIGIT抗体为抗体h10D8OF或h10D8OFKF，抗体h10D8OF和h10D8OFKF的重链包含如SEQ ID NO:9所示的氨基酸序列，抗体h10D8OF和h10D8OFKF的轻链包含如SEQ ID NO:10所示的氨基酸序列；抗体h10D8OF和h10D8OFKF分别含有两条序列相同的重链和两条序列相同的轻链。
在一些实施方案中，所述抗TIGIT抗体(例如抗体h10D8OFKF)或抗原结合片段的岩藻糖基化水平为0-10％。在一些实施方案中，所述抗TIGIT抗体(例如抗体h10D8OFKF)或抗原结合片段的岩藻糖基化水平为0-5％。在一些实施方案中，所述抗TIGIT抗体(例如抗体h10D8OFKF)或抗原结合片段的岩藻糖基化水平为约0、约0.1％、约0.5％、约0.8％、约1％、约1.3％、约1.6％、约2.1％、2.9％、约3％、约3.3％、3.8％、约4％、约4.2％、4.3％、约4.6％、约5％或这些数值中任何两个值之间的范围(包括端点)或其中任何值。在一些实施方案中，所述抗TIGIT抗体(例如抗体h10D8OFKF)或抗原结合片段没有结合岩藻糖。在一些实施方案中，所述抗TIGIT抗体(例如抗体h10D8OFKF)或抗原结合片段具有增强的ADCC效应(antibody-dependent cell-mediated cytotoxicity)。
抗TIGIT抗体或抗原结合片段可以通过基因工程在CHO细胞或HEK293细胞中表达，并通过纯化获得；纯化可以采用常规方法进行，例如先离心细胞悬液并收集上清液，再次离心进一步去除杂质。ProteinA亲和柱和离子交换柱等方法可以用于纯化抗体蛋白。
在一些实施方案中，低岩藻糖基化或无岩藻糖基化的抗TIGIT抗体或抗原结合片段由α-(1,6)-岩藻糖基转移酶基因敲除的细胞系表达。在一些实施方案中，抗体h10D8OFKF由α-(1,6)-岩藻糖基转移酶基因敲除的细胞系表达，例如α-(1,6)-岩藻糖基转移酶基因敲除的CHO细胞。
在一些实施方案中，所述方法或用途包括：向有需要的患者施用有效量的抗TIGIT抗体或抗原结合片段。在一些实施方案中，所述抗TIGIT抗体为抗体h10D8OF或h10D8OFKF。在一些实施方案中，抗TIGIT抗体或抗原结合片段施用的有效剂量为每剂约9mg至1200mg。
在一些实施方案中，患者患有肿瘤或癌症。在一些实施方案中，肿瘤和癌症包括但不限于血液癌症、实体瘤。在一些实施方案中，血液癌症包括但不限于白血病、淋巴瘤和骨髓瘤。在一些实施方案中，白血病包括急性淋巴细胞性白血病(ALL)、急性骨髓性白血病(AML)、慢性淋巴细胞性白血病(CLL)、慢性骨髓性白血病(CML)和骨髓性增生疾病/肿瘤(MPDS)。在一些实施方案中，淋巴瘤包括霍奇金淋巴瘤、无痛性和侵袭性非霍奇金淋巴瘤、伯基特淋巴瘤和滤泡性淋巴瘤(小细胞和大细胞)。在一些实施方案中，骨髓瘤包括多发性骨髓瘤(MM)、巨细胞骨髓瘤、重链骨髓瘤和轻链或本斯-琼斯骨髓瘤。在一些实施方案中，实体瘤包括乳腺癌、卵巢癌、胰腺癌、前列腺癌、黑素瘤、结直肠癌、结肠癌、肺癌、头颈癌、膀胱癌、食道癌、肝癌和肾癌。在一些实施方案中，肿瘤和癌症为尚无有效治疗手段的经病理学确诊的局部晚期或转移性恶性实体肿瘤。
在一些实施方案中，本发明公开了一种用于治疗有需要患者的肿瘤或癌症的方法，其包括施用有效量的抗TIGIT抗体或抗原结合片段，其中抗TIGIT抗体或抗原结合片段施用的有效量为约9mg至1200mg每个治疗周期。在一些实施方案中，一个治疗周期为1周、2周、3周、4周、1个月、5周、6周、7周，或这些数值中的任何两个值之间的范围(包括端点)或其中任何值。在一些实施方案中，所述抗TIGIT抗体为抗体h10D8OF或h10D8OFKF。
在一些实施方案中，可以将抗TIGIT抗体配制成药物组合物，并以适合于所选给药途径的多种形式向患者给药，给药途径例如肠胃外、静脉内(iv)、肌肉内、局部或皮下。在一些实施方案中，可以将抗TIGIT抗体静脉输注。抗TIGIT抗体的给药量将取决于药物的性质、细胞表面触发药物的内在化、运输和释放的程度，以及所治疗的疾病和患者的状况(如年龄、性别、体重等)。
在一些实施方案中，每次施用的抗TIGIT抗体(如抗体h10D8OF或h10D8OFKF)或抗原结合片段为0.01mg/kg至26mg/kg或含此剂量抗TIGIT抗体或抗原结合片段的制剂。在一些实施方案中，每次施用的抗TIGIT抗体或抗原结合片段为约0.01mg/kg、约0.02mg/kg、约0.03mg/kg、约0.06mg/kg、约0.08mg/kg、约0.1mg/kg、约0.2mg/kg、约0.3mg/kg、约0.5mg/kg、约0.9mg/kg、约1mg/kg、约1.6mg/kg、约2mg/kg、约2.5mg/kg、约3mg/kg、约4mg/kg、约5mg/kg、约6mg/kg、约7mg/kg、约8mg/kg、约9mg/kg、约10mg/kg、约11mg/kg、约12mg/kg、约13mg/kg、约14mg/kg、约15mg/kg、约16mg/kg、约17mg/kg、约18mg/kg、约19mg/kg、约20mg/kg、约21mg/kg、约22mg/kg、约23mg/kg、约24mg/kg、约25mg/kg、约26mg/kg，或这些数值中任何两个值之间的范围(包括端点)或其中任何值，或含此剂量抗TIGIT抗体或抗原结合片段的制剂。在一些实施方案中，约1周、约2周、约3周、约4周、约5周、约6周或约7周给药1次。
在一些实施方案中，本发明公开了一种用于治疗肿瘤或癌症的方法，所述方法包括：每3周向有需要的患者给予约0.01mg/kg、约0.02mg/kg、约0.03mg/kg、约0.06mg/kg、约0.08mg/kg、约0.1mg/kg、约0.2mg/kg、约0.3mg/kg、约0.5mg/kg、约0.9mg/kg、约1mg/kg、约1.6mg/kg、约2mg/kg、约2.5mg/kg、约3mg/kg、约4mg/kg、约5mg/kg、约6mg/kg、约7mg/kg、约8mg/kg、约9mg/kg、约10mg/kg、约11mg/kg、约12mg/kg、约13mg/kg、约14mg/kg、约15mg/kg、约16mg/kg、约17mg/kg、约18mg/kg、约19mg/kg、约20mg/kg、约21mg/kg、约22mg/kg、约23mg/kg、约24mg/kg、约25mg/kg、约26mg/kg的抗TIGIT抗体或抗原结合片段，或这些数值中任何两个值之间的范围(包括端点)或其中任何值，或含此剂量抗TIGIT抗体或抗原结合片段的制剂。
在一些实施方案中，本发明公开了一种用于治疗肿瘤或癌症的方法，所述方法包括：每3周向有需要的患者给予约0.01mg/kg、约0.02mg/kg、约0.03mg/kg、约0.06mg/kg、约0.08mg/kg、约0.1mg/kg、约0.2mg/kg、约0.3mg/kg、约0.5mg/kg、约0.9mg/kg、约1mg/kg、约1.6mg/kg、约2mg/kg、约2.5mg/kg、约3mg/kg、约4mg/kg、约5mg/kg、约6mg/kg、约7mg/kg、约8mg/kg、约9mg/kg、约10mg/kg、约11mg/kg、约12mg/kg、约13mg/kg、约14mg/kg、约15mg/kg、约16mg/kg、约17mg/kg、约18mg/kg、约19mg/kg、约20mg/kg、约21mg/kg、约22mg/kg、约23mg/kg、约24mg/kg、约25mg/kg、约26mg/kg的抗体h10D8OF，或含此剂量抗体h10D8OF的制剂。
在一些实施方案中，本发明公开了一种用于治疗肿瘤或癌症的方法，所述方法包括：每3周向有需要的患者给予约0.01mg/kg、约0.02mg/kg、约0.03mg/kg、约0.06mg/kg、约0.08mg/kg、约0.1mg/kg、约0.2mg/kg、约0.3mg/kg、约0.5mg/kg、约0.9mg/kg、约1mg/kg、约1.6mg/kg、约2mg/kg、约2.5mg/kg、约3mg/kg、约4mg/kg、约5mg/kg、约6mg/kg、约7mg/kg、约8mg/kg、约9mg/kg、约10mg/kg、约11mg/kg、约12mg/kg、约13mg/kg、约14mg/kg、约15mg/kg、约16mg/kg、约17mg/kg、约18mg/kg、约19mg/kg、约20mg/kg、约21mg/kg、约22mg/kg、约23mg/kg、约24mg/kg、约25mg/kg、约26mg/kg的抗体h10D8OFKF，或含此剂量抗体h10D8OFKF的制剂。
在一些实施方案中，本发明公开了一种治疗肿瘤或癌症的方法，其包括向有需要的患者施用有效量的抗TIGIT抗体或抗原结合片段(或制剂)；其中，抗TIGIT抗体或抗原结合片段的有效量为单次给药9mg至1200mg(或含此剂量抗TIGIT抗体或抗原结合片段的制剂)。剂量时间表和给药方式取决于某些患者群中的抗TIGIT抗体或抗原结合片段(或制剂)的获益风险评估和一般临床实践指南。
在一些实施方案中，患者每个治疗周期内抗TIGIT抗体或抗原结合片段施用的有效量为9mg至1200mg(或含此剂量抗TIGIT抗体或抗原结合片段的制剂)。
[根据细则26改正23.03.2022]　
在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体(如抗体h10D8OF或h10D8OFKF)或抗原结合片段的有效量为约9mg、约10mg、约18mg、约26mg、约30mg、约38mg、约45mg、约60mg、约70mg、约80mg、约100mg、约120mg、约180mg、约200mg、约250mg、约290mg、约300mg、约330mg、约380mg、约400mg、约434mg、约480mg、约500mg、约567mg、约580mg、约600mg、约700mg、约800mg、约900mg，或这些数值中任何两个值之间的范围(包括端点)或其中任何值，或含此剂量抗TIGIT抗体或抗原结合片段的制剂。在一些实施方案中，一个治疗周期为1周至7周给药1次。在一些实施方案中，每个治疗周期内施用抗TIGIT抗体或抗原结合片段的有效量为约10mg至900mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；其中，一个治疗周期为约1周、约2周、约3周、约4周、约5周、约6周、约7周，或这些数值中的任何两个值之间的范围(包括端点)或其中任何值。在一些实施方案中，一个治疗周期为约1周、约2周、约3周或约4周。在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体或抗原结合片段的有效量为约10mg至约300mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；其中，一个治疗周期为约1周、约2周、约3周、或约4周。在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体或抗原结合片段的有效量为约10mg、约30mg、约40mg、约50mg、约78mg、约82mg、约96mg、约100mg、约124mg、约143mg、约157mg、约180mg、约196mg、约200mg、约226mg、约240mg、约260mg、约286mg、约300mg，或这些数值中的任何两个值之间的范围(包括端点)或其中任何值，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；其中，一个治疗周期为约1周、约2周、约3周或约4周。在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体或抗原结合片段的有效量为约300mg至600mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；其中，一个治疗周期为约1周、约2周、约3周、或约4周。在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体或抗原结合片段的有效量为约300mg、约355mg、约360mg、约380mg、约390mg、约400mg、约420mg、约460mg、约480mg、约500mg、约520mg、约540mg、约580mg、约600mg，或这些数值中的任何两个值之间的范围(包括端点)或其中任何值，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；其中，一个治疗周期为约1周、约2周、约3周或约4周。在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体或抗原结合片段的有效量为约600mg至900mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；其中，一个治疗周期为约1周、约2周、约3周或约4周。在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体或抗原结合片段的有效量为约600mg、约630mg、约642mg、 约660mg、约680mg、约700mg、约734mg、约750mg、约780mg、约790mg、约800mg、约820mg、约844mg、约878mg、约900mg，或这些数值中的任何两个值之间的范围(包括端点)或其中任何值，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；其中，一个治疗周期为约1周、约2周、约3周或约4周。
在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体(如抗体h10D8OF或h10D8OFKF)或抗原结合片段的有效量为约9mg至13mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；比如约10mg给药1次。在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体或抗原结合片段的有效量为约10mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；其中，一个治疗周期为约1周、约2周、约3周或约4周。
在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体(如抗体h10D8OF或h10D8OFKF)或抗原结合片段的有效量为约25mg至33mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；比如约30mg给药1次。在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体或抗原结合片段的有效量为约30mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；其中，一个治疗周期为约1周、约2周、约3周或约4周。
在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体(如抗体h10D8OF或h10D8OFKF)或抗原结合片段的有效量为约89mg至120mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；比如约100mg给药1次。在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体或抗原结合片段的有效量为约100mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；其中，一个治疗周期为约1周、约2周、约3周或约4周。
在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体(如抗体h10D8OF或h10D8OFKF)或抗原结合片段的有效量为约289mg至314mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；比如约300mg给药1次。在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体或抗原结合片段的有效量为约300mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；其中，一个治疗周期为约1周、约2周、约3周或约4周。
在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体(如抗体h10D8OF或h10D8OFKF)或抗原结合片段的有效量为约390mg至410mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；比如约400mg给药1次。在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体或抗原结合片段的有效量为约400mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；其中，一个治疗周期为约1周、约2周、约3周 或约4周。
在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体(如抗体h10D8OF或h10D8OFKF)或抗原结合片段的有效量为约580mg至740mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；比如约600mg给药1次。在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体或抗原结合片段的有效量为约600mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；其中，一个治疗周期为约1周、约2周、约3周或约4周。
在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体(如抗体h10D8OF或h10D8OFKF)或抗原结合片段的有效量为约748mg至833mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；比如约800mg给药1次。在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体或抗原结合片段的有效量为约800mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；其中，一个治疗周期为约1周、约2周、约3周或约4周。
在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体(如抗体h10D8OF或h10D8OFKF)或抗原结合片段的有效量为约872mg至943mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；比如约900mg给药1次。在一些实施方案中，患者每个治疗周期内施用抗TIGIT抗体或抗原结合片段的有效量为约900mg，或含此剂量抗TIGIT抗体或抗原结合片段的制剂；其中，一个治疗周期为约1周、约2周、约3周或约4周。
在一些实施方案中，抗TIGIT抗体(如抗体h10D8OF或h10D8OFKF)或抗原结合片段施用的有效量为9mg至600mg每3周一次。在一些实施方案中，抗TIGIT抗体施用的有效量为约9mg、约10mg、约16mg、约18mg、约30mg、约60mg、约100mg、约180mg、约200mg、约300mg、约400mg、约500mg或约600mg每3周一次。在一些实施方案中，抗TIGIT抗体或抗原结合片段施用的有效量为约10mg、约30mg、约100mg、约300mg或约600mg每3周一次。在一些实施方案中，抗TIGIT抗体或抗原结合片段施用的有效量为约600mg至1200mg每3周一次。在一些实施方案中，抗TIGIT抗体或抗原结合片段施用的有效量为约700mg、约800mg、900mg或约1200每3周一次。
在一些实施方案中，患者每个治疗周期内给药一次抗TIGIT抗体或抗原结合片段(或制剂)。在一些实施方案中，每个治疗周期内多次给药抗TIGIT抗体或抗原结合片段(或制剂)，例如2次、3次、4次或5次。在一些实施方案中，患者每个治疗周期只能给药1次或4次。
在一些实施方案中，患者接受一个治疗周期治疗。在一些实施方案中，患者接受多个(例如2个、3个或4个)治疗周期治疗。在一些实施方案中，患者接受治疗直至病症得到缓解而不再需要治疗。
在一些实施方案中，本发明公开了一种用于治疗肿瘤或癌症的方法，所述方法包括：每3周向有需要的患者给予约9mg至30mg、约30mg至100mg、约100mg至300mg、约300mg至600mg、约600mg至900mg或约900mg至1200mg，比如约9mg、约10mg、约18mg、约20mg、约30mg、约60mg、100mg、约180mg、约200mg、约300mg、约400mg、约500mg、约600mg、约900mg或约1200mg(或这些数值中的任何两个值之间的范围(包括端点)或其中任何值)的抗TIGIT抗体或抗原结合片段，或含此剂量抗TIGIT抗体或抗原结合片段的制剂。在一些实施方案中，抗TIGIT抗体为抗体h10D8OF。在一些实施方案中，抗TIGIT抗体为抗体h10D8OFKF。
在一些实施方案中，每3周一次给药抗TIGIT抗体或抗原结合片段约9mg。在一些实施方案中，每3周一次给药抗TIGIT抗体或抗原结合片段约10mg。在一些实施方案中，每3周一次给药抗TIGIT抗体或抗原结合片段约18mg。在一些实施方案中，每3周一次给药抗TIGIT抗体或抗原结合片段约20mg。在一些实施方案中，每3周一次给药抗TIGIT抗体或抗原结合片段约30mg。在一些实施方案中，每3周一次给药抗TIGIT抗体或抗原结合片段约60mg。在一些实施方案中，每3周一次给药抗TIGIT抗体或抗原结合片段约100mg。在一些实施方案中，每3周一次给药抗TIGIT抗体或抗原结合片段约300mg。在一些实施方案中，每3周一次给药抗TIGIT抗体或抗原结合片段约600mg。在一些实施方案中，每3周一次给药抗TIGIT抗体或抗原结合片段约900mg。在一些实施方案中，每3周一次给药抗TIGIT抗体或抗原结合片段约1200mg。
在一些实施方案中，单剂量给药后，患者的症状得到缓解。在一些实施方案中，单剂量给药后，患者后的症状未得到预期缓解，再对患者给药约9mg至1200mg或约10mg至900mg抗TIGIT抗体或抗原结合片段，直至患者的症状得到缓解。
在一些实施方案中，抗TIGIT抗体或抗原结合片段(或制剂)是通过皮下(s.c.)注射、腹膜内(i.p.)注射、肠胃外注射、动脉内注射或静脉内(i.v.)注射等方式进行给药。在一些实施方案中，抗TIGIT抗体或抗原结合片段(或制剂)是输液方式进行给药。在一些实施方案中，抗TIGIT抗体或抗原结合片段(或制剂)是推注方式进行给药。
在一些实施方案中，抗TIGIT抗体或抗原结合片段(或制剂)是通过静脉内(i.v.)输液方式(即静脉输注)进行给药。在一些实施方案中，静脉内输液持续时间为约50分钟、约55分钟、约60分钟、约65分钟、约70分钟、约75分钟、约81分钟、约 87分钟、约90分钟、约95分钟，或这些数值中任何两个值之间的范围(包括端点)或其中任何值。在一些实施方案中，静脉输液持续时间≥60分钟。
在一些实施方案中，抗TIGIT抗体或抗原结合片段(或制剂)与其他治疗方法联合用于治疗肿瘤或癌症，例如化疗、放疗、免疫治疗、激素治疗、靶向治疗、生物治疗和手术治疗等。在一些实施方案中，抗TIGIT抗体或抗原结合片段(或制剂)与其他肿瘤或癌症治疗剂联合治疗肿瘤或癌症，如激素、治疗肿瘤或癌症的抗体等。
另一方面，本发明公开了抗TIGIT抗体或抗原结合片段在制备用于治疗肿瘤或癌症的药物中的应用。在一些实施方案中，用于治疗肿瘤或癌症的药物包括抗TIGIT抗体或抗原结合片段。在一些实施方案中，抗TIGIT抗体为抗体h10D8OF。在一些实施方案中，抗TIGIT抗体为抗体h10D8OFKF。在一些实施方案中，抗体h10D8OFKF岩藻糖基化水平为0-10％。
另一方面，本发明还公开了一种试剂盒，试剂盒包含抗TIGIT抗体或抗原结合片段(或制剂)和用于指导有需要患者给药抗TIGIT抗体或抗原结合片段(或制剂)的说明书。在一些实施方案中，抗TIGIT抗体为抗体h10D8OF。在一些实施方案中，抗TIGIT抗体为抗体h10D8OFKF。在一些实施方案中，抗体h10D8OFKF岩藻糖基化水平为0-10％。
另一方面，本发明还公开了包含抗TIGIT抗体或抗原结合片段的适合注射用的药物组合物，如推注型药物组合物或输液(滴注)型药物组合物。适于注射用途的药物组合物包括无菌水性溶液(在此是水溶性的)或分散体以及用于即时制备无菌注射液或分散体的无菌粉末。对于静脉内施用，合适的载体包括生理盐水、抑菌水或磷酸盐缓冲盐水(PBS)、乙醇、多元醇(例如，甘油、丙二醇和液体聚乙二醇等)的溶剂或分散介质，及其适宜的混合物。在一些实施方案中，药物组合物还包括药学可接受的载体。在一些实施方案中，药学上可接受的载体可以包含抗细菌剂和/或抗真菌剂，如对羟基苯甲酸酯、氯代丁醇、苯酚、抗坏血酸、硫柳汞等来实现。在一些实施方案中，药学上可接受的载体可以包含等渗剂，如糖、多元醇(诸如甘露糖醇、山梨醇)、氯化钠。在一些实施方案中，药物组合物至少包含0.1％的抗TIGIT抗体或抗原结合片段。抗体的百分比可以变化，并且为给定剂型重量的约2％至90％之间。这种治疗上有用的药物组合物中抗TIGIT抗体或抗原结合片段的量可以为给药的有效量。在一些实施方案中，抗TIGIT抗体为抗体h10D8OF。在一些实施方案中，抗TIGIT抗体为抗体h10D8OFKF。在一些实施方案中，抗体h10D8OFKF由α-(1,6)-岩藻糖基转移酶基因敲除的细胞系表达。在一些实施方案中，抗体h10D8OFKF由α-(1,6)-岩藻糖基转移酶基因敲除的CHO细胞表达。
另一方面，本发明还公开了上述药物组合物的制备方法：分别将本文所述的抗 TIGIT抗体或抗原结合片段与药学上可接受的适合注射用的载体(例如注射用水，生理盐水等)混合。上述抗TIGIT抗体或抗原结合片段与药学上可接受的载体的混合方法是本领域通常已知的。
本发明抗TIGIT抗体或抗原结合片段(或制剂)可用于肿瘤或癌症的治疗。
附图说明
图1A和1B显示抗TIGIT抗体抑制肿瘤细胞的增殖；其中，Isotype IgG 30mg/kg表示G1组，h10D8OF 30mg/kg表示G2组，h10D8OFKF 10mg/kg表示G3组，h10D8OFKF 30mg/kg表示G4组，Tiragolumab 30mg/kg表示G5组。
术语
除非另作说明，否则下列的每一个术语应当具有下文所述的含义。
定义
应当注意的是，术语“一种”实体是指一种或多种该实体，例如“一种抗体”应当被理解为一种或多种抗体，因此，术语“一种”(或“一个”)、“一种或多种”和“至少一种”可以在本文中互换使用。
本文所用的术语“包含”或“包括”意味着组合物和方法等包括所列举的元素，例如组份或步骤，但不排除其它。“基本上由……组成”意味着组合物和方法排除对组合的特征有根本影响的其它元素，但不排除对组合物或方法无本质上影响的元素。“由……组成”意味着排除未特别列举的元素。
术语“多肽”旨在涵盖单数的“多肽”以及复数的“多肽”，并且是指由通过酰胺键(也称为肽键)线性连接的氨基酸单体构成的分子。术语“多肽”是指两个或更多个氨基酸的任何单条链或多条链，并且不涉及产物的特定长度。因此，“多肽”的定义中包括肽、二肽、三肽、寡肽、“蛋白质”、“氨基酸链”或用于指两个或多个氨基酸链的任何其他术语，并且术语“多肽”可以用来代替上述任何一个术语，或者与上述任何一个术语交替使用。术语“多肽”也意在指多肽表达后修饰的产物，包括但不限于糖基化、乙酰化、磷酸化、酰胺化、通过已知的保护/封闭基团衍生化、蛋白水解切割或非天然发生的氨基酸修饰。多肽可以源自天然生物来源或通过重组技术产生，但其不必从指定的核酸序列翻译所得，它可能以包括化学合成的任何方式产生。
“氨基酸”是指既含氨基又含羧基的有机化合物，比如α-氨基酸，其可直接或以前体的形式由核酸编码。单个氨基酸由三个核苷酸(所谓的密码子或碱基三联体)组成的核酸编码。每一个氨基酸由至少一个密码子编码。相同氨基酸由不同密码子编码称为“遗传密码的简并性”。氨基酸包括天然氨基酸和非天然氨基酸。天然氨基酸包括丙氨酸(三字母代码：ala，一字母代码：A)、精氨酸(arg，R)、天冬酰胺(asn，N)、天冬 氨酸(asp，D)、半胱氨酸(cys，C)、谷氨酰胺(gln，Q)、谷氨酸(glu，E)、甘氨酸(gly，G)、组氨酸(his，H)、异亮氨酸(ile，I)、亮氨酸(leu，L)、赖氨酸(lys，K)、甲硫氨酸(met，M)、苯丙氨酸(phe，F)、脯氨酸(pro，P)、丝氨酸(ser，S)、苏氨酸(thr，T)、色氨酸(trp，W)、酪氨酸(tyr，Y)和缬氨酸(val，V)。
“保守氨基酸取代”是指一个氨基酸残基被另一个含有化学性质(例如电荷或疏水性)相似的侧链(R基团)的氨基酸残基所取代。一般而言，保守氨基酸取代不大会在实质上改变蛋白质的功能性质。含有化学性质相似侧链的氨基酸类别的实例包括：1)脂族侧链：甘氨酸、丙氨酸、缬氨酸、亮氨酸和异亮氨酸；2)脂族羟基侧链：丝氨酸和苏氨酸；3)含酰胺的侧链：天冬酰胺和谷氨酰胺；4)芳族侧链：苯丙氨酸、酪氨酸和色氨酸；5)碱性侧链：赖氨酸、精氨酸和组氨酸；6)酸性侧链：天冬氨酸和谷氨酸。
“VL、VH的保守氨基酸取代”的氨基酸数目可为约1个、约2个、约3个、约4个、约5个、约6个、约8个、约9个、约10个、约11个、约13个、约14个、约15个保守氨基酸取代，或这些数值中的任何两个值之间的范围(包括端点)或其中任何值。“重链或轻链的保守氨基酸取代”的氨基酸数目可为约1个、约2个、约3个、约4个、约5个、约6个、约8个、约9个、约10个、约11个、约13个、约14个、约15个、约18个、约19个、约22个、约24个、约25个、约29个、约31个、约35个、约38个、约41个、约45个保守氨基酸取代，或这些数值中的任何两个值之间的范围(包括端点)或其中任何值。
术语“编码”应用于多聚核苷酸时，是指被称为“编码”多肽的多聚核苷酸，在其天然状态或当通过本领域技术人员公知的方法操作时，经转录和/或翻译可以产生该多肽和/或其片段。
本发明公开的抗体、抗原结合片段或衍生物包括但不限于多克隆、单克隆、多特异性、全人源、人源化、灵长类化、嵌合抗体、单链抗体、表位结合片段(例如类Fab、类Fab'和类F(ab') 2)、类单链Fvs(scFv)。
术语“重组”涉及多肽或多聚核苷酸，意指天然不存在的多肽或多聚核苷酸的形式，不受限制的实施例可以通过组合产生通常并不存在的多聚核苷酸或多肽。
“同一性”是指两个肽之间或两个核酸分子之间的序列相似性。可以通过比较每个序列中可以比对的位置来确定同源性。当被比较的序列中的位置被相同的碱基或氨基酸占据时，则分子在该位置是同源的。序列之间的同源程度是由序列共有的匹配或同源位置的数目组成的一个函数。
“至少80％同一性”为约80％同一性、约81％同一性、约82％同一性、约83％同 一性、约85％同一性、约86％同一性、约87％同一性、约88％同一性、约90％同一性、约91％同一性、约92％同一性、约94％同一性、约95％同一性、约98％同一性、约99％同一性，或这些数值中的任何两个值之间的范围(包括端点)或其中任何值。
多聚核苷酸或多聚核苷酸序列(或多肽或抗体序列)与另一序列有具有一定百分比(例如90％、95％、98％或者99％)的“同一性”或“序列同一性”是指当序列比对时，所比较的两个序列中该百分比的碱基(或氨基酸)相同。可以使用目测或本领域已知的软件程序来确定该比对同一性百分比或序列同一性，比如Ausubel et al.eds.(2007)在Current Protocols in Molecular Biology中所述的软件程序。优选使用默认参数进行比对。其中一种比对程序是使用默认参数的BLAST，例如BLASTN和BLASTP，两者使用下列默认参数：Geneticcode＝standard；filter＝none；strand＝both；cutoff＝60；expect＝10；Matrix＝BLOSUM62；Descriptions＝50sequences；sortby＝HIGHSCORE；Databases＝non-redundant；GenBank+EMBL+DDBJ+PDB+GenBankCDStranslations+SwissProtein+SPupdate+PIR。生物学上等同的多聚核苷酸是具有上述指定百分比的同一性并编码具有相同或相似生物学活性的多肽的多聚核苷酸。
“抗体”、“抗原结合片段”是指特异性识别和结合抗原的多肽或多肽复合物。抗体可以是完整的抗体及其任何抗原结合片段或其单链。因此术语“抗体”包括分子中含有具有与抗原结合的生物学活性的免疫球蛋白分子的至少一部分的任何蛋白质或肽。抗体和抗原结合片段包括但不局限重链或轻链或其配体结合部分的互补决定区(CDR)、重链可变区(VH)、轻链可变区(VL)、重链恒定区(CH)、轻链恒定区(CL)、框架区(FR)或其任何部分，或结合蛋白的至少一部分。CDR区包括轻链的CDR区(LCDR1-3)和重链的CDR区(HCDR1-3)。
术语“抗体”包括可以在生物化学上区分的各种广泛种类的多肽。本领域技术人员将会理解，重链的类别包括gamma、mu、alpha、delta或epsilon(γ、μ、α、δ、ε)，其中还有一些亚类(例如γ1-γ4)。该链的性质决定了抗体的“种类”分别为IgG、IgM、IgA、IgG或IgE。免疫球蛋白亚类(同种型)，例如IgG1、IgG2、IgG3、IgG4、IgG5等已被充分表征并且赋予的功能特异性也已知。所有的免疫球蛋白种类都在本发明公开的保护范围内。在一些实施方案中，免疫球蛋白分子为IgG种类。
轻链可以分为kappa(κ)或lambda(λ)。每个重链可以与κ或λ轻链结合。一般来说，当由杂交瘤，B细胞或基因工程宿主细胞生产免疫球蛋白时，其轻链和重链通过共价键结合，两条重链的“尾巴”部分通过共价二硫键或非共价键结合。在重链中，氨基酸序列从Y构型的叉状末端的N末端延伸至每条链底部的C末端。免疫球蛋白κ轻链可变区为Vκ；免疫球蛋白λ轻链可变区为V λ。
轻链和重链都分成结构和功能同源性的区域。术语“恒定的”和“可变的”根据功能 被使用。轻链可变区(VL)和重链可变区(VH)决定了抗原识别和特异性。轻链恒定区(CL)和重链恒定区(CH)赋予重要的生物学性质，如分泌、经胎盘移动、Fc受体结合、补体结合等。按照惯例，恒定区的编号随着它们变得更远离抗体的抗原结合位点或氨基末端而增加。N端部分是可变区，C端部分是恒定区；CH3和CL结构域实际上分别包含重链和轻链的羧基端。
在本领域中使用和/或接受的术语有两个或多个定义的情况下，除非明确地对立指出，否则本文使用的术语的定义包括所有这些含义。一个具体的例子是使用“互补决定区”(“CDR”)一词来描述在重链和轻链多肽的可变区内发现的非连续的抗原结合位点。这一特定区域在Kabat et al.,U.S.Dept.of Health and Human Services,Sequences of Proteins of Immunological Interest(1983)和Chothia等在J.Mol.Biol.196:901-917(1987)有相关描述，其通过引用全部并入本文。
根据Kabat和Chothia定义的CDR包括相互比较时的氨基酸残基的重叠或子集。尽管如此，应用任一定义来指代抗体或其变体的CDR都在本发明范围内。包含特定CDR的确切残基编号将根据CDR的序列和大小而变化。本领域技术人员通常可以根据抗体的可变区氨基酸序列确定出CDR包含哪些特定的残基。
Kabat等人还定义了适用于任何抗体的可变区序列的编号系统。本领域普通技术人员可以不依赖于序列本身以外的其他实验数据将该“Kabat编号”系统应用到任何可变区序列。“Kabat编号”是指由Kabat et al.,U.S.Dept.of Health and Human Services在“Sequence of Proteinsof Immunological Interest”(1983)提出的编号系统。抗体还可以用EU或Chothia编号系统。
“治疗”是指治疗性治疗和预防性或防治性措施，其目的是预防、减缓、改善和停止不良的生理改变或紊乱，例如疾病的进程，包括但不限于以下无论是可检测还是不可检测的结果，症状的缓解、疾病程度的减小、疾病状态的稳定(即不恶化)、疾病进展的延迟或减缓、疾病状态的改善或缓和，减轻或消失(无论是部分还是全部)、延长与不接受治疗时预期的生存期限等。需要治疗的患者包括已经患有病症或紊乱的患者，容易患有病症或紊乱的患者，或者需要预防该病症或紊乱的患者，可以或预期从施用本发明公开的抗体或组合物用于检测、诊断过程和/或治疗中受益的患者。
“患者”指需要诊断、预后或治疗的任何哺乳动物，包括人类、狗、猫、豚鼠、兔子、大鼠、小鼠、马、牛等。在一些实施方案中，患者为人。
“约”指相关技术领域技术人员容易知道的相应数值的常规误差范围。在一些实施方式中，本文中提到“约”指所描述的数值以及其±10％、±5％或±1％的范围。
“有效量”是指活性化合物或药剂的量，其能引起组织、系统、动物、个体或人类 的生物学或医学反应；有效量由研究人员、兽医、医生或其他临床医生寻求的。
“IC 50”表示50％抑制浓度，即对指定的生物过程抑制一半时所需的药物或者抑制剂的浓度。
如本文所用，术语“有需要”是指已将患者鉴定为需要特定方法或治疗。在一些实施例中，可以通过任何诊断方式进行识别。在本文描述的任何方法和治疗中，患者可能需要。
可以按常规方法根据本文所述抗体氨基酸序列设计合成编码抗体的DNA，将其置入表达载体中，然后转染宿主细胞，在培养基中培养被转染的宿主细胞产生单克隆抗体。在一些实施方案中，表达抗体载体包括至少一个启动子元件，抗体编码序列，转录终止信号和polyA尾。其他元件包括增强子，Kozak序列及插入序列两侧RNA剪接的供体和受体位点。可以通过SV40的前期和后期启动子，来自逆转录病毒的长末端重复序列如RSV、HTLV1、HIVI及巨细胞病毒的早期启动子来获得高效的转录，也可应用其它一些细胞的启动子如肌动蛋白启动子。合适的表达载体可包括pIRES1neo，pRetro-Off，pRetro-On，PLXSN，或者Plncx，pcDNA3.1(+/-)，pcDNA/Zeo(+/-)，pcDNA3.1/Hygro(+/-)，PSVL，PMSG，pRSVcat，pSV2dhfr，pBC12MI和pCS2等。常使用的哺乳动物细胞包括HEK293细胞，Cos1细胞，Cos7细胞，CV1细胞，鼠L细胞和CHO细胞等。
具体实施方式
以下通过具体的实施例进一步说明本发明的技术方案，具体实施例不代表对本发明保护范围的限制。其他人根据本发明理念所做出的一些非本质的修改和调整仍属于本发明的保护范围。
下述实施例中所用的材料、试剂等，如无特殊说明，均可从商业途径得到。
实施例1抗体的制备方法
根据CHO密码子偏爱性优化抗体的重链和轻链DNA序列。用PCR引物修饰DN A序列的5’端，在轻链和重链DNA序列的5’端添加kozak序列及信号肽DNA序列，再克隆到现有表达载体中，通过测序分析验证重组质粒的正确构建。将上述重组质粒转染表达细胞中进行表达，收集上清液、纯化获得抗体蛋白样品，并用于下面各种实施例。
其中，1)抗体h10D8OF制备过程中，采用的表达载体为pCDNA3.1 TM(+)(Invitr ogen公司，货号为V79020)，表达细胞为CHO细胞；2)抗体h10D8OFKF制备过程中，采用的表达载体为pCDNA3.1 TM(+)，表达细胞为α-(1,6)-岩藻糖基转移酶基因敲除的CHO细胞，经测试岩藻糖基化水平约为0；3)参照抗体Tiragolumab制备过程中， 采用的表达载体为pCDNA3.1 TM(+)，表达细胞为CHO细胞。抗体h10D8OF和h10D8OFKF的氨基酸序列见表1(抗体h10D8OF和h10D8OFKF的序列是相同的)，参照抗体Tiragolumab的氨基酸序列见表2，抗体h10D8OF和h10D8OFKF的DNA序列见表3(抗体h10D8OF和h10D8OFKF的序列是相同的)。
抗体h10D8OF和h10D8OFKF的轻链DNA序列的5’端添加的DNA序列为 gccgc caccatggactttcaggtgcagatcatctccttcctgctgatcagcgcctccgtgatcatgtccaggggc，如SEQ ID NO:19所示，kozak序列用下划线示出，信号肽用斜体示出；重链DNA序列的5’端添加的DNA序列为 gccgccaccatgggctggagcctgatcctgctgttcctggtggccgtggccaccagagtgctgtcc，如SEQ ID NO:20所示，kozak序列用下划线示出，信号肽用斜体示出。参照抗体Tira golumab的轻链DNA序列的5’端添加的DNA序列为 gccgccaccatggacatgagggtgctggccc agctgctgggactgctgctgctgtgcttcccaggcgccagatgc，如SEQ ID NO:21所示，kozak序列用下划线示出，信号肽用斜体示出；重链DNA序列的5’端添加的DNA序列为 gccgccac catggagtttgggctgagctgggttttccttgttgctatattaaaaggtgtccagtgt，如SEQ ID NO:22所示，ko zak序列用下划线示出，信号肽用斜体示出。
表1抗体h10D8OF和h10D8OFKF的氨基酸序列
Figure PCTCN2022078683-appb-000001
Figure PCTCN2022078683-appb-000002
表2参照抗体Tiragolumab的氨基酸序列
Figure PCTCN2022078683-appb-000003
表3抗体h10D8OF和h10D8OFKF的DNA序列
Figure PCTCN2022078683-appb-000004
Figure PCTCN2022078683-appb-000005
实施例2阻断TIGIT与配体PVR结合试验
采用流式细胞术检测抗TIGIT抗体阻断游离的PVR-Fc与TIGIT-Jurkat细胞表面的TIGIT的结合。试验步骤为：取活力良好(细胞活力大于90％)的TIGIT-Jurkat细胞，离心后用PBS重悬为密度1000万/ml，加入到96孔尖底板中，每孔50μl，即每孔细胞数量为50万；取适量的生物素化的PVR-Fc(即PVR-Fc-bio)，用PBS(磷酸缓冲液)进行稀释，配制成PVR-Fc-bio稀释液，终浓度为12.5nM；取适量抗TIGIT抗体或参照抗体Tiragolumab，用PVR-Fc-bio稀释液进行稀释，抗体起始浓度为200nM，2倍梯度稀释，共10个浓度梯度，每个浓度点设3个复孔；2-8℃条件下孵育1个小时，然后用PBS洗涤2次，加入1:1000稀释的荧光二抗Streptavidin-PE(eBioscience，CAT#12-4317-87)稀释液，每孔100μl，2-8℃孵育30min；然后PBS洗涤2次，采用流式分析仪检测荧光强度(Mean PE-A)。
PVR-Fc-bio的制备方法为：人PVR胞外区的核酸序列添加酶切位点(HindIII和EcoRI)，通过连接子与人IgG1重链恒定区的核酸序列进行融合；融合之后的序列被插入到pCDNA3.1 TM(+)载体中，然后瞬时转染HEK293F细胞；培养完成的细胞上清通过ProteinA亲和层析纯化，纯化得到的融合蛋白命名为PVR-Fc；取适量PVR-Fc蛋白，使用Thermo scientific公司的生物素标记试剂盒(EZ-Link
Figure PCTCN2022078683-appb-000006
HSulfo-NHS-LC-Biotinylation Kit，货号：21435)，按照说明书中的操作步骤对PVR-Fc进行生物素化标记，标记后的蛋白命名为PVR-Fc-bio。其中，人PVR胞外区的氨基序列如SEQ ID NO:13所示，连接子的核苷酸序列如SEQ ID NO:14所示，连接子的氨基酸序列如SEQ ID NO:23所示，人IgG1重链恒定区的氨基酸序列如SEQ ID NO:15所示，PVR-Fc的氨基酸序列如SEQ ID NO:24所示(见表4)。
TIGIT-Jurkat细胞的制备方法为：用人全长TIGIT基因替换pCMV2-CFD-Flag(义翘神州，货号：HG10160-M-F)上的目的基因得到重组质粒，用限制性内切酶ClaI(Bsu15I)将重组质粒线性化之后采用电穿孔法进行转染Jurkat细胞系(ATCC，Clone E6-1，TIB-152 TM)。筛选压为潮霉素，得到阳性细胞株再进行亚克隆进而得到可以稳定表达人TIGIT细胞系，即：TIGIT-Jurkat细胞。其中，人全长TIGIT基因的序列如SEQ ID NO:16所示(见表4)。
1)在同样条件下，抗体h10D8OF和抗体Tiragolumab均可以有效地阻断TIGIT与PVR-Fc的结合，两者的IC 50值分别为0.4409nM和2.820nM；抗体h10D8OF的阻断能力要优于抗体Tiragolumab。
2)在同样条件下，抗体h10D8OFKF和抗体Tiragolumab均可以有效地阻断TIGIT与PVR-Fc的结合，两者的IC 50值分别为0.742nM和2.820nM；抗体h10D8OFKF的阻断能力要优于抗体Tiragolumab。
表4相关序列
Figure PCTCN2022078683-appb-000007
Figure PCTCN2022078683-appb-000008
实施例3体内药效试验
人源化小鼠BALB/c-hPD1/hTIGIT(江苏集萃药康生物科技股份有限公司)皮下接种CT26结肠癌肿瘤细胞；接种肿瘤细胞后，当小鼠的平均肿瘤体积为79.65mm 3时，进行分组，每组10只。分组当天定义为D0天，并于D0天、D4天、D7天、D11天、 D14天、D18天采用腹腔注射(I.P.)方式进行给药，给药的剂量为10mg/kg或30mg/kg。给药方案见表5。
表5给药方案
Figure PCTCN2022078683-appb-000009
细胞接种后，每周常规监测肿瘤对动物正常行为的影响；具体指标包括：小鼠的活动性，摄食和饮水情况，体重增加或降低情况，眼睛、被毛及其它异常情况。肿瘤体积(mm 3)为0.5×(肿瘤长径×肿瘤短径 2)，各组小鼠的肿瘤体积以平均值±标准误差(mean±SEM)表示。TGItw(肿瘤重量的抑制率)计算公式为：
TGItw＝(1-(mean TW给药组)/(mean TW 对照组))×100％；Mean TW给药组：给药组小鼠终点处理时肿瘤重量的平均值，Mean TW 对照组：对照组小鼠终点处理时肿瘤重量的平均值。
1)如图1A所示，抗体h10D8OF和抗体Tiragolumab都可以抑制CT26结肠癌增长，并且抗体h10D8OF抑制肿瘤增长的效果优于抗体Tiragolumab；在30mg/kg剂量下，试验终点(D20天)时相对肿瘤抑制率TGItw(％)前者为92.89％，后者为71.07％。
2)如图1B所示，抗体h10D8OFKF和抗体Tiragolumab都可以抑制CT26结肠癌增长，并且抗体h10D8OFKF抑制肿瘤增长的效果优于抗体Tiragolumab。h10D8OFKF在10mg/kg和30mg/kg剂量下以及Tiragolumab在30mg/kg剂量下，试验终点时(D20天)相对肿瘤抑制率TGItw(％)分别为92.39％、95.94％和71.07％。
实施例4药代动力学和毒理试验
4a)本研究采用18只食蟹猴(雌雄各半)，随机分为3组，每组雌性和雄性动物各3只；每组动物未禁食单次静脉输注分别给予2、12和24mg/kg抗体(抗体h10D8OF或h10D8OFKF)注射液，共给药1次，静脉输注30min±10s；给药结束后血样采集至29天。通过血药浓度数据，运用Phoenix
Figure PCTCN2022078683-appb-000010
7.0软件采用非房室模型计算其主要药代动力学参数。
如表4和表5所示，在2-24mg/kg给药剂量范围内，未禁食后单次静脉输注给予食蟹猴注射液后，血清中抗体(抗体h10D8OF和h10D8OFKF)的系统暴露量无明显性别差异，并且血清中系统暴露量与给药剂量呈正相关。
表4食蟹猴给予抗体h10D8OF注射液后的药代动力学参数
Figure PCTCN2022078683-appb-000011
表5食蟹猴给予抗体h10D8OFKF注射液后的药代动力学参数
Figure PCTCN2022078683-appb-000012
Figure PCTCN2022078683-appb-000013
4b)考察食蟹猴静脉输注给予抗TIGIT抗体后急性毒性反应。试验共设计4组，每组雌性各1只。给药剂量分别为0、100、200、400mg/kg，静脉输注给药，共给药一次，输注30min±10s；给药后连续观察22天。
食蟹猴单次静脉输注30min给予100、200、400mg/kg抗体(抗体h10D8OF或h10D8OFKF)注射液后，食蟹猴能较好耐受；与同期对照组相比，抗体h10D8OFKF注射液所有剂量组中，食蟹猴在给药后出现与抗体供试品相关的耗食量下降，后呈现恢复趋势且缺乏关联性，不被认为是有害的；抗体h10D8OF各剂量组未见耗食量明显异常变化；无其它与抗体供试品相关的生存率、临床观察、体重、体温、心电图、临床病理及肉眼解剖变化出现。因此，本试验条件下，抗体注射液静脉单次给药的最大耐受剂量(MTD)大于400mg/kg。
4c)食蟹猴每周静脉输注给予抗体(抗体h10D8OF或h10D8OFKF)注射液一次，共重复五次后评估毒性反应及其毒代动力学；恢复四周后观察损害的可逆情况或可能的迟发毒性反应。给药结束后每组每性别解剖3只动物，恢复期结束每组每性别解剖剩余2只动物。
如表6和表7所示，首次给药及第4次给药后血清中抗体(抗体h10D8OF或h10D8OFK)暴露量(以AUC (0-t)计)随剂量的增加而增加。试验期间，试验组动物均未观察到与给予抗体(抗体h10D8OF或h10D8OFK)相关的体重、耗食量、给药部位刺激、眼科、呼吸、血液学、血清生化、凝血、尿液、细胞因子、淋巴细胞表型及脏器重量等指标的异常变化。
静脉给予10、30、100mg/kg的抗体h10D8OF或h10D8OFKF注射液5次后，抗体供试品未引起明显或严重的毒性反应，推测抗体h10D8OF和h10D8OFKF注射液未观察到有损伤作用的剂量水平(NOAEL)为100mg/kg。
免疫原性结果表明，1)食蟹猴在重复给药毒性试验中给予10、30、100mg/kg的 抗体h10D8OF后，10、30mg/kg剂量组抗药物抗体(ADA)的阳性率分别为50％和25％，高剂量组未检测到ADA；2)食蟹猴在重复给药毒性试验中给予10、30、100mg/kg的抗体h10D8OFKF后，10、30、100mg/kg剂量组抗药物抗体的阳性率分别为70％、33％、50％。
表6表 食蟹猴长期毒性试验中抗体h10D8OF的毒代动力参数
Figure PCTCN2022078683-appb-000014
表7表 食蟹猴长期毒性试验中h10D8OFKF的毒代动力参数
Figure PCTCN2022078683-appb-000015
4d)采用新西兰兔血红细胞进行溶血试验。当抗体(抗体h10D8OF或h10D8OFKF)注射液(25mg/mL)稀释超过10倍时，不会引起兔红细胞的溶血或凝集。
实施例5抗体h10D8OF和h10D8OFKF的临床研究
本研究是两项分别评价抗体h10D8OF和h10D8OFKF注射液在晚期恶性实体肿瘤 患者中的安全性、耐受性、药代动力学(PK)特征和初步临床有效性的多中心、开放性、剂量递增的I期临床试验；探索最大耐受剂量(MTD)或最大给药剂量(MAD)。
抗体h10D8OF临床研究分为两个阶段，第一阶段是基于加速滴定方法和“3+3”的剂量递增规则来探索安全剂量范围，主要分为：30mg(起始剂量)组、100mg组采用加速滴定方法进行剂量递增研究；300mg组、600mg组、900mg组按标准“3+3”规则进行剂量递增研究；第二阶段，剂量扩展研究，进一步研究抗体注射液的安全性和临床有效性。
抗体h10D8OFKF临床研究分为两个阶段，第一阶段是基于加速滴定方法和“3+3”的剂量递增规则来探索安全剂量范围，主要分为：10mg(起始剂量)组、30mg组、100mg组采用加速滴定方法进行剂量递增研究；300mg组、600mg组、900mg组按标准“3+3”规则进行剂量递增研究；第二阶段，剂量扩展研究，进一步研究抗体注射液的安全性和临床有效性。
三组采用加速滴定的剂量递增方案即队列规模＝1。首先纳入1例受试者，在DLT评估期内如果未观察到DLT事件，则可直接进入下一组进行剂量递增，以此规则(“1+1”)类推，加速递增至(300mg)剂量组。在加速递增过程中，若在某一组别观察到与研究药物相关的2级及以上毒性反应，则停止加速剂量递增，转为标准“3+3”剂量递增(即队列规模＝3)规则。
每3周以静脉输注给药1次，为1个周期(疗程)，输注时长可≥60分钟。如果患者出现输液相关反应并能够继续治疗，可基于临床实际情况可以使用苯海拉明或对乙酰氨基酚等进行预防给药。研究药物治疗应持续至出现：疾病进展、或不可耐受的毒性而退出、或因无疗效获益而接受新的抗肿瘤治疗、或撤回知情同意以及其它原因主动退出、或最长17个周期(约1年)，以先出现者为准。DLT评估期：为第1个治疗周期(从首次给药至给药后21天)。
耐受性评价指标涉及：剂量限制性毒性(DLT)事件及其发生率；安全性评价指标涉及：生命体征与体格检查、实验室检查(血常规、血生化、甲状腺功能、凝血常规、尿常规、便常规、妊娠试验)、ECOG评分、心电图、不良事件(包括免疫相关不良事件)等。
药效学受体占有率(RO)研究：抗体注射液的受体占有率研究是通过检测外周血中T细胞表面TIGIT受体结合来实现的。所有剂量组的受试者在治疗期间的特定时间点需要收集血样。药效学受体占有率研究只在剂量递增受试者中开展。每个时间点计划采集2mL血样，分别于第1周期给药前、给药结束时、168h、336h密集采样，第2至第6个周期给药前采样。
免疫原性评价指标涉及：抗药抗体(ADA)的样品阳性率和个体阳性率，ADA阳性样品的滴度，ADA阳性的样本将继续检测是否为中和抗体(nab)。
所有剂量组的受试者在治疗期间(前3个治疗周期)的规定时间点需要收集血样。给药前60分钟内监测血浆浓度(C trough)。每个时间点计划采2mL血样，检测血清药物的浓度水平，研究药代动力学(PK)特征。PK涉及的参数有：单次给药时，参数包括C max、T max、T 1/2、CL、Vd、Ke、MRT、AUC (0-τ)、AUC (0-∞)；多次给药时，参数包括C max,ss、C avg,ss、C min,ss、AUC (0-τ),ss、AU C(0-∞),ss、Tmax,ss、T 1/2,ss、CL、Vss、Ke、MRT、蓄积指数(Rac)、波动指数DF。
临床有效性评价：客观缓解率(ORR)、缓解持续时间(DOR)、疾病控制率(DCR)、无进展生存期(PFS)及总生存期(OS)。
客观缓解率(Objective Response Rate,ORR)：为完全缓解(CR)和部分缓解(PR)的受试者的比例；缓解持续时间(Duration of Response,DOR)：为肿瘤第一次评估为客观缓解(Objective Response，OR)至第一次评估为PD(Progressive Disease)或PD前任何原因死亡的时间，反映ORR的持续时间；无进展生存期(Progression-Free Survival：PFS)：为从首次给药至出现肿瘤客观进展或全因死亡的时间(以先发生者为准)。“MTD”为某一剂量组在DLT评估期内观察到≤1/6的受试者中探索到的DLT的最高剂量水平。
“剂量限制性毒性(DLT)”为在DLT观察期发生的不良事件(AE)并被认为至少可能与研究药物相关，具体如下：
·5级毒性；
■血液学毒性：
·4级血液学毒性(注：3～4级淋巴细胞减少症不计入DLT)；
·持续任何时间的4级血小板减少症；
·伴有出血倾向的或需要输注血小板的3级血小板减少症；
·持续任何时间的4级中性粒细胞减少症；
·3级的中性粒细胞减少伴感染或持续时间≥7天或3级中性粒细胞减少伴发热；
■非血液学毒性：
·3、4级非血液学毒性(注：①3级恶心、呕吐、皮疹除外；②经过临床干预后可控制在3天以内的3级腹泻除外；③持续时间在7天以内疲劳/乏力除外)；
·符合任一以下条件的、因临床实验室检查发现的3、4级非血液学毒性：①需要进行临床干预的；②导致住院治疗的；③持续时间≥7天的；④所有导致受试者在第1周期中停止治疗的；⑤任何导致第2周期给药延迟超过2周的治疗相关毒性；
·≥2级的脑部病变；
·需进行系统治疗的眼毒性(注：任何出现视力变化或眼部毒性达2级或更高的病人都将由眼科医生进行评估)；
在DLT判定中，一些明显且无可争议的由于疾病进展或DLT以外的原因而导致的AE则不应计入DLT。在为期21天的DLT观察期内，由于非DLT原因(如与基础疾病、疾病进展、伴随用药或伴随疾病明显相关的不良事件)而退出的受试者将被视为不可评估，并将被替换。若受试者是主动退出研究，须尽快、按要求完成末次访视。被替补的受试者在统计时纳入SS集，但在计算DLT事件发生比例时不计入该组别总人数中。经申办方与研究者一致同意后，允许对部分受试者进行重复筛选，需重新分配筛选号。

Claims (10)

  1. 一种用于治疗肿瘤或癌症的方法，其包括：向有需要的患者给药有效量的抗TIGIT抗体或抗原结合片段，所述有效量为每个治疗周期9mg至1200mg；
    所述抗TIGIT抗体或抗原结合片段包含SEQ ID NO:1所示的HCDR1、SEQ ID NO:2所示的HCDR2、SEQ ID NO:3所示的HCDR3、SEQ ID NO:4所示的LCDR1、SEQ ID NO:5所示的LCDR2和SEQ ID NO:6所示的LCDR3。
  2. 如权利要求1所述的方法，所述抗TIGIT抗体或抗原结合片段包含重链可变区和轻链可变区，其中：
    所述重链可变区包含SEQ ID NO:7所示的氨基酸序列，或与SEQ ID NO:7所示序列相比具有至少80％同一性的氨基酸序列，或与SEQ ID NO:7所示序列相比具有一个或多个保守氨基酸取代的氨基酸序列；和/或
    所述轻链可变区包含SEQ ID NO:8所示的序列，或与SEQ ID NO:8所示序列相比具有至少80％同一性的氨基酸序列，或与SEQ ID NO:8所示序列相比具有一个或多个保守氨基酸取代的氨基酸序列。
  3. 如权利要求1或2所述的方法，所述抗TIGIT抗体的重链包含SEQ ID NO:9所示的氨基酸序列，或与SEQ ID NO:9所示序列相比具有至少80％同一性的氨基酸序列，或与SEQ ID NO:9所示序列相比具有一个或多个保守氨基酸取代的氨基酸序列；和/或
    所述抗TIGIT抗体的轻链包含SEQ ID NO:10所示的氨基酸序列，或与SEQ ID NO:10所示序列相比具有至少80％同一性的氨基酸序列，或与SEQ ID NO:10所示序列相比具有一个或多个保守氨基酸取代的氨基酸序列。
  4. 如权利要求1-3任一项所述的方法，所述抗TIGIT抗体或抗原结合片段的岩藻糖基化水平为0-10％。
  5. 如权利要求1-4任一项所述的方法，所述抗TIGIT抗体或抗原结合片段由α-(1,6)-岩藻糖基转移酶基因敲除的细胞表达；或者，所述抗TIGIT抗体或抗原结合片段由α-(1,6)-岩藻糖基转移酶基因敲除的CHO细胞表达。
  6. 如权利要求1-5任一项所述的方法，所述肿瘤或癌症选自于急性淋巴细胞性白血病、急性骨髓性白血病、慢性淋巴细胞性白血病、慢性骨髓性白血病、骨髓性增生 疾病/肿瘤、霍奇金淋巴瘤、无痛性和侵袭性非霍奇金淋巴瘤、伯基特淋巴瘤、滤泡性淋巴瘤、多发性骨髓瘤、巨细胞骨髓瘤、重链骨髓瘤、轻链或本斯-琼斯骨髓瘤、乳腺癌、卵巢癌、胰腺癌、前列腺癌、黑素瘤、结直肠癌、结肠癌、肺癌、头颈癌、膀胱癌、食道癌、肝癌和肾癌。
  7. 如权利要求1-6任一项所述的方法，一个治疗周期为1周、2周、3周、4周、5周、6周或7周。
  8. 如权利要求1-7任一项所述的方法，所述抗TIGIT抗体或抗原结合片段的单次给药量为0.01mg/kg-26mg/kg。
  9. 如权利要求1-8任一项所述的方法，给药方式为静脉注射或皮下注射。
  10. 一种试剂盒，其包含抗TIGIT抗体或抗原结合片段和用于指导有需要患者给药抗TIGIT抗体或抗原结合片段的说明书；
    所述抗TIGIT抗体或抗原结合片段包含SEQ ID NO:1所示的HCDR1、SEQ ID NO:2所示的HCDR2、SEQ ID NO:3所示的HCDR3、SEQ ID NO:4所示的LCDR1、SEQ ID NO:5所示的LCDR2和SEQ ID NO:6所示的LCDR3。
PCT/CN2022/078683 2021-03-02 2022-03-01 抗tigit抗体在治疗肿瘤或癌症中的应用 WO2022184068A1 (zh)

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