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WO2023274275A1 - 药物组合及其用途 - Google Patents

药物组合及其用途 Download PDF

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Publication number
WO2023274275A1
WO2023274275A1 PCT/CN2022/102169 CN2022102169W WO2023274275A1 WO 2023274275 A1 WO2023274275 A1 WO 2023274275A1 CN 2022102169 W CN2022102169 W CN 2022102169W WO 2023274275 A1 WO2023274275 A1 WO 2023274275A1
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Prior art keywords
amino acid
acid sequence
seq
sequence shown
terminal
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Ceased
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PCT/CN2022/102169
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English (en)
French (fr)
Inventor
王晨光
张晓娇
党晓萌
张晋
付雨婷
王一涵
王超
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Tianjin Lipogen Technology Co Ltd
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Tianjin Lipogen Technology Co Ltd
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Application filed by Tianjin Lipogen Technology Co Ltd filed Critical Tianjin Lipogen Technology Co Ltd
Priority to CN202280045663.8A priority Critical patent/CN117881419A/zh
Priority to US18/574,085 priority patent/US20250270316A1/en
Publication of WO2023274275A1 publication Critical patent/WO2023274275A1/zh
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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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
    • 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

  • This application relates to the field of biomedicine, in particular to the development and application of a drug combination.
  • the PD-L1/PD-1 signaling pathway is a very important co-inhibitory signaling pathway in the immune response.
  • ITAM immunoreceptor tyrosine activation motif
  • PD-1 is found to be highly expressed on the surface of CD8+ T cells that specifically recognize HIV, and the virus activates the PD-L1/PD-1 signaling pathway to make CD8+ T cells that specifically recognize HIV
  • the activity is inhibited, the secretion ability of cytokines and the proliferation ability of T cells are greatly weakened, and the adaptive immune function defect is caused.
  • TLRs Toll-like receptors
  • PRRs pattern recognition receptors
  • PAMPs pathogen-associated molecular patterns
  • DAMP damage-related molecular patterns
  • chemokines and inflammatory cytokines such as IL-1, IL-6, TNF- ⁇ , etc.
  • TLRs play an important role in antiviral innate immunity.
  • TLRs related to virus recognition and body antiviral immunity mainly include TLR2, TLR3, TLR4, TLR7, TLR8, and TLR9.
  • TLR2 and TLR4 located on the surface of the cell membrane mainly recognize the envelope protein of the virus;
  • TLR3, TLR7, TLR8 and TLR9 located in the cell mainly recognize the viral nucleic acid.
  • TLR7/8 is one of the members of Toll like receptor (TLR), which is mainly distributed in plasma dendritic cells (pDC) and B cells. TLR7/8 mainly recognizes ssRNA viruses and recognizes and clears them in the human body important role in pathogenic microorganisms.
  • TLR Toll like receptor
  • PRRs pattern recognition receptors
  • Toll-like receptors 7 and 8 are important PRRs among them, which can stimulate Antigen-presenting cells induce dendritic cells to secrete a variety of cytokines and express a variety of co-stimulatory molecules to stimulate interferon- ⁇ , tumor necrosis factor (TNF) and interleukins (IL-1, IL-6 and IL-8, etc.
  • TNF tumor necrosis factor
  • IL-1, IL-6 and IL-8 interleukins
  • synthesis thereby activating the body's innate immune response, and at the same time, it can also activate pDC, improve the anti-presentation ability of pDC, promote the proliferation of CD4+T cells, and further activate CD8+T cells, kill tumor cells, and enhance the body's anti-virus and antitumor effects.
  • the application provides a drug combination and its application in antitumor drugs.
  • the drug combination consists mainly of two parts:
  • Immune checkpoint inhibitors such as PD-1/PD-L1
  • TLR agonists such as imidazoquinoline derivatives.
  • the drug combination of the application can prepare high-efficiency, low-toxicity antitumor drugs.
  • Combining PD-L1/PD-1 inhibitors with TLR agonists on the one hand, releases inhibitory signals, enhances T cell activation, and promotes the adaptive immune system response; on the other hand, induces the expression of INF and cytokines, and promotes T cell Aggregation, activation of the innate immune system response, and a two-pronged approach to enhance the response of cytotoxic cells to tumors have high clinical prospects and application value.
  • the present application provides a pharmaceutical combination, which comprises a programmed cell death protein 1 (PD-1) inhibitor and/or a programmed death ligand 1 (PD-L1) inhibitor, and a TLR agonist.
  • a pharmaceutical combination which comprises a programmed cell death protein 1 (PD-1) inhibitor and/or a programmed death ligand 1 (PD-L1) inhibitor, and a TLR agonist.
  • TLRs include TLR1, TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9 and/or TLR10.
  • the TLR agonist is selected from one or more of TLR7 agonists, TLR8 agonists, and TLR9 agonists.
  • the TLR agonist comprises a dual agonist of TLR7 and TLR8 (TLR7/TLR8 agonist).
  • TLR agonists include dsRNA, ssRNA, CpG DNA, imidazoquinoline derivatives and/or guanosine analogs.
  • the TLR agonist comprises an imidazoquinoline derivative.
  • TLR agonist is selected from the group consisting of Imiquimod, Gardiquimod, Resiquimod, 1V209, Selgantolimod (GS-9688), Vesatolimod ( GS-9620), Sumanirole, PF-4878691, derivatives thereof, and analogs thereof.
  • the TLR agonist includes imiquimod, resiquimod or a pharmaceutically acceptable salt thereof.
  • the TLR agonist is selected from LHC-165, NKTR-262, DN1508052-01, SHR2150, CL307, CL264, loxoribine, exartoribine, DSR-6434, GSK2245035, One or more of SM-276001, SM-324405, SM-324406, AZ12441970 and AZ12443988, their derivatives, and their analogs.
  • the PD-1 inhibitor has one or more of the following characteristics:
  • PD-1 Inhibit or reduce the expression of PD-1, such as the transcription or translation of PD-1;
  • Inhibiting or reducing PD-1 activity such as inhibiting or reducing the binding of PD-1 to its cognate ligand, such as PD-L1 or PD-L2;
  • the PD-1 inhibitor comprises an anti-PD-1 antibody or an antigen-binding fragment thereof.
  • the anti-PD-1 antibody is selected from Pembrolizumab, Nivolumab, Pidilizumab, Tislelizumab, Camrelizumab (SHR-1210), Sintilimab, Toripalimab, MEDI0680, BGB-A317, TSR-042, REGN2810, PF- 06801591, RB0004, analogs thereof, and combinations thereof.
  • the anti-PD-1 antibody comprises at least one CDR in the antibody heavy chain variable region (VH)
  • the VH comprises the amino acid sequence shown in SEQ ID NO:8.
  • the anti-PD-1 antibody comprises VH
  • the VH comprises HCDR3
  • the HCDR3 comprises the amino acid sequence shown in SEQ ID NO:3.
  • VH further comprises HCDR2, wherein said HCDR2 comprises the amino acid sequence shown in SEQ ID NO:2.
  • VH further comprises HCDR1, wherein said HCDR1 comprises the amino acid sequence shown in SEQ ID NO:1.
  • the VH comprises HCDR1, HCDR2 and HCDR3, wherein the HCDR3 comprises the amino acid sequence shown in SEQ ID NO:3, and the HCDR2 comprises the amino acid sequence shown in SEQ ID NO:2, the Said HCDR1 comprises the amino acid sequence shown in SEQ ID NO:1.
  • the C-terminal of the HFR1 is directly or indirectly linked to the N-terminal of the HCDR1, and the HFR1 comprises the amino acid sequence shown in SEQ ID NO: 4 or An amino acid sequence having at least about 70% sequence identity to the amino acid sequence set forth in SEQ ID NO:4.
  • the VH includes a framework region HFR2
  • the N-terminal of the HFR2 is directly or indirectly connected to the C-terminal of the HCDR1
  • the C-terminal of the HFR2 is directly or indirectly connected to the N-terminal of the HCDR2 Indirectly connected
  • the HFR2 comprises the amino acid sequence shown in SEQ ID NO: 5 or an amino acid sequence having at least about 70% sequence identity with the amino acid sequence shown in SEQ ID NO: 5.
  • the VH includes a framework region HFR3
  • the N-terminal of the HFR3 is directly or indirectly connected to the C-terminal of the HCDR2
  • the C-terminal of the HFR3 is directly or indirectly connected to the N-terminal of the HCDR3 Indirectly linked
  • the HFR3 comprises the amino acid sequence shown in SEQ ID NO: 6 or an amino acid sequence having at least about 70% sequence identity with the amino acid sequence shown in SEQ ID NO: 6.
  • the N-terminal of the HFR4 is directly or indirectly connected to the C-terminal of the HCDR3, and the HFR4 comprises the amino acid sequence shown in SEQ ID NO: 7 or An amino acid sequence having at least about 70% sequence identity to the amino acid sequence set forth in SEQ ID NO:7.
  • the C-terminus of HFR1 is directly or indirectly connected to the N-terminus of the HCDR1, and the N-terminus of the HFR2 is connected to the N-terminus of the HCDR1
  • the C-terminal of the HFR2 is directly or indirectly connected to the C-terminal of the HCDR2
  • the C-terminal of the HFR2 is directly or indirectly connected to the N-terminal of the HCDR2
  • the N-terminal of the HFR3 is directly or indirectly connected to the C-terminal of the HCDR2
  • the HFR3 The C-terminal is directly or indirectly connected to the N-terminal of the HCDR3, and the N-terminal of the HFR4 is directly or indirectly connected to the C-terminal of the HCDR3;
  • the HFR1 comprises the amino acid sequence shown in SEQ ID NO: 4 or with The amino acid sequence shown in SEQ ID NO: 4 has an amino acid sequence
  • said anti-PD-1 antibody comprises VH
  • said VH comprises the amino acid sequence shown in SEQ ID NO:8.
  • the anti-PD-1 antibody comprises an antibody heavy chain (HC)
  • the HC comprises an amino acid sequence shown in SEQ ID NO:9.
  • the anti-PD-1 antibody comprises at least one CDR in the antibody light chain variable region (VL)
  • the VL comprises the amino acid sequence shown in SEQ ID NO:17.
  • said anti-PD-1 antibody comprises at least one CDR in VH
  • said VH comprises the amino acid sequence shown in SEQ ID NO: 8
  • said anti-PD-1 antibody comprises at least one CDR in VL At least one CDR
  • the VL comprises the amino acid sequence shown in SEQ ID NO:17.
  • the anti-PD-1 antibody comprises a VL
  • the VL comprises LCDR1
  • the LCDR1 comprises the amino acid sequence shown in SEQ ID NO:10.
  • said VL further comprises LCDR2, wherein said LCDR2 comprises the amino acid sequence shown in SEQ ID NO: 11.
  • said VL further comprises LCDR3, wherein said LCDR3 comprises the amino acid sequence shown in SEQ ID NO:12.
  • the VL comprises LCDR1, LCDR2 and LCDR3, wherein the LCDR1 comprises the amino acid sequence shown in SEQ ID NO: 10, and the LCDR2 comprises the amino acid sequence shown in SEQ ID NO: 11, the Said LCDR3 comprises the amino acid sequence shown in SEQ ID NO:12.
  • the anti-PD-1 antibody comprises VH and antibody VL
  • the VH comprises HCDR1, HCDR2 and HCDR3, wherein the HCDR3 comprises the amino acid sequence shown in SEQ ID NO: 3, and the HCDR2 comprises SEQ ID NO: 3
  • the HCDR1 comprises the amino acid sequence shown in SEQ ID NO:1
  • the VL comprises LCDR1, LCDR2 and LCDR3, wherein the LCDR1 comprises the amino acid shown in SEQ ID NO:10 Sequence
  • the LCDR2 comprises the amino acid sequence shown in SEQ ID NO:11
  • the LCDR3 comprises the amino acid sequence shown in SEQ ID NO:12.
  • the C-terminus of the LFR1 is directly or indirectly connected to the N-terminus of the LCDR1, and the LFR1 comprises the amino acid sequence shown in SEQ ID NO: 13 or An amino acid sequence having at least about 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 13.
  • the N-terminal of the LFR2 is directly or indirectly connected to the C-terminal of the LCDR1, and the C-terminal of the LFR2 is directly or indirectly connected to the N-terminal of the LCDR2.
  • the LFR2 comprises the amino acid sequence shown in SEQ ID NO: 14 or an amino acid sequence having at least about 70% sequence identity with the amino acid sequence shown in SEQ ID NO: 14.
  • the VL includes a framework region LFR3
  • the N-terminal of the LFR3 is directly or indirectly connected to the C-terminal of the LCDR2
  • the C-terminal of the LFR3 is directly or indirectly connected to the N-terminal of the LCDR3 Indirectly connected
  • the LFR3 comprises the amino acid sequence shown in SEQ ID NO: 15 or an amino acid sequence having at least about 70% sequence identity with the amino acid sequence shown in SEQ ID NO: 15.
  • the N-terminal of the LFR4 is directly or indirectly connected to the C-terminal of the LCDR3, and the LFR4 comprises the amino acid sequence shown in SEQ ID NO: 16 or An amino acid sequence having at least about 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 16.
  • the VL includes framework regions LFR1, LFR2, LFR3 and LFR4, the C-terminus of the LFR1 is directly or indirectly connected to the N-terminus of the LCDR1, and the N-terminus of the LFR2 is connected to the LCDR1
  • the C-terminal of the LFR2 is directly or indirectly connected to the C-terminal of the LCDR2
  • the C-terminal of the LFR2 is directly or indirectly connected to the N-terminal of the LCDR2
  • the N-terminal of the LFR3 is directly or indirectly connected to the C-terminal of the LCDR2
  • the LFR3 is directly or indirectly connected to the C-terminal of the LCDR2
  • the LFR3 The C-terminus is directly or indirectly connected to the N-terminus of the LCDR3, and the N-terminus of the LFR4 is directly or indirectly connected to the C-terminus of the LCDR3;
  • the LFR1 comprises the amino acid sequence shown in SEQ ID NO: 13 or with The amino acid sequence shown in SEQ ID
  • the VL comprises the amino acid sequence shown in SEQ ID NO:17.
  • the anti-PD-1 antibody comprises VH and VL
  • the VH comprises the amino acid sequence shown in SEQ ID NO:8
  • the VL comprises the amino acid sequence shown in SEQ ID NO:17 .
  • said anti-PD-1 antibody comprises an antibody light chain (LC)
  • said LC comprises an amino acid sequence shown in SEQ ID NO:18.
  • the anti-PD-1 antibody comprises HC and LC
  • the HC comprises the amino acid sequence shown in SEQ ID NO:9
  • the LC comprises the amino acid sequence shown in SEQ ID NO:18.
  • the PD-L1 inhibitor has one or more of the following characteristics:
  • PD-L1 Inhibit or reduce the expression of PD-L1, such as the transcription or translation of PD-L1;
  • Inhibiting or reducing PD-L1 activity such as inhibiting or reducing the binding of PD-L1 to its cognate receptor, such as PD-1;
  • the PD-L1 inhibitor comprises an anti-PD-L1 antibody or an antigen-binding fragment thereof.
  • the anti-PD-L1 antibody is selected from Durvalumab, Atezolizumab, Avelumab, MDX-1105, YW243.55.S70, MDPL3280A, AMP-224, LY3300054, RB0005, analogs thereof, and combinations thereof .
  • said anti-PD-L1 antibody comprises at least one CDR in VH
  • said VH comprises the amino acid sequence shown in SEQ ID NO:25.
  • the anti-PD-L1 antibody comprises VH
  • the VH comprises HCDR3
  • the HCDR3 comprises the amino acid sequence shown in SEQ ID NO:21.
  • VH further comprises HCDR2, wherein said HCDR2 comprises the amino acid sequence shown in SEQ ID NO: 20.
  • VH further comprises HCDR1, wherein said HCDR1 comprises the amino acid sequence shown in SEQ ID NO: 19.
  • the VH comprises HCDR1, HCDR2 and HCDR3, wherein the HCDR3 comprises the amino acid sequence shown in SEQ ID NO: 21, and the HCDR2 comprises the amino acid sequence shown in SEQ ID NO: 20, the Said HCDR1 comprises the amino acid sequence shown in SEQ ID NO:19.
  • the C-terminal of the HFR1 is directly or indirectly connected to the N-terminal of the HCDR1, and the HFR1 comprises the amino acid sequence shown in SEQ ID NO: 22 or An amino acid sequence having at least about 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 22.
  • the N-terminal of the HFR2 is directly or indirectly connected to the C-terminal of the HCDR1
  • the C-terminal of the HFR2 is directly or indirectly connected to the N-terminal of the HCDR2 Indirectly linked
  • the HFR2 comprises the amino acid sequence shown in SEQ ID NO: 23 or an amino acid sequence having at least about 70% sequence identity with the amino acid sequence shown in SEQ ID NO: 23.
  • the VH includes a framework region HFR3
  • the N-terminal of the HFR3 is directly or indirectly connected to the C-terminal of the HCDR2
  • the C-terminal of the HFR3 is directly or indirectly connected to the N-terminal of the HCDR3 Indirectly connected
  • the HFR3 comprises the amino acid sequence shown in SEQ ID NO: 24 or an amino acid sequence having at least about 70% sequence identity with the amino acid sequence shown in SEQ ID NO: 24.
  • the N-terminal of the HFR4 is directly or indirectly connected to the C-terminal of the HCDR3, and the HFR4 comprises the amino acid sequence shown in SEQ ID NO: 7 or An amino acid sequence having at least about 70% sequence identity to the amino acid sequence set forth in SEQ ID NO:7.
  • the C-terminus of HFR1 is directly or indirectly connected to the N-terminus of the HCDR1, and the N-terminus of the HFR2 is connected to the N-terminus of the HCDR1
  • the C-terminal of the HFR2 is directly or indirectly connected to the C-terminal of the HCDR2
  • the C-terminal of the HFR2 is directly or indirectly connected to the N-terminal of the HCDR2
  • the N-terminal of the HFR3 is directly or indirectly connected to the C-terminal of the HCDR2
  • the HFR3 The C-terminal is directly or indirectly connected to the N-terminal of the HCDR3, and the N-terminal of the HFR4 is directly or indirectly connected to the C-terminal of the HCDR3;
  • the HFR1 comprises the amino acid sequence shown in SEQ ID NO: 22 or with The amino acid sequence shown in SEQ ID NO:22 has an amino acid sequence
  • said anti-PD-L1 antibody comprises VH
  • said VH comprises the amino acid sequence shown in SEQ ID NO:25.
  • said anti-PD-L1 antibody comprises HC
  • said HC comprises the amino acid sequence shown in SEQ ID NO:26.
  • the VL comprises the amino acid sequence shown in SEQ ID NO:37.
  • said anti-PD-L1 antibody comprises at least one CDR in VH
  • said VH comprises the amino acid sequence shown in SEQ ID NO: 25
  • said anti-PD-L1 antibody comprises at least one CDR in VL At least one CDR
  • the VL comprises the amino acid sequence shown in SEQ ID NO:37.
  • said anti-PD-L1 antibody comprises at least one CDR in VH
  • said VH comprises the amino acid sequence shown in SEQ ID NO: 25
  • said anti-PD-L1 antibody comprises at least one CDR in VL At least one CDR
  • the VL comprises the amino acid sequence shown in SEQ ID NO:38, SEQ ID NO:39 or SEQ ID NO:40.
  • the anti-PD-L1 antibody comprises a VL
  • the VL comprises LCDR1
  • the LCDR1 comprises the amino acid sequence shown in SEQ ID NO:27.
  • the anti-PD-L1 antibody comprises VL
  • the VL comprises LCDR1
  • the LCDR1 comprises the amino acid sequence shown in SEQ ID NO:28, SEQ ID NO:29 or SEQ ID NO:30 .
  • said VL further comprises LCDR2, wherein said LCDR2 comprises the amino acid sequence shown in SEQ ID NO:31.
  • said VL further comprises LCDR3, wherein said LCDR3 comprises the amino acid sequence shown in SEQ ID NO:32.
  • said VL comprises LCDR1, LCDR2 and LCDR3, wherein said LCDR1 comprises the amino acid sequence shown in SEQ ID NO: 27, and said LCDR2 comprises the amino acid sequence shown in SEQ ID NO: 31, said Said LCDR3 comprises the amino acid sequence shown in SEQ ID NO:32.
  • said VL comprises LCDR1, LCDR2 and LCDR3, wherein said LCDR1 comprises the amino acid sequence shown in SEQ ID NO: 28, and said LCDR2 comprises the amino acid sequence shown in SEQ ID NO: 31, said Said LCDR3 comprises the amino acid sequence shown in SEQ ID NO:32;
  • the LCDR1 comprises the amino acid sequence shown in SEQ ID NO:29
  • the LCDR2 comprises the amino acid sequence shown in SEQ ID NO:31
  • the LCDR3 comprises the amino acid sequence shown in SEQ ID NO:32;
  • the LCDR1 comprises the amino acid sequence shown in SEQ ID NO:30
  • the LCDR2 comprises the amino acid sequence shown in SEQ ID NO:31
  • the LCDR3 comprises the amino acid sequence shown in SEQ ID NO:32.
  • the anti-PD-L1 antibody comprises VH and antibody VL
  • the VH comprises HCDR1, HCDR2 and HCDR3, wherein the HCDR3 comprises the amino acid sequence shown in SEQ ID NO: 21, and the HCDR2 Comprising the amino acid sequence shown in SEQ ID NO:20
  • the HCDR1 includes the amino acid sequence shown in SEQ ID NO:19
  • the VL includes LCDR1, LCDR2 and LCDR3, wherein the LCDR1 includes the amino acid sequence shown in SEQ ID NO:27
  • the amino acid sequence of the LCDR2 comprises the amino acid sequence shown in SEQ ID NO:31
  • the LCDR3 comprises the amino acid sequence shown in SEQ ID NO:32.
  • the VH comprises HCDR1, HCDR2 and HCDR3, wherein the HCDR3 comprises the amino acid sequence shown in SEQ ID NO: 21, and the HCDR2 Comprising the amino acid sequence shown in SEQ ID NO: 20, the HCDR1 includes the amino acid sequence shown in SEQ ID NO: 19; and the VL includes LCDR1, LCDR2 and LCDR3, wherein the LCDR1 includes SEQ ID NO: 28, SEQ ID NO: The amino acid sequence shown in ID NO: 29 or SEQ ID NO: 30, the LCDR2 includes the amino acid sequence shown in SEQ ID NO: 31, and the LCDR3 includes the amino acid sequence shown in SEQ ID NO: 32.
  • the C-terminus of the LFR1 is directly or indirectly connected to the N-terminus of the LCDR1, and the LFR1 comprises the amino acid sequence shown in SEQ ID NO: 33 or An amino acid sequence having at least about 70% sequence identity to the amino acid sequence set forth in SEQ ID NO:33.
  • the N-terminal of the LFR2 is directly or indirectly connected to the C-terminal of the LCDR1, and the C-terminal of the LFR2 is directly or indirectly connected to the N-terminal of the LCDR2.
  • the LFR2 comprises the amino acid sequence shown in SEQ ID NO: 34 or an amino acid sequence having at least about 70% sequence identity with the amino acid sequence shown in SEQ ID NO: 34.
  • the VL includes a framework region LFR3
  • the N-terminal of the LFR3 is directly or indirectly connected to the C-terminal of the LCDR2
  • the C-terminal of the LFR3 is directly or indirectly connected to the N-terminal of the LCDR3 Indirectly linked
  • the LFR3 comprises the amino acid sequence shown in SEQ ID NO: 35 or an amino acid sequence having at least about 70% sequence identity with the amino acid sequence shown in SEQ ID NO: 35.
  • the N-terminal of the LFR4 is directly or indirectly connected to the C-terminal of the LCDR3, and the LFR4 comprises the amino acid sequence shown in SEQ ID NO: 36 or An amino acid sequence having at least about 70% sequence identity to the amino acid sequence set forth in SEQ ID NO:36.
  • the VL includes framework regions LFR1, LFR2, LFR3 and LFR4, the C-terminus of the LFR1 is directly or indirectly connected to the N-terminus of the LCDR1, and the N-terminus of the LFR2 is connected to the LCDR1
  • the C-terminal of the LFR2 is directly or indirectly connected to the C-terminal of the LCDR2
  • the C-terminal of the LFR2 is directly or indirectly connected to the N-terminal of the LCDR2
  • the N-terminal of the LFR3 is directly or indirectly connected to the C-terminal of the LCDR2
  • the LFR3 is directly or indirectly connected to the C-terminal of the LCDR2
  • the LFR3 The C-terminus is directly or indirectly connected to the N-terminus of the LCDR3, and the N-terminus of the LFR4 is directly or indirectly connected to the C-terminus of the LCDR3;
  • the LFR1 comprises the amino acid sequence shown in SEQ ID NO: 33 or with The amino acid sequence shown in SEQ ID
  • the anti-PD-L1 antibody comprises a VL
  • the VL comprises the amino acid sequence shown in SEQ ID NO:37.
  • the anti-PD-L1 antibody comprises a VL
  • the VL comprises the amino acid sequence shown in SEQ ID NO:38, SEQ ID NO:39 or SEQ ID NO:40.
  • the anti-PD-L1 antibody comprises VH and VL
  • the VH comprises the amino acid sequence shown in SEQ ID NO:25
  • the VL comprises the amino acid sequence shown in SEQ ID NO:37 .
  • the anti-PD-L1 antibody comprises VH and VL
  • the VH comprises the amino acid sequence shown in SEQ ID NO:25
  • the VL comprises SEQ ID NO:38, SEQ ID NO: 39 or the amino acid sequence shown in SEQ ID NO:40.
  • said anti-PD-L1 antibody comprises LC
  • said LC comprises the amino acid sequence shown in SEQ ID NO:41.
  • the LC comprises the amino acid sequence shown in SEQ ID NO:42, SEQ ID NO:43 or SEQ ID NO:44.
  • the anti-PD-L1 antibody comprises HC and LC
  • the HC comprises the amino acid sequence shown in SEQ ID NO:26
  • the LC comprises the amino acid sequence shown in SEQ ID NO:41.
  • the anti-PD-L1 antibody comprises HC and LC
  • the HC comprises the amino acid sequence shown in SEQ ID NO:26
  • the LC comprises SEQ ID NO:42, SEQ ID NO:43 Or the amino acid sequence shown in SEQ ID NO:44.
  • i) the PD-1 inhibitor and/or PD-L1 inhibitor, and ii) the TLR agonist in the drug combination are not mixed with each other in the drug combination.
  • the PD-1 inhibitor and/or PD-L1 inhibitor and ii) the TLR agonist in the pharmaceutical combination are in a single dosage form.
  • the pharmaceutical combination is formulated as a pharmaceutical composition.
  • the pharmaceutical composition includes a PD-1 inhibitor or PD-L1 inhibitor, and a TLR agonist.
  • TLR agonist is present in an amount from about 0.0001 mg/kg to about 200 mg/kg.
  • the PD-1 inhibitor or PD-L1 inhibitor is present in an amount of 0.0001 mg/kg to about 200 mg/kg.
  • the pharmaceutical composition further comprises one or more pharmaceutically acceptable carriers.
  • the present application also provides the use of the aforementioned drug combination in the preparation of medicines for treating neoplastic diseases.
  • the neoplastic disease comprises tumor and/or wart disease.
  • the present application also provides the aforementioned drug combination for treating neoplastic diseases.
  • the present application also provides a drug for treating neoplastic diseases, which comprises the aforementioned drug combination.
  • the present application also provides a method for treating neoplastic diseases, which comprises administering an effective amount of the aforementioned drug combination to a subject in need.
  • the neoplasm comprises tumors and/or warts.
  • administering comprises topical, intraneoplastic (eg, intratumor or intrawart) or systemic administration.
  • intraneoplastic eg, intratumor or intrawart
  • systemic administration e.g, topical, intraneoplastic (eg, intratumor or intrawart) or systemic administration.
  • administering comprises intravenous injection, intravenous drip, intramuscular injection, subcutaneous injection and/or intraneoplastic injection.
  • a PD-1 inhibitor or a PD-L1 inhibitor, and ii) a TLR agonist in the drug combination are administered by the same or different routes of administration.
  • it comprises injecting said TLR agonist into the neoplasm.
  • it also includes injecting or systemically infusing the PD-1 inhibitor or PD-L1 inhibitor into the neoplasm.
  • it comprises injecting i) the PD-1 inhibitor or PD-L1 inhibitor, and ii) the TLR agonist of the drug combination into the neoplasm.
  • i) the PD-1 inhibitor or PD-L1 inhibitor and ii) the TLR agonist in the pharmaceutical combination are administered simultaneously or at different times.
  • the PD-1 inhibitor or PD-L1 inhibitor is administered before and/or after administration of the TLR agonist.
  • the method comprises: i) injecting the TLR agonist into the neoplasm; ii) injecting or systemically infusing the PD-1 inhibitory agent into the neoplasm after administering the TLR agonist agents or PD-L1 inhibitors.
  • the method comprises: i) injecting the STING pathway agonist into the neoplasm; ii) systemically infusing the PD-1 inhibitor or PD- L1 inhibitor.
  • the PD-1 inhibitor or PD-L1 inhibitor is administered about 2 hours to about 72 hours after the TLR agonist is administered.
  • the PD-1 inhibitory agent is administered at about 2h, about 4h, about 8h, about 16h, about 24h, about 36h, about 48h, about 60h, or about 72h after administration of the TLR agonist agents or PD-L1 inhibitors.
  • the method comprises: i) injecting the STING pathway agonist into the neoplasm; ii) injecting or systemically infusing the STING pathway agonist into the neoplasm about 48 hours after administration of the STING pathway agonist.
  • the PD-1 inhibitor or PD-L1 inhibitor in the drug combination is administered simultaneously with ii) the TLR agonist.
  • the PD-1 inhibitor or PD-L1 inhibitor in the drug combination is administered simultaneously with ii) the TLR agonist by intraneoplastic injection, and the PD- 1 inhibitor or PD-L1 inhibitor and said TLR agonist are located in the same dosage form.
  • the PD-1 inhibitor or PD-L1 inhibitor in the drug combination is administered simultaneously with ii) the TLR agonist by intraneoplastic injection, and the PD- 1 inhibitor or PD-L1 inhibitor in a separate dosage form from said TLR agonist.
  • the present application provides a kit comprising the pharmaceutical combination described in the present application.
  • Figure 1 shows the in vivo drug efficacy study of the PD-L1 inhibitor RB0005 and Imiquimod drug combination described in the application-the body weight changes of mice in each group during the administration period;
  • Figure 2 shows the in vivo drug efficacy study of the PD-L1 inhibitor RB0005 and Imiquimod drug combination described in the application - the change in the survival rate of mice in each group during the administration period;
  • Figure 3A shows the in vivo pharmacodynamic study of the PD-L1 inhibitor RB0005 and Imiquimod drug combination described in this application - the changes in tumor proliferation of mice in each group during the administration period;
  • Figure 3B shows the in vivo efficacy study of the PD-L1 inhibitor RB0005 and Imiquimod drug combination described in this application - the changes in tumor proliferation of mice in each group on day 15 after treatment;
  • Figure 4 shows the in vivo efficacy study of the PD-1 inhibitor RB0004 and Imiquimod drug combination described in this application - the change in the survival rate of mice in each group during the administration period;
  • Figure 5A shows the in vivo efficacy study of the PD-1 inhibitor RB0004 and Imiquimod drug combination described in this application - the changes in tumor proliferation of mice in each group during administration;
  • Figure 5B shows the in vivo pharmacodynamic study of the PD-1 inhibitor RB0004 and Imiquimod drug combination described in this application - the changes in tumor proliferation in mice in each group on day 15 after treatment;
  • Figure 6 shows the in vivo pharmacodynamic study of the PD-1 inhibitor RB0004 and Imiquimod drug combination described in this application - the inhibition rate of tumor weight in mice dissected after the observation of administration;
  • Figure 7 shows the in vivo pharmacodynamic study of the PD-L1 inhibitor RB0005 described in this application and different doses of Imiquimod drug combinations - changes in the survival rate of mice in each group during the administration period;
  • Figure 8 shows the in vivo pharmacodynamic study of the PD-L1 inhibitor RB0005 described in this application and different doses of Imiquimod drug combination - the inhibition rate of tumor weight in mice dissected after the observation of administration;
  • Figure 9 shows the in vivo pharmacodynamic study of the PD-L1 inhibitor RB0005 described in the present application and different doses of Imiquimod drug combinations - changes in body weight of mice in each group during administration;
  • Figure 10 shows the in vivo drug efficacy study of the drug combination of PD-L1 inhibitor RB0005 described in the present application and different doses of Imiquimod (including 5% Imiquimod cream) - changes in the survival rate of mice in each group during administration;
  • Figure 11 shows the in vivo efficacy study of the drug combination of the PD-L1 inhibitor RB0005 described in this application and different doses of Imiquimod (including 5% Imiquimod cream) - the inhibition rate of tumor weight in mice dissected after the observation of administration;
  • Figure 12 shows the in vivo pharmacodynamic study of the drug combination of PD-1 inhibitor RB0004 described in the present application and different doses of Imiquimod (including 5% Imiquimod cream) - changes in the survival rate of mice in each group during administration;
  • Figure 13 shows the in vivo efficacy study of the drug combination of the PD-1 inhibitor RB0004 described in the present application and different doses of Imiquimod (including 5% Imiquimod cream) - the inhibition rate of tumor weight in mice dissected after administration observation.
  • Figure 14 shows the trend of tumor proliferation in each group of mice during the administration period in Example 6 of the present application.
  • Figure 15 shows the survival curves of mice in each group during the administration period in Example 6 of the present application.
  • Figure 16 shows the average tumor mass of mice in each group during the administration period in Example 6 of the present application.
  • Figure 17 shows the trend of tumor proliferation in each group of mice during the administration period in Example 7 of the present application.
  • Figure 18 shows the survival curves of mice in each group during the administration period in Example 7 of the present application.
  • Figure 19 shows the average tumor mass of mice in each group during the administration period in Example 7 of the present application.
  • Figure 20 shows the trend of tumor proliferation in each group of mice during the administration period in Example 8 of the present application.
  • Figure 21 shows the survival curves of mice in each group during the administration period in Example 8 of the present application.
  • Figure 22 shows the average tumor mass of mice in each group during the administration period in Example 8 of the present application.
  • Figure 23 shows the trend of tumor proliferation in each group of mice during the administration period in Example 9 of the present application.
  • Figure 24 shows the survival curves of mice in each group during the administration period in Example 9 of the present application.
  • Figure 25 shows the average tumor mass of mice in each group during the administration period in Example 9 of the present application.
  • Figure 26 shows the trend of tumor proliferation in each group of mice during the administration period in Example 10.1 of the present application.
  • Figure 27 shows the survival curves of mice in each group during the administration period in Example 10.1 of the present application.
  • Figure 28 shows the average tumor mass of mice in each group during the administration period in Example 10.1 of the present application.
  • Figure 29 shows the trend of tumor proliferation in each group of mice during the administration period in Example 10.2 of the present application.
  • Figure 30 shows the survival curves of mice in each group during the administration period in Example 10.2 of the present application.
  • Figure 31 shows the average tumor mass of mice in each group during the administration period in Example 10.2 of the present application.
  • Figure 32 shows the trend of body weight changes of mice in each group during the administration period in Example 11 of the present application.
  • Figure 33 shows the trend of tumor proliferation in mice in each group during the administration period in Example 11 of the present application.
  • Figure 34 shows the average tumor mass of mice in each group during the administration period in Example 11 of the present application.
  • Figure 35 shows a schematic diagram of sequential administration in Example 12 of the present application.
  • Figure 36 shows the trend of tumor proliferation in each group of mice during the administration period in Example 12 of the present application.
  • Figure 37 shows the survival curves of mice in each group during the administration period in Example 12 of the present application.
  • Figure 38 shows the average tumor mass of mice in each group during the administration period in Example 12 of the present application.
  • PD-1 generally refers to programmed cell death protein 1, a 288 amino acid type I membrane protein first described in 1992 (Ishida et al., EMBO J., 11 (1992), 3887-3895).
  • PD-1 is a member of the expanded CD28/CTLA-4 family of T cell regulators and has two ligands, PD-L1 (B7-H1, CD274) and PD-L2 (B7-DC, CD273).
  • the protein's structure includes an extracellular IgV domain, followed by a transmembrane region and an intracellular tail.
  • the intracellular tail contains two phosphorylation sites located in the immunoreceptor tyrosine-based inhibitory motif and the immunoreceptor tyrosine-based switch motif, suggesting that PD-1 negatively regulates TCR signaling. This is consistent with the binding of SHP-1 and SHP-2 phosphatases to the cytoplasmic tail of PD-1 after ligand binding.
  • PD-1 is not expressed on naive T cells, it is upregulated following T cell receptor (TCR)-mediated activation and is observed on both activated and exhausted T cells (Agata et al., Int. Immunology 8 (1996), 765-772). These exhausted T cells have a dysfunctional phenotype and are unable to respond appropriately.
  • the PD-1 may include human PD-1 (hPD-1) or its variants, isotypes and species homologues, and analogs having at least one common epitope with hPD-1.
  • the amino acid sequence of an exemplary hPD-1 can be found under GenBank Accession No. U64863.
  • the term "PD-L1" generally refers to programmed cell death 1 ligand 1, also known as B7 homolog 1, B7-H1, cluster of differentiation 274, (3)274 or CD274, which is related to PD-1 binding downregulates T cell activation and cytokine secretion.
  • P-L1 includes any native PD-L1 of any vertebrate origin, including mammals, such as primates (e.g., humans and cynomolgus monkeys) and rodents (e.g., mice and rats) ).
  • the term encompasses "full length", unprocessed PD-L1 as well as any form of PD-L1 produced by cellular processing.
  • PD-L1 can exist as a transmembrane protein or as a soluble protein.
  • "PD-L1" includes intact PD-L1 and its fragments, and also includes functional variants, isoforms, species homologues, derivatives, analogs of PD-L1, and PD-L1 with at least one Epitope analogs.
  • the basic structure of PD-L1 includes four domains: extracellular Ig-like V-type domain and Ig-like C2-type domain, transmembrane domain and cytoplasmic domain.
  • Exemplary human PD-L1 amino acid sequences can be found under NCBI Accession No. NP_001254653 or UniProt Accession No. Q9NZQ7.
  • the term "inhibitor” generally refers to the ability to completely or partially prevent or reduce one or more specific biomolecules (for example, proteins (such as PD-1 or PD-L1), polypeptides, lipopolysaccharides, glycoproteins , ribonucleoprotein complexes, etc.) physiologically functional compounds/substances or compositions.
  • the reduction of the physiological function of one or more specific proteins may include the reduction of the activity of the protein itself (such as the ability to bind to other molecules, etc.) or the reduction of the existing amount of the protein itself.
  • Suitable inhibitor molecules may include antagonist antibodies or antibody fragments, fragments or derivatives of small molecules, peptides, antisense oligonucleotides, small organic molecules, and the like.
  • the inhibitor is capable of blocking activation of a cell signaling pathway.
  • the PD-1/PD-L1 inhibitor is an anti-PD-1/PD-L1 antibody or antigen-binding fragment thereof.
  • the terms "Pembrolizumab”, “Nivolumab”, “Pidilizumab”, “Tislelizumab”, “Camrelizumab (SHR-1210)", “Sintilimab”, “Toripalimab”, “MEDI0680”, “BGB-A317”, “ TSR-042”, “REGN2810”, “PF-06801591”, “Durvalumab”, “Atezolizumab”, “Avelumab”, “MDX-1105”, “YW243.55.S70”, “MDPL3280A”, “AMP-224" , "LY3300054”, “RB0004", “RB0005" are used according to their plain and common meanings as understood in the art.
  • TLR Toll-like receptor
  • TLR polypeptides share a characteristic structure that includes an extracellular domain with leucine-rich repeats, a transmembrane domain, and an intracellular domain involved in TLR signaling.
  • Toll-like receptor 7 and “TLR7” generally refer to a sequence that shares at least 70% with publicly available TLR7 sequences, such as GenBank accession number AAZ99026 for human TLR7 polypeptides, or GenBank accession number AAK62676 for murine TLR7 polypeptides. , 80%, 90%, 95%, 96%, 97%, 98%, 99% or more sequence identity nucleic acid or polypeptide.
  • Toll-like receptor 8 and “TLR8” generally refer to a sequence that shares at least 70% with publicly available TLR7 sequences, such as GenBank accession number AAZ95441 for human TLR8 polypeptides, or GenBank accession number AAK62677 for murine TLR8 polypeptides. , 80%, 90%, 95%, 96%, 97%, 98%, 99% or more sequence identity nucleic acid or polypeptide.
  • TLR9 or “Toll-like receptor 9” (also known as CD289, UNQ5798 or PRO19605) generally refers to activation by unmethylated cytosine-phosphate-guanine (CpG) dinucleotide Nucleotide-sensitive TLRs.
  • TLR9 include, but are not limited to, human TLR9, a 1032 amino acid long protein encoded by a 3922 nucleotide long mRNA transcript (NM_017442.3). The exemplified amino acid sequence of human TLR9 is represented by GenBank Accession No. NP_059138.1.
  • TLR9 includes TLR9 homologues from species other than humans, such as Macaca Fascicularis (cynomolgus monkeys) or Pantroglodytes (chimpanzees).
  • TLR9 includes proteins comprising mutations of full-length wild-type TLR9, such as point mutations, fragments, insertions, deletions, and splice variants.
  • TLR9 also encompasses post-translational modifications of the amino acid sequence of TLR9.
  • agonist generally refers to a molecule (ie, a modulator) that directly or indirectly modulates other molecules (eg, TLRs) and increases the activity, activation or function of other molecules.
  • Agonists may include proteins, nucleic acids, carbohydrates, organic molecules, small organic molecules (with or without organic moieties), or other molecules.
  • a modulator that enhances the gene transcription, biological activity or biochemical function of a protein is a substance that enhances the transcription of said protein or stimulates its biochemical properties or activity.
  • an agonist induces, stimulates, increases, activates, facilitates, enhances, or upregulates the activity of a receptor, such activity being referred to as "agonistic activity.”
  • a "TLR agonist” is a substance that directly or indirectly binds a TLR (eg TLR7 and/or TLR8) to induce TLR signaling. Any detectable difference in TLR signaling may indicate that an agonist stimulates or activates a TLR. Signaling differences can manifest, for example, as changes in the expression of target genes, changes in the phosphorylation of signaling components, changes in the intracellular localization of downstream elements such as nuclear factor- ⁇ B (NF- ⁇ B), certain components (such as changes in the association of IL-1 receptor-associated kinase (IRAK) with other proteins or intracellular structures, or changes in the biochemical activity of components such as kinases such as mitogen-activated protein kinase (MAPK).
  • TLR agonist refers to any compound that acts as an agonist of a TLR.
  • derivative generally refers to a chemical substance that is structurally related to another chemical substance, or that can be derived from another chemical substance (i.e., a chemical substance from which the chemical substance is derived).
  • Derivatives of organic molecules include, but are not limited to, modified molecules, eg, molecules modified by the addition or deletion of hydroxyl, methyl, ethyl, carboxyl, nitro or amino groups.
  • organic molecules can also be esterified, alkylated and/or phosphorylated.
  • the term "pharmaceutically acceptable derivatives” includes isomers, salts, esters, enol ethers, enol esters, acetals, ketals, orthoesters, hemiacetals, hemiketals, Acids, bases, solvates, hydrates or prodrugs thereof.
  • Such derivatives can be readily prepared by those skilled in the art using known methods for such derivatizations.
  • the term “isomer” generally refers to different compounds having the same molecular formula but differing in the arrangement and configuration of the atoms.
  • the term “isomer” includes, but is not limited to, optical isomers and analogs, structural isomers and analogs, conformational isomers and analogs, and the like.
  • analogue generally refers to its art-recognized meaning.
  • the term “analogue” generally refers to a second organic or inorganic molecule that has a similar or identical function to, and is structurally similar to, a first organic or inorganic molecule .
  • the term “analog” generally refers to a peptide/protein in which one or more amino acid residues have been replaced by other amino acid residues and/or in which one or more amino acid residues have been derived from the peptide/protein A modified peptide or protein that has been deleted from and/or in which one or more amino acid residues have been added to the peptide/protein.
  • the term "pharmaceutically acceptable salt” generally refers to a pharmaceutically acceptable organic or inorganic salt of a compound.
  • Exemplary salts include, but are not limited to, sulfate, citrate, acetate, oxalate, chloride, bromide, iodide, nitrate, bisulfate, phosphate, acid phosphate, isonicotinate , lactate, salicylate, acid citrate, tartrate, oleate, tannin, pantothenate, bitartrate, ascorbate, succinate, maleate, gentian salt, fumarate, gluconate, glucuronate, saccharate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonate, pamoate (i.e.
  • a pharmaceutically acceptable salt may involve the inclusion of another molecule such as acetate, succinate or other counterion.
  • the counterion can be any organic or inorganic moiety which stabilizes the charge on the parent compound.
  • pharmaceutically acceptable salts can have more than one charged atom in their structure. In instances where multiple charged atoms are part of a pharmaceutically acceptable salt, the salt can have multiple counterions. Thus, a pharmaceutically acceptable salt may have one or more charged atoms and/or one or more counterions.
  • antibody is generally used in the broadest sense and specifically covers monoclonal antibodies, polyclonal antibodies, dimers, multimers, multispecific antibodies (e.g., bispecific antibodies), and antibody fragments as long as they show the desired biological activity (Miller et al (2003) Jour. of Immunology 170:4854-4861).
  • Antibodies can be murine, human, humanized, chimeric, or derived from other species.
  • a full-length antibody typically refers to an antibody consisting of two “full-length antibody heavy chains” and two “full-length antibody light chains”.
  • a “full-length antibody heavy chain” is generally a polypeptide consisting, in the N-terminal to C-terminal direction, of an antibody heavy chain variable domain (VH), an antibody constant heavy chain domain 1 (CH1), an antibody hinge region (HR) , antibody heavy chain constant domain 2 (CH2), and antibody heavy chain constant domain 3 (CH3), abbreviated as VH-CH1-HR-CH2-CH3; and in the case of antibodies of the IgE subclass, optionally It also includes the antibody heavy chain constant domain 4 (CH4).
  • VH antibody heavy chain variable domain
  • CH1 antibody constant heavy chain domain 1
  • HR antibody hinge region
  • CH2 antibody heavy chain constant domain 2
  • CH3 antibody heavy chain constant domain 3
  • a "full-length antibody heavy chain” is a polypeptide consisting of VH, CH1, HR, CH2 and CH3 in an N-terminal to C-terminal direction.
  • a “full-length antibody light chain” is generally a polypeptide consisting of an antibody light chain variable domain (VL) and an antibody light chain constant domain (CL) in the N-terminal to C-terminal direction, abbreviated as VL-CL.
  • the antibody light chain constant domain (CL) may be kappa (kappa) or lambda (lambda).
  • the two full-length antibody chains are linked together by an inter-polypeptide disulfide bond between the CL domain and the CH1 domain and between the hinge region of the full-length antibody heavy chain.
  • Typical examples of full-length antibodies are natural antibodies such as IgG (eg, IgGl and IgG2), IgM, IgA, IgD, and IgE).
  • an antigen binding domain generally refers to a portion of an antibody molecule comprising the amino acids responsible for the specific binding between the antibody and the antigen.
  • the portion of an antigen that is specifically recognized and bound by an antibody is called an "epitope" as described above.
  • An antigen binding domain will typically comprise an antibody light chain variable region (VL) and an antibody heavy chain variable region (VH); however, it need not comprise both.
  • Fd fragments for example, have two VH regions and typically retain some antigen-binding function of the full antigen-binding domain.
  • antigen-binding fragments of antibodies include (1) Fab fragments, monovalent fragments having VL, VH, constant light chain (CL) and CH1 domains; (2) F(ab') 2 fragments, having two Bivalent fragment of two Fab fragments connected by sulfur bridge; (3) Fd fragment with two VH and CH1 domains; (4) Fv fragment with VL and VH domains of antibody single arm, (5) dAb fragment (Ward et al., "Binding Activities of a Repertoire of Single Immunoglobulin Variable Domains Secreted From Escherichia coli," Nature 341:544-546 (1989), which is hereby incorporated by reference in its entirety), which has a VH domain; (6) Isolated Complementarity Determining Regions (CDRs); (7) Single-chain Fv (scFv), for example derived from a scFv-library.
  • Fab fragments monovalent fragments having VL, VH, constant light chain (CL) and CH1 domains
  • the two domains VL and VH of the Fv fragment are encoded by separate genes, they can be joined using recombinant methods by a synthetic linker that allows it to be produced as a single protein in which the VL and VH regions pair to form a monovalent molecule chain (termed single-chain Fv (scFv)) (see, e.g., Huston et al., "Protein Engineering of Antibody Binding Sites: Recovery of Specific Activity in an Anti-Digoxin Single-Chain Fv Analogue Produced in Escherichia coli," Proc.
  • scFv single-chain Fv
  • VHH relates to variable antigens from heavy chain antibodies of the family Camelidae (camel, dromedary, llama, alpaca, etc.) Binding domain (see Nguyen VK et al., 2000, The EMBO Journal, 19, 921-930; Muyldermans S., 2001, J Biotechnol., 74, 277-302 and review Vanlandschoot P. et al., 2011, Antiviral Research 92 , 389-407). VHHs may also be referred to as Nanobodies (Nb) and/or Single Domain Antibodies. These antibody fragments are obtained using conventional techniques known to those skilled in the art, and the function of the fragments is evaluated in the same manner as intact antibodies.
  • variable region or “variable domain” generally refers to a region in which some segments of the variable domain may have large differences in sequence between antibodies.
  • a “variable region” in a light chain may comprise the light chain variable region VL; a “variable region” in a heavy chain may comprise the heavy chain variable region VH.
  • the variable domains mediate antigen binding and determine the specificity of a particular antibody for its particular antigen.
  • the variability is not evenly distributed throughout the variable domains. It is usually concentrated in three segments called hypervariable regions (CDRs or HVRs) in the light and heavy chain variable domains. The more highly conserved portions of variable domains are called the framework regions (FR).
  • variable domains of native heavy and light chains each comprise four FR regions, most adopting a ⁇ -sheet configuration, connected by three CDRs, which form a circular connection and in some cases form part of the ⁇ -sheet structure .
  • the CDRs in each chain are held in close proximity by the FR regions, and the CDRs from the other chain together contribute to the formation of the antibody's antigen-binding site (see Kabat et al, Sequences of Immunological Interest, Fifth Edition, National Institute of Health, Bethesda, Md. (1991)).
  • the terms "VH" and "VH domain” are used interchangeably to refer to the heavy chain variable region of an antibody or antigen-binding molecule thereof.
  • CDR generally refers to the complementarity determining regions within the variable sequences of antibodies.
  • CDR1, CDR2 and CDR3 are 3 CDRs in each variable region of the heavy and light chains, which are called CDR1, CDR2 and CDR3 for each variable region.
  • the precise boundaries of these CDRs have been defined differently according to different systems.
  • the system described by Kabat Kabat (Kabat et al., Sequences of Proteins of Immunological Interest (National Institutes of Health, Bethesda, Md. (1987) and (1991)) not only provides an unambiguous residue numbering system applicable to any variable region of an antibody , but also provides the precise residue limits that define these three CDRs.
  • These CDRs can be referred to as Kabat CDRs.
  • Padlan FASEB J.9:133 -139 (1995)
  • MacCallum JMoI Biol 262 (5): 732-45 (1996)
  • CDR bounds may not strictly follow one of the above-mentioned systems, but Still overlapping with the Kabat CDRs, although they could be shortened or lengthened according to the following predictions or experimental findings, specific residues or groups of residues or even the entire CDR did not significantly affect antigen binding.
  • CDR CDR
  • HCDR1 CDR1
  • HCDR2 CDR3
  • LCDR1 CDR2
  • LCDR3 CDR3
  • percent (%) sequence identity generally refers to the amino acids with which two or more aligned amino acid sequences are identical compared to the number of amino acid residues that make up the total length of these amino acid sequences The number of matches ("hits") for .
  • alignment is used, for two or more sequences, when the sequences are compared and aligned for maximum correspondence (as measured using sequence comparison algorithms known in the art), or when manually aligned and visually Upon inspection, the percentage of amino acid residues that are identical (eg, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% sequence identity) can be determined.
  • sequences compared to determine sequence identity can be distinguished by one or more amino acid substitutions, additions or deletions.
  • Suitable programs for aligning protein sequences are known to those skilled in the art.
  • the percent sequence identity of protein sequences can be determined, for example, with programs such as CLUSTALW, Clustal Omega, FASTA or BLAST, for example using the NCBI BLAST algorithm (AltschulSF et al. (1997), Nucleic Acids Res. 25:3389-3402) .
  • antibody analogue is generally used in the broadest sense and encompasses in particular molecules which specifically bind a target molecule with a monospecificity and which differ structurally from natural antibodies.
  • antibody analog refers to an antibody comprising a segment of substantial identity to a portion of the amino acid sequence and having at least one of the following properties : (1) specific binding to PD-1 or PD-L1 under appropriate binding conditions, (2) ability to inhibit at least one biological activity of PD-1 or PD-L1.
  • antibody analogs contain conservative amino acid substitutions (or insertions or deletions) relative to the native sequence.
  • Analogs are typically at least 20 or 25 amino acids in length, at least 50, 60, 70, 80, 90, 100, 150, or 200 amino acids in length or longer, and typically can be as long as a full-length heavy or light chain of an antibody .
  • Some examples include antibody analogs having 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or 17 substitutions compared to the germline amino acid sequence .
  • each active ingredient/therapeutic agent can be prepared as a separate formulation (solid, liquid, gel, etc.), and in some embodiments, each active ingredient/therapeutic agent can be present in a different container, It can also be formulated into a desired preparation with a suitable carrier simultaneously or separately when needed; in some embodiments, each active ingredient/therapeutic agent can be from different sources (such as prepared, produced or sold by different merchants); in some In an embodiment, each active ingredient/therapeutic agent may be mixed to form a pharmaceutical composition.
  • compositions generally refers to a preparation that is in a form that permits the biological activity of the active ingredient to be effective and that does not contain additional substances that are unacceptably toxic to the subject to which the composition is to be administered.
  • Such compositions may be sterile, and may contain a pharmaceutically acceptable carrier, such as physiological saline.
  • Suitable pharmaceutical compositions may comprise one or more buffers (e.g. acetate, phosphate or citrate buffers), surfactants (e.g. polysorbates), stabilizers (e.g. human albumin), preservatives, agents (such as benzyl alcohol), absorption enhancers for enhanced bioavailability, and/or other conventional solubilizing or dispersing agents.
  • buffers e.g. acetate, phosphate or citrate buffers
  • surfactants e.g. polysorbates
  • stabilizers e.g. human albumin
  • preservatives agents (such as benzyl alcohol), absorption enhancers for enhanced bioavailability,
  • the term "pharmaceutically acceptable carrier” generally refers to one or more non-toxic materials that do not interfere with the effectiveness of the biological activity of the active ingredient.
  • Such formulations may conventionally contain salts, buffers, preservatives, compatible carriers, and optionally other therapeutic agents.
  • Such pharmaceutically acceptable formulations may also contain compatible solid or liquid fillers, diluents or encapsulating substances suitable for human administration.
  • Other contemplated carriers, excipients, and/or additives that may be used in the formulations described herein may include, for example, flavoring agents, antimicrobial agents, sweeteners, antioxidants, antistatic agents, lipids , protein excipients (such as serum albumin, gelatin, casein), salt-forming counterions (such as sodium), etc.
  • Neoplastic generally refers to cells undergoing new and abnormal proliferation, especially a disease in which proliferation is uncontrolled and progresses, resulting in a neoplasm.
  • Neoplastic cells can be malignant, ie invasive and metastatic, or benign.
  • neoplastic generally refers to an abnormal mass of tissue in which the growth of the mass exceeds that of normal tissue and is not coordinated with the growth of normal tissue.
  • a “neoplastic” can be defined as “benign” or “malignant”, depending on the following characteristics: degree of cellular differentiation, including morphology and function, rate of growth, local invasion and metastasis.
  • benign neoplasms are generally well differentiated, have characteristically slower growth than malignant neoplasms, and remain localized to the site of origin. Furthermore, benign tumors do not have the ability to infiltrate, invade, or metastasize to distant sites.
  • Malignant neoplasms are usually poorly differentiated (regressive), with characteristic rapid growth with progressive infiltration, invasion, and destruction of surrounding tissue. In addition, malignant neoplasms have the ability to metastasize to distant sites.
  • tumor or cancer
  • cancer generally refers to any medical condition characterized by the growth, proliferation or metastasis of neoplastic or malignant cells, and the tumor may be a solid tumor or a non-solid tumor.
  • wart generally refers to a type of superficial benign skin neoplasms caused by human papilloma virus (HPV), mainly caused by cell proliferation.
  • HPV belongs to the A genus of Papovaviridae in the DNA virus.
  • the term "administer" and similar terms are generally not limited to bodily administration, suitable methods include in vitro, ex vivo or in vivo methods.
  • suitable methods include in vitro, ex vivo or in vivo methods.
  • any method of administration known to those skilled in the art for contacting cells, organs or tissues with the composition may be employed.
  • the compounds may be introduced into the body of a subject in need of treatment by any route of introduction or delivery.
  • the compositions of the present application may be administered orally, topically, intranasally, intramuscularly, subcutaneously, intradermally, intrathecally, intraperitoneally, transdermally, or intratumorally.
  • the term “effective amount” or “effective dose” generally refers to an amount sufficient to achieve, or at least partially achieve, the desired effect.
  • a “therapeutically effective amount” or “therapeutically effective dose” of a drug or therapeutic agent is typically one that, when used alone or in combination with another therapeutic agent, promotes regression of disease (by reducing the severity of disease symptoms, frequency of asymptomatic periods of disease), any amount of drug that is evidenced by an increase in the degree and duration of the disease, or by the prevention of impairment or disability due to the presence of a disease.
  • the term “treating” generally refers to slowing or ameliorating the progression, severity, and and/or duration, or amelioration of one or more symptoms (eg, one or more identifiable symptoms) of a proliferative disorder.
  • the term “treating” may also refer to the improvement of at least one measurable physical parameter of a proliferative disorder, such as tumor growth, not necessarily discernible by the patient.
  • the term “treating” in this application may also refer to inhibiting the progression of a proliferative disorder physically, eg, by stabilizing discernible symptoms, physiologically, eg, by stabilizing physical parameters, or both. In certain instances, the term “treating” may refer to reducing or stabilizing tumor size or cancer cell count.
  • the term “synergy” generally means that the combined effect of two or more active drugs is greater than the sum of each active drug alone.
  • the combination of two or more drugs results in “synergistic inhibition” of an activity or process, such as tumor growth, it means that the inhibition of that activity or process is greater than the sum of the inhibitory effects of the individual active drugs.
  • subject generally refers to human or non-human animals, including but not limited to cats, dogs, horses, pigs, cows, sheep, rabbits, mice, rats or monkeys.
  • the term "about” generally refers to a range of 0.5%-10% above or below the specified value, such as about 0.5%, about 1%, about 1.5%, about 2% above or below the specified value , about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, about 5%, about 5.5%, about 6%, about 6.5%, about 7%, about 7.5%, about 8%, about 8.5%, about 9%, about 9.5%, or about 10%.
  • the present application provides an agonist that induces the expression of INF and cytokines, activates the innate immune system response, and is used as a drug combination with an immune checkpoint inhibitor, including a TLR (eg, TLR7/TLR8) agonist and an immune checkpoint (eg, PD-1 or PD-L1) inhibitors, etc.
  • an immune checkpoint inhibitor including a TLR (eg, TLR7/TLR8) agonist and an immune checkpoint (eg, PD-1 or PD-L1) inhibitors, etc.
  • the present application provides a pharmaceutical combination, which may comprise a programmed cell death protein 1 (PD-1) inhibitor and/or a programmed death ligand 1 (PD-L1) inhibitor, and a TLR agonist.
  • a pharmaceutical combination which may comprise a programmed cell death protein 1 (PD-1) inhibitor and/or a programmed death ligand 1 (PD-L1) inhibitor, and a TLR agonist.
  • the TLR agonist comprises TLR1, TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9 and/or TLR10.
  • the TLR agonist is selected from one or more of TLR7 agonists, TLR8 agonists, and TLR9 agonists.
  • the TLR agonist comprises a dual agonist of TLR7 and TLR8 (TLR7/TLR8 agonist).
  • the TLR agonist comprises an imidazoquinoline derivative.
  • the TLR agonist is selected from the group consisting of Imiquimod, Gardiquimod, Resiquimod, 1V209, Selgantolimod (GS-9688), Vesatolimod ( GS-9620), Sumanirole, PF-4878691, and one or more of pharmaceutically acceptable derivatives thereof.
  • the TLR agonist includes imiquimod, resiquimod or a pharmaceutically acceptable salt thereof.
  • the TLR agonist is selected from LHC-165, NKTR-262, DN1508052-01, SHR2150, CL307, CL264, loxoribine, exartoribine, DSR-6434, GSK2245035, One or more of SM-276001, SM-324405, SM-324406, AZ12441970, AZ12443988 and pharmaceutically acceptable derivatives thereof.
  • the PD-1 inhibitor has one or more of the following characteristics:
  • PD-1 Inhibit or reduce the expression of PD-1, such as the transcription or translation of PD-1;
  • Inhibiting or reducing PD-1 activity such as inhibiting or reducing the binding of PD-1 to its cognate ligand, such as PD-L1 or PD-L2;
  • the PD-1 inhibitor comprises an anti-PD-1 antibody or an antigen-binding fragment thereof.
  • the pharmaceutical combination may comprise: 1) an anti-PD-1 antibody or antigen-binding fragment thereof; and 2) a TLR7 and/or TLR8 agonist.
  • the pharmaceutical combination may comprise: 1) an anti-PD-1 antibody or an antigen-binding fragment thereof; and 2) an imidazoquinoline derivative.
  • the anti-PD-1 antibody is selected from Pembrolizumab (Pembrolizumab), Nivolumab (Navolumab), Pidilizumab, Tislelizumab (Tislelizumab), SHR-1210 (Incyte /Jiangsu Hengrui Pharmaceutical Co., Ltd.), MEDI0680 (also known as AMP-514; Amplimmune Inc./Medimmune), BGB-A317 (BeiGene Ltd.), TSR-042 (also known as ANB011; AnaptysBio/Tesaro, Inc.) , REGN2810 (Regeneron Pharmaceuticals, Inc./Sanofi-Aventis), PF-06801591 (Pfizer), RB0004, analogs thereof, and combinations thereof.
  • the drug combination may include: 1) anti-PD-1 antibody, the anti-PD-1 antibody may be selected from Pembrolizumab (pembrolizumab), Nivolumab (nivolumab), Pidilizumab, Tislelizumab, SHR-1210 (Incyte/Jiangsu Hengrui Pharmaceutical Co., Ltd.), MEDI0680 (also known as AMP-514; Amplimmune Inc./Medimmune), BGB-A317 (BeiGene Ltd.), TSR-042 (also known as ANB011; AnaptysBio/Tesaro, Inc.
  • Pembrolizumab pembrolizumab
  • Nivolumab nivolumab
  • Pidilizumab Tislelizumab
  • SHR-1210 Incyte/Jiangsu Hengrui Pharmaceutical Co., Ltd.
  • MEDI0680 also known as AMP-514; Amplimmune Inc./Medimm
  • the anti-PD-1 antibody is RB0004.
  • RB0004 and other humanized anti-PD-1 monoclonal antibodies are disclosed in CN201610345750.1, WO2017201766A1.
  • the pharmaceutical combination comprises: 1) an anti-PD-1 antibody, the anti-PD-1 antibody is RB0004 or an analog thereof and combinations thereof; and 2) imidazoquinoline derivatives, the imidazoquinoline derivatives It can be Imiquimod or its derivatives and combinations thereof.
  • the antibody is PD-1 and Pembrolizumab (trade name KEYTRUDA, formerly known as Lambrolizumab (Lambrolizumab), also known as Merck 3745, MK-3475 or SCH-900475) is a combination with PD -1 binding humanized IgG4 monoclonal antibody.
  • Pembrolizumab is disclosed, for example, in Hamid, et al. (2013) New England Journal of Medicine 369(2):134-44, WO2009/114335 and US 8,354,509.
  • the pharmaceutical combination comprises: 1) an anti-PD-1 antibody, the anti-PD-1 antibody can be pembrolizumab or its biological enhancer, and its biological equivalent, and a combination thereof; and 2) imidazoquine Line derivatives, the imidazoquinoline derivatives can be Imiquimod or its derivatives and combinations thereof.
  • the anti-PD-1 antibody is Nivolumab (CAS registry number: 946414-94-4, alternative names include MDX-1106, MDX-1106-04, ONO-4538 or BMS-936558).
  • Nivolumab is a fully human IgG4 monoclonal antibody that specifically blocks PD-1.
  • Nivolumab (clone 5C4) and other human monoclonal antibodies that specifically bind PD-1 are disclosed in US8,008,449 and WO2006/121168.
  • the pharmaceutical combination comprises: 1) an anti-PD-1 antibody, the anti-PD-1 antibody can be Nivolumab or its analogs and combinations thereof; and 2) imidazoquinoline derivatives, the imidazoquinoline derivatives
  • the drug can be Imiquimod or its derivatives and combinations thereof.
  • the anti-PD-1 antibody is Pidilizumab.
  • Pidilizumab CT-011; Cure Tech
  • CT-011 Cure Tech
  • IgG1 monoclonal antibody that binds to PD-1.
  • Pidilizumab and other humanized anti-PD-1 monoclonal antibodies are disclosed in WO2009/101611.
  • Other anti-Rpd-1 antibodies are disclosed in US 8,609,089, US2010028330 and/or US20120114649.
  • Other anti-PD-1 antibodies include AMP514 (Amplimmune).
  • the pharmaceutical combination comprises: 1) an anti-PD-1 antibody, the anti-PD-1 antibody can be Pidilizumab or analogs and combinations thereof; and 2) imidazoquinoline derivatives, the imidazoquinoline derivatives It can be Imiquimod or its derivatives and combinations thereof.
  • the anti-PD-1 antibody comprises at least one CDR in the antibody heavy chain variable region (VH)
  • the VH comprises the amino acid sequence shown in SEQ ID NO:8.
  • the anti-PD-1 antibody comprises VH
  • the VH comprises HCDR3
  • the HCDR3 comprises the amino acid sequence shown in SEQ ID NO: 3 or the amino acid sequence shown in SEQ ID NO: 3 having at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, Amino acid sequences having about 98%, or about 99% sequence identity.
  • said VH further comprises HCDR2
  • said HCDR2 comprises or has at least about 70%, about 75% of the amino acid sequence set forth in SEQ ID NO:2 %, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% Amino acid sequences with sequence identity.
  • the VH further comprises HCDR1, wherein the HCDR1 comprises the amino acid sequence shown in SEQ ID NO: 1 or has at least about 70%, about 75% of the amino acid sequence shown in SEQ ID NO: 1 %, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% Amino acid sequences with sequence identity.
  • said VH comprises HCDR1, HCDR2 and HCDR3, wherein said HCDR3 comprises or has at least about 70% of the amino acid sequence set forth in SEQ ID NO:3 , about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or An amino acid sequence with about 99% sequence identity
  • the HCDR2 comprises the amino acid sequence shown in SEQ ID NO: 2 or has at least about 70%, about 75%, about 80%, Amino acid sequences having about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity
  • the HCDR1 comprises the amino acid sequence shown in SEQ ID NO: 1 or has at least about 70%, about 75%, about 80%, about 85%, about 90%, about Amino acid sequences having 91%, about 92%,
  • the C-terminal of the HFR1 is directly or indirectly linked to the N-terminal of the HCDR1, and the HFR1 comprises the amino acid sequence shown in SEQ ID NO: 4 or At least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about Amino acid sequences having 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the N-terminal of the HFR2 is directly or indirectly connected to the C-terminal of the HCDR1
  • the C-terminal of the HFR2 is directly or indirectly connected to the N-terminal of the HCDR2 Indirectly linked
  • the HFR2 comprises the amino acid sequence shown in SEQ ID NO:5 or has at least about 70%, about 75%, about 80%, about 85%, about 90% of the amino acid sequence shown in SEQ ID NO:5 Amino acid sequences having %, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the VH includes a framework region HFR3
  • the N-terminal of the HFR3 is directly or indirectly connected to the C-terminal of the HCDR2
  • the C-terminal of the HFR3 is directly or indirectly connected to the N-terminal of the HCDR3 Indirectly linked
  • the HFR3 comprises the amino acid sequence shown in SEQ ID NO:6 or has at least about 70%, about 75%, about 80%, about 85%, about 90% of the amino acid sequence shown in SEQ ID NO:6 Amino acid sequences having %, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the N-terminal of the HFR4 is directly or indirectly connected to the C-terminal of the HCDR3, and the HFR4 comprises the amino acid sequence shown in SEQ ID NO: 7 or At least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about Amino acid sequences having 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the C-terminus of HFR1 is directly or indirectly connected to the N-terminus of the HCDR1, and the N-terminus of the HFR2 is connected to the N-terminus of the HCDR1
  • the C-terminal of the HFR2 is directly or indirectly connected to the C-terminal of the HCDR2
  • the C-terminal of the HFR2 is directly or indirectly connected to the N-terminal of the HCDR2
  • the N-terminal of the HFR3 is directly or indirectly connected to the C-terminal of the HCDR2
  • the HFR3 The C-terminal is directly or indirectly connected to the N-terminal of the HCDR3, and the N-terminal of the HFR4 is directly or indirectly connected to the C-terminal of the HCDR3;
  • the HFR1 comprises the amino acid sequence shown in SEQ ID NO: 4 or with The amino acid sequence shown in SEQ ID NO: 4 has at least about 70%
  • said anti-PD-1 antibody comprises VH
  • said VH comprises the amino acid sequence shown in SEQ ID NO:8 or has at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about Amino acid sequences with 99% sequence identity.
  • said anti-PD-1 antibody comprises an antibody heavy chain (HC)
  • said HC comprises an amino acid sequence shown in SEQ ID NO: 9 or has at least the same amino acid sequence as shown in SEQ ID NO: 9 About 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98 %, or about 99% sequence identity of amino acid sequences.
  • the anti-PD-1 antibody comprises at least one CDR in the antibody light chain variable region (VL)
  • the VL comprises the amino acid sequence shown in SEQ ID NO:17.
  • said anti-PD-1 antibody comprises at least one CDR in VH
  • said VH comprises the amino acid sequence shown in SEQ ID NO: 8
  • said anti-PD-1 antibody comprises at least one CDR in VL At least one CDR
  • the VL comprises the amino acid sequence shown in SEQ ID NO:17.
  • the anti-PD-1 antibody comprises VL
  • the VL comprises LCDR1
  • the LCDR1 comprises the amino acid sequence shown in SEQ ID NO: 10 or the amino acid sequence shown in SEQ ID NO: 10 having at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, Amino acid sequences having about 98%, or about 99% sequence identity.
  • said VL further comprises LCDR2, wherein said LCDR2 comprises the amino acid sequence shown in SEQ ID NO: 11 or has at least about 70%, about 75% of the amino acid sequence shown in SEQ ID NO: 11 %, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% Amino acid sequences with sequence identity.
  • said VL further comprises LCDR3, wherein said LCDR3 comprises the amino acid sequence shown in SEQ ID NO: 12 or has at least about 70%, about 75% of the amino acid sequence shown in SEQ ID NO: 12 %, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% Amino acid sequences with sequence identity.
  • said VL comprises LCDR1, LCDR2 and LCDR3, wherein said LCDR1 comprises or has at least about 70% of the amino acid sequence set forth in SEQ ID NO:10 , about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or An amino acid sequence with about 99% sequence identity
  • said LCDR2 comprising the amino acid sequence shown in SEQ ID NO: 11 or having at least about 70%, about 75%, about 80%, Amino acid sequences having about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity
  • the LCDR3 comprises the amino acid sequence shown in SEQ ID NO: 12 or has at least about 70%, about 75%, about 80%, about 85%, about 90%, about 80% of the amino acid sequence shown in SEQ ID NO: 12 Amino acid
  • the VH comprises HCDR1, HCDR2 and HCDR3, wherein the HCDR3 comprises the amino acid sequence shown in SEQ ID NO: 3 or the same as SEQ ID
  • the amino acid sequence shown in NO:3 has at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, An amino acid sequence of about 96%, about 97%, about 98%, or about 99% sequence identity
  • the HCDR2 comprises the amino acid sequence shown in SEQ ID NO: 2 or has the amino acid sequence shown in SEQ ID NO: 2
  • the HCDR1 comprises the amino acid sequence shown in SEQ ID NO: 1 or has
  • the anti-PD-1 antibody can comprise VH and antibody VL
  • the VH can comprise HCDR1, HCDR2 and HCDR3, wherein the HCDR3 can comprise the amino acid sequence shown in SEQ ID NO: 3, and the HCDR2 can comprise SEQ ID NO: 3
  • the HCDR1 may comprise the amino acid sequence shown in SEQ ID NO:1
  • the VL may comprise LCDR1, LCDR2 and LCDR3, wherein the LCDR1 may comprise SEQ ID NO:10
  • the amino acid sequence shown, the LCDR2 can include the amino acid sequence shown in SEQ ID NO: 11, and the LCDR3 can include the amino acid sequence shown in SEQ ID NO: 12.
  • the drug combination may include: 1) anti-PD-1 antibody, the anti-PD-1 antibody may include VH and antibody VL, the VH may include HCDR1, HCDR2 and HCDR3, wherein the HCDR3 may include SEQ
  • the HCDR2 can comprise the amino acid sequence shown in SEQ ID NO:2
  • the HCDR1 can comprise the amino acid sequence shown in SEQ ID NO:1
  • the VL can comprise LCDR1, LCDR2 and LCDR3, wherein the LCDR1 can comprise the amino acid sequence shown in SEQ ID NO:10
  • the LCDR2 can comprise the amino acid sequence shown in SEQ ID NO:11
  • the LCDR3 can comprise the amino acid sequence shown in SEQ ID NO:12 Amino acid sequence
  • imidazoquinoline derivatives the imidazoquinoline derivatives can be Imiquimod or its derivatives and combinations thereof.
  • the C-terminus of the LFR1 is directly or indirectly connected to the N-terminus of the LCDR1, and the LFR1 comprises the amino acid sequence shown in SEQ ID NO: 13 or At least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about Amino acid sequences having 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the N-terminal of the LFR2 is directly or indirectly connected to the C-terminal of the LCDR1, and the C-terminal of the LFR2 is directly or indirectly connected to the N-terminal of the LCDR2.
  • the LFR2 comprises the amino acid sequence shown in SEQ ID NO: 14 or has at least about 70%, about 75%, about 80%, about 85%, about 90% of the amino acid sequence shown in SEQ ID NO: 14 Amino acid sequences having %, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the VL includes a framework region LFR3
  • the N-terminal of the LFR3 is directly or indirectly connected to the C-terminal of the LCDR2
  • the C-terminal of the LFR3 is directly or indirectly connected to the N-terminal of the LCDR3 Indirectly linked
  • the LFR3 comprises the amino acid sequence shown in SEQ ID NO: 15 or has at least about 70%, about 75%, about 80%, about 85%, about 90% of the amino acid sequence shown in SEQ ID NO: 15
  • Amino acid sequences having %, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the N-terminal of the LFR4 is directly or indirectly connected to the C-terminal of the LCDR3, and the LFR4 comprises the amino acid sequence shown in SEQ ID NO: 16 or At least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about Amino acid sequences having 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the VL includes framework regions LFR1, LFR2, LFR3 and LFR4, the C-terminus of the LFR1 is directly or indirectly connected to the N-terminus of the LCDR1, and the N-terminus of the LFR2 is connected to the LCDR1
  • the C-terminal of the LFR2 is directly or indirectly connected to the C-terminal of the LCDR2
  • the N-terminal of the LFR3 is directly or indirectly connected to the C-terminal of the LCDR2
  • the C-terminal of the LFR3 is directly or indirectly connected to the C-terminal of the LCDR2
  • the C-terminus is directly or indirectly connected to the N-terminus of the LCDR3, and the N-terminus of the LFR4 is directly or indirectly connected to the C-terminus of the LCDR3
  • the LFR1 comprises the amino acid sequence shown in SEQ ID NO: 13 or with The amino acid sequence shown in SEQ ID NO: 13 has at least about 70%, about 75%, about 80%, about 85%, about 90%, about 9
  • the VL comprises the amino acid sequence shown in SEQ ID NO: 17 or has at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about Amino acid sequences with 99% sequence identity.
  • said anti-PD-1 antibody comprises VH and VL
  • said VH comprises the amino acid sequence shown in SEQ ID NO:8 or has at least about 70 %, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, Or an amino acid sequence with about 99% sequence identity
  • the VL comprises the amino acid sequence shown in SEQ ID NO: 17 or has at least about 70%, about 75%, about 80% of the amino acid sequence shown in SEQ ID NO: 17 %, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity amino acid sequence.
  • the anti-PD-1 antibody may comprise VH and VL
  • the VH may comprise the amino acid sequence shown in SEQ ID NO:8
  • the VL may comprise the amino acid sequence shown in SEQ ID NO:17.
  • the drug combination may include: 1) anti-PD-1 antibody, wherein the anti-PD-1 antibody may include VH and VL, and the VH may include the amino acid sequence shown in SEQ ID NO: 8, and the Said VL may comprise the amino acid sequence shown in SEQ ID NO: 17; 2) imidazoquinoline derivatives, said imidazoquinoline derivatives may be Imiquimod or derivatives thereof and combinations thereof.
  • the anti-PD-1 antibody comprises an antibody light chain (LC)
  • the LC comprises an amino acid sequence shown in SEQ ID NO: 18 or has at least the same amino acid sequence as shown in SEQ ID NO: 18 About 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98 %, or about 99% sequence identity of amino acid sequences.
  • the anti-PD-1 antibody comprises HC and LC
  • the HC comprises or has at least about 70% of the amino acid sequence set forth in SEQ ID NO:9 , about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or An amino acid sequence with about 99% sequence identity
  • the LC comprises the amino acid sequence shown in SEQ ID NO: 18 or has at least about 70%, about 75%, about 80% with the amino acid sequence shown in SEQ ID NO: 18 Amino acids with about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity sequence.
  • the anti-PD-1 antibody may comprise HC and LC
  • the HC may comprise the amino acid sequence shown in SEQ ID NO:9
  • the LC may comprise the amino acid sequence shown in SEQ ID NO:18.
  • the drug combination may comprise: 1) anti-PD-1 antibody, wherein the anti-PD-1 antibody may comprise HC and LC, the HC may comprise the amino acid sequence shown in SEQ ID NO: 9, and the LC can comprise the aminoacid sequence shown in SEQ ID NO:18; 2) imidazoquinoline derivatives, described imidazoquinoline derivatives can be Imiquimod or its derivatives and their combination.
  • the PD-L1 inhibitor has one or more of the following characteristics:
  • PD-L1 Inhibit or reduce the expression of PD-L1, such as the transcription or translation of PD-L1;
  • Inhibiting or reducing PD-L1 activity such as inhibiting or reducing the binding of PD-L1 to its cognate receptor, such as PD-1;
  • the PD-L1 inhibitor comprises an anti-PD-L1 antibody or an antigen-binding fragment thereof.
  • the pharmaceutical combination may comprise: 1) an anti-PD-L1 antibody or antigen-binding fragment thereof; and 2) a TLR7 and/or TLR8 agonist.
  • the pharmaceutical combination may comprise: 1) an anti-PD-L1 antibody or an antigen-binding fragment thereof; and 2) an imidazoquinoline derivative.
  • the anti-PD-L1 antibody is selected from Durvalumab (MEDI4736, disclosed in US2013/0034559A1), Atezolizumab (MPDL3280A, disclosed in US8,217,149), Avelumab (MSB0010718C, disclosed in US2014/0341917A1), MDX-1105, YW243.55.S70, MDPL3280A, AMP-224 (Amplimmune, GlaxoSmithKline), LY3300054 (Eli Lilly and Co.), RB0005, analogs thereof, and combinations thereof.
  • Durvalumab MEDI4736, disclosed in US2013/0034559A1
  • Atezolizumab MPDL3280A, disclosed in US8,217,149
  • Avelumab MSB0010718C, disclosed in US2014/0341917A1
  • MDX-1105 YW243.55.S70
  • MDPL3280A AMP-224
  • LY3300054 Eli Lilly and Co.
  • the pharmaceutical combination may comprise: 1) anti-PD-L1 antibody, wherein the anti-PD-L1 antibody is selected from Durvalumab (MEDI4736, disclosed in US2013/0034559A1), Atezolizumab (MPDL3280A, disclosed in US8,217,149), Avelumab (MSB0010718C, disclosed in US2014/0341917A1), MDX-1105, YW243.55.S70, MDPL3280A, AMP-224, LY3300054, RB0005, their analogs and combinations thereof; and 2) imidazoquinoline derivatives, the imidazole
  • the quinoline derivatives may be selected from Imiquimod, Gardiquimod, Resiquimod, derivatives thereof and combinations thereof.
  • the anti-PD-L1 antibody is RB0005.
  • RB0005 and other humanized anti-PD-L1 monoclonal antibodies are disclosed in CN201610340678.3, WO2017197667A1.
  • the pharmaceutical combination comprises: 1) an anti-PD-L1 antibody, the anti-PD-1 antibody is RB0005 or analogs and combinations thereof; and 2) imidazoquinoline derivatives, the imidazoquinoline derivatives can be is Imiquimod or its derivatives and combinations thereof.
  • the PD-L1 inhibitor is MDX-1105.
  • MDX-1105 also known as BMS-936559, is an anti-PD-L1 antibody described in WO2007/005874.
  • the pharmaceutical combination comprises: 1) an anti-PD-L1 antibody, the anti-PD-1 antibody is MDX-1105 or its analogs and combinations thereof; and 2) imidazoquinoline derivatives, the imidazoquinoline The derivative may be Imiquimod or its derivatives and combinations thereof.
  • the PD-L1 inhibitor is YW243.55.S70.
  • the YW243.55.S70 antibody is an anti-PD-L1 antibody described in WO2010/077634.
  • the PD-L1 inhibitor is MDPL3280A (Genentech/Roche).
  • MDPL3280A is a human Fc-optimized IgG1 monoclonal antibody that binds to PD-L1.
  • Other human monoclonal antibodies to MDPL3280A and PD-L1 are disclosed in US Patent No. 7,943,743 and US Patent Publication No. 20120039906.
  • said anti-PD-L1 antibody comprises at least one CDR in VH
  • said VH comprises the amino acid sequence shown in SEQ ID NO:25.
  • the VH comprises HCDR3
  • the HCDR3 comprises the amino acid sequence shown in SEQ ID NO: 21 or the amino acid sequence shown in SEQ ID NO: 21 having at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, Amino acid sequences having about 98%, or about 99% sequence identity.
  • said VH further comprises HCDR2, wherein said HCDR2 comprises the amino acid sequence shown in SEQ ID NO: 20 or has at least about 70%, about 75% of the amino acid sequence shown in SEQ ID NO: 20 %, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% Amino acid sequences with sequence identity.
  • the VH further comprises HCDR1, wherein the HCDR1 comprises the amino acid sequence shown in SEQ ID NO: 19 or has at least about 70%, about 75% of the amino acid sequence shown in SEQ ID NO: 19 %, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% Amino acid sequences with sequence identity.
  • said VH comprises HCDR1, HCDR2 and HCDR3, wherein said HCDR3 comprises or has at least about 70% of the amino acid sequence set forth in SEQ ID NO:21 , about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or An amino acid sequence with about 99% sequence identity
  • the HCDR2 comprises the amino acid sequence shown in SEQ ID NO: 20 or has at least about 70%, about 75%, about 80%, Amino acid sequences having about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity
  • the HCDR1 comprises the amino acid sequence shown in SEQ ID NO: 19 or has at least about 70%, about 75%, about 80%, about 85%, about 90%, about Amino acid sequences having 91%, about 92%,
  • said VH can comprise HCDR1, HCDR2 and HCDR3, wherein said HCDR3 comprises the amino acid sequence shown in SEQ ID NO: 21, said HCDR2 comprises the amino acid sequence shown in SEQ ID NO: 20, and said HCDR1 comprises SEQ ID NO: Amino acid sequence shown in ID NO:19.
  • the C-terminal of the HFR1 is directly or indirectly connected to the N-terminal of the HCDR1, and the HFR1 comprises the amino acid sequence shown in SEQ ID NO: 22 or At least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about Amino acid sequences having 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the N-terminal of the HFR2 is directly or indirectly connected to the C-terminal of the HCDR1
  • the C-terminal of the HFR2 is directly or indirectly connected to the N-terminal of the HCDR2 Indirectly connected
  • the HFR2 comprises the amino acid sequence shown in SEQ ID NO:23 or has at least about 70%, about 75%, about 80%, about 85%, about 90% of the amino acid sequence shown in SEQ ID NO:23 Amino acid sequences having %, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the VH includes a framework region HFR3
  • the N-terminal of the HFR3 is directly or indirectly connected to the C-terminal of the HCDR2
  • the C-terminal of the HFR3 is directly or indirectly connected to the N-terminal of the HCDR3 Indirectly connected
  • the HFR3 comprises the amino acid sequence shown in SEQ ID NO:24 or has at least about 70%, about 75%, about 80%, about 85%, about 90% of the amino acid sequence shown in SEQ ID NO:24 Amino acid sequences having %, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the N-terminal of the HFR4 is directly or indirectly connected to the C-terminal of the HCDR3, and the HFR4 comprises the amino acid sequence shown in SEQ ID NO: 7 or
  • the amino acid sequence shown in SEQ ID NO: 7 has at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95% %, about 96%, about 97%, about 98%, or about 99% sequence identity of amino acid sequences.
  • the C-terminus of HFR1 is directly or indirectly connected to the N-terminus of the HCDR1, and the N-terminus of the HFR2 is connected to the N-terminus of the HCDR1
  • the C-terminal of the HFR2 is directly or indirectly connected to the C-terminal of the HCDR2
  • the C-terminal of the HFR2 is directly or indirectly connected to the N-terminal of the HCDR2
  • the N-terminal of the HFR3 is directly or indirectly connected to the C-terminal of the HCDR2
  • the HFR3 The C-terminal is directly or indirectly connected to the N-terminal of the HCDR3, and the N-terminal of the HFR4 is directly or indirectly connected to the C-terminal of the HCDR3;
  • the HFR1 comprises the amino acid sequence shown in SEQ ID NO: 22 or with The amino acid sequence shown in SEQ ID NO: 22 has at least about 70%
  • the VH comprises the amino acid sequence shown in SEQ ID NO: 25 or has at least about 70% of the amino acid sequence shown in SEQ ID NO: 25, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about Amino acid sequences with 99% sequence identity.
  • said anti-PD-L1 antibody comprises HC
  • said HC comprises the amino acid sequence shown in SEQ ID NO:26 or has at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% Amino acid sequences for % sequence identity.
  • the VL comprises the amino acid sequence shown in SEQ ID NO:37.
  • said anti-PD-L1 antibody comprises at least one CDR in VH
  • said VH comprises the amino acid sequence shown in SEQ ID NO: 25
  • said anti-PD-L1 antibody comprises at least one CDR in VL At least one CDR
  • the VL comprises the amino acid sequence shown in SEQ ID NO:37.
  • said anti-PD-L1 antibody comprises at least one CDR in VH
  • said VH comprises the amino acid sequence shown in SEQ ID NO: 25
  • said anti-PD-L1 antibody comprises at least one CDR in VL At least one CDR
  • the VL comprises the amino acid sequence shown in SEQ ID NO:38, SEQ ID NO:39 or SEQ ID NO:40.
  • the anti-PD-L1 antibody comprises VL
  • the VL comprises LCDR1
  • the LCDR1 comprises the amino acid sequence shown in SEQ ID NO:27 or the amino acid sequence shown in SEQ ID NO:27 having at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, Amino acid sequences having about 98%, or about 99% sequence identity.
  • the anti-PD-L1 antibody comprises VL
  • the VL comprises LCDR1
  • the LCDR1 comprises the amino acid sequence shown in SEQ ID NO:28, SEQ ID NO:29 or SEQ ID NO:30 or at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, Amino acid sequences having about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • said VL further comprises LCDR2, wherein said LCDR2 comprises the amino acid sequence shown in SEQ ID NO: 31 or has at least about 70%, about 75% of the amino acid sequence shown in SEQ ID NO: 31 %, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% Amino acid sequences with sequence identity.
  • said VL further comprises LCDR3, wherein said LCDR3 comprises the amino acid sequence shown in SEQ ID NO:32 or has at least about 70%, about 75% of the amino acid sequence shown in SEQ ID NO:32 %, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% Amino acid sequences with sequence identity.
  • said VL comprises LCDR1, LCDR2 and LCDR3, wherein said LCDR1 comprises or has at least about 70% of the amino acid sequence set forth in SEQ ID NO:27 , about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or An amino acid sequence with about 99% sequence identity
  • said LCDR2 comprising the amino acid sequence shown in SEQ ID NO: 31 or having at least about 70%, about 75%, about 80%, Amino acid sequences having about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity
  • the LCDR3 comprises the amino acid sequence shown in SEQ ID NO:32 or has at least about 70%, about 75%, about 80%, about 85%, about 90%, about Amino acid sequences having 91%, about 92%, about 93%, about
  • said VL comprises LCDR1, LCDR2 and LCDR3, wherein said LCDR1 comprises the amino acid sequence shown in SEQ ID NO:28 or has at least about 70% of the amino acid sequence shown in SEQ ID NO:28 , about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or An amino acid sequence with about 99% sequence identity, said LCDR2 comprising the amino acid sequence shown in SEQ ID NO: 31 or having at least about 70%, about 75%, about 80%, Amino acid sequences having about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity , the LCDR3 comprises the amino acid sequence shown in SEQ ID NO:32 or has at least about 70%, about 75%, about 80%, about 85%, about 90%, about Amino acid sequences having 9
  • said VL comprises LCDR1, LCDR2 and LCDR3, wherein said LCDR1 comprises or has at least about 70% of the amino acid sequence set forth in SEQ ID NO:29 , about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or An amino acid sequence with about 99% sequence identity
  • said LCDR2 comprising the amino acid sequence shown in SEQ ID NO: 31 or having at least about 70%, about 75%, about 80%, Amino acid sequences having about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity
  • the LCDR3 comprises the amino acid sequence shown in SEQ ID NO:32 or has at least about 70%, about 75%, about 80%, about 85%, about 90%, about Amino acid sequences with 91%, about 92%, about 93%, about
  • said VL comprises LCDR1, LCDR2 and LCDR3, wherein said LCDR1 comprises the amino acid sequence shown in SEQ ID NO:30 or has at least about 70% of the amino acid sequence shown in SEQ ID NO:30 , about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or An amino acid sequence with about 99% sequence identity, said LCDR2 comprising the amino acid sequence shown in SEQ ID NO: 31 or having at least about 70%, about 75%, about 80%, Amino acid sequences having about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity , the LCDR3 comprises the amino acid sequence shown in SEQ ID NO:32 or has at least about 70%, about 75%, about 80%, about 85%, about 90%, about Amino acid sequences having 9
  • the VH comprises HCDR1, HCDR2 and HCDR3, wherein the HCDR3 comprises the amino acid sequence shown in SEQ ID NO: 21 or the same as SEQ ID
  • the amino acid sequence shown in NO:21 has at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, An amino acid sequence of about 96%, about 97%, about 98%, or about 99% sequence identity
  • the HCDR2 comprises the amino acid sequence shown in SEQ ID NO:20 or has the amino acid sequence shown in SEQ ID NO:20 At least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about An amino acid sequence of 98%, or about 99% sequence identity
  • said HCDR1 comprises the amino acid sequence shown in SEQ ID NO: 19
  • the anti-PD-L1 antibody may comprise VH and antibody VL
  • the VH may comprise HCDR1, HCDR2 and HCDR3, wherein the HCDR3 may comprise the amino acid sequence shown in SEQ ID NO: 21, and the HCDR2 may comprise The amino acid sequence shown in SEQ ID NO:20
  • the HCDR1 can comprise the amino acid sequence shown in SEQ ID NO:19
  • the VL can comprise LCDR1, LCDR2 and LCDR3, wherein the LCDR1 can comprise SEQ ID NO:27
  • the LCDR2 may comprise the amino acid sequence shown in SEQ ID NO:31
  • the LCDR3 may comprise the amino acid sequence shown in SEQ ID NO:32.
  • the drug combination may include: 1) anti-PD-L1 antibody, wherein the anti-PD-L1 antibody may include VH and antibody VL, and the VH may include HCDR1, HCDR2 and HCDR3, wherein the HCDR3 may include
  • the amino acid sequence shown in SEQ ID NO:21 the HCDR2 can include the amino acid sequence shown in SEQ ID NO:20
  • the HCDR1 can include the amino acid sequence shown in SEQ ID NO:19
  • the VL can include LCDR1 , LCDR2 and LCDR3, wherein the LCDR1 can comprise the amino acid sequence shown in SEQ ID NO:27
  • the LCDR2 can comprise the amino acid sequence shown in SEQ ID NO:31
  • the LCDR3 can comprise the amino acid sequence shown in SEQ ID NO:32 2) imidazoquinoline derivatives, the imidazoquinoline derivatives can be Imiquimod or its derivatives and combinations thereof.
  • the VH comprises HCDR1, HCDR2 and HCDR3, wherein the HCDR3 comprises the amino acid sequence shown in SEQ ID NO: 21 or the same as SEQ ID
  • the amino acid sequence shown in NO:21 has at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, An amino acid sequence of about 96%, about 97%, about 98%, or about 99% sequence identity
  • the HCDR2 comprises the amino acid sequence shown in SEQ ID NO:20 or has the amino acid sequence shown in SEQ ID NO:20 At least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about An amino acid sequence of 98%, or about 99% sequence identity
  • said HCDR1 comprises the amino acid sequence shown in SEQ ID NO: 19
  • the anti-PD-L1 antibody may comprise VH and antibody VL
  • the VH may comprise HCDR1, HCDR2 and HCDR3, wherein the HCDR3 may comprise the amino acid sequence shown in SEQ ID NO: 21, and the HCDR2 may comprise The amino acid sequence shown in SEQ ID NO:20
  • the HCDR1 may comprise the amino acid sequence shown in SEQ ID NO:19
  • the VL comprises LCDR1, LCDR2 and LCDR3, wherein the LCDR1 may comprise SEQ ID NO:28
  • the LCDR2 can include the amino acid sequence shown in SEQ ID NO:31
  • the LCDR3 can include the amino acid sequence shown in SEQ ID NO:32.
  • the drug combination may comprise: 1) anti-PD-L1 antibody, wherein the anti-PD-L1 antibody comprises VH and antibody VL, the VH may comprise HCDR1, HCDR2 and HCDR3, wherein the HCDR3 may comprise SEQ
  • the HCDR2 can comprise the amino acid sequence shown in SEQ ID NO:20
  • the HCDR1 can comprise the amino acid sequence shown in SEQ ID NO:19
  • the VL can comprise LCDR1, LCDR2 and LCDR3, wherein said LCDR1 can comprise the amino acid sequence shown in SEQ ID NO:28, SEQ ID NO:29 or SEQ ID NO:30
  • said LCDR2 can comprise the amino acid sequence shown in SEQ ID NO:31
  • Said LCDR3 can comprise the amino acid sequence shown in SEQ ID NO:32
  • imidazoquinoline derivatives said imidazoquinoline derivatives can be Imiquimod or derivatives thereof and combinations thereof.
  • the C-terminus of the LFR1 is directly or indirectly connected to the N-terminus of the LCDR1, and the LFR1 comprises the amino acid sequence shown in SEQ ID NO: 33 or At least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about Amino acid sequences having 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the N-terminal of the LFR2 is directly or indirectly connected to the C-terminal of the LCDR1, and the C-terminal of the LFR2 is directly or indirectly connected to the N-terminal of the LCDR2.
  • the LFR2 comprises the amino acid sequence shown in SEQ ID NO:34 or has at least about 70%, about 75%, about 80%, about 85%, about 90% of the amino acid sequence shown in SEQ ID NO:34 Amino acid sequences having %, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the VL includes a framework region LFR3
  • the N-terminal of the LFR3 is directly or indirectly connected to the C-terminal of the LCDR2
  • the C-terminal of the LFR3 is directly or indirectly connected to the N-terminal of the LCDR3 Indirectly linked
  • the LFR3 comprises the amino acid sequence shown in SEQ ID NO:35 or has at least about 70%, about 75%, about 80%, about 85%, about 90% of the amino acid sequence shown in SEQ ID NO:35 Amino acid sequences having %, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the N-terminal of the LFR4 is directly or indirectly connected to the C-terminal of the LCDR3, and the LFR4 comprises the amino acid sequence shown in SEQ ID NO: 36 or At least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about Amino acid sequences having 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the VL includes framework regions LFR1, LFR2, LFR3 and LFR4, the C-terminus of the LFR1 is directly or indirectly connected to the N-terminus of the LCDR1, and the N-terminus of the LFR2 is connected to the LCDR1
  • the C-terminal of the LFR2 is directly or indirectly connected to the C-terminal of the LCDR2
  • the N-terminal of the LFR3 is directly or indirectly connected to the C-terminal of the LCDR2
  • the C-terminal of the LFR3 is directly or indirectly connected to the C-terminal of the LCDR2
  • the C-terminus is directly or indirectly connected to the N-terminus of the LCDR3, and the N-terminus of the LFR4 is directly or indirectly connected to the C-terminus of the LCDR3
  • the LFR1 comprises the amino acid sequence shown in SEQ ID NO: 33 or with The amino acid sequence shown in SEQ ID NO: 33 has at least about 70%, about 75%, about 80%, about 85%, about 90%, about 9
  • the VL comprises the amino acid sequence shown in SEQ ID NO: 37 or has at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about Amino acid sequences with 99% sequence identity.
  • the VL comprises the amino acid sequence shown in SEQ ID NO:38, SEQ ID NO:39 or SEQ ID NO:40 or with SEQ ID NO: 38.
  • the amino acid sequence shown in SEQ ID NO:39 or SEQ ID NO:40 has at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93% Amino acid sequences having %, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • said VH comprises the amino acid sequence shown in SEQ ID NO:25 or has at least about 70% of the amino acid sequence shown in SEQ ID NO:25 %, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, Or an amino acid sequence with about 99% sequence identity
  • the VL comprises the amino acid sequence shown in SEQ ID NO: 37 or has at least about 70%, about 75%, about 80% of the amino acid sequence shown in SEQ ID NO: 37 %, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity amino acid sequence.
  • the anti-PD-L1 antibody may comprise VH and VL
  • the VH may comprise the amino acid sequence shown in SEQ ID NO:25
  • the VL may comprise the amino acid sequence shown in SEQ ID NO:37.
  • the drug combination may include: 1) anti-PD-L1 antibody, wherein the anti-PD-L1 antibody may include VH and VL, and the VH may include the amino acid sequence shown in SEQ ID NO: 25, and the Said VL can comprise the amino acid sequence shown in SEQ ID NO:37; 2) imidazoquinoline derivatives, said imidazoquinoline derivatives can be Imiquimod or derivatives thereof and combinations thereof.
  • said VH comprises the amino acid sequence shown in SEQ ID NO:25 or has at least about 70% of the amino acid sequence shown in SEQ ID NO:25 %, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, Or an amino acid sequence with about 99% sequence identity
  • the VL comprises the amino acid sequence shown in SEQ ID NO:38, SEQ ID NO:39 or SEQ ID NO:40 or with SEQ ID NO:38, SEQ ID NO: 39 or the amino acid sequence shown in SEQ ID NO: 40 has at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, Amino acid sequences having about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity.
  • the anti-PD-L1 antibody may comprise VH and VL
  • the VH may comprise the amino acid sequence shown in SEQ ID NO:25
  • the VL may comprise SEQ ID NO:38, SEQ ID NO:39 or Amino acid sequence shown in SEQ ID NO:40.
  • the drug combination may include: 1) anti-PD-L1 antibody, wherein the anti-PD-L1 antibody may include VH and VL, and the VH may include the amino acid sequence shown in SEQ ID NO: 25, and the Said VL can comprise the amino acid sequence shown in SEQ ID NO:38, SEQ ID NO:39 or SEQ ID NO:40; 2) imidazoquinoline derivatives, said imidazoquinoline derivatives can be Imiquimod or its derivatives and their combination.
  • the LC comprises the amino acid sequence shown in SEQ ID NO:41 or has at least about 70%, about 70% of the amino acid sequence shown in SEQ ID NO:41 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% Amino acid sequences for % sequence identity.
  • said LC comprises the amino acid sequence shown in SEQ ID NO:42, SEQ ID NO:43 or SEQ ID NO:44 or with SEQ ID NO:42 , SEQ ID NO:43 or the amino acid sequence shown in SEQ ID NO:44 has at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93% , about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity of amino acid sequences.
  • the anti-PD-L1 antibody comprises HC and LC
  • the HC comprises or has at least about 70% of the amino acid sequence set forth in SEQ ID NO:26 , about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or An amino acid sequence with about 99% sequence identity
  • the LC comprises the amino acid sequence shown in SEQ ID NO: 41 or has at least about 70%, about 75%, about 80%, Amino acid sequences having about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity .
  • the anti-PD-L1 antibody may comprise HC and LC
  • the HC may comprise the amino acid sequence shown in SEQ ID NO:26
  • the LC may comprise the amino acid sequence shown in SEQ ID NO:41.
  • the drug combination may comprise: 1) anti-PD-L1 antibody, wherein the anti-PD-L1 antibody may comprise HC and LC, the HC may comprise the amino acid sequence shown in SEQ ID NO: 26, and the LC can comprise the aminoacid sequence shown in SEQ ID NO:41; 2) imidazoquinoline derivatives, and described imidazoquinoline derivatives can be Imiquimod or derivatives thereof and combinations thereof.
  • the anti-PD-L1 antibody comprises HC and LC
  • the HC comprises or has at least about 70% of the amino acid sequence set forth in SEQ ID NO:26 , about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or An amino acid sequence with about 99% sequence identity
  • the LC comprises the amino acid sequence shown in SEQ ID NO: 42, SEQ ID NO: 43 or SEQ ID NO: 44 or is the same as SEQ ID NO: 42, SEQ ID NO: 43 or
  • the amino acid sequence shown in SEQ ID NO: 44 has at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95% %, about 96%, about 97%, about 98%, or about 99% sequence identity of amino acid sequences.
  • the anti-PD-L1 antibody can comprise HC and LC
  • the HC can comprise the amino acid sequence shown in SEQ ID NO:26
  • the LC can comprise SEQ ID NO:42, SEQ ID NO:43 or SEQ ID NO:43 Amino acid sequence shown in ID NO:44.
  • the drug combination may comprise: 1) anti-PD-L1 antibody, wherein the anti-PD-L1 antibody may comprise HC and LC, the HC may comprise the amino acid sequence shown in SEQ ID NO: 26, and the LC can comprise the aminoacid sequence shown in SEQ ID NO:42, SEQ ID NO:43 or SEQ ID NO:44; 2) imidazoquinoline derivatives, described imidazoquinoline derivatives can be Imiquimod or its derivatives and their combination.
  • the PD-1 inhibitor and/or PD-L1 inhibitor, and ii) the TLR agonist in the drug combination are not mixed with each other in the drug combination, i.e. i) said PD-1 inhibitor and/or ii) PD-L1 inhibitor and TLR agonist are in separate dosage forms.
  • the PD-1 inhibitor and/or PD-L1 inhibitor and ii) the TLR agonist in the pharmaceutical combination are in a single dosage form.
  • the pharmaceutical combination is formulated into a pharmaceutical composition (eg, a compound preparation).
  • the pharmaceutical composition can be injected directly into a large tumor without affecting normal (surrounding) tissue, enabling the killing of cancer cells, enabling the arrest or delay of the growth of a malignant mass (e.g., making the mass smaller or shrinking the tumor), And enabling terminal cancer patients to live with tumors (in a similar way to human patients living with parasites).
  • the drug combination When the drug combination is injected into the tumor, the drug can travel along blood vessels or lymphatic vessels to the metastases, and it will kill the metastatic cells. Injection of the drug combination into the tumor results in little trauma to the patient and can be repeated several times, eg monthly. Direct injections can also be given to both the primary tumor and secondary tumors to which the cancer has metastasized.
  • the pharmaceutical composition includes a PD-1 inhibitor or PD-L1 inhibitor, and a TLR agonist.
  • the pharmaceutical composition can include a PD-1 inhibitor and a TLR agonist.
  • the pharmaceutical composition may include an anti-PD-1 antibody or an antigen-binding fragment thereof and an imidazoquinoline derivative.
  • the pharmaceutical composition can include a PD-L1 inhibitor and a TLR agonist.
  • the pharmaceutical composition may include an anti-PD-L1 antibody or an antigen-binding fragment thereof and an imidazoquinoline derivative.
  • the TLR agonist is present in an amount from about 0.0001 mg/kg to about 200 mg/kg.
  • the TLR agonist can be present in the combinations described herein relative to the subject's body weight (ie, mg/kg).
  • the TLR agonist is present in an amount equal to about 0.0001 mg/kg to about 200 mg/kg, 0.001 mg/kg to about 200 mg/kg, 0.01 mg/kg to about 200 mg/kg, 0.01 mg/kg to about 150 mg /kg, 0 01 mg/kg to about 100 mg/kg, 0.01 mg/kg to about 50 mg/kg, 0.01 mg/kg to about 25 mg/kg, 0.01 mg/kg to about 10 mg/kg, or 0.01 mg/kg to about 5 mg /kg, 0.05mg/kg to about 200mg/kg, 0.05mg/kg to about 150mg/kg, 0.05mg/kg to about 100mg/kg, 0.05mg/kg to about 50mg/kg, 0.05mg/kg to about 25mg /kg, 0.05mg/kg to about 10mg/kg, or 0.05mg/kg to about 5mg/kg, 0.5mg/kg to about 200mg/kg, 0.5mg/kg to about 150m
  • the TLR agonist is present in an amount equal to about 1 mg/kg to about 200 mg/kg, 1 mg/kg to about 150 mg/kg, 1 mg/kg to about 100 mg/kg, 1 mg/kg to about 50 mg/kg, 1 mg /kg to about 25 mg/kg, 1 mg/kg to about 10 mg/kg, or 1 mg/kg to about 5 mg/kg.
  • the TLR agonist can be about 1 mg, 5 mg, 10 mg, 15 mg, 20 mg, 25 mg, 30 mg, 40 mg, 50 mg, 60 mg, 70 mg, 75 mg, 80 mg, 90 mg, 100 mg, 150 mg, 200 mg, 250 mg, 300 mg, 400 mg, 500 mg, Amounts of 600 mg, 700 mg, 800 mg, 900 mg, 1000 mg, 1100 mg, 1200 mg, 1300 mg, 1400 mg, 1500 mg, 1600 mg, 1700 mg, 1800 mg, 1900 mg or 2000 mg are present in combination.
  • the TLR agonist can be from about 1 mg to about 10 mg, 10 mg to about 20 mg, 25 mg to about 50 mg, 30 mg to about 60 mg, 40 mg to about 50 mg, 50 mg to about 100 mg, 75 mg to about 150 mg, 100 mg to about 200 mg, 200 mg to about 500 mg, Amounts from 500 mg to about 1000 mg, 1000 mg to about 1200 mg, 1000 mg to about 1500 mg, 1200 mg to about 1500 mg, or 1500 mg to about 2000 mg are present in combination.
  • the TLR agonist can be about 0.1 mg/mL, 0.5 mg/mL, 1 mg/mL, 2 mg/mL, 3 mg/mL, 4 mg/mL, 5 mg/mL, 6 mg/mL, 7 mg/mL, 8 mg/mL, 9mg/mL, 10mg/mL, 15mg/mL, 20mg/mL, 25mg/mL, 30mg/mL, 40mg/mL, 50mg/mL, 60mg/mL, 70mg/mL, 80mg/mL, 90mg/mL, 100mg/mL Amounts of mL, 150 mg/mL, 200 mg/mL, 250 mg/mL, 300 mg/mL, 400 mg/mL or 500 mg/mL are present in combination.
  • the TLR agonist is administered at about 1 mg/mL to about 10 mg/mL, 5 mg/mL to about 10 mg/mL, 5 mg/mL to about 15 mg/mL, 10 mg/mL to about 25 mg/mL, 20 mg/mL An amount of up to about 30 mg/mL, 25 mg/mL to about 50 mg/mL, or 50 mg/mL to about 100 mg/mL is present in combination.
  • the PD-1 inhibitor or PD-L1 inhibitor eg, anti-PD-1/PD-L1 antibody
  • the PD-1 inhibitor or PD-L1 inhibitor can be present in the combinations described herein relative to the subject's body weight (ie, mg/kg).
  • the PD-1 inhibitor or PD-L1 inhibitor is present in an amount equal to about 0.0001 mg/kg to about 200 mg/kg, 0.001 mg/kg to about 200 mg/kg, 0.01 mg/kg to about 200 mg/kg , 0.01 mg/kg to about 150 mg/kg, 0 01 mg/kg to about 100 mg/kg, 0.01 mg/kg to about 50 mg/kg, 0.01 mg/kg to about 25 mg/kg, 0.01 mg/kg to about 10 mg/kg or 0.01 mg/kg to about 5 mg/kg, 0.05 mg/kg to about 200 mg/kg, 0.05 mg/kg to about 150 mg/kg, 0.05 mg/kg to about 100 mg/kg, 0.05 mg/kg to about 50 mg/kg , 0.05 mg/kg to about 25 mg/kg, 0.05 mg/kg to about 10 mg/kg, or 0.05 mg/kg to about 5 mg/kg, 0.5 mg/kg to about 200 mg/kg, 0.5 mg/kg to about 150 mg/kg , 0.5 mg/kg to about 100 mg/kg
  • the PD-1 inhibitor or PD-L1 inhibitor is present in an amount equal to about 1 mg/kg to about 200 mg/kg, 1 mg/kg to about 150 mg/kg, 1 mg/kg to about 100 mg/kg, 1 mg/kg kg to about 50 mg/kg, 1 mg/kg to about 25 mg/kg, 1 mg/kg to about 10 mg/kg, or 1 mg/kg to about 5 mg/kg.
  • the anti-PD-1/PD-L1 antibody can be about 1 mg, 5 mg, 10 mg, 15 mg, 20 mg, 25 mg, 30 mg, 40 mg, 50 mg, 60 mg, 70 mg, 75 mg, 80 mg, 90 mg, 100 mg, 150 mg, 200 mg, 250 mg, Amounts of 300 mg, 400 mg, 500 mg, 600 mg, 700 mg, 800 mg, 900 mg, 1000 mg, 1100 mg, 1200 mg, 1300 mg, 1400 mg, 1500 mg, 1600 mg, 1700 mg, 1800 mg, 1900 mg or 2000 mg are present in combination.
  • the anti-PD-1/PD-L1 antibody can be about 1 mg to about 10 mg, 10 mg to about 20 mg, 25 mg to about 50 mg, 30 mg to about 60 mg, 40 mg to about 50 mg, 50 mg to about 100 mg, 75 mg to about 150 mg, 100 mg to about 200 mg , 200 mg to about 500 mg, 500 mg to about 1000 mg, 1000 mg to about 1200 mg, 1000 mg to about 1500 mg, 1200 mg to about 1500 mg, or 1500 mg to about 2000 mg are present in combination.
  • the anti-PD-1/PD-L1 antibody can be about 0.1mg/mL, 0.5mg/mL, 1mg/mL, 2mg/mL, 3mg/mL, 4mg/mL, 5mg/ml, 6mg/mL, 7mg/mL mL, 8mg/mL, 9mg/mL, 10mg/mL, 15mg/mL, 20mg/mL, 25mg/mL, 30mg/mL, 40mg/mL, 50mg/mL, 60mg/mL, 70mg/mL, 80mg/mL, Amounts of 90 mg/mL, 100 mg/mL, 150 mg/mL, 200 mg/mL, 250 mg/mL, 300 mg/mL, 400 mg/mL or 500 mg/mL are present in combination.
  • the anti-PD-1/PD-L1 antibody is administered at about 1 mg/mL to about 10 mg/mL, 5 mg/mL to about 10 mg/mL, 5 mg/mL to about 15 mg/mL, 10 mg/mL to about 25 mg /mL, 20 mg/mL to about 30 mg/mL, 25 mg/mL to about 50 mg/mL, or 50 mg/mL to about 100 mg/mL is present in combination.
  • the TLR agonist may be provided in an amount synergistic with the amount of the PD-1/PD-L1 inhibitor.
  • the dosage administered will of course vary with known factors such as the pharmacokinetic properties of the particular agent, and its mode and route of administration; the age, health, and weight of the recipient; the nature and extent of symptoms , the type of concurrent treatment, the frequency of treatment, and the expected effect.
  • the pharmaceutical composition further comprises one or more pharmaceutically acceptable carriers.
  • examples of pharmaceutically acceptable carriers may include one or more of the following: water, saline, phosphate-buffered saline, dextrose, glycerol, ethanol, etc., and combinations thereof.
  • isotonic agents such as sugars, polyalcohols (eg, mannitol, sorbitol), or sodium chloride, may be included in the composition.
  • Pharmaceutically acceptable carriers may further include minor amounts of auxiliary substances that enhance the shelf-life or effectiveness of the binding protein or binding protein portion, such as wetting or emulsifying agents, preservatives or buffers.
  • the present application also provides the use of the aforementioned drug combination in the preparation of medicines for treating neoplastic diseases.
  • the neoplastic disease comprises tumor and/or wart disease.
  • the tumor includes, but is not limited to: hepatocellular carcinoma, liver metastatic cancer, advanced hepatocellular carcinoma, pancreatic cancer, adenocarcinoma, mast cell tumor or mast cell tumor, ovarian cancer, Non-small cell lung cancer, small cell lung cancer, melanoma, retinoblastoma, breast tumor, colorectal cancer, histiocytic sarcoma, brain tumor, astrocytoma, glioblastoma, neuroma, neuroblastoma , colon cancer, cervical cancer, sarcoma, prostate tumor, bladder tumor, reticuloendothelial tumor, Wilms tumor, ovarian cancer, bone cancer, osteosarcoma, kidney cancer, or cancer of the head and neck, oral cavity, larynx, or Oropharyngeal cancer, breast cancer, genitourinary tract cancer, lung cancer, gastrointestinal cancer, epidermoid cancer, melanoma.
  • the present application also provides the aforementioned drug combination for treating neoplastic diseases.
  • the present application also provides a drug for treating neoplastic diseases, which comprises the aforementioned drug combination.
  • the present application also provides a method for treating neoplastic diseases, which comprises administering an effective amount of the aforementioned drug combination to a subject in need.
  • neoplasm located in a mammal in a location selected from:
  • Brain Brain, head, eyes, nasopharynx, mouth, tongue, neck, thyroid, gastrointestinal system, liver, pancreas, gallbladder, lungs, respiratory system, genitourinary system, kidneys, bladder, breast, lymphatic system, cardiovascular system , nervous system, skin, thoracic cavity, pleura, musculoskeletal system, abdomen, of a primary or secondary nature.
  • the neoplasm comprises tumors and/or warts.
  • administering comprises topical, intraneoplastic (eg, intratumor or intrawart) or systemic administration.
  • intraneoplastic eg, intratumor or intrawart
  • systemic administration e.g, topical, intraneoplastic (eg, intratumor or intrawart) or systemic administration.
  • intratumoral or intrawart injections the approach both allows for less trauma to the patient and allows the killing of cancer cells, but not normal cells. Injecting the drug combination directly into the malignancy also greatly reduces or eliminates many common side effects.
  • administering comprises intravenous injection, intravenous drip, intramuscular injection, subcutaneous injection and/or intraneoplastic injection.
  • the tumor comprises:
  • Carcinoma or sarcoma of the esophagus, stomach, duodenum, small intestine, and the drug combination is injected into the tumor with a needle through the enteroscope, or through a hole made in the patient's abdominal wall during laparoscopic surgery. Injection into the tumor with a long syringe, or through a hole made in the patient's chest wall during thoracoscopic surgery;
  • cancer of the lung and the drug combination is injected with a syringe under the use of ultrasound, x-ray, CT scan or MR scan, or through a hole made in the patient's chest wall during thoracoscopic surgery; or
  • the neoplasm comprises a wart
  • the drug combination is injected through the patient's skin into the wart using a needle.
  • i) the PD-1 inhibitor or PD-L1 inhibitor and ii) the TLR agonist in the drug combination are administered by the same or different administration routes.
  • it comprises injecting said TLR agonist into the neoplasm.
  • it also includes injection or systemic infusion (eg, intravenous injection, intravenous infusion) of the PD-1 inhibitor or PD-L1 inhibitor into the neoplasm.
  • injection or systemic infusion eg, intravenous injection, intravenous infusion
  • it comprises injecting i) the PD-1 inhibitor or PD-L1 inhibitor, and ii) the TLR agonist of the drug combination into the neoplasm.
  • i) the PD-1 inhibitor or PD-L1 inhibitor and ii) the TLR agonist in the pharmaceutical combination are administered simultaneously or at different times.
  • the PD-1 inhibitor or PD-L1 inhibitor is administered before and/or after administration of the TLR agonist.
  • the PD-1 inhibitor or PD-L1 inhibitor in the drug combination and ii) the TLR agonist are administered simultaneously by intraneoplastic injection.
  • the PD-1 inhibitor or PD-L1 inhibitor in the drug combination is administered simultaneously with ii) the TLR agonist by intraneoplastic injection, and i) the PD -1 inhibitor or PD-L1 inhibitor and ii) said TLR agonist are located in the same dosage form.
  • the PD-1 inhibitor or PD-L1 inhibitor in the drug combination is injected intravenously, and the TLR agonist is administered simultaneously by intratumor injection, and the PD- 1 inhibitor or PD-L1 inhibitor and said TLR agonist are in separate dosage forms.
  • the present application provides a kit comprising the aforementioned drug combination.
  • each component of the pharmaceutical combination in the kit may be provided in a separate individual container.
  • the components of the pharmaceutical combinations described herein may be provided in a single container.
  • the container may be a container ready to be administered to a patient in need thereof, such as an IV bag, ampoule, or syringe.
  • the TLR agonist in the kit is formulated for intratumoral or intrawart injection administration.
  • the PD-1/PD-L1 inhibitor can be provided, for example, in the form of a powder (e.g., a lyophilized powder) or a solution for parenteral administration.
  • the PD-1/PD-L1 inhibitor can be an anti-PD-1/PD-L1 antibody as described herein formulated for administration by, for example, intravenous administration or intratumoral or intrawart injection.
  • the PD-1 or PD-L1 inhibitor and TLR agonist are formulated as a combined preparation, and used for intratumoral or intrawart injection.
  • the co-formulation is configured as a liquid formulation.
  • the compound formulation is provided in the form of a stable solution.
  • the compound preparation is provided as a dosage form that can be used for direct injection (ie, no dilution is required before use).
  • the compound preparation is configured as a solid preparation (eg, in a freeze-dried form).
  • kits described herein can be provided in sterile form.
  • the kit and its contents can be provided ready to be administered to a subject in need thereof.
  • the components of the kit are provided as formulations, and optionally in administration devices, such that administration requires little further action by the user.
  • administration devices such devices include those known and understood by those skilled in the art for the routes of administration described herein, such as, but not limited to, syringes, pumps, bags, cups, inhalers, drops, tube, patch, cream, or injector.
  • Kits described herein may also include instructions containing information on, for example, usage, dosage, administration, contraindications, and/or warnings regarding the use of such drugs.
  • mice C57/BL6JNifdc mice, female, weighing 17-22 g, 6-8 weeks old, SPF grade, were purchased from Beijing Weitong Lihua Experimental Animal Technology Co., Ltd. [Experimental animal quality certificate number: SCXK (Beijing) 2016-0006] .
  • mice All mice were free to forage and drink, and were raised at room temperature (25 ⁇ 2)°C. Feed and water were all processed by autoclaving, and all experimental feeding processes were SPF grade.
  • Tumor cell lines are selected from: mouse colorectal cancer cell lines CT26, MC38, mouse lung cancer Lewis tumor line LLC1, mouse melanoma cell line B16, mouse prostate cancer cell lines RM-1, TRAMP-C1, mouse mammary gland Cancer cell lines 4T1, NAFA, MET-1, etc.
  • Tumor cells were cultured and passaged, and the cells were collected in the logarithmic phase of the cells to make a cell suspension with a concentration of (1.0 ⁇ 10 7 ) per milliliter, and inject 0.1ml of the cell suspension under the right flank of the mouse (the number of cells was 1.0 ⁇ 10 6 per mouse), the tumor grew to a diameter of about 5 mm in about 10 days, the model was established successfully, and the tumors were randomly divided into 8.
  • Tumor weight inhibition rate [1-experimental group (G2/G3/G4) average tumor weight/negative control group G1 average tumor weight)] ⁇ 100%.
  • the data are represented by x ⁇ s, processed by using SPSS10.0 software, and adopt one-way ANOVA (one-way ANOVA). After statistical processing, the P value of the data difference among the groups was determined, and P ⁇ 0.05 was judged as having a significant difference between the groups.
  • the xenograft tumor model was used to test the anti-tumor effect of the drug combination composed of PD-L1 inhibitor RB0005 and TLR7/TLR8 agonist Imiquimod (LPG2005), that is, the inhibitory effect on the growth of subcutaneous xenograft tumor in mice.
  • mice After successfully establishing the subcutaneous colorectal cancer MC38 xenograft tumor model in mice, they were randomly divided into 6-8 mice in each group. Administer once every two days, and observe up to 15 days after a total of 3 administrations.
  • Figures 1-4 show that the new drug combination and each single drug can significantly inhibit tumor growth, and the anatomical tumor weight is significantly lower than that of the negative control group (P ⁇ 0.01, P ⁇ 0.001), and the efficacy of the new drug combination is better than that of Imiquimod or PD
  • the -L1 antibody RB0005 was used alone, indicating that the novel drug combination has greater advantages.
  • mice in the solvent control group began to appear "death" on the 9th day (when the tumor volume of a single mouse exceeded 2000mm 3 , the mouse could be considered dead in the statistical analysis of the survival rate of the mice. (It was not excluded in the curve analysis of tumor volume change))", the survival rate of the mice was 0 on the 11th day after the start of administration; the survival rate of the mice in the RB0005 single drug group was 33.3% until the end. The survival rate of mice in the Imiquimod monotherapy group was 50%. The survival rate of the mice in the single preparation group was 83.3%. Among them, the tumor disappeared in 1 mouse, and the tumor volume in 2 mice continued to decrease (tumor volume ⁇ 100mm 3 ).
  • the xenograft tumor model was used to test the anti-tumor effect of the drug combination consisting of the PD-1 inhibitor RB0004 and the TLR7/TLR8 agonist Imiquimod, that is, the inhibitory effect on the growth of subcutaneous xenograft tumors in mice.
  • mice After the humanized PD-1 mouse colorectal cancer MC38 subcutaneous xenograft tumor model was successfully established, they were randomly divided into 8 mice in each group and administered once every two days.
  • the survival rate of the mice in the solvent control group was 0 on the 13th day after the start of administration; the survival rate of the mice in the RB0004 single drug group was 37.5% until the end.
  • the survival rate of mice in the Imiquimod group was 40%.
  • the survival rate of the mice in the combined administration group (G4) was 87.5%, and the tumor volume of one mouse in the group continued to decrease (tumor volume ⁇ 100mm 3 ).
  • the tumor volume of mice in the solvent control group gradually increased over time; in the RB0004 single-drug group and LPG2005 single-drug group, the growth of tumor volume was slower than that in the solvent control group; The volume growth slowed down significantly, and the difference was statistically significant compared with the control group (P ⁇ 0.001).
  • mice colorectal cancer MC38 xenograft model was used to test the anti-tumor effect of the drug combination composed of PD-L1 inhibitor RB0005 and different doses of TLR7/TLR8 agonist Imiquimod, that is, the inhibitory effect on the growth of subcutaneous xenograft tumor in mice.
  • mice After the successful establishment of the subcutaneous xenograft tumor model in mice, they were randomly divided into groups of 8 mice and administered once every two days.
  • mice The subcutaneous transplanted tumor model in mice was successfully established. Among the three single dosage forms, the G8 group had a significant tumor growth inhibitory effect, and the tumor weight inhibition rate was above 75% (P ⁇ 0.001), while the G6 and G7 groups did not. There was a tumor inhibitory effect (P>0.05). The specific results are shown in Figure 7-8.
  • the xenograft tumor model was used to test the anti-tumor effect of the drug combination composed of PD-L1 inhibitor RB0005 and different doses of TLR7/TLR8 agonist Imiquimod, that is, the inhibitory effect on the growth of subcutaneous xenograft tumor in mice.
  • mice After the successful establishment of the subcutaneous xenograft tumor model in mice, they were randomly divided into groups of 8 mice and administered once every two days.
  • mice in groups G6, G7 and G8 were all 100% until the end of observation.
  • the tumor volume of 2 mice in the G7 group continued to decrease
  • the tumor volume of 3 mice in the G8 group continued to decrease (tumor volume ⁇ 100mm 3 ).
  • the survival rate of the mice until the end of the observation day was 100%.
  • the tumor weight inhibition rates of G6 group, G7 group and G8 group were 73.87%, 70.47% and 65.97%, respectively, which were statistically significant (P ⁇ 0.001), but the CDI index showed , G6 group and G7 group were 0.61 and 0.86 respectively, while the CDI of G8 group was 1.09, indicating that G6 group and G7 group had a synergistic drug effect.
  • the xenograft tumor model was used to test the anti-tumor effect of the drug combination composed of PD-1 inhibitor RB0004 and different doses of TLR7/TLR8 agonist Imiquimod, that is, the inhibitory effect on the growth of subcutaneous xenograft tumor in mice.
  • mice After the successful establishment of the subcutaneous xenograft tumor model in mice, they were randomly divided into groups of 8 mice and administered once every two days.
  • the tumor weight inhibition rate of the G7 group was 59.25%, which was statistically significant.
  • Example 6 The anti-tumor effect of the drug combination LPG2006 (single dosage form) composed of PD-L1 inhibitor RB0005 and different doses of TLR7/TLR8 agonist Imiquimod
  • mice After successful establishment of mouse colorectal cancer MC38 humanized CD274 cell subcutaneous xenograft tumor model, the mice were randomly divided into 6-8 groups, administered once every two days, and administered 3 times in total.
  • a single dosage form of TLR7/TLR8 agonist Imiquimod and PD-L1 monoclonal antibody the dose of Imiquimod in this dosage form is 2-6.25mpk, has tumor inhibitory effect and high survival rate (P ⁇ 0.05), the efficacy of the new drug combination is better than The PD-L1 antibody RB0005 was used alone, indicating that the new drug combination has greater advantages.
  • G4 2.0 58.5 the G5 1.8 47.4 the G6 0.9 73.7 0.6( ⁇ 1) G7 1.0 70.8 0.8( ⁇ 1) G8 1.2 64.9 >1
  • Example 7 The anti-tumor effect of the compound preparation (single dosage form and separate dosage form) composed of PD-L1 inhibitor RB0005 and TLR7/8 agonist Imiquimod
  • mice After successful establishment of mouse colorectal cancer MC38 humanized CD274 cell subcutaneous xenograft tumor model, the mice were randomly divided into 6-8 groups, administered once every two days, and administered 3 times in total.
  • the single dosage form (G7 ⁇ G8) of the new compound preparation was administered intraperitoneally and intratumorally.
  • the tumor reinhibition rate of intratumoral administration was 77%.
  • the tumor inhibitory effect was superior to that of intraperitoneal administration.
  • a synergistic drug effect was shown, and the CDI value was 0.7.
  • the single dosage form of Imiquimod and PD-L1 monoclonal antibody, the intratumoral administration (local) method has better tumor suppression effect, and the survival rate is high (P ⁇ 0.05), and it is better than the intraperitoneal administration of the compound preparation and the single use of each single drug. It shows that the new drug combination has greater advantages.
  • mice After successful establishment of mouse colorectal cancer MC38 humanized CD274 cell subcutaneous xenograft tumor model, the mice were randomly divided into 6-8 groups, administered once every two days, and administered 3 times in total.
  • the results are shown in Figure 20-22 and Table 7.
  • the tumor reinhibition rate of RB0005 single drug was 34%
  • the tumor reinhibition rate of Gardiquimod single drug was 42%
  • the tumor reinhibition rate of the new compound preparation single dosage form (G4) was 72%.
  • the survival rate of the compound preparation mice was 100%.
  • Intratumoral administration of a single dosage form of the compound preparation composed of Gardiquimod showed a synergistic drug effect, and the CDI value was ⁇ 1.
  • the single dosage form composed of TLR7/TLR8 agonist Gardiquimod and PD-L1 monoclonal antibody has tumor suppressive effect and shows drug synergy, and the survival rate in the test is high (P ⁇ 0.05), and the combination drug effect is better than single drug, indicating that the new drug Combinations have greater advantages.
  • mice After successful establishment of mouse colorectal cancer MC38 humanized CD274 cell subcutaneous xenograft tumor model, the mice were randomly divided into 6-8 groups, administered once every two days, and administered 3 times in total.
  • the tumor weight inhibition rate of Resiquimod single drug was 40%
  • the tumor weight inhibition rate of the new compound preparation single dosage form (G4) was 73%
  • the survival rate of the compound preparation mice was 85%
  • the single dosage form composed of TLR7/TLR8 agonist Resiquimod and PD-L1 monoclonal antibody has tumor suppressive effect and shows drug synergy, and the survival rate in the test is high (P ⁇ 0.05), and the combined drug effect is better than that of single drug.
  • the single dosage form of the new compound preparation (G4 ⁇ G6 ⁇ G8 ⁇ G10), among which the tumor reinhibition rate of intratumoral administration in the G8 group (Sintili) and G10 group (Tripril) was 35% and 60%, showing a synergistic Drug effect (CDI value ⁇ 1), and the survival rate of mice at the end point of the test is over 60%.
  • Intratumoral administration of Tislelizumab in the marketed PD-1 inhibitor single drug has a better tumor inhibitory effect, and the compound preparation composed of TLR7/8 agonist Imiquimod contains sintilimab and Terry respectively
  • the compound preparation of pulimumab has tumor inhibitory effect and high survival rate (P ⁇ 0.05), and this drug combination has great advantages.
  • mice After successful establishment of mouse colorectal cancer MC38 humanized CD274 cell subcutaneous xenograft tumor model, the mice were randomly divided into 6-8 groups, administered once every two days, and administered 3 times in total.
  • the results are shown in Figures 29-31 and Table 10.
  • the tumor weight inhibition rate of the marketed PD-L1 inhibitor Durvalumab single drug (G3) was 21%, and the new compound preparation composed of the TLR7/8 agonist Imiquimod (G4)
  • the tumor weight inhibition rate was 69%, showing synergistic drug effects, and the survival rate of the compound preparation was above 80%;
  • the tumor weight inhibition rate of the new composite preparation (G6) composed of RB0005 and the TLR7/8 agonist Imiquimod was 78%, and it showed a significant synergistic drug effect (CDI value ⁇ 0.5), and the survival rate of the composite preparation at the experimental end point was 100%;
  • the compound preparation containing RB0005 has a more significant tumor inhibitory effect.
  • Example 11 PD-L1 inhibitor RB0005 combined with Imiquimod LPG2006 (single dosage form), CT26 tumor model
  • mice After successful establishment of mouse colorectal cancer CT26 humanized CD274 cell subcutaneous xenograft tumor model, the mice were randomly divided into 6-8 groups, administered once every two days, and administered 3 times in total.
  • the CT26 model is not sensitive to RB0005 single drug, but the single dosage form composed of RB0005 and TLR7/TLR8 agonist Imiquimod has tumor inhibitory effect and shows drug synergy, indicating that the combined drug effect is better than single drug and improves the tumor model's PD-L1 Inhibitor sensitivity.
  • Example 12 Inhibition of MC38 tumor proliferation by sequential administration of Imiquimod and PD-L1 monoclonal antibody
  • mice After successful establishment of mouse colorectal cancer MC38 humanized CD274 cell subcutaneous xenograft tumor model, the mice were randomly divided into 6-8 groups, administered once every two days, and administered 3 times in total.
  • the sequential administration group (G4) had a significant inhibitory effect on tumor proliferation, and the tumor volume increased slowly during the observation period after the administration. Compared with the control group and the RB0005 single-drug group, the sequential administration group (G4) had a statistically significant difference (ANOVA, P ⁇ 0.001), and the tumor weight inhibition rate was 68%.
  • Intratumoral administration of Imiquimod and PD-L1 monoclonal antibody by first administering small molecules for 48 hours and then administering PD-L1 inhibitors has a significant tumor inhibitory effect, showing a synergistic drug effect, and improving the survival rate of mice (P ⁇ 0.05), sequential administration has significant advantages compared with single drug.

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Abstract

一种药物组合及其应用。该药物组合包含PD-1抑制剂和/或PD-L1免疫检查点抑制剂；以及Toll样受体激动剂。该药物组合抗肿瘤药效比单独使用免疫检查点抑制剂或单独使用Toll样受体激动剂具有更好的抗肿瘤药效，实现了机体固有免疫通路激活和防止免疫逃逸双管齐下进行治疗，具有高效、低毒的临床应用前景。

Description

药物组合及其用途 技术领域
本申请涉及生物医药领域，具体的涉及一种药物组合的开发及其应用。
背景技术
过去十年来，靶向PD-1和PD-L1为癌症临床治疗开辟了新时代。然而，对于许多肿瘤类型，免疫检查点抑制剂单药效果不佳。即便对于免疫应答型肿瘤，大部分患者也没有持久的临床获益。绝大多数患者发生了原发性或后天性治疗耐药性。在多数情况下，免疫治疗耐药性可归因于免疫抑制性TME的存在和体内免疫细胞数量下降不足以激活T细胞抗肿瘤效应。因此，与单药治疗相比，积极寻求能够减少肿瘤环境中的免疫抑制或增强细胞毒性细胞对肿瘤反应的新的治疗策略尤为重要。
PD-L1/PD-1信号通路是免疫反应中非常重要的共抑制信号途径。研究表明，当PD-L1与PD-1结合后，将会通过补充具有SH2结构域的蛋白酪氨酸磷酸酶SHP-1和SHP-2。这两种磷酸酶能够使CD3ζ链的免疫受体酪氨酸活化基序(ITAM)磷酸化程度降低，削弱ZAP-70活化，并抑制TCR下游信号传递，从而起到共抑制T细胞活化的作用，通过这种负向调节效应来防止效应T细胞过度激活而导致自身免疫损伤。
研究表明一些病毒感染也与PD-L1/PD-1信号通路息息相关。例如，在慢性HIV感染中，PD-1被发现在特异性识别HIV的CD8+T细胞表面高表达，病毒通过激活PD-L1/PD-1信号途径，使得特异性识别HIV的CD8+T细胞活性受到抑制，细胞因子的分泌能力及T细胞自身的增殖能力大大削弱，引发适应性免疫功能缺陷。
Toll样受体(TLRs)是天然免疫中重要的模式识别受体(PRRs)，特异性识别病原微生物表面保守的病原体相关分子模式(PAMP)及一些内源性配体，即损伤相关的分子模式(DAMP)。通过MyD88依赖或者非依赖途径，引发信号转导，诱导机体产生I型干扰素和多种趋化因子及炎症细胞因子(如IL-1、IL-6、TNF-α等)，启动机体天然免疫应答，并最终激活获得性免疫系统，在特异性与非特异性免疫应答中扮演重要角色。
TLRs在抗病毒天然免疫中扮演着重要角色。研究发现，与病毒识别和机体抗病毒免疫有关的TLRs主要包括TLR2、TLR3、TLR4、TLR7、TLR8、TLR9。其中定位于细胞膜表面的TLR2、TLR4主要识别病毒的包膜蛋白；位于细胞内的TLR3、TLR7、TLR8、TLR9主要识别病毒核酸。
TLR7/8是Toll样受体(Toll like receptor，TLR)成员之一，主要分布在浆树突状细胞 (pDC)、B细胞的胞内，TLR7/8主要识别ssRNA病毒，在人体识别和清除病微生物过程中发挥重要作用。在识别“病原体相关分子模式”后，通过启动信号级联发挥作用，其中模式识别受体(PRRs)是这个级联反应的关键，而Toll样受体7和8是其中重要的PRRs，可以激发抗原提呈细胞，诱发树突状细胞分泌多种细胞因子以及表达多种共刺激分子，刺激干扰素-α、肿瘤坏死因子(TNF)和白介素(IL-1、IL-6和IL-8等)的合成，进而活化机体先天免疫反应，同时，还可激活pDC，提高pDC的抗提呈能力，促进CD4+T细胞的增殖，并进一步激活CD8+T细胞，杀伤肿瘤细胞，增强机体抗病毒和抗肿瘤效应。
发明内容
本申请提供了一种药物组合及其在抗肿瘤药物中的应用。该药物组合主要由两部分组成：
(1)免疫检查点抑制剂(如PD-1/PD-L1)；(2)TLR激动剂(如咪唑喹啉衍生物)。本申请的药物组合可以制备高效、低毒的抗肿瘤药物。PD-L1/PD-1抑制剂与TLR激动剂联用，一方面解除抑制性信号，增强T细胞活化，促进适应性免疫系统反应；另一方面诱导INF和细胞因子的表达，促进T细胞的聚集，激活固有免疫系统反应，双管齐下增强细胞毒性细胞对肿瘤等的反应，具有很高的临床前景及应用价值。
一方面，本申请提供一种药物组合，其包含程序性细胞死亡蛋白1(PD-1)抑制剂和/或程序性死亡配体1(PD-L1)抑制剂，以及TLR激动剂。
在某些实施方式中，其中所述TLR包括TLR1，TLR2，TLR3，TLR4，TLR5，TLR6，TLR7，TLR8，TLR9和/或TLR10。
在某些实施方式中，其中所述TLR激动剂选自：TLR7激动剂，TLR8激动剂，TLR9激动剂中的一种或多种。
在某些实施方式中，其中所述TLR激动剂包括TLR7和TLR8双重激动剂(TLR7/TLR8激动剂)。
在某些实施方式中，其中所述TLR激动剂包括dsRNA、ssRNA、CpG DNA、咪唑喹啉衍生物和/或鸟苷类似物。
在某些实施方式中，其中所述TLR激动剂包括咪唑喹啉衍生物。
在某些实施方式中，其中所述TLR激动剂选自咪喹莫特(Imiquimod)、嘎德莫特(Gardiquimod)、瑞喹莫特(Resiquimod)、1V209、Selgantolimod(GS-9688)、Vesatolimod(GS-9620)、Sumanirole、PF-4878691、其衍生物、及其类似物中的一种或多种。
在某些实施方式中，其中所述TLR激动剂包括咪喹莫特、瑞喹莫特或其药学上可接受的盐。
在某些实施方式中，其中所述TLR激动剂选自LHC-165、NKTR-262、DN1508052-01、SHR2150、CL307、CL264、洛索立宾、艾沙托立宾、DSR-6434、GSK2245035、SM-276001、SM-324405、SM-324406、AZ12441970和AZ12443988其衍生物、及其类似物中的一种或多种。
在某些实施方式中，其中所述PD-1抑制剂具有以下一个或多个特征：
a.抑制或减少PD-1表达，例如PD-1的转录或翻译；
b.抑制或降低PD-1活性，例如抑制或降低PD-1与其同源配体，例如PD-L1或PD-L2的结合；和
c.结合PD-1或其一个或多个配体，例如PD-L1或PD-L2。
在某些实施方式中，其中所述PD-1抑制剂包括抗PD-1抗体或其抗原结合片段。
在某些实施方式中，其中所述抗PD-1抗体选自Pembrolizumab，Nivolumab，Pidilizumab，Tislelizumab，Camrelizumab(SHR-1210)，Sintilimab，Toripalimab，MEDI0680，BGB-A317，TSR-042，REGN2810、PF-06801591、RB0004、其类似物及它们的组合。
在某些实施方式中，其中所述抗PD-1抗体包含抗体重链可变区(VH)中的至少一个CDR，所述VH包含SEQ ID NO:8所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含VH，所述VH包含HCDR3，所述HCDR3包含SEQ ID NO:3所示的氨基酸序列。
在某些实施方式中，其中所述VH还包含HCDR2，其中所述HCDR2包含SEQ ID NO:2所示的氨基酸序列。
在某些实施方式中，其中所述VH还包含HCDR1，其中所述HCDR1包含SEQ ID NO:1所示的氨基酸序列。
在某些实施方式中，其中所述VH包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:3所示的氨基酸序列，所述HCDR2包含SEQ ID NO:2所示的氨基酸序列，所述HCDR1包含SEQ ID NO:1所示的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR1，所述HFR1的C末端与所述HCDR1的N末端直接或间接相连，且所述HFR1包含SEQ ID NO:4所示的氨基酸序列或与SEQ ID NO:4所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR2，所述HFR2的N末端与所述HCDR1的C末端直接或间接相连，且所述HFR2的C末端与所述HCDR2的N末端直接或间接相连；且所述HFR2包含SEQ ID NO:5所示的氨基酸序列或与SEQ ID NO:5所示的氨基酸序 列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR3，所述HFR3的N末端与所述HCDR2的C末端直接或间接相连，且所述HFR3的C末端与所述HCDR3的N末端直接或间接相连；且所述HFR3包含SEQ ID NO:6所示的氨基酸序列或与SEQ ID NO:6所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR4，所述HFR4的N末端与所述HCDR3的C末端直接或间接相连，且所述HFR4包含SEQ ID NO:7所示的氨基酸序列或与SEQ ID NO:7所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR1，HFR2，HFR3和HFR4，所述HFR1的C末端与所述HCDR1的N末端直接或间接相连，所述HFR2的N末端与所述HCDR1的C末端直接或间接相连，且所述HFR2的C末端与所述HCDR2的N末端直接或间接相连，所述HFR3的N末端与所述HCDR2的C末端直接或间接相连，且所述HFR3的C末端与所述HCDR3的N末端直接或间接相连，所述HFR4的N末端与所述HCDR3的C末端直接或间接相连；其中，所述HFR1包含SEQ ID NO:4所示的氨基酸序列或与SEQ ID NO:4所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述HFR2包含SEQ ID NO:5所示的氨基酸序列或与SEQ ID NO:5所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述HFR3包含SEQ ID NO:6所示的氨基酸序列或与SEQ ID NO:6所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述HFR4包含SEQ ID NO:7所示的氨基酸序列或与SEQ ID NO:7所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含VH，所述VH包含SEQ ID NO:8所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含抗体重链(HC)，所述HC包含SEQ ID NO:9所示氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含抗体轻链可变区(VL)中的至少一个CDR，所述VL包含SEQ ID NO:17所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含VH中的至少一个CDR，所述VH包含SEQ ID NO:8所示的氨基酸序列，且所述抗PD-1抗体包含VL中的至少一个CDR，所述VL包含SEQ ID NO:17所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含VL，所述VL包含LCDR1，所述LCDR1包含SEQ ID NO:10所示的氨基酸序列。
在某些实施方式中，其中所述VL还包含LCDR2，其中所述LCDR2包含SEQ ID NO:11所示的氨基酸序列。
在某些实施方式中，其中所述VL还包含LCDR3，其中所述LCDR3包含SEQ ID NO:12所示的氨基酸序列。
在某些实施方式中，其中所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:10所示的氨基酸序列，所述LCDR2包含SEQ ID NO:11所示的氨基酸序列，所述LCDR3包含SEQ ID NO:12所示的氨基酸序列。
在某些实施方式中，其中抗PD-1抗体包含VH以及抗体VL，所述VH包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:3所示的氨基酸序列，所述HCDR2包含SEQ ID NO:2所示的氨基酸序列，所述HCDR1包含SEQ ID NO:1所示的氨基酸序列；且所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:10所示的氨基酸序列，所述LCDR2包含SEQ ID NO:11所示的氨基酸序列，所述LCDR3包含SEQ ID NO:12所示的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR1，所述LFR1的C末端与所述LCDR1的N末端直接或间接相连，且所述LFR1包含SEQ ID NO:13所示的氨基酸序列或与SEQ ID NO:13所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR2，所述LFR2的N末端与所述LCDR1的C末端直接或间接相连，且所述LFR2的C末端与所述LCDR2的N末端直接或间接相连；且所述LFR2包含SEQ ID NO:14所示的氨基酸序列或与SEQ ID NO:14所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR3，所述LFR3的N末端与所述LCDR2的C末端直接或间接相连，且所述LFR3的C末端与所述LCDR3的N末端直接或间接相连；且所述LFR3包含SEQ ID NO:15所示的氨基酸序列或与SEQ ID NO:15所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR4，所述LFR4的N末端与所述LCDR3的C末端直接或间接相连，且所述LFR4包含SEQ ID NO:16所示的氨基酸序列或与SEQ ID NO:16所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR1，LFR2，LFR3和LFR4，所述LFR1的C末端与所述LCDR1的N末端直接或间接相连，所述LFR2的N末端与所述LCDR1的C末端直接或间接相连，且所述LFR2的C末端与所述LCDR2的N末端直接或间接相连， 所述LFR3的N末端与所述LCDR2的C末端直接或间接相连，且所述LFR3的C末端与所述LCDR3的N末端直接或间接相连，所述LFR4的N末端与所述LCDR3的C末端直接或间接相连；其中，所述LFR1包含SEQ ID NO:13所示的氨基酸序列或与SEQ ID NO:13所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述LFR2包含SEQ ID NO:14所示的氨基酸序列或与SEQ ID NO:14所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述LFR3包含SEQ ID NO:15所示的氨基酸序列或与SEQ ID NO:15所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述LFR4包含SEQ ID NO:16所示的氨基酸序列或与SEQ ID NO:16所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含VL，所述VL包含SEQ ID NO:17所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含VH以及VL，所述VH包含SEQ ID NO:8所示的氨基酸序列，且所述VL包含SEQ ID NO:17所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含抗体轻链(LC)，所述LC包含SEQ ID NO:18所示氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含HC和LC，所述HC包含SEQ ID NO:9所示氨基酸序列，且所述LC包含如SEQ ID NO:18所示氨基酸序列。
在某些实施方式中，其中所述PD-L1抑制剂具有以下一个或多个特征：
a.抑制或减少PD-L1表达，例如PD-L1的转录或翻译；
b.抑制或降低PD-L1活性，例如抑制或降低PD-L1与其关联受体，例如PD-1的结合；和
c.结合PD-L1或其受体，例如PD-1。
在某些实施方式中，其中所述PD-L1抑制剂包括抗PD-L1抗体或其抗原结合片段。
在某些实施方式中，其中所述抗PD-L1抗体选自Durvalumab，Atezolizumab，Avelumab，MDX-1105，YW243.55.S70，MDPL3280A，AMP-224、LY3300054、RB0005、其类似物及它们的组合。
在某些实施方式中，其中所述抗PD-L1抗体包含VH中的至少一个CDR，所述VH包含SEQ ID NO:25所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VH，所述VH包含HCDR3，所述HCDR3包含SEQ ID NO:21所示的氨基酸序列。
在某些实施方式中，其中所述VH还包含HCDR2，其中所述HCDR2包含SEQ ID NO: 20所示的氨基酸序列。
在某些实施方式中，其中所述VH还包含HCDR1，其中所述HCDR1包含SEQ ID NO:19所示的氨基酸序列。
在某些实施方式中，其中所述VH包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:21所示的氨基酸序列，所述HCDR2包含SEQ ID NO:20所示的氨基酸序列，所述HCDR1包含SEQ ID NO:19所示的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR1，所述HFR1的C末端与所述HCDR1的N末端直接或间接相连，且所述HFR1包含SEQ ID NO:22所示的氨基酸序列或与SEQ ID NO:22所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR2，所述HFR2的N末端与所述HCDR1的C末端直接或间接相连，且所述HFR2的C末端与所述HCDR2的N末端直接或间接相连；且所述HFR2包含SEQ ID NO:23所示的氨基酸序列或与SEQ ID NO:23所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR3，所述HFR3的N末端与所述HCDR2的C末端直接或间接相连，且所述HFR3的C末端与所述HCDR3的N末端直接或间接相连；且所述HFR3包含SEQ ID NO:24所示的氨基酸序列或与SEQ ID NO:24所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR4，所述HFR4的N末端与所述HCDR3的C末端直接或间接相连，且所述HFR4包含SEQ ID NO:7所示的氨基酸序列或SEQ ID NO:7所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR1，HFR2，HFR3和HFR4，所述HFR1的C末端与所述HCDR1的N末端直接或间接相连，所述HFR2的N末端与所述HCDR1的C末端直接或间接相连，且所述HFR2的C末端与所述HCDR2的N末端直接或间接相连，所述HFR3的N末端与所述HCDR2的C末端直接或间接相连，且所述HFR3的C末端与所述HCDR3的N末端直接或间接相连，所述HFR4的N末端与所述HCDR3的C末端直接或间接相连；其中，所述HFR1包含SEQ ID NO:22所示的氨基酸序列或与SEQ ID NO:22所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述HFR2包含SEQ ID NO:23所示的氨基酸序列或与SEQ ID NO:23所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述HFR3包含SEQ ID NO:24所示的氨基酸序列或与SEQ ID NO:24所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述HFR4包含SEQ ID NO:7所示的氨基 酸序列或与SEQ ID NO:7所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VH，所述VH包含SEQ ID NO:25所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含HC，所述HC包含SEQ ID NO:26所示氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VL中的至少一个CDR，所述VL包含SEQ ID NO:37所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VH中的至少一个CDR，所述VH包含SEQ ID NO:25所示的氨基酸序列，且所述抗PD-L1抗体包含VL中的至少一个CDR，所述VL包含SEQ ID NO:37所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VH中的至少一个CDR，所述VH包含SEQ ID NO:25所示的氨基酸序列，且所述抗PD-L1抗体包含VL中的至少一个CDR，所述VL包含SEQ ID NO:38、SEQ ID NO:39或SEQ ID NO:40所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VL，所述VL包含LCDR1，所述LCDR1包含SEQ ID NO:27所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VL，所述VL包含LCDR1，所述LCDR1包含SEQ ID NO:28、SEQ ID NO:29或SEQ ID NO:30所示的氨基酸序列。
在某些实施方式中，其中所述VL还包含LCDR2，其中所述LCDR2包含SEQ ID NO:31所示的氨基酸序列。
在某些实施方式中，其中所述VL还包含LCDR3，其中所述LCDR3包含SEQ ID NO:32所示的氨基酸序列。
在某些实施方式中，其中所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:27所示的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列。
在某些实施方式中，其中所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:28所示的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列；
所述LCDR1包含SEQ ID NO:29所示的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列；或
所述LCDR1包含SEQ ID NO:30所示的氨基酸序列，所述LCDR2包含SEQ ID NO:31 所示的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VH以及抗体VL，所述VH包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:21所示的氨基酸序列，所述HCDR2包含SEQ ID NO:20所示的氨基酸序列，所述HCDR1包含SEQ ID NO:19所示的氨基酸序列；且所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:27所示的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VH以及抗体VL，所述VH包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:21所示的氨基酸序列，所述HCDR2包含SEQ ID NO:20所示的氨基酸序列，所述HCDR1包含SEQ ID NO:19所示的氨基酸序列；且所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:28、SEQ ID NO:29或SEQ ID NO:30所示的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR1，所述LFR1的C末端与所述LCDR1的N末端直接或间接相连，且所述LFR1包含SEQ ID NO:33所示的氨基酸序列或与SEQ ID NO:33所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR2，所述LFR2的N末端与所述LCDR1的C末端直接或间接相连，且所述LFR2的C末端与所述LCDR2的N末端直接或间接相连；且所述LFR2包含SEQ ID NO:34所示的氨基酸序列或与SEQ ID NO:34所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR3，所述LFR3的N末端与所述LCDR2的C末端直接或间接相连，且所述LFR3的C末端与所述LCDR3的N末端直接或间接相连；且所述LFR3包含SEQ ID NO:35所示的氨基酸序列或与SEQ ID NO:35所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR4，所述LFR4的N末端与所述LCDR3的C末端直接或间接相连，且所述LFR4包含SEQ ID NO:36所示的氨基酸序列或与SEQ ID NO:36所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR1，LFR2，LFR3和LFR4，所述LFR1的C末端与所述LCDR1的N末端直接或间接相连，所述LFR2的N末端与所述LCDR1的C末端直接或间接相连，且所述LFR2的C末端与所述LCDR2的N末端直接或间接相连， 所述LFR3的N末端与所述LCDR2的C末端直接或间接相连，且所述LFR3的C末端与所述LCDR3的N末端直接或间接相连，所述LFR4的N末端与所述LCDR3的C末端直接或间接相连；其中，所述LFR1包含SEQ ID NO:33所示的氨基酸序列或与SEQ ID NO:33所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述LFR2包含SEQ ID NO:34所示的氨基酸序列或与SEQ ID NO:34所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述LFR3包含SEQ ID NO:35所示的氨基酸序列或与SEQ ID NO:35所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述LFR4包含SEQ ID NO:36所示的氨基酸序列或与SEQ ID NO:36所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VL，所述VL包含SEQ ID NO:37所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VL，所述VL包含SEQ ID NO:38、SEQ ID NO:39或SEQ ID NO:40所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VH以及VL，所述VH包含SEQ ID NO:25所示的氨基酸序列，且所述VL包含SEQ ID NO:37所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VH以及VL，所述VH包含SEQ ID NO:25所示的氨基酸序列，且所述VL包含SEQ ID NO:38、SEQ ID NO:39或SEQ ID NO:40所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含LC，所述LC包含SEQ ID NO:41所示氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含LC，所述LC包含SEQ ID NO:42、SEQ ID NO:43或SEQ ID NO:44所示氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含HC和LC，所述HC包含SEQ ID NO:26所示氨基酸序列，且所述LC包含SEQ ID NO:41所示氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含HC和LC，所述HC包含SEQ ID NO:26所示氨基酸序列，且所述LC包含SEQ ID NO:42、SEQ ID NO:43或SEQ ID NO:44所示氨基酸序列。
在某些实施方式中，所述药物组合中的i)所述PD-1抑制剂和/或PD-L1抑制剂，与ii)所述TLR激动剂在所述药物组合中彼此不混合。
在某些实施方式中，所述药物组合中的i)所述PD-1抑制剂和/或PD-L1抑制剂，与ii)所述TLR激动剂位于单一的剂型中。
在某些实施方式中，其中所述药物组合被配制成药物组合物。
在某些实施方式中，其中所述药物组合物包括PD-1抑制剂或PD-L1抑制剂，以及TLR激动剂。
在某些实施方式中，其中所述TLR激动剂的存在量为约0.0001mg/kg至约200mg/kg。
在某些实施方式中，其中所述PD-1抑制剂或PD-L1抑制剂的存在量为0.0001mg/kg至约200mg/kg。
在某些实施方式中，其中所述药物组合物还包含一种或多种药学上可接受的载体。
另一方面，本申请还提供了前述药物组合在制备药物中的用途，所述药物用于治疗赘生性疾病。
在某些实施方式中，其中所述赘生性疾病包括肿瘤和/或疣类疾病。
另一方面，本申请还提供了前述药物组合用于治疗赘生性疾病。
另一方面，本申请还提供了一种治疗赘生性疾病的药物，其包含前述药物组合。
另一方面，本申请还提供了一种治疗赘生性疾病的方法，其包括向有需要的受试者施用有效量的前述药物组合。
在某些实施方式中，其中所述受试者患有赘生物。
在某些实施方式中，其中所述赘生物包括肿瘤和/或疣。
在某些实施方式中，其中所述施用包括局部、赘生物内(例如，肿瘤内或疣体内)或全身性施用。
在某些实施方式中，其中所述施用包括静脉注射、静脉滴注、肌肉注射、皮下注射和/或赘生物内注射。
在某些实施方式中，其中采取相同或不同的给药途径所述药物组合中的i)PD-1抑制剂或PD-L1抑制剂，与ii)TLR激动剂。
在某些实施方式中，其包括向赘生物中注射所述TLR激动剂。
在某些实施方式中，其还包括向赘生物中注射或全身输注所述PD-1抑制剂或PD-L1抑制剂。
在某些实施方式中，其包括向赘生物中注射所述药物组合中的i)所述PD-1抑制剂或PD-L1抑制剂，与ii)所述TLR激动剂。
在某些实施方式中，其中同时或不同时施用所述药物组合中的i)PD-1抑制剂或PD-L1抑制剂，与ii)所述TLR激动剂。
在某些实施方式中，其中在所述TLR激动剂施用之前和/或施用之后施用所述PD-1抑制 剂或PD-L1抑制剂。
在某些实施方式中，其中在所述TLR激动剂施用之后施用所述PD-1抑制剂或PD-L1抑制剂。
在某些实施方式中，所述方法包括：i)向赘生物中注射所述TLR激动剂；ii)在施用所述TLR激动剂之后向赘生物中注射或全身输注所述PD-1抑制剂或PD-L1抑制剂。
在某些实施方式中，所述方法包括：i)向赘生物中注射所述STING通路激动剂；ii)在施用所述STING通路激动剂之后全身输注所述PD-1抑制剂或PD-L1抑制剂。
在某些实施方式中，其中在所述TLR激动剂施用之后约2h至约72h施用所述PD-1抑制剂或PD-L1抑制剂。
在某些实施方式中，其中在所述TLR激动剂施用之后约2h，约4h，约8h，约16h，约24h，约36h，约48h，约60h，或约72h施用所述PD-1抑制剂或PD-L1抑制剂。
在某些实施方式中，所述方法包括：i)向赘生物中注射所述STING通路激动剂；ii)在施用所述STING通路激动剂之后约48h后向赘生物中注射或全身输注所述PD-1抑制剂或PD-L1抑制剂。
在某些实施方式中，其中所述药物组合中的i)PD-1抑制剂或PD-L1抑制剂，与ii)所述TLR激动剂同时施用。
在某些实施方式中，其中所述药物组合中的i)PD-1抑制剂或PD-L1抑制剂，与ii)所述TLR激动剂通过赘生物内注射方式同时施用，且所述PD-1抑制剂或PD-L1抑制剂与所述TLR激动剂位于同一剂型内。
在某些实施方式中，其中所述药物组合中的i)PD-1抑制剂或PD-L1抑制剂，与ii)所述TLR激动剂通过赘生物内注射方式同时施用，且所述PD-1抑制剂或PD-L1抑制剂与所述TLR激动剂位于分开的剂型内。
另一方面，本申请提供一种药盒，其包含本申请所述的药物组合。
本领域技术人员能够从下文的详细描述中容易地洞察到本申请的其它方面和优势。下文的详细描述中仅显示和描述了本申请的示例性实施方式。如本领域技术人员将认识到的，本申请的内容使得本领域技术人员能够对所公开的具体实施方式进行改动而不脱离本申请所涉及发明的精神和范围。相应地，本申请的附图和说明书中的描述仅仅是示例性的，而非为限制性的。
附图说明
本申请所涉及的发明的具体特征如所附权利要求书所显示。通过参考下文中详细描述的 示例性实施方式和附图能够更好地理解本申请所涉及发明的特点和优势。对附图简要说明如下：
图1显示的是本申请所述PD-L1抑制剂RB0005与Imiquimod药物组合体内药效研究-给药期间各组小鼠体重变化；
图2显示的是本申请所述PD-L1抑制剂RB0005与Imiquimod药物组合体内药效研究-给药期间各组小鼠生存率变化；
图3A显示的是本申请所述PD-L1抑制剂RB0005与Imiquimod药物组合体内药效研究-给药期间各组小鼠肿瘤增殖变化；
图3B显示的是本申请所述PD-L1抑制剂RB0005与Imiquimod药物组合体内药效研究-治疗后第15天各组小鼠肿瘤增殖变化；
图4显示的是本申请所述PD-1抑制剂RB0004与Imiquimod药物组合体内药效研究-给药期间各组小鼠生存率变化；
图5A显示的是本申请所述PD-1抑制剂RB0004与Imiquimod药物组合体内药效研究-给药期间各组小鼠肿瘤增殖变化；
图5B显示的是本申请所述PD-1抑制剂RB0004与Imiquimod药物组合体内药效研究-治疗后第15天各组小鼠肿瘤增殖变化；
图6显示的是本申请所述PD-1抑制剂RB0004与Imiquimod药物组合体内药效研究-给药观察结束后解剖所得小鼠瘤重抑制率；
图7显示的是本申请所述PD-L1抑制剂RB0005与不同剂量的Imiquimod药物组合体内药效研究-给药期间各组小鼠生存率变化；
图8显示的是本申请所述PD-L1抑制剂RB0005与不同剂量的Imiquimod药物组合体内药效研究-给药观察结束后解剖所得小鼠瘤重抑制率；
图9显示的是本申请所述PD-L1抑制剂RB0005与不同剂量的Imiquimod药物组合体内药效研究-给药期间各组小鼠体重变化；
图10显示的是本申请所述PD-L1抑制剂RB0005与不同剂量的Imiquimod(包含5％Imiquimod乳膏)药物组合体内药效研究-给药期间各组小鼠生存率变化；
图11显示的是本申请所述PD-L1抑制剂RB0005与不同剂量的Imiquimod(包含5％Imiquimod乳膏)药物组合体内药效研究-给药观察结束后解剖所得小鼠瘤重抑制率；
图12显示的是本申请所述PD-1抑制剂RB0004与不同剂量的Imiquimod(包含5％Imiquimod乳膏)药物组合体内药效研究-给药期间各组小鼠生存率变化；
图13显示的是本申请所述PD-1抑制剂RB0004与不同剂量的Imiquimod(包含5％Imiquimod乳膏)药物组合体内药效研究-给药观察结束后解剖所得小鼠瘤重抑制率。
图14显示的是本申请实施例6中给药期间各组小鼠肿瘤增殖趋势。
图15显示的是本申请实施例6中给药期间各组小鼠生存曲线。
图16显示的是本申请实施例6中给药期间各组小鼠平均肿瘤质量。
图17显示的是本申请实施例7中给药期间各组小鼠肿瘤增殖趋势。
图18显示的是本申请实施例7中给药期间各组小鼠生存曲线。
图19显示的是本申请实施例7中给药期间各组小鼠平均肿瘤质量。
图20显示的是本申请实施例8中给药期间各组小鼠肿瘤增殖趋势。
图21显示的是本申请实施例8中给药期间各组小鼠生存曲线。
图22显示的是本申请实施例8中给药期间各组小鼠平均肿瘤质量。
图23显示的是本申请实施例9中给药期间各组小鼠肿瘤增殖趋势。
图24显示的是本申请实施例9中给药期间各组小鼠生存曲线。
图25显示的是本申请实施例9中给药期间各组小鼠平均肿瘤质量。
图26显示的是本申请实施例10.1中给药期间各组小鼠肿瘤增殖趋势。
图27显示的是本申请实施例10.1中给药期间各组小鼠生存曲线。
图28显示的是本申请实施例10.1中给药期间各组小鼠平均肿瘤质量。
图29显示的是本申请实施例10.2中给药期间各组小鼠肿瘤增殖趋势。
图30显示的是本申请实施例10.2中给药期间各组小鼠生存曲线。
图31显示的是本申请实施例10.2中给药期间各组小鼠平均肿瘤质量。
图32显示的是本申请实施例11中给药期间各组小鼠体重变化趋势。
图33显示的是本申请实施例11中给药期间各组小鼠小鼠肿瘤增殖趋势。
图34显示的是本申请实施例11中给药期间各组小鼠平均肿瘤质量。
图35显示的是本申请实施例12中序贯给药的示意图。
图36显示的是本申请实施例12中给药期间各组小鼠肿瘤增殖趋势。
图37显示的是本申请实施例12中给药期间各组小鼠生存曲线。
图38显示的是本申请实施例12中给药期间各组小鼠平均肿瘤质量。
具体实施方式
以下由特定的具体实施例说明本申请发明的实施方式，熟悉此技术的人士可由本说明书所公开的内容容易地了解本申请发明的其他优点及效果。
术语定义
在本申请中，术语“PD-1”通常是指编程性细胞死亡蛋白1，是一种288个氨基酸的I型膜蛋白，首次在1992年有描述(Ishida et al.,EMBO J.,11(1992)，3887-3895)。PD-1是扩大的CD28/CTLA-4T细胞调节物家族的一个成员且具有两种配体，PD-L1(B7-H1，CD274)和PD-L2(B7-DC，CD273)。该蛋白质的结构包括一个胞外IgV域，接着是一个跨膜区和一个胞内尾。胞内尾含有位于免疫受体基于酪氨酸的抑制性基序和免疫受体基于酪氨酸的转换基序中的两个磷酸化位点，提示PD-1阴性负调节TCR信号。这与配体结合后SHP-1和SHP-2磷酸酶对PD-1的胞质尾的结合一致。虽然PD-1不在幼稚T细胞上表达，但是它在T细胞受体(TCR)介导的活化后上调且在活化的和耗尽的T细胞二者上观察到(Agata et al.,Int.Immunology 8(1996),765-772)。这些耗尽的T细胞具有功能障碍性表型且不能够恰当地响应。虽然PD-1具有相对广泛的表达样式，但是它最重要的作用很可能是作为T细胞上的共抑制性受体(Chinai et al.,Trends in Pharmacological Sciences 36(2015),587-595)。当前的治疗办法因而聚焦于阻断PD-1与它的配体的相互作用以增强T细胞应答。在本申请中，所述PD-1可以包括人PD-1(hPD-1)或其变体、同种型以及物种同源物，以及与hPD-1具有至少一个共同表位的类似物。示例性的hPD-1的氨基酸序列可以在GenBank登录号U64863下找到。
在本申请中，术语“PD-L1”通常是指程序性细胞死亡1配体1，也可称为B7同源物1、B7-H1、分化簇274、(3)274或CD274，其与PD-1结合后下调T细胞活化和细胞因子分泌。“PD-L1”包括任何脊椎动物来源的任何天然PD-L1，所述任何脊椎动物来源包括哺乳动物，诸如灵长类(例如，人和食蟹猴)和啮齿类(例如，小鼠和大鼠)。所述术语涵盖“全长”、未加工的PD-L1以及由细胞加工所产生的任何形式的PD-L1。PD-L1可作为跨膜蛋白或作为可溶性蛋白存在。“PD-L1”包括完整的PD-L1及其片段，还包括PD-L1的功能性变体、同工型、物种同源物、衍生物、类似物，以及具有至少一个与PD-L1共同表位的类似物。PD-L1的基本结构包括4个结构域：胞外Ig样V型结构域和Ig样C2型结构域、跨膜结构域以及细胞质结构域。示例性的人PD-L1氨基酸序列可在NCBI登录号NP_001254653或UniProt登录号Q9NZQ7下找到。
在本申请中，术语“抑制剂”通常指能够完全或部分地预防或降低一种或多种特定生物分子(例如，蛋白质(例如PD-1或PD-L1)、多肽、脂多糖、糖蛋白、核糖核蛋白复合体等)的生理功能的化合物/物质或组合物。所述降低一种或多种特定蛋白质的生理功能可以包含蛋白质本身活性(例如与其他分子结合的能力等)的降低或者本身存在量的降低。合适的抑制剂分子可以包括拮抗剂抗体或抗体片段、小分子的片段或衍生物、肽、反义寡核苷酸、小的 有机分子等。在某些实施方式中，所述抑制剂能够阻碍细胞信号通路的激活。在某些实施方式中，所述PD-1/PD-L1抑制剂是抗PD-1/PD-L1抗体或其抗原结合片段。
在本申请中，术语“Pembrolizumab”，“Nivolumab”，“Pidilizumab”，“Tislelizumab”，“Camrelizumab(SHR-1210)”，“Sintilimab”，“Toripalimab”，“MEDI0680”，“BGB-A317”，“TSR-042”，“REGN2810”、“PF-06801591”、“Durvalumab”，“Atezolizumab”，“Avelumab”，“MDX-1105”，“YW243.55.S70”，“MDPL3280A”，“AMP-224”、“LY3300054”，“RB0004”，“RB0005”根据其在本领域中理解的一般和普通含义使用。
在本申请中，术语“Toll样受体”和“TLR”通常是指高度保守的哺乳动物蛋白家族的任何成员，其识别病原体相关的分子模式并且在先天免疫中充当关键的信号传导元件。TLR多肽共有特征性结构，其包括具有富含亮氨酸重复的细胞外结构域、跨膜结构域和参与TLR信号传导的细胞内结构域。
在本申请中，术语“Toll样受体7”和“TLR7”通常是指与可公开获得的TLR7序列例如人类TLR7多肽的GenBank登录号AAZ99026、或鼠TLR7多肽的GenBank登录号AAK62676共有至少70％、80％、90％、95％、96％、97％、98％、99％或更多序列同一性的核酸或多肽。
在本申请中，术语“Toll样受体8”和“TLR8”通常是指与可公开获得的TLR7序列例如人类TLR8多肽的GenBank登录号AAZ95441、或鼠TLR8多肽的GenBank登录号AAK62677共有至少70％、80％、90％、95％、96％、97％、98％、99％或更多序列同一性的核酸或多肽。
在本申请中，术语“TLR9”或“Toll样受体9”(也称为CD289、UNQ5798或PRO19605)通常是指通过非甲基化胞嘧啶-磷酸-鸟嘌呤(CpG)二核苷酸激活的核苷酸敏感TLR。TLR9的示例包括但不限于人TLR9，即由3922个核苷酸长的mRNA转录物(NM_017442.3)编码的1032个氨基酸长的蛋白质。所例示的人TLR9的氨基酸序列以GenBank登录号NP_059138.1表示。在本申请中，术语“TLR9”包括来自人以外的物种，诸如Macaca Fascicularis(食蟹猴)或Pantroglodytes(黑猩猩)的TLR9同源物。术语“TLR9”包括包含全长野生型TLR9的突变，例如点突变、片段、插入、缺失和剪接变体的蛋白质。术语“TLR9”还涵盖TLR9氨基酸序列的翻译后修饰。
在本申请中，术语“激动剂”通常是指分子(即，调节剂)，其直接或间接调节其它分子(例如，TLR)并且增加其它分子的活性、激活或功能。激动剂可以包括蛋白质、核酸、糖类、有机分子、小有机分子(具有或没有有机部分)或其他分子。例如，增强蛋白质的基因转录、生物 活性或者生物化学功能的调节剂是增强所述蛋白质的转录或刺激其生物化学性质或活性的物质。例如，激动剂可诱导、刺激、增大、激活、促进、增强、或上调受体的活性，此类活性称为“激动活性”。
在本申请中，“TLR激动剂”是直接或间接结合TLR(例如TLR7和/或TLR8)以诱导TLR信号传导的物质。TLR信号传导中的任何可检测出的差异可以表明激动剂刺激或活化TLR。信号传导差异可以表现为，例如，靶基因表达的变化、信号转导组分的磷酸化的变化、下游元件如核因子-κB(NF-κB)的细胞内定位的变化、某些组分(如IL-1受体相关激酶(IRAK))与其它蛋白或细胞内结构的缔合的变化、或组分如激酶(如促分裂原活化蛋白激酶(MAPK))的生化活性的变化。在本申请中，术语“TLR激动剂”是指充当TLR的激动剂的任何化合物。
在本申请中，术语“衍生物”通常是指与一种化学物质(chemical substance)在结构上相关的另一种化学物质，或可由另一种化学物质(即，衍生出该化学物质的化学物质)制备的一种化学物质，例如通过化学或酶的修饰。有机分子的衍生物包括但不限于修饰的分子，例如通过添加或删除羟基、甲基、乙基、羧基、硝基或氨基而修饰的分子。例如，有机分子也可以被酯化、烷基化和/或磷酸化。
在本申请中，术语“药学上可接受的衍生物”包括异构体、盐、酯、烯醇醚、烯醇酯、缩醛、缩酮、原酸酯、半缩醛、半缩酮、酸、碱、溶剂化物、水合物或其前药。本领域技术人员可以使用用于此类衍生化的已知方法容易地制得此类衍生物。
在本申请中，术语“异构体”通常是指具有相同分子式但是在原子排列和构型方面不同的不同化合物。术语“异构体”包括，但不局限于光学异构体和类似物、结构异构体和类似物、构象异构体和类似物等。
在本申请中，术语“类似物”通常是指其如本领域公认的意义。在非蛋白类似物的上下文中，术语“类似物”通常是指具有与第一有机或无机分子类似或相同的功能，并且结构上与该第一有机或无机分子类似的第二有机或无机分子。在提及多肽或蛋白质时，术语“类似物”通常指其中肽/蛋白质的一个或多个氨基酸残基已经被其它氨基酸残基替换和/或其中一个或多个氨基酸残基已经从肽/蛋白质中删除和/或其中已经在肽/蛋白质中添加一个或多个氨基酸残基的经修饰的肽或蛋白质。
在本申请中，术语“药学上可接受的盐”通常是指化合物的药学上可接受的有机盐或无机盐。实例性盐包括但不限于硫酸盐、柠檬酸盐、乙酸盐、草酸盐、氯化物、溴化物、碘化物、硝酸盐、硫酸氢盐、磷酸盐、酸式磷酸盐、异烟酸盐、乳酸盐、水杨酸盐、酸式柠檬酸盐、酒石酸盐、油酸盐、丹宁酸盐、泛酸盐、酒石酸氢盐、抗坏血酸盐、琥珀酸盐、马来酸盐、龙胆 酸盐、富马酸盐、葡萄糖酸盐、葡萄糖醛酸盐、糖酸盐、甲酸盐、苯甲酸盐、谷氨酸盐、甲磺酸盐、乙磺酸盐、苯磺酸盐、对甲苯磺酸盐、双羟萘酸盐(即1,1'-亚甲基-双-(2-羟基-3-萘甲酸)盐)、碱金属(例如钠和钾)盐、碱土金属(例如镁)盐和铵盐。药学上可接受的盐可涉及包含另一种分子，该分子例如乙酸根离子、琥珀酸根离子或其它抗衡离子。该抗衡离子可为任何有机或无机部分，其稳定所述母体化合物上的电荷。此外，药学上可接受的盐在其结构中可具有多于一个带电的原子。在多个带电原子为药学上可接受的盐的一部分的实例中，该盐可以具有多个抗衡离子。因此，药学上可接受的盐可具有一个或多个带电原子和/或一个或多个抗衡离子。
在本申请中，术语“抗体”通常是指在最广泛的意义上加以使用并且具体地涵盖单克隆抗体、多克隆抗体、二聚体、多聚体、多特异性抗体(例如，双特异性抗体)、和抗体片段，只要它们显示所期望的生物活性(Milleretal(2003)Jour.ofImmunology170:4854-4861)。抗体可以是鼠、人、人源化、嵌合抗体，或源于其它物种。
全长抗体典型地是指由两条“全长抗体重链”和两条“全长抗体轻链”组成的抗体。“全长抗体重链”通常是这样的多肽，其在N端到C端方向由抗体重链可变结构域(VH)、抗体恒定重链结构域1(CH1)，抗体铰链区(HR)，抗体重链恒定结构域2(CH2)，和抗体重链恒定结构域3(CH3)组成，缩写为VH-CH1-HR-CH2-CH3；并且在IgE亚类的抗体的情形中，任选地还包括抗体重链恒定结构域4(CH4)。在一些实施方式中，“全长抗体重链”是在N端到C端方向由VH，CH1，HR，CH2和CH3组成的多肽。“全长抗体轻链”通常是在N端到C端方向由抗体轻链可变结构域(VL)，和抗体轻链恒定结构域(CL)组成的多肽，缩写为VL-CL。所述抗体轻链恒定结构域(CL)可以是κ(kappa)或λ(lambda)。两条全长抗体链通过在CL结构域和CH1结构域之间的多肽间二硫键和全长抗体重链的铰链区之间的多肽间二硫键连接在一起。典型的全长抗体的实例是天然抗体如IgG(例如，IgG1和IgG2)，IgM，IgA，IgD，和IgE)。
在本申请中，术语“抗原结合片段”通常是指抗体分子的一部分，其包含负责抗体与抗原之间的特异性结合的氨基酸。抗原中由抗体特异性地识别和结合的部分是称作如上文所述的“表位”。抗原结合结构域可典型地包含抗体轻链可变区(VL)和抗体重链可变区(VH)；然而，其并非必须包含两者。Fd片段例如具有两个VH区并且通常保留完整抗原结合结构域的一些抗原结合功能。抗体的抗原结合片段的实例包括(1)Fab片段，具有VL、VH、恒定轻链(CL)和CH1结构域的单价片段；(2)F(ab’) 2片段，具有由铰链区的二硫桥连接的两个Fab片段的二价片段；(3)具有两个VH和CH1结构域的Fd片段；(4)具有抗体单臂的VL和VH结构域的 Fv片段，(5)dAb片段(Ward等人,“Binding Activities of a Repertoire of Single Immunoglobulin Variable Domains Secreted From Escherichia coli,”Nature 341:544-546(1989)，其以引用的方式整体并入本申请)，其具有VH结构域；(6)分离的互补决定区(CDR)；(7)单链Fv(scFv)，例如源于scFV-文库。尽管Fv片段的两个结构域VL和VH是由独立基因编码，但其可通过合成连接子使用重组方法接合，合成连接子使得其被制备为其中VL和VH区配对以形成单价分子的单一蛋白链(称为单链Fv(scFv))(可参见例如Huston等人,“Protein Engineering of Antibody Binding Sites:Recovery of Specific Activity in an Anti-Digoxin Single-Chain Fv Analogue Produced in Escherichia coli,”Proc.Natl.Acad.Sci.USA 85:5879-5883(1988))；和(8)VHH，“VHH”涉及来自骆驼科(骆驼、单峰骆驼、美洲驼、羊驼等)重链抗体的可变抗原结合结构域(参见Nguyen V.K.等人，2000，The EMBO Journal，19，921-930；Muyldermans S.，2001，J Biotechnol.，74，277-302以及综述Vanlandschoot P.等人，2011，Antiviral Research 92，389-407)。VHH也可称为纳米抗体(Nanobody)(Nb)和/或单域抗体。这些抗体片段使用所属领域的技术人员已知的常规技术获得，且以与完整抗体相同的方式评估所述片段的功能。
在本申请中，术语“可变区”或“可变结构域”通常指在抗体之间可变结构域的某些区段在序列上可能存在较大差异的区域。轻链中的“可变区”可以包含轻链可变区VL；重链中的“可变区”可以包含重链可变区VH。可变结构域介导抗原结合并决定特定抗体对其特定抗原的特异性。然而，可变性并非在整个可变结构域范围内均匀分布。它通常集中在轻链和重链可变结构域中称为高变区(CDR或HVR)的三个区段中。可变结构域的更高度保守的部分称为框架区(FR)。天然重链和轻链的可变结构域各自包含四个FR区，大多数采用β-折叠构型，通过三个CDR连接，其形成环形连接，并且在一些情况下形成β-折叠结构的一部分。每条链中的CDR通过FR区紧密靠近地保持在一起，并且来自另一条链的CDR一同促进抗体的抗原结合位点的形成(参见Kabat et al,Sequences of Immunological Interest,Fifth Edition,National Institute of Health,Bethesda,Md.(1991))。术语“VH”和“VH结构域”可互换使用以指抗体或其抗原结合分子的重链可变区。
在本申请中，术语“CDR”通常是指抗体可变序列内的互补性决定区。在重链和轻链的每个可变区中存在3个CDR，对于每个可变区，其称为CDR1、CDR2和CDR3。这些CDR的精确界限已经根据不同系统进行了不同定义。Kabat描述的系统(Kabat等人，Sequences of Proteins of Immunological Interest(National Institutes ofHealth，Bethesda，Md.(1987)和(1991))不仅提供了适用于抗体的任何可变区的明确的残基编号系统，而且还提供了定义这三个CDR的精确残基界限。这些CDR可称为Kabat CDR。Chothia及其同事(Chothia&Lesk， J.MoI.Biol.196:901-917(1987)和Chothia等人，Nature 342:877-883(1989))发现，Kabat CDR内的一些亚部分几乎采取了相同的肽骨架构象，尽管在氨基酸序列水平上具有很大差异。这些亚部分指定为L1、L2和L3，或者H1、H2和H3，其中“L”和“H”分别是指轻链和重链区域。这些区域可以称为Chothia CDR，其具有与Kabat CDR重叠的界限。Padlan(FASEB J.9:133-139(1995))和MacCallum(JMoI Biol 262(5):732-45(1996))中已经描述了定义与Kabat CDR重叠的CDR的其他界限。其他CDR界限可能没有严格遵照一个上述系统，但是仍然与Kabat CDR重叠，尽管根据以下预测或实验发现，他们可以被缩短或延长，特定残基或残基组或甚至整个CDR没有显著影响抗原结合。除非在说明书中另有明确说明，如本申请所用，术语“CDR”、“HCDR1”、“HCDR2”、“HCDR3”、“LCDR1”、“LCDR2”和“LCDR3”包括上文所述的任何方法(Kabat、Chothia或IMGT)所定义的CDR。
在本申请中，术语“百分比(％)序列同一性”通常是指两个或更多个经比对的氨基酸序列与组成这些氨基酸序列的总长度的氨基酸残基数相比而言一致的氨基酸的匹配(“命中”)数。换言之，使用比对，对于两个或更多个序列，当将这些序列针对最大对应(如使用本领域已知的序列比较算法测量的)进行比较和比对时，或者当手动比对和视觉检查时，可以确定相同的氨基酸残基的百分比(例如70％、75％、80％、85％、90％、95％、96％、97％、98％或99％序列同一性)。因此，进行比较以确定序列一致性的序列可以通过一个或多个氨基酸取代、添加或缺失来区分。用于比对蛋白序列的适合程序是本领域技术人员已知的。蛋白序列的百分比序列一致性可以例如用程序如CLUSTALW、Clustal Omega、FASTA或BLAST来确定，例如使用NCBI BLAST算法(AltschulSF等人(1997),Nucleic Acids Res.[核酸研究]25:3389-3402)。
在本申请中，术语“抗体类似物”通常以最广意义使用并且特别地涵盖以单特异性特异地结合靶分子并且结构上与天然抗体不同的分子。例如，在描述抗PD-1抗体或抗PD-L1抗体的上下文中，术语“抗体类似物”是指包含与氨基酸序列的部分具有大体上的同一性的区段并且具有至少一个下列性质的抗体：(1)在适当的结合条件下特异性结合PD-1或PD-L1，(2)抑制PD-1或PD-L1的至少一个生物活性的能力。通常，抗体类似物包含相对于天然序列的保守氨基酸置换(或插入或缺失)。类似物通常为至少20或25个氨基酸长，至少50、60、70、80、90、100、150或200个氨基酸长或更长，并且通常可与抗体的全长重链或轻链一样长。一些例子包括与种系氨基酸序列相比具有1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16或17个置换的抗体类似物。
在本申请中，术语“药物组合”通常是指至少包含两种活性成分/治疗剂的组合。在一些实施方式中，各个活性成分/治疗剂可以各自制备成独立的制剂(固体、液体、凝胶体等)，在一 些实施方式中，各个活性成分/治疗剂可以存在于不同的容器中，还可以在需要的时候同时或分别与合适的载体配制成期望的制剂；在一些实施方式中，各个活性成分/治疗剂可以是不同来源的(例如不同的商家制备生产或销售的)；在一些实施方式中，各个活性成分/治疗剂可以以混合的形式形成药物组合物。
在本申请中，术语“药物组合物”通常是指一种制剂，该制剂处于允许活性成分的生物活性有效的形式并且不含有对该组合物将要给予的受试者具有不可接受的毒性的另外的组分。这种组合物可以是无菌的，并且可以包含药学上可接受的载体，例如生理盐水。合适的药物组合物可以包含一种或多种缓冲液(例如乙酸盐、磷酸盐或柠檬酸盐缓冲液)、表面活性剂(例如聚山梨酯)、稳定剂(例如人类白蛋白)、防腐剂(例如苯甲醇)、用于增强生物利用度的吸收促进剂、和/或其他常规的增溶剂或分散剂。本申请的药物组合物包括但不限于液体、冷冻和冻干组合物。
在本申请中，术语“药学上可接受的载体”通常是指不干扰活性成分的生物活性的有效性的一种或多种非毒性材料。这类制剂常规地可以含有盐、缓冲剂、防腐剂、相容的载体、以及任选地其他治疗剂。这类药学上可接受的制剂还可以含有适合于给予人的相容的固体或液体填料、稀释剂或包封物质。可以用于在此所描述的配制品中的其他设想的载体、赋形剂、和/或添加剂可以包括：例如，调味剂、抗微生物剂、增甜剂、抗氧化剂、抗静电剂、脂质、蛋白质赋形剂(如血清白蛋白、明胶、酪蛋白)、成盐平衡离子(如钠)等等。适合用于在此所描述的配制品中的这些和另外已知的药物载体、赋形剂和/或添加剂是本领域中已知的。
在本申请中，术语“赘生性”通常表示进行新的和非正常增殖的细胞，特别是其中增殖不受控制和进展，导致赘生物的疾病。赘生性细胞可以是恶性的，即侵袭性的和转移性的，或良性的。
在本申请中，术语“赘生物”通常是指组织的异常肿块，其中肿块的生长超过正常组织的生长，且不与正常组织的生长协调。“赘生物”可被定义为“良性”或“恶性”，其取决于以下特征：细胞分化的程度，包括形态和功能，生长的速率，局部侵袭和转移。“良性赘生物”通常是分化良好的，具有比恶性赘生物特征性更慢的生长，并且保持定位至来源部位。此外，良性肿瘤不具有渗透、侵袭或转移至远端部位的能力。“恶性赘生物”通常是低分化的(退行发育)，具有特征性快速生长，伴随周围组织的进行性渗透、侵袭和破坏。此外，恶性赘生物具有转移至远端部位的能力。
在本申请中，术语“肿瘤”或“癌症”通常是指以赘生性或恶性细胞生长、增殖或转移为特征的任何医学疾患，肿瘤可以是实体瘤或非实体瘤。
在本申请中，术语“疣”通常是指人类头瘤病毒(human papilloma Virus，HPV)引起的以细胞增生反应为主的一类皮肤浅表性良性赘生物。HPV属DNA病毒中乳多空病毒科A属，HPV分80余种亚型，与不同类型的疣有关。术语“疣(wart)”，除非另有说明，通常是指所有类型的疣，包括但不限于足底疣、普通疣、和生殖器疣。
在本申请中，术语“施用(administer)”和类似术语通常不限于身体施用，合适的方法包括体外、先体外后体内或体内方法。例如，可以采用本领域技术人员已知的用于使细胞、器官或组织与组合物接触的任何施用方法。例如，可以通过任意引入或递送途径将所述化合物引入需要治疗的受试者的身体中。在一些实施方案中，本申请的组合物可以口服、局部、鼻内、肌内、皮下、皮内、鞘内、腹膜内、经皮或瘤内施用。
在本申请中，术语“有效量”或“有效剂量”通常是指足以实现或至少部分实现所需效果的量。药物或治疗剂的“治疗有效量”或“治疗有效剂量”通常是当单独使用或与另一种治疗剂组合使用时促进疾病消退(这通过疾病症状严重程度的降低、疾病无症状期的频度和持续时间的增加、或者由于罹患疾病而引起的损害或残疾的预防来证明)的任何药物量。
在本申请中，术语“治疗”通常是指由于施用一种或多种疗法(例如一种或多种治疗剂比如本申请的药物组合物)减慢或改善增生性病症的进展、严重程度和/或持续时间，或改善增生性病症的一种或多种症状(例如，一种或多种可辨别的症状)。在本申请中，术语“治疗”还可指改善增生性病症的至少一种可测量的物理参数比如肿瘤生长，不必是患者可辨别的。本申请中的术语“治疗”还可指通过例如稳定可辨别的症状以物理方式、通过例如稳定物理参数以生理学方式或两者抑制增生性病症的进展。在某些情形中，术语“治疗”可以指减小或稳定肿瘤尺寸或癌细胞计数。
在本申请中，术语“协同”通常是指两种或多种活性药物的联用的效力大于各活性药物单用之和。因此，当两种或多种药物的联用导致某活性或过程如肿瘤生长的“协同抑制”时，则指对该活性或过程的抑制作用大于各活性药物的抑制作用之和。
在本申请中，术语“受试者”通常是指人类或非人类动物，包括但不限于猫、狗、马、猪、奶牛、羊、兔、小鼠、大鼠或猴等。
在本申请中，术语“约”通常是指在指定数值以上或以下0.5％-10％的范围内变动，例如在指定数值以上或以下约0.5％、约1％、约1.5％、约2％、约2.5％、约3％、约3.5％、约4％、约4.5％、约5％、约5.5％、约6％、约6.5％、约7％、约7.5％、约8％、约8.5％、约9％、约9.5％、或约10％的范围内变动。
在本申请中，术语“包含”和其变形形式，包括“含有”、“包括”等其它形式，通常是指包含 其它组分、元素、数值、步骤等。
发明详述
本申请提供一种诱导INF和细胞因子的表达，激活固有免疫系统反应的激动剂与免疫检查点抑制剂作为药物组合使用，包含TLR(例如，TLR7/TLR8)激动剂和免疫检查点(例如，PD-1或PD-L1)抑制剂，等等。
一方面，本申请提供一种药物组合，其可以包含程序性细胞死亡蛋白1(PD-1)抑制剂和/或程序性死亡配体1(PD-L1)抑制剂，以及TLR激动剂。
在某些实施方式中，其中所述TLR激动剂包括TLR1，TLR2，TLR3，TLR4，TLR5，TLR6，TLR7，TLR8，TLR9和/或TLR10。
在某些实施方式中，其中所述TLR激动剂选自：TLR7激动剂，TLR8激动剂，TLR9激动剂中的一种或多种。
在某些实施方式中，其中所述TLR激动剂包括TLR7和TLR8双重激动剂(TLR7/TLR8激动剂)。
在某些实施方式中，其中所述TLR激动剂包括咪唑喹啉衍生物。
在某些实施方式中，其中所述TLR激动剂选自咪喹莫特(Imiquimod)、嘎德莫特(Gardiquimod)、瑞喹莫特(Resiquimod)、1V209、Selgantolimod(GS-9688)、Vesatolimod(GS-9620)、Sumanirole、PF-4878691、及其药学上可接受的衍生物中的一种或多种。
在某些实施方式中，其中所述TLR激动剂包括咪喹莫特、瑞喹莫特或其药学上可接受的盐。
在某些实施方式中，其中所述TLR激动剂选自LHC-165、NKTR-262、DN1508052-01、SHR2150、CL307、CL264、洛索立宾、艾沙托立宾、DSR-6434、GSK2245035、SM-276001、SM-324405、SM-324406、AZ12441970、AZ12443988及其药学上可接受的衍生物中的一种或多种。
在某些实施方式中，其中所述PD-1抑制剂具有以下一个或多个特征：
a.抑制或减少PD-1表达，例如PD-1的转录或翻译；
b.抑制或降低PD-1活性，例如抑制或降低PD-1与其同源配体，例如PD-L1或PD-L2的结合；和
c.结合PD-1或其一个或多个配体，例如PD-L1或PD-L2。
在某些实施方式中，其中所述PD-1抑制剂包括抗PD-1抗体或其抗原结合片段。
例如，所述药物组合可以包含：1)抗PD-1抗体或其抗原结合片段；和2)TLR7和/或 TLR8激动剂。
又例如，例如，所述药物组合可以包含：1)抗PD-1抗体或其抗原结合片段；和2)咪唑喹啉衍生物。
在某些实施方式中，其中所述抗PD-1抗体选自Pembrolizumab(派姆单抗)，Nivolumab(纳武单抗)，Pidilizumab，Tislelizumab(替雷利珠单抗)，SHR-1210(Incyte/江苏恒瑞医药有限公司)，MEDI0680(也称为AMP-514；Amplimmune Inc./Medimmune)，BGB-A317(BeiGene Ltd.)，TSR-042(也称为ANB011；AnaptysBio/Tesaro，Inc.)，REGN2810(Regeneron Pharmaceuticals，Inc./Sanofi-Aventis)、PF-06801591(Pfizer)、RB0004、其类似物及它们的组合。
例如，所述药物组合可以包含：1)抗PD-1抗体，所述抗PD-1抗体可以选自Pembrolizumab(派姆单抗)，Nivolumab(纳武单抗)，Pidilizumab，Tislelizumab，SHR-1210(Incyte/江苏恒瑞医药有限公司)，MEDI0680(也称为AMP-514；Amplimmune Inc./Medimmune)，BGB-A317(BeiGene Ltd.)，TSR-042(也称为ANB011；AnaptysBio/Tesaro，Inc.)，REGN2810(Regeneron Pharmaceuticals，Inc./Sanofi-Aventis)、PF-06801591(Pfizer)、RB0004、其类似物及它们的组合；和2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以选自Imiquimod、Gardiquimod、Resiquimod、其衍生物及它们的组合。
在一些实施方案中，抗PD-1抗体是RB0004。RB0004和其它人源化抗PD-1单克隆抗体公开于CN201610345750.1，WO2017201766A1。例如，所述药物组合其包含：1)抗PD-1抗体，所述抗PD-1抗体是RB0004或其类似物及其组合；和2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以是Imiquimod或其衍生物及它们的组合。
在另一些实施方式中，所述抗体是PD-1是Pembrolizumab(商品名KEYTRUDA，以前的兰布罗珠单抗(Lambrolizumab)，也称为Merck 3745、MK-3475或SCH-900475)是与PD-1结合的人源化IgG4单克隆抗体。Pembrolizumab公开于例如Hamid，等人(2013)New England Journal of Medicine 369(2):134-44，W02009/114335和US 8,354,509。例如，所述药物组合其包含：1)抗PD-1抗体，所述抗PD-1抗体可以是Pembrolizumab或其其生物增强物，和其生物等效物、及其组合；和2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以是Imiquimod或其衍生物及它们的组合。
在另一些实施方案中，所述抗PD-1抗体是Nivolumab(CAS登记号：946414-94-4，替代名称包括MDX-1106、MDX-1106-04、ONO-4538或BMS-936558)。Nivolumab是完全人IgG4单克隆抗体，其特异性阻断PD-1。US8,008,449和W02006/121168中公开了Nivolumab(克隆 5C4)和其它特异性结合PD-1的人单克隆抗体。例如，所述药物组合其包含：1)抗PD-1抗体，所述抗PD-1抗体可以是Nivolumab或其类似物及其组合；和2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以是Imiquimod或其衍生物及它们的组合。
在另一些实施方案中，抗PD-1抗体是Pidilizumab。Pidilizumab(CT-011；Cure Tech)是与PD-1结合的人源化IgGl单克隆抗体。Pidilizumab和其它人源化抗PD-1单克隆抗体公开于W02009/101611中。US 8,609,089、US2010028330和/或US20120114649中公开了其它抗Rpd-1抗体。其它抗PD-1抗体包括AMP514(Amplimmune)。例如，所述药物组合其包含：1)抗PD-1抗体，所述抗PD-1抗体可以是Pidilizumab或类似物及其组合；和2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以是Imiquimod或其衍生物及它们的组合。
在某些实施方式中，其中所述抗PD-1抗体包含抗体重链可变区(VH)中的至少一个CDR，所述VH包含SEQ ID NO:8所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含VH，所述VH包含HCDR3，所述HCDR3包含SEQ ID NO:3所示的氨基酸序列或与SEQ ID NO:3所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH还包含HCDR2，其中所述HCDR2包含SEQ ID NO:2所示的氨基酸序列或与SEQ ID NO:2所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH还包含HCDR1，其中所述HCDR1包含SEQ ID NO:1所示的氨基酸序列或与SEQ ID NO:1所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:3所示的氨基酸序列或与SEQ ID NO:3所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HCDR2包含SEQ ID NO:2所示的氨基酸序列或与SEQ ID NO:2所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HCDR1包含SEQ ID NO:1所示的氨基酸序列或与 SEQ ID NO:1所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR1，所述HFR1的C末端与所述HCDR1的N末端直接或间接相连，且所述HFR1包含SEQ ID NO:4所示的氨基酸序列或与SEQ ID NO:4所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR2，所述HFR2的N末端与所述HCDR1的C末端直接或间接相连，且所述HFR2的C末端与所述HCDR2的N末端直接或间接相连；且所述HFR2包含SEQ ID NO:5所示的氨基酸序列或与SEQ ID NO:5所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR3，所述HFR3的N末端与所述HCDR2的C末端直接或间接相连，且所述HFR3的C末端与所述HCDR3的N末端直接或间接相连；且所述HFR3包含SEQ ID NO:6所示的氨基酸序列或与SEQ ID NO:6所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR4，所述HFR4的N末端与所述HCDR3的C末端直接或间接相连，且所述HFR4包含SEQ ID NO:7所示的氨基酸序列或与SEQ ID NO:7所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR1，HFR2，HFR3和HFR4，所述HFR1的C末端与所述HCDR1的N末端直接或间接相连，所述HFR2的N末端与所述HCDR1的C末端直接或间接相连，且所述HFR2的C末端与所述HCDR2的N末端直接或间接相连，所述HFR3的N末端与所述HCDR2的C末端直接或间接相连，且所述HFR3的C末端与所述HCDR3的N末端直接或间接相连，所述HFR4的N末端与所述HCDR3的C末端直接或间接相连；其中，所述HFR1包含SEQ ID NO:4所示的氨基酸序列或与SEQ ID NO:4所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约 93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HFR2包含SEQ ID NO:5所示的氨基酸序列或与SEQ ID NO:5所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HFR3包含SEQ ID NO:6所示的氨基酸序列或与SEQ ID NO:6所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HFR4包含SEQ ID NO:7所示的氨基酸序列或与SEQ ID NO:7所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含VH，所述VH包含SEQ ID NO:8所示的氨基酸序列或与SEQ ID NO:8所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含抗体重链(HC)，所述HC包含SEQ ID NO:9所示氨基酸序列或与SEQ ID NO:9所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含抗体轻链可变区(VL)中的至少一个CDR，所述VL包含SEQ ID NO:17所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含VH中的至少一个CDR，所述VH包含SEQ ID NO:8所示的氨基酸序列，且所述抗PD-1抗体包含VL中的至少一个CDR，所述VL包含SEQ ID NO:17所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含VL，所述VL包含LCDR1，所述LCDR1包含SEQ ID NO:10所示的氨基酸序列或与SEQ ID NO:10所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL还包含LCDR2，其中所述LCDR2包含SEQ ID NO:11所示的氨基酸序列或与SEQ ID NO:11所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、 或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL还包含LCDR3，其中所述LCDR3包含SEQ ID NO:12所示的氨基酸序列或与SEQ ID NO:12所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:10所示的氨基酸序列或与SEQ ID NO:10所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR2包含SEQ ID NO:11所示的氨基酸序列或与SEQ ID NO:11所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR3包含SEQ ID NO:12所示的氨基酸序列或与SEQ ID NO:12所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含VH以及抗体VL，所述VH包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:3所示的氨基酸序列或与SEQ ID NO:3所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HCDR2包含SEQ ID NO:2所示的氨基酸序列或与SEQ ID NO:2所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HCDR1包含SEQ ID NO:1所示的氨基酸序列或与SEQ ID NO:1所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列；且所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:10所示的氨基酸序列或与SEQ ID NO:10所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR2包含SEQ ID NO:11所示的氨基酸序列或与SEQ ID NO:11所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约 96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR3包含SEQ ID NO:12所示的氨基酸序列或与SEQ ID NO:12所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
例如，所述抗PD-1抗体可以包含VH以及抗体VL，所述VH可以包含HCDR1，HCDR2和HCDR3，其中所述HCDR3可以包含SEQ ID NO:3所示的氨基酸序列，所述HCDR2可以包含SEQ ID NO:2所示的氨基酸序列，所述HCDR1可以包含SEQ ID NO:1所示的氨基酸序列；且所述VL可以包含LCDR1，LCDR2和LCDR3，其中所述LCDR1可以包含SEQ ID NO:10所示的氨基酸序列，所述LCDR2可以包含SEQ ID NO:11所示的氨基酸序列，所述LCDR3可以包含SEQ ID NO:12所示的氨基酸序列。
又例如，所述药物组合可以包含：1)抗PD-1抗体，所述抗PD-1抗体可以包含VH以及抗体VL，所述VH可以包含HCDR1，HCDR2和HCDR3，其中所述HCDR3可以包含SEQ ID NO:3所示的氨基酸序列，所述HCDR2可以包含SEQ ID NO:2所示的氨基酸序列，所述HCDR1可以包含SEQ ID NO:1所示的氨基酸序列；且所述VL可以包含LCDR1，LCDR2和LCDR3，其中所述LCDR1可以包含SEQ ID NO:10所示的氨基酸序列，所述LCDR2可以包含SEQ ID NO:11所示的氨基酸序列，所述LCDR3可以包含SEQ ID NO:12所示的氨基酸序列；2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以是Imiquimod或其衍生物及它们的组合。
在某些实施方式中，其中所述VL包括框架区LFR1，所述LFR1的C末端与所述LCDR1的N末端直接或间接相连，且所述LFR1包含SEQ ID NO:13所示的氨基酸序列或与SEQ ID NO:13所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR2，所述LFR2的N末端与所述LCDR1的C末端直接或间接相连，且所述LFR2的C末端与所述LCDR2的N末端直接或间接相连；且所述LFR2包含SEQ ID NO:14所示的氨基酸序列或与SEQ ID NO:14所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR3，所述LFR3的N末端与所述LCDR2的C末端直接或间接相连，且所述LFR3的C末端与所述LCDR3的N末端直接或间接相连；且所述LFR3包含SEQ ID NO:15所示的氨基酸序列或与SEQ ID NO:15所示的氨基酸序列 具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR4，所述LFR4的N末端与所述LCDR3的C末端直接或间接相连，且所述LFR4包含SEQ ID NO:16所示的氨基酸序列或与SEQ ID NO:16所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR1，LFR2，LFR3和LFR4，所述LFR1的C末端与所述LCDR1的N末端直接或间接相连，所述LFR2的N末端与所述LCDR1的C末端直接或间接相连，且所述LFR2的C末端与所述LCDR2的N末端直接或间接相连，所述LFR3的N末端与所述LCDR2的C末端直接或间接相连，且所述LFR3的C末端与所述LCDR3的N末端直接或间接相连，所述LFR4的N末端与所述LCDR3的C末端直接或间接相连；其中，所述LFR1包含SEQ ID NO:13所示的氨基酸序列或与SEQ ID NO:13所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LFR2包含SEQ ID NO:14所示的氨基酸序列或与SEQ ID NO:14所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LFR3包含SEQ ID NO:15所示的氨基酸序列或与SEQ ID NO:15所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LFR4包含SEQ ID NO:16所示的氨基酸序列或与SEQ ID NO:16所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含VL，所述VL包含SEQ ID NO:17所示的氨基酸序列或与SEQ ID NO:17所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含VH以及VL，所述VH包含SEQ ID NO:8所示的氨基酸序列或与SEQ ID NO:8所示的氨基酸序列具有至少约70％、约75％、约80％、 约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，且所述VL包含SEQ ID NO:17所示的氨基酸序列或与SEQ ID NO:17所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
例如，其中所述抗PD-1抗体可以包含VH以及VL，所述VH可以包含SEQ ID NO:8所示的氨基酸序列，且所述VL可以包含SEQ ID NO:17所示的氨基酸序列。
又例如，所述药物组合可以包含：1)抗PD-1抗体，其中所述抗PD-1抗体可以包含VH以及VL，所述VH可以包含SEQ ID NO:8所示的氨基酸序列，且所述VL可以包含SEQ ID NO:17所示的氨基酸序列；2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以是Imiquimod或其衍生物及它们的组合。
在某些实施方式中，其中所述抗PD-1抗体包含抗体轻链(LC)，所述LC包含SEQ ID NO:18所示氨基酸序列或与SEQ ID NO:18所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-1抗体包含HC和LC，所述HC包含SEQ ID NO:9所示氨基酸序列或与SEQ ID NO:9所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，且所述LC包含如SEQ ID NO:18所示氨基酸序列或与SEQ ID NO:18所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
例如，其中所述抗PD-1抗体可以包含HC和LC，所述HC可以包含SEQ ID NO:9所示氨基酸序列，且所述LC可以包含如SEQ ID NO:18所示氨基酸序列。
又例如，所述药物组合可以包含：1)抗PD-1抗体，其中所述抗PD-1抗体可以包含HC和LC，所述HC可以包含SEQ ID NO:9所示氨基酸序列，且所述LC可以包含如SEQ ID NO:18所示氨基酸序列；2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以是Imiquimod或其衍生物及它们的组合。
在某些实施方式中，其中所述PD-L1抑制剂具有以下一个或多个特征：
a.抑制或减少PD-L1表达，例如PD-L1的转录或翻译；
b.抑制或降低PD-L1活性，例如抑制或降低PD-L1与其关联受体，例如PD-1的结合；和
c.结合PD-L1或其受体，例如PD-1。
在某些实施方式中，其中所述PD-L1抑制剂包括抗PD-L1抗体或其抗原结合片段。
例如，所述药物组合可以包含：1)抗PD-L1抗体或其抗原结合片段；和2)TLR7和/或TLR8激动剂。
又例如，所述药物组合可以包含：1)抗PD-L1抗体或其抗原结合片段；和2)咪唑喹啉衍生物。
在某些实施方式中，其中所述抗PD-L1抗体选自Durvalumab(MEDI4736，公开于US2013/0034559A1)，Atezolizumab(MPDL3280A，公开于US8,217,149)，Avelumab(MSB0010718C，公开于US2014/0341917A1)，MDX-1105，YW243.55.S70，MDPL3280A，AMP-224(Amplimmune,GlaxoSmithKline)、LY3300054(Eli Lilly and Co.)、RB0005、其类似物及它们的组合。
例如，所述药物组合可以包含：1)抗PD-L1抗体，其中所述抗PD-L1抗体选自Durvalumab(MEDI4736，公开于US2013/0034559A1)，Atezolizumab(MPDL3280A，公开于US8,217,149)，Avelumab(MSB0010718C，公开于US2014/0341917A1)，MDX-1105，YW243.55.S70，MDPL3280A，AMP-224、LY3300054、RB0005、其类似物及它们的组合；和2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以选自Imiquimod、Gardiquimod、Resiquimod、其衍生物及它们的组合。
在一些实施方案中，抗PD-L1抗体是RB0005。RB0005和其它人源化抗PD-L1单克隆抗体公开于CN201610340678.3，WO2017197667A1。例如，所述药物组合其包含：1)抗PD-L1抗体，所述抗PD-1抗体是RB0005或类似物及其组合；和2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以是Imiquimod或其衍生物及它们的组合。
在另一些实施方案中，PD-Ll抑制剂是MDX-1105。MDX-1105也称为BMS-936559，是W02007/005874中描述的抗PD-Ll抗体。例如，所述药物组合其包含：1)抗PD-L1抗体，所述抗PD-1抗体是MDX-1105或其类似物及其组合；和2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以是Imiquimod或其衍生物及它们的组合。
在另一些实施方案中，PD-Ll抑制剂是YW243.55.S70。YW243.55.S70抗体是W02010/077634中描述的抗PD-L1抗体。
在另一些实施方案中，PD-Ll抑制剂是MDPL3280A(Genentech/Roche)。MDPL3280A是 与PD-Ll结合的人Fc优化IgGl单克隆抗体。美国专利号7,943,743和美国专利公开号20120039906中公开了MDPL3280A和PD-L1的其它人单克隆抗体。
在某些实施方式中，其中所述抗PD-L1抗体包含VH中的至少一个CDR，所述VH包含SEQ ID NO:25所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VH，所述VH包含HCDR3，所述HCDR3包含SEQ ID NO:21所示的氨基酸序列或与SEQ ID NO:21所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH还包含HCDR2，其中所述HCDR2包含SEQ ID NO:20所示的氨基酸序列或与SEQ ID NO:20所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH还包含HCDR1，其中所述HCDR1包含SEQ ID NO:19所示的氨基酸序列或与SEQ ID NO:19所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:21所示的氨基酸序列或与SEQ ID NO:21所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HCDR2包含SEQ ID NO:20所示的氨基酸序列或与SEQ ID NO:20所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HCDR1包含SEQ ID NO:19所示的氨基酸序列或与SEQ ID NO:19所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
例如，其中所述VH可以包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:21所示的氨基酸序列，所述HCDR2包含SEQ ID NO:20所示的氨基酸序列，所述HCDR1包含SEQ ID NO:19所示的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR1，所述HFR1的C末端与所述HCDR1 的N末端直接或间接相连，且所述HFR1包含SEQ ID NO:22所示的氨基酸序列或与SEQ ID NO:22所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR2，所述HFR2的N末端与所述HCDR1的C末端直接或间接相连，且所述HFR2的C末端与所述HCDR2的N末端直接或间接相连；且所述HFR2包含SEQ ID NO:23所示的氨基酸序列或与SEQ ID NO:23所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR3，所述HFR3的N末端与所述HCDR2的C末端直接或间接相连，且所述HFR3的C末端与所述HCDR3的N末端直接或间接相连；且所述HFR3包含SEQ ID NO:24所示的氨基酸序列或与SEQ ID NO:24所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR4，所述HFR4的N末端与所述HCDR3的C末端直接或间接相连，且所述HFR4包含SEQ ID NO:7所示的氨基酸序列或SEQ ID NO:7所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VH包括框架区HFR1，HFR2，HFR3和HFR4，所述HFR1的C末端与所述HCDR1的N末端直接或间接相连，所述HFR2的N末端与所述HCDR1的C末端直接或间接相连，且所述HFR2的C末端与所述HCDR2的N末端直接或间接相连，所述HFR3的N末端与所述HCDR2的C末端直接或间接相连，且所述HFR3的C末端与所述HCDR3的N末端直接或间接相连，所述HFR4的N末端与所述HCDR3的C末端直接或间接相连；其中，所述HFR1包含SEQ ID NO:22所示的氨基酸序列或与SEQ ID NO:22所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HFR2包含SEQ ID NO:23所示的氨基酸序列或与SEQ ID NO:23所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HFR3包含SEQ ID NO:24所示的氨基酸序列或与SEQ ID NO:24所示的氨基酸序列具有至少约70％、约75％、 约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HFR4包含SEQ ID NO:7所示的氨基酸序列或与SEQ ID NO:7所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VH，所述VH包含SEQ ID NO:25所示的氨基酸序列或与SEQ ID NO:25所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含HC，所述HC包含SEQ ID NO:26所示氨基酸序列或与SEQ ID NO:26所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VL中的至少一个CDR，所述VL包含SEQ ID NO:37所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VH中的至少一个CDR，所述VH包含SEQ ID NO:25所示的氨基酸序列，且所述抗PD-L1抗体包含VL中的至少一个CDR，所述VL包含SEQ ID NO:37所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VH中的至少一个CDR，所述VH包含SEQ ID NO:25所示的氨基酸序列，且所述抗PD-L1抗体包含VL中的至少一个CDR，所述VL包含SEQ ID NO:38、SEQ ID NO:39或SEQ ID NO:40所示的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VL，所述VL包含LCDR1，所述LCDR1包含SEQ ID NO:27所示的氨基酸序列或与SEQ ID NO:27所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VL，所述VL包含LCDR1，所述LCDR1包含SEQ ID NO:28、SEQ ID NO:29或SEQ ID NO:30所示的氨基酸序列或与SEQ ID NO:28、SEQ ID NO:29或SEQ ID NO:30所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL还包含LCDR2，其中所述LCDR2包含SEQ ID NO:31所示的氨基酸序列或与SEQ ID NO:31所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL还包含LCDR3，其中所述LCDR3包含SEQ ID NO:32所示的氨基酸序列或与SEQ ID NO:32所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:27所示的氨基酸序列或与SEQ ID NO:27所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列或与SEQ ID NO:31所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列或与SEQ ID NO:32所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:28所示的氨基酸序列或与SEQ ID NO:28所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列或与SEQ ID NO:31所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列或与SEQ ID NO:32所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列；
在某些实施方式中，其中所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:29所示的氨基酸序列或与SEQ ID NO:29所示的氨基酸序列具有至少约70％、 约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列或与SEQ ID NO:31所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列或与SEQ ID NO:32所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列；或
在某些实施方式中，其中所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:30所示的氨基酸序列或与SEQ ID NO:30所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列或与SEQ ID NO:31所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列或与SEQ ID NO:32所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VH以及抗体VL，所述VH包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:21所示的氨基酸序列或与SEQ ID NO:21所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HCDR2包含SEQ ID NO:20所示的氨基酸序列或与SEQ ID NO:20所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HCDR1包含SEQ ID NO:19所示的氨基酸序列或与SEQ ID NO:19所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列；且所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:27所示的氨基酸序列或与SEQ ID NO:27所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约 93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列或与SEQ ID NO:31所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列或与SEQ ID NO:32所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
例如，其中所述抗PD-L1抗体可以包含VH以及抗体VL，所述VH可以包含HCDR1，HCDR2和HCDR3，其中所述HCDR3可以包含SEQ ID NO:21所示的氨基酸序列，所述HCDR2可以包含SEQ ID NO:20所示的氨基酸序列，所述HCDR1可以包含SEQ ID NO:19所示的氨基酸序列；且所述VL可以包含LCDR1，LCDR2和LCDR3，其中所述LCDR1可以包含SEQ ID NO:27所示的氨基酸序列，所述LCDR2可以包含SEQ ID NO:31所示的氨基酸序列，所述LCDR3可以包含SEQ ID NO:32所示的氨基酸序列。
又例如，所述药物组合可以包含：1)抗PD-L1抗体，其中所述抗PD-L1抗体可以包含VH以及抗体VL，所述VH可以包含HCDR1，HCDR2和HCDR3，其中所述HCDR3可以包含SEQ ID NO:21所示的氨基酸序列，所述HCDR2可以包含SEQ ID NO:20所示的氨基酸序列，所述HCDR1可以包含SEQ ID NO:19所示的氨基酸序列；且所述VL可以包含LCDR1，LCDR2和LCDR3，其中所述LCDR1可以包含SEQ ID NO:27所示的氨基酸序列，所述LCDR2可以包含SEQ ID NO:31所示的氨基酸序列，所述LCDR3可以包含SEQ ID NO:32所示的氨基酸序列；2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以是Imiquimod或其衍生物及它们的组合。
在某些实施方式中，其中所述抗PD-L1抗体包含VH以及抗体VL，所述VH包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:21所示的氨基酸序列或与SEQ ID NO:21所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HCDR2包含SEQ ID NO:20所示的氨基酸序列或与SEQ ID NO:20所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述HCDR1包含SEQ ID NO:19所示的氨基酸序列或与SEQ ID NO:19所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约 96％、约97％、约98％、或约99％序列同一性的氨基酸序列；且所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:28、SEQ ID NO:29或SEQ ID NO:30所示的氨基酸序列或与SEQ ID NO:28、SEQ ID NO:29或SEQ ID NO:30所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列或与SEQ ID NO:31所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列或与SEQ ID NO:32所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
例如，其中所述抗PD-L1抗体可以包含VH以及抗体VL，所述VH可以包含HCDR1，HCDR2和HCDR3，其中所述HCDR3可以包含SEQ ID NO:21所示的氨基酸序列，所述HCDR2可以包含SEQ ID NO:20所示的氨基酸序列，所述HCDR1可以包含SEQ ID NO:19所示的氨基酸序列；且所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1可以包含SEQ ID NO:28、SEQ ID NO:29或SEQ ID NO:30所示的氨基酸序列，所述LCDR2可以包含SEQ ID NO:31所示的氨基酸序列，所述LCDR3可以包含SEQ ID NO:32所示的氨基酸序列。
又例如，所述药物组合可以包含：1)抗PD-L1抗体，其中所述抗PD-L1抗体包含VH以及抗体VL，所述VH可以包含HCDR1，HCDR2和HCDR3，其中所述HCDR3可以包含SEQ ID NO:21所示的氨基酸序列，所述HCDR2可以包含SEQ ID NO:20所示的氨基酸序列，所述HCDR1可以包含SEQ ID NO:19所示的氨基酸序列；且所述VL可以包含LCDR1，LCDR2和LCDR3，其中所述LCDR1可以包含SEQ ID NO:28、SEQ ID NO:29或SEQ ID NO:30所示的氨基酸序列，所述LCDR2可以包含SEQ ID NO:31所示的氨基酸序列，所述LCDR3可以包含SEQ ID NO:32所示的氨基酸序列；2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以是Imiquimod或其衍生物及它们的组合。
在某些实施方式中，其中所述VL包括框架区LFR1，所述LFR1的C末端与所述LCDR1的N末端直接或间接相连，且所述LFR1包含SEQ ID NO:33所示的氨基酸序列或与SEQ ID NO:33所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸 序列。
在某些实施方式中，其中所述VL包括框架区LFR2，所述LFR2的N末端与所述LCDR1的C末端直接或间接相连，且所述LFR2的C末端与所述LCDR2的N末端直接或间接相连；且所述LFR2包含SEQ ID NO:34所示的氨基酸序列或与SEQ ID NO:34所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR3，所述LFR3的N末端与所述LCDR2的C末端直接或间接相连，且所述LFR3的C末端与所述LCDR3的N末端直接或间接相连；且所述LFR3包含SEQ ID NO:35所示的氨基酸序列或与SEQ ID NO:35所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR4，所述LFR4的N末端与所述LCDR3的C末端直接或间接相连，且所述LFR4包含SEQ ID NO:36所示的氨基酸序列或与SEQ ID NO:36所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述VL包括框架区LFR1，LFR2，LFR3和LFR4，所述LFR1的C末端与所述LCDR1的N末端直接或间接相连，所述LFR2的N末端与所述LCDR1的C末端直接或间接相连，且所述LFR2的C末端与所述LCDR2的N末端直接或间接相连，所述LFR3的N末端与所述LCDR2的C末端直接或间接相连，且所述LFR3的C末端与所述LCDR3的N末端直接或间接相连，所述LFR4的N末端与所述LCDR3的C末端直接或间接相连；其中，所述LFR1包含SEQ ID NO:33所示的氨基酸序列或与SEQ ID NO:33所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LFR2包含SEQ ID NO:34所示的氨基酸序列或与SEQ ID NO:34所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LFR3包含SEQ ID NO:35所示的氨基酸序列或与SEQ ID NO:35所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，所述LFR4包含SEQ ID NO:36所示的氨基酸 序列或与SEQ ID NO:36所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VL，所述VL包含SEQ ID NO:37所示的氨基酸序列或与SEQ ID NO:37所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VL，所述VL包含SEQ ID NO:38、SEQ ID NO:39或SEQ ID NO:40所示的氨基酸序列或与SEQ ID NO:38、SEQ ID NO:39或SEQ ID NO:40所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含VH以及VL，所述VH包含SEQ ID NO:25所示的氨基酸序列或与SEQ ID NO:25所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，且所述VL包含SEQ ID NO:37所示的氨基酸序列或与SEQ ID NO:37所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
例如，其中所述抗PD-L1抗体可以包含VH以及VL，所述VH可以包含SEQ ID NO:25所示的氨基酸序列，且所述VL可以包含SEQ ID NO:37所示的氨基酸序列。
又例如，所述药物组合可以包含：1)抗PD-L1抗体，其中所述抗PD-L1抗体可以包含VH以及VL，所述VH可以包含SEQ ID NO:25所示的氨基酸序列，且所述VL可以包含SEQ ID NO:37所示的氨基酸序列；2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以是Imiquimod或其衍生物及它们的组合。
在某些实施方式中，其中所述抗PD-L1抗体包含VH以及VL，所述VH包含SEQ ID NO:25所示的氨基酸序列或与SEQ ID NO:25所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，且所述VL包含SEQ ID NO:38、SEQ ID NO:39或SEQ ID NO:40所示的氨基酸序列或与SEQ ID NO:38、SEQ ID NO:39或SEQ ID NO:40 所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
例如，其中所述抗PD-L1抗体可以包含VH以及VL，所述VH可以包含SEQ ID NO:25所示的氨基酸序列，且所述VL可以包含SEQ ID NO:38、SEQ ID NO:39或SEQ ID NO:40所示的氨基酸序列。
又例如，所述药物组合可以包含：1)抗PD-L1抗体，其中所述抗PD-L1抗体可以包含VH以及VL，所述VH可以包含SEQ ID NO:25所示的氨基酸序列，且所述VL可以包含SEQ ID NO:38、SEQ ID NO:39或SEQ ID NO:40所示的氨基酸序列；2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以是Imiquimod或其衍生物及它们的组合。
在某些实施方式中，其中所述抗PD-L1抗体包含LC，所述LC包含SEQ ID NO:41所示氨基酸序列或与SEQ ID NO:41所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含LC，所述LC包含SEQ ID NO:42、SEQ ID NO:43或SEQ ID NO:44所示氨基酸序列或与SEQ ID NO:42、SEQ ID NO:43或SEQ ID NO:44所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
在某些实施方式中，其中所述抗PD-L1抗体包含HC和LC，所述HC包含SEQ ID NO:26所示氨基酸序列或与SEQ ID NO:26所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，且所述LC包含SEQ ID NO:41所示氨基酸序列或与SEQ ID NO:41所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
例如，其中所述抗PD-L1抗体可以包含HC和LC，所述HC可以包含SEQ ID NO:26所示氨基酸序列，且所述LC可以包含SEQ ID NO:41所示氨基酸序列。
又例如，所述药物组合可以包含：1)抗PD-L1抗体，其中所述抗PD-L1抗体可以包含HC和LC，所述HC可以包含SEQ ID NO:26所示氨基酸序列，且所述LC可以包含SEQ ID NO:41所示氨基酸序列；2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以是Imiquimod或其衍 生物及它们的组合。
在某些实施方式中，其中所述抗PD-L1抗体包含HC和LC，所述HC包含SEQ ID NO:26所示氨基酸序列或与SEQ ID NO:26所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列，且所述LC包含SEQ ID NO:42、SEQ ID NO:43或SEQ ID NO:44所示氨基酸序列或与SEQ ID NO:42、SEQ ID NO:43或SEQ ID NO:44所示的氨基酸序列具有至少约70％、约75％、约80％、约85％、约90％、约91％、约92％、约93％、约94％、约95％、约96％、约97％、约98％、或约99％序列同一性的氨基酸序列。
例如，其中所述抗PD-L1抗体可以包含HC和LC，所述HC可以包含SEQ ID NO:26所示氨基酸序列，且所述LC可以包含SEQ ID NO:42、SEQ ID NO:43或SEQ ID NO:44所示氨基酸序列。
又例如，所述药物组合可以包含：1)抗PD-L1抗体，其中所述抗PD-L1抗体可以包含HC和LC，所述HC可以包含SEQ ID NO:26所示氨基酸序列，且所述LC可以包含SEQ ID NO:42、SEQ ID NO:43或SEQ ID NO:44所示氨基酸序列；2)咪唑喹啉衍生物，所述咪唑喹啉衍生物可以是Imiquimod或其衍生物及它们的组合。
在某些实施方式中，所述药物组合中的i)所述PD-1抑制剂和/或PD-L1抑制剂，与ii)所述TLR激动剂在所述药物组合中彼此不混合，即i)所述PD-1抑制剂和/或ii)PD-L1抑制剂与TLR激动剂位于分开的剂型中。
在某些实施方式中，所述药物组合中的i)所述PD-1抑制剂和/或PD-L1抑制剂，与ii)所述TLR激动剂位于单一的剂型中。
在某些实施方式中，其中所述药物组合被配制成药物组合物(例如复合制剂)。该药物组合物可以直接注射到大肿瘤内，而不影响正常(周围)组织，使得能够杀死癌细胞，使得能够阻止或延迟恶性肿块的生长(例如，使肿块变小或使肿瘤缩小)，并且使晚期癌症患者能够与肿瘤一起生活(以与寄生虫一起生活的人患者类似的方式)。当将药物组合注射到肿瘤内时，该药物可以沿着血管或淋巴管流动至转移灶，并且它将杀死转移细胞。药物组合注射到肿瘤内导致对患者的很少创伤，并且可以例如每月重复多次。直接注射也可以同时施用于原发性肿瘤和癌症已转移到的继发性肿瘤。
在某些实施方式中，其中所述药物组合物包括PD-1抑制剂或PD-L1抑制剂，以及TLR激动剂。
例如，所述药物组合物可以包括PD-1抑制剂和TLR激动剂。又例如，所述药物组合物 可以包括抗PD-1抗体或其抗原结合片段和咪唑喹啉衍生物。
例如，所述药物组合物可以包括PD-L1抑制剂和TLR激动剂。又例如，所述药物组合物可以包括抗PD-L1抗体或其抗原结合片段和咪唑喹啉衍生物。
在某些实施方式中，其中所述TLR激动剂的存在量为约0.0001mg/kg至约200mg/kg。例如，所述TLR激动剂可以相对于受试者的体重存在于本文所述的组合中(即mg/kg)。在一些情况下，TLR激动剂的存在量等于约0.0001mg/kg至约200mg/kg、0.001mg/kg至约200mg/kg、0.01mg/kg至约200mg/kg、0.01mg/kg至约150mg/kg、0 01mg/kg至约100mg/kg、0.01mg/kg至约50mg/kg、0.01mg/kg至约25mg/kg、0.01mg/kg至约10mg/kg或0.01mg/kg至约5mg/kg、0.05mg/kg至约200mg/kg、0.05mg/kg至约150mg/kg、0.05mg/kg至约100mg/kg、0.05mg/kg至约50mg/kg、0.05mg/kg至约25mg/kg、0.05mg/kg至约10mg/kg或0.05mg/kg至约5mg/kg、0.5mg/kg至约200mg/kg、0.5mg/kg至约150mg/kg、0.5mg/kg至约100mg/kg、0.5mg/kg至约50mg/kg、0.5mg/kg至约25mg/kg、0.5mg/kg至约10mg/kg或0.5mg/kg至约5mg/kg。在其它情况下，TLR激动剂的存在量等于约1mg/kg至约200mg/kg、1mg/kg至约150mg/kg、1mg/kg至约100mg/kg、1mg/kg至约50mg/kg、1mg/kg至约25mg/kg、1mg/kg至约10mg/kg或1mg/kg至约5mg/kg。
又例如，TLR激动剂可以约1mg、5mg、10mg、15mg、20mg、25mg、30mg、40mg、50mg、60mg、70mg、75mg、80mg、90mg、100mg、150mg、200mg、250mg、300mg、400mg、500mg、600mg、700mg、800mg、900mg、1000mg、1100mg、1200mg、1300mg、1400mg、1500mg、1600mg、1700mg、1800mg、1900mg或2000mg的量存在于组合中。TLR激动剂可以约1mg至约10mg、10mg至约20mg、25mg至约50mg、30mg至约60mg、40mg至约50mg、50mg至约100mg、75mg至约150mg、100mg至约200mg、200mg至约500mg、500mg至约1000mg、1000mg至约1200mg、1000mg至约1500mg、1200mg至约1500mg或1500mg至约2000mg的量存在于组合中。
又例如，TLR激动剂可以约0.1mg/mL、0.5mg/mL、1mg/mL、2mg/mL、3mg/mL、4mg/mL、5mg/ml、6mg/mL、7mg/mL、8mg/mL、9mg/mL、10mg/mL、15mg/mL、20mg/mL、25mg/mL、30mg/mL、40mg/mL、50mg/mL、60mg/mL、70mg/mL、80mg/mL、90mg/mL、100mg/mL、150mg/mL、200mg/mL、250mg/mL、300mg/mL、400mg/mL或500mg/mL的量存在于组合中。在一些实施例中，TLR激动剂以约1mg/mL至约10mg/mL、5mg/mL至约10mg/mL、5mg/mL至约15mg/mL、10mg/mL至约25mg/mL、20mg/mL至约30mg/mL、25mg/mL至约50mg/mL或50mg/mL至约100mg/mL的量存在于组合中。
在某些实施方式中，其中所述PD-1抑制剂或PD-L1抑制剂(例如抗PD-1/PD-L1抗体)的存在量为约0.0001mg/kg至约200mg/kg。例如，所述PD-1抑制剂或PD-L1抑制剂可以相对于受试者的体重存在于本文所述的组合中(即mg/kg)。在一些情况下，PD-1抑制剂或PD-L1抑制剂的存在量等于约0.0001mg/kg至约200mg/kg、0.001mg/kg至约200mg/kg、0.01mg/kg至约200mg/kg、0.01mg/kg至约150mg/kg、0 01mg/kg至约100mg/kg、0.01mg/kg至约50mg/kg、0.01mg/kg至约25mg/kg、0.01mg/kg至约10mg/kg或0.01mg/kg至约5mg/kg、0.05mg/kg至约200mg/kg、0.05mg/kg至约150mg/kg、0.05mg/kg至约100mg/kg、0.05mg/kg至约50mg/kg、0.05mg/kg至约25mg/kg、0.05mg/kg至约10mg/kg或0.05mg/kg至约5mg/kg、0.5mg/kg至约200mg/kg、0.5mg/kg至约150mg/kg、0.5mg/kg至约100mg/kg、0.5mg/kg至约50mg/kg、0.5mg/kg至约25mg/kg、0.5mg/kg至约10mg/kg或0.5mg/kg至约5mg/kg。在其它情况下，PD-1抑制剂或PD-L1抑制剂的存在量等于约1mg/kg至约200mg/kg、1mg/kg至约150mg/kg、1mg/kg至约100mg/kg、1mg/kg至约50mg/kg、1mg/kg至约25mg/kg、1mg/kg至约10mg/kg或1mg/kg至约5mg/kg。
又例如，抗PD-1/PD-L1抗体可以约1mg、5mg、10mg、15mg、20mg、25mg、30mg、40mg、50mg、60mg、70mg、75mg、80mg、90mg、100mg、150mg、200mg、250mg、300mg、400mg、500mg、600mg、700mg、800mg、900mg、1000mg、1100mg、1200mg、1300mg、1400mg、1500mg、1600mg、1700mg、1800mg、1900mg或2000mg的量存在于组合中。抗PD-1/PD-L1抗体可以约1mg至约10mg、10mg至约20mg、25mg至约50mg、30mg至约60mg、40mg至约50mg、50mg至约100mg、75mg至约150mg、100mg至约200mg、200mg至约500mg、500mg至约1000mg、1000mg至约1200mg、1000mg至约1500mg、1200mg至约1500mg或1500mg至约2000mg的量存在于组合中。
又例如，抗PD-1/PD-L1抗体可以约0.1mg/mL、0.5mg/mL、1mg/mL、2mg/mL、3mg/mL、4mg/mL、5mg/ml、6mg/mL、7mg/mL、8mg/mL、9mg/mL、10mg/mL、15mg/mL、20mg/mL、25mg/mL、30mg/mL、40mg/mL、50mg/mL、60mg/mL、70mg/mL、80mg/mL、90mg/mL、100mg/mL、150mg/mL、200mg/mL、250mg/mL、300mg/mL、400mg/mL或500mg/mL的量存在于组合中。在一些实施例中，抗PD-1/PD-L1抗体以约1mg/mL至约10mg/mL、5mg/mL至约10mg/mL、5mg/mL至约15mg/mL、10mg/mL至约25mg/mL、20mg/mL至约30mg/mL、25mg/mL至约50mg/mL或50mg/mL至约100mg/mL的量存在于组合中。
在某些实施方式中，所述的TLR激动剂可以与所述PD-1/PD-L1抑制剂的量协同的量提供。给药的剂量毫无疑问随已知的因素而变，该因素比如是特定试剂的药物动力学特性、和 其给药模式和途径；受体的年龄、健康状况、和体重；症状性质和程度、并发治疗的种类、治疗频率、和预期效果。
在某些实施方式中，其中所述药物组合物还包含一种或多种药学上可接受的载体。
例如，药学上可接受的载体的实例可以包括以下一个或多个：水、盐水、磷酸盐缓冲盐水、右旋糖、甘油、乙醇等以及其组合。在一些实施例中，组合物中可包括等张剂，例如糖、多元醇(如甘露糖醇、山梨糖醇)或氯化钠。药学上可接受的载体可进一步包括少量增强结合蛋白或结合蛋白部分的存放期或有效性的辅助物质，如湿润剂或乳化剂、防腐剂或缓冲剂。
另一方面，本申请还提供了前述药物组合在制备药物中的用途，所述药物用于治疗赘生性疾病。
在某些实施方式中，其中所述赘生性疾病包括肿瘤和/或疣类疾病。
在某些实施方案中，所述肿瘤(或癌症)包括但不限于：肝细胞癌、肝转移性癌症、晚期肝细胞癌、胰腺癌、腺癌、肥大细胞瘤或肥大细胞肿瘤、卵巢癌、非小细胞肺癌、小细胞肺癌、黑素瘤、视网膜母细胞瘤、乳腺肿瘤、结肠直肠癌、组织细胞肉瘤、脑肿瘤、星形细胞瘤、胶质母细胞瘤、神经瘤、神经母细胞瘤、结肠癌、子宫颈癌、肉瘤、前列腺肿瘤、膀胱肿瘤、网状内皮组织肿瘤、肾母细胞瘤、卵巢癌、骨癌、骨肉瘤、肾癌、或头颈癌、口腔癌、喉癌、或口咽癌、乳腺癌、泌尿生殖道癌、肺癌、胃肠癌、表皮样癌、黑素瘤。
另一方面，本申请还提供了前述药物组合用于治疗赘生性疾病。
另一方面，本申请还提供了一种治疗赘生性疾病的药物，其包含前述药物组合。
另一方面，本申请还提供了一种治疗赘生性疾病的方法，其包括向有需要的受试者施用有效量的前述药物组合。
在某些实施方式中，其中所述受试者患有赘生物。
在某些实施方式中，其中所述赘生物位于哺乳动物中选自以下的位置中：
脑、头部、眼、鼻咽、口、舌、颈部、甲状腺、胃肠系统、肝脏、胰腺、胆囊、肺、呼吸系统、泌尿生殖系统、肾脏、膀胱、乳房、淋巴系统、心血管系统、神经系统、皮肤、胸腔、胸膜、肌肉骨骼系统、腹部，具有原发性或继发性性质。
在某些实施方式中，其中所述赘生物包括肿瘤和/或疣。
在某些实施方式中，其中所述施用包括局部、赘生物内(例如，肿瘤内或疣体内)或全身性施用。例如，采用瘤内或疣内注射，该方法既允许对患者的更少创伤，又允许杀死癌细胞，而不是正常细胞。药物组合直接注射入恶性肿瘤内也极大地降低或消除了许多常见的副作用。
在某些实施方式中，其中所述施用包括静脉注射、静脉滴注、肌肉注射、皮下注射和/或赘生物内注射。
在某些实施方式中，所述肿瘤包括：
(i)皮肤、眼、舌、口、甲状腺、乳房、子宫颈、子宫、肛门、前列腺、阴道、骨肉瘤、尿道癌、阴茎、睾丸和附睾的浅表恶性疾病，并且所述药物组合无需稀释用注射器直接注射到瘤体内；或
(ii)鼻咽的癌症，并且所述药物组合通过鼻咽镜用注射器或针注射到瘤体内；或
(iii)肝脏、肾脏和胆囊的癌症，并且所述药物组合使用注射器在超声的辅助下通过皮肤注射到瘤体内，或通过在腹腔镜手术过程中在患者的腹壁中形成的孔注射到瘤体内；或
(iv)卵巢、输卵管、胰腺、淋巴结转移或腹腔的直接腹膜侵入、腹部淋巴瘤的癌症，并且所述药物组合通过在腹腔镜手术过程中在患者的腹壁中形成的孔，用注射器注射到瘤体内；或
(v)食道、胃、十二指肠、小肠的癌或肉瘤，并且所述药物组合通过肠镜用针注射到瘤体内，或通过在腹腔镜手术过程中在患者的腹壁中形成的孔经由长注射器注射到瘤体内，或通过在胸腔镜手术过程中在患者的胸壁中形成的孔进行注射；
(vi)大肠和直肠的癌或肉瘤，并且所述药物组合通过结肠镜检查用针注射到瘤体内，或通过在腹腔镜手术过程中在患者的腹壁中形成的孔使用注射器进行注射；或
(vii)肺和气管的癌或肉瘤，并且所述药物组合使用纤维支气管镜的针注射到瘤体内；或
(viii)肺的癌，并且所述药物组合在超声、x射线、CT扫描或MR扫描的使用下用注射器进行注射，或通过在胸腔镜手术过程中在患者的胸壁中形成的孔进行注射；或
(ix)膀胱的癌或肉瘤，并且所述药物组合通过膀胱镜用针注射到瘤体内，或通过在腹腔镜手术过程中在患者的腹壁中形成的孔进行注射；或
(x)子宫的癌或肉瘤，并且所述药物组合的可注射制剂用注射器或宫腔镜的针注射到瘤体内；或通过在腹腔镜手术过程中在患者的腹壁中形成的孔进行注射；或
(xi)鼻咽和喉的癌或肉瘤，并且所述药物组合通过喉镜用针注射到瘤体内；或
(xii)脑的癌，并且所述药物组合在X射线、CT扫描或MR扫描的使用下，在相应的颅骨中钻洞后，用注射器或纤维镜的针注射到瘤体内；或
(xiii)恶性淋巴瘤或具有转移的淋巴结，并且所述药物组合使用针通过患者的皮肤注射到瘤体内，或者通过在腹腔镜手术过程中在患者的腹壁中形成的孔、或通过在胸腔镜手术过 程中在患者的胸壁中形成的孔进行注射。
在某些实施方式中，所述赘生物包括疣，并且所述药物组合使用针通过患者的皮肤注射到疣体内。
在某些实施方式中，其中采取相同或不同的给药途径施用所述药物组合中的i)PD-1抑制剂或PD-L1抑制剂，与ii)TLR激动剂。
在某些实施方式中，其包括向赘生物中注射所述TLR激动剂。
在某些实施方式中，其还包括向赘生物中注射或全身输注(例如静脉注射、静脉滴注)所述PD-1抑制剂或PD-L1抑制剂。
在某些实施方式中，其包括向赘生物中注射所述药物组合中的i)所述PD-1抑制剂或PD-L1抑制剂，与ii)所述TLR激动剂。
在某些实施方式中，其中同时或不同时施用所述药物组合中的i)PD-1抑制剂或PD-L1抑制剂，与ii)所述TLR激动剂。
在某些实施方式中，其中在所述TLR激动剂施用之前和/或施用之后施用所述PD-1抑制剂或PD-L1抑制剂。
例如，在TLR激动剂施用之前和/或施用之后至少5min，10min，20min，40min，1h，2h，4h，8h，12h，16h，1天，2天，3天，4天，5天，6天，7天或2周，施用所述PD-1/PD-L1抑制剂。
在某些实施方式中，所述药物组合中的i)PD-1抑制剂或PD-L1抑制剂，与ii)所述TLR激动剂通过赘生物内注射方式同时施用。
在某些实施方式中，所述药物组合中的i)PD-1抑制剂或PD-L1抑制剂，与ii)所述TLR激动剂通过赘生物内注射方式同时施用，且i)所述PD-1抑制剂或PD-L1抑制剂与ii)所述TLR激动剂位于同一剂型内。
在某些实施方式中，其中所述药物组合中的PD-1抑制剂或PD-L1抑制剂通过静脉注射方式，且所述TLR激动剂通过赘生物内注射方式同时施用，且所述PD-1抑制剂或PD-L1抑制剂与所述TLR激动剂位于分开剂型内。
另一方面，本申请提供了一种药盒，其包含前述药物组合。
在某些实施方式中，所述药盒中的药物组合的每种组分可以在分离的单独容器中提供。或者，本文所述药物组合的组分可以在单个容器中提供。在此类情况下，容器可以是准备好施用至有需要的患者的容器，例如IV袋、安瓿或注射器。在一些实施例中，药盒中的TLR激动剂被配制用于瘤内或疣内注射施用。PD-1/PD-L1抑制剂可以例如粉末(例如冻干粉末)或肠 胃外施用的溶液的形式提供。在某些情况下，PD-1/PD-L1抑制剂可以是如本文所述的抗PD-1/PD-L1抗体，其配制用于通过例如静脉内施用或瘤内或疣内注射施用。在一些实施例中，所述PD-1或PD-L1抑制剂以及TLR激动剂被配置为复合制剂，且用于瘤内或疣内注射施用。在一些实施方式中，所述复合制剂被配置为液体制剂。例如，将复合制剂提供为稳定的溶液的形式。又例如，将所述复合制剂提供为可用于直接注射(即在使用前不需要稀释)的剂型。在另一些实施方式中，所述复合制剂被配置成固体制剂(例如冻干形式)。
本文所述的药盒的内容物可以无菌形式提供。药盒和其内容物可以准备好向有需要的对象施用的形式提供。在此类情况下，药盒中的组分作为制剂提供，并且任选地在施用装置中提供，使得施用对于使用者几乎不需要进一步的行动。在药盒包括施用装置的情况下，此类装置包括本领域技术人员已知和理解的用于本文所述的施用途径的装置，如但不限于注射器、泵、袋、杯、吸入器、滴管、贴片、乳膏或注入器。
本文所述的药盒的还可以包含说明书，其含有关于例如用法、剂量、施用、禁忌症和/或关于使用此类药物的警告的信息。
不欲被任何理论所限，下文中的实施例仅仅是为了阐释本申请的药物组合和用途等，而不用于限制本申请发明的范围。
实施例
动物种属、品系、性别、体重、来源、合格证
C57/BL6JNifdc小鼠，雌性，体重17-22g，6-8周龄，SPF级，购于北京维通利华实验动物技术有限公司[实验动物质量合格证号：SCXK(京)2016-0006]。
饲养条件
所有小鼠均自由觅食和饮水，在室温(25±2)℃下饲养。饲料及水均经高压灭菌处理，全部实验饲养过程为SPF级。
肿瘤细胞株
小鼠结直肠癌细胞株MC38、CT26，小鼠肺癌Lewis瘤株LLC1，小鼠黑色素瘤细胞株B16，小鼠前列腺癌细胞株RM-1、TRAMP-C1，小鼠乳腺癌细胞株4T1、NAFA、MET-1等。
皮下移植瘤模型的建立
肿瘤细胞株选自：小鼠结直肠癌细胞株CT26、MC38，小鼠肺癌Lewis瘤株LLC1，小鼠黑色素瘤细胞株B16，小鼠前列腺癌细胞株RM-1、TRAMP-C1，小鼠乳腺癌细胞株4T1、NAFA、MET-1等
肿瘤细胞培养，传代，在细胞对数期收集细胞，做成浓度为(1.0×10 7)每毫升的细胞悬液， 小鼠右侧胁肋部下注射0.1ml细胞悬液(细胞数目为1.0×10 6个/只)，10天左右肿瘤长至直径约5mm，模型建立成功，随机均分为8。
药物评价指标
小鼠体重、肿瘤体积变化趋势，小鼠生存率，解剖瘤重抑制率。
瘤重抑制率＝[1-实验组(G2/G3/G4)平均瘤重/阴性对照组G1组平均瘤重)]×100％。
统计分析
数据用x±s表示，利用SPSS10.0软件进行处理，采用单因素方差分析(one-way ANOVA)。经统计学处理各组间数据差异性P值大小，P<0.05判定为组间具有显著性差异。
两药相互作用系数(coefficient of drug in interaction CDI/CI)评价。
CDI(CI)按下列公式计算:CDI(CI)＝AB/A*B。根据解剖瘤重进行计算，AB是两药联合组与对照组的比值。A或B是各药单独使用组与对照组的比值。如CDI＜1，证明两药作用性质为协同；如CDI＝1，则两药作用性质为相加；如CDI>1，则两药作用性质为拮抗。(Pikman,Y.et al,Synergistic Drug Combinations with a CDK4/6 Inhibitor in T-cell Acute Lymphoblastic Leukemia,(2017)Clin Cancer Res 23,1012-1024.PMID:28151717 PMCID:PMC5432118 DOI:10.1158/1078-0432.CCR-15-2869.)
实施例1
采用移植瘤模型测试PD-L1抑制剂RB0005与TLR7/TLR8激动剂Imiquimod(LPG2005)组成的药物组合抗肿瘤作用即对小鼠皮下移植瘤生长的抑制作用。
试验主要步骤
1.1小鼠移植瘤模型
成功建立小鼠皮下结直肠癌MC38移植瘤模型后，随机分组，每组6～8只。每两天给药1次，共计给药3次后观察至15天。
Figure PCTCN2022102169-appb-000001
1.2结果
图1-4显示，新型药物组合及各单药均可明显抑制肿瘤生长，解剖瘤重均显著低于阴性对照组(P<0.01，P<0.001)，新型药物组合药效优于Imiquimod或PD-L1抗体RB0005单独用药，表明新型药物组合具有更大优势。
如图2所示，溶剂对照组小鼠在第9天开始出现“死亡(当单只小鼠肿瘤体积超过2000mm 3时，在小鼠生存率统计分析过程中可将该只小鼠视为死亡(肿瘤体积变化曲线分析中并未将其排除))”，开始给药后第11天时，小鼠生存率为0；RB0005单药组截止到最后，小鼠生存率为33.3％。Imiquimod单药组小鼠生存率为50％。单一制剂组小鼠生存率为83.3％，其中组内有1只小鼠肿瘤消失，2只小鼠肿瘤体积持续减小(肿瘤体积<100mm 3)。
如图3A-3B所示，单一制剂组(G4组)小鼠肿瘤重量明显减小，肿瘤抑制率为87.9％，统计学上具有显著性差异(P<0.001)。
实施例2
采用移植瘤模型测试PD-1抑制剂RB0004与TLR7/TLR8激动剂Imiquimod组成的药物组合抗肿瘤作用即对小鼠皮下移植瘤生长的抑制作用。
2.1小鼠移植瘤模型
成功建立人源化PD-1小鼠结直肠癌MC38皮下移植瘤模型后，随机分组，每组8只，每两天给药1次。
Figure PCTCN2022102169-appb-000002
2.2结果
结果：新型药物组合(分开剂型)及各单药均可抑制肿瘤生长，解剖瘤重均显著低于阴性对照组(P<0.05，P<0.001)，新型药物组合药效优于Imiquimod或PD-1抗体RB0004单独用药，表明新型药物组合具有更大优势。
如图4所示，溶剂对照组开始给药后第13天时，小鼠生存率为0；RB0004单药组截止到最后，小鼠生存率为37.5％。Imiquimod组小鼠生存率为40％。联合给药组(G4)小鼠生存 率为87.5％，其中组内有1只小鼠肿瘤体积持续减小(肿瘤体积<100mm 3)。
如图5所示，溶剂对照组小鼠肿瘤体积随时间逐渐增大；RB0004单药组及LPG2005单药组，肿瘤体积增长较溶剂对照组缓慢；联合给药组(G4)给药后，肿瘤体积增长明显减缓，与对照组相比统计学差异显著(P＜0.001)。
如图6和表1所示，G4组小鼠肿瘤重量明显减小，瘤重抑制率为72.99％，统计学上具有显著性差异(P<0.001)且CDI＝0.83，说明二药有协同药物作用效果。
表1
组别 瘤重(g) IR TW％ CDI(相互作用指数)
G1:Vel 6.32 0.00 /
G2:LPG2005 3.62 42.71 /
G3:RB0004 3.60 42.98 /
G4:RB0004+LPG2005 1.71 72.99 0.83
实施例3
采用小鼠结直肠癌MC38移植瘤模型测试PD-L1抑制剂RB0005与不同剂量的TLR7/TLR8激动剂Imiquimod组成的药物组合抗肿瘤作用即对小鼠皮下移植瘤生长的抑制作用。
3.1小鼠移植瘤模型
成功建立小鼠皮下移植瘤模型后，随机分组，每组8只，每两天给药1次。
Figure PCTCN2022102169-appb-000003
Figure PCTCN2022102169-appb-000004
3.2结果
结果：成功建立了小鼠皮下移植瘤模型，三种单一剂型中G8组具有显著的肿瘤增殖抑制效果，瘤重抑制率均在75％以上(P<0.001)，而G6组和G7组没有表现出肿瘤抑制效果(P＞0.05)。具体结果图7-8。
如图7所示，G8组小鼠生存率为100％。
如图8和表2所示，G8组小鼠肿瘤重量明显减小，瘤重抑制率为75.09％，统计学上具有显著性差异(P<0.001)且CDI＝0.48，说明该复合制剂有显著的协同药物作用效果。
表2
组别 瘤重(g) IR TW％ CDI(相互作用指数)
G1:Vel 3.53 0.00 /
G2:RB0005 2.38 32.61 /
G5:LPG2005 2.70 23.35 /
G8:RB0005+LPG2005 0.88 75.09 0.48
实施例4
采用移植瘤模型测试PD-L1抑制剂RB0005与不同剂量的TLR7/TLR8激动剂Imiquimod组成的药物组合抗肿瘤作用即对小鼠皮下移植瘤生长的抑制作用。
4.1小鼠移植瘤模型
成功建立小鼠皮下移植瘤模型后，随机分组，每组8只，每两天给药1次。
Figure PCTCN2022102169-appb-000005
4.2结果
结果：成功建立了小鼠皮下移植瘤模型，三种单一剂型中G6组和G7组具有显著的肿瘤增殖抑制效果，瘤重抑制率均在70％以上(P<0.001)，而G8组没有表现出肿瘤抑制效果(P＞0.05)。具体结果图9-11。
如图10所示，G6组，G7组和G8组小鼠生存率到观察结束日均为100％。其中G7组内有2只小鼠肿瘤体积持续减小，G8组内有3只小鼠肿瘤体积持续减小(肿瘤体积<100mm 3)。RB0005联合5％Imiquimod Cream组(G10组)，到小鼠观察结束日生存率为100％。其中组内有2只小鼠肿瘤体积持续减小(肿瘤体积<100mm 3)。
如图11和表3所示，G6组，G7组和G8组的瘤重抑制率分别为73.87％、70.47％、65.97％，统计学上具有显著性差异(P<0.001)，但CDI指数显示，G6组和G7组分别为0.61和0.86，而G8组的CDI为1.09，说明G6组和G7组具有协同药物作用效果。RB0005联合5％Imiquimod Cream组(G10)的瘤重抑制率均在55.77％以上(P<0.05)，CDI＝0.86，说明二者联合有协同作用药物效果。
表3
组别 瘤重(g) IR TW％ CDI(相互作用指数)
G6 1.01 73.87 0.61
G7 1.16 70.47 0.86
G8 2.99 65.97 1.09
G10 1.51 55.77 0.86
实施例5
采用移植瘤模型测试PD-1抑制剂RB0004与不同剂量的TLR7/TLR8激动剂Imiquimod组成的药物组合抗肿瘤作用即对小鼠皮下移植瘤生长的抑制作用。
5.1小鼠移植瘤模型
成功建立小鼠皮下移植瘤模型后，随机分组，每组8只，每两天给药1次。
Figure PCTCN2022102169-appb-000006
Figure PCTCN2022102169-appb-000007
5.2结果
结果：G7组均有显著的肿瘤抑制效果(P<0.05)，瘤重抑制率为59.25％。具体结果图12-图13。
如图12所示，RB0004联合LPG2005-1、3.16mpk，到观察结束日小鼠生存率均高于50％。RB0004联合5％Cream组，到小鼠观察结束日生存率为83.3％。
如图13和表4所示，G7组与对照组相比，瘤重抑制率为59.25％，统计学上有显著性差异。RB0004联合5％Cream组(G10组)，瘤重抑制率为67.82％且CDI＝0.78，说明二者联合有协同作用药物效果。
表4
组别 瘤重(g) IR TW％ CDI(相互作用指数)
G10 1.72 67.82 0.775
实施例6 PD-L1抑制剂RB0005与不同剂量的TLR7/TLR8激动剂Imiquimod组成的药物组合LPG2006(单一剂型)抗肿瘤作用
6.1小鼠移植瘤模型
成功建立小鼠结直肠癌MC38人源化CD274细胞皮下移植瘤模型后，随机分组，每组6～8只，每两天给药1次，共计给药3次。
6.2分组
Figure PCTCN2022102169-appb-000008
Figure PCTCN2022102169-appb-000009
6.3结果
结果如图14-16及表5所示，RB0005单药有肿瘤抑制效果，瘤重抑制率为约40％；新型复合制剂组RB0005+Imiquimod-2mpk(G6)、RB0005+Imiquimod-6.32mpk(G7)对肿瘤有显著抑制作用，瘤重抑制率分别为73％、71％且表现出药物协同作用CDI值<1，差异有统计学意义(P<0.05)。
TLR7/TLR8激动剂Imiquimod与PD-L1单抗的单一剂型，该剂型中Imiquimod的剂量为2～6.25mpk，具有肿瘤抑制效果，且生存率高(P<0.05)，新型药物组合药效优于PD-L1抗体RB0005单独用药，表明新型药物组合具有更大优势。
表5
组别 Tumor Weight(g) IR TW％ CDI(相互作用指数)
G1 3.4 -  
G2 2.0 41.2  
G3 2.5 27.5  
G4 2.0 58.5  
G5 1.8 47.4  
G6 0.9 73.7 0.6(＜1)
G7 1.0 70.8 0.8(＜1)
G8 1.2 64.9 ＞1
实施例7 PD-L1抑制剂RB0005与TLR7/8激动剂Imiquimod组成的复合制剂(单一剂型和分开剂型)抗肿瘤作用
7.1小鼠移植瘤模型
成功建立小鼠结直肠癌MC38人源化CD274细胞皮下移植瘤模型后，随机分组，每组6～8只，每两天给药1次，共计给药3次。
7.2分组
Figure PCTCN2022102169-appb-000010
Figure PCTCN2022102169-appb-000011
7.3结果
结果如图17-19及表6所示，RB0005单药腹腔和瘤内(G3\G4)给药瘤重抑制率为43％、56％；Imiquimod单药腹腔和瘤内(G5\G6)，瘤内给药肿瘤抑制作用较好，且与Imiquimod腹腔给药相比，差异有统计学意义(P<0.05)。
新型复合制剂单一剂型(G7\G8)腹腔和瘤内给药，瘤内给药瘤重抑制率为77％，肿瘤抑制作用优与腹腔给药，试验终点小鼠生存率为100％，且表现出协同药物作用效果，CDI值为0.7。
Imiquimod与PD-L1单抗的单一剂型，瘤内给药(局部)方式肿瘤抑制效果较好，且生存率高(P<0.05)，优与复合制剂的腹腔给药及各单药单独使用，表明新型药物组合具有更大优势。
表6
组别 Tumor Weight(g) IR TW％ CDI(相互作用指数)
G1 3.17 -  
G2 3.4 -  
G3 1.8 43.2  
G4 1.5 55.9  
G5 3.74 -  
G6 2.6 23.5  
G7 2.16 31.9 ＞1
G8 0.78 77.1 0.7(＜1)
实施例8 PD-L1抑制剂RB0005与其他类TLR7/8激动剂(Gardiquimod)组成的药品组合抗肿瘤作用
8.1小鼠移植瘤模型
成功建立小鼠结直肠癌MC38人源化CD274细胞皮下移植瘤模型后，随机分组，每组6～8只，每两天给药1次，共计给药3次。
8.2分组
Figure PCTCN2022102169-appb-000012
Figure PCTCN2022102169-appb-000013
8.3结果
结果如图20-22及表7所示，RB0005单药瘤重抑制率为34％，Gardiquimod单药瘤重抑制率为42％，新型复合制剂单一剂型(G4)瘤重抑制率为72％，复合制剂小鼠生存率为100％。
与Gardiquimod组成的复合制剂单一剂型瘤内给药表现出协同药物作用效果，CDI值＜1。
TLR7/TLR8激动剂Gardiquimod与PD-L1单抗组成的单一剂型，具有肿瘤抑制效果且表现出药物协同作用，试验中生存率高(P<0.05)，组合药效优于单独用药，表明新型药物组合具有更大优势。
表7
Figure PCTCN2022102169-appb-000014
实施例9 PD-L1抑制剂RB0005与其他类TLR7/8激动剂(Resiquimod)组成的药品组合抗肿瘤作用
9.1小鼠移植瘤模型
成功建立小鼠结直肠癌MC38人源化CD274细胞皮下移植瘤模型后，随机分组，每组 6～8只，每两天给药1次，共计给药3次。
9.2分组
Figure PCTCN2022102169-appb-000015
9.3结果
结果如图23-25及表8所示，Resiquimod单药瘤重抑制率为40％，新型复合制剂单一剂型(G4)瘤重抑制率为73％，复合制剂小鼠生存率为85％，且表现出协同药物作用效果，CDI值＜1。
TLR7/TLR8激动剂Resiquimod与PD-L1单抗组成的单一剂型，具有肿瘤抑制效果且表现出药物协同作用，试验中生存率高(P<0.05)，组合药效优于单独用药。
表8
Figure PCTCN2022102169-appb-000016
实施例10
10.1上市类PD-1或PD-L1抑制剂与TLR7/8激动剂Imiquimod组成的复合制剂(单一剂型)抗肿瘤作用
10.1.1小鼠移植瘤模型
成功建立结直肠癌MC38人源化PD1小鼠皮下移植瘤模型后，随机分组，每组6～8只，每两天给药1次，共计给药3次。
10.1.2分组
Figure PCTCN2022102169-appb-000017
10.1.3结果
结果如图26-28及表9所示，上市类PD-1抑制剂单药(G3\G5\G7\G9)瘤内给药方式，G3 组(替雷利珠)肿瘤抑制效果较好，瘤重抑制率为61.4％；
新型复合制剂单一剂型(G4\G6\G8\G10)，其中G8组(信迪利)及G10组(特瑞普利)瘤内给药瘤重抑制率为35％和60％，表现出协同药物作用效果(CDI值＜1)，且试验终点小鼠生存率为60％以上。
上市类PD-1抑制剂单药中替雷利珠单抗瘤内给药对肿瘤抑制作用较好，与TLR7/8激动剂Imiquimod组成的复合制剂中，分别含信迪利单抗和特瑞普利单抗的复合制剂具有肿瘤抑制效果，且生存率高(P<0.05)，该药物组合具有较大优势。
表9
Figure PCTCN2022102169-appb-000018
10.2上市类PD-L1抑制剂与TLR7/8激动剂Imiquimod组成的复合制剂(单一剂型)抗肿瘤作用
10.2.1小鼠移植瘤模型
成功建立小鼠结直肠癌MC38人源化CD274细胞皮下移植瘤模型后，随机分组，每组6～8只，每两天给药1次，共计给药3次。
10.2.2分组
Figure PCTCN2022102169-appb-000019
Figure PCTCN2022102169-appb-000020
10.2.3结果
结果如图29-31及表10所示，上市类PD-L1抑制剂Durvalumab单药(G3)瘤内给药方式瘤重抑制率为21％，与TLR7/8激动剂Imiquimod组成的新型复合制剂(G4)瘤重抑制率为69％，表现出协同药物作用效果，且复合制剂生存率在80％以上；
RB0005与TLR7/8激动剂Imiquimod组成的新型复合制剂(G6)瘤重抑制率为78％，且表现出显著的协同药物作用效果(CDI值≤0.5)，实验终点复合制剂生存率100％；
新型复合制剂单一剂型中，含RB0005的复合制剂肿瘤抑制作用更显著。
上市类PD-L1抑制剂Durvalumab及RB0005分别与TLR7/8激动剂Imiquimod组成的复合制剂，均表现出肿瘤抑制效果及内含两种药物成分的协同作用，且生存率高(P<0.05)，表明该药物组合较单药更具有抗肿瘤优势，尤其是RB0005的药物组合。
表10
Figure PCTCN2022102169-appb-000021
Figure PCTCN2022102169-appb-000022
实施例11 PD-L1抑制剂RB0005与Imiquimod组合LPG2006(单一剂型)，CT26肿瘤模型
11.1小鼠移植瘤模型
成功建立小鼠结直肠癌CT26人源化CD274细胞皮下移植瘤模型后，随机分组，每组6～8只，每两天给药1次，共计给药3次。
11.2分组
Figure PCTCN2022102169-appb-000023
Figure PCTCN2022102169-appb-000024
11.3结果
结果如图32-34及表11所示，RB0005单药在CT26模型中未表现出肿瘤抑制作用，但新型复合制剂单一剂型(G6)瘤重抑制率为51％，且表现出协同药物作用效果，CDI值＜1。
CT26模型对RB0005单药不敏感，但RB0005与TLR7/TLR8激动剂Imiquimod组成的单一剂型，具有肿瘤抑制效果且表现出药物协同作用，表明组合药效优于单独用药且提高肿瘤模型对PD-L1抑制剂的敏感性。
表11
组别 Tumor Weight(g) IR TW％ CDI(相互作用指数)
G3 1.84    
G4 1.91 -  
G5 1.05 42  
G6 0.92 51 0.8(＜1)
实施例12 Imiquimod与PD-L1单抗序贯给药对MC38肿瘤增值抑制情况
12.1小鼠移植瘤模型
成功建立小鼠结直肠癌MC38人源化CD274细胞皮下移植瘤模型后，随机分组，每组6～8只，每两天给药1次，共计给药3次。
12.2分组
Figure PCTCN2022102169-appb-000025
Figure PCTCN2022102169-appb-000026
12.3结果
结果如图36-38及表12所示，序贯给药组(G4)肿瘤增值抑制作用明显，给药结束后观察期内，肿瘤体积增值缓慢。序贯给药组(G4)与对照及RB0005单药组相比，差异有统计学意义(ANOVA，P＜0.001)，瘤重抑制率为68％。
序贯联合(先给小分子，后大分子)，肿瘤抑制作用显著，且表现出协同药物作用效果，CDI＝0.49。实验终点，序贯联合给药组小鼠生存率为80％以上，与对照组及RB0005单药组相比，显著提高了小鼠生存率。
Imiquimod与PD-L1单抗，以先给小分子48h后给PD-L1抑制剂的方式瘤内给药，肿瘤抑制效果显著，表现出协同药物作用效果，且提高了小鼠生存率(P＜0.05)，序贯给药与单药相比具有显著优势。
表12
Figure PCTCN2022102169-appb-000027

Claims (111)

  1. 药物组合，其包含程序性细胞死亡蛋白1(PD-1)抑制剂和/或程序性死亡配体1(PD-L1)抑制剂，以及Toll样受体(TLR)激动剂。
  2. 根据权利要求1所述的药物组合，其中所述TLR包括TLR1，TLR2，TLR3，TLR4，TLR5，TLR6，TLR7，TLR8，TLR9和/或TLR10。
  3. 根据权利要求1-2中任一项所述的药物组合，其中所述TLR激动剂选自：TLR7激动剂，TLR8激动剂，TLR9激动剂中的一种或多种。
  4. 根据权利要求1-3中任一项所述的药物组合，其中所述TLR激动剂包括TLR7和TLR8双重激动剂(TLR7/TLR8激动剂)。
  5. 根据权利要求1-4中任一项所述的组合物，其中所述TLR激动剂包括dsRNA、ssRNA、CpG DNA、咪唑喹啉衍生物和/或鸟苷类似物。
  6. 根据权利要求1-5中任一项所述的组合物，其中所述TLR激动剂包括咪唑喹啉衍生物。
  7. 根据权利要求1-6中任一项所述的组合物，其中所述TLR激动剂选自咪喹莫特(Imiquimod)、嘎德莫特(Gardiquimod)、瑞喹莫特(Resiquimod)、1V209、Selgantolimod(GS-9688)、Vesatolimod(GS-9620)、Sumanirole、PF-4878691、及其药学上可接受的衍生物中的一种或多种。
  8. 根据权利要求1-7中任一所述的药物组合，其中所述TLR激动剂包括咪喹莫特、瑞喹莫特或其药学上可接受的盐。
  9. 根据权利要求1-5中任一项所述的组合物，其中所述TLR激动剂选自LHC-165、NKTR-262、DN1508052-01、SHR2150、CL307、CL264、洛索立宾、艾沙托立宾、DSR-6434、GSK2245035、SM-276001、SM-324405、SM-324406、AZ12441970、AZ12443988及其药学上可接受的衍生物中的一种或多种。
  10. 根据权利要求1-9中任一项所述的药物组合，其中所述PD-1抑制剂具有以下一个或多个特征：
    a.抑制或减少PD-1表达，例如PD-1的转录或翻译；
    b.抑制或降低PD-1活性，例如抑制或降低PD-1与其同源配体，例如PD-L1或PD-L2的结合；和
    c.结合PD-1或其一个或多个配体，例如PD-L1或PD-L2。
  11. 根据权利要求1-10中任一项所述的药物组合，其中所述PD-1抑制剂包括抗PD-1抗体或其抗原结合片段。
  12. 根据权利要求11所述的药物组合，其中所述抗PD-1抗体选自Pembrolizumab，Nivolumab， Pidilizumab，Tislelizumab，Camrelizumab(SHR-1210)，Sintilimab，Toripalimab，MEDI0680，BGB-A317，TSR-042，REGN2810、PF-06801591、RB0004、其类似物及它们的组合。
  13. 根据权利要求11-12中任一项所述的药物组合，其中所述抗PD-1抗体包含抗体重链可变区(VH)中的至少一个CDR，所述VH包含SEQ ID NO:8所示的氨基酸序列。
  14. 根据权利要求11-13中任一项所述的药物组合，其中所述抗PD-1抗体包含VH，所述VH包含HCDR3，所述HCDR3包含SEQ ID NO:3所示的氨基酸序列。
  15. 根据权利要求14所述的药物组合，其中所述VH还包含HCDR2，其中所述HCDR2包含SEQ ID NO:2所示的氨基酸序列。
  16. 根据权利要求14-15中任一项所述的药物组合，其中所述VH还包含HCDR1，其中所述HCDR1包含SEQ ID NO:1所示的氨基酸序列。
  17. 根据权利要求14-16中任一项所述的药物组合，其中所述VH包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:3所示的氨基酸序列，所述HCDR2包含SEQ ID NO:2所示的氨基酸序列，所述HCDR1包含SEQ ID NO:1所示的氨基酸序列。
  18. 根据权利要求14-17中任一项所述的药物组合，其中所述VH包括框架区HFR1，所述HFR1的C末端与所述HCDR1的N末端直接或间接相连，且所述HFR1包含SEQ ID NO:4所示的氨基酸序列或与SEQ ID NO:4所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  19. 根据权利要求14-18中任一项所述的药物组合，其中所述VH包括框架区HFR2，所述HFR2的N末端与所述HCDR1的C末端直接或间接相连，且所述HFR2的C末端与所述HCDR2的N末端直接或间接相连；且所述HFR2包含SEQ ID NO:5所示的氨基酸序列或与SEQ ID NO:5所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  20. 根据权利要求14-19中任一项所述的药物组合，其中所述VH包括框架区HFR3，所述HFR3的N末端与所述HCDR2的C末端直接或间接相连，且所述HFR3的C末端与所述HCDR3的N末端直接或间接相连；且所述HFR3包含SEQ ID NO:6所示的氨基酸序列或与SEQ ID NO:6所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  21. 根据权利要求14-20中任一项所述的药物组合，其中所述VH包括框架区HFR4，所述HFR4的N末端与所述HCDR3的C末端直接或间接相连，且所述HFR4包含SEQ ID NO:7所示的氨基酸序列或与SEQ ID NO:7所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  22. 根据权利要求14-21中任一项所述的药物组合，其中所述VH包括框架区HFR1，HFR2， HFR3和HFR4，所述HFR1的C末端与所述HCDR1的N末端直接或间接相连，所述HFR2的N末端与所述HCDR1的C末端直接或间接相连，且所述HFR2的C末端与所述HCDR2的N末端直接或间接相连，所述HFR3的N末端与所述HCDR2的C末端直接或间接相连，且所述HFR3的C末端与所述HCDR3的N末端直接或间接相连，所述HFR4的N末端与所述HCDR3的C末端直接或间接相连；其中，所述HFR1包含SEQ ID NO:4所示的氨基酸序列或与SEQ ID NO:4所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述HFR2包含SEQ ID NO:5所示的氨基酸序列或与SEQ ID NO:5所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述HFR3包含SEQ ID NO:6所示的氨基酸序列或与SEQ ID NO:6所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述HFR4包含SEQ ID NO:7所示的氨基酸序列或与SEQ ID NO:7所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  23. 根据权利要求11-22中任一项所述的药物组合，其中所述抗PD-1抗体包含VH，所述VH包含SEQ ID NO:8所示的氨基酸序列。
  24. 根据权利要求11-23中任一项所述的药物组合，其中所述抗PD-1抗体包含抗体重链(HC)，所述HC包含SEQ ID NO:9所示氨基酸序列。
  25. 根据权利要求11-24中任一项所述的药物组合，其中所述抗PD-1抗体包含抗体轻链可变区(VL)中的至少一个CDR，所述VL包含SEQ ID NO:17所示的氨基酸序列。
  26. 根据权利要求11-25中任一项所述的药物组合，其中所述抗PD-1抗体包含VH中的至少一个CDR，所述VH包含SEQ ID NO:8所示的氨基酸序列，且所述抗PD-1抗体包含VL中的至少一个CDR，所述VL包含SEQ ID NO:17所示的氨基酸序列。
  27. 根据权利要求11-26中任一项所述的药物组合，其中所述抗PD-1抗体包含VL，所述VL包含LCDR1，所述LCDR1包含SEQ ID NO:10所示的氨基酸序列。
  28. 根据权利要求27所述的药物组合，其中所述VL还包含LCDR2，其中所述LCDR2包含SEQ ID NO:11所示的氨基酸序列。
  29. 根据权利要求27-28中任一项所述的药物组合，其中所述VL还包含LCDR3，其中所述LCDR3包含SEQ ID NO:12所示的氨基酸序列。
  30. 根据权利要求27-29中任一项所述的药物组合，其中所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:10所示的氨基酸序列，所述LCDR2包含SEQ ID NO:11所示的氨基酸序列，所述LCDR3包含SEQ ID NO:12所示的氨基酸序列。
  31. 根据权利要求11-30中任一项所述的药物组合，其中所述抗PD-1抗体包含VH以及抗体 VL，所述VH包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:3所示的氨基酸序列，所述HCDR2包含SEQ ID NO:2所示的氨基酸序列，所述HCDR1包含SEQ ID NO:1所示的氨基酸序列；且所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:10所示的氨基酸序列，所述LCDR2包含SEQ ID NO:11所示的氨基酸序列，所述LCDR3包含SEQ ID NO:12所示的氨基酸序列。
  32. 根据权利要求27-31中任一项所述的药物组合，其中所述VL包括框架区LFR1，所述LFR1的C末端与所述LCDR1的N末端直接或间接相连，且所述LFR1包含SEQ ID NO:13所示的氨基酸序列或与SEQ ID NO:13所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  33. 根据权利要求27-32中任一项所述的药物组合，其中所述VL包括框架区LFR2，所述LFR2的N末端与所述LCDR1的C末端直接或间接相连，且所述LFR2的C末端与所述LCDR2的N末端直接或间接相连；且所述LFR2包含SEQ ID NO:14所示的氨基酸序列或与SEQ ID NO:14所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  34. 根据权利要求27-33中任一项所述的药物组合，其中所述VL包括框架区LFR3，所述LFR3的N末端与所述LCDR2的C末端直接或间接相连，且所述LFR3的C末端与所述LCDR3的N末端直接或间接相连；且所述LFR3包含SEQ ID NO:15所示的氨基酸序列或与SEQ ID NO:15所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  35. 根据权利要求27-34中任一项所述的药物组合，其中所述VL包括框架区LFR4，所述LFR4的N末端与所述LCDR3的C末端直接或间接相连，且所述LFR4包含SEQ ID NO:16所示的氨基酸序列或与SEQ ID NO:16所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  36. 根据权利要求27-35中任一项所述的药物组合，其中所述VL包括框架区LFR1，LFR2，LFR3和LFR4，所述LFR1的C末端与所述LCDR1的N末端直接或间接相连，所述LFR2的N末端与所述LCDR1的C末端直接或间接相连，且所述LFR2的C末端与所述LCDR2的N末端直接或间接相连，所述LFR3的N末端与所述LCDR2的C末端直接或间接相连，且所述LFR3的C末端与所述LCDR3的N末端直接或间接相连，所述LFR4的N末端与所述LCDR3的C末端直接或间接相连；其中，所述LFR1包含SEQ ID NO:13所示的氨基酸序列或与SEQ ID NO:13所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述LFR2包含SEQ ID NO:14所示的氨基酸序列或与SEQ ID NO:14所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述LFR3包含SEQ ID NO:15所示 的氨基酸序列或与SEQ ID NO:15所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述LFR4包含SEQ ID NO:16所示的氨基酸序列或与SEQ ID NO:16所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  37. 根据权利要求11-36中任一项所述的药物组合，其中所述抗PD-1抗体包含VL，所述VL包含SEQ ID NO:17所示的氨基酸序列。
  38. 根据权利要求11-37中任一项所述的药物组合，其中所述抗PD-1抗体包含VH以及VL，所述VH包含SEQ ID NO:8所示的氨基酸序列，且所述VL包含SEQ ID NO:17所示的氨基酸序列。
  39. 根据权利要求11-38中任一项所述的药物组合，其中所述抗PD-1抗体包含抗体轻链(LC)，所述LC包含SEQ ID NO:18所示氨基酸序列。
  40. 根据权利要求11-39中任一项所述的药物组合，其中所述抗PD-1抗体包含HC和LC，所述HC包含SEQ ID NO:9所示氨基酸序列，且所述LC包含如SEQ ID NO:18所示氨基酸序列。
  41. 根据权利要求1所述的药物组合，其中所述PD-L1抑制剂具有以下一个或多个特征：
    a.抑制或减少PD-L1表达，例如PD-L1的转录或翻译；
    b.抑制或降低PD-L1活性，例如抑制或降低PD-L1与其关联受体，例如PD-1的结合；和
    c.结合PD-L1或其受体，例如PD-1。
  42. 根据权利要求1-41中任一项所述的药物组合，其中所述PD-L1抑制剂包括抗PD-L1抗体或其抗原结合片段。
  43. 根据权利要求42所述的药物组合，其中所述抗PD-L1抗体选自Durvalumab，Atezolizumab，Avelumab，MDX-1105，YW243.55.S70，MDPL3280A，AMP-224、LY3300054、RB0005、其类似物及它们的组合。
  44. 根据权利要求42-43中任一项所述的药物组合，其中所述抗PD-L1抗体包含VH中的至少一个CDR，所述VH包含SEQ ID NO:25所示的氨基酸序列。
  45. 根据权利要求42-44中任一项所述的药物组合，其中所述抗PD-L1抗体包含VH，所述VH包含HCDR3，所述HCDR3包含SEQ ID NO:21所示的氨基酸序列。
  46. 根据权利要求45所述的药物组合，其中所述VH还包含HCDR2，其中所述HCDR2包含SEQ ID NO:20所示的氨基酸序列。
  47. 根据权利要求45-46中任一项所述的药物组合，其中所述VH还包含HCDR1，其中所述 HCDR1包含SEQ ID NO:19所示的氨基酸序列。
  48. 根据权利要求45-47中任一项所述的药物组合，其中所述VH包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:21所示的氨基酸序列，所述HCDR2包含SEQ ID NO:20所示的氨基酸序列，所述HCDR1包含SEQ ID NO:19所示的氨基酸序列。
  49. 根据权利要求45-48中任一项所述的药物组合，其中所述VH包括框架区HFR1，所述HFR1的C末端与所述HCDR1的N末端直接或间接相连，且所述HFR1包含SEQ ID NO:22所示的氨基酸序列或与SEQ ID NO:22所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  50. 根据权利要求45-49中任一项所述的药物组合，其中所述VH包括框架区HFR2，所述HFR2的N末端与所述HCDR1的C末端直接或间接相连，且所述HFR2的C末端与所述HCDR2的N末端直接或间接相连；且所述HFR2包含SEQ ID NO:23所示的氨基酸序列或与SEQ ID NO:23所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  51. 根据权利要求45-50中任一项所述的药物组合，其中所述VH包括框架区HFR3，所述HFR3的N末端与所述HCDR2的C末端直接或间接相连，且所述HFR3的C末端与所述HCDR3的N末端直接或间接相连；且所述HFR3包含SEQ ID NO:24所示的氨基酸序列或与SEQ ID NO:24所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  52. 根据权利要求45-51中任一项所述的药物组合，其中所述VH包括框架区HFR4，所述HFR4的N末端与所述HCDR3的C末端直接或间接相连，且所述HFR4包含SEQ ID NO:7所示的氨基酸序列或SEQ ID NO:7所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  53. 根据权利要求45-52中任一项所述的药物组合，其中所述VH包括框架区HFR1，HFR2，HFR3和HFR4，所述HFR1的C末端与所述HCDR1的N末端直接或间接相连，所述HFR2的N末端与所述HCDR1的C末端直接或间接相连，且所述HFR2的C末端与所述HCDR2的N末端直接或间接相连，所述HFR3的N末端与所述HCDR2的C末端直接或间接相连，且所述HFR3的C末端与所述HCDR3的N末端直接或间接相连，所述HFR4的N末端与所述HCDR3的C末端直接或间接相连；其中，所述HFR1包含SEQ ID NO:22所示的氨基酸序列或与SEQ ID NO:22所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述HFR2包含SEQ ID NO:23所示的氨基酸序列或与SEQ ID NO:23所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述HFR3包含SEQ ID NO:24所示的氨基酸序列或与SEQ ID NO:24所示的氨基酸序列具有至少约70％序列同一性 的氨基酸序列，所述HFR4包含SEQ ID NO:7所示的氨基酸序列或与SEQ ID NO:7所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  54. 根据权利要求42-53中任一项所述的药物组合，其中所述抗PD-L1抗体包含VH，所述VH包含SEQ ID NO:25所示的氨基酸序列。
  55. 根据权利要求42-54中任一项所述的药物组合，其中所述抗PD-L1抗体包含HC，所述HC包含SEQ ID NO:26所示氨基酸序列。
  56. 根据权利要求42-55中任一项所述的药物组合，其中所述抗PD-L1抗体包含VL中的至少一个CDR，所述VL包含SEQ ID NO:37所示的氨基酸序列。
  57. 根据权利要求42-56中任一项所述的药物组合，其中所述抗PD-L1抗体包含VH中的至少一个CDR，所述VH包含SEQ ID NO:25所示的氨基酸序列，且所述抗PD-L1抗体包含VL中的至少一个CDR，所述VL包含SEQ ID NO:37所示的氨基酸序列。
  58. 根据权利要求42-57中任一项所述的药物组合，其中所述抗PD-L1抗体包含VH中的至少一个CDR，所述VH包含SEQ ID NO:25所示的氨基酸序列，且所述抗PD-L1抗体包含VL中的至少一个CDR，所述VL包含SEQ ID NO:38、SEQ ID NO:39或SEQ ID NO:40所示的氨基酸序列。
  59. 根据权利要求42-58中任一项所述的药物组合，其中所述抗PD-L1抗体包含VL，所述VL包含LCDR1，所述LCDR1包含SEQ ID NO:27所示的氨基酸序列。
  60. 根据权利要求59所述的药物组合，其中所述抗PD-L1抗体包含VL，所述VL包含LCDR1，所述LCDR1包含SEQ ID NO:28、SEQ ID NO:29或SEQ ID NO:30所示的氨基酸序列。
  61. 根据权利要求59-60中任一项所述的药物组合，其中所述VL还包含LCDR2，其中所述LCDR2包含SEQ ID NO:31所示的氨基酸序列。
  62. 根据权利要求59-61中任一项所述的药物组合，其中所述VL还包含LCDR3，其中所述LCDR3包含SEQ ID NO:32所示的氨基酸序列。
  63. 根据权利要求59-62中任一项所述的药物组合，其中所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:27所示的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列。
  64. 根据权利要求63所述的药物组合，其中所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:28所示的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列；
    所述LCDR1包含SEQ ID NO:29所示的氨基酸序列，所述LCDR2包含SEQ ID NO:31 所示的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列；或
    所述LCDR1包含SEQ ID NO:30所示的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列。
  65. 根据权利要求42-64中任一项所述的药物组合，其中抗PD-L1抗体包含VH以及抗体VL，所述VH包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:21所示的氨基酸序列，所述HCDR2包含SEQ ID NO:20所示的氨基酸序列，所述HCDR1包含SEQ ID NO:19所示的氨基酸序列；且所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:27所示的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列。
  66. 根据权利要求65所述的药物组合，其中所述抗PD-L1抗体包含VH以及抗体VL，所述VH包含HCDR1，HCDR2和HCDR3，其中所述HCDR3包含SEQ ID NO:21所示的氨基酸序列，所述HCDR2包含SEQ ID NO:20所示的氨基酸序列，所述HCDR1包含SEQ ID NO:19所示的氨基酸序列；且所述VL包含LCDR1，LCDR2和LCDR3，其中所述LCDR1包含SEQ ID NO:28、SEQ ID NO:29或SEQ ID NO:30所示的氨基酸序列，所述LCDR2包含SEQ ID NO:31所示的氨基酸序列，所述LCDR3包含SEQ ID NO:32所示的氨基酸序列。
  67. 根据权利要求59-66中任一项所述的药物组合，其中所述VL包括框架区LFR1，所述LFR1的C末端与所述LCDR1的N末端直接或间接相连，且所述LFR1包含SEQ ID NO:33所示的氨基酸序列或与SEQ ID NO:33所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  68. 根据权利要求59-67中任一项所述的药物组合，其中所述VL包括框架区LFR2，所述LFR2的N末端与所述LCDR1的C末端直接或间接相连，且所述LFR2的C末端与所述LCDR2的N末端直接或间接相连；且所述LFR2包含SEQ ID NO:34所示的氨基酸序列或与SEQ ID NO:34所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  69. 根据权利要求59-68中任一项所述的药物组合，其中所述VL包括框架区LFR3，所述LFR3的N末端与所述LCDR2的C末端直接或间接相连，且所述LFR3的C末端与所述LCDR3的N末端直接或间接相连；且所述LFR3包含SEQ ID NO:35所示的氨基酸序列或与SEQ ID NO:35所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  70. 根据权利要求59-69中任一项所述的药物组合，其中所述VL包括框架区LFR4，所述LFR4的N末端与所述LCDR3的C末端直接或间接相连，且所述LFR4包含SEQ ID NO:36所 示的氨基酸序列或与SEQ ID NO:36所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  71. 根据权利要求59-70中任一项所述的药物组合，其中所述VL包括框架区LFR1，LFR2，LFR3和LFR4，所述LFR1的C末端与所述LCDR1的N末端直接或间接相连，所述LFR2的N末端与所述LCDR1的C末端直接或间接相连，且所述LFR2的C末端与所述LCDR2的N末端直接或间接相连，所述LFR3的N末端与所述LCDR2的C末端直接或间接相连，且所述LFR3的C末端与所述LCDR3的N末端直接或间接相连，所述LFR4的N末端与所述LCDR3的C末端直接或间接相连；其中，所述LFR1包含SEQ ID NO:33所示的氨基酸序列或与SEQ ID NO:33所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述LFR2包含SEQ ID NO:34所示的氨基酸序列或与SEQ ID NO:34所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述LFR3包含SEQ ID NO:35所示的氨基酸序列或与SEQ ID NO:35所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列，所述LFR4包含SEQ ID NO:36所示的氨基酸序列或与SEQ ID NO:36所示的氨基酸序列具有至少约70％序列同一性的氨基酸序列。
  72. 根据权利要求42-71中任一项所述的药物组合，其中所述抗PD-L1抗体包含VL，所述VL包含SEQ ID NO:37所示的氨基酸序列。
  73. 根据权利要求72所述的药物组合，其中所述抗PD-L1抗体包含VL，所述VL包含SEQ ID NO:38、SEQ ID NO:39或SEQ ID NO:40所示的氨基酸序列。
  74. 根据权利要求42-73中任一项所述的药物组合，其中所述抗PD-L1抗体包含VH以及VL，所述VH包含SEQ ID NO:25所示的氨基酸序列，且所述VL包含SEQ ID NO:37所示的氨基酸序列。
  75. 根据权利要求74所述的药物组合，其中所述抗PD-L1抗体包含VH以及VL，所述VH包含SEQ ID NO:25所示的氨基酸序列，且所述VL包含SEQ ID NO:38、SEQ ID NO:39或SEQ ID NO:40所示的氨基酸序列。
  76. 根据权利要求42-75中任一项所述的药物组合，其中所述抗PD-L1抗体包含LC，所述LC包含SEQ ID NO:41所示氨基酸序列。
  77. 根据权利要求76所述的药物组合，其中所述抗PD-L1抗体包含LC，所述LC包含SEQ ID NO:42、SEQ ID NO:43或SEQ ID NO:44所示氨基酸序列。
  78. 根据权利要求42-77中任一项所述的药物组合，其中所述抗PD-L1抗体包含HC和LC，所述HC包含SEQ ID NO:26所示氨基酸序列，且所述LC包含SEQ ID NO:41所示氨基 酸序列。
  79. 根据权利要求78所述的药物组合，其中所述抗PD-L1抗体包含HC和LC，所述HC包含SEQ ID NO:26所示氨基酸序列，且所述LC包含SEQ ID NO:42、SEQ ID NO:43或SEQ ID NO:44所示氨基酸序列。
  80. 根据权利要求1-79中任一项所述的药物组合，所述药物组合中的i)所述PD-1抑制剂和/或PD-L1抑制剂，与ii)所述TLR激动剂在所述药物组合中彼此不混合。
  81. 根据权利要求1-79中任一项所述的药物组合，所述药物组合中的i)所述PD-1抑制剂和/或PD-L1抑制剂，与ii)所述TLR激动剂位于单一的剂型中。
  82. 根据权利要求81所述的药物组合，其中所述药物组合被配制成药物组合物。
  83. 根据权利要求82所述的药物组合，其中所述药物组合物包括PD-1抑制剂或PD-L1抑制剂，以及TLR激动剂。
  84. 根据权利要求82-83中任一项所述的药物组合，其中所述TLR激动剂的存在量为约0.0001mg/kg至约200mg/kg。
  85. 根据权利要求82-84中任一项所述的药物组合，其中所述PD-1抑制剂或PD-L1抑制剂的存在量为约0.0001mg/kg至约200mg/kg。
  86. 根据权利要求82-85中任一项所述的药物组合，其中所述药物组合物还包含一种或多种药学上可接受的载体。
  87. 权利要求1-86中任一项所述的药物组合在制备药物中的用途，所述药物用于治疗赘生性疾病。
  88. 根据权利要求87所述的用途，其中所述赘生性疾病包括肿瘤和/或疣类疾病。
  89. 权利要求1-86中任一项所述的药物组合用于治疗赘生性疾病。
  90. 治疗赘生性疾病的药物，其包含权利要求1-86中任一项所述的药物组合。
  91. 治疗赘生性疾病的方法，其包括向有需要的受试者施用有效量的权利要求1-86中任一项所述的药物组合。
  92. 根据权利要求91所述的方法，其中所述受试者患有赘生物。
  93. 根据权利要求92所述的方法，其中所述赘生物包括肿瘤和/或疣。
  94. 根据权利要求91-93中任一项所述的方法，其中所述施用包括局部、赘生物内或全身性施用。
  95. 根据权利要求91-94中任一项所述的方法，其中所述施用包括静脉注射、静脉滴注、肌肉注射、皮下注射和/或赘生物内注射。
  96. 根据权利要求91-95中任一项所述的方法，其中采取相同或不同的给药途径施用所述药物组合中的i)PD-1抑制剂或PD-L1抑制剂，与ii)TLR激动剂。
  97. 根据权利要求91-96中任一项所述的方法，其包括向赘生物中注射所述TLR激动剂。
  98. 根据权利要求91-97中任一项所述的方法，其还包括向赘生物中注射或全身输注所述PD-1抑制剂或PD-L1抑制剂。
  99. 根据权利要求91-98中任一项所述的方法，其包括向赘生物中注射所述药物组合中的i)所述PD-1抑制剂或PD-L1抑制剂，与ii)所述TLR激动剂。
  100. 根据权利要求91-99中任一项所述的方法，其中同时或不同时施用所述药物组合中的i)PD-1抑制剂或PD-L1抑制剂，与ii)所述TLR激动剂。
  101. 根据权利要求100所述的方法，其中在所述TLR激动剂施用之前和/或施用之后施用所述PD-1抑制剂或PD-L1抑制剂。
  102. 根据权利要求101所述的方法，其中在所述TLR激动剂施用之后施用所述PD-1抑制剂或PD-L1抑制剂。
  103. 根据权利要求102所述的方法，所述方法包括：i)向赘生物中注射所述TLR激动剂；ii)在施用所述TLR激动剂之后向赘生物中注射或全身输注所述PD-1抑制剂或PD-L1抑制剂。
  104. 根据权利要求104所述的方法，所述方法包括：i)向赘生物中注射所述STING通路激动剂；ii)在施用所述STING通路激动剂之后全身输注所述PD-1抑制剂或PD-L1抑制剂。
  105. 根据权利要求100-104中任一项所述的方法，其中在所述TLR激动剂施用之后约2h至约72h施用所述PD-1抑制剂或PD-L1抑制剂。
  106. 根据权利要求100-105中任一项所述的方法，其中在所述TLR激动剂施用之后约2h，约4h，约8h，约16h，约24h，约36h，约48h，约60h，或约72h施用所述PD-1抑制剂或PD-L1抑制剂。
  107. 根据权利要求100-106中任一项所述的方法，所述方法包括：i)向赘生物中注射所述STING通路激动剂；ii)在施用所述STING通路激动剂之后约48h后向赘生物中注射或全身输注所述PD-1抑制剂或PD-L1抑制剂。
  108. 根据权利要求100所述的方法，其中所述药物组合中的i)PD-1抑制剂或PD-L1抑制剂，与ii)所述TLR激动剂同时施用。
  109. 根据权利要求108所述的方法，其中所述药物组合中的i)PD-1抑制剂或PD-L1抑制剂，与ii)所述TLR激动剂通过赘生物内注射方式同时施用，且所述PD-1抑制剂或 PD-L1抑制剂与所述TLR激动剂位于同一剂型内。
  110. 根据权利要求108所述的方法，其中所述药物组合中的i)PD-1抑制剂或PD-L1抑制剂，与ii)所述TLR激动剂通过赘生物内注射方式同时施用，且所述PD-1抑制剂或PD-L1抑制剂与所述TLR激动剂位于分开的剂型内。
  111. 药盒，其包含权利要求1-86中任一项所述的药物组合。
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