Fusion Polypeptides and Uses Thereof

Abstract

Fusion peptides and uses thereof are provided. The fusion peptide of the present application may have the function of blocking PD-1 / PD-L1 signaling and / or activating immune responses, and the present application also provides uses of the peptide.

Description

[Technical Field]

[0001] The present application relates to the field of biomedicine, and in particular to fusion polypeptides and uses thereof. [Background technology]

[0002] Immune checkpoints are a class of immune-related molecules, including inhibitory signaling molecules, that primarily function by regulating immune responses in peripheral tissues. These include immune defense, immune tolerance, and immune tissue damage. Immune checkpoint-targeting therapies aim to treat diseases by intervening in the immune system by modulating immune responses, including innate and adaptive immune responses. Tumor cells can exploit the inhibitory signaling pathways of immune checkpoints to evade the immune system. Binding of programmed death receptor-1 (PD-1) to its ligand, programmed death ligand-1 (PD-L1), is a key event in suppressing antitumor immune responses. PD-L1 is expressed on various tumor cells, including colon cancer, lung cancer, ovarian cancer, and multiple myeloma, and PD-L1 expression has been associated with the prognosis of various cancers. PD-L1-PD-1 binding can induce effects such as T cell apoptosis, suppression of immune signals, cell exhaustion, and secretion of immunosuppressive factors. As a result, the function of tumor-infiltrating T cells is neutralized, helping tumor cells evade immune system surveillance.

[0003] Currently, commercially available PD-L1-targeting inhibitors include Atezolizumab, developed by Roche; Avelumab, co-developed by Merck KGaA / Pfizer; and Durvalumab, developed by AstraZeneca. Commercially available PD-1-targeting inhibitors include Nivolumab, developed by Bristol-Myers Squibb (BMS); Pembrolizumab, developed by Merck; Camrelizumab, developed by Hengrui Pharmaceutical; Trelizumab, developed by Baekje Shenzhou; and Sintilizumab, developed by Cinda Biosciences. While these antibodies have demonstrated efficacy in tumor treatment, the average therapeutic response rate is only approximately 20%, and a significant number of tumor patients remain unresponsive to anti-PD-L1 and anti-PD-1 antibody treatments. Therefore, improving the efficacy of tumor treatments remains a critical issue in current tumor treatment. Summary of the Invention [Problem to be solved by the invention]

[0004] Given the current limitations in the effectiveness of tumor treatment based on blocking the PD-1 / PD-L1 pathway to regulate T cell activation, this application proposes developing a novel drug that coordinates PD-1 / PD-L1 pathway blockade to enhance T cell activation and applying it to clinical tumor treatment. The development of a novel drug that coordinates PD-1 / PD-L1 pathway blockade to enhance T cell activation and further improve antigen-presenting cell activation and its use in clinical tumor treatment may bring further benefits to more tumor patients. [Means for solving the problem]

[0005] The present application provides fusion peptides, which can include: (i) a portion derived from an antigen-binding fragment associated with an anti-programmed death ligand-1 (PD-L1) antibody / anti-programmed death receptor-1 (PD-1) antibody, or / and (ii) an immunoglobulin Fc domain, or / and (iii) a fusion peptide composed of the CD80 extracellular domain (ECD), or / and (iv) a fusion peptide composed of the LAG3 extracellular domain. The present application also provides polynucleotides expressing the fusion peptides. The present application also provides methods for inducing and / or enhancing immunity and treating diseases (e.g., cancer) using the fusion peptides.

[0006] The present application provides a dual- and / or multifunctional fusion peptide that simultaneously interferes with, suppresses, or blocks the PD-1 / PD-L1 signaling pathway, and / or coordinately stimulates the activation of antigen-presenting cells and / or coordinately stimulates the activation of T cells. The fusion peptide can effectively activate T cells to enhance immune responses and / or activate antigen-presenting cells, potentially resulting in tumor-killing effects. This can be used to treat diseases caused by suppressed T cell function, such as tumors, and can improve therapeutic effects, particularly in tumor patients who are unresponsive or poorly responsive to anti-PD-1 antibodies and / or anti-PD-L1 antibodies.

[0007] Meanwhile, the present application provides a fusion polypeptide, which comprises a first domain and a second domain, wherein the first domain is capable of inhibiting PD-1 / PD-L1 signaling, and the second domain comprises CD86 or a fragment thereof having functional activity.

[0008] In some embodiments, the fusion polypeptide has a first domain capable of binding to PD-L1 and / or PD-1.

[0009] In one embodiment, the fusion polypeptide comprises the first domain comprising HCDR1, HCDR2 and / or HCDR3 of an antibody heavy chain, wherein the antibody heavy chain comprises an amino acid sequence selected from the group consisting of: SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0010] In one embodiment, the fusion polypeptide comprises a heavy chain variable region (VH) of an antibody heavy chain, wherein the antibody heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15, and SEQ ID NO:17.

[0011] In one embodiment, the fusion polypeptide comprises an antibody heavy chain, and the antibody heavy chain comprises an amino acid sequence selected from the group consisting of: SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15, and SEQ ID NO:17.

[0012] In one embodiment, the fusion polypeptide comprises the first domain comprising LCDR1, LCDR2 and / or LCDR3 of an antibody light chain, wherein the antibody light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0013] In some embodiments, the fusion polypeptide includes a first domain comprising an antibody or antigen-binding fragment thereof. In some embodiments, the fusion polypeptide includes a light chain variable region (VL) of an antibody light chain, wherein the antibody light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16, and SEQ ID NO:18.

[0014] In one embodiment, the fusion polypeptide comprises a first domain comprising an antibody light chain, wherein the antibody light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16, and SEQ ID NO:18.

[0015] In some embodiments of the fusion polypeptide, the antibody is selected from the group consisting of recombinant antibodies, single domain antibodies, heavy chain antibodies, chimeric antibodies, and bispecific antibodies.

[0016] In certain embodiments, the fusion polypeptide comprises one or more antigen-binding fragments selected from the group consisting of Fab, Fab', Fv fragment, F(ab')2, F(ab)2, scFv, di-scFv, VHH, and dAb.

[0017] In one embodiment of the fusion polypeptide, the first domain comprises an antibody or antigen-binding fragment thereof selected from the group consisting of Sugemalimab, Durvalumab, Atezolizumab, Avelumab, Envafolimab, Permbrolizumab, and Nivolumab.

[0018] In one embodiment, the fusion polypeptide comprises a first domain comprising sugemalimab or an antigen-binding fragment thereof.

[0019] In one embodiment of the fusion polypeptide, the second domain is selected from the group consisting of CD86 of human origin or a functionally active fragment thereof, CD86 of mouse origin or a functionally active fragment thereof.

[0020] In one embodiment of the fusion polypeptide, the second domain is capable of binding to CD28 and / or CTLA4.

[0021] In one embodiment, the fusion polypeptide second domain comprises the IgV domain of CD86 or a functionally active fragment thereof.

[0022] In one embodiment of the fusion polypeptide, the second domain comprises the extracellular domain of CD86 or a functionally active fragment thereof.

[0023] In one embodiment of the fusion polypeptide, the second domain comprises the amino acid sequence set forth in SEQ ID NO:2 or SEQ ID NO:3.

[0024] In one embodiment, the fusion polypeptide comprises a direct or indirect link between the first domain and the second domain.

[0025] In one embodiment, the fusion polypeptide comprises a heavy chain of the first domain linked directly or indirectly to the second domain.

[0026] In one embodiment, the fusion polypeptide comprises a C-terminus of the heavy chain of the first domain linked directly or indirectly to the N-terminus of the second domain.

[0027] In one embodiment, the fusion polypeptide comprises the N-terminus of the heavy chain of the first domain linked directly or indirectly to the C-terminus of the second domain.

[0028] In one embodiment, the fusion polypeptide comprises a light chain of the first domain linked directly or indirectly to the second domain.

[0029] In one embodiment, the fusion polypeptide comprises the C-terminus of the light chain of the first domain linked directly or indirectly to the N-terminus of the second domain.

[0030] In one embodiment, the fusion polypeptide comprises the N-terminus of the light chain of the first domain linked directly or indirectly to the C-terminus of the second domain.

[0031] In one embodiment of the fusion polypeptide, the indirect connection comprises a connection via a linker.

[0032] In one embodiment of the fusion polypeptide, the linker comprises an amino acid sequence selected from the group consisting of: SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34 and SEQ ID NO:35.

[0033] In one embodiment of the fusion polypeptide, the fusion polypeptide comprises an amino acid sequence selected from the group consisting of SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, SEQ ID NO:42, SEQ ID NO:43, SEQ ID NO:44, SEQ ID NO:45, SEQ ID NO:46, SEQ ID NO:47, SEQ ID NO:48, SEQ ID NO:49, SEQ ID NO:50, SEQ ID NO:51, SEQ ID NO:52, SEQ ID NO:53, SEQ ID NO:54, SEQ ID NO:55, SEQ ID NO:56, SEQ ID NO:57, SEQ ID NO:58, SEQ ID NO:59, SEQ ID NO:60, SEQ ID NO:61, SEQ ID NO:62, SEQ ID NO:63, SEQ ID NO:64, SEQ ID NO:65, SEQ ID NO:66, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:74, SEQ ID NO:75, SEQ ID NO:76, SEQ ID NO:77, SEQ ID NO:78, SEQ ID NO:79, SEQ ID NO:80, SEQ ID NO:81, SEQ ID NO:82, SEQ ID NO:83, SEQ ID NO:84, SEQ ID NO:85, SEQ ID NO:86 and SEQ ID NO:87.

[0034] In one embodiment of the fusion polypeptide, the fusion polypeptide comprises an amino acid sequence selected from the group consisting of: SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, SEQ ID NO:42, SEQ ID NO:43, SEQ ID NO:44 and SEQ ID NO:77.

[0035] On the other hand, the present application provides a fusion polypeptide, which comprises a first domain, a second domain, and a third domain, wherein the first domain is capable of inhibiting PD-1 / PD-L1 signaling, the second domain comprises CD86 or a fragment thereof having functional activity, and the third domain is capable of activating an innate immune response.

[0036] In one embodiment of the fusion polypeptide, the third domain is capable of binding to an MHC II molecule on an antigen-presenting cell.

[0037] In one embodiment, the fusion polypeptide, the third domain comprises LAG3 or a functionally active fragment thereof.

[0038] In some embodiments, the fusion polypeptide has a first domain capable of binding to PD-L1 and / or PD-1.

[0039] In one embodiment of the fusion polypeptide, the first domain comprises HCDR1, HCDR2, and / or HCDR3 of an antibody heavy chain, wherein the antibody heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15, and SEQ ID NO:17.

[0040] In one embodiment of the fusion polypeptide, the first domain comprises a heavy chain variable region, VH, of an antibody heavy chain, wherein the antibody heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0041] In one embodiment of the fusion polypeptide, the first domain comprises an antibody heavy chain, and the antibody heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0042] In one embodiment of the fusion polypeptide, the first domain comprises LCDR1, LCDR2 and / or LCDR3 of the light chain of an antibody light chain, wherein the antibody light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0043] In one embodiment of the fusion polypeptide, the first domain comprises a light chain variable region, VL, of an antibody light chain, wherein the antibody light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0044] In one embodiment of the fusion polypeptide, the first domain comprises an antibody light chain, and the antibody light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0045] In one embodiment of the fusion polypeptide, the first domain comprises an antibody or antigen-binding fragment thereof.

[0046] In one embodiment of the fusion polypeptide, said antibody is selected from the group consisting of a recombinant antibody, a single domain antibody, a heavy chain antibody, a chimeric antibody, and a bispecific antibody.

[0047] In one embodiment of the fusion polypeptide, the antigen-binding fragment is selected from one or more of the following group: Fab, Fab', Fv fragment, F(ab')2, F(ab)2, scFv, di-scFv, VHH, and dAb.

[0048] In one embodiment of the fusion polypeptide, the first domain comprises an antibody or antigen-binding fragment thereof selected from the group consisting of sugemalimab, durvalumab, atezolizumab, avelumab, embafolimab, permbrolizumab, and nivolumab.

[0049] In one embodiment of the fusion polypeptide, the first domain comprises sugemalimab or an antigen-binding fragment thereof.

[0050] In one embodiment of the fusion polypeptide, said second domain is selected from the group consisting of CD86 of human origin or a functionally active fragment thereof, and CD86 of mouse origin or a functionally active fragment thereof.

[0051] In one embodiment of the fusion polypeptide, the second domain is capable of binding to CD28 and / or CTLA4.

[0052] In one embodiment of the fusion polypeptide, said second domain comprises the IgV domain of CD86 or a functionally active fragment thereof.

[0053] In one embodiment of the fusion polypeptide, said second domain comprises the extracellular domain of CD86 or a functionally active fragment thereof.

[0054] In one embodiment of the fusion polypeptide, the second domain comprises the amino acid sequence set forth in SEQ ID NO:2 or SEQ ID NO:3.

[0055] In one embodiment of the fusion polypeptide, said third domain is selected from the group consisting of LAG3 of human origin, or a functionally active fragment thereof, and LAG3 of mouse origin, or a functionally active fragment thereof.

[0056] In one embodiment of the fusion polypeptide, the third domain comprises the extracellular domain of LAG3 or a functionally active fragment thereof.

[0057] In one embodiment of the fusion polypeptide, said third domain comprises IgD1, IgD2, IgD3 and / or IgD4 of LAG3 or a functionally active fragment thereof.

[0058] In one embodiment of the fusion polypeptide, the third domain comprises IgD1, IgD1-IgD2, IgD1-IgD2-IgD3, and / or IgD1-IgD2-IgD3-IgD4 of LAG3 or a functionally active fragment thereof.

[0059] In one embodiment of the fusion polypeptide, the third domain comprises an amino acid sequence selected from the group consisting of SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27 and SEQ ID NO:28.

[0060] In one embodiment, the fusion polypeptide comprises a direct or indirect link between the first domain and the second domain.

[0061] In one embodiment, the fusion polypeptide comprises a heavy chain of the first domain linked directly or indirectly to the second domain.

[0062] In one embodiment, the fusion polypeptide comprises a C-terminus of the heavy chain of the first domain linked directly or indirectly to the N-terminus of the second domain.

[0063] In one embodiment, the fusion polypeptide comprises the N-terminus of the heavy chain of the first domain linked directly or indirectly to the C-terminus of the second domain.

[0064] In one embodiment, the fusion polypeptide comprises a light chain of the first domain linked directly or indirectly to the second domain. In one embodiment, the fusion polypeptide comprises the C-terminus of the light chain of the first domain linked directly or indirectly to the N-terminus of the second domain.

[0065] In one embodiment, the fusion polypeptide comprises the N-terminus of the light chain of the first domain linked directly or indirectly to the C-terminus of the second domain.

[0066] In one embodiment, the fusion polypeptide comprises a first domain, a heavy chain, and a second domain, wherein the C-terminus of the heavy chain of the first domain is linked to the N-terminus of the second domain, and the light chain and the C-terminus of the light chain of the first domain are linked to the N-terminus of the second domain.

[0067] In one embodiment, the fusion polypeptide comprises a first domain, a heavy chain, and a second domain, wherein the N-terminus of the heavy chain of the first domain is linked to the C-terminus of the second domain, and the light chain of the first domain is linked to the C-terminus of the second domain.

[0068] In one embodiment, the fusion polypeptide comprises a direct or indirect link between the first domain and the third domain.

[0069] In one embodiment, the fusion polypeptide comprises a heavy chain of the first domain linked directly or indirectly to the third domain.

[0070] In one embodiment, the fusion polypeptide comprises the C-terminus of the heavy chain of the first domain linked directly or indirectly to the N-terminus of the third domain.

[0071] In one embodiment, the fusion polypeptide comprises the N-terminus of the heavy chain of the first domain linked directly or indirectly to the C-terminus of the third domain.

[0072] In one embodiment, the fusion polypeptide comprises a light chain of the first domain linked directly or indirectly to the third domain.

[0073] In one embodiment, the fusion polypeptide comprises a C-terminus of the light chain of the first domain linked directly or indirectly to the N-terminus of the third domain.

[0074] In one embodiment, the fusion polypeptide comprises the N-terminus of the light chain of the first domain linked directly or indirectly to the C-terminus of the third domain.

[0075] In one embodiment, the fusion polypeptide includes a direct or indirect link between the second domain and the third domain.

[0076] In one embodiment, the fusion polypeptide has the C-terminus of the second domain linked directly or indirectly to the N-terminus of the third domain.

[0077] In one embodiment, the fusion polypeptide has the N-terminus of the second domain linked directly or indirectly to the C-terminus of the third domain.

[0078] In one embodiment, the fusion polypeptide has the first domain linked directly or indirectly to the second domain, which is further linked directly or indirectly to the third domain.

[0079] In one embodiment, the fusion polypeptide comprises a first domain whose C-terminus is linked, directly or indirectly, to the N-terminus of the second domain, and a second domain whose C-terminus is linked, directly or indirectly, to the N-terminus of the third domain.

[0080] In one embodiment, the fusion polypeptide comprises a first domain, wherein the C-terminus of the heavy chain of the first domain is linked, directly or indirectly, to the N-terminus of the second domain, and a second domain, wherein the C-terminus of the second domain is linked, directly or indirectly, to the N-terminus of the third domain.

[0081] In one embodiment, the fusion polypeptide comprises a first domain whose light chain C-terminus is linked, directly or indirectly, to the second domain's N-terminus, and a second domain whose light chain C-terminus is linked, directly or indirectly, to the third domain's N-terminus.

[0082] In one embodiment, the fusion polypeptide comprises a first domain whose N-terminus is linked, directly or indirectly, to a second domain whose C-terminus is linked, directly or indirectly, to a third domain whose N-terminus is linked, directly or indirectly, to a third domain whose C-terminus is linked, directly or indirectly, to a third domain.

[0083] In one embodiment, the fusion polypeptide comprises a first domain, wherein the N-terminus of the heavy chain of the first domain is linked, directly or indirectly, to the C-terminus of the second domain, and a second domain, wherein the N-terminus of the second domain is linked, directly or indirectly, to the C-terminus of the third domain.

[0084] In one embodiment, the fusion polypeptide comprises a first domain whose light chain is linked, directly or indirectly, at its N-terminus to the C-terminus of the second domain, and a second domain whose light chain is linked, directly or indirectly, at its N-terminus to the C-terminus of the third domain.

[0085] In one embodiment, the fusion polypeptide has the first domain linked directly or indirectly to the third domain, which is further linked directly or indirectly to the second domain.

[0086] In one embodiment, the fusion polypeptide has the C-terminus of the first domain linked directly or indirectly to the N-terminus of the third domain, and the C-terminus of the third domain linked directly or indirectly to the N-terminus of the second domain.

[0087] In one embodiment, the fusion polypeptide comprises a first domain, wherein the C-terminus of the heavy chain of the first domain is linked, directly or indirectly, to the N-terminus of the third domain, and a third domain, wherein the C-terminus of the third domain is linked, directly or indirectly, to the N-terminus of the second domain.

[0088] In one embodiment, the fusion polypeptide comprises a first domain whose light chain C-terminus is linked, directly or indirectly, to the N-terminus of the third domain, and a third domain whose C-terminus is linked, directly or indirectly, to the N-terminus of the second domain.

[0089] In one embodiment, the fusion polypeptide comprises the first domain having an N-terminus linked directly or indirectly to the C-terminus of the third domain, and the third domain having an N-terminus linked directly or indirectly to the C-terminus of the second domain.

[0090] In one embodiment, the fusion polypeptide comprises a first domain, wherein the N-terminus of the heavy chain of the first domain is linked, directly or indirectly, to the C-terminus of the third domain, and a third domain, wherein the N-terminus of the third domain is linked, directly or indirectly, to the C-terminus of the second domain.

[0091] In one embodiment, the fusion polypeptide comprises a first domain whose light chain is linked, directly or indirectly, at its N-terminus to the C-terminus of the third domain, and a third domain whose light chain is linked, directly or indirectly, at its N-terminus to the C-terminus of the second domain.

[0092] In one embodiment, the fusion polypeptide has the first domain linked directly or indirectly to the second domain, and the first domain linked directly or indirectly to the third domain.

[0093] In one embodiment, the fusion polypeptide comprises a first domain whose C-terminus is linked, directly or indirectly, to the second domain whose N-terminus is linked, directly or indirectly, to the third domain whose C-terminus is linked, directly or indirectly, to the third domain.

[0094] In one embodiment, the fusion polypeptide comprises a first domain, wherein the C-terminus of the heavy chain of the first domain is linked, directly or indirectly, to the N-terminus of the second domain, and a first domain, wherein the N-terminus of the heavy chain of the first domain is linked, directly or indirectly, to the C-terminus of the third domain.

[0095] In one embodiment, the fusion polypeptide comprises a first domain, a light chain, and a third domain, wherein the C-terminus of the light chain of the first domain is linked, directly or indirectly, to the N-terminus of the second domain, and the N-terminus of the light chain of the first domain is linked, directly or indirectly, to the C-terminus of the third domain.

[0096] In one embodiment, the fusion polypeptide comprises a first domain, wherein the C-terminus of the heavy chain is directly or indirectly linked to the N-terminus of the second domain, and a first domain, wherein the N-terminus of the light chain is directly or indirectly linked to the C-terminus of the third domain.

[0097] In one embodiment, the fusion polypeptide comprises a first domain, a light chain, and a heavy chain, the C-terminus of which is linked directly or indirectly to the N-terminus of the second domain, and a first domain, a heavy chain, and a third domain, the C-terminus of which is linked directly or indirectly to the N-terminus of the third domain.

[0098] In one embodiment, the fusion polypeptide comprises a first domain whose N-terminus is linked, directly or indirectly, to a second domain whose C-terminus is linked, directly or indirectly, to a third domain whose N-terminus is linked, directly or indirectly, to a third ... and indirectly, to a third domain whose C-terminus is linked, and indirectly, to a third domain whose N-terminus is linked, and indirectly, to a third

[0099] In one embodiment, the fusion polypeptide comprises a first domain, wherein the N-terminus of the heavy chain of the first domain is linked, directly or indirectly, to the C-terminus of the second domain, and a first domain, wherein the C-terminus of the heavy chain of the first domain is linked, directly or indirectly, to the N-terminus of the third domain.

[0100] In one embodiment, the fusion polypeptide comprises a first domain, a light chain, and a third domain, wherein the N-terminus of the light chain of the first domain is linked, directly or indirectly, to the C-terminus of the second domain, and the C-terminus of the light chain of the first domain is linked, directly or indirectly, to the N-terminus of the third domain.

[0101] In one embodiment, the fusion polypeptide comprises a first domain, wherein the N-terminus of the heavy chain is directly or indirectly linked to the C-terminus of the second domain, and a first domain, wherein the C-terminus of the light chain is directly or indirectly linked to the N-terminus of the third domain.

[0102] In one embodiment, the fusion polypeptide comprises the N-terminus of the light chain of the first domain linked directly or indirectly to the C-terminus of the second domain, and the C-terminus of the heavy chain of the first domain linked directly or indirectly to the N-terminus of the third domain.

[0103] In one embodiment of the fusion polypeptide, the indirect connection comprises a connection via a linker.

[0104] In one embodiment of the fusion polypeptide, the linker is selected from the amino acid sequences set forth in the following group: SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34 and SEQ ID NO:35.

[0105] In one embodiment, the fusion polypeptide is selected from the amino acid sequences set forth in the following group: SEQ ID NO:88, SEQ ID NO:89, SEQ ID NO:90, SEQ ID NO:91, SEQ ID NO:92, SEQ ID NO:93, SEQ ID NO:94, SEQ ID NO:95, SEQ ID NO:96, SEQ ID NO:97, SEQ ID NO:98, SEQ ID NO:99, SEQ ID NO:100, SEQ ID NO:101, SEQ ID NO:102, SEQ ID NO:103, SEQ ID NO:104, SEQ ID NO:105, SEQ ID NO:106, SEQ ID NO:107, SEQ ID NO:108, SEQ ID NO:109, SEQ ID NO: Number 110, SEQ ID NO:111, SEQ ID NO:112, SEQ ID NO:113, SEQ ID NO:114, SEQ ID NO:115, SEQ ID NO:116, SEQ ID NO:117, SEQ ID NO:118, SEQ ID NO:119, SEQ ID NO:120, SEQ ID NO:121, SEQ ID NO:122, SEQ ID NO:123, SEQ ID NO:124, SEQ ID NO:125, SEQ ID NO:126, SEQ ID NO:127, SEQ ID NO:128, SEQ ID NO:129, SEQ ID NO:130, SEQ ID NO:131, SEQ ID NO:132, SEQ ID NO:133, SEQ ID NO:134, SEQ ID NO:135, SEQ ID NO:171, SEQ ID NO:173.

[0106] In one embodiment, the fusion polypeptide is selected from the amino acid sequences set forth in the following group: SEQ ID NO:104, SEQ ID NO:107, SEQ ID NO:108, SEQ ID NO:112, SEQ ID NO:114, SEQ ID NO:115, SEQ ID NO:116, SEQ ID NO:171, SEQ ID NO:173.

[0107] On the other hand, the present application provides CD86 mutant polypeptides, which comprise amino acid substitution mutations of humanized CD86.

[0108] In one embodiment, the CD86 variant polypeptide comprises an amino acid substitution variant of the extracellular IgV domain of humanized CD86.

[0109] In one embodiment, the CD86 mutant polypeptide comprises an amino acid substitution mutant in the IgV domain of CD86, and the mutation sites of the amino acid substitution mutant in the IgV domain of CD86 include one or more of the amino acid site mutations shown in the following group: A13I, A13L, A13F, A13M, Q25A, Q25I, Q25V, Q25F, Q25M, F33A, F33L, F33V, F33M, M60R, I89A, I89L, I89V, I89F, I89M, H90I, H90V, H90M, and H90F. In this specification, unless otherwise clearly stated, the designation of the mutation site is based on the wild-type sequence. For example, "the mutation site A13I of the amino acid substitution mutant in the IgV domain of CD86" indicates that the 13th amino acid A in the IgV domain of wild-type CD86 has been mutated to I.

[0110] In one embodiment of a CD86 variant polypeptide, the CD86 variant polypeptide comprises an amino acid substitution variant of the IgV domain of CD86, wherein the amino acid substitution variant comprises a combination selected from the group consisting of Q25I / F33L / H90I, Q25V / F33L / H90I, Q25I / F33V / H90I, Q25V / F33V / H90I, Q25I / F33V / H90I, F33L / H90V, Q25V / F33L / H90V, Q25I / F33V / H90V, Q25V / F33V / H90V, A13L / Q25I / F33L / H90I, A13I / Q25I / F33L / H90I, A13L / Q25V / F33L / H90I, A13I / Q25V / F33L / H90I, Q25I / F33L / I89L / H90I, Q25I / F33L / M60R / H90I, Q25V / F33L / I89L / H90I, Q25V / F33L / M60R / H90I, Q25F / F33L / H90I, Q25I / F33L / H90F, Q25I / F33A / H90I, Q25I / H90I, Q25I, H90I, Q25V / H90V, Q25V / H90I, Q25F / H90I, Q25F / H90V, A13F / Q25I / F33L / H90I, A13M / Q25I / F33L / H90I, Q25A / F33L / H90I, Q25M / F33L / H90I, Q25I / F33M / H90I, Q25I / F33L / I89V / H90I, Q25I / F33L / I89F / H90I, Q25I / F33L / I89M / H90I sum Q25I / F33L / H90M.

[0111] In one embodiment, the CD86 mutant polypeptide comprises the amino acid substitution mutants Q25I / F33L / H90I in the IgV domain of CD86.

[0112] In one embodiment, the CD86 variant polypeptide comprises an amino acid substitution variant of the extracellular domain of humanized CD86.

[0113] In one embodiment, the CD86 mutant polypeptide comprises an amino acid substitution mutant of the extracellular domain of CD86, wherein the amino acid substitution mutant of the extracellular domain of CD86 comprises one or more mutations at amino acid positions shown in the following group: A13I, A13L, A13F, A13M, Q25A, Q25I, Q25V, Q25F, Q25M, F33A, F33L, F33V, F33M, M60R, I89A, I89L, I89V, I89F, I89M, H90I, H90V, H90M, H90F.

[0114] In one embodiment, the CD86 variant polypeptide comprises an amino acid substitution variant of the extracellular domain of CD86, the amino acid substitution variants comprising a combination selected from the following group: Q25I / F33L / H90I, Q25V / F33L / H90I, Q25I / F33V / H90I, Q25V / F33V / H90I, Q25I / F33 L / H90V, Q25V / F33L / H90V, Q25I / F33V / H90V, Q25V / F33V / H90V, A13L / Q25I / F33L / H90I, A13I / Q2 5I / F33L / H90I, A13L / Q25V / F33L / H90I, A13I / Q25V / F33L / H90I, Q25I / F33L / I89L / H90I, Q25I / F3 3L / M60R / H90I, Q25V / F33L / I89L / H90I, Q25V / F33L / M60R / H90I, Q25F / F33L / H90I, Q25I / F33L / H 90F, Q25I / F33A / H90I, Q25I / H90I, Q25I, H90I, Q25V / H90V, Q25V / H90I, Q25F / H90I, Q25F / H90V, A 13F / Q25I / F33L / H90I, A13M / Q25I / F33L / H90I, Q25A / F33L / H90I, Q25M / F33L / H90I, Q25I / F33M / H90I, Q25I / F33L / I89V / H90I, Q25I / F33L / I89F / H90I, Q25I / F33L / I89M / H90I sum Q25I / F33L / H90M.

[0115] In one embodiment, the CD86 mutant polypeptide comprises the amino acid substitution mutants Q25I / F33L / H90I in the extracellular domain of CD86.

[0116] In one embodiment, the CD86 variant polypeptide comprises an amino acid sequence selected from SEQ ID NO:178 to SEQ ID NO:243.

[0117] On the other hand, the present application provides a fusion polypeptide, the fusion polypeptide comprising a first domain and a second domain, wherein the first domain comprises a CD86 variant polypeptide described in the present application, and the second domain comprises an antibody or antigen-binding fragment thereof, or an immunoglobulin structural domain.

[0118] In one embodiment of the fusion polypeptide, the second domain comprises an antibody or antigen-binding fragment thereof.

[0119] In one embodiment, the fusion polypeptide, the second domain is capable of binding to one or more of the targets set forth in the following group: PD-L1, PD-L2, PD-1, OX40, 4-1BB, ICOS, TIGIT, CTLA4, LAG3, CD3, VEGF, VEGFR, CD47, HGF, Trop2, EpCAM, CCR8, CCR4, CCR5, GPRC5D, BCMA, CD19, CD20, HER-2 neu, DLL1, HER-3, HER-4, EGFR, PSMA, CEA, MUC-1 (mucin), MUC2, MUC3, MUC4, MUC5AC, MUC5B, MUC7, CD123, CD33, CD30, CD38, NKG2A, Nkp36, and Tim3.

[0120] In one embodiment of the fusion polypeptide, the second domain is capable of binding to PD-L1, PD-1 and / or PD-L2.

[0121] In one embodiment, in the fusion polypeptide, the second domain comprises HCDR1, HCDR2 and / or HCDR3 of an antibody heavy chain, and the antibody heavy chain comprises an amino acid sequence selected from the following group: SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0122] In one embodiment of the fusion polypeptide, the second domain comprises a heavy chain variable region VH of an antibody heavy chain, and the antibody heavy chain comprises an amino acid sequence selected from the following group: SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0123] In one embodiment, the fusion polypeptide comprises an antibody heavy chain, and the antibody heavy chain comprises an amino acid sequence selected from the following group: SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15, and SEQ ID NO:17.

[0124] In one embodiment, in the fusion polypeptide, the second domain comprises LCDR1, LCDR2 and / or LCDR3 of an antibody light chain, and the antibody light chain comprises an amino acid sequence selected from the following group: SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0125] In one embodiment of the fusion polypeptide, the second domain comprises a light chain variable region VL of an antibody light chain, wherein the antibody light chain comprises an amino acid sequence selected from the following group: SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0126] In one embodiment, the fusion polypeptide comprises an antibody light chain, and the antibody light chain comprises an amino acid sequence selected from the following group: SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16, and SEQ ID NO:18.

[0127] In one embodiment of the fusion polypeptide, the second domain comprises an antibody or antigen-binding fragment thereof from the following group: Sugemalimab, Durvalumab, Atezolizumab, Avelumab, Envafolimab, Permbrolizumab, and Nivolumab.

[0128] In one embodiment of the fusion polypeptide, the second domain comprises sugemalimab or an antigen-binding fragment thereof.

[0129] In one embodiment of the fusion polypeptide, the second domain is capable of binding to CD3.

[0130] In one embodiment of the fusion polypeptide, the second domain comprises a CD3B219 antibody or an antigen-binding fragment thereof.

[0131] In one embodiment of the fusion polypeptide, the second domain comprises HCDR1, HCDR2 and / or HCDR3 of an antibody heavy chain, wherein the antibody heavy chain comprises the amino acid sequence set forth in SEQ ID NO:616.

[0132] In one embodiment of the fusion polypeptide, the second domain comprises a heavy chain variable region, VH, of an antibody, wherein the antibody heavy chain comprises the amino acid sequence set forth in SEQ ID NO:616.

[0133] In one embodiment of the fusion polypeptide, the second domain comprises a heavy chain of an antibody, and the antibody heavy chain comprises the amino acid sequence set forth in SEQ ID NO:616.

[0134] In one embodiment, the fusion polypeptide comprises the second domain comprising LCDR1, LCDR2 and / or LCDR3 of an antibody light chain, wherein the antibody light chain comprises the amino acid sequence set forth in SEQ ID NO:617.

[0135] In one embodiment of the fusion polypeptide, the second domain comprises a light chain variable region, VL, of an antibody, wherein the antibody light chain comprises the amino acid sequence set forth in SEQ ID NO:617.

[0136] In one embodiment of the fusion polypeptide, the second domain comprises a light chain of an antibody, and the antibody light chain comprises the amino acid sequence set forth in SEQ ID NO:617.

[0137] In one embodiment of the fusion polypeptide, the antigen-binding fragment is selected from one or more of the following group: Fab, Fab', Fv fragment, F(ab')2, F(ab)2, scFv, di-scFv, VHH, and dAb.

[0138] In one embodiment of the fusion polypeptide, the second domain comprises an immunoglobulin structural domain.

[0139] In one embodiment of the fusion polypeptide, the second domain comprises an immunoglobulin IgG antibody.

[0140] In one embodiment, the fusion polypeptide comprises one or more of the following: human IgG1, human IgG2, human IgG3, human IgG4, mouse IgG1, mouse IgG2a, mouse IgG2b, and mouse IgG3.

[0141] In one embodiment of the fusion polypeptide, the second domain comprises an immunoglobulin Fc structural domain.

[0142] In one embodiment of the fusion polypeptide, the second domain comprises an Fc structural domain of an immunoglobulin IgG antibody.

[0143] In one embodiment of the fusion polypeptide, the second domain comprises an Fc structural domain of an immunoglobulin IgG antibody, and the Fc structural domain of the immunoglobulin IgG antibody is selected from the following: a human IgG1 Fc structural domain, a human IgG2 Fc structural domain, a human IgG3 Fc structural domain, a human IgG4 Fc structural domain, a mouse IgG1 Fc structural domain, a mouse IgG2a Fc structural domain, a mouse IgG2b Fc structural domain, or a mouse IgG3 Fc structural domain.

[0144] In one embodiment of the fusion polypeptide, the second domain comprises an Fc structural domain of human-derived immunoglobulin IgG4.

[0145] In one embodiment of the fusion polypeptide, the second domain is selected from the amino acid sequences set forth in the following group: SEQ ID NO: 4 to SEQ ID NO: 18, SEQ ID NO: 29 to SEQ ID NO: 30, SEQ ID NO: 634 to SEQ ID NO: 639, SEQ ID NO: 616 and SEQ ID NO: 617.

[0146] In one embodiment, the fusion polypeptide has the first domain linked directly or indirectly to the second domain.

[0147] In one embodiment, the fusion polypeptide comprises the first domain linked directly or indirectly to the N-terminus of the second domain.

[0148] In one embodiment of the fusion polypeptide, the first domain is linked directly or indirectly to the C-terminus of the second domain.

[0149] In one embodiment of the fusion polypeptide, the indirect connection comprises a connection via a linker.

[0150] In one embodiment of the fusion polypeptide, the linker is selected from the amino acid sequences set forth in the following group: SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34 and SEQ ID NO:35.

[0151] In one embodiment, the fusion polypeptide is selected from the amino acid sequences set forth in the following group: SEQ ID NO: 350 to SEQ ID NO: 382, ​​SEQ ID NO: 384 to SEQ ID NO: 416, SEQ ID NO: 418 to SEQ ID NO: 450, SEQ ID NO: 451 to SEQ ID NO: 483, SEQ ID NO: 484 to SEQ ID NO: 615, SEQ ID NO: 618 to SEQ ID NO: 624, and SEQ ID NO: 626 to SEQ ID NO: 632.

[0152] Meanwhile, the present application provides a fusion polypeptide, which comprises a first domain, a second domain, and a third domain, wherein the first domain comprises a CD86 variant disclosed herein, the second domain comprises an antibody or an antigen-binding fragment thereof, or an immunoglobulin antibody, and the third domain is the same or different from the second domain, an antigen-binding fragment thereof, or a functional protein or an active fragment thereof.

[0153] In some embodiments, the second domain comprises an antibody or an antigen-binding fragment thereof.

[0154] In some embodiments, the second domain is capable of binding to one or more of the targets set forth in the following group: PD-L1, PD-L2, PD-1, OX40, 4-1BB, ICOS, TIGIT, CTLA4, LAG3, CD3, VEGF, VEGFR, CD47, HGF, Trop2, EpCAM, CCR8, CCR4, CCR5, GPRC5D, BCMA, CD19, CD20, HER-2 neu, DLL1, HER-3, HER-4, EGFR, PSMA, CEA, MUC-1 (mucin), MUC2, MUC3, MUC4, MUC5AC, MUC5B, MUC7, CD123, CD33, CD30, CD38, NKG2A, Nkp36, and Tim3.

[0155] In some embodiments, the second domain is capable of binding to PD-L1, PD-1 and / or PD-L2.

[0156] In some embodiments, the second domain comprises HCDR1, HCDR2 and / or HCDR3 of an antibody heavy chain, and the antibody heavy chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0157] In some embodiments, the second domain comprises a heavy chain variable region VH of an antibody heavy chain, and the antibody heavy chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0158] In some embodiments, the second domain comprises an antibody heavy chain, and the antibody heavy chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15, and SEQ ID NO:17.

[0159] In some embodiments, the second domain comprises LCDR1, LCDR2 and / or LCDR3 of an antibody light chain, and the antibody light chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0160] In some embodiments, the second domain comprises a light chain variable region VL of an antibody light chain, and the antibody light chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16, and SEQ ID NO:18.

[0161] In some embodiments, the second domain comprises an antibody light chain, and the antibody light chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16, and SEQ ID NO:18.

[0162] In one embodiment of the fusion polypeptide, the second domain comprises an antibody or antigen-binding fragment thereof selected from the group consisting of Sugemalimab, Durvalumab, Atezolizumab, Avelumab, Envafolimab, Permbrolizumab, and Nivolumab.

[0163] In one embodiment of the fusion polypeptide, the second domain comprises sugemalimab or an antigen-binding fragment thereof.

[0164] In one embodiment of the fusion polypeptide, the second domain is capable of binding to CD3.

[0165] In one embodiment of the fusion polypeptide, the second domain comprises a CD3B219 antibody or an antigen-binding fragment thereof.

[0166] In one embodiment of the fusion polypeptide, the second domain comprises HCDR1, HCDR2 and / or HCDR3 of an antibody heavy chain, wherein the antibody heavy chain comprises the amino acid sequence set forth in SEQ ID NO:616.

[0167] In one embodiment of the fusion polypeptide, the second domain comprises a heavy chain variable region, VH, of an antibody, wherein the antibody heavy chain comprises the amino acid sequence set forth in SEQ ID NO:616.

[0168] In one embodiment of the fusion polypeptide, the second domain comprises a heavy chain of an antibody, and the antibody heavy chain comprises the amino acid sequence set forth in SEQ ID NO:616.

[0169] In one embodiment, the fusion polypeptide comprises the second domain comprising LCDR1, LCDR2 and / or LCDR3 of an antibody light chain, wherein the antibody light chain comprises the amino acid sequence set forth in SEQ ID NO:617.

[0170] In one embodiment of the fusion polypeptide, the second domain comprises a light chain variable region, VL, of an antibody, wherein the antibody light chain comprises the amino acid sequence set forth in SEQ ID NO:617.

[0171] In some embodiments of the fusion polypeptide, the second domain comprises a light chain of an antibody, and the antibody light chain comprises the amino acid sequence set forth in SEQ ID NO: 617. In some embodiments of the fusion polypeptide, the antibody is selected from the group consisting of immunoglobulin antibodies, recombinant antibodies, chimeric antibodies, heavy chain antibodies, single domain antibodies, and bispecific antibodies.

[0172] In certain embodiments, in the fusion polypeptide, the antigen-binding fragment is selected from one or more of a Fab, a Fab', an Fv fragment, a F(ab')2, a F(ab)2, a scFv, a di-scFv, a VHH, and a dAb.

[0173] In some embodiments, the second domain in the fusion polypeptide can comprise an immunoglobulin domain.

[0174] In certain embodiments, in the fusion polypeptide, the second domain comprises an immunoglobulin IgG antibody, and the immunoglobulin IgG antibody can comprise one or more of human IgG1, human IgG2, human IgG3, human IgG4, mouse IgG1, mouse IgG2a, mouse IgG2b, and mouse IgG3.

[0175] In some embodiments, the second domain in the fusion polypeptide can comprise an immunoglobulin Fc domain.

[0176] In some embodiments, the second domain in the fusion polypeptide can comprise the Fc domain of an immunoglobulin IgG antibody.

[0177] In some embodiments, in the fusion polypeptide, the second domain comprises an Fc domain of an immunoglobulin IgG antibody, and the Fc domain of the immunoglobulin IgG antibody can comprise a human IgG1 Fc domain, a human IgG2 Fc domain, a human IgG3 Fc domain, a human IgG4 Fc domain, a mouse IgG1 Fc domain, a mouse IgG2a Fc domain, a mouse IgG2b Fc domain, or a mouse IgG3 Fc domain.

[0178] In some embodiments, the second domain of the fusion polypeptide can comprise the Fc domain of human immunoglobulin IgG4.

[0179] In some embodiments of the fusion polypeptide, the third domain is a functional protein or an active fragment thereof, wherein the functional protein or an active fragment thereof is capable of activating an innate immune response.

[0180] In some embodiments, the third domain of the fusion polypeptide is capable of binding to an MHC II molecule on an antigen-presenting cell.

[0181] In some embodiments, in the fusion polypeptide, the third domain can comprise LAG3 or a functionally active fragment thereof.

[0182] In one embodiment of the fusion polypeptide, the third domain is selected from the group consisting of: LAG3 of human origin, or a functionally active fragment thereof, and LAG3 of mouse origin, or a functionally active fragment thereof.

[0183] In some embodiments, the third domain in the fusion polypeptide can comprise the extracellular domain of LAG3 or a functionally active fragment thereof.

[0184] In one embodiment of the fusion polypeptide, the third domain can comprise IgD1, IgD2, IgD3 and / or IgD4 of LAG3 or functionally active fragments thereof.

[0185] In certain embodiments, in the fusion polypeptide, the third domain can comprise IgD1, IgD1-IgD2, IgD1-IgD2-IgD3 and / or IgD1-IgD2-IgD3-IgD4 of LAG3, or functionally active fragments thereof.

[0186] In some embodiments, in the fusion polypeptide, the third domain can comprise a LAG3 mutant polypeptide.

[0187] In one embodiment of the fusion polypeptide, the third domain comprises a LAG3 variant polypeptide, which comprises a truncation and / or mutation based on human-derived LAG3.

[0188] In one embodiment of the fusion polypeptide, the third domain comprises a LAG3 mutant polypeptide, which comprises a truncation based on the extracellular domain of wild-type human-derived LAG3.

[0189] In one embodiment of the fusion polypeptide, the third domain comprises a LAG3 mutant polypeptide, which is based on the extracellular domain polypeptide of wild-type human-derived LAG3 and has an N-terminus shortened by 0 to 124 amino acids.

[0190] In one embodiment, the fusion polypeptide comprises a third domain comprising a LAG3 mutant polypeptide, which is based on the extracellular domain polypeptide of wild-type human-derived LAG3 and is N-terminally shortened by 0, 5, 21, 37, 45, 60, 71, 74, 79, 84, 89, 94, 99, 104, 109, or 114 amino acids.

[0191] In one embodiment, the fusion polypeptide comprises a third domain comprising a LAG3 mutant polypeptide, which is based on the extracellular domain polypeptide of wild-type human-derived LAG3 and is N-terminally shortened by 0, 5, 21, 37, 45, 60, 71, or 74 amino acids.

[0192] In one embodiment, the fusion polypeptide comprises a third domain comprising a LAG3 mutant polypeptide, which is based on the extracellular domain polypeptide of wild-type human-derived LAG3 and whose C-terminus terminates between amino acid positions 122 and 167.

[0193] In one embodiment, the fusion polypeptide comprises a third domain comprising a LAG3 mutant polypeptide based on the extracellular domain of wild-type human LAG3, the C-terminus of which terminates at amino acid position 122, 129, 136, 146, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, or 167.

[0194] In one embodiment, the fusion polypeptide comprises a third domain comprising a LAG3 mutant polypeptide based on the extracellular domain of wild-type human LAG3, and the C-terminus of the LAG3 mutant polypeptide terminates at amino acid position 122, 129, 136, 146, 156, 161, or 167.

[0195] In one embodiment, the fusion polypeptide comprises a third domain comprising a LAG3 mutant polypeptide based on the extracellular domain of wild-type human-derived LAG3, the C-terminus of which terminates at amino acid position 156, 161, or 167.

[0196] In one embodiment, the fusion polypeptide comprises a third domain comprising a LAG3 mutant polypeptide, which is based on the extracellular domain polypeptide of wild-type human-derived LAG3 and is N-terminally shortened by 0, 5, 21, 37, 45, 60, 71, or 74 amino acids, and C-terminally terminates at amino acid position 122, 129, 136, 146, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, or 167.

[0197] In one embodiment of the fusion polypeptide, the third domain comprises a LAG3 mutant polypeptide, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site.

[0198] In one embodiment of the fusion polypeptide, the third domain comprises a LAG3 mutant polypeptide, the LAG3 mutant polypeptide comprising a mutation at an amino acid site, the mutation site being the 97th Arg amino acid.

[0199] In one embodiment, the fusion polypeptide comprises a third domain comprising a LAG3 mutant polypeptide, the LAG3 mutant polypeptide comprising a mutation at an amino acid site, where the 97th Arg amino acid is mutated to a Glu amino acid.

[0200] In one embodiment of the fusion polypeptide, the third domain comprises an amino acid sequence selected from SEQ ID NO:20 to SEQ ID NO:28 and SEQ ID NO:244 to SEQ ID NO:349.

[0201] In one embodiment, the fusion polypeptide has the first domain linked directly or indirectly to the second domain.

[0202] In one embodiment, the fusion polypeptide has the first domain linked directly or indirectly to the third domain.

[0203] In one embodiment, the fusion polypeptide has the second domain linked directly or indirectly to the third domain.

[0204] In one embodiment, the fusion polypeptide has the first domain linked directly or indirectly to the second domain, and the first domain linked directly or indirectly to the third domain.

[0205] In one embodiment, the fusion polypeptide has the first domain linked directly or indirectly to the second domain, which is further linked directly or indirectly to the third domain.

[0206] In one embodiment, the fusion polypeptide has the first domain linked directly or indirectly to the third domain, and the second domain linked directly or indirectly to the third domain.

[0207] In one embodiment of the fusion polypeptide, the indirect connection comprises a connection via a linker.

[0208] In one embodiment of the fusion polypeptide, the linker comprises an amino acid sequence selected from the amino acid sequences set forth in SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34 and SEQ ID NO:35.

[0209] In one embodiment, the fusion polypeptide comprises an amino acid sequence selected from SEQ ID NO:751 to SEQ ID NO:1024, 1249, 1251, 1253 and 1255.

[0210] The present application provides LAG3 mutant polypeptides, which contain truncations and / or mutations based on human-derived LAG3. The present application also provides methods for innovatively modifying LAG3 polypeptides to obtain functional LAG3 mutant polypeptides, and methods for using the same to regulate the immune response of antigen-presenting cells, thereby enabling the treatment or prevention of diseases such as infections and tumors caused by suppressed immune cell function.

[0211] In one embodiment, the LAG3 variant polypeptide comprises a truncation based on the extracellular domain of wild-type human-derived LAG3.

[0212] In one embodiment, the LAG3 mutant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3, and the N-terminus thereof is shortened by 0 to 124 amino acids, for example, 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, or 34. , 35 pieces, 36 pieces, 37 pieces, 38 pieces, 39 pieces, 40 pieces, 41 pieces, 42 pieces, 43 pieces, 44 pieces, 45 pieces, 46 pieces, 47 pieces, 48 ​​pieces, 49 pieces, 50 pieces, 51 pieces, 52 pieces, 53 pieces, 54 pieces, 55 pieces, 56 pieces, 57 pieces, 58 pieces, 59 pieces, 60 pieces, 61 pieces, 62 pieces, 63 pieces, 64 pieces, 65 pieces, 66 pieces, 67 pieces, 6 8 pieces, 69 pieces, 70 pieces, 71 pieces, 72 pieces, 73 pieces, 74 pieces, 75 pieces, 76 pieces, 77 pieces, 78 pieces, 79 pieces, 80 pieces, 81 pieces, 82 pieces, 83 pieces, 84 pieces, 85 pieces, 86 pieces, 87 pieces, 88 pieces, 89 pieces, 90 pieces, 91 pieces, 92 pieces, 93 pieces, 94 pieces, 95 pieces, 96 pieces, 97 pieces, 98 pieces, 99 pieces, 100 pieces, 10 The amino acids are truncated by 1, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, or 124 amino acids.Additionally, the C-terminus terminates at the following positions in the extracellular domain of wild-type LAG3: amino acid positions 122-167, e.g., 122-167, 123-167, 124-167, 125-167, 126-167, 127-167, 128-167, 129-167, 130-167, 131-167, 132-167, 133-167, 134-167, 135-167, 136-167, 137-167, 138-167, 139-167, 140-167, 141-167, 142-167, 143-167, 144-167, 145-167, 146-167, 147-167, 148-167, 149-167, 150-167, 151-167, 152-167, 153-167, 154-167, 155-167, 156-167, 157-167, 158-167, 159-167, 160-167, 161-167, 162-167, 163-167, 164-167, 165-167, 166-167, 167-167, 168-167, 169-167, 170-167, 171-167, 172-167, 173-167 Corresponds to one of the positions: 0 to 167, 141 to 167, 142 to 167, 143 to 167, 144 to 167, 145 to 167, 146 to 167, 147 to 167, 148 to 167, 149 to 167, 150 to 167, 151 to 167, 152 to 167, 153 to 167, 154 to 167, 155 to 167, 156 to 167, 157 to 167, 158 to 167, 159 to 167, 160 to 167, 161 to 167, 162 to 167, 163 to 167, 164 to 167, 165 to 167, 166 to 167. For example, the C-terminus may terminate at the following positions: amino acid position 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, or 167 of the wild-type LAG3 extracellular domain polypeptide.

[0213] In some preferred embodiments, the LAG3 mutant polypeptide is based on the extracellular domain of wild-type human LAG3, and is N-terminally shortened by 0, 5, 21, 37, 45, 60, 71, 74, 79, 84, 89, 94, 99, 104, 109, or 114 amino acids, and further terminates at the C-terminus corresponding to the following positions in the extracellular domain of wild-type LAG3: 122, 129, 136, 146, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, or 167.

[0214] In one embodiment, the LAG3 variant polypeptide comprises a mutation at an amino acid site, and the amino acid mutation site is the 97th Arg amino acid. In one embodiment, the LAG3 mutant polypeptide comprises a mutation at an amino acid site where the 97th Arg amino acid is mutated to a Glu amino acid.

[0215] In some preferred embodiments, the truncated LAG3 variant polypeptide has at least one of the following characteristics compared to the wild-type LAG3 extracellular domain polypeptide:

[0216] (1) The N-terminus is shortened by 74 amino acids, and the C-terminus terminates at the following positions: amino acid positions 122, 129, 136, 146, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, or 167 of the extracellular domain of wild-type LAG3. However, preferably, the C-terminus terminates at the following positions: amino acid positions 122, 129, 136, 146, 156, or 161 of the extracellular domain of wild-type LAG3.

[0217] (2) The N-terminus is shortened by 74 amino acids, the 97th Arg amino acid is mutated to a Glu amino acid, and the C-terminus terminates at the following positions: amino acid positions 122, 129, 136, 146, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, or 167 of the extracellular domain polypeptide of wild-type LAG3. However, preferably, the C-terminus terminates at the following positions: amino acid positions 122, 129, 136, 146, 156, or 161 of the extracellular domain polypeptide of wild-type LAG3.

[0218] (3) The N-terminus is shortened by 5 amino acids, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0219] (4) The N-terminus is shortened by five amino acids, the 97th Arg amino acid is mutated to a Glu amino acid, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0220] (5) The N-terminus is shortened by 21 amino acids, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0221] (6) The N-terminus is shortened by 21 amino acids, the 97th Arg amino acid is mutated to a Glu amino acid, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0222] (7) The N-terminus is shortened by 37 amino acids, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0223] (8) The N-terminus is shortened by 37 amino acids, the 97th Arg amino acid is mutated to a Glu amino acid, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0224] (9) The N-terminus is shortened by 45 amino acids, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0225] (10) The N-terminus is shortened by 45 amino acids, the 97th Arg amino acid is mutated to a Glu amino acid, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0226] (11) The N-terminus is shortened by 60 amino acids, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0227] (12) The N-terminus is shortened by 60 amino acids, the 97th Arg amino acid is mutated to a Glu amino acid, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0228] (13) The N-terminus is shortened by 71 amino acids, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0229] (14) The N terminus is shortened by 71 amino acids, the 97th Arg amino acid is mutated to a Glu amino acid, and the C terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0230] (15) The N-terminus is shortened by 79 amino acids, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0231] (16) The N-terminus is shortened by 84 amino acids, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0232] (17) The N-terminus is shortened by 89 amino acids, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0233] (18) The N-terminus is shortened by 94 amino acids, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0234] (19) The N-terminus is shortened by 99 amino acids, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0235] (20) The N-terminus is shortened by 104 amino acids, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0236] (21) The N-terminus is shortened by 109 amino acids, and the C-terminus terminates at the following positions: amino acid position 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0237] (22) The N-terminus is shortened by 114 amino acids, and the C-terminus terminates at the following positions: amino acid positions 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0238] (23) The N-terminus is shortened by 0 amino acids, and the C-terminus terminates at the following positions: amino acid positions 122, 129, 136, 146, 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0239] (24) The N-terminus is shortened by 0 amino acids, the 97th Arg amino acid is mutated to a Glu amino acid, and the C-terminus terminates at the following positions: amino acid positions 122, 129, 136, 146, 156, 161, or 167 of the extracellular domain polypeptide of wild-type LAG3.

[0240] In one embodiment, the LAG3 variant polypeptide is selected from the amino acid sequences set forth in SEQ ID NO:244 to SEQ ID NO:349.

[0241] On the other hand, the present application provides LAG3 mutant polypeptides, which include truncations and / or mutations based on human-derived LAG3.

[0242] In some embodiments, the LAG3 variant polypeptide comprises a truncation based on the extracellular domain of wild-type human-derived LAG3 polypeptide.

[0243] In some embodiments, the LAG3 mutant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3, and is N-terminally shortened by 0 to 124 amino acids and C-terminally terminated between amino acid positions 121 and 167.

[0244] In some embodiments, the LAG3 mutant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3 and includes an N-terminal truncation of 0, 5, 21, 37, 45, 60, 71, 74, 79, 81, 84, 89, 94, 99, 104, 109, or 121 amino acids.

[0245] In some embodiments, the LAG3 mutant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3 and includes an N-terminal truncation of 0, 5, 21, 37, 45, 60, 71, 74, 81, or 99 amino acids.

[0246] In some embodiments, the LAG3 variant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3 and terminates C-terminally at amino acid position 121, 122, 129, 136, 146, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, or 167.

[0247] In some embodiments, the LAG3 variant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3 and terminates C-terminally at amino acid position 121, 122, 129, 136, 146, 156, 161, or 167.

[0248] In some embodiments, the LAG3 variant polypeptide is based on the extracellular domain of wild-type human-derived LAG3 and terminates C-terminally at amino acid position 121, 146, 156, 161, or 167.

[0249] In some embodiments, the LAG3 mutant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3 and comprises an N-terminal truncation of 0, 5, 21, 37, 45, 60, 71, 74, 81, or 99 amino acids and a C-terminal termination at amino acid position 121, 122, 129, 136, 146, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, or 167.

[0250] In some embodiments, the LAG3 mutant polypeptide comprises a mutation at an amino acid site.

[0251] In some embodiments, the LAG3 mutant polypeptide comprises a mutation at an amino acid site, wherein the amino acid mutation site is R110, R113, R119, R129, G130, and / or R141.

[0252] In some embodiments, the LAG3 mutant polypeptide is N-terminally shortened by 81 amino acids, C-terminally terminated at amino acid position 121, and mutated at R110 and / or R113 and / or R119.

[0253] In some embodiments, the LAG3 mutant polypeptide is N-terminally shortened by 99 amino acids, C-terminally terminated at amino acid position 146, and mutated at R129 and / or G130 and / or R141.

[0254] In some embodiments, the LAG3 mutant polypeptide is shortened by 81 amino acids at the N-terminus, terminates at amino acid position 146 at the C-terminus, and has mutations at R110 and / or R113, and / or R119, and / or R129, and / or G130, and / or R141.

[0255] In some embodiments, the LAG3 mutant polypeptide comprises a mutation at an amino acid site, wherein the R110 site is mutated to K110.

[0256] In some embodiments, the LAG3 mutant polypeptide comprises a mutation at an amino acid site, wherein the R113 site is mutated to K113.

[0257] In some embodiments, the LAG3 mutant polypeptide comprises a mutation at an amino acid site, wherein the R119 site is mutated to K119.

[0258] In some embodiments, the LAG3 mutant polypeptide comprises a mutation at an amino acid site, wherein the R129 site is mutated to K129.

[0259] In some embodiments, the LAG3 mutant polypeptide comprises a mutation at an amino acid site, wherein the G130 site is mutated to P130, A130, T130, Y130, or S130.

[0260] In some embodiments, the LAG3 mutant polypeptide comprises a mutation at an amino acid site, wherein the R141 site is mutated to K141.

[0261] In certain embodiments, the amino acid sequence of the LAG3 variant polypeptide is selected from SEQ ID NO:1285 to SEQ ID NO:1318.

[0262] On the other hand, the present application provides a fusion polypeptide comprising a first domain and a second domain, wherein the first domain comprises a LAG3 mutant polypeptide described in the present application, and the second domain comprises an antibody or antigen-binding fragment thereof or an immunoglobulin domain.

[0263] In some embodiments, the second domain comprises an antibody or an antigen-binding fragment thereof.

[0264] In some embodiments, the second domain is capable of binding to one or more of the targets set forth in the following group: PD-L1, PD-L2, PD-1, OX40, 4-1BB, ICOS, TIGIT, CTLA4, LAG3, CD3, VEGF, VEGFR, CD47, HGF, Trop2, EpCAM, CCR8, CCR4, CCR5, GPRC5D, BCMA, CD19, CD20, HER-2 neu, DLL1, HER-3, HER-4, EGFR, PSMA, CEA, MUC-1 (mucin), MUC2, MUC3, MUC4, MUC5AC, MUC5B, MUC7, CD123, CD33, CD30, CD38, NKG2A, Nkp36, and Tim3.

[0265] In some embodiments, the second domain is capable of binding to PD-L1, PD-1 and / or PD-L2.

[0266] In some embodiments, the second domain comprises HCDR1, HCDR2 and / or HCDR3 of an antibody heavy chain, and the antibody heavy chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0267] In some embodiments, the second domain comprises a heavy chain variable region VH of an antibody heavy chain, and the antibody heavy chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0268] In some embodiments, the second domain comprises an antibody heavy chain, and the antibody heavy chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15, and SEQ ID NO:17.

[0269] In some embodiments, the second domain comprises LCDR1, LCDR2 and / or LCDR3 of an antibody light chain, and the antibody light chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0270] In some embodiments, the second domain comprises a light chain variable region VL of an antibody light chain, and the antibody light chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16, and SEQ ID NO:18.

[0271] In some embodiments, the second domain comprises an antibody light chain, and the antibody light chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16, and SEQ ID NO:18.

[0272] In one embodiment of the fusion polypeptide, the second domain comprises an antibody or antigen-binding fragment thereof selected from the group consisting of Sugemalimab, Durvalumab, Atezolizumab, Avelumab, Envafolimab, Permbrolizumab, and Nivolumab.

[0273] In one embodiment of the fusion polypeptide, the second domain comprises sugemalimab or an antigen-binding fragment thereof. In some embodiments, the fusion polypeptide, the antigen-binding fragment is selected from one or more of the following group: Fab, Fab', Fv fragment, F(ab')2, F(ab)2, scFv, di-scFv, VHH, and dAb.

[0274] In one embodiment of the fusion polypeptide, the second domain comprises an immunoglobulin domain.

[0275] In one embodiment, the fusion polypeptide comprises an immunoglobulin IgG antibody, wherein the immunoglobulin IgG is selected from the group consisting of one or more of human IgG1, human IgG2, human IgG3, human IgG4, mouse IgG1, mouse IgG2a, mouse IgG2b, and mouse IgG3.

[0276] In one embodiment of the fusion polypeptide, the second domain comprises an immunoglobulin Fc domain.

[0277] In one embodiment, the fusion polypeptide comprises an immunoglobulin IgG Fc domain, and the immunoglobulin Fc domain is selected from the group consisting of a human IgG1 Fc domain, a human IgG2 Fc domain, a human IgG3 Fc domain, a human IgG4 Fc domain, a mouse IgG1 Fc domain, a mouse IgG2a Fc domain, a mouse IgG2b Fc domain, and a mouse IgG3 Fc domain.

[0278] In one embodiment, the fusion polypeptide comprises an immunoglobulin Fc domain comprising the Fc domain of human immunoglobulin IgG4.

[0279] In one embodiment of the fusion polypeptide, the second domain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:4 to SEQ ID NO:18, SEQ ID NO:29 to SEQ ID NO:30, SEQ ID NO:634 to SEQ ID NO:639.

[0280] In one embodiment, the fusion polypeptide has the first domain linked directly or indirectly to the second domain.

[0281] In one embodiment, the fusion polypeptide comprises the first domain linked directly or indirectly to the N-terminus of the second domain.

[0282] In one embodiment of the fusion polypeptide, the first domain is linked directly or indirectly to the C-terminus of the second domain.

[0283] In one embodiment of the fusion polypeptide, the indirect connection comprises a connection via a linker.

[0284] In one embodiment of the fusion polypeptide, the linker is selected from the amino acid sequences set forth in the following group: SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34 and SEQ ID NO:35.

[0285] In one embodiment of the fusion polypeptide, the fusion polypeptide is selected from the amino acid sequences set forth in the following group: SEQ ID NO:640 to SEQ ID NO:745 and SEQ ID NO:747 to SEQ ID NO:750.

[0286] In some embodiments, the fusion polypeptide comprises an antibody heavy chain, and preferably the antibody heavy chain polypeptide comprises the amino acid sequence set forth in SEQ ID NO:4; the amino acid sequence comprising the antibody heavy chain is selected from those set forth in SEQ ID NO:1353 to SEQ ID NO:1386.

[0287] In some embodiments, the fusion polypeptide comprises a light chain, wherein the light chain comprises an antibody light chain polypeptide, preferably the antibody light chain polypeptide comprises the amino acid sequence set forth in SEQ ID NO:5.

[0288] In some embodiments, the heavy chain of the fusion polypeptide and the light chain of the fusion polypeptide combine to form the fusion polypeptide.

[0289] In some embodiments, the first domain is directly or indirectly linked to the second domain.

[0290] In some embodiments, the first domain is directly or indirectly linked to the N-terminus of the second domain.

[0291] In some embodiments, the first domain is directly or indirectly linked to the C-terminus of the second domain.

[0292] In some embodiments, the second domain can comprise an antibody heavy chain, and the LAG3 mutant polypeptide is directly or indirectly bound to the antibody heavy chain of the second domain.

[0293] In some embodiments, the second domain can comprise an antibody heavy chain, and the LAG3 mutant polypeptide is linked directly or indirectly to the C-terminus of the antibody heavy chain of the second domain.

[0294] In some embodiments, the second domain can comprise an antibody heavy chain, and the N-terminus of the LAG3 mutant polypeptide is linked directly or indirectly to the C-terminus of the antibody heavy chain of the second domain.

[0295] In some embodiments, the second domain can comprise an antibody heavy chain, and the LAG3 mutant polypeptide is linked directly or indirectly to the N-terminus of the antibody heavy chain of the second domain.

[0296] In some embodiments, the second domain can comprise an antibody heavy chain, and the C-terminus of the LAG3 mutant polypeptide is directly or indirectly linked to the N-terminus of the antibody heavy chain of the second domain.

[0297] In some embodiments, the second domain can comprise an antibody light chain, and the LAG3 mutant polypeptide is directly or indirectly bound to the antibody light chain of the second domain.

[0298] In some embodiments, the second domain can comprise an antibody light chain, and the LAG3 mutant polypeptide is linked directly or indirectly to the C-terminus of the antibody light chain of the second domain.

[0299] In some embodiments, the second domain can comprise an antibody light chain, and the N-terminus of the LAG3 mutant polypeptide is linked directly or indirectly to the C-terminus of the antibody light chain of the second domain.

[0300] In some embodiments, the second domain can comprise an antibody light chain, and the LAG3 mutant polypeptide is linked directly or indirectly to the N-terminus of the antibody light chain of the second domain.

[0301] In some embodiments, the second domain can comprise an antibody light chain, and the C-terminus of the LAG3 mutant polypeptide is directly or indirectly linked to the N-terminus of the antibody light chain of the second domain.

[0302] In some embodiments, the indirect attachment is via a linker.

[0303] In some embodiments, the second domain comprises an immunoglobulin Fc domain.

[0304] In some embodiments, the second domain comprises the Fc domain of an immunoglobulin IgG antibody.

[0305] In some embodiments, the second domain comprises an Fc domain of an immunoglobulin IgG antibody, and the Fc domain of the immunoglobulin IgG antibody is selected from the group consisting of a human IgG1 Fc domain, a human IgG2 Fc domain, a human IgG3 Fc domain, a human IgG4 Fc domain, a mouse IgG1 Fc domain, a mouse IgG2a Fc domain, a mouse IgG2b Fc domain, or a mouse IgG3 Fc domain.

[0306] In some embodiments, the second domain comprises the Fc domain of human-derived immunoglobulin IgG4.

[0307] In some embodiments, the LAG3 mutant polypeptide is covalently linked to the second domain with or without a linker peptide.

[0308] In some embodiments, the second domain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:1313 to SEQ ID NO:1352.

[0309] In some embodiments, the fusion polypeptide further comprises a third domain, which is the same or different from the second domain, an antibody, an antigen-binding fragment, or a functional protein or active fragment thereof.

[0310] On the other hand, the present application provides a fusion polypeptide, the fusion polypeptide comprising a first domain, a second domain and a third domain, wherein the first domain comprises a LAG3 mutant polypeptide described in the present application, the second domain comprises an antibody or an antigen-binding fragment thereof, or an immunoglobulin domain, and the third domain is an antibody, antigen-binding fragment, or functional protein or active fragment thereof that is the same as or different from the second domain.

[0311] In some embodiments, the second domain comprises an antibody or an antigen-binding fragment thereof.

[0312] In some embodiments, the second domain is capable of binding to one or more of the targets set forth in the following group: PD-L1, PD-L2, PD-1, OX40, 4-1BB, ICOS, TIGIT, CTLA4, LAG3, CD3, VEGF, VEGFR, CD47, HGF, Trop2, EpCAM, CCR8, CCR4, CCR5, GPRC5D, BCMA, CD19, CD20, HER-2 neu, DLL1, HER-3, HER-4, EGFR, PSMA, CEA, MUC-1 (mucin), MUC2, MUC3, MUC4, MUC5AC, MUC5B, MUC7, CD123, CD33, CD30, CD38, NKG2A, Nkp36, and Tim3.

[0313] In some embodiments, the second domain is capable of binding to PD-L1, PD-1, and / or PD-L2. In some embodiments, the second domain comprises HCDR1, HCDR2, and / or HCDR3 of an antibody heavy chain, and the antibody heavy chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15, and SEQ ID NO:17.

[0314] In some embodiments, the second domain comprises a heavy chain variable region VH of an antibody heavy chain, and the antibody heavy chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0315] In some embodiments, the second domain comprises an antibody heavy chain, and the antibody heavy chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15, and SEQ ID NO:17.

[0316] In some embodiments, the second domain comprises LCDR1, LCDR2 and / or LCDR3 of an antibody light chain, and the antibody light chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0317] In some embodiments, the second domain comprises a light chain variable region VL of an antibody light chain, and the antibody light chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16, and SEQ ID NO:18.

[0318] In some embodiments, the second domain comprises an antibody light chain, and the antibody light chain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16, and SEQ ID NO:18.

[0319] In one embodiment of the fusion polypeptide, the second domain comprises an antibody or antigen-binding fragment thereof selected from the group consisting of sugemalimab, durvalumab, atezolizumab, avelumab, embafolimab, permbrolizumab, and nivolumab.

[0320] In one embodiment of the fusion polypeptide, the second domain comprises sugemalimab or an antigen-binding fragment thereof.

[0321] In some embodiments of the fusion polypeptide, the antibody is selected from the group consisting of immunoglobulin antibodies, recombinant antibodies, chimeric antibodies, heavy chain antibodies, single domain antibodies, and bispecific antibodies.

[0322] In one embodiment of the fusion polypeptide, the antigen-binding fragment is selected from one or more of the following group: Fab, Fab', Fv fragment, F(ab')2, F(ab)2, scFv, di-scFv, VHH, and dAb.

[0323] In one embodiment of the fusion polypeptide, the second domain comprises an immunoglobulin domain.

[0324] In one embodiment, the fusion polypeptide comprises an immunoglobulin IgG antibody, wherein the immunoglobulin IgG is selected from the group consisting of one or more of human IgG1, human IgG2, human IgG3, human IgG4, mouse IgG1, mouse IgG2a, mouse IgG2b, and mouse IgG3.

[0325] In one embodiment of the fusion polypeptide, the second domain comprises an immunoglobulin Fc domain.

[0326] In one embodiment, the fusion polypeptide comprises an immunoglobulin IgG Fc domain, and the immunoglobulin Fc domain is selected from the group consisting of a human IgG1 Fc domain, a human IgG2 Fc domain, a human IgG3 Fc domain, a human IgG4 Fc domain, a mouse IgG1 Fc domain, a mouse IgG2a Fc domain, a mouse IgG2b Fc domain, and a mouse IgG3 Fc domain.

[0327] In one embodiment, the fusion polypeptide comprises an immunoglobulin Fc domain comprising the Fc domain of human immunoglobulin IgG4.

[0328] In one embodiment of the fusion polypeptide, the second domain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:4 to SEQ ID NO:18, SEQ ID NO:29 to SEQ ID NO:30, SEQ ID NO:634 to SEQ ID NO:639.

[0329] In one embodiment of the fusion polypeptide, the third domain is a functional protein or an active fragment thereof, wherein the functional protein or an active fragment thereof is capable of activating an innate immune response.

[0330] In one embodiment of the fusion polypeptide, the third domain is capable of binding to CD28 and / or CTLA4.

[0331] In one embodiment of the fusion polypeptide, the third domain comprises CD80, CD86, or an active fragment thereof.

[0332] In one embodiment, the fusion polypeptide has a third domain selected from the group consisting of CD86 or a functionally active fragment thereof of human origin, CD86 or a functionally active fragment thereof of mouse origin, CD80 or a functionally active fragment thereof of human origin, or CD80 or a functionally active fragment thereof of mouse origin.

[0333] In one embodiment of the fusion polypeptide, the third domain comprises the IgV domain of CD80, the IgV domain of CD86, or an active fragment thereof.

[0334] In one embodiment of the fusion polypeptide, the third domain comprises the extracellular domain of CD80, the extracellular domain of CD86, or an active fragment thereof.

[0335] In one embodiment of the fusion polypeptide, the third domain comprises a CD86 mutant polypeptide.

[0336] In one embodiment of the fusion polypeptide, the third domain comprises a CD86 variant polypeptide, wherein the variant comprises an amino acid substitution mutation of humanized CD86.

[0337] In one embodiment of the fusion polypeptide, the third domain comprises a CD86 mutant polypeptide, and the CD86 mutant polypeptide comprises an IgV domain amino acid substitution mutant of CD86 and / or an extracellular domain amino acid substitution mutant of humanized CD86.

[0338] In one embodiment, the fusion polypeptide comprises a CD86 mutant polypeptide, wherein the CD86 mutant polypeptide comprises an amino acid substitution mutant of the IgV domain of CD86, and the mutation sites of the amino acid substitution mutant of the IgV domain of CD86 include one or more of the amino acid site mutations shown in the following group: A13I, A13L, A13F, A13M, Q25A, Q25I, Q25V, Q25F, Q25M, F33A, F33L, F33V, F33M, M60R, I89A, I89L, I89V, I89F, I89M, H90I, H90V, H90M, H90F.

[0339] In one embodiment, the fusion polypeptide comprises a CD86 variant polypeptide, wherein the CD86 variant polypeptide comprises an extracellular domain amino acid substitution variant of CD86, and the extracellular domain amino acid substitution variant of CD86 comprises one or more amino acid site mutations shown in the following group: A13I, A13L, A13F, A13M, Q25A, Q25I, Q25V, Q25F, Q25M, F33A, F33L, F33V, F33M, M60R, I89A, I89L, I89V, I89F, I89M, H90I, H90V, H90M, H90F.

[0340] In one embodiment of the fusion polypeptide, the third domain is selected from the amino acid sequences set forth in the following group: SEQ ID NO:2 to SEQ ID NO:3, SEQ ID NO:178 to SEQ ID NO:243.

[0341] In some embodiments, the amino acid substitution mutants in the IgV or extracellular domain of CD86 may be selected from the combinations of amino acid mutation sites shown in the following group: Q25I / F33L / H90I, Q25V / F33L / H90I, Q25I / F33V / H90I, Q25V / F33V / H90I, Q25I / F33L / H90V , Q25V / F33L / H90V, Q25I / F33V / H90V, Q25V / F33V / H90V, A13L / Q25I / F33L / H90I, A13I / Q25I / F3 3L / H90I, A13L / Q25V / F33L / H90I, A13I / Q25V / F33L / H90I, Q25I / F33L / I89L / H90I, Q25I / F33L / M 60R / H90I, Q25V / F33L / I89L / H90I, Q25V / F33L / M60R / H90I, Q25F / F33L / H90I, Q25I / F33L / H90F , Q25I / F33A / H90I, Q25I / H90I, Q25I, H90I, Q25V / H90V, Q25V / H90I, Q25F / H90I, Q25F / H90V, A13 F / Q25I / F33L / H90I, A13M / Q25I / F33L / H90I, Q25A / F33L / H90I, Q25M / F33L / H90I, Q25I / F33M / H 90I, Q25I / F33L / I89V / H90I, Q25I / F33L / I89F / H90I, Q25I / F33L / I89M / H90I sum Q25I / F33L / H90M.

[0342] In some embodiments, the amino acid substitution mutant of the IgV or extracellular domain of CD86 comprises the following amino acid mutation sites: Q25I / F33L / H90I.

[0343] In one embodiment, the fusion polypeptide comprises a direct or indirect link between the first domain and the second domain.

[0344] In one embodiment, the fusion polypeptide has the first domain linked directly or indirectly to the third domain.

[0345] In one embodiment, the fusion polypeptide includes a direct or indirect link between the second domain and the third domain.

[0346] In one embodiment, the fusion polypeptide has the first domain linked directly or indirectly to the second domain, and the first domain linked directly or indirectly to the third domain.

[0347] In one embodiment, the fusion polypeptide has the first domain linked directly or indirectly to the second domain, which is further linked directly or indirectly to the third domain.

[0348] In one embodiment, the fusion polypeptide has the first domain linked directly or indirectly to the third domain, and the second domain linked directly or indirectly to the third domain.

[0349] In one embodiment of the fusion polypeptide, the indirect connection comprises a connection via a linker.

[0350] In one embodiment of the fusion polypeptide, the linker comprises an amino acid sequence selected from the group consisting of SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34 and SEQ ID NO:35.

[0351] In one embodiment, the fusion polypeptide comprises an amino acid sequence selected from among SEQ ID NO:751 to SEQ ID NO:1024, 1249, 1251, 1253, and 1255.

[0352] In some embodiments, the fusion polypeptide comprises the heavy chain of a multifunctional LAG3 mutant fusion Sugemalimab-CD86 IgV Q25I / F33L / H90I antibody polypeptide complex, wherein the amino acid sequence of the heavy chain is selected from among SEQ ID NO: 1387 to SEQ ID NO: 1420.

[0353] In some embodiments, the fusion polypeptide comprises the heavy chain of a multifunctional LAG3 mutant fusion Sugemalimab-CD86 IgV Q25I / F33L / H90I antibody polypeptide complex, wherein the amino acid sequence of the heavy chain is selected from among SEQ ID NO: 1421 and SEQ ID NO: 1426.

[0354] In some embodiments, the nucleic acid encoding the fusion polypeptide comprises a nucleotide sequence set forth in any of SEQ ID NOs: 1422-1425, 1427-1433.

[0355] On the other hand, the present application provides an immunoconjugate, which comprises a fusion polypeptide described in the present application, a CD86 mutant polypeptide described in the present application, and / or a LAG3 mutant described in the present application.

[0356] On the other hand, the present application provides nucleic acid molecules, which encode the fusion polypeptides described in the present application, the CD86 mutant polypeptides described in the present application, and / or the LAG3 mutants described in the present application.

[0357] On the other hand, the present application provides a vector, which comprises a nucleic acid molecule described in the present application.

[0358] On the other hand, the present application provides cells that contain and / or express a fusion polypeptide described in the present application, a CD86 mutant polypeptide described in the present application, a LAG3 mutant described in the present application, an immunoconjugate described in the present application, a nucleic acid molecule described in the present application, and / or a vector described in the present application.

[0359] On the other hand, the present application provides a composition, which comprises a fusion polypeptide described in the present application, an immunoconjugate described in the present application, a nucleic acid molecule described in the present application, a vector described in the present application, and / or a cell described in the present application, and optionally further comprises a pharmaceutically acceptable carrier.

[0360] On the other hand, the present application provides methods for producing the fusion polypeptides described in the present application, the CD86 mutant polypeptides described in the present application, and the LAG3 mutants described in the present application, which methods comprise culturing the cells described in the present application under conditions that allow the fusion polypeptides to be expressed.

[0361] On the other hand, the present application provides a method for inhibiting the interaction of PD-L1 protein and PD-1, the method comprising administering an effective amount of a fusion polypeptide described in the present application, a CD86 mutant polypeptide described in the present application, a LAG3 mutant described in the present application, an immunoconjugate described in the present application, a nucleic acid molecule described in the present application, a vector described in the present application, a cell described in the present application and / or a composition described in the present application.

[0362] On the other hand, the present application provides a method for stimulating antigen-presenting cells and / or promoting T cell activation, the method comprising administering an effective amount of a fusion polypeptide described in the present application, a CD86 mutant polypeptide described in the present application, a LAG3 mutant described in the present application, an immunoconjugate described in the present application, a nucleic acid molecule described in the present application, a vector described in the present application, a cell described in the present application and / or a composition described in the present application.

[0363] In the methods described in the present application, the stimulation of the antigen-presenting cells includes those selected from the group consisting of increasing the expression of costimulatory molecules in the antigen-presenting cells, maturing the morphological changes of the antigen-presenting cells, increasing the secretion of chemokines in the antigen-presenting cells, and enhancing the phagocytic ability of the antigen-presenting cells.

[0364] On the other hand, the present application provides a method for inhibiting the growth and / or proliferation of a tumor or tumor cells, the method comprising administering an effective amount of a fusion polypeptide described in the present application, a CD86 mutant polypeptide described in the present application, a LAG3 mutant described in the present application, an immunoconjugate described in the present application, a nucleic acid molecule described in the present application, a vector described in the present application, a cell described in the present application and / or a composition described in the present application.

[0365] On the other hand, the present application provides the use of a fusion polypeptide described in the present application, a CD86 mutant polypeptide described in the present application, a LAG3 mutant described in the present application, an immunoconjugate described in the present application, a nucleic acid molecule described in the present application, a vector described in the present application, a cell described in the present application and / or a composition described in the present application in the manufacture of a medicament for use in the prevention, amelioration and / or treatment of a tumor.

[0366] According to the uses described in this application, tumors include solid tumors and / or hematological tumors.

[0367] According to the use described in the present application, the tumor is selected from the group consisting of colon tumor, breast tumor, lung tumor, gastric tumor, melanoma, head and neck tumor, lymphoma, nasopharyngeal tumor, cervical tumor, esophageal tumor, kidney tumor, cutaneous squamous cell carcinoma, endometrial tumor, liver tumor, bladder tumor, urothelial tumor and skin tumor.

[0368] Meanwhile, the present application provides a fusion polypeptide described herein, an immunoconjugate described herein, a nucleic acid molecule described herein, a vector described herein, a cell described herein, and / or a composition described herein, which are used for the prevention, amelioration, and / or treatment of tumors.

[0369] However, the present application also provides a fusion polypeptide described herein, an immunoconjugate described herein, a nucleic acid molecule described herein, a vector described herein, a cell described herein, and / or a composition described herein, for use in the prevention, amelioration, and / or treatment of tumors, including solid tumors and hematological tumors.

[0370] On the other hand, the present application provides a fusion polypeptide described herein, an immunoconjugate described herein, a nucleic acid molecule described herein, a vector described herein, a cell described herein, and / or a composition described herein, for use in the prevention, amelioration, and / or treatment of a tumor, wherein the tumor is selected from the following group: colon tumor, breast tumor, lung tumor, gastric tumor, melanoma, head and neck tumor, lymphoma, nasopharyngeal tumor, cervical tumor, esophageal tumor, kidney tumor, cutaneous squamous cell carcinoma, endometrial tumor, liver tumor, bladder tumor, urothelial tumor, and skin tumor.

[0371] Meanwhile, the present application provides a method for preventing, ameliorating, and / or treating tumors. The method may include administering to a subject in need thereof a fusion polypeptide described herein, an immunoconjugate described herein, a nucleic acid molecule described herein, a vector described herein, a cell described herein, and / or a composition described herein.

[0372] According to the methods described in this application, the tumors include solid tumors and hematological tumors.

[0373] According to the methods described in the present application, the tumor is selected from the group consisting of colon tumor, breast tumor, lung tumor, gastric tumor, melanoma, head and neck tumor, lymphoma, nasopharyngeal tumor, cervical tumor, esophageal tumor, kidney tumor, cutaneous squamous cell carcinoma, endometrial tumor, liver tumor, bladder tumor, urothelial tumor and skin tumor.

[0374] Those skilled in the art will be able to readily understand other aspects and advantages of the present application from the following detailed description. In the following detailed description, only exemplary embodiments of the present application are shown and described. As those skilled in the art will understand, the present application allows modifications to be made to the disclosed specific embodiments without departing from the spirit and scope of the present invention. Accordingly, the drawings and description of the present application are merely illustrative and not restrictive. [Brief explanation of the drawings]

[0375] Specific features of the invention of this application are set forth in the appended claims. A better understanding of the features and advantages of the invention can be obtained by reference to the examples set forth in the following detailed description and the accompanying drawings, a brief description of which follows: [Figure 1] Figure 1 shows an exemplary structural diagram of the fusion polypeptide described in the present application. The present application provides the following exemplary structures: a first domain comprising one or more subunits derived from the antigen recognition site (i) of an anti-PD-L1 and / or anti-PD-1 antibody, an immunoglobulin Fc region (ii), a second domain (iii), and / or a third domain (iv). (Although a structure having one subunit in the second domain (iii) and / or the third domain (iv) is shown here, tandem structures of two or more of the same subunit are also contemplated. This can also be extended to tandem structures with mixed combinations of any subunits in (iv).) The first domain is linked to the second domain via the immunoglobulin Fc region by a linker or a covalent bond, and similarly, the subunits of the third domain and the second domain can also be linked by a linker or a covalent bond. [Figure 2] FIG. 1 shows the results of detecting the expression status of the fusion polypeptide complex of the present application using SDS-PAGE. [Figure 3] Figure 1 shows the results of detecting the binding status of the fusion polypeptide complex of the present application to human PD-L1 protein using ELISA. [Figure 4] FIG. 1 shows the results of detecting the binding status between the fusion polypeptide complex of the present application and human CTLA4 protein using ELISA. [Figure 5] FIG. 1 shows the results of detecting the binding status between the fusion polypeptide complex of the present application and human CD28 protein using ELISA. [Figure 6] 1 shows the results of the fusion polypeptide complex of the present application regulating the activation of Jurkat T cells. [Figure 7] 1 shows results demonstrating that the fusion polypeptide complex of the present application upregulates the expression of costimulatory molecules in antigen-presenting cells. [Figure 8] 1 shows the results of the fusion polypeptide complex of the present application regulating the morphological differentiation of antigen-presenting cells. [Figure 9] 1 shows the results of the fusion polypeptide complex of the present application regulating the secretion of CCL4 in antigen-presenting cells. [Figure 10] FIG. 1 shows the results of detecting the expression status of CD86 mutant fusion polypeptide complexes using SDS-PAGE. [Figure 11] FIG. 1 shows the results of detecting the expression status of CD86 mutant polypeptide fusion polypeptide complexes using AKTA. [Figure 12] 1 shows the results of the fusion polypeptide complex of the present application regulating the secretion of CCL4 in antigen-presenting cells. [Figure 13] 1 shows the tumor growth suppressive effect of the fusion polypeptide complex of the present application. [Figure 14] FIG. 1 shows the results of detecting the expression status of the LAG3 mutant polypeptide complex of the present application using SDS-PAGE. [Figure 15] 1 shows the results of the fusion polypeptide complex of the present application regulating the secretion of CCL4 in antigen-presenting cells. [Figure 16]1 shows the results of the fusion polypeptide complex of the present application regulating the secretion of CCL4 in primary PBMC cells. [Figure 17] FIG. 1 shows the results of the fusion polypeptide complex of the present application regulating the morphological differentiation of primary DCs cells. [Figure 18] FIG. 1 shows the results of the fusion polypeptide complex of the present application upregulating the expression of costimulatory molecules in primary DCs cells. [Figure 19] FIG. 1 shows the results of the fusion polypeptide complex of the present application regulating the secretion of TNFα in primary DCs cells. [Figure 20] 1 shows results demonstrating how the fusion polypeptide complexes of the present application regulate T cell activation. [Figure 21] FIG. 1 shows the results of the LAG3 mutant polypeptide complex of the present application regulating the morphological differentiation of primary DCs cells. [Figure 22] FIG. 1 shows the results of the LAG3 mutant polypeptide complex of the present application upregulating the expression of costimulatory molecules in primary DCs cells. [Figure 23] FIG. 1 shows the results of the LAG3 mutant polypeptide complex of the present application regulating the secretion of TNFα in primary DCs cells. [Figure 24] 1 shows deglycosylated, reduced LC-MS mass spectrometry of the LAG3 mutant polypeptide complex of the present application. DETAILED DESCRIPTION OF THE INVENTION

[0376] The following describes the embodiments of the present invention using specific examples, and those skilled in the art can easily understand other advantages and effects of the present invention from the contents disclosed herein. Definition of Terms

[0377] In the present application, the term "modulating a T cell immune response" generally refers to the modulation of lymphatic T cell activity through the action of the bi- and / or multi-functional fusion polypeptides disclosed herein. This also includes the expansion of T cell responses, such as the expression of NFAT transcription factors, the secretion of cytokines IL-2, IFN-γ, TNFα, and Granzyme B, and actions against target cells (e.g., tumor cells) including cell proliferation and cytotoxicity in T cells treated with other related T cell modulating agents. The T cells include T cell lineages and / or primary T cells, including, but not limited to, CD4 T cells and CD4 T cell subpopulations (Th1, Th2, Th9, Th17, TGF-β, and / or Treg cells). They also include CD8 T cells, including, but not limited to, tumor tissue-infiltrating CD8 T cells, effector CD8 T cells, and immune memory CD8 T cells.

[0378] In the present application, the term "stimulation of antigen-presenting cells" generally refers to the modulation of antigen-presenting cell activation by the bi- and / or multi-functional fusion polypeptides disclosed herein. For example, this can be extended to include the effects of other antigen-presenting cell-modulating agents on the treated antigen-presenting cells, including, but not limited to, the expression of costimulatory molecules (ICOSL, CD40L, CD137L, OX40L, CD80, CD83, and / or CD86), activation and differentiation / maturation of antigen-presenting cells, antigen phagocytosis and presentation, production of chemokines such as CCL4, CCL22, and CCL17, expression of cytokines such as IL-1beta, TNFα, IFNγ, IL-6, and IL-12, induction and recruitment of T cells for migration to secondary lymphoid organs, and enhancement of T cell immune responses. The antigen-presenting cells include, but are not limited to, B lymphocytes and primary B lymphocytes, monocytes and primary monocytes, macrophages and primary macrophages, dendritic cells (DC) and primary DC cells, etc. The phagocytosis of the antigen includes, but is not limited to, the phagocytosis of bacteria, viruses, proteins, polysaccharides, etc.

[0379] As used herein, the term "relieving PD-L1 / PD-1-mediated T cell inhibition" generally means that the bi- and / or multi-functional fusion polypeptides disclosed herein bind to PD-L1 protein and inhibit the binding of PD-L1 to PD-1, thereby improving PD-L1 / PD-1-mediated T cell inhibition, including, but not limited to, increasing NFAT transcription factor expression in T cells, secretion of cytokines such as IL-2, IFN-γ, TNFα, and Granzyme B, cell proliferation, and cytotoxic effects on target cells (e.g., tumor cells).

[0380] In this application, the term "anti-tumor activity" generally refers to a biological activity that reduces or prevents the growth or viability of tumor cells in vivo and / or in vitro. In one embodiment, the anti-tumor activity refers to the anti-tumor effect of the bi- and / or multi-functional fusion polypeptides described herein.

[0381] In this application, the term "CD86" generally refers to a molecule of activated cells. For example, CD86 in this application includes its full-length form, variants, and / or functionally active fragments. In this application, CD86 can refer to a polypeptide or fragment thereof that has at least about 85% amino acid identity with the protein encoded by the gene with NCBI accession number Gene ID: 942 and has binding activity to CD28 (the protein encoded by the gene with NCBI accession number Gene ID: 940) and / or CTLA4 (the protein encoded by the gene with NCBI accession number Gene ID: 1493). An exemplary human CD86 amino acid sequence (SEQ ID NO: 1) is provided below. The terms "CD86 extracellular region," "CD86 extracellular domain," and "CD86 ECD region" refer to the amino acid sequence of a domain in the extracellular region of the CD86 protein, and refer to a polypeptide or fragment thereof that has at least about 85% amino acid identity and has binding activity to CD28 and / or CTLA4. An exemplary amino acid sequence of the CD86 extracellular region (SEQ ID NO: 2) is provided below. The terms "CD86 extracellular structural region IgV domain" and "CD86 IgV domain" refer to the amino acid sequence of an IgV-like structural domain in the structural region of the extracellular region of the CD86 protein, and refer to a polypeptide or fragment thereof having at least about 85% amino acid identity and having binding activity to CD28 and / or CTLA4. An exemplary amino acid sequence of a CD86 extracellular structural region IgV domain (SEQ ID NO: 3) is provided below. The number of linked CD86 extracellular structural regions or CD86 extracellular structural region IgV domains disclosed in the present invention is not limited to the number shown as an example, and any linked combination comprising this functional structural region is considered to be within the scope of the present invention.

[0382] As used herein, the term "anti-PD-L1 antibody" refers to an antibody that selectively binds to and inhibits PD-L1 peptide activity. For example, Chinese Patent No. CN102245640(B), U.S. Patent Publication No. 7,943,743, U.S. Patent Publication No. 8,779,108, and U.S. Patent Publication No. 7,943,743 are all cited herein. Anti-PD-L1 antibodies include, but are not limited to, Durvalumab (MEDI4736) developed by AstraZeneca, Sugemalimab (CS1001) developed by Basigo Pharmaceutical, JS003 developed by Junshi Biosciences, Envafolimab (KN035) developed by Corning Incorporated, Atezolizumab (MPDL3280A) developed by Roche, and Avelumab (MSB0010718C) jointly developed by Merck KGaA of Germany and Pfizer. Provided below are the heavy chain multipeptide amino acid sequence (SEQ ID NO: 4) and light chain multipeptide amino acid sequence (SEQ ID NO: 5) of exemplary anti-PD-L1 antibodies Sugemalimab (CS1001), the heavy chain multipeptide amino acid sequence (SEQ ID NO: 6) and light chain multipeptide amino acid sequence (SEQ ID NO: 7) of JS003, the heavy chain multipeptide amino acid sequence (SEQ ID NO: 8) and light chain multipeptide amino acid sequence (SEQ ID NO: 9) of Durvalumab (MEDI4736), the heavy chain multipeptide amino acid sequence (SEQ ID NO: 10) and light chain multipeptide amino acid sequence (SEQ ID NO: 11) of atezolizumab (MPDL3280A), the heavy chain multipeptide amino acid sequence (SEQ ID NO: 12) and light chain multipeptide amino acid sequence (SEQ ID NO: 13) of Avelumab (MSB0010718C), and the amino acid sequence of Envafolimab (KN035) (SEQ ID NO: 14).

[0383] As used herein, the term "anti-PD-1 antibody" refers to an antibody that selectively binds to and inhibits PD-1 multi-peptide activity. For example, U.S. Patent Application Publication Nos. 8,354,509 and 7,488,802 are incorporated herein by reference. Anti-PD-1 antibodies include Permbrolizumab (MK-3475) developed by Merck and Nivolumab (BMS-936558) developed by Biogen. The heavy chain multi-peptide amino acid sequence (SEQ ID NO: 15) and light chain multi-peptide amino acid sequence (SEQ ID NO: 16) of exemplary anti-PD-1 antibodies Permbrolizumab (MK-3475) and the heavy chain multi-peptide amino acid sequence (SEQ ID NO: 17) and light chain multi-peptide amino acid sequence (SEQ ID NO: 18) of Nivolumab (BMS-936558) are provided below.

[0384] In the present application, the term "LAG3" refers to a type of protein or polypeptide. For example, LAG3 in the present application includes its full-length, variants, and / or functionally active fragments. For example, LAG3 can refer to a polypeptide or fragment thereof that has at least about 85% amino acid identity with the protein encoded by LAG3 (Lymphocyte activation gene 3) under NCBI accession number Gene ID:3902, and that exhibits binding activity to MHCII (or HLA-DR) and / or FGL1 (protein encoded by NCBI accession number Gene ID:2267), a complex protein composed of an α subunit (protein encoded by NCBI accession number Gene ID:3122) and a β subunit (protein encoded by NCBI accession number Gene ID:3123 or NCBI accession number Gene ID:3125), and / or activates antigen-presenting cells. The following is an exemplary amino acid sequence of human LAG3 (SEQ ID NO:19): The term "LAG3 extracellular domain" refers to the amino acid sequence of the extracellular domain of the full-length LAG3 protein (a polypeptide consisting of amino acids 23-450 in the LAG3 protein, NCBI numbering system; the extracellular domain begins at amino acid 23, so if 74 amino acids are truncated, the truncated form can begin at amino acid 97), which has at least about 85% amino acid identity and includes a polypeptide or fragment thereof that exhibits MHCII and / or FGL1 binding activity and activates antigen-presenting cells. The following provides an exemplary human-derived LAG3 extracellular domain amino acid sequence (SEQ ID NO: 20) and a tandem human-derived LAG3 extracellular domain amino acid sequence covalently linked by a linker (SEQ ID NO: 21); the number of tandem domains disclosed in the present invention is not limited to the exemplary number, and any tandem combination containing this functional domain is considered within the scope of the present invention.

[0385] As used herein, the term "LAG3-IgD1" refers to the amino acid sequence of the Ig-like V-type domain in the extracellular region of human-derived LAG3 and a polypeptide or fragment thereof having at least about 85% amino acid identity. An exemplary human-derived LAG3-IgD1 amino acid sequence (SEQ ID NO: 22) is provided below. The term "LAG3-IgD2" refers to the amino acid sequence of the Ig-like C2-type 1 domain in the extracellular region of human-derived LAG3 and a polypeptide or fragment thereof having at least about 85% amino acid identity. An exemplary human-derived LAG3-IgD2 amino acid sequence (SEQ ID NO: 23) is provided below. The term "LAG3-IgD3" refers to the amino acid sequence of the Ig-like C2-type 2 domain in the extracellular region of human-derived LAG3 and a polypeptide or fragment thereof having at least about 85% amino acid identity. An exemplary human-derived LAG3-IgD3 amino acid sequence (SEQ ID NO: 24) is provided below. The term "LAG3-IgD4" refers to a polypeptide or fragment thereof that has an amino acid sequence containing the Ig-like C2-type 3 domain of the human-derived LAG3 extracellular region and has at least about 85% amino acid identity. An exemplary human-derived LAG3-IgD4 amino acid sequence (SEQ ID NO: 25) is provided below. The term "LAG3-IgD1 / D2" refers to a polypeptide or fragment thereof that contains the amino acid sequence of human-derived LAG3-IgD1 and human-derived LAG3-IgD2, has at least about 85% amino acid identity, and exhibits MHCII and / or FGL1 binding activity or activates antigen-presenting cells. An exemplary human-derived LAG3-D1 / D2 amino acid sequence (SEQ ID NO: 26) is provided below. The number of linkages disclosed in the present invention is not limited to the exemplary numbers shown, and any combination of linkages containing the functional domains LAG3-D1 / D2 is considered to be within the scope of the present invention. The term "LAG3-D1 / D2 / D3" refers to a polypeptide or fragment thereof having an amino acid sequence including human-derived LAG3-IgD1, LAG3-IgD2, and LAG3-IgD3, which has at least about 85% amino acid identity and has MHCII and / or FGL1 binding activity.An exemplary amino acid sequence of human-derived LAG3-D1 / D2 / D3 (SEQ ID NO: 27) is provided below. The number of linkages disclosed in the present invention is not limited to the exemplary number, and any combination of linkages comprising the functional domains LAG3-D1 / D2 / D3 is considered to be within the scope of the present invention. The term "LAG3-D1 / D2 / D3 / D4" refers to an amino acid sequence comprising human-derived LAG3-IgD1, LAG3-IgD2, LAG3-IgD3, and LAG3-IgD4, which has at least about 85% amino acid identity and exhibits MHCII and / or FGL1 binding activity and / or activates antigen-presenting cells, or a fragment thereof. An exemplary amino acid sequence of human-derived LAG3-D1 / D2 / D3 / D4 (SEQ ID NO: 28) is provided below. The number of linkages disclosed in the present invention is not limited to the exemplary number, and any combination of linkages comprising the functional domains LAG3-D1 / D2 / D3 / D4 is considered to be within the scope of the present invention.

[0386] As used herein, the term "fusion polypeptide" generally refers to a polypeptide obtained by fusing two or more proteins or polypeptides. As used herein, "fusion polypeptide" can be used interchangeably with "fusion protein." Fusion polypeptides can include fusion polypeptide complexes. Fusion polypeptides can be artificially produced by recombinant DNA technology. For example, genes or nucleic acid molecules encoding the two or more proteins or polypeptides can be linked to each other to form a fusion gene or fusion nucleic acid molecule, which can encode the fusion polypeptide. Translation of the fusion gene produces a single polypeptide, which can have the properties of at least one of the two or more proteins or polypeptides before fusion, and each of them can also have the properties of the other.

[0387] As used herein, the term "bi- and / or multi-functional fusion polypeptide" generally refers to a polypeptide or protein in which one or more polypeptides or domains thereof of different origins are fused. This functional fusion polypeptide comprises an anti-PD-L1 antibody and / or an anti-PD-1 antibody covalently linked via a linker to a functional domain comprising at least the CD86 extracellular domain, or / and at least the LAG3 extracellular domain, or / and at least one LAG3 extracellular structure, wherein two or more of the functional domains comprising LAG3 extracellular domains or / and at least one LAG3 extracellular structure disclosed herein are arranged in tandem with the functional domain comprising LAG3 extracellular domains or / and at least one LAG3 extracellular structure, and the two or more functional domains comprising LAG3 extracellular domains or / and at least one LAG3 extracellular structure are linked via the linker. The bi- and / or multi-functional fusion polypeptides disclosed in the present application, including but not limited to the anti-PD-L1 antibodies and / or anti-PD-1 antibodies, are considered to be within the scope of the present application when any form of anti-PD-L1 antibody and / or anti-PD-1 antibody is covalently linked via a linker to a functional fusion polypeptide comprising the CD86 extracellular domain, and / or the LAG3 extracellular domain, and / or at least one subunit functional domain of the LAG3 extracellular structure, or a combination thereof.

[0388] In this application, the term "antibody-related antigen-binding fragment" generally refers to an amino acid fragment within an antibody that is responsible for specific binding to an antigen. This may be a fragment that determines the primary difference of an antibody molecule and is also referred to as an antigen-binding domain, as well as an "epitope" or "antigenic determinant." An antigen-binding domain is typically composed of an antibody's heavy chain variable region (VH) and light chain variable region (VL), but does not necessarily include both. The antigen-binding domain of the antibody disclosed in this application is not limited to domains composed of conventional VH and VL, but also includes, but is not limited to, antigen-binding domains contained in recombinant antibodies, single-domain antibodies, heavy-chain antibodies, chimeric antibodies, bispecific antibodies, and other atypical antibodies and combinations thereof.

[0389] Furthermore, the anti-PD-1 antibodies and / or anti-PD-L1 antibodies disclosed in the present application are not limited to conventional naturally occurring antibodies, but include any type of antibody having anti-PD-1 and / or anti-PD-L1 properties, such as, but not limited to, recombinant antibodies, single domain antibodies, heavy chain antibodies, chimeric antibodies, bispecific antibodies, other atypical antibodies, and combinations thereof.

[0390] In this application, the term "Fab" refers to an antibody fragment composed of the VL, VH, CL and CH1 domains.

[0391] As used herein, the term "Fab'" refers to an antibody fragment that has a few additional residues at the carboxy terminus of the CH1 domain compared to a Fab fragment. For example, Fab' can include one or more cysteines from the antibody hinge region.

[0392] In this application, the term "F(ab)2" refers to an antigen-binding fragment derived from a pair of Fab fragments linked by cysteines.

[0393] In the present application, the term "dAb fragment" refers to an antibody fragment consisting of only the VH domain (Ward et al., Nature 341: 544-546 (1989)).

[0394] In this application, the term "complementarity-determining region (CDR)" refers to three hypervariable regions (HVRs) present in the light chain variable region (VL) and the heavy chain variable region (VH). Because these regions bind to antigen determinants in a precise complementary manner in a three-dimensional structure, the hypervariable regions are also called complementarity-determining regions.

[0395] In this application, the term "Fv fragment" refers to an antibody fragment consisting of the VL and VH domains that constitute a single arm of an antibody.

[0396] In this application, the term "scFv" refers to a molecule in which the heavy chain variable region and light chain variable region of an antibody are linked by a short peptide linker, also referred to as a single-chain antibody.

[0397] In this application, the term "immunoglobulin" refers to an antibody comprising an amino acid fragment responsible for antigen-specific binding and a constant region fragment. The antigen-specific binding region is a fragment that provides crucial differences in an immunoglobulin and is also referred to as an antigen-binding domain, as well as an "epitope" or "antigenic determinant." An antigen-binding domain is typically composed of an antibody heavy chain variable region (VH) and light chain variable region (VL), but does not necessarily contain both. The antigen-binding domain of the antibody disclosed in the present invention is not limited to conventional domains consisting of VH and VL, but also encompasses antigen-binding domains contained in recombinant antibodies, single-domain antibodies, heavy-chain antibodies, chimeric antibodies, bispecific antibodies, and other non-conventional antibodies, as well as combinations thereof. The constant region refers to a structural region common to immunoglobulins and includes the light chain constant region and heavy chain constant region of an antibody.

[0398] In the present application, the term "immunoglobulin Fc domain" refers to one Fc fragment and two identical Fab fragments produced after conventional IgG antibody is hydrolyzed with the enzyme papain. The Fc domain can include antibody heavy chain CH2, CH3, and hinge region fragments. Conventional Fc fragments induce relevant biological effects through binding to Fc fragment receptors, but their biological function can be influenced by changing their binding ability to the corresponding target receptors through site-directed mutation. The immunoglobulin Fc domains disclosed in the present application are not limited to conventional Fc fragments, but also include all types of Fc variants. In the present application, biological functions that the immunoglobulin Fc domain may provide include, but are not limited to, extending half-life, improving molecular stability, facilitating expression and detection of fusion proteins, mediating passage through placenta and mucosal barriers, mediating antibody-dependent cellular cytotoxicity (ADCC), mediating inflammatory responses, mediating antibody-dependent cellular phagocytosis (ADCP), mediating complement-dependent cytotoxicity (CDC), mediating promotion of dendritic cell (DC) maturation, regulating cytokine secretion, regulating B cell proliferation and differentiation, etc. Exemplary Fc domains of human-derived immunoglobulin IgG1 (SEQ ID NO: 20) and human-derived immunoglobulin IgG4 (SEQ ID NO: 21) are provided below.

[0399] In the present invention, the terms "polypeptide" and "protein" have the same meaning and can be used interchangeably. Also, in the present invention, amino acids are generally represented by one-letter and three-letter abbreviations known in the art. For example, alanine can be represented as A or Ala.

[0400] The proteins, polypeptides and / or amino acid sequences related to the present application are understood to encompass at least the following scope: variants or homologs having the same or similar functions as the proteins or polypeptides in question.

[0401] In this application, the term "variant" generally refers to a peptide essentially similar to the peptide, and can include proteins or polypeptides in which one or more amino acids have been substituted, deleted, or added in the amino acid sequence of a protein and / or polypeptide, such as a specific binding protein antibody or fragment thereof. For example, a functional variant can include a protein or polypeptide having amino acid changes, such as at least one, e.g., 1 to 30, 1 to 20, or 1 to 10, and even more e.g., 1, 2, 3, 4, or 5 amino acids substituted, deleted, and / or inserted. Such functional variants can essentially retain the biological properties of the protein or polypeptide prior to the alteration (e.g., substitution, deletion, or addition). For example, functional variants can retain at least 60%, 70%, 80%, 90%, or 100% of the biological activity (e.g., antigen-binding ability) of the protein or polypeptide prior to the alteration. For example, such substitutions can be conservative substitutions.

[0402] As used herein, the homolog can be a protein or polypeptide that has at least about 85% (e.g., at least about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or more) sequence homology with the amino acid sequence of the protein and / or polypeptide (e.g., an antibody or fragment thereof of a specific binding protein).

[0403] As used herein, homology generally refers to the similarity, similar properties, or relatedness between two or more sequences. The "percentage of sequence identity" can be calculated as follows: Align the two sequences to be compared within a comparison window, and determine the number of positions in the two sequences where the same nucleotide base (e.g., A, T, C, G, I) or the same amino acid residue (e.g., Ala, Pro, Ser, Thr, Gly, Val, Leu, Ile, Phe, Tyr, Trp, Lys, Arg, His, Asp, Glu, Asn, Gln, Cys, Met) occurs. This determines the number of matching positions. Then, divide the number of matching positions by the total number of positions within the comparison window (i.e., the window size), and multiply the result by 100 to calculate the percentage of sequence identity. Comparisons to determine the percentage of sequence identity can be performed using various methods known in the art. For example, publicly available computer software such as BLAST, BLAST-2, ALIGN, or Megalign (DNASTAR) software can be used. Those skilled in the art can set parameters appropriate for the sequences to be compared, including any algorithms necessary to achieve maximum alignment within the full length sequence span or target sequence region to be compared. The homology can also be measured by methods such as, but not limited to, FASTA and BLAST.

[0404] In this application, a chain structure can be formed in a multi-peptide chain by bonding an amino group to a carboxyl group in another peptide chain, but amino acid residues that do not form peptide bonds remain at both ends of the protein, one end of the multi-peptide chain having a free amino group, and the other end of the multi-peptide chain having a free carboxyl group. In this application, the term "N-terminus" generally refers to the end of a multi-peptide chain where an amino acid residue with a free amino group is located. In this application, the term "C-terminus" generally refers to the end of a multi-peptide chain where an amino acid residue with a free carboxyl group is located.

[0405] In this application, the term "nucleic acid molecule" generally refers to nucleotides, deoxyribonucleotides, or ribonucleotides, or analogs thereof, in isolated form of any length, either separated from their natural environment or artificially synthesized.

[0406] In this application, the term "immunoconjugate" generally refers to a polypeptide molecule having one or more heterologous molecules (including, but not limited to, cytotoxins) attached thereto.

[0407] In the present invention, the term "vector" refers to a nucleic acid delivery tool into which a polynucleotide encoding a specific protein is inserted so that the protein can be expressed.

[0408] In this application, the term "linker" refers to a molecule that connects one or more polypeptides or domains thereof. For example, the linker is structurally flexible and can include a short peptide chain formed by a combination of glycine (Gly, G) and serine (Ser, S) residues. In this case, the ratio of the number of glycines (Gly) to the number of serines (Ser) can be ≧1. The linkers disclosed in this application can be extended to any short peptide that has this property. The amino acid sequences of exemplary linkers are shown below, but are not limited to these: GGGGS (SEQ ID NO: 22), GGGGSGGGGS (SEQ ID NO: 23), GGGGSGGGGSGGGGS (SEQ ID NO: 24), GGGGSGGGGSGGGGSGGGGS (SEQ ID NO: 25), GGGGSGGGS (SEQ ID NO: 26). In this application, the term "CD3" generally refers to a protein complex composed of four independent chains. In mammals, this complex includes one CD3γ chain, one CD3δ chain, and two CD3ε chains. These chains bind to the T cell receptor (TCR) and ζ chain to generate an activation signal in T lymphocytes. The TCR, ζ chain, and a portion of the CD3 molecule form the TCR complex. As used herein, the term "CD3" is intended to encompass any native CD3 of human origin. This term encompasses not only the "full-length" and unprocessed protein, but also all intracellularly processed forms of the protein and one or more CD3 chains (polypeptides) (e.g., mature polypeptides). It also encompasses naturally occurring variants and isoforms of CD3, such as splice variants and allelic variants. For example, www.uniprot.org / uniprot / P04234, www.uniprot.org / uniprot / P07766, www.uniprot.org / uniprot / P09693 provide descriptions of the CD3 gamma, delta, and epsilon chains and their sequences.

[0409] In this application, the term "anti-CD3 antibody" includes not only antibodies and antigen-binding fragments thereof that specifically recognize a single CD3 subunit (e.g., β, δ, γ, or ζ), but also antibodies and antigen-binding fragments thereof that specifically recognize two CD3 subunits (e.g., γ / ε, δ / ε, and ζ / ζ CD3 dimers). For example, monoclonal antibodies that target the CD3 receptor within the T cell antigen receptor of mature T cells may include human, humanized, chimeric, or murine antibodies. For example, the anti-CD3 antibody may be a CD3B219 antibody or an antigen-binding fragment thereof.

[0410] In this application, the term "amino acid mutation" generally refers to amino acid substitution, deletion, insertion, and modification. Any combination of substitution, deletion, insertion, and modification can be used to obtain a final construct with desired properties. In one embodiment, the amino acid mutation is a substitution. The term "amino acid mutation at a specific position" refers to the substitution or deletion of a specific residue, or the insertion of at least one amino acid residue at a position adjacent to the specific residue. Amino acid substitutions can be conservative or non-conservative. For example, "A13I" indicates that alanine (A, Ala) at position 13 is substituted with isoleucine (I, Ile).

[0411] In this application, the term "active fragment" generally refers to a nucleic acid or amino acid fragment or variant thereof that has a particular biological activity or function. For example, a functionally active fragment can retain or partially retain the ability of the full-length protein to bind to other molecules.

[0412] In the present application, the term "CD86 active fragment" refers to a polypeptide or a fragment thereof, which is a CD86 IgV or CD86 ECD protein or its derived mutant polypeptide described in the present invention, having binding activity to CD28 and / or CTLA4, including, but not limited to, properties such as regulating T cell activity, activating the immune system, and antitumor activity.

[0413] In this application, "LAG3 active fragment" refers to a polypeptide or fragment thereof that activates antigen-presenting cells, such as the LAG3 protein and its derived mutant polypeptides described in the present invention, and includes, but is not limited to, properties such as regulating the activity of antigen-presenting cells, activating the immune system, and anti-tumor activity.

[0414] The term "CD86 mutant polypeptide" refers to a protein or polypeptide in which one or more amino acids have been substituted, deleted, or added in the amino acid sequence of wild-type CD86. For example, such functional variants can include proteins or polypeptides having amino acid changes due to substitution, deletion, and / or insertion of at least one, e.g., 1-30, 1-20, or 1-10, and even more particularly, 1, 2, 3, 4, or 5 amino acids. Such functional variants can essentially retain the biological properties of CD86 prior to the alteration (e.g., substitution, deletion, or addition). For example, such functional variants can retain at least 60%, 70%, 80%, 90%, or 100% of the biological activity (e.g., antigen-binding ability) of the protein or polypeptide prior to the alteration. For example, such substitutions may be conservative. A "CD86 mutant polypeptide" refers to an active fragment derived from the CD86 protein and possessing functional properties of the CD86 protein, including, but not limited to, CD28 and / or CTLA4 binding activity.

[0415] The term "mutant polypeptide fusion polypeptide complex" is also referred to herein as "mutant polypeptide complex," "polypeptide complex," "fusion polypeptide complex," or "fusion complex." A "CD86 mutant fusion polypeptide complex" refers to a functional fusion polypeptide complex in which a CD86 ECD mutant or a CD86 IgV mutant is linked, with or without a linker, to an antibody or antigen-binding fragment thereof, an immunoglobulin Fc domain, or a functional polypeptide fragment. In functional fusion polypeptide complexes in which the CD86 variants disclosed in the present invention are covalently linked to an antibody or its antigen-binding fragment with or without a linker, the antibody includes, but is not limited to, an anti-PD-L1 antibody, an anti-PD-1 antibody, an anti-TIGIT antibody, an anti-CTLA4 antibody, an anti-CEA antibody, an anti-BCMA antibody, an anti-LAG3 antibody, an anti-CD3 antibody, an anti-Her2 antibody, an anti-Her3 antibody, an anti-VEGF antibody, an anti-VEGFR antibody, an anti-EGFR antibody, an anti-c-Met antibody, an anti-CD19 antibody, an anti-CD20 antibody, an anti-CD38 antibody, an anti-TROP-2 antibody, an anti-CD40 antibody, an anti-4-1BB antibody, an anti-CD30 antibody, etc. Functional fusion proteins in which any type of antibody is covalently linked with the CD86 ECD variants or CD86 IgV variants disclosed in the present invention with or without a linker, or combinations thereof, are considered to be within the scope of the present invention.

[0416] The term "LAG3 variant polypeptide" refers to a protein or polypeptide in which one or more amino acids have been substituted, deleted, or added in the amino acid sequence of wild-type LAG3. For example, such functional variants can include proteins or polypeptides in which at least one, e.g., 1-30, 1-20, or 1-10 amino acids have been substituted, deleted, and / or inserted, or in which one, two, three, four, or five amino acids have been changed. Such functional variants can essentially retain the biological properties of LAG3 prior to the substitution, deletion, or addition. For example, such functional variants can maintain at least 60%, 70%, 80%, 90%, or 100% of the biological activity (e.g., antigen-binding ability) of the protein or polypeptide prior to the alteration. For example, such substitutions can be conservative. A "LAG3 variant polypeptide" refers to an active fragment derived from a LAG3 protein and possessing functional properties of the LAG3 protein, including, but not limited to, promoting the activation and maturation of antigen-presenting cells. The term "LAG3 variant polypeptide" refers to an active fragment derived from the LAG3 protein and having functional properties of the LAG3 protein, including, but not limited to, binding to MHCII (or HLA-DR) and / or FGL1, and promoting the activation and maturation of antigen-presenting cells.

[0417] As used herein, the term "LAG3 mutant fusion polypeptide complex" generally refers to a polypeptide or protein to which one or more derived polypeptides or domains thereof are fused. The term "fusion polypeptide complex of mutant polypeptides" is also referred to herein as a "mutant polypeptide complex," "polypeptide complex," "fusion polypeptide complex," or "fusion complex." The LAG3 mutant polypeptide contains at least an Fc domain, with or without a linker, and / or at least an antibody or antigen-binding fragment thereof. The LAG3 mutant fusion polypeptide complexes disclosed herein include, but are not limited to, a LAG3 mutant polypeptide and any type of antibody, functional protein fragment, and / or functional fusion polypeptide in which at least one subunit functional domain of the Fc domain is covalently linked, or a combination thereof. All of these are intended to be within the scope of this application.

[0418] The activation and maturation of antigen-presenting cells in the present invention includes, but is not limited to, effects such as increased expression of costimulatory receptors in antigen-presenting cells, improved antigen phagocytic ability, and enhanced cytokine and chemokine secretion.

[0419] Costimulatory receptors in the present invention include, but are not limited to, ICOSL, CD40L, CD137L, OX40L, CD80, CD83, CD86, and the like.

[0420] In the present invention, phagocytosis of antigens includes, but is not limited to, phagocytosis of bacteria, viruses, proteins, polysaccharides, and the like.

[0421] Cytokines in the present invention include, but are not limited to, IL-1beta, TNFα, IFNγ, IL-6, IL-12, and the like.

[0422] Chemokines in the present invention include, but are not limited to, CCL1, CCL2, CCL3, CCL4, CCL5, CXCL1, CXCL2, CXCL3, CXCL4, CXCL5, and the like.

[0423] In the present application, the expression "X amino acids truncated at the N-terminus" refers to the deletion of amino acid residues 1 to X at the N-terminus of the extracellular domain of a protein. For example, the expression "LAG3 extracellular domain polypeptide truncated by 74 amino acids at the N-terminus" can refer to a mutant polypeptide in which the amino acid residues at positions 23-96 in the NCBI numbering system of a wild-type LAG3 extracellular domain polypeptide are deleted at the N-terminus. For example, the expression "LAG3 extracellular domain polypeptide truncated by 74 amino acids at the N-terminus" can refer to a polypeptide starting from amino acid 97 of a wild-type human-derived LAG3 polypeptide. For example, a "LAG3 extracellular domain polypeptide truncated by 74 amino acids at the N-terminus" can be expressed as "LAG3 97-450."

[0424] As used herein, the phrase "the C-terminus ends at amino acid position X" means that all amino acid residues from amino acid position X onward (i.e., amino acid position X+1 onward) are deleted. For example, the phrase "the C-terminus ends at amino acid position 156 of the LAG3 extracellular region polypeptide" can refer to a mutant polypeptide obtained by deleting all amino acid residues from amino acid position NCBI number 156 onward (i.e., amino acid position 157 onward) of a wild-type LAG3 extracellular region polypeptide. For example, the phrase "the C-terminus ends at amino acid position 156 of the LAG3 extracellular region polypeptide" can mean that the C-terminus of the wild-type LAG3 extracellular region polypeptide has been shortened by 294 amino acids. For example, the phrase "the C-terminus ends at amino acid position 156 of the LAG3 extracellular region polypeptide" can be expressed as "LAG3 23-156." Furthermore, the phrase "the N-terminus is shortened by 74 amino acids, and the C-terminus ends at amino acid position 156" can be expressed as "LAG3 97-156."

[0425] As used herein, the term "comprising" means including the features explicitly stated but not excluding other elements. As used herein, the term "about" means varying within 0.5%-10% above or below the specified numerical value, including, for example, varying within 0.5%, 1%, 1.5%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%, 5%, 5.5%, 6%, 6.5%, 7%, 7.5%, 8%, 8.5%, 9%, 9.5% or 10% above or below the specified numerical value.

[0426] On the other hand, the present application provides a fusion polypeptide, which can comprise a first domain and a second domain, wherein the first domain can block PD-1 / PD-L1 signaling, and the second domain comprises CD86 or a functionally active fragment thereof. For example, the fusion polypeptide can comprise one or more of the first domains and one or more of the second domains.

[0427] For example, the first domain can bind to PD-L1 and / or PD-1.

[0428] For example, the first domain may comprise an HCDR3 of an antibody heavy chain, and the antibody heavy chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0429] For example, the first domain may comprise HCDR2 of an antibody heavy chain, and the antibody heavy chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0430] For example, the first domain may comprise HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0431] For example, the first domain may comprise a heavy chain variable region VH of an antibody heavy chain, and the antibody heavy chain variable region may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0432] For example, the first domain may comprise an antibody heavy chain, and the antibody heavy chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0433] For example, the first domain may comprise an LCDR3 of an antibody light chain, and the antibody light chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0434] For example, the first domain may comprise LCDR2 of an antibody light chain, and the antibody light chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0435] For example, the first domain may comprise LCDR1 of an antibody light chain, and the antibody light chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0436] For example, the first domain may comprise a light chain variable region VL of an antibody light chain, and the antibody light chain variable region may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0437] For example, the first domain may comprise an antibody light chain, and the antibody light chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0438] For example, the first domain may comprise HCDR3, HCDR2 and HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 4. For example, the antibody may be Sugemalimab.

[0439] For example, the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, and the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 5. For example, the antibody may be Sugemalimab.

[0440] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, wherein the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 4, and the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, wherein the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 5. For example, the antibody may be sugemalimab.

[0441] For example, the first domain may comprise a heavy chain variable region VH of an antibody heavy chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 4, and the first domain may comprise a light chain variable region VL of an antibody light chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 5. For example, the antibody may be sugemalimab.

[0442] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 6. For example, the antibody may be JS003.

[0443] For example, the first domain can include LCDR3, LCDR2, and LCDR1 of an antibody light chain, and the antibody light chain can include the amino acid sequence set forth in SEQ ID NO: 7. For example, the antibody can be JS003.

[0444] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, where the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 6, and the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, where the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 7. For example, the antibody may be JS003.

[0445] For example, the first domain may comprise a heavy chain variable region VH of an antibody heavy chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 6, and the first domain may comprise a light chain variable region VL of an antibody light chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 7. For example, the antibody may be JS003.

[0446] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 8. For example, the antibody may be durvalumab.

[0447] For example, the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, and the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 9. For example, the antibody may be durvalumab.

[0448] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, wherein the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 8, and the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, wherein the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 9. For example, the antibody may be durvalumab.

[0449] For example, the first domain may comprise a heavy chain variable region, VH, of an antibody heavy chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 8, and the first domain may comprise a light chain variable region, VL, of an antibody light chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 9. For example, the antibody may be durvalumab.

[0450] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 10. For example, the antibody may be Atezolizumab.

[0451] For example, the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, and the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 11. For example, the antibody may be Atezolizumab.

[0452] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, wherein the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 10, and the first domain may comprise antibody LCDR3, LCDR2, and HCDR1, wherein the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 11. For example, the antibody may be atezolizumab.

[0453] For example, the first domain may comprise a heavy chain variable region, VH, of an antibody heavy chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 10, and the first domain may comprise a light chain variable region, VL, of an antibody light chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 11. For example, the antibody may be atezolizumab.

[0454] For example, the first domain may comprise HCDR3, HCDR2 and HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 12. For example, the antibody may be Avelumab.

[0455] For example, the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, and the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 13. For example, the antibody may be Avelumab.

[0456] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, wherein the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 12, and the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, wherein the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 13. For example, the antibody may be Avelumab.

[0457] For example, the first domain may comprise a heavy chain variable region, VH, of an antibody heavy chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 12, and the first domain may comprise a light chain variable region, VL, of an antibody light chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 13. For example, the antibody may be Avelumab.

[0458] For example, the first domain may comprise HCDR3, HCDR2 and HCDR1 of an antibody, and the antibody may comprise the amino acid sequence set forth in SEQ ID NO: 14. For example, the antibody may be Envafolimab.

[0459] For example, the first domain may comprise HCDR3, HCDR2 and HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 15. For example, the antibody may be Permbrolizumab.

[0460] For example, the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, and the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 16. For example, the antibody may be Permbrolizumab.

[0461] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, wherein the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 15, and the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, wherein the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 16. For example, the antibody may be permbrolizumab.

[0462] For example, the first domain may comprise a heavy chain variable region, VH, of an antibody heavy chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 15, and the first domain may comprise a light chain variable region, VL, of an antibody light chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 16. For example, the antibody may be permbrolizumab.

[0463] For example, the first domain may comprise HCDR3, HCDR2 and HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 17. For example, the antibody may be Nivolumab.

[0464] For example, the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, and the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 18. For example, the antibody may be Nivolumab.

[0465] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, wherein the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 17, and the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, wherein the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 18. For example, the antibody may be Nivolumab.

[0466] For example, the first domain may comprise a heavy chain variable region, VH, of an antibody heavy chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 17, and the first domain may comprise a light chain variable region, VL, of an antibody light chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 18. For example, the antibody may be Nivolumab.

[0467] For example, the first domain may comprise an antibody or an antigen-binding fragment thereof.

[0468] For example, the antibody may be selected from the group consisting of a recombinant antibody, a single domain antibody, a heavy chain antibody, a chimeric antibody, and a bispecific antibody.

[0469] For example, the antigen-binding fragment may be selected from one or more of the following group: Fab, Fab', Fv fragment, F(ab')2, F(ab)2, scFv, di-scFv, VHH and dAb.

[0470] For example, the first domain is an antibody or an antigen thereof selected from the group consisting of: Binding fragments may include: sugemalimab, durvalumab, atezolizumab, avelumab, embafolimab, permbrolizumab and nivolumab.

[0471] For example, the first domain may comprise sugemalimab or an antigen-binding fragment thereof.

[0472] For example, the second domain may comprise CD86 or a functionally active fragment thereof.

[0473] For example, the second domain may be selected from the group consisting of CD86 of human origin, or a functionally active fragment thereof, and CD86 of mouse origin, or a functionally active fragment thereof.

[0474] For example, the second domain may bind to CD28 and / or CTLA4.

[0475] For example, the second domain may comprise the IgV domain of CD86 or a functionally active fragment thereof.

[0476] For example, the second domain may comprise the extracellular domain of CD86 or a functionally active fragment thereof.

[0477] For example, the second domain may comprise the amino acid sequence shown in SEQ ID NO:2 or SEQ ID NO:3.

[0478] For example, the first domain and the second domain may be directly or indirectly connected.

[0479] For example, the N-terminus of the first domain and the N-terminus of the second domain may be directly or indirectly connected.

[0480] For example, the C-terminus of the first domain and the C-terminus of the second domain may be directly or indirectly connected.

[0481] For example, the first domain may comprise an antibody heavy chain, and the antibody heavy chain of the first domain is directly or indirectly bound to the second domain.

[0482] For example, the first domain may comprise an antibody heavy chain, and the C-terminus of the antibody heavy chain of the first domain is connected directly or indirectly to the N-terminus of the second domain.

[0483] For example, the first domain can comprise an antibody heavy chain, and the N-terminus of the antibody heavy chain of the first domain is connected directly or indirectly to the C-terminus of the second domain.

[0484] For example, the first domain may comprise an antibody light chain, and the first domain antibody light chain and the second domain may be directly or indirectly connected.

[0485] For example, the first domain comprises an antibody light chain, and the C-terminus of the antibody light chain of the first domain is connected directly or indirectly to the N-terminus of the second domain.

[0486] For example, the first domain comprises an antibody light chain, and the N-terminus of the antibody light chain of the first domain is connected directly or indirectly to the C-terminus of the second domain.

[0487] For example, the indirect connection can include a connection via a linker.

[0488] For example, the linker may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34, and SEQ ID NO:35.

[0489] For example, the fusion polypeptide may comprise an amino acid sequence selected from the following group: SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, SEQ ID NO:42, SEQ ID NO:43, SEQ ID NO:44, SEQ ID NO:45, SEQ ID NO:46, SEQ ID NO:47, SEQ ID NO:48, SEQ ID NO:49, SEQ ID NO:50, SEQ ID NO:51, SEQ ID NO:52, SEQ ID NO:53, SEQ ID NO:54, SEQ ID NO:55, SEQ ID NO:56, SEQ ID NO:57, SEQ ID NO:58, SEQ ID NO:59, SEQ ID NO:60, SEQ ID NO:61, SEQ ID NO:62, SEQ ID NO:63, SEQ ID NO:64, SEQ ID NO:65, SEQ ID NO:66, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:74, SEQ ID NO:75, SEQ ID NO:76, SEQ ID NO:77, SEQ ID NO:78, SEQ ID NO:79, SEQ ID NO:80, SEQ ID NO:81, SEQ ID NO:82, SEQ ID NO:83, SEQ ID NO:84, SEQ ID NO:85, SEQ ID NO:86 and SEQ ID NO:87.

[0490] For example, the fusion polypeptide can comprise an amino acid sequence selected from the group consisting of SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, SEQ ID NO:42, SEQ ID NO:43, SEQ ID NO:44, and SEQ ID NO:77.

[0491] Meanwhile, the present application further provides a fusion polypeptide, which may comprise a first domain, a second domain, and a third domain. The first domain can inhibit PD-1 / PD-L1 signaling, the second domain comprises CD86 or a fragment thereof having functional activity, and the third domain can activate an innate immune response. For example, the fusion polypeptide may comprise one or more of the first domains, one or more of the second domains, and one or more of the third domains.

[0492] For example, the first domain can bind to PD-L1 and / or PD-1.

[0493] For example, the first domain may comprise an HCDR3 of an antibody heavy chain, and the antibody heavy chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0494] For example, the first domain may comprise HCDR2 of an antibody heavy chain, and the antibody heavy chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0495] For example, the first domain may comprise HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0496] For example, the first domain may comprise a heavy chain variable region VH of an antibody heavy chain, and the antibody heavy chain variable region may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17. For example, the first domain may comprise an antibody heavy chain, and the antibody heavy chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:17.

[0497] For example, the first domain may comprise an LCDR3 of an antibody light chain, and the antibody light chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0498] For example, the first domain may comprise LCDR2 of an antibody light chain, and the antibody light chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0499] For example, the first domain may comprise LCDR1 of an antibody light chain, and the antibody light chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0500] For example, the first domain may comprise a light chain variable region VL of an antibody light chain, and the antibody light chain variable region may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0501] For example, the first domain may comprise an antibody light chain, and the antibody light chain may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:18.

[0502] For example, the first domain may comprise HCDR3, HCDR2 and HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 4. For example, the antibody may be Sugemalimab.

[0503] For example, the first domain can comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, and the antibody light chain can comprise the amino acid sequence set forth in SEQ ID NO: 5. For example, the antibody can be Sugemalimab.

[0504] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, wherein the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 4, and the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, wherein the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 5. For example, the antibody may be sugemalimab.

[0505] For example, the first domain may comprise a heavy chain variable region VH of an antibody heavy chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 4, and the first domain may comprise a light chain variable region VL of an antibody light chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 5. For example, the antibody may be sugemalimab.

[0506] For example, the first domain may comprise HCDR3, HCDR2 and HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 6. For example, the antibody may be JS003.

[0507] For example, the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, and the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 7. For example, the antibody may be JS003.

[0508] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, where the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 6, and the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, where the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 7. For example, the antibody may be JS003.

[0509] For example, the first domain may comprise a heavy chain variable region VH of an antibody heavy chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 6, and the first domain may comprise a light chain variable region VL of an antibody light chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 7. For example, the antibody may be JS003.

[0510] For example, the first domain may comprise HCDR3, HCDR2 and HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 8. For example, the antibody may be durvalumab.

[0511] For example, the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, and the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 9. For example, the antibody may be durvalumab.

[0512] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, wherein the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 8, and the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, wherein the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 9. For example, the antibody may be durvalumab.

[0513] For example, the first domain may comprise a heavy chain variable region, VH, of an antibody heavy chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 8, and the first domain may comprise a light chain variable region, VL, of an antibody light chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 9. For example, the antibody may be durvalumab.

[0514] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 10. For example, the antibody may be Atezolizumab.

[0515] For example, the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, and the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 11. For example, the antibody may be Atezolizumab.

[0516] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, wherein the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 10, and the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, wherein the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 11. For example, the antibody may be atezolizumab.

[0517] For example, the first domain may comprise a heavy chain variable region, VH, of an antibody heavy chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 10, and the first domain may comprise a light chain variable region, VL, of an antibody light chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 11. For example, the antibody may be atezolizumab.

[0518] For example, the first domain may comprise HCDR3, HCDR2 and HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 12. For example, the antibody may be Avelumab.

[0519] For example, the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, and the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 13. For example, the antibody may be Avelumab.

[0520] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, wherein the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 12, and the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, wherein the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 13. For example, the antibody may be Avelumab.

[0521] For example, the first domain may comprise a heavy chain variable region, VH, of an antibody heavy chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 12, and the first domain may comprise a light chain variable region, VL, of an antibody light chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 13. For example, the antibody may be Avelumab.

[0522] For example, the first domain may comprise HCDR3, HCDR2 and HCDR1 of an antibody, and the antibody may comprise the amino acid sequence set forth in SEQ ID NO: 14. For example, the antibody may be Envafolimab.

[0523] For example, the first domain may comprise HCDR3, HCDR2 and HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 15. For example, the antibody may be Permbrolizumab.

[0524] For example, the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, and the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 16. For example, the antibody may be Permbrolizumab.

[0525] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, wherein the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 15, and the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, wherein the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 16. For example, the antibody may be permbrolizumab.

[0526] For example, the first domain may comprise a heavy chain variable region, VH, of an antibody heavy chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 15, and the first domain may comprise a light chain variable region, VL, of an antibody light chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 16. For example, the antibody may be permbrolizumab.

[0527] For example, the first domain may comprise HCDR3, HCDR2 and HCDR1 of an antibody heavy chain, and the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 17. For example, the antibody may be Nivolumab.

[0528] For example, the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, and the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 18. For example, the antibody may be Nivolumab.

[0529] For example, the first domain may comprise HCDR3, HCDR2, and HCDR1 of an antibody heavy chain, wherein the antibody heavy chain may comprise the amino acid sequence set forth in SEQ ID NO: 17, and the first domain may comprise LCDR3, LCDR2, and LCDR1 of an antibody light chain, wherein the antibody light chain may comprise the amino acid sequence set forth in SEQ ID NO: 18. For example, the antibody may be Nivolumab.

[0530] For example, the first domain may comprise a heavy chain variable region, VH, of an antibody heavy chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 17, and the first domain may comprise a light chain variable region, VL, of an antibody light chain, which may comprise the amino acid sequence set forth in SEQ ID NO: 18. For example, the antibody may be Nivolumab.

[0531] For example, the first domain may comprise an antibody or an antigen-binding fragment thereof.

[0532] For example, the antibody may be selected from the group consisting of a recombinant antibody, a single domain antibody, a heavy chain antibody, a chimeric antibody, and a bispecific antibody.

[0533] For example, the antigen-binding fragment can be one or more selected from the following group: Fab, Fab', Fv fragment, F(ab')2, F(ab)2, scFv, di-scFv, VHH, and dAb.

[0534] For example, the first domain can comprise an antibody or antigen-binding fragment thereof selected from the group consisting of Sugemalimab, Durvalumab, Atezolizumab, Avelumab, Envafolimab, Pembrolizumab, and Nivolumab.

[0535] For example, the first domain may comprise sugemalimab or an antigen-binding fragment thereof.

[0536] For example, the second domain may comprise CD86 or a functionally active fragment thereof.

[0537] For example, the second domain may comprise one selected from the following group: CD86 of human origin or a functionally active fragment thereof, CD86 of mouse origin or a functionally active fragment thereof.

[0538] For example, the second domain can bind to CD28 and / or CTLA4.

[0539] For example, the second domain may comprise the IgV domain of CD86 or a functionally active fragment thereof.

[0540] For example, the second domain may comprise the extracellular domain of CD86 or a functionally active fragment thereof.

[0541] For example, the second domain may comprise the amino acid sequence shown in SEQ ID NO:2 or SEQ ID NO:3.

[0542] For example, the third domain can bind to an MHC II molecule on an antigen-presenting cell and / or activate antigen presentation.

[0543] For example, the third domain may comprise LAG3 or a functionally active fragment thereof.

[0544] For example, the third domain may be selected from the group consisting of: LAG3 of human origin or a functionally active fragment thereof, and LAG3 of mouse origin or a functionally active fragment thereof.

[0545] For example, the third domain may comprise the extracellular domain of LAG3 or a functionally active fragment thereof.

[0546] For example, the third domain can include IgD1, IgD2, IgD3, and / or IgD4 of LAG3 or functionally active fragments thereof.

[0547] For example, the third domain may comprise IgD1, IgD1-IgD2, IgD1-IgD2-IgD3, and / or IgD1-IgD2-IgD3-IgD4 of LAG3 or functionally active fragments thereof.

[0548] For example, the third domain may be selected from the amino acid sequences set forth in the following group: SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, and SEQ ID NO:28.

[0549] For example, the first domain and the second domain can be linked directly or indirectly.

[0550] For example, the first domain comprises an antibody heavy chain, and the antibody heavy chain of the first domain can bind directly or indirectly to the second domain.

[0551] For example, the first domain may comprise an antibody heavy chain, and the C-terminus of the antibody heavy chain of the first domain may be directly or indirectly linked to the N-terminus of the second domain.

[0552] For example, the first domain may comprise an antibody heavy chain, and the N-terminus of the antibody heavy chain of the first domain may be directly or indirectly linked to the C-terminus of the second domain.

[0553] For example, the first domain comprises an antibody light chain, and the antibody light chain of the first domain can bind directly or indirectly to the second domain.

[0554] For example, the first domain may comprise an antibody light chain, and the C-terminus of the antibody light chain of the first domain may be directly or indirectly linked to the N-terminus of the second domain.

[0555] For example, the first domain may comprise an antibody light chain, and the N-terminus of the antibody light chain of the first domain may be linked directly or indirectly to the C-terminus of the second domain.

[0556] For example, the first domain may comprise an antibody heavy chain and an antibody light chain, and the C-terminus of the antibody heavy chain of the first domain may be directly or indirectly bound to the N-terminus of the second domain, and the C-terminus of the antibody light chain of the first domain may be directly or indirectly bound to the N-terminus of the second domain.

[0557] For example, the first domain may comprise an antibody heavy chain and an antibody light chain, and the N-terminus of the antibody heavy chain of the first domain may be directly or indirectly bound to the C-terminus of the second domain, and the N-terminus of the antibody light chain of the first domain may be directly or indirectly bound to the C-terminus of the second domain.

[0558] For example, the first domain can bind directly or indirectly to the third domain.

[0559] For example, the first domain comprises an antibody heavy chain, and the antibody heavy chain of the first domain can bind directly or indirectly to the third domain.

[0560] For example, the first domain comprises an antibody heavy chain, and the C-terminus of the antibody heavy chain of the first domain can be directly or indirectly linked to the N-terminus of the third domain.

[0561] For example, the first domain may comprise an antibody heavy chain, and the N-terminus of the antibody heavy chain of the first domain may be directly or indirectly linked to the C-terminus of the third domain.

[0562] For example, the first domain comprises an antibody light chain, and the antibody light chain of the first domain can bind directly or indirectly to the third domain.

[0563] For example, the first domain may comprise an antibody light chain, and the C-terminus of the antibody light chain of the first domain may be directly or indirectly linked to the N-terminus of the third domain.

[0564] For example, the first domain may comprise an antibody light chain, and the N-terminus of the antibody light chain of the first domain may be directly or indirectly linked to the C-terminus of the third domain.

[0565] For example, the second domain and the third domain can be linked directly or indirectly.

[0566] For example, the C-terminus of the second domain can be directly or indirectly linked to the N-terminus of the third domain.

[0567] For example, the N-terminus of the second domain can be directly or indirectly linked to the C-terminus of the third domain.

[0568] For example, the first domain can bind directly or indirectly to the second domain, and the second domain can bind directly or indirectly to the third domain.

[0569] For example, the C-terminus of the first domain can be directly or indirectly linked to the N-terminus of the second domain, and the C-terminus of the second domain can be directly or indirectly linked to the N-terminus of the third domain.

[0570] For example, the first domain may comprise an antibody heavy chain, the C-terminus of the antibody heavy chain of the first domain may be directly or indirectly linked to the N-terminus of the second domain, and the C-terminus of the second domain may be directly or indirectly linked to the N-terminus of the third domain.

[0571] For example, the first domain may comprise an antibody light chain, the C-terminus of the antibody light chain of the first domain may be directly or indirectly linked to the N-terminus of the second domain, and the C-terminus of the second domain may be directly or indirectly linked to the N-terminus of the third domain.

[0572] For example, the N-terminus of the first domain can be directly or indirectly linked to the C-terminus of the second domain, and the N-terminus of the second domain can be directly or indirectly linked to the C-terminus of the third domain.

[0573] For example, the first domain may comprise an antibody heavy chain, the N-terminus of the antibody heavy chain of the first domain may be directly or indirectly linked to the C-terminus of the second domain, and the N-terminus of the second domain may be directly or indirectly linked to the C-terminus of the third domain.

[0574] For example, the first domain may comprise an antibody light chain, the N-terminus of the antibody light chain of the first domain may be directly or indirectly linked to the C-terminus of the second domain, and the N-terminus of the second domain may be directly or indirectly linked to the C-terminus of the third domain.

[0575] For example, the first domain can bind directly or indirectly to the third domain, and the third domain can bind directly or indirectly to the second domain.

[0576] For example, the C-terminus of the first domain can be directly or indirectly linked to the N-terminus of the third domain, and the C-terminus of the third domain can be directly or indirectly linked to the N-terminus of the second domain.

[0577] For example, the first domain may comprise an antibody heavy chain, the C-terminus of the antibody heavy chain of the first domain may be directly or indirectly bound to the N-terminus of the third domain, and the C-terminus of the third domain may be directly or indirectly bound to the N-terminus of the second domain.

[0578] For example, the first domain may comprise an antibody light chain, the C-terminus of the antibody light chain of the first domain may be directly or indirectly linked to the N-terminus of the third domain, and the C-terminus of the third domain may be directly or indirectly linked to the N-terminus of the second domain.

[0579] For example, the N-terminus of the first domain can be directly or indirectly linked to the C-terminus of the third domain, and the N-terminus of the third domain can be directly or indirectly linked to the C-terminus of the second domain.

[0580] For example, the first domain may comprise an antibody heavy chain, the N-terminus of the antibody heavy chain of the first domain may be directly or indirectly linked to the C-terminus of the third domain, and the N-terminus of the third domain may be directly or indirectly linked to the C-terminus of the second domain.

[0581] For example, the first domain may comprise an antibody light chain, the N-terminus of the antibody light chain of the first domain may be directly or indirectly linked to the C-terminus of the third domain, and the N-terminus of the third domain may be directly or indirectly linked to the C-terminus of the second domain.

[0582] For example, the first domain can bind directly or indirectly to the second domain, and the first domain can bind directly or indirectly to the third domain.

[0583] For example, the C-terminus of the first domain can be directly or indirectly linked to the N-terminus of the second domain, and the N-terminus of the first domain can be directly or indirectly linked to the C-terminus of the third domain.

[0584] For example, the first domain may comprise an antibody heavy chain, the C-terminus of the antibody heavy chain of the first domain may be directly or indirectly bound to the N-terminus of the second domain, and the N-terminus of the antibody heavy chain of the first domain may be directly or indirectly bound to the C-terminus of the third domain.

[0585] For example, the first domain comprises an antibody light chain, the C-terminus of the antibody light chain of the first domain can be directly or indirectly linked to the N-terminus of the second domain, and the N-terminus of the antibody light chain of the first domain can be directly or indirectly linked to the C-terminus of the third domain.

[0586] For example, the first domain may comprise an antibody heavy chain, the C-terminus of the antibody heavy chain of the first domain may be directly or indirectly bound to the N-terminus of the second domain, and the N-terminus of the antibody light chain of the first domain may be directly or indirectly bound to the C-terminus of the third domain.

[0587] For example, the first domain may comprise an antibody light chain, the C-terminus of the antibody light chain of the first domain may be directly or indirectly linked to the N-terminus of the second domain, and the N-terminus of the antibody heavy chain of the first domain may be directly or indirectly linked to the C-terminus of the third domain.

[0588] For example, the N-terminus of the first domain can be directly or indirectly linked to the C-terminus of the second domain, and the C-terminus of the first domain can be directly or indirectly linked to the N-terminus of the third domain.

[0589] For example, the first domain may comprise an antibody heavy chain, the N-terminus of the antibody heavy chain of the first domain may be directly or indirectly bound to the C-terminus of the second domain, and the C-terminus of the antibody heavy chain of the first domain may be directly or indirectly bound to the N-terminus of the third domain.

[0590] For example, the first domain may comprise an antibody light chain, the N-terminus of the antibody light chain of the first domain may be directly or indirectly linked to the C-terminus of the second domain, and the C-terminus of the antibody light chain of the first domain may be directly or indirectly linked to the N-terminus of the third domain.

[0591] For example, the first domain may comprise an antibody heavy chain and an antibody light chain, and the N-terminus of the antibody heavy chain of the first domain may be directly or indirectly bound to the C-terminus of the second domain, and the C-terminus of the antibody light chain of the first domain may be directly or indirectly bound to the N-terminus of the third domain.

[0592] For example, the first domain may comprise an antibody heavy chain and an antibody light chain, and the N-terminus of the antibody light chain of the first domain may be directly or indirectly bound to the C-terminus of the second domain, and the C-terminus of the antibody heavy chain of the first domain may be directly or indirectly bound to the N-terminus of the third domain.

[0593] For example, the indirect attachment may be via a linker.

[0594] For example, the linker can comprise an amino acid sequence selected from the group consisting of SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34 and SEQ ID NO:35.

[0595] For example, the fusion polypeptide may be selected from the group consisting of SEQ ID NO:88, SEQ ID NO:89, SEQ ID NO:90, SEQ ID NO:91, SEQ ID NO:92, SEQ ID NO:93, SEQ ID NO:94, SEQ ID NO:95, SEQ ID NO:96, SEQ ID NO:97, SEQ ID NO:98, SEQ ID NO:99, SEQ ID NO:100, SEQ ID NO:101, SEQ ID NO:102, SEQ ID NO:103, SEQ ID NO:104, SEQ ID NO:105, SEQ ID NO:106, SEQ ID NO:107, SEQ ID NO:108, SEQ ID NO:109, SEQ ID NO:110, SEQ ID NO:111, SEQ ID NO:112, SEQ ID NO:113, SEQ ID NO: The amino acid sequence may comprise an amino acid sequence selected from the group consisting of SEQ ID NO:114, SEQ ID NO:115, SEQ ID NO:116, SEQ ID NO:117, SEQ ID NO:118, SEQ ID NO:119, SEQ ID NO:120, SEQ ID NO:121, SEQ ID NO:122, SEQ ID NO:123, SEQ ID NO:124, SEQ ID NO:125, SEQ ID NO:126, SEQ ID NO:127, SEQ ID NO:128, SEQ ID NO:129, SEQ ID NO:130, SEQ ID NO:131, SEQ ID NO:132, SEQ ID NO:133, SEQ ID NO:134, SEQ ID NO:135, SEQ ID NO:171, SEQ ID NO:173.

[0596] For example, the fusion polypeptide can comprise an amino acid sequence selected from the group consisting of SEQ ID NO:104, SEQ ID NO:107, SEQ ID NO:108, SEQ ID NO:112, SEQ ID NO:114, SEQ ID NO:115, SEQ ID NO:116, SEQ ID NO:171 and SEQ ID NO:173.

[0597] In one embodiment, the present application discloses bi- and / or multi-functional fusion polypeptides, including two-chain fusion polypeptides. The fusion polypeptides comprise (i) an antigen-binding fragment that specifically binds to a first cellular target, and / or (ii) a first domain, with or without an Fc domain, and (iii) a second domain that specifically binds to a second cellular target, and / or (iv) a third domain that specifically binds to a third cellular target. The second domain that binds to the second cellular target is covalently linked via the polypeptide C-terminus of the first domain or the polypeptide N-terminus of the first domain, with or without a linker. The third domain that binds to the third cellular target is covalently linked via the polypeptide C-terminus of the first domain, or the C-terminus of the second domain, or the polypeptide N-terminus of the first domain, or the N-terminus of the second domain, with or without a linker. The first domains disclosed in the present invention may also be extended to anti-PD-L1 antibodies and / or anti-PD-1 antibodies that do not have an Fc domain, including, but not limited to, single-domain antibodies, recombinant antibodies, and single-chain antibodies. For example, the present invention is considered to be within the scope of the claims when the C-terminus of the heavy or light chain polypeptide of the first domain is covalently linked to the second and / or third domain via a linker. Similarly, the present invention is also considered to be within the scope of the claims when the N-terminus of the heavy or light chain polypeptide of the first domain is covalently linked to the second and / or third domain via a linker. Thus, the bi- and / or multi-functional fusion polypeptide is capable of simultaneously binding to a first cellular target, a second cellular target, and / or a third cellular target.

[0598] In one embodiment, the present invention provides a specific polypeptide complex comprising a bi- and / or multi-functional fusion polypeptide, which comprises a first polypeptide and a second polypeptide comprising the polypeptide complex, although it should be clear that in certain antibodies, the second polypeptide may be absent.

[0599] Meanwhile, the second polypeptide of the polypeptide complex of the present invention is the light chain of the first domain. The first polypeptide of the polypeptide complex of the present invention is composed of, from the N-terminus to the C-terminus of the polypeptide, the third domain and / or the second domain, a linker, the heavy chain of the first domain, a linker, and the third domain and / or the second domain, which are covalently bound together. The first and second polypeptides thus constitute a specific polypeptide complex of a bi- and / or multifunctional fusion polypeptide. Here, the linkers may be present independently or may not be present. Furthermore, under certain circumstances, only one type of second domain or one type of third domain may be present.

[0600] Meanwhile, the second polypeptide of the polypeptide complex of the present invention is the light chain of the first domain. The first polypeptide of the polypeptide complex of the present invention is composed of, from the N-terminus to the C-terminus of the polypeptide, the heavy chain of the first domain, a linker, a third domain and / or a second domain, a linker, and a third domain and / or a second domain, which are covalently bound together. Thus, the first and second polypeptides constitute a specific polypeptide complex of a bi- and / or multifunctional fusion polypeptide. Here, the linkers may be present independently or may not be present. Furthermore, under certain circumstances, only one type of second domain or one type of third domain may be present.

[0601] Meanwhile, the second polypeptide of the polypeptide complex of the present invention is the light chain of the first domain. The first polypeptide of the polypeptide complex of the present invention is composed of, in order from the N-terminus to the C-terminus of the polypeptide, the third domain and / or the second domain, a linker, the third domain and / or the second domain, a linker, and the heavy chain of the first domain, which are covalently bound to each other. Thus, the first and second polypeptides constitute a specific polypeptide complex of a bi- and / or multifunctional fusion polypeptide. Here, the linkers may be present independently or may not be present. Furthermore, under certain circumstances, only one type of second domain or one type of third domain may be present.

[0602] Meanwhile, the first polypeptide of the polypeptide complex of the present invention is the heavy chain of the first domain. The second polypeptide of the polypeptide complex of the present invention is composed of, in order from the N-terminus to the C-terminus of the polypeptide, the light chain of the first domain, a linker, the third domain and / or the second domain, a linker, and the third domain and / or the second domain, which are covalently bound to each other. Thus, the first and second polypeptides constitute a specific polypeptide complex of a bi- and / or multifunctional fusion polypeptide. Here, the linkers may be present independently or may not be present. Furthermore, under certain circumstances, only one type of second domain or one type of third domain may be present.

[0603] Meanwhile, the first polypeptide of the polypeptide complex of the present invention is the heavy chain of the first domain. The second polypeptide of the polypeptide complex of the present invention is composed of, in order from the N-terminus to the C-terminus of the polypeptide, the light chain of the first domain, a linker, the third domain and / or the second domain, a linker, and the third domain and / or the second domain, which are covalently bound to each other. Thus, the first and second polypeptides constitute a specific polypeptide complex of a bi- and / or multifunctional fusion polypeptide. Here, the linkers may be present independently or may not be present. Furthermore, under certain circumstances, only one type of second domain or one type of third domain may be present.

[0604] Meanwhile, the first polypeptide of the polypeptide complex of the present invention is the heavy chain of the first domain. The second polypeptide of the polypeptide complex of the present invention is composed of, in order from the N-terminus to the C-terminus of the polypeptide, the third domain and / or the second domain, a linker, the third domain and / or the second domain, a linker, and the light chain of the first domain, covalently bound to each other. Thus, the first and second polypeptides constitute a specific polypeptide complex of a bi- and / or multifunctional fusion polypeptide. Here, the linkers may be present independently or may not be present. Furthermore, under certain circumstances, only one type of second domain or one type of third domain may be present.

[0605] On the other hand, the first polypeptide of the polypeptide complex of the present invention is composed of, in order from the N-terminus to the C-terminus of the polypeptide, a third domain and / or a second domain, a linker, and a heavy chain of the first domain, which are covalently bonded. The second polypeptide of the polypeptide complex of the present invention is composed of, in order from the N-terminus to the C-terminus of the polypeptide, a light chain of the first domain, a linker, and a third domain and / or a second domain, which are covalently bonded. Here, the linkers may be present independently or may not be present. Furthermore, under certain circumstances, only one type of second domain or third domain may be present.

[0606] On the other hand, the first polypeptide of the polypeptide complex of the present invention is composed of, in order from the N-terminus to the C-terminus of the polypeptide, a third domain and / or a second domain, a linker, and a heavy chain of the first domain, which are covalently bonded. The second polypeptide of the polypeptide complex of the present invention is composed of, in order from the N-terminus to the C-terminus of the polypeptide, a third domain and / or a second domain, a linker, and a light chain of the first domain, which are covalently bonded. Here, the linkers may be present independently or may not be present. Furthermore, under certain circumstances, only one type of second domain or third domain may be present.

[0607] On the other hand, the first polypeptide of the polypeptide complex of the present invention is composed of the heavy chain of the first domain, a linker, a third domain, and / or a second domain, which are covalently bonded in order from the N-terminus to the C-terminus of the polypeptide. The second polypeptide of the polypeptide complex of the present invention is composed of the third domain and / or the second domain, a linker, and a light chain of the first domain, which are covalently bonded in order from the N-terminus to the C-terminus of the polypeptide. Here, the linkers may be present independently or may not be present. Furthermore, under certain circumstances, only one type of second domain or third domain may be present.

[0608] On the other hand, the first polypeptide of the polypeptide complex of the present invention is composed of the heavy chain of the first domain, a linker, a third domain, and / or a second domain, which are covalently bound in order from the N-terminus to the C-terminus of the polypeptide. The second polypeptide of the polypeptide complex of the present invention is composed of the light chain of the first domain, a linker, a third domain, and / or a second domain, which are covalently bound in order from the N-terminus to the C-terminus of the polypeptide. Here, the linkers may be present independently or may not be present. Furthermore, under certain circumstances, only one type of second domain or third domain may be present.

[0609] On the other hand, the polypeptide complexes of the present invention comprise only a first polypeptide structure, which is composed of, from the N-terminus to the C-terminus of the polypeptide, the heavy chain of the first domain, a linker, a third domain and / or a second domain, a linker, a third domain and / or a second domain, which are covalently linked to form a specific polypeptide complex of a bi- and / or multifunctional fusion polypeptide. Here, the linkers may be present independently or may be absent. Also, under certain circumstances, only one type of second domain or one type of third domain may be present.

[0610] On the other hand, the polypeptide complexes of the present invention comprise only a first polypeptide structure, which is composed of, from the N-terminus to the C-terminus of the polypeptide, the heavy chain of the first domain, a linker, a third domain and / or a second domain, a linker, a third domain and / or a second domain, which are covalently linked to form a specific polypeptide complex of a bi- and / or multifunctional fusion polypeptide. Here, the linkers may be present independently or may be absent. Also, under certain circumstances, only one type of second domain or one type of third domain may be present.

[0611] On the other hand, the polypeptide complexes of the present invention comprise only the structure of a first polypeptide, which is composed of, from the N-terminus to the C-terminus of the polypeptide, the covalently linked third and / or second domains, a linker, the third and / or second domains, a linker, and the heavy chain of the first domain, forming a specific polypeptide complex of a bi- and / or multifunctional fusion polypeptide. Here, the linkers may be present independently or may be absent. Also, under certain circumstances, only one type of second or third domain may be present.

[0612] The bi- and / or multi-functional fusion polypeptides of the present invention comprise (a) an antigen-binding fragment contained in the first domain, which may be derived from an anti-PD-L1 antibody and / or anti-PD-1 antibody, including antibodies that inhibit or reduce the binding of PD-L1 to its receptor, thereby relieving the immunosuppressive effects of PD-L1, including existing published anti-PD-L1 antibodies and / or anti-PD-1 antibodies, as well as anti-PD-L1 antibodies and / or anti-PD-1 antibodies developed in the future.

[0613] The bi- and / or multifunctional fusion polypeptides of the present invention include, as the second domain, the extracellular domain of one or more ligands. This includes at least a polypeptide containing the CD86 extracellular domain or the CD86 extracellular domain IgV region. For example, CD86 is a protein belonging to the B7 molecule family and is involved in the costimulatory activation of T cells. The mature CD86 molecule is composed of an extracellular domain (ECD), a transmembrane domain, and an intracellular domain. The CD86 extracellular domain (ECD) is the main region that binds to its corresponding receptor on T cells and contains an immunoglobulin-like V (IgV) region and an immunoglobulin-like C2 (IgC2) region. The IgV domain is the main structural region directly involved in receptor binding, and the IgV region of the CD86 extracellular domain binds to CD28 and is involved in T cell activation, proliferation, and induction of effector function. CTLA4 also binds to the IgV region of CD86 and is involved in immunosuppressive regulation.

[0614] The bi- and / or multifunctional fusion polypeptide of the present invention includes, as a third domain, the extracellular domain of one or more types of ligands. This includes the LAG3 extracellular domain or one or more subunits of a polypeptide comprised of IgD1, IgD2, IgD3, and IgD4 contained in the LAG3 extracellular domain. For example, LAG3 (Lymphocyte Activation Gene-3) is a type I transmembrane protein involved in immune checkpoints and is primarily expressed in activated NK cells and T cells. LAG3 activates tumor-infiltrating, exhausted CD8 + LAG3 is expressed in T cells and is a promising target for tumor immunotherapy. Its function is dual: on the one hand, it acts as a negative regulator of T cell proliferation and activation, suppressing T cell activation and function by binding to its ligands, major histocompatibility complex II (MHCII) or fibrinogen-like protein 1 (FGL1). On the other hand, LAG3 supports T cell activation by inducing antigen-presenting cell activation through binding to MHCII molecules on antigen-presenting cells, upregulating CD80 / CD83 / CD86 expression, enhancing antigen-presenting capacity, and promoting T cell recruitment via chemokine secretion. Antibodies that block LAG3 have the effect of relieving LAG3-mediated T cell suppression, but they also inhibit LAG3-mediated antigen-presenting cell activation, preventing full enhancement of LAG3-related immune activation. On the other hand, the LAG3 extracellular domain chimeric protein provided by the present invention can simultaneously address both T cell immune suppression and antigen-presenting cell activation, potentially demonstrating its potential for tumor therapy.

[0615] Provided below is an exemplary anti-PD-L1 antibody, sugemalimab, linked via an Fc domain to a linker peptide and covalently linked to a CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 36) of a bifunctional fusion polypeptide complex, wherein the first polypeptide and a sugemalimab light chain second polypeptide (SEQ ID NO: 5) constitute the bifunctional antibody polypeptide complex. Sugemalimab is linked via an Fc domain to a linker peptide and covalently linked to a CD86 extracellular domain IgV region to form the first polypeptide amino acid sequence (SEQ ID NO: 37) of a bifunctional fusion polypeptide complex, wherein the first polypeptide and a sugemalimab light chain second polypeptide constitute the bifunctional antibody polypeptide complex. Sugemalimab is linked via the N-terminus of the heavy chain of the antigen-binding site to a linker peptide and covalently linked to a CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 38) of a bifunctional fusion polypeptide complex, wherein the first polypeptide and a sugemalimab light chain second polypeptide constitute the bifunctional fusion polypeptide complex. Sugemalimab is bound to a linker peptide via the N-terminus of its heavy chain in the antigen-binding site and covalently bound to the IgV region of the CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 39) of the bifunctional fusion polypeptide complex, and the first polypeptide and the sugemalimab light chain second polypeptide form the bifunctional fusion polypeptide complex. Sugemalimab is bound to a linker peptide via the C-terminus of its light chain in the antigen-binding site and covalently bound to the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO: 40) of the bifunctional fusion polypeptide complex, and the second polypeptide and the sugemalimab heavy chain first polypeptide (SEQ ID NO: 4) form the bifunctional fusion polypeptide complex. Sugemalimab is bound to a linker peptide via the C-terminus of its light chain in the antigen-binding site and covalently bound to the IgV region of the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO: 41) of the bifunctional fusion polypeptide complex, and the second polypeptide and the sugemalimab heavy chain first polypeptide form the bifunctional fusion polypeptide complex.Sugemalimab is bound to a linker peptide via the N-terminus of the light chain of the antigen-binding site and is further covalently bound to the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO: 42) of the bifunctional fusion polypeptide complex, and the second polypeptide and the sugemalimab heavy chain first polypeptide form the bifunctional fusion polypeptide complex. Sugemalimab is bound to a linker peptide via the N-terminus of the light chain of the antigen-binding site and is further covalently bound to the CD86 extracellular domain IgV region to form the second polypeptide amino acid sequence (SEQ ID NO: 43) of the bifunctional fusion polypeptide complex, and the second polypeptide and the sugemalimab heavy chain first polypeptide form the bifunctional fusion polypeptide complex.

[0616] Provided below is an exemplary anti-PD-L1 antibody, Durvalumab, linked to a linker peptide via an Fc domain and covalently linked to a CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 44) of a bifunctional fusion polypeptide complex, where this first polypeptide, together with a Durvalumab light chain second polypeptide (SEQ ID NO: 9), constitutes the bifunctional fusion polypeptide complex. Durvalumab is linked to a linker peptide via an Fc domain and covalently linked to a CD86 extracellular domain IgV region to form the first polypeptide amino acid sequence (SEQ ID NO: 45) of a bifunctional fusion polypeptide complex, where the first polypeptide and the Durvalumab light chain second polypeptide constitute the bifunctional antibody polypeptide complex. Durvalumab is linked to a linker peptide via the N-terminus of the heavy chain in the antigen-binding site and covalently linked to a CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 46) of a bifunctional fusion polypeptide complex, where the first polypeptide and the Durvalumab light chain second polypeptide constitute the bifunctional fusion polypeptide complex. Durvalumab is bound to a linker peptide via the N-terminus of the antigen-binding site of its heavy chain and covalently linked to the IgV region of the CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO:47) of the bifunctional fusion polypeptide complex, and the first polypeptide and the durvalumab light chain second polypeptide form the bifunctional fusion polypeptide complex. Durvalumab is bound to a linker peptide via the C-terminus of its antigen-binding site of its light chain and covalently linked to the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO:48) of the bifunctional fusion polypeptide complex, and the second polypeptide and the durvalumab heavy chain first polypeptide (SEQ ID NO:8) form the bifunctional fusion polypeptide complex. Durvalumab is bound to a linker peptide via the C-terminus of its antigen-binding site of its light chain and covalently linked to the IgV region of the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO:49) of the bifunctional fusion polypeptide complex, and the second polypeptide and the durvalumab heavy chain first polypeptide form the bifunctional fusion polypeptide complex.Durvalumab is bound to a linker peptide via the N-terminus of the antigen-binding light chain and covalently linked to the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO: 50) of the bifunctional fusion polypeptide complex, and the second polypeptide and the durvalumab heavy chain first polypeptide form the bifunctional fusion polypeptide complex. Durvalumab is bound to a linker peptide via the N-terminus of the antigen-binding light chain and covalently linked to the CD86 extracellular domain IgV region to form the second polypeptide amino acid sequence (SEQ ID NO: 51) of the bifunctional fusion polypeptide complex, and the second polypeptide and the durvalumab heavy chain first polypeptide form the bifunctional fusion polypeptide complex.

[0617] Provided below is an exemplary anti-PD-L1 antibody, atezolizumab, linked via an Fc domain to a linker peptide and covalently linked to a CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 52) of a bifunctional fusion polypeptide complex, where the first polypeptide and the atezolizumab light chain second polypeptide (SEQ ID NO: 11) comprise the bifunctional fusion polypeptide complex. Atezolizumab is linked via an Fc domain to a linker peptide and covalently linked to a CD86 extracellular domain IgV region to form the first polypeptide amino acid sequence (SEQ ID NO: 53) of a bifunctional fusion polypeptide complex, where the first polypeptide and the atezolizumab light chain second polypeptide comprise the bifunctional antibody polypeptide complex. Atezolizumab is linked via the N-terminus of the heavy chain in the antigen-binding site to a linker peptide and covalently linked to a CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 54) of a bifunctional fusion polypeptide complex, where the first polypeptide and the atezolizumab light chain second polypeptide comprise the bifunctional fusion polypeptide complex. Atezolizumab is bound to a linker peptide via the N-terminus of the antigen-binding site of its heavy chain and covalently bound to the IgV region of the CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 55) of the bifunctional fusion polypeptide complex, and the first polypeptide and the atezolizumab light chain second polypeptide form the bifunctional fusion polypeptide complex. Atezolizumab is bound to a linker peptide via the C-terminus of its antigen-binding site of its light chain and covalently bound to the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO: 56) of the bifunctional fusion polypeptide complex, and the second polypeptide and the atezolizumab heavy chain first polypeptide (SEQ ID NO: 10) form the bifunctional fusion polypeptide complex. Atezolizumab is bound to a linker peptide via the C-terminus of its antigen-binding site of its light chain and covalently bound to the IgV region of the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO: 57) of the bifunctional fusion polypeptide complex, and the second polypeptide and the atezolizumab heavy chain first polypeptide form the bifunctional fusion polypeptide complex.The second polypeptide and the atezolizumab heavy chain first polypeptide form a bifunctional antibody polypeptide complex. Atezolizumab is bound to a linker peptide via the N-terminus of the light chain of the antigen-binding site and is further covalently bound to a CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO: 58) of the bifunctional fusion polypeptide complex, and the second polypeptide and the atezolizumab heavy chain first polypeptide form the bifunctional fusion polypeptide complex. Atezolizumab is bound to a linker peptide via the N-terminus of the light chain of the antigen-binding site and is further covalently bound to a CD86 extracellular domain IgV region to form the second polypeptide amino acid sequence (SEQ ID NO: 59) of the bifunctional fusion polypeptide complex, and the second polypeptide and the atezolizumab heavy chain first polypeptide form the bifunctional fusion polypeptide complex.

[0618] Provided below is an exemplary anti-PD-L1 antibody, avelumab, linked via an Fc domain to a linker peptide and covalently linked to a CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 60) of a bifunctional fusion polypeptide complex, where the first polypeptide and the avelumab light chain second polypeptide (SEQ ID NO: 13) comprise the bifunctional fusion polypeptide complex. Avelumab is linked via an Fc domain to a linker peptide and covalently linked to a CD86 extracellular domain IgV region to form the first polypeptide amino acid sequence (SEQ ID NO: 61) of a bifunctional fusion polypeptide complex, where the first polypeptide and the avelumab light chain second polypeptide comprise the bifunctional antibody polypeptide complex. Avelumab is linked via the N-terminus of the heavy chain in the antigen-binding site to a linker peptide and covalently linked to a CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 62) of a bifunctional fusion polypeptide complex, where the first polypeptide and the avelumab light chain second polypeptide comprise the bifunctional fusion polypeptide complex. Avelumab is bound to a linker peptide via the N-terminus of the antigen-binding site of its heavy chain and covalently linked to the IgV region of the CD86 extracellular domain to form the first polypeptide amino acid sequence of the bifunctional fusion polypeptide complex (SEQ ID NO: 63), and the first polypeptide and the avelumab light chain second polypeptide constitute the bifunctional fusion polypeptide complex. Avelumab is bound to a linker peptide via the C-terminus of its antigen-binding site of its light chain and covalently linked to the CD86 extracellular domain to form the second polypeptide amino acid sequence of the bifunctional fusion polypeptide complex (SEQ ID NO: 64), and the second polypeptide and the avelumab heavy chain first polypeptide (SEQ ID NO: 12) constitute the bifunctional fusion polypeptide complex. Avelumab is bound to a linker peptide via the C-terminus of its antigen-binding site of its light chain and covalently linked to the IgV region of the CD86 extracellular domain to form the second polypeptide amino acid sequence of the bifunctional fusion polypeptide complex (SEQ ID NO: 65), and the second polypeptide and the avelumab heavy chain first polypeptide constitute the bifunctional fusion polypeptide complex. The second polypeptide and the avelumab heavy chain first polypeptide constitute the bifunctional antibody polypeptide complex.Avelumab is bound to a linker peptide via the N-terminus of the antigen-binding light chain and covalently linked to the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO: 66) of the bifunctional fusion polypeptide complex, and the second polypeptide and the avelumab heavy chain first polypeptide form the bifunctional fusion polypeptide complex. Avelumab is bound to a linker peptide via the N-terminus of the antigen-binding light chain and covalently linked to the CD86 extracellular domain IgV region to form the second polypeptide amino acid sequence (SEQ ID NO: 67) of the bifunctional fusion polypeptide complex, and the second polypeptide and the avelumab heavy chain first polypeptide form the bifunctional fusion polypeptide complex.

[0619] Provided below is an exemplary anti-PD-L1 antibody, envafolimab, linked via an Fc domain to a linker peptide and covalently linked to a CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 68) of a bifunctional fusion polypeptide complex, the first polypeptide comprising the bifunctional fusion polypeptide complex. Envafolimab is linked via an Fc domain to a linker peptide and covalently linked to a CD86 extracellular domain IgV region to form the first polypeptide amino acid sequence (SEQ ID NO: 69) of the bifunctional fusion polypeptide complex, the first polypeptide comprising the bifunctional antibody polypeptide complex. Envafolimab is linked via the N-terminus of the heavy chain of the antigen-binding site to a linker peptide and covalently linked to a CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 70) of the bifunctional fusion polypeptide complex. Embafolimab is bound to a linker peptide via the N-terminus of the heavy chain of the antigen-binding site and is further covalently bound to the IgV region of the CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 71) of the bifunctional fusion polypeptide complex, and the first polypeptide constitutes the bifunctional fusion polypeptide complex.

[0620] Provided below is an exemplary anti-PD-1 antibody, pembrolizumab, linked via an Fc domain to a linker peptide and covalently linked to a CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 72) of the bifunctional fusion polypeptide complex, where the first polypeptide and a pembrolizumab light chain second polypeptide (SEQ ID NO: 16) constitute the bifunctional fusion polypeptide complex. Pembrolizumab is linked via an Fc domain to a linker peptide and covalently linked to a CD86 extracellular domain IgV region to form the first polypeptide amino acid sequence (SEQ ID NO: 73) of the bifunctional fusion polypeptide complex, where the first polypeptide and a pembrolizumab light chain second polypeptide constitute the bifunctional antibody polypeptide complex. Pembrolizumab is linked via the heavy chain N-terminus of the antigen-binding site to a linker peptide and covalently linked to a CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 74) of the bifunctional fusion polypeptide complex, where the first polypeptide and a pembrolizumab light chain second polypeptide constitute the bifunctional fusion polypeptide complex. Pembrolizumab is bound to a linker peptide via the N-terminus of the heavy chain of the antigen-binding site and is further covalently bound to the IgV region of the CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 75) of the bifunctional fusion polypeptide complex, and the first polypeptide and the pembrolizumab light chain second polypeptide constitute the bifunctional fusion polypeptide complex. Pembrolizumab is bound to a linker peptide via the C-terminus of the light chain of the antigen-binding site and is further covalently bound to the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO: 76) of the bifunctional fusion polypeptide complex, and the second polypeptide and the pembrolizumab heavy chain first polypeptide (SEQ ID NO: 15) constitute the bifunctional fusion polypeptide complex.Pembrolizumab is bound to a linker peptide via the C-terminus of the light chain in the antigen-binding site and covalently bound to the IgV region of the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO: 77) of the bifunctional fusion polypeptide complex, and the second polypeptide and the pembrolizumab heavy chain first polypeptide form the bifunctional antibody polypeptide complex. Pembrolizumab is bound to a linker peptide via the N-terminus of the light chain in the antigen-binding site and covalently bound to the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO: 78) of the bifunctional fusion polypeptide complex, and the second polypeptide and the pembrolizumab heavy chain first polypeptide form the bifunctional fusion polypeptide complex. Pembrolizumab is bound to a linker peptide via the N-terminus of the light chain in the antigen-binding site and covalently bound to the IgV region of the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO: 79) of the bifunctional fusion polypeptide complex, and the second polypeptide and the pembrolizumab heavy chain first polypeptide form the bifunctional fusion polypeptide complex.

[0621] Provided below is an exemplary anti-PD-1 antibody, nivolumab, linked via an Fc domain to a linker peptide and covalently linked to a CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 80) of the bifunctional fusion polypeptide complex, wherein the first polypeptide and a nivolumab light chain second polypeptide (SEQ ID NO: 18) constitute the bifunctional fusion polypeptide complex. Nivolumab is linked via an Fc domain to a linker peptide and covalently linked to a CD86 extracellular domain IgV region to form the first polypeptide amino acid sequence (SEQ ID NO: 81) of the bifunctional fusion polypeptide complex, wherein the first polypeptide and a nivolumab light chain second polypeptide constitute the bifunctional antibody polypeptide complex. Nivolumab is linked via the heavy chain N-terminus of the antigen-binding site to a linker peptide and covalently linked to a CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 82) of the bifunctional fusion polypeptide complex, wherein the first polypeptide and a nivolumab light chain second polypeptide constitute the bifunctional fusion polypeptide complex. Nivolumab is bound to a linker peptide via the N-terminus of its heavy chain in the antigen-binding site and covalently bound to the IgV region of the CD86 extracellular domain to form the first polypeptide amino acid sequence (SEQ ID NO: 83) of the bifunctional fusion polypeptide complex, and the first polypeptide and the nivolumab light chain second polypeptide constitute the bifunctional fusion polypeptide complex. Nivolumab is bound to a linker peptide via the C-terminus of its light chain in the antigen-binding site and covalently bound to the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO: 84) of the bifunctional fusion polypeptide complex, and the second polypeptide and the nivolumab heavy chain first polypeptide (SEQ ID NO: 17) constitute the bifunctional fusion polypeptide complex. Nivolumab is bound to a linker peptide via the C-terminus of its light chain in the antigen-binding site and covalently bound to the IgV region of the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO: 85) of the bifunctional fusion polypeptide complex, and the second polypeptide and the nivolumab heavy chain first polypeptide constitute the bifunctional antibody polypeptide complex.Nivolumab is bound to a linker peptide via the N-terminus of the light chain of the antigen-binding site and is further covalently bound to the CD86 extracellular domain to form the second polypeptide amino acid sequence (SEQ ID NO: 86) of the bifunctional fusion polypeptide complex, and the second polypeptide and the nivolumab heavy chain first polypeptide constitute the bifunctional fusion polypeptide complex. Nivolumab is bound to a linker peptide via the N-terminus of the light chain of the antigen-binding site and is further covalently bound to the CD86 extracellular domain IgV region to form the second polypeptide amino acid sequence (SEQ ID NO: 87) of the bifunctional fusion polypeptide complex, and the second polypeptide and the nivolumab heavy chain first polypeptide constitute the bifunctional fusion polypeptide complex.

[0622] Provided below is an exemplary anti-PD-L1 antibody, Sugimalimab, in which the heavy chain is covalently bound to a CD86 extracellular domain via a linker peptide through the C-terminus of the Fc domain, and the CD86 extracellular domain is covalently bound to a linker peptide through its C-terminus and covalently bound to a LAG3 extracellular domain, forming a first polypeptide amino acid sequence (SEQ ID NO: 88) of a multifunctional fusion polypeptide complex, and the first polypeptide and the Sugimalimab light chain second polypeptide together constitute the multifunctional fusion polypeptide complex. The Sugimalimab heavy chain is covalently bound to a CD86 extracellular domain via a linker peptide through the C-terminus of the Fc domain, and the CD86 extracellular domain is covalently bound to a linker peptide through its C-terminus and covalently bound to a LAG3 extracellular domain D1 / D2 / D3, forming a first polypeptide amino acid sequence (SEQ ID NO: 89) of a multifunctional fusion polypeptide complex, and the first polypeptide and the Sugimalimab light chain second polypeptide together constitute the multifunctional fusion polypeptide complex. The Sugimalimab heavy chain is covalently bound to the CD86 extracellular domain via a linker peptide at the C-terminus of the Fc domain, the CD86 extracellular domain is covalently bound to the linker peptide at the C-terminus and further covalently bound to the LAG3 extracellular domain D1 / D2, forming the first polypeptide amino acid sequence (SEQ ID NO: 90) of the multifunctional fusion polypeptide complex, and the first polypeptide and the Sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The Sugimalimab heavy chain is covalently bound to the CD86 extracellular domain via a linker peptide at the C-terminus of the Fc domain, and further covalently bound to the CD86 extracellular domain, the CD86 extracellular domain is covalently bound to the linker peptide at the C-terminus and further covalently bound to the LAG3 extracellular domain D1, forming the first polypeptide amino acid sequence (SEQ ID NO: 91) of the multifunctional fusion polypeptide complex, and the first polypeptide and the Sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex.The Sugimalimab heavy chain is bound to a linker peptide via the C-terminus of the Fc domain and is covalently bound to a LAG3 extracellular domain, which is covalently bound to the linker peptide via its C-terminus and covalently bound to a CD86 extracellular domain, forming a first polypeptide amino acid sequence (SEQ ID NO: 92) of a multifunctional fusion polypeptide complex, and the first polypeptide and the Sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The Sugimalimab heavy chain is bound to a linker peptide via the C-terminus of the Fc domain and is covalently bound to a LAG3 extracellular domain D1 / D2 / D3, which is covalently bound to a linker peptide via its C-terminus and covalently bound to a CD86 extracellular domain, forming a first polypeptide amino acid sequence (SEQ ID NO: 93) of a multifunctional fusion polypeptide complex, and the first polypeptide and the Sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The first polypeptide and the Sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The Sugimalimab heavy chain is bound to a linker peptide via the C-terminus of the Fc domain and is covalently bound to the LAG3 extracellular domains D1 / D2, the LAG3 extracellular domains D1 / D2 are covalently bound to the linker peptide via their C-terminus and are covalently bound to the CD86 extracellular domain, forming the first polypeptide amino acid sequence (SEQ ID NO: 94) of a multifunctional fusion polypeptide complex, and the first polypeptide and the Sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The Sugimalimab heavy chain is bound to a linker peptide via the C-terminus of the Fc domain and is covalently bound to the LAG3 extracellular domain D1, the LAG3 extracellular domain D1 is covalently bound to the linker peptide via their C-terminus and are covalently bound to the CD86 extracellular domain, forming the first polypeptide amino acid sequence (SEQ ID NO: 95) of a multifunctional fusion polypeptide complex, and the first polypeptide and the Sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex.

[0623] Provided below is an exemplary anti-PD-L1 antibody, Sugimalimab, heavy chain, which is linked to a linker peptide via the C-terminus of the Fc domain, and covalently linked to the IgV region of the CD86 extracellular domain, which is covalently linked to the linker peptide via its C-terminus and to a LAG3 extracellular domain, forming a first polypeptide amino acid sequence (SEQ ID NO: 96) of a multifunctional fusion polypeptide complex, and the first polypeptide and Sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The Sugimalimab heavy chain, which is linked to a linker peptide via the C-terminus of the Fc domain, and covalently linked to the IgV region of the CD86 extracellular domain, which is covalently linked to the linker peptide via its C-terminus and to a LAG3 extracellular domain D1 / D2 / D3, forms a first polypeptide amino acid sequence (SEQ ID NO: 97) of a multifunctional fusion polypeptide complex, and the first polypeptide and Sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The Sugimalimab heavy chain is bound to a linker peptide via the C-terminus of the Fc domain, and is covalently bound to the IgV region of the CD86 extracellular domain, which is covalently bound to the linker peptide via its C-terminus and further covalently bound to the LAG3 extracellular domain D1 / D2, forming the first polypeptide amino acid sequence (SEQ ID NO: 98) of a multifunctional fusion polypeptide complex, and the first polypeptide and the Sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The Sugimalimab heavy chain is bound to a linker peptide via the C-terminus of the Fc domain, and is covalently bound to the IgV region of the CD86 extracellular domain, which is covalently bound to the linker peptide via its C-terminus and further covalently bound to the LAG3 extracellular domain D1, forming the first polypeptide amino acid sequence (SEQ ID NO: 99) of a multifunctional fusion polypeptide complex, and the first polypeptide and the Sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex.The Sugimalimab heavy chain is bound to a linker peptide via the C-terminus of the Fc domain and is covalently bound to a LAG3 extracellular domain, the LAG3 extracellular domain is covalently bound to the linker peptide via its C-terminus and is covalently bound to the IgV region of the CD86 extracellular domain, forming a first polypeptide amino acid sequence (SEQ ID NO: 100) of a multifunctional fusion polypeptide complex, and the first polypeptide and the Sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The Sugimalimab heavy chain is bound to a linker peptide via the C-terminus of the Fc domain and is covalently bound to a LAG3 extracellular domain D1 / D2 / D3, the LAG3 extracellular domain D1 / D2 / D3 is covalently bound to the linker peptide via its C-terminus and is covalently bound to the IgV region of the CD86 extracellular domain, forming a first polypeptide amino acid sequence (SEQ ID NO: 101) of a multifunctional fusion polypeptide complex, and the first polypeptide and the Sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The Sugimalimab heavy chain is bound to a linker peptide via the C-terminus of the Fc domain, and is covalently bound to the LAG3 extracellular domain D1 / D2, which is covalently bound to the linker peptide via its C-terminus and further covalently bound to the IgV region of the CD86 extracellular domain, forming a first polypeptide amino acid sequence (SEQ ID NO: 102) of a multifunctional fusion polypeptide complex, and the first polypeptide and the Sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The Sugimalimab heavy chain is bound to a linker peptide via the C-terminus of the Fc domain, and is covalently bound to the LAG3 extracellular domain D1, which is covalently bound to the linker peptide via its C-terminus and further covalently bound to the IgV region of the CD86 extracellular domain, forming a first polypeptide amino acid sequence (SEQ ID NO: 103) of a multifunctional fusion polypeptide complex, and the first polypeptide and the Sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex.

[0624] Provided below is an exemplary multifunctional fusion polypeptide complex in which the C-terminus of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, and the C-terminus of sugimalimab is covalently linked to the LAG3 extracellular domain via a linker peptide, forming a first polypeptide amino acid sequence (SEQ ID NO: 104) of the multifunctional fusion polypeptide complex, and the first polypeptide and the sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The C-terminus of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, and the C-terminus of the sugimalimab heavy chain is covalently linked to the LAG3 extracellular domain D1 / D2 / D3 via a linker peptide, forming a first polypeptide amino acid sequence (SEQ ID NO: 105) of the multifunctional fusion polypeptide complex, and the first polypeptide and the sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The C-terminus of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, and the C-terminus of the heavy chain of sugimalimab is covalently linked to the LAG3 extracellular domain D1 / D2 via a linker peptide to form a first polypeptide amino acid sequence (SEQ ID NO: 106) of a multifunctional fusion polypeptide complex, and the first polypeptide and the second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, and the C-terminus of the heavy chain of sugimalimab is covalently linked to the LAG3 extracellular domain D1 via a linker peptide to form a first polypeptide amino acid sequence (SEQ ID NO: 107) of a multifunctional fusion polypeptide complex, and the first polypeptide and the second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the LAG3 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, and the C-terminus of sugimalimab is covalently linked to the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex (SEQ ID NO: 108), and the first polypeptide and a second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the LAG3 extracellular domain D1 / D2 / D3 is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, and the C-terminus of sugimalimab is covalently linked to the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex (SEQ ID NO: 109), and the first polypeptide and a second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the LAG3 extracellular domain D1 / D2 is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, and the C-terminus of sugimalimab is covalently linked to the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence (SEQ ID NO: 110) of a multifunctional fusion polypeptide complex, and the first polypeptide and a second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the LAG3 extracellular domain D1 is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, and the C-terminus of sugimalimab is covalently linked to the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence (SEQ ID NO: 111) of a multifunctional fusion polypeptide complex, and the first polypeptide and a second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex.

[0625] Provided below is an exemplary multifunctional fusion polypeptide complex in which the C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, and the C-terminus of sugimalimab is covalently linked to the LAG3 extracellular domain via a linker peptide, forming a first polypeptide amino acid sequence (SEQ ID NO: 112) of the multifunctional fusion polypeptide complex, and the first polypeptide and the second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, and the C-terminus of the sugimalimab heavy chain is covalently linked to the LAG3 extracellular domain D1 / D2 / D3 via a linker peptide, forming a first polypeptide amino acid sequence (SEQ ID NO: 113) of the multifunctional fusion polypeptide complex, and the first polypeptide and the second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, and the C-terminus of the heavy chain of sugimalimab is covalently linked to the LAG3 extracellular domain D1 / D2 via a linker peptide to form a first polypeptide amino acid sequence (SEQ ID NO: 114) of a multifunctional fusion polypeptide complex, and the first polypeptide and the second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, and the C-terminus of the heavy chain of sugimalimab is covalently linked to the LAG3 extracellular domain D1 via a linker peptide to form a first polypeptide amino acid sequence (SEQ ID NO: 115) of a multifunctional fusion polypeptide complex, and the first polypeptide and the second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the LAG3 extracellular domain is covalently linked to the anti-PD-L1 antibody sugimalimab heavy chain via a linker peptide, and the C-terminus of sugimalimab is covalently linked to the IgV region of the CD86 extracellular domain via a linker peptide to form the first polypeptide amino acid sequence (SEQ ID NO: 116) of a multifunctional fusion polypeptide complex, and the first polypeptide and the sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex.The C-terminus of the LAG3 extracellular domain D1 / D2 / D3 is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, and the C-terminus of sugimalimab is covalently linked to the IgV region of the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex (SEQ ID NO: 117), and the first polypeptide and a second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the LAG3 extracellular domain D1 / D2 is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, and the C-terminus of sugimalimab is covalently linked to the IgV region of the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex (SEQ ID NO: 118), and the first polypeptide and a second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the LAG3 extracellular domain D1 is covalently linked to the anti-PD-L1 antibody sugimalimab heavy chain via a linker peptide, and the C-terminus of sugimalimab is covalently linked to the IgV region of the CD86 extracellular domain via a linker peptide to form the first polypeptide amino acid sequence (SEQ ID NO: 119) of a multifunctional fusion polypeptide complex, and the first polypeptide and the sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex.

[0626] Provided below is an exemplary multifunctional fusion polypeptide complex in which the C-terminus of the CD86 extracellular domain is covalently linked to the LAG3 extracellular domain D1 / D2 / D3 via a linker peptide, and the C-terminus of the LAG3 extracellular domain D1 / D2 / D3 is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence (SEQ ID NO: 121) of the multifunctional fusion polypeptide complex, and the first polypeptide and the sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The C-terminus of the CD86 extracellular domain is covalently linked to the LAG3 extracellular domain D1 / D2 via a linker peptide, and the C-terminus of the LAG3 extracellular domain D1 / D2 is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence (SEQ ID NO: 122) of the multifunctional fusion polypeptide complex, and the first polypeptide and the sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The C-terminus of the CD86 extracellular domain is covalently linked to the LAG3 extracellular domain D1 via a linker peptide, and the C-terminus of the LAG3 extracellular domain D1 is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence (SEQ ID NO: 123) of a multifunctional fusion polypeptide complex, and the first polypeptide and the sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex. The C-terminus of the LAG3 extracellular domain is covalently linked to the CD86 extracellular domain via a linker peptide, and the C-terminus of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence (SEQ ID NO: 124) of a multifunctional fusion polypeptide complex, and the first polypeptide and the sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex.The C-terminus of the LAG3 extracellular domain D1 / D2 / D3 is covalently linked to the CD86 extracellular domain via a linker peptide, and the C-terminus of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex (SEQ ID NO: 125), and the first polypeptide and a second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the LAG3 extracellular domain D1 / D2 is covalently linked to the CD86 extracellular domain via a linker peptide, and the C-terminus of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex (SEQ ID NO: 126), and the first polypeptide and a second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the LAG3 extracellular domain D1 is covalently linked to the CD86 extracellular domain via a linker peptide, and the C-terminus of the CD86 extracellular domain is covalently linked to the anti-PD-L1 antibody sugimalimab heavy chain via a linker peptide to form the first polypeptide amino acid sequence (SEQ ID NO: 127) of a multifunctional fusion polypeptide complex, and the first polypeptide and the sugimalimab light chain second polypeptide constitute the multifunctional fusion polypeptide complex.

[0627] Provided below is an exemplary multifunctional fusion polypeptide complex in which the C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the LAG3 extracellular domain via a linker peptide, and the C-terminus of the LAG3 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence (SEQ ID NO: 128) of the multifunctional fusion polypeptide complex, and the first polypeptide and the second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the LAG3 extracellular domain D1 / D2 / D3 via a linker peptide, and the C-terminus of the LAG3 extracellular domain D1 / D2 / D3 is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence (SEQ ID NO: 129) of the multifunctional fusion polypeptide complex, and the first polypeptide and the second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the LAG3 extracellular domain D1 / D2 via a linker peptide, and the C-terminus of the LAG3 extracellular domain D1 / D2 is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex (SEQ ID NO: 130), and the first polypeptide and a second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the LAG3 extracellular domain D1 via a linker peptide, and the C-terminus of the LAG3 extracellular domain D1 is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex (SEQ ID NO: 131), and the first polypeptide and a second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex.The C-terminus of the LAG3 extracellular domain is covalently linked to the IgV region of the CD86 extracellular domain via a linker peptide, and the C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form the first polypeptide amino acid sequence of the multifunctional fusion polypeptide complex (SEQ ID NO: 132), and the first polypeptide and the second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the LAG3 extracellular domain D1 / D2 / D3 is covalently linked to the IgV region of the CD86 extracellular domain via a linker peptide, and the C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form the first polypeptide amino acid sequence of the multifunctional fusion polypeptide complex (SEQ ID NO: 133), and the first polypeptide and the second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the LAG3 extracellular domain D1 / D2 is covalently linked to the IgV region of the CD86 extracellular domain via a linker peptide, and the C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex (SEQ ID NO: 134), and the first polypeptide and a second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex. The C-terminus of the LAG3 extracellular domain D1 is covalently linked to the IgV region of the CD86 extracellular domain via a linker peptide, and the C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex (SEQ ID NO: 135), and the first polypeptide and a second polypeptide of the sugimalimab light chain constitute the multifunctional fusion polypeptide complex.

[0628] Provided below is an exemplary multifunctional fusion polypeptide complex in which the C-terminus of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form the first polypeptide amino acid sequence (SEQ ID NO: 38) of the multifunctional fusion polypeptide complex, and the C-terminus of the first polypeptide and the LAG3 extracellular domain are covalently linked to the second light chain polypeptide of sugimalimab via a linker peptide (SEQ ID NO: 136) of the multifunctional fusion polypeptide complex. The C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form the first polypeptide amino acid sequence (SEQ ID NO: 39) of the multifunctional fusion polypeptide complex, and the C-terminus of the first polypeptide and the LAG3 extracellular domain are covalently linked to the second light chain polypeptide of sugimalimab via a linker peptide to form the multifunctional fusion polypeptide complex. The C-terminus of the PD-L1 antibody sugimalimab heavy chain is covalently linked to the CD86 extracellular domain via a linker peptide to form the first polypeptide amino acid sequence (SEQ ID NO: 40) of the multifunctional fusion polypeptide complex, and the C-terminus of the first polypeptide and the LAG3 extracellular domain are covalently linked to the sugimalimab light chain second polypeptide via a linker peptide to form the multifunctional fusion polypeptide complex. The C-terminus of the PD-L1 antibody sugimalimab heavy chain is covalently linked to the IgV region of the CD86 extracellular domain via a linker peptide to form the first polypeptide amino acid sequence (SEQ ID NO: 41) of the multifunctional fusion polypeptide complex, and the C-terminus of the first polypeptide and the LAG3 extracellular domain are covalently linked to the sugimalimab light chain second polypeptide via a linker peptide to form the multifunctional fusion polypeptide complex.

[0629] Provided below are exemplary multifunctional fusion polypeptide complexes in which the C-terminus of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence, and the C-terminus of the first polypeptide and the LAG3 extracellular domain D1 / D2 / D3 are covalently linked to a second polypeptide of the light chain of sugimalimab (SEQ ID NO: 137) via a linker peptide to form the multifunctional fusion polypeptide complex. The C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence of the multifunctional fusion polypeptide complex, and the first polypeptide forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of the LAG3 extracellular domain D1 / D2 / D3 to the light chain of sugimalimab via a linker peptide. The C-terminus of the heavy chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex, and this first polypeptide forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of the LAG3 extracellular domain D1 / D2 / D3 to the light chain of sugimalimab via a linker peptide. The C-terminus of the heavy chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the IgV region of the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex, and this first polypeptide forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of the LAG3 extracellular domain D1 / D2 / D3 to the light chain of sugimalimab via a linker peptide.

[0630] Provided below are exemplary multifunctional fusion polypeptide complexes in which the C-terminus of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence, which forms a multifunctional fusion polypeptide complex with a second polypeptide in which the C-terminus of the LAG3 extracellular domain D1 / D2 is covalently linked to the light chain of sugimalimab (SEQ ID NO: 138) via a linker peptide. The C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a first polypeptide amino acid sequence, which forms a multifunctional fusion polypeptide complex with a second polypeptide in which the C-terminus of the LAG3 extracellular domain D1 / D2 is covalently linked to the light chain of sugimalimab via a linker peptide. The C-terminus of the heavy chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex, and this first polypeptide forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of the LAG3 extracellular domain D1 / D2 to the light chain of sugimalimab via a linker peptide. The C-terminus of the heavy chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the IgV region of the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex, and this first polypeptide forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of the LAG3 extracellular domain D1 / D2 to the light chain of sugimalimab via a linker peptide.

[0631] Provided below are exemplary amino acid sequences of a first polypeptide of a multifunctional fusion polypeptide complex formed by covalently linking the C-terminus of a CD86 extracellular domain to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, which forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of a LAG3 extracellular domain D1 to the light chain of sugimalimab (SEQ ID NO: 139) via a linker peptide. The C-terminus of a CD86 extracellular domain IgV region is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, which forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of a LAG3 extracellular domain D1 to the light chain of sugimalimab via a linker peptide. The C-terminus of the heavy chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex, and the first polypeptide forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of the LAG3 extracellular domain D1 to the light chain of sugimalimab via a linker peptide. The C-terminus of the heavy chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the IgV region of the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex, and the first polypeptide forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of the LAG3 extracellular domain D1 to the light chain of sugimalimab via a linker peptide.

[0632] Provided below are exemplary amino acid sequences of a first polypeptide of a multifunctional fusion polypeptide complex formed by covalently linking the C-terminus of the CD86 extracellular domain to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, which forms a multifunctional fusion polypeptide complex with a second polypeptide (SEQ ID NO: 140) formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain via a linker peptide. The C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, which forms a multifunctional fusion polypeptide complex with a second polypeptide (SEQ ID NO: 140) formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain via a linker peptide. The C-terminus of the heavy chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex, and the first polypeptide forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain via a linker peptide. The C-terminus of the heavy chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the IgV region of the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex, and the first polypeptide forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain via a linker peptide.

[0633] Provided below are exemplary amino acid sequences of a first polypeptide of a multifunctional fusion polypeptide complex formed by covalently linking the C-terminus of the CD86 extracellular domain to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, which forms a multifunctional fusion polypeptide complex with a second polypeptide (SEQ ID NO: 141) formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain D1 / D2 / D3 via a linker peptide. Provided below are exemplary amino acid sequences of a first polypeptide of a multifunctional fusion polypeptide complex formed by covalently linking the C-terminus of the IgV region of the CD86 extracellular domain to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, which forms a multifunctional fusion polypeptide complex with a second polypeptide (SEQ ID NO: 141) formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain D1 / D2 / D3 via a linker peptide. The C-terminus of the heavy chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex, and the first polypeptide forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain D1 / D2 / D3 via a linker peptide. The C-terminus of the heavy chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the CD86 extracellular domain IgV region via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex, and the first polypeptide forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain D1 / D2 / D3 via a linker peptide.

[0634] Provided below are exemplary amino acid sequences of a first polypeptide of a multifunctional fusion polypeptide complex formed by covalently linking the C-terminus of the CD86 extracellular domain to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, which forms a multifunctional fusion polypeptide complex with a second polypeptide (SEQ ID NO: 142) formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain D1 / D2 via a linker peptide. Provided below are exemplary amino acid sequences of a first polypeptide of a multifunctional fusion polypeptide complex formed by covalently linking the C-terminus of the IgV region of the CD86 extracellular domain to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, which forms a multifunctional fusion polypeptide complex with a second polypeptide (SEQ ID NO: 142) formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain D1 / D2 via a linker peptide. The C-terminus of the heavy chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex, and the first polypeptide forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain D1 / D2 via a linker peptide. The C-terminus of the heavy chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the CD86 extracellular domain IgV region via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex, and the first polypeptide forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain D1 / D2 via a linker peptide.

[0635] Provided below are exemplary amino acid sequences of a first polypeptide of a multifunctional fusion polypeptide complex formed by covalently linking the C-terminus of the CD86 extracellular domain to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, which forms a multifunctional fusion polypeptide complex with a second polypeptide (SEQ ID NO: 143) formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain D1 via a linker peptide. Provided below are exemplary amino acid sequences of a first polypeptide of a multifunctional fusion polypeptide complex formed by covalently linking the C-terminus of the IgV region of the CD86 extracellular domain to the heavy chain of the anti-PD-L1 antibody sugimalimab via a linker peptide, which forms a multifunctional fusion polypeptide complex with a second polypeptide (SEQ ID NO: 143) formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain D1 via a linker peptide. The C-terminus of the heavy chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the CD86 extracellular domain via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex, and the first polypeptide forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain D1 via a linker peptide. The C-terminus of the heavy chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the CD86 extracellular domain IgV region via a linker peptide to form a first polypeptide amino acid sequence of a multifunctional fusion polypeptide complex, and the first polypeptide forms a multifunctional fusion polypeptide complex with a second polypeptide formed by covalently linking the C-terminus of the light chain of sugimalimab to the LAG3 extracellular domain D1 via a linker peptide.

[0636] Provided below is an exemplary multifunctional fusion polypeptide complex in which the C-terminus of a CD86 extracellular domain is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a second polypeptide amino acid sequence (SEQ ID NO: 42), which together with a first polypeptide (SEQ ID NO: 144) (formed by covalently linking the C-terminus of a LAG3 extracellular domain to the heavy chain of sugimalimab via a linker peptide) constitutes the multifunctional fusion polypeptide complex. The C-terminus of the IgV region of a CD86 extracellular domain is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a second polypeptide amino acid sequence (SEQ ID NO: 43), which together with a first polypeptide (formed by covalently linking the C-terminus of a LAG3 extracellular domain to the heavy chain of sugimalimab via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the CD86 extracellular domain via a linker peptide to form the second polypeptide amino acid sequence (SEQ ID NO: 40) of the multifunctional fusion polypeptide complex, and the second polypeptide and the first polypeptide (formed by covalently linking the C-terminus of the LAG3 extracellular domain to the sugimalimab heavy chain via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently linked to the CD86 extracellular domain IgV region via a linker peptide to form the second polypeptide amino acid sequence (SEQ ID NO: 41) of the multifunctional fusion polypeptide complex, and the second polypeptide and the first polypeptide (formed by covalently linking the C-terminus of the LAG3 extracellular domain to the sugimalimab heavy chain via a linker peptide) constitute the multifunctional fusion polypeptide complex.

[0637] Provided below are exemplary multifunctional fusion polypeptide complexes in which the C-terminus of a CD86 extracellular domain is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a second polypeptide amino acid sequence, and the second polypeptide and a first polypeptide (SEQ ID NO: 145) (formed by the C-terminus of the LAG3 extracellular domain D1 / D2 / D3 covalently linked to the heavy chain of sugimalimab via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the IgV region of a CD86 extracellular domain is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a second polypeptide amino acid sequence, and the second polypeptide and a first polypeptide (formed by the C-terminus of the LAG3 extracellular domain D1 / D2 / D3 covalently linked to the heavy chain of sugimalimab via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently bound to the CD86 extracellular domain via a linker peptide to form the second polypeptide amino acid sequence of the multifunctional fusion polypeptide complex, and the second polypeptide (formed by covalently binding the C-terminus of the LAG3 extracellular domain D1 / D2 / D3 to the heavy chain of sugimalimab via a linker peptide) and the first polypeptide constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently bound to the IgV region of the CD86 extracellular domain via a linker peptide to form the second polypeptide amino acid sequence of the multifunctional fusion polypeptide complex, and the second polypeptide (formed by covalently binding the C-terminus of the LAG3 extracellular domain D1 / D2 / D3 to the heavy chain of sugimalimab via a linker peptide to form the first polypeptide) constitutes the multifunctional fusion polypeptide complex.

[0638] Provided below are exemplary multifunctional fusion polypeptide complexes in which the C-terminus of the CD86 extracellular domain is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a second polypeptide amino acid sequence, and the second polypeptide and a first polypeptide (SEQ ID NO: 146) (formed by the C-terminus of the LAG3 extracellular domain D1 / D2 covalently linked to the heavy chain of sugimalimab via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a second polypeptide amino acid sequence, and the second polypeptide and a first polypeptide (formed by the C-terminus of the LAG3 extracellular domain D1 / D2 covalently linked to the heavy chain of sugimalimab via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently bound to the CD86 extracellular domain via a linker peptide to form the second polypeptide amino acid sequence of the multifunctional fusion polypeptide complex, and the second polypeptide (formed by covalently binding the C-terminus of the LAG3 extracellular domain D1 / D2 to the heavy chain of sugimalimab via a linker peptide) and the first polypeptide constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently bound to the CD86 extracellular domain IgV region via a linker peptide to form the second polypeptide amino acid sequence of the multifunctional fusion polypeptide complex, and the second polypeptide (formed by covalently binding the C-terminus of the LAG3 extracellular domain D1 / D2 to the heavy chain of sugimalimab via a linker peptide to form the first polypeptide) constitutes the multifunctional fusion polypeptide complex.

[0639] Provided below are exemplary multifunctional fusion polypeptide complexes in which the C-terminus of the CD86 extracellular domain is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a second polypeptide amino acid sequence, and the second polypeptide and a first polypeptide (SEQ ID NO: 147) (formed by the C-terminus of the LAG3 extracellular domain D1 covalently linked to the heavy chain of sugimalimab via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the CD86 extracellular domain IgV region is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a second polypeptide amino acid sequence, and the second polypeptide and a first polypeptide (formed by the C-terminus of the LAG3 extracellular domain D1 covalently linked to the heavy chain of sugimalimab via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently bound to the CD86 extracellular domain via a linker peptide to form the second polypeptide amino acid sequence of the multifunctional fusion polypeptide complex, and the second polypeptide and the first polypeptide (formed by covalently binding the C-terminus of the LAG3 extracellular domain D1 to the heavy chain of sugimalimab via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently bound to the CD86 extracellular domain IgV region via a linker peptide to form the second polypeptide amino acid sequence of the multifunctional fusion polypeptide complex, and the second polypeptide and the first polypeptide (formed by covalently binding the C-terminus of the LAG3 extracellular domain D1 to the heavy chain of sugimalimab via a linker peptide) constitute the multifunctional fusion polypeptide complex.

[0640] Provided below are exemplary multifunctional fusion polypeptide complexes in which the C-terminus of the CD86 extracellular domain is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a second polypeptide amino acid sequence, and the second polypeptide and a first polypeptide (SEQ ID NO: 148) (formed by the C-terminus of the heavy chain of sugimalimab covalently linked to the LAG3 extracellular domain via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a second polypeptide amino acid sequence, and the second polypeptide and a first polypeptide (formed by the C-terminus of the heavy chain of sugimalimab covalently linked to the LAG3 extracellular domain via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently bound to the CD86 extracellular domain via a linker peptide to form the second polypeptide amino acid sequence of the multifunctional fusion polypeptide complex, and the second polypeptide and the first polypeptide (formed by the C-terminus of the heavy chain of sugimalimab covalently bound to the LAG3 extracellular domain via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently bound to the IgV region of the CD86 extracellular domain via a linker peptide to form the second polypeptide amino acid sequence of the multifunctional fusion polypeptide complex, and the second polypeptide and the first polypeptide (formed by the C-terminus of the heavy chain of sugimalimab covalently bound to the LAG3 extracellular domain via a linker peptide) constitute the multifunctional fusion polypeptide complex.

[0641] Provided below is an exemplary multifunctional fusion polypeptide complex in which the C-terminus of the CD86 extracellular domain is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form the second polypeptide amino acid sequence, and the second polypeptide and the first polypeptide (SEQ ID NO: 149) (formed by the C-terminus of the heavy chain of sugimalimab covalently linked to the LAG3 extracellular domain D1 / D2 / D3 via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form the second polypeptide amino acid sequence, and the second polypeptide and the first polypeptide (formed by the C-terminus of the heavy chain of sugimalimab covalently linked to the LAG3 extracellular domain D1 / D2 / D3 via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently bound to the CD86 extracellular domain via a linker peptide to form the second polypeptide amino acid sequence of the multifunctional fusion polypeptide complex, and the second polypeptide and the first polypeptide (formed by the C-terminus of the heavy chain of sugimalimab covalently bound to the LAG3 extracellular domain D1 / D2 / D3 via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently bound to the CD86 extracellular domain IgV region via a linker peptide to form the second polypeptide amino acid sequence of the multifunctional fusion polypeptide complex, and the second polypeptide and the first polypeptide (formed by the C-terminus of the heavy chain of sugimalimab covalently bound to the LAG3 extracellular domain D1 / D2 / D3 via a linker peptide) constitute the multifunctional fusion polypeptide complex.

[0642] Provided below are exemplary multifunctional fusion polypeptide complexes in which the C-terminus of the CD86 extracellular domain is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a second polypeptide amino acid sequence, and the second polypeptide and a first polypeptide (SEQ ID NO: 150) (formed by the C-terminus of the heavy chain of sugimalimab covalently linked to the LAG3 extracellular domain D1 / D2 via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form a second polypeptide amino acid sequence, and the second polypeptide and a first polypeptide (formed by the C-terminus of the heavy chain of sugimalimab covalently linked to the LAG3 extracellular domain D1 / D2 via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently bound to the CD86 extracellular domain via a linker peptide to form the second polypeptide amino acid sequence of the multifunctional fusion polypeptide complex, and the second polypeptide and the first polypeptide (formed by covalently binding the C-terminus of the heavy chain of sugimalimab to the LAG3 extracellular domain D1 / D2 via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently bound to the CD86 extracellular domain IgV region via a linker peptide to form the second polypeptide amino acid sequence of the multifunctional fusion polypeptide complex, and the second polypeptide and the first polypeptide (formed by covalently binding the C-terminus of the heavy chain of sugimalimab to the LAG3 extracellular domain D1 / D2 via a linker peptide) constitute the multifunctional fusion polypeptide complex.

[0643] Provided below is an exemplary multifunctional fusion polypeptide complex in which the C-terminus of the CD86 extracellular domain is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form the second polypeptide amino acid sequence, and the second polypeptide and a first polypeptide (SEQ ID NO: 151) (formed by the C-terminus of the heavy chain of sugimalimab covalently linked to the LAG3 extracellular domain D1 via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the IgV region of the CD86 extracellular domain is covalently linked to the light chain of the anti-PD-L1 antibody sugimalimab via a linker peptide to form the second polypeptide amino acid sequence, and the second polypeptide and a first polypeptide (formed by the C-terminus of the heavy chain of sugimalimab covalently linked to the LAG3 extracellular domain D1 via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently bound to the CD86 extracellular domain via a linker peptide to form the second polypeptide amino acid sequence of the multifunctional fusion polypeptide complex, and the second polypeptide and the first polypeptide (formed by the C-terminus of the heavy chain of sugimalimab covalently bound to the LAG3 extracellular domain D1 via a linker peptide) constitute the multifunctional fusion polypeptide complex. The C-terminus of the light chain of the anti-PD-L1 antibody sugimalimab is covalently bound to the CD86 extracellular domain IgV region via a linker peptide to form the second polypeptide amino acid sequence of the multifunctional fusion polypeptide complex, and the second polypeptide and the first polypeptide (formed by the C-terminus of the heavy chain of sugimalimab covalently bound to the LAG3 extracellular domain D1 via a linker peptide) constitute the multifunctional fusion polypeptide complex.

[0644] For example, antibody AN_Bif_Abs_0016 is composed of the Sugemalimab heavy chain antibody AN_Bif_Abs_0016 first polypeptide (SEQ ID NO: 4) and the Sugemalimab light chain second polypeptide (SEQ ID NO: 5). The AN_Bif_Abs_0016 antibody is assembled by introducing a vector containing a multi-nucleic acid sequence (SEQ ID NO: 152) that expresses the first polypeptide of AN_Bif_Abs_0016 and a vector containing a multi-nucleic acid sequence (SEQ ID NO: 153) that expresses the second polypeptide of AN_Bif_Abs_0016 into a host cell and expressing them.

[0645] For example, the bifunctional antibody AN_Bif_Abs_0220 is composed of a chimeric antibody AN_Bif_Abs_0220 first polypeptide (SEQ ID NO: 38) formed by covalently linking the C-terminus of the CD86 extracellular domain to the N-terminus of the sugemalimab heavy chain via a linker peptide (SEQ ID NO: 34), and a sugemalimab light chain second polypeptide (SEQ ID NO: 5). The AN_Bif_Abs_0220 bifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 154) that expresses the first polypeptide of AN_Bif_Abs_0220 and a vector containing a multinucleic acid sequence (SEQ ID NO: 153) that expresses the second polypeptide of AN_Bif_Abs_0220 into host cells and expressing them.

[0646] For example, the bifunctional antibody AN_Bif_Abs_0222 is composed of a chimeric antibody AN_Bif_Abs_0222 first polypeptide (SEQ ID NO: 36) formed by covalently linking the Sugemalimab heavy chain Fc domain to the N-terminus of the CD86 extracellular domain via a linker peptide (SEQ ID NO: 34), and a Sugemalimab light chain second polypeptide (SEQ ID NO: 5). The AN_Bif_Abs_0222 bifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 155) that expresses the first polypeptide of AN_Bif_Abs_0222 and a vector containing a multinucleic acid sequence (SEQ ID NO: 153) that expresses the second polypeptide of AN_Bif_Abs_0222 into host cells and expressing them.

[0647] For example, the bifunctional antibody AN_Bif_Abs_0703 is composed of a chimeric antibody AN_Bif_Abs_0703 first polypeptide (SEQ ID NO: 37) formed by covalently linking the Sugemalimab heavy chain Fc domain to the CD86 extracellular domain IgV region via a linker peptide (SEQ ID NO: 34), and a Sugemalimab light chain second polypeptide (SEQ ID NO: 5). The AN_Bif_Abs_0703 bifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 156) that expresses the first polypeptide of AN_Bif_Abs_0703 and a vector containing a multinucleic acid sequence (SEQ ID NO: 153) that expresses the second polypeptide of AN_Bif_Abs_0703 into host cells and expressing them.

[0648] For example, the bifunctional antibody AN_Bif_Abs_0158 is composed of a chimeric antibody AN_Bif_Abs_0158 first polypeptide (SEQ ID NO: 39) formed by covalently linking the C-terminus of the CD86 extracellular domain IgV region to the N-terminus of the sugemalimab heavy chain via a linker peptide (SEQ ID NO: 34), and a sugemalimab light chain second polypeptide (SEQ ID NO: 5). The AN_Bif_Abs_0158 bifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 157) that expresses the first polypeptide of AN_Bif_Abs_0158 and a vector containing a multinucleic acid sequence (SEQ ID NO: 153) that expresses the second polypeptide of AN_Bif_Abs_0158 into host cells and expressing them.

[0649] For example, the bifunctional antibody AN_Bif_Abs_0675 is composed of a sugemalimab heavy chain first polypeptide (SEQ ID NO: 4) and a chimeric antibody AN_Bif_Abs_0675 second polypeptide (SEQ ID NO: 43) formed by covalently linking the CD86 extracellular domain IgV region to the sugemalimab light chain via a linker peptide (SEQ ID NO: 34). The AN_Bif_Abs_0675 bifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 152) that expresses the first polypeptide of AN_Bif_Abs_0675 and a vector containing a multinucleic acid sequence (SEQ ID NO: 158) that expresses the second polypeptide of AN_Bif_Abs_0675 into host cells and expressing them.

[0650] For example, the bifunctional antibody AN_Bif_Abs_0676 is composed of a chimeric antibody AN_Bif_Abs_0676 second polypeptide (SEQ ID NO: 41) formed by covalently linking the Sugemalimab heavy chain first polypeptide (SEQ ID NO: 4) to the CD86 extracellular domain IgV region via a linker peptide (SEQ ID NO: 34). The AN_Bif_Abs_0676 bifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 152) that expresses the first polypeptide of AN_Bif_Abs_0676 and a vector containing a multinucleic acid sequence (SEQ ID NO: 159) that expresses the second polypeptide of AN_Bif_Abs_0676 into host cells and expressing them.

[0651] For example, the multifunctional antibody AN_Bif_Abs_0159 is composed of a chimeric antibody AN_Bif_Abs_0159 first polypeptide (SEQ ID NO: 39) formed by covalently linking the C-terminus of the CD86 extracellular domain IgV region to the N-terminus of the sugemalimab heavy chain via a linker peptide (SEQ ID NO: 34), and a second polypeptide (SEQ ID NO: 138) formed by covalently linking the C-terminus of LAG3 D1 / D2 to the sugemalimab light chain via a linker peptide (SEQ ID NO: 34). The AN_Bif_Abs_0159 multifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 157) that expresses the first polypeptide of AN_Bif_Abs_0159 and a vector containing a multinucleic acid sequence (SEQ ID NO: 160) that expresses the second polypeptide of AN_Bif_Abs_0159 into host cells and expressing them.

[0652] For example, the multifunctional antibody AN_Bif_Abs_0160 is composed of a chimeric antibody AN_Bif_Abs_0160 first polypeptide (SEQ ID NO: 39) formed by covalently linking the C-terminus of the CD86 extracellular domain IgV region to the N-terminus of the sugemalimab heavy chain via a linker peptide (SEQ ID NO: 34), and a second polypeptide (SEQ ID NO: 142) formed by covalently linking the C-terminus of the sugemalimab light chain to LAG3 D1 / D2 via a linker peptide (SEQ ID NO: 34). The AN_Bif_Abs_0160 multifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 157) that expresses the first polypeptide of AN_Bif_Abs_0160 and a vector containing a multinucleic acid sequence (SEQ ID NO: 161) that expresses the second polypeptide of AN_Bif_Abs_0160 into host cells and expressing them.

[0653] For example, the multifunctional antibody AN_Bif_Abs_0161 is composed of a chimeric antibody AN_Bif_Abs_0161 first polypeptide (SEQ ID NO: 114) formed by covalently linking the C-terminus of the CD86 extracellular domain IgV region to the N-terminus of the sugemalimab heavy chain via a linker peptide (SEQ ID NO: 34), and a sugemalimab light chain second polypeptide (SEQ ID NO: 5). The AN_Bif_Abs_0161 multifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 162) that expresses the first polypeptide of AN_Bif_Abs_0161 and a vector containing a multinucleic acid sequence (SEQ ID NO: 153) that expresses the second polypeptide of AN_Bif_Abs_0161 into host cells and expressing them.

[0654] For example, the multifunctional antibody AN_Bif_Abs_0644 is composed of a chimeric antibody AN_Bif_Abs_0644 first polypeptide (SEQ ID NO: 107) formed by covalently linking the C-terminus of the CD86 extracellular domain to the N-terminus of the sugemalimab heavy chain via a linker peptide (SEQ ID NO: 34) and further covalently linking the C-terminus of the sugemalimab heavy chain to LAG3 D1 via a linker peptide (SEQ ID NO: 34), and a sugemalimab light chain second polypeptide (SEQ ID NO: 5). The AN_Bif_Abs_0644 multifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 163) expressing the first polypeptide of AN_Bif_Abs_0644 and a vector containing a multinucleic acid sequence (SEQ ID NO: 153) expressing the second polypeptide of AN_Bif_Abs_0644 into host cells and expressing them.

[0655] For example, the multifunctional antibody AN_Bif_Abs_0646 is composed of a chimeric antibody AN_Bif_Abs_0646 first polypeptide (SEQ ID NO: 115) formed by covalently linking the C-terminus of the CD86 extracellular domain IgV region to the N-terminus of the sugemalimab heavy chain via a linker peptide (SEQ ID NO: 34) and further covalently linking the C-terminus of the sugemalimab heavy chain to LAG3 D1 via a linker peptide (SEQ ID NO: 34), and a sugemalimab light chain second polypeptide (SEQ ID NO: 5). The AN_Bif_Abs_0646 multifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 164) expressing the first polypeptide of AN_Bif_Abs_0646 and a vector containing a multinucleic acid sequence (SEQ ID NO: 153) expressing the second polypeptide of AN_Bif_Abs_0646 into host cells and expressing them.

[0656] For example, the multifunctional antibody AN_Bif_Abs_0162 is composed of a chimeric antibody AN_Bif_Abs_0162 first polypeptide (SEQ ID NO: 112) formed by covalently linking the C-terminus of the IgV region of the CD86 extracellular domain to the N-terminus of the sugemalimab heavy chain via a linker peptide (SEQ ID NO: 34), and a sugemalimab light chain second polypeptide (SEQ ID NO: 5). The AN_Bif_Abs_0162 multifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 165) expressing the first polypeptide of AN_Bif_Abs_0162 and a vector containing a multinucleic acid sequence (SEQ ID NO: 153) expressing the second polypeptide of AN_Bif_Abs_0162 into host cells and expressing them.

[0657] For example, the multifunctional antibody AN_Bif_Abs_0163 is composed of a chimeric antibody AN_Bif_Abs_0163 first polypeptide (SEQ ID NO: 104) formed by covalently linking the C-terminus of the CD86 extracellular domain to the N-terminus of the sugemalimab heavy chain via a linker peptide (SEQ ID NO: 34) and further covalently linking the C-terminus of the sugemalimab heavy chain to the LAG3 extracellular domain via a linker peptide (SEQ ID NO: 34), and a sugemalimab light chain second polypeptide (SEQ ID NO: 5). The AN_Bif_Abs_0163 multifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 166) that expresses the first polypeptide of AN_Bif_Abs_0163 and a vector containing a multinucleic acid sequence (SEQ ID NO: 153) that expresses the second polypeptide of AN_Bif_Abs_0163 into host cells and expressing them.

[0658] For example, the multifunctional antibody AN_Bif_Abs_0164 is composed of a chimeric antibody AN_Bif_Abs_0164 first polypeptide (SEQ ID NO: 108) formed by covalently linking the C-terminus of the LAG3 extracellular domain to the N-terminus of the sugemalimab heavy chain via a linker peptide (SEQ ID NO: 34), and a sugemalimab light chain second polypeptide (SEQ ID NO: 5). The AN_Bif_Abs_0164 multifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 167) that expresses the first polypeptide of AN_Bif_Abs_0164 and a vector containing a multinucleic acid sequence (SEQ ID NO: 153) that expresses the second polypeptide of AN_Bif_Abs_0164 into host cells and expressing them.

[0659] For example, the multifunctional antibody AN_Bif_Abs_0166 is composed of a chimeric antibody AN_Bif_Abs_0166 first polypeptide (SEQ ID NO: 116) formed by covalently linking the C-terminus of the LAG3 extracellular domain to the N-terminus of the sugemalimab heavy chain via a linker peptide (SEQ ID NO: 34), and a sugemalimab light chain second polypeptide (SEQ ID NO: 5). The AN_Bif_Abs_0166 multifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 168) that expresses the first polypeptide of AN_Bif_Abs_0166 and a vector containing a multinucleic acid sequence (SEQ ID NO: 153) that expresses the second polypeptide of AN_Bif_Abs_0166 into host cells and expressing them.

[0660] For example, antibody AN_Bif_Abs_0031 is composed of antibody AN_Bif_Abs_0031 first polypeptide (SEQ ID NO: 14) formed from envafolimab. The AN_Bif_Abs_0031 antibody is assembled by introducing a vector containing a polynucleotide sequence (SEQ ID NO: 169) that expresses the first polypeptide of AN_Bif_Abs_0031 into a host cell and expressing the vector.

[0661] For example, the bifunctional antibody AN_Bif_Abs_0129 is composed of a chimeric antibody AN_Bif_Abs_0129 first polypeptide (SEQ ID NO: 71) formed by covalently linking the C-terminus of the CD86 extracellular domain IgV region to the N-terminus of envafolimab via a linker peptide (SEQ ID NO: 34). The AN_Bif_Abs_0129 bifunctional antibody is assembled by introducing a vector containing a polynucleotide sequence (SEQ ID NO: 170) that expresses the first polypeptide of AN_Bif_Abs_0129 into a host cell and expressing it.

[0662] For example, the multifunctional antibody AN_Bif_Abs_0130 is composed of a chimeric antibody AN_Bif_Abs_0130 first polypeptide (SEQ ID NO: 171) formed by covalently linking the C-terminus of the CD86 extracellular domain IgV region to the N-terminus of LAG3 D1 / D2 via a linker peptide (SEQ ID NO: 34), and further covalently linking the C-terminus of LAG3 D1 / D2 to the N-terminus of envafolimab via a linker peptide (SEQ ID NO: 34).The AN_Bif_Abs_0130 multifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 172) that expresses the first polypeptide of AN_Bif_Abs_0130 into a host cell and expressing it.

[0663] For example, the multifunctional antibody AN_Bif_Abs_0131 is composed of a chimeric antibody AN_Bif_Abs_0131 first polypeptide (SEQ ID NO: 173) formed by covalently linking the C-terminus of the CD86 extracellular domain IgV region to the N-terminus of Envafolimab via a linker peptide (SEQ ID NO: 34), and further covalently linking the C-terminus of Envafolimab to LAG3 D1 / D2 via a linker peptide (SEQ ID NO: 34). The AN_Bif_Abs_0131 multifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 174) that expresses the first polypeptide of AN_Bif_Abs_0131 into a host cell and expressing it.

[0664] For example, the bifunctional antibody AN_Bif_Abs_0012 is composed of a chimeric antibody AN_Bif_Abs_0012 first polypeptide (SEQ ID NO: 44) formed by covalently linking the C-terminus of the durvalumab heavy chain to the CD86 extracellular domain via a linker peptide (SEQ ID NO: 34), and a durvalumab light chain second polypeptide (SEQ ID NO: 9). The AN_Bif_Abs_0012 multifunctional antibody is assembled by introducing a vector containing a multinucleic acid sequence (SEQ ID NO: 175) that expresses the first polypeptide of AN_Bif_Abs_0012 and a vector containing a multinucleic acid sequence (SEQ ID NO: 176) that expresses the second polypeptide of AN_Bif_Abs_0012 into host cells and expressing them.

[0665] For example, antibody AN_Bif_Abs_0001 is composed of the durvalumab heavy chain antibody AN_Bif_Abs_0001 first polypeptide (SEQ ID NO: 8) and the durvalumab light chain second polypeptide (SE...

Claims

1. A fusion polypeptide comprising a first domain capable of inhibiting PD-1 / PD-L1 signaling and a second domain comprising CD86 or a functionally active fragment thereof.

2. 2. The fusion polypeptide of claim 1, wherein the first domain is capable of binding to PD-L1 and / or PD-1.

3. The fusion polypeptide of claim 1 or 2, wherein the first domain comprises an antibody or an antigen-binding fragment thereof.

4. The fusion polypeptide of claim 3, wherein the antibody is selected from the group consisting of a recombinant antibody, a single domain antibody, a heavy chain antibody, a chimeric antibody, and a bispecific antibody.

5. The fusion polypeptide of claim 3, wherein the antigen-binding fragment is one or more selected from the group consisting of Fab, Fab', Fv fragment, F(ab')2, F(ab)2, scFv, di-scFv, VHH, and dAb.

6. 6. The fusion polypeptide of any one of claims 1 to 5, wherein the first domain comprises an antibody or antigen-binding fragment thereof selected from the group consisting of Sugemalimab, Durvalumab, Atezolizumab, Avelumab, Envafolimab, Pembrolizumab and Nivolumab.

7. 7. The fusion polypeptide of claim 1, wherein the first domain comprises HCDR1, HCDR2 and / or HCDR3 of an antibody heavy chain, and the antibody heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 15 and SEQ ID NO:

17.

8. 8. The fusion polypeptide of claim 1, wherein the first domain comprises an antibody heavy chain variable region VH, and the antibody heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 15 and SEQ ID NO:

17.

9. 9. The fusion polypeptide of claim 1, wherein the first domain comprises an antibody heavy chain, and the antibody heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 15 and SEQ ID NO:

17.

10. 10. The fusion polypeptide of any one of claims 1 to 9, wherein the first domain comprises LCDR1, LCDR2 and / or LCDR3 of an antibody light chain, and the antibody light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 7, SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 16 and SEQ ID NO:

18.

11. 11. The fusion polypeptide of claim 1, wherein the first domain comprises an antibody light chain variable region (VL), and the antibody light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 7, SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 16 and SEQ ID NO:

18.

12. 12. The fusion polypeptide of any one of claims 1 to 11, wherein the first domain comprises an antibody light chain, and the antibody light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 7, SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 16 and SEQ ID NO:

18.

13. The fusion polypeptide of any one of claims 1 to 12, wherein the first domain comprises Sugemalimab or an antigen-binding fragment thereof.

14. The fusion polypeptide of any one of claims 1 to 13, wherein the second domain is capable of binding to CD28 and / or CTLA4.

15. The fusion polypeptide of any one of claims 1 to 14, wherein the second domain is selected from the group consisting of human-derived CD86 or a functionally active fragment thereof, or mouse-derived CD86 or a functionally active fragment thereof.

16. The fusion polypeptide of any one of claims 1 to 15, wherein the second domain comprises the IgV domain of CD86 or a functionally active fragment thereof.

17. The fusion polypeptide of any one of claims 1 to 16, wherein the second domain comprises the extracellular domain of CD86 or a functionally active fragment thereof.

18. The fusion polypeptide of any one of claims 1 to 17, wherein the second domain comprises the amino acid sequence shown in SEQ ID NO: 2 or SEQ ID NO:

3.

19. The fusion polypeptide of any one of claims 1 to 15, wherein the CD86 or functionally active fragment thereof comprises a CD86 mutant polypeptide.

20. The fusion polypeptide of claim 19, wherein the CD86 mutant polypeptide comprises an amino acid substitution mutation of humanized CD86.

21. The CD86 mutant polypeptide of claim 20, wherein the CD86 mutant polypeptide comprises an extracellular domain IgV domain amino acid substitution mutant of humanized CD86.

22. The fusion polypeptide of any one of claims 19 to 21, wherein the CD86 mutant polypeptide comprises an amino acid substitution mutant of the IgV domain of CD86, and the mutation site of the amino acid substitution mutant of the IgV domain of CD86 comprises one or more amino acid mutations selected from the group consisting of A13I, A13L, A13F, A13M, Q25A, Q25I, Q25V, Q25F, Q25M, F33A, F33L, F33V, F33M, M60R, I89A, I89L, I89V, I89F, I89M, H90I, H90V, H90M, and H90F.

23. 23. The fusion polypeptide of any one of claims 19 to 22, wherein the CD86 variant polypeptide comprises one, two, three, four, five or more combinations of amino acid mutations selected from the group consisting of A13I, A13L, A13F, A13M, Q25A, Q25I, Q25V, Q25F, Q25M, F33A, F33L, F33V, F33M, M60R, I89A, I89L, I89V, I89F, I89M, H90I, H90V, H90M, H90F.

24. The CD86 mutant polypeptide comprises an amino acid substitution mutant of the IgV domain of CD86, and the amino acid substitution mutant is Q25I / F33L / H90I, Q25V / F33L / H90I, Q25I / F33V / H90I, Q25V / F33V / H90I, Q25I / F33L / H90V, Q25V / F33L / H90V, Q25I / F33V / H90V, Q25V / F33V / H90V, A13L / Q25I / F33L / H90I, A13I / Q25I / F33L / H90I, A13L / Q25V / F33L / H90I, A13I / Q25V / F 33L / H90I, Q25I / F33L / I89L / H90I, Q25I / F33L / M60R / H90I, Q25V / F33L / I89L / H90I, Q25V / F33L / M60 R / H90I, Q25F / F33L / H90I, Q25I / F33L / H90F, Q25I / F33A / H90I, Q25I / H90I, Q25I, H90I, Q25V / H90V, Q25V / H90I, Q25F / H90I, Q25F / H90V, A13F / Q25I / F33L / H90I, A13M / Q25I / F33L / H90I, Q25A / F33L / H9 24. The fusion polypeptide of any one of claims 19 to 23, comprising one or more combinations selected from the group consisting of Q25M / F33L / H90I, Q25I / F33M / H90I, Q25I / F33L / I89V / H90I, Q25I / F33L / I89F / H90I, Q25I / F33L / I89M / H90I, and Q25I / F33L / H90M.

25. The fusion polypeptide of any one of claims 19 to 24, wherein the CD86 mutant polypeptide comprises the IgV domain amino acid substitution mutants Q25I / F33L / H90I of CD86.

26. The fusion polypeptide of any one of claims 19 to 25, wherein the CD86 mutant polypeptide comprises an extracellular domain amino acid substitution mutant of humanized CD86.

27. The fusion polypeptide of any one of claims 19 to 26, wherein the CD86 mutant polypeptide comprises an amino acid substitution mutant in the extracellular domain of CD86, and the mutation site of the amino acid substitution mutant in the extracellular domain of CD86 comprises one or more amino acid mutations selected from the group consisting of A13I, A13L, A13F, A13M, Q25A, Q25I, Q25V, Q25F, Q25M, F33A, F33L, F33V, F33M, M60R, I89A, I89L, I89V, I89F, I89M, H90I, H90V, H90M, and H90F.

28. The CD86 mutant polypeptide comprises an amino acid substitution mutant of the extracellular domain of CD86, and the amino acid substitution mutants are Q25I / F33L / H90I, Q25V / F33L / H90I, Q25I / F33V / H90I, Q25V / F33V / H90I, Q25I / F33L / H90V, Q25V / F33L / H90V, Q25I / F33V / H90V, Q25V / F33V / H90V, A13L / Q25I / F33L / H90I, A13I / Q25I / F33L / H90I, A13L / Q25V / F33L / H90I, A13I / Q25V / F 33L / H90I, Q25I / F33L / I89L / H90I, Q25I / F33L / M60R / H90I, Q25V / F33L / I89L / H90I, Q25V / F33L / M60 R / H90I, Q25F / F33L / H90I, Q25I / F33L / H90F, Q25I / F33A / H90I, Q25I / H90I, Q25I, H90I, Q25V / H90V, Q25V / H90I, Q25F / H90I, Q25F / H90V, A13F / Q25I / F33L / H90I, A13M / Q25I / F33L / H90I, Q25A / F33L / H9 28. The fusion polypeptide of any one of claims 19 to 27, comprising one or more combinations selected from the group consisting of Q25M / F33L / H90I, Q25I / F33M / H90I, Q25I / F33L / I89V / H90I, Q25I / F33L / I89F / H90I, Q25I / F33L / I89M / H90I, and Q25I / F33L / H90M.

29. The fusion polypeptide of any one of claims 19 to 28, wherein the CD86 mutant polypeptide comprises the extracellular domain amino acid substitution mutants Q25I / F33L / H90I of CD86.

30. The fusion polypeptide of any one of claims 19 to 29, wherein the CD86 variant polypeptide comprises an amino acid sequence selected from the amino acid sequences set forth in SEQ ID NO: 178 to SEQ ID NO:

243.

31. The fusion polypeptide according to any one of claims 1 to 30, wherein the first domain and the second domain are directly or indirectly linked.

32. The fusion polypeptide of any one of claims 1 to 31, wherein the first domain comprises an antibody heavy chain, and the antibody heavy chain of the first domain and the second domain are directly or indirectly linked.

33. 33. The fusion polypeptide of any one of claims 1 to 32, wherein the first domain comprises an antibody heavy chain, and the C-terminus of the antibody heavy chain of the first domain is directly or indirectly linked to the N-terminus of the second domain.

34. 34. The fusion polypeptide of any one of claims 1 to 33, wherein the first domain comprises an antibody heavy chain, and the N-terminus of the antibody heavy chain of the first domain is directly or indirectly linked to the C-terminus of the second domain.

35. The fusion polypeptide of any one of claims 1 to 34, wherein the first domain comprises an antibody light chain, and the antibody light chain of the first domain and the second domain are directly or indirectly linked.

36. 36. The fusion polypeptide of any one of claims 1 to 35, wherein the first domain comprises an antibody light chain, and the C-terminus of the antibody light chain of the first domain is directly or indirectly linked to the N-terminus of the second domain.

37. 37. The fusion polypeptide of any one of claims 1 to 36, wherein the first domain comprises an antibody light chain, and the N-terminus of the antibody light chain of the first domain is directly or indirectly linked to the C-terminus of the second domain.

38. 38. The fusion polypeptide of any one of claims 1 to 37, wherein the first domain comprises an antibody heavy chain and an antibody light chain, and the C-terminus of the antibody heavy chain of the first domain is directly or indirectly linked to the N-terminus of the second domain, and the C-terminus of the antibody light chain of the first domain is directly or indirectly linked to the N-terminus of the second domain.

39. 39. The fusion polypeptide of any one of claims 1 to 38, wherein the first domain comprises an antibody heavy chain and an antibody light chain, and the N-terminus of the antibody heavy chain of the first domain is directly or indirectly linked to the C-terminus of the second domain, and the N-terminus of the antibody light chain of the first domain is directly or indirectly linked to the C-terminus of the second domain.

40. The fusion polypeptide of any one of claims 1 to 39, wherein the indirect linkage comprises a linkage via a linker.

41. 41. The fusion polypeptide of claim 40, wherein the linker comprises an amino acid sequence selected from the group consisting of SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34 and SEQ ID NO:

35.

42. 42. The fusion polypeptide of any one of claims 1 to 41, wherein the fusion polypeptide comprises an amino acid sequence selected from the group consisting of SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, SEQ ID NO:41, SEQ ID NO:42, SEQ ID NO:43, SEQ ID NO:44, SEQ ID NO:45, SEQ ID NO:46, SEQ ID NO:47, SEQ ID NO:48, SEQ ID NO:49, SEQ ID NO:50, SEQ ID NO:51, SEQ ID NO:52, SEQ ID NO:53, SEQ ID NO:54, SEQ ID NO:55, SEQ ID NO:56, SEQ ID NO:57, SEQ ID NO:58, SEQ ID NO:59, SEQ ID NO:60, SEQ ID NO:61, SEQ ID NO:62, SEQ ID NO:63, SEQ ID NO:64, SEQ ID NO:65, SEQ ID NO:66, SEQ ID NO:67, SEQ ID NO:68, SEQ ID NO:69, SEQ ID NO:70, SEQ ID NO:71, SEQ ID NO:72, SEQ ID NO:73, SEQ ID NO:74, SEQ ID NO:75, SEQ ID NO:76, SEQ ID NO:77, SEQ ID NO:78, SEQ ID NO:79, SEQ ID NO:80, SEQ ID NO:81, SEQ ID NO:82, SEQ ID NO:83, SEQ ID NO:84, SEQ ID NO:85, SEQ ID NO:86 and SEQ ID NO:

87.

43. 43. The fusion polypeptide of any one of claims 1 to 42, wherein the fusion polypeptide comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 418 to 450, 451 to 483, and 484 to 615.

44. The fusion polypeptide of any one of claims 1 to 43, wherein the fusion polypeptide further comprises a third domain capable of activating an innate immune response.

45. 45. The fusion polypeptide of claim 44, wherein the third domain is capable of binding to an MHC II molecule on an antigen-presenting cell.

46. The fusion polypeptide of any one of claims 44 to 45, wherein the third domain comprises LAG3 or a functionally active fragment thereof.

47. 47. The fusion polypeptide of any one of claims 44 to 46, wherein the third domain is selected from the group consisting of LAG3 of human origin or a functionally active fragment thereof, and LAG3 of mouse origin or a functionally active fragment thereof.

48. 48. The fusion polypeptide of any one of claims 44 to 47, wherein the third domain comprises the extracellular domain of LAG3 or a functionally active fragment thereof.

49. The fusion polypeptide of any one of claims 44 to 48, wherein the third domain comprises IgD1, IgD2, IgD3 and / or IgD4 of LAG3 or a functionally active fragment thereof.

50. 50. The fusion polypeptide of any one of claims 44 to 49, wherein the third domain comprises IgD1, IgD1-IgD2, IgD1-IgD2-IgD3, and / or IgD1-IgD2-IgD3-IgD4 of LAG3 or a functionally active fragment thereof.

51. 51. The fusion polypeptide of any one of claims 44 to 50, wherein the third domain comprises an amino acid sequence selected from the group consisting of SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27 and SEQ ID NO:

28.

52. A fusion polypeptide according to any one of claims 44 to 51, wherein the LAG3 or a functionally active fragment thereof comprises a LAG3 mutant polypeptide, the LAG3 mutant polypeptide comprising a truncation and / or mutation based on human-derived LAG3.

53. The fusion polypeptide of claim 52, wherein the LAG3 mutant polypeptide comprises a truncation based on the extracellular domain polypeptide of wild-type human-derived LAG3.

54. A fusion polypeptide described in any one of claims 52 to 53, wherein the LAG3 mutant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3, and is shortened by 0 to 124 amino acids at the N-terminus and ends at between amino acid positions 121 and 167 at the C-terminus.

55. The fusion polypeptide of any one of claims 52 to 54, wherein the LAG3 mutant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3 and is shortened at the N-terminus by 0, 5, 21, 37, 45, 60, 71, 74, 79, 84, 89, 94, 99, 104, 109 or 114 amino acids.

56. The fusion polypeptide of any one of claims 52 to 55, wherein the LAG3 mutant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3 and is shortened at the N-terminus by 0, 5, 21, 37, 45, 60, 71, 74, 81, or 99 amino acids.

57. The fusion polypeptide of any one of claims 52 to 56, wherein the LAG3 mutant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3 and is shortened at the N-terminus by 0, 5, 21, 37, 45, 60, 71, or 74 amino acids.

58. The fusion polypeptide of any one of claims 52 to 57, wherein the LAG3 mutant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3 and the C-terminus ends at amino acid position 121, 122, 129, 136, 146, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 or 167.

59. A fusion polypeptide described in any one of claims 52 to 58, wherein the LAG3 mutant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3, and the C-terminus ends at amino acid position 121, 122, 129, 136, 146, 156, 161 or 167.

60. A fusion polypeptide described in any one of claims 52 to 59, wherein the LAG3 mutant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3, and the C-terminus ends at amino acid position 121, 146, 156, 161 or 167.

61. A fusion polypeptide described in any one of claims 52 to 60, wherein the LAG3 mutant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3, and the C-terminus ends at amino acid position 156, 161 or 167.

62. The fusion polypeptide of any one of claims 52 to 61, wherein the LAG3 mutant polypeptide is based on the extracellular domain polypeptide of wild-type human LAG3 and is shortened by 0, 5, 21, 37, 45, 60, 71, 74, 81, or 99 amino acids at the N-terminus and ends at amino acid position 121, 122, 129, 136, 146, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, or 167 at the C-terminus.

63. The fusion polypeptide of any one of claims 52 to 62, wherein the LAG3 mutant polypeptide is based on the extracellular domain polypeptide of wild-type human-derived LAG3 and is shortened by 0, 5, 21, 37, 45, 60, 71, or 74 amino acids at the N-terminus and ends at amino acid position 122, 129, 136, 146, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, or 167 at the C-terminus.

64. The fusion polypeptide of any one of claims 52 to 63, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site.

65. 65. The fusion polypeptide of any one of claims 52 to 64, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site, and the mutation at the amino acid site is arginine (Arg) at residue 97.

66. 66. The fusion polypeptide of any one of claims 52 to 65, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site, wherein the mutation at the amino acid site mutates arginine (Arg) at residue 97 to glutamic acid (Glu).

67. 67. The fusion polypeptide of any one of claims 52 to 66, wherein the LAG3 mutant polypeptide is selected from the amino acid sequences set forth in SEQ ID NO: 244 to SEQ ID NO:

349.

68. 68. The fusion polypeptide of any one of claims 52 to 67, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site, wherein the mutation at the amino acid site is at R110, R113, R119, R129, G130 and / or R141.

69. 69. The fusion polypeptide of any one of claims 52 to 68, wherein the LAG3 mutant polypeptide is N-terminally shortened by 81 amino acids and C-terminally terminates at amino acid position 121, with mutations introduced at positions R110 and / or R113 and / or R119.

70. 70. The fusion polypeptide of any one of claims 52 to 69, wherein the LAG3 mutant polypeptide is N-terminally shortened by 99 amino acids and C-terminally terminates at amino acid position 146, with mutations introduced at positions R129 and / or G130 and / or R141.

71. 71. The fusion polypeptide of any one of claims 52 to 70, wherein the LAG3 mutant polypeptide is N-terminally shortened by 81 amino acids and C-terminally terminates at amino acid position 146, and mutations are introduced at positions R110 and / or R113 and / or R119 and / or R129 and / or G130 and / or R141.

72. 72. The fusion polypeptide of any one of claims 52 to 71, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid position where amino acid R110 is mutated to K110.

73. 73. The fusion polypeptide of any one of claims 52 to 72, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid position where amino acid R113 is mutated to K113.

74. 74. The fusion polypeptide of any one of claims 52 to 73, wherein the LAG3 mutant polypeptide comprises a mutation at the amino acid position where amino acid R119 is mutated to K119.

75. 75. The fusion polypeptide of any one of claims 52 to 74, wherein the LAG3 mutant polypeptide comprises a mutation at the amino acid position where amino acid R129 is mutated to K129.

76. 76. The fusion polypeptide of any one of claims 52 to 75, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site where amino acid G130 is mutated to P130, or A130, or T130, or Y130, or S130.

77. 77. The fusion polypeptide of any one of claims 52 to 76, wherein the LAG3 mutant polypeptide comprises a mutation at the amino acid position where amino acid R141 is mutated to K141.

78. The fusion polypeptide of any one of claims 52 to 77, wherein the amino acid sequence of the LAG3 mutant polypeptide is selected from the amino acid sequences of any of SEQ ID NO: 1285 to SEQ ID NO: 1318.

79. 79. The fusion polypeptide of any one of claims 44 to 78, wherein the first domain is directly or indirectly linked to the third domain.

80. 80. The fusion polypeptide of any one of claims 44 to 79, wherein the first domain comprises an antibody heavy chain, and the antibody heavy chain of the first domain is directly or indirectly linked to the third domain.

81. 81. The fusion polypeptide of any one of claims 44 to 80, wherein the first domain comprises an antibody heavy chain, and the C-terminus of the antibody heavy chain of the first domain is linked directly or indirectly to the N-terminus of the third domain.

82. 82. The fusion polypeptide of any one of claims 44 to 81, wherein the first domain comprises an antibody heavy chain, and the N-terminus of the antibody heavy chain of the first domain is linked directly or indirectly to the C-terminus of the third domain.

83. 83. The fusion polypeptide of any one of claims 44 to 82, wherein the first domain comprises an antibody light chain, and the light chain of the first domain is linked directly or indirectly to the third domain.

84. 84. The fusion polypeptide of any one of claims 44 to 83, wherein the first domain comprises an antibody light chain, and the C-terminus of the light chain of the first domain is linked directly or indirectly to the N-terminus of the third domain.

85. 85. The fusion polypeptide of any one of claims 44 to 84, wherein the first domain comprises an antibody light chain, and the N-terminus of the light chain of the first domain is linked directly or indirectly to the C-terminus of the third domain.

86. 86. The fusion polypeptide of any one of claims 44 to 85, wherein the second domain is directly or indirectly linked to the third domain.

87. 87. The fusion polypeptide of any one of claims 44 to 86, wherein the C-terminus of the second domain is linked directly or indirectly to the N-terminus of the third domain.

88. 88. The fusion polypeptide of any one of claims 44 to 87, wherein the N-terminus of the second domain is linked directly or indirectly to the C-terminus of the third domain.

89. 89. The fusion polypeptide of any one of claims 44 to 88, wherein the first domain is directly or indirectly linked to the second domain, and the second domain is directly or indirectly linked to the third domain.

90. 90. The fusion polypeptide of any one of claims 44 to 89, wherein the C-terminus of the first domain is linked, directly or indirectly, to the N-terminus of the second domain, and the C-terminus of the second domain is linked, directly or indirectly, to the N-terminus of the third domain.

91. 91. The fusion polypeptide of any one of claims 44 to 90, wherein the first domain comprises an antibody heavy chain, the C-terminus of the antibody heavy chain of the first domain is linked, directly or indirectly, to the N-terminus of the second domain, and the C-terminus of the second domain is linked, directly or indirectly, to the N-terminus of the third domain.

92. 92. The fusion polypeptide of any one of claims 44 to 91, wherein the first domain comprises an antibody light chain, the C-terminus of the antibody light chain of the first domain is linked, directly or indirectly, to the N-terminus of the second domain, and the C-terminus of the second domain is linked, directly or indirectly, to the N-terminus of the third domain.

93. 93. The fusion polypeptide of any one of claims 44 to 92, wherein the N-terminus of the first domain is linked, directly or indirectly, to the C-terminus of the second domain, and the N-terminus of the second domain is linked, directly or indirectly, to the C-terminus of the third domain.

94. 94. The fusion polypeptide of any one of claims 44 to 93, wherein the first domain comprises an antibody heavy chain, the N-terminus of the antibody heavy chain of the first domain is linked, directly or indirectly, to the C-terminus of the second domain, and the N-terminus of the second domain is linked, directly or indirectly, to the C-terminus of the third domain.

95. 95. The fusion polypeptide of any one of claims 44 to 94, wherein the first domain comprises an antibody light chain, the N-terminus of the antibody light chain of the first domain being linked, directly or indirectly, to the C-terminus of the second domain, and the N-terminus of the second domain being linked, directly or indirectly, to the C-terminus of the third domain.

96. 96. The fusion polypeptide of any one of claims 44 to 95, wherein the first domain is directly or indirectly linked to the third domain, and the third domain is directly or indirectly linked to the second domain.

97. 97. The fusion polypeptide of any one of claims 44 to 96, wherein the C-terminus of the first domain is linked, directly or indirectly, to the N-terminus of the third domain, and the C-terminus of the third domain is linked, directly or indirectly, to the N-terminus of the second domain.

98. 98. The fusion polypeptide of any one of claims 44 to 97, wherein the first domain comprises an antibody heavy chain, the C-terminus of the antibody heavy chain of the first domain is linked, directly or indirectly, to the N-terminus of the third domain, and the C-terminus of the third domain is linked, directly or indirectly, to the N-terminus of the second domain.

99. 99. The fusion polypeptide of any one of claims 44 to 98, wherein the first domain comprises an antibody light chain, the C-terminus of the antibody light chain of the first domain is linked, directly or indirectly, to the N-terminus of the third domain, and the C-terminus of the third domain is linked, directly or indirectly, to the N-terminus of the second domain.

100. 99. The fusion polypeptide of any one of claims 44 to 99, wherein the N-terminus of the first domain is linked, directly or indirectly, to the C-terminus of the third domain, and the N-terminus of the third domain is linked, directly or indirectly, to the C-terminus of the second domain.

101. 101. The fusion polypeptide of any one of claims 44 to 100, wherein the first domain comprises an antibody heavy chain, the N-terminus of the antibody heavy chain of the first domain being linked, directly or indirectly, to the C-terminus of the third domain, and the N-terminus of the third domain being linked, directly or indirectly, to the C-terminus of the second domain.

102. 102. The fusion polypeptide of any one of claims 44 to 101, wherein the first domain comprises an antibody light chain, the N-terminus of the antibody light chain of the first domain being linked, directly or indirectly, to the C-terminus of the third domain, and the N-terminus of the third domain being linked, directly or indirectly, to the C-terminus of the second domain.

103. 103. The fusion polypeptide of any one of claims 44 to 102, wherein the first domain is directly or indirectly linked to the second domain, and the first domain is directly or indirectly linked to the third domain.

104. 104. The fusion polypeptide of any one of claims 44 to 103, wherein the C-terminus of the first domain is linked, directly or indirectly, to the N-terminus of the second domain, and the N-terminus of the first domain is linked, directly or indirectly, to the C-terminus of the third domain.

105. 105. The fusion polypeptide of any one of claims 44 to 104, wherein the first domain comprises an antibody heavy chain, the C-terminus of the antibody heavy chain of the first domain being linked, directly or indirectly, to the N-terminus of the second domain, and the N-terminus of the antibody heavy chain of the first domain being linked, directly or indirectly, to the C-terminus of the third domain.

106. 106. The fusion polypeptide of any one of claims 44 to 105, wherein the first domain comprises an antibody light chain, the C-terminus of the antibody light chain of the first domain being linked, directly or indirectly, to the N-terminus of the second domain, and the N-terminus of the antibody light chain of the first domain being linked, directly or indirectly, to the C-terminus of the third domain.

107. 107. The fusion polypeptide of any one of claims 44 to 106, wherein the first domain comprises an antibody heavy chain, the C-terminus of the antibody heavy chain of the first domain being linked, directly or indirectly, to the N-terminus of the second domain, and the N-terminus of the antibody light chain of the first domain being linked, directly or indirectly, to the C-terminus of the third domain.

108. 108. The fusion polypeptide of any one of claims 44 to 107, wherein the first domain comprises an antibody light chain, the C-terminus of the antibody light chain of the first domain being linked, directly or indirectly, to the N-terminus of the second domain, and the N-terminus of the antibody heavy chain of the first domain being linked, directly or indirectly, to the C-terminus of the third domain.

109. 109. The fusion polypeptide of any one of claims 44 to 108, wherein the N-terminus of the first domain is linked, directly or indirectly, to the C-terminus of the second domain, and the C-terminus of the first domain is linked, directly or indirectly, to the N-terminus of the third domain.

110. 110. The fusion polypeptide of any one of claims 44 to 109, wherein the first domain comprises an antibody heavy chain, the N-terminus of the antibody heavy chain of the first domain being linked, directly or indirectly, to the C-terminus of the second domain, and the C-terminus of the antibody heavy chain of the first domain being linked, directly or indirectly, to the N-terminus of the third domain.

111. 111. The fusion polypeptide of any one of claims 44 to 110, wherein the first domain comprises an antibody light chain, the N-terminus of the antibody light chain of the first domain being linked, directly or indirectly, to the C-terminus of the second domain, and the C-terminus of the antibody light chain of the first domain being linked, directly or indirectly, to the N-terminus of the third domain.

112. 112. The fusion polypeptide of any one of claims 44 to 111, wherein the first domain comprises an antibody heavy chain and an antibody light chain, and the N-terminus of the antibody heavy chain of the first domain is linked, directly or indirectly, to the C-terminus of the second domain, and the C-terminus of the antibody light chain of the first domain is linked, directly or indirectly, to the N-terminus of the third domain.

113. 113. The fusion polypeptide of any one of claims 44 to 112, wherein the first domain comprises an antibody heavy chain and an antibody light chain, the N-terminus of the antibody light chain of the first domain being linked, directly or indirectly, to the C-terminus of the second domain, and the C-terminus of the antibody heavy chain of the first domain being linked, directly or indirectly, to the N-terminus of the third domain.

114. The fusion polypeptide according to any one of claims 44 to 113, wherein the indirect linkage includes linkage via a linker.

115. 115. The fusion polypeptide of claim 114, wherein the linker comprises an amino acid sequence selected from the group consisting of SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34 and SEQ ID NO:

35.

116. The fusion polypeptide may comprise SEQ ID NO:88, SEQ ID NO:89, SEQ ID NO:90, SEQ ID NO:91, SEQ ID NO:92, SEQ ID NO:93, SEQ ID NO:94, SEQ ID NO:95, SEQ ID NO:96, SEQ ID NO:97, SEQ ID NO:98, SEQ ID NO:99, SEQ ID NO:100, SEQ ID NO:101, SEQ ID NO:102, SEQ ID NO:103, SEQ ID NO:104, SEQ ID NO:105, SEQ ID NO:106, SEQ ID NO:107, SEQ ID NO:108, SEQ ID NO:109, SEQ ID NO:110, SEQ ID NO:111, SEQ ID NO:112, SEQ ID NO:113, SEQ ID NO:114, SEQ ID NO:115, 16. The fusion polypeptide of any one of claims 44 to 115, comprising an amino acid sequence selected from the group consisting of: SEQ ID NO:116, SEQ ID NO:117, SEQ ID NO:118, SEQ ID NO:119, SEQ ID NO:120, SEQ ID NO:121, SEQ ID NO:122, SEQ ID NO:123, SEQ ID NO:124, SEQ ID NO:125, SEQ ID NO:126, SEQ ID NO:127, SEQ ID NO:128, SEQ ID NO:129, SEQ ID NO:130, SEQ ID NO:131, SEQ ID NO:132, SEQ ID NO:133, SEQ ID NO:134, SEQ ID NO:135, SEQ ID NO:171, SEQ ID NO:

173.

117. The fusion polypeptide of any one of claims 44 to 116, wherein the fusion polypeptide comprises an amino acid sequence selected from SEQ ID NOs:751 to 1024, 1249, 1251, 1253 and 1255.

118. The fusion polypeptide of any one of claims 44 to 117, wherein the fusion polypeptide comprises the heavy chain of a multifunctional LAG3 mutant fusion Sugimalimab-CD86 IgV Q25I / F33L / H90I antibody polypeptide complex, and the amino acid sequence of the heavy chain is selected from SEQ ID NOs: 1387 to 1420.

119. The fusion polypeptide of any one of claims 44 to 118, wherein the fusion polypeptide comprises the sequence of any of SEQ ID NOs: 1421 and 1426.

120. A CD86 mutant polypeptide comprising an amino acid substitution mutation of humanized CD86.

121. The CD86 mutant polypeptide of claim 120, wherein the CD86 mutant polypeptide comprises an amino acid substitution mutant of the extracellular domain IgV domain of humanized CD86.

122. The CD86 mutant polypeptide according to any one of claims 120 to 121, wherein the CD86 mutant polypeptide comprises an amino acid substitution mutant in the IgV domain of CD86, and the mutation site of the amino acid substitution mutant in the IgV domain of CD86 comprises one or more amino acid mutations selected from the group consisting of A13I, A13L, A13F, A13M, Q25A, Q25I, Q25V, Q25F, Q25M, F33A, F33L, F33V, F33M, M60R, I89A, I89L, I89V, I89F, I89M, H90I, H90V, H90M, and H90F.

123. 123. The CD86 variant polypeptide of any one of claims 120 to 122, wherein the CD86 variant polypeptide comprises one, two, three, four, five or more combinations of amino acid mutations selected from the group consisting of A13I, A13L, A13F, A13M, Q25A, Q25I, Q25V, Q25F, Q25M, F33A, F33L, F33V, F33M, M60R, I89A, I89L, I89V, I89F, I89M, H90I, H90V, H90M and H90F.

124. The CD86 mutant polypeptide comprises an amino acid substitution mutant of the IgV domain of CD86, and the amino acid substitution mutants include Q25I / F33L / H90I, Q25V / F33L / H90I, Q25I / F33V / H90I, Q25V / F33V / H90I, Q25I / F33L / H90V, Q25V / F33L / H90V, Q25I / F33V / H90V, Q25V / F33V / H90V, A13L / Q25I / F33L / H90I, A13I / Q25I / F33L / H90I, A13L / Q25V / F33L / H90I, A13I / Q25V / F33 L / H90I, Q25I / F33L / I89L / H90I, Q25I / F33L / M60R / H90I, Q25V / F33L / I89L / H90I, Q25V / F33L / M60R / H9 0I, Q25F / F33L / H90I, Q25I / F33L / H90F, Q25I / F33A / H90I, Q25I / H90I, Q25I, H90I, Q25V / H90V, Q25V / H90I, Q25F / H90I, Q25F / H90V, A13F / Q25I / F33L / H90I, A13M / Q25I / F33L / H90I, Q25A / F33L / H90I, Q25 124. The CD86 variant polypeptide of any one of claims 120 to 123, comprising one or more combinations selected from the group consisting of M / F33L / H90I, Q25I / F33M / H90I, Q25I / F33L / I89V / H90I, Q25I / F33L / I89F / H90I, Q25I / F33L / I89M / H90I and Q25I / F33L / H90M.

125. The CD86 mutant polypeptide of any one of claims 120 to 124, wherein the CD86 mutant polypeptide comprises the amino acid substitution mutations Q25I / F33L / H90I in the IgV domain of CD86.

126. The CD86 mutant polypeptide of any one of claims 120 to 125, wherein the CD86 mutant polypeptide comprises an amino acid substitution mutant of the extracellular domain of humanized CD86.

127. 127. The CD86 variant polypeptide of any one of claims 120 to 126, wherein the CD86 variant polypeptide comprises an amino acid substitution variant of the extracellular domain of CD86, wherein the amino acid substitution variant of the extracellular domain of CD86 comprises one or more mutations of amino acid residues selected from the group consisting of: A13I, A13L, A13F, A13M, Q25A, Q25I, Q25V, Q25F, Q25M, F33A, F33L, F33V, F33M, M60R, I89A, I89L, I89V, I89F, I89M, H90I, H90V, H90M, H90F.

128. The CD86 variant polypeptide of any one of claims 120 to 127, wherein the CD86 variant polypeptide comprises amino acid substitution variants of the extracellular domain of CD86, and the amino acid substitution variants comprise one or more sets of combinations selected from the group consisting of: Q25I / F33L / H90I, Q25V / F33L / H90I, Q25I / F33V / H90I, Q25V / F33V / H90I, Q25I / F33 L / H90V, Q25V / F33L / H90V, Q25I / F33V / H90V, Q25V / F33V / H90V, A13L / Q25I / F33L / H 90I, A13I / Q25I / F33L / H90I, A13L / Q25V / F33L / H90I, A13I / Q25V / F33L / H90I, Q25I / F33L / I89L / H90I, Q25I / F33L / M60R / H90I, Q25V / F33L / I89L / H90I, Q25V / F33L / M6 0R / H90I, Q25F / F33L / H90I, Q25I / F33L / H90F, Q25I / F33A / H90I, Q25I / H90I, Q25I, H90I, Q25V / H90V, Q25V / H90I, Q25F / H90I, Q25F / H90V, A13F / Q25I / F33L / H90I, A13 M / Q25I / F33L / H90I, Q25A / F33L / H90I, Q25M / F33L / H90I, Q25I / F33M / H90I, Q25I / F 33L / I89V / H90I, Q25I / F33L / I89F / H90I, Q25I / F33L / I89M / H90I, Q25I / F33L / H90M.

129. 129. The CD86 mutant polypeptide of any one of claims 120 to 128, wherein the CD86 mutant polypeptide comprises the amino acid substitution mutations Q25I / F33L / H90I in the extracellular domain of CD86.

130. 130. The CD86 variant polypeptide of any one of claims 120 to 129, wherein said CD86 variant polypeptide comprises an amino acid sequence selected from SEQ ID NO: 178 to SEQ ID NO:

243.

131. 131. A fusion polypeptide comprising a first domain and a second domain, wherein the first domain comprises a CD86 mutant polypeptide according to any one of claims 120 to 130, and the second domain comprises an antibody, an antigen-binding fragment of an antibody, or an immunoglobulin Fc domain.

132. 132. The fusion polypeptide of claim 131, wherein the second domain comprises an antibody or an antigen-binding fragment thereof.

133. The fusion polypeptide of any one of claims 131 to 132, wherein the antibody is an antibody selected from the group consisting of an immunoglobulin IgG antibody, a recombinant antibody, a chimeric antibody, a heavy chain antibody, a single domain antibody, and a bispecific antibody.

134. the second domain is selected from the group consisting of PD-L1, PD-L2, PD-1, OX40, 4-1BB, ICOS, TIGIT, CTLA4, LAG3, CD3, VEGF, VEGFR, CD47, HGF, Trop2, EpCAM, CCR8, CCR4, CCR5, GPRC5D, BCMA, CD19, CD20, and HER-2 134. The fusion polypeptide of any one of claims 131 to 133, which is capable of binding to one or more of the targets set forth in the group consisting of neu, DLL1, HER-3, HER-4, EGFR, PSMA, CEA, MUC-1 (mucin), MUC2, MUC3, MUC4, MUC5AC, MUC5B, MUC7, CD123, CD33, CD30, CD38, NKG2A, Nkp36, and Tim3.

135. 135. The fusion polypeptide of any one of claims 131 to 134, wherein the second domain is capable of binding to CD3.

136. 136. The fusion polypeptide of any one of claims 131 to 135, wherein the second domain comprises a CD3B219 antibody or an antigen-binding fragment thereof.

137. 137. The fusion polypeptide of any one of claims 131 to 136, wherein the second domain comprises HCDR1, HCDR2 and / or HCDR3 of an antibody heavy chain, and the antibody heavy chain comprises the amino acid sequence set forth in SEQ ID NO:

616.

138. 138. The fusion polypeptide of any one of claims 131 to 137, wherein the second domain comprises a heavy chain variable region VH of an antibody, and the antibody heavy chain comprises the amino acid sequence set forth in SEQ ID NO:

616.

139. 139. The fusion polypeptide of any one of claims 131 to 138, wherein the second domain comprises a heavy chain of an antibody, and the antibody heavy chain comprises the amino acid sequence set forth in SEQ ID NO:

616.

140. 140. The fusion polypeptide of any one of claims 131 to 139, wherein the second domain comprises LCDR1, LCDR2 and / or LCDR3 of an antibody light chain, and wherein the antibody light chain comprises the amino acid sequence set forth in SEQ ID NO:

617.

141. 141. The fusion polypeptide of any one of claims 131 to 140, wherein the second domain comprises a light chain variable region VL of an antibody, and the antibody light chain comprises the amino acid sequence set forth in SEQ ID NO:

617.

142. 142. The fusion polypeptide of any one of claims 131 to 141, wherein the second domain comprises a light chain of an antibody, and the antibody light chain comprises the amino acid sequence set forth in SEQ ID NO:

617.

143. 143. The fusion polypeptide of any one of claims 131-142, wherein the immunoglobulin Fc domain comprises an IgG antibody Fc domain, and the immunoglobulin IgG antibody comprises one or more selected from human IgG1, human IgG2, human IgG3, human IgG4, mouse IgG1, mouse IgG2a, mouse IgG2b, and mouse IgG3.

144. 144. The fusion polypeptide of any one of claims 131 to 143, wherein the fusion polypeptide comprises an amino acid sequence selected from SEQ ID NO: 350 to SEQ ID NO: 382, ​​SEQ ID NO: 384 to SEQ ID NO: 416, SEQ ID NO: 618 to SEQ ID NO: 624 and SEQ ID NO: 626 to SEQ ID NO:

632.

145. 145. The fusion polypeptide of any one of claims 131 to 144, wherein the fusion polypeptide further comprises a third domain, the third domain being the same as or different from the second domain, an antibody or antigen-binding fragment thereof, or a functional protein or active fragment thereof.

146. 146. The fusion polypeptide of any one of claims 131 to 145, wherein the third domain is a functional protein or an active fragment thereof, and wherein the functional protein or an active fragment thereof is capable of activating an innate immune response.

147. The fusion polypeptide of any one of claims 131 to 146, wherein the third domain is capable of binding to an MHCII molecule on an antigen-presenting cell.

148. 148. The fusion polypeptide of any one of claims 131 to 147, wherein the third domain comprises LAG3 or a functionally active fragment thereof.

149. A LAG3 mutant polypeptide, wherein said LAG3 mutant polypeptide comprises truncations and / or mutations based on human-derived LAG3.

150. The fusion polypeptide or LAG3 variant polypeptide of claim 149, wherein the LAG3 variant polypeptide comprises a truncation based on a wild-type human-derived LAG3 extracellular domain polypeptide.

151. The LAG3 mutant polypeptide of any one of claims 149 to 150, wherein the N-terminus of the LAG3 mutant polypeptide is shortened by 0 to 124 amino acids based on the wild-type human-derived LAG3 extracellular domain polypeptide, and the C-terminus of the LAG3 mutant polypeptide terminates between amino acid positions 121 and 167 based on the wild-type human-derived LAG3 extracellular domain polypeptide.

152. The LAG3 mutant polypeptide of any one of claims 149 to 151, wherein the LAG3 mutant polypeptide comprises an N-terminal truncation of 0, 5, 21, 37, 45, 60, 71, 74, 79, 84, 89, 94, 99, 104, 109, or 114 amino acids based on the wild-type human-derived LAG3 extracellular domain polypeptide.

153. 153. The LAG3 mutant polypeptide of any one of claims 149 to 152, wherein the LAG3 mutant polypeptide comprises an N-terminal truncation of 0, 5, 21, 37, 45, 60, 71, 74, 81, or 99 amino acids based on the wild-type human-derived LAG3 extracellular domain polypeptide.

154. The LAG3 mutant polypeptide of any one of claims 149 to 153, wherein the LAG3 mutant polypeptide comprises an N-terminal truncation of 0, 5, 21, 37, 45, 60, 71, or 74 amino acids based on the wild-type human-derived LAG3 extracellular domain polypeptide.

155. The LAG3 mutant polypeptide according to any one of claims 149 to 154, wherein the C-terminus of the LAG3 mutant polypeptide terminates at amino acid position 121, 122, 129, 136, 146, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 or 167 based on the wild-type human-derived LAG3 extracellular domain polypeptide.

156. The LAG3 mutant polypeptide of any one of claims 149 to 155, wherein the C-terminus of the LAG3 mutant polypeptide terminates at amino acid position 121, 122, 129, 136, 146, 156, 161 or 167 based on the wild-type human-derived LAG3 extracellular domain polypeptide.

157. The LAG3 mutant polypeptide according to any one of claims 149 to 156, wherein the C-terminus of the LAG3 mutant polypeptide terminates at amino acid position 121, 146, 156, 161 or 167 based on the wild-type human-derived LAG3 extracellular domain polypeptide.

158. The LAG3 mutant polypeptide of any one of claims 149 to 157, wherein the C-terminus of the LAG3 mutant polypeptide terminates at amino acid position 156, 161 or 167 based on the wild-type human-derived LAG3 extracellular domain polypeptide.

159. 159. The LAG3 mutant polypeptide of any one of claims 149 to 158, wherein the LAG3 mutant polypeptide comprises an N-terminal truncation of 0, 5, 21, 37, 45, 60, 71, 74, 81, or 99 amino acids based on the wild-type human-derived LAG3 extracellular domain polypeptide, and terminates at amino acid position 121, 122, 129, 136, 146, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, or 167 at the C-terminus.

160. 160. The LAG3 mutant polypeptide of any one of claims 149 to 159, wherein the LAG3 mutant polypeptide comprises an N-terminal truncation of 0, 5, 21, 37, 45, 60, 71, or 74 amino acids based on the wild-type human-derived LAG3 extracellular domain polypeptide, and the C-terminal ends at amino acid position 122, 129, 136, 146, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, or 167.

161. 161. The LAG3 mutant polypeptide of any one of claims 149 to 160, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site.

162. 162. The LAG3 mutant polypeptide of any one of claims 149 to 161, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site, and the amino acid mutation site is an Arg amino acid at position 97.

163. 163. The LAG3 mutant polypeptide of any one of claims 149 to 162, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site, and the amino acid mutation site is a mutation from an Arg amino acid at position 97 to a Glu amino acid.

164. 164. The LAG3 mutant polypeptide of any one of claims 149 to 163, wherein the LAG3 mutant polypeptide is selected from the amino acid sequences set forth in SEQ ID NO: 244 to SEQ ID NO:

349.

165. 165. The LAG3 mutant polypeptide of any one of claims 149 to 164, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site, and the amino acid mutation site is R110, R113, R119, R129, G130 and / or R141.

166. 166. The LAG3 mutant polypeptide of any one of claims 149 to 165, wherein the LAG3 mutant polypeptide is truncated by 81 amino acids at the N-terminus, terminates at amino acid position 121 at the C-terminus, and further contains mutations at R110, R113, and / or R119.

167. 166. The LAG3 mutant polypeptide of any one of claims 149 to 165, wherein the LAG3 mutant polypeptide is truncated by 99 amino acids at the N-terminus, terminates at amino acid position 146 at the C-terminus, and further contains mutations at R129, G130, and / or R141.

168. 166. The LAG3 mutant polypeptide of any one of claims 149 to 165, wherein the LAG3 mutant polypeptide is truncated by 81 amino acids at the N-terminus, terminates at amino acid position 146 at the C-terminus, and further contains mutations at R110, R113, R119, R129, G130, and / or R141.

169. 169. The LAG3 mutant polypeptide of any one of claims 149 to 168, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site, wherein the R110 site is mutated to K110.

170. 170. The LAG3 mutant polypeptide of any one of claims 149 to 169, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site, wherein the R113 site is mutated to K113.

171. 171. The LAG3 mutant polypeptide of any one of claims 149 to 170, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site, and the R119 site is mutated to K119.

172. 172. The LAG3 mutant polypeptide of any one of claims 149 to 171, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site, and the R129 site is mutated to K129.

173. 173. The LAG3 mutant polypeptide of any one of claims 149 to 172, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site, and the G130 site is mutated to P130, A130, T130, Y130, or S130.

174. 174. The LAG3 mutant polypeptide of any one of claims 149 to 173, wherein the LAG3 mutant polypeptide comprises a mutation at an amino acid site, wherein the R141 site is mutated to K141.

175. 175. The LAG3 mutant polypeptide of any one of claims 149 to 174, wherein the amino acid sequence of the LAG3 mutant polypeptide is any one of SEQ ID NO: 1285 to SEQ ID NO: 1318.

176. 176. A fusion polypeptide comprising a first domain and a second domain, wherein the first domain comprises the LAG3 mutant polypeptide of any one of claims 149 to 175, and the second domain comprises an antibody or antigen-binding fragment thereof, or an immunoglobulin Fc domain.

177. 177. The fusion polypeptide of claim 176, wherein the second domain comprises an antibody or an antigen-binding fragment thereof.

178. The fusion polypeptide of any one of claims 176 to 177, wherein the antibody is selected from an immunoglobulin antibody, a recombinant antibody, a chimeric antibody, a heavy chain antibody, a single domain antibody or a bispecific antibody.

179. The fusion polypeptide of any one of claims 176 to 178, wherein the antigen-binding fragment is any one or more of a Fab, a Fab', an Fv fragment, F(ab')2, F(ab)2, a scFv, a di-scFv, a VHH, and a dAb.

180. the second domain is selected from the group consisting of PD-L1, PD-L2, PD-1, OX40, 4-1BB, ICOS, TIGIT, CTLA4, LAG3, CD3, VEGF, VEGFR, CD47, HGF, Trop2, EpCAM, CCR8, CCR4, CCR5, GPRC5D, BCMA, CD19, CD20, and HER-2 180. The fusion polypeptide of any one of claims 176 to 179, which is capable of binding to any one or more of neu, DLL1, HER-3, HER-4, EGFR, PSMA, CEA, MUC-1 (mucin), MUC2, MUC3, MUC4, MUC5AC, MUC5B, MUC7, CD123, CD33, CD30, CD38, NKG2A, Nkp36, and Tim3.

181. 181. The fusion polypeptide of any one of claims 176 to 180, wherein the second domain is capable of binding to PD-L1, PD-1 and / or PD-L2.

182. 182. The fusion polypeptide of any one of claims 176 to 181, wherein the second domain comprises HCDR1, HCDR2 and / or HCDR3 of an antibody heavy chain, preferably wherein the antibody heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:

17.

183. 183. The fusion polypeptide of any one of claims 176 to 182, wherein the second domain comprises the variable region (VH) of an antibody heavy chain, preferably wherein the antibody heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:

17.

184. 184. The fusion polypeptide of any one of claims 176 to 183, wherein the second domain comprises a heavy chain of an antibody, preferably wherein the heavy chain of the antibody comprises an amino acid sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:10, SEQ ID NO:12, SEQ ID NO:14, SEQ ID NO:15 and SEQ ID NO:

17.

185. The fusion polypeptide of any one of claims 176 to 184, wherein the second domain comprises LCDR1, LCDR2 and / or LCDR3 of an antibody light chain, preferably wherein the antibody light chain comprises an amino acid sequence selected from the group consisting of: SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:

18.

186. The fusion polypeptide of any one of claims 176 to 185, wherein the second domain comprises the variable region (VL) of an antibody light chain, preferably wherein the antibody light chain comprises an amino acid sequence selected from the group consisting of: SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:

18.

187. 187. The fusion polypeptide of any one of claims 176 to 186, wherein the second domain comprises a light chain of an antibody, and preferably the light chain of the antibody comprises an amino acid sequence selected from the group consisting of SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:16 and SEQ ID NO:

18.

188. The fusion polypeptide of any one of claims 176 to 187, wherein the second domain comprises an antibody or antigen-binding fragment thereof selected from the group consisting of Sugemalimab, Durvalumab, Atezolizumab, Avelumab, Envafolimab, Permbrolizumab, and Nivolumab.

189. The fusion polypeptide of any one of claims 176 to 188, wherein the second domain comprises sugemalimab or an antigen-binding fragment thereof.

190. The fusion polypeptide of any one of claims 176 to 189, wherein the fusion polypeptide comprises a heavy chain, and the amino acid sequence of the heavy chain is any one of SEQ ID NO: 1353 to SEQ ID NO: 1386.

191. 191. The fusion polypeptide of any one of claims 176 to 190, wherein the fusion polypeptide comprises a light chain, wherein the light chain comprises an antibody light chain polypeptide, and preferably the sequence of the light chain of the fusion polypeptide is selected from the group comprising SEQ ID NO:

5.

192. 192. The fusion polypeptide of any one of claims 176 to 191, wherein the heavy chain of the fusion polypeptide and the light chain of the fusion polypeptide associate to form the fusion polypeptide.

193. 193. The fusion polypeptide of any one of claims 176 to 192, wherein the first domain and the second domain are linked directly or indirectly.

194. The fusion polypeptide of any one of claims 176 to 193, wherein the first domain and the second domain are directly or indirectly linked at their N-termini.

195. 195. The fusion polypeptide of any one of claims 176 to 194, wherein the first domain and the second domain are directly or indirectly linked at their C-terminus.

196. 196. The fusion polypeptide of any one of claims 176 to 195, wherein the second domain comprises an antibody heavy chain, and the LAG3 mutant polypeptide is directly or indirectly linked to the antibody heavy chain of the second domain.

197. 197. The fusion polypeptide of any one of claims 176 to 196, wherein the second domain comprises an antibody heavy chain, and the LAG3 mutant polypeptide is linked directly or indirectly to the C-terminus of the antibody heavy chain of the second domain.

198. 198. The fusion polypeptide of any one of claims 176 to 197, wherein the second domain comprises an antibody heavy chain, and the N-terminus of the LAG3 mutant polypeptide is linked directly or indirectly to the C-terminus of the antibody heavy chain of the second domain.

199. 199. The fusion polypeptide of any one of claims 176 to 198, wherein the second domain comprises an antibody heavy chain, and the LAG3 mutant polypeptide is linked directly or indirectly to the N-terminus of the antibody heavy chain of the second domain.

200. 200. The fusion polypeptide of any one of claims 176 to 199, wherein the second domain comprises an antibody heavy chain, and the C-terminus of the LAG3 mutant polypeptide is linked directly or indirectly to the N-terminus of the antibody heavy chain of the second domain.

201. 201. The fusion polypeptide of any one of claims 176 to 200, wherein the second domain comprises an antibody light chain, and the LAG3 mutant polypeptide is directly or indirectly linked to the antibody light chain of the second domain.

202. 202. The fusion polypeptide of any one of claims 176 to 201, wherein the second domain comprises an antibody light chain, and the LAG3 mutant polypeptide is linked directly or indirectly to the C-terminus of the antibody light chain of the second domain.

203. 203. The fusion polypeptide of any one of claims 176-202, wherein the second domain comprises an antibody light chain, and the N-terminus of the LAG3 mutant polypeptide is linked directly or indirectly to the C-terminus of the antibody light chain of the second domain.

204. 204. The fusion polypeptide of any one of claims 176 to 203, wherein the second domain comprises an antibody light chain, and the LAG3 mutant polypeptide is linked directly or indirectly to the N-terminus of the antibody light chain of the second domain.

205. 205. The fusion polypeptide of any one of claims 176 to 204, wherein the second domain comprises an antibody light chain, and the C-terminus of the LAG3 mutant polypeptide is linked directly or indirectly to the N-terminus of the antibody light chain of the second domain.

206. The fusion polypeptide of any one of claims 176 to 205, wherein the indirect linkage comprises a linkage via a linker.

207. 207. The fusion polypeptide of any one of claims 176 to 206, wherein the second domain comprises an immunoglobulin Fc domain.

208. The fusion polypeptide of claim 207, wherein the immunoglobulin Fc domain comprises the Fc domain of an IgG antibody, and the immunoglobulin IgG antibody is selected from any one or more of human IgG1, human IgG2, human IgG3, human IgG4, mouse IgG1, mouse IgG2a, mouse IgG2b, and mouse IgG3.

209. The fusion polypeptide of any one of claims 176 to 208, wherein the second domain comprises an immunoglobulin Fc domain.

210. 210. The fusion polypeptide of any one of claims 176 to 209, wherein the second domain comprises the Fc domain of an immunoglobulin IgG antibody.

211. 211. The fusion polypeptide of any one of claims 176-210, wherein the second domain comprises an Fc domain of an immunoglobulin IgG antibody, wherein the Fc domain of the immunoglobulin IgG antibody comprises a human IgG1 Fc domain, a human IgG2 Fc domain, a human IgG3 Fc domain, a human IgG4 Fc domain, a mouse IgG1 Fc domain, a mouse IgG2a Fc domain, a mouse IgG2b Fc domain, or a mouse IgG3 Fc domain.

212. The fusion polypeptide of any one of claims 176 to 211, wherein the second domain comprises the Fc domain of human-derived immunoglobulin IgG4.

213. The fusion polypeptide of any one of claims 176 to 212, wherein the second domain comprises an amino acid sequence selected from the following amino acid sequences: SEQ ID NO: 4 to SEQ ID NO: 18, SEQ ID NO: 29 to SEQ ID NO: 30, SEQ ID NO: 634 to SEQ ID NO:

639.

214. 214. The fusion polypeptide of any one of claims 176 to 213, wherein the second domain comprises an amino acid sequence selected from the following amino acid sequences: SEQ ID NO: 1319 to SEQ ID NO: 1352.

215. 215. The fusion polypeptide of any one of claims 176 to 214, wherein the first domain is directly or indirectly linked to the second domain.

216. 216. The fusion polypeptide of any one of claims 176 to 215, wherein the first domain is linked directly or indirectly to the N-terminus of the second domain.

217. 217. The fusion polypeptide of any one of claims 176 to 216, wherein the first domain is linked directly or indirectly to the C-terminus of the second domain.

218. The fusion polypeptide of any one of claims 176 to 217, wherein the indirect linkage comprises a linkage via a linker.

219. The fusion polypeptide of claim 218, wherein the linker comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, and SEQ ID NO:

35.

220. The fusion polypeptide of any one of claims 176 to 219, wherein the fusion polypeptide comprises an amino acid sequence selected from SEQ ID NO: 640 to SEQ ID NO: 745, SEQ ID NO: 747 to SEQ ID NO: 750, SEQ ID NO: 1353 to SEQ ID NO: 1386.

221. The fusion polypeptide of any one of claims 176 to 220, further comprising a third domain, wherein the third domain is the same as or different from the second domain, an antibody, antigen-binding fragment, functional protein, or an active fragment thereof.

222. 222. The fusion polypeptide of claim 221, wherein the third domain is a functional protein or an active fragment thereof, and the functional protein or active fragment thereof is capable of activating an innate immune response.

223. 223. The fusion polypeptide of any one of claims 221 to 222, wherein the third domain is capable of binding to CD28 and / or CTLA4.

224. 224. The fusion polypeptide of any one of claims 221 to 223, wherein the third domain comprises CD80, CD86, or an active fragment thereof.

225. An immunoconjugate comprising a fusion polypeptide according to any one of claims 1 to 119, 131 to 148 and 176 to 224, a CD86 variant polypeptide according to any one of claims 120 to 130, and / or a LAG3 variant according to any one of claims 149 to 175.

226. A nucleic acid molecule encoding a fusion polypeptide according to any one of claims 1 to 119, 131 to 148 and 176 to 224, a CD86 variant polypeptide according to any one of claims 120 to 130, and / or a LAG3 variant according to any one of claims 149 to 175.

227. A vector comprising the nucleic acid molecule of claim 226.

228. A cell comprising and / or expressing a fusion polypeptide according to any one of claims 1 to 119, 131 to 148 and 176 to 224, a CD86 variant polypeptide according to any one of claims 120 to 130, a LAG3 variant according to any one of claims 149 to 175, an immunoconjugate according to claim 225, a nucleic acid molecule according to claim 226, a vector according to claim 227.

229. 228. A composition comprising a fusion polypeptide according to any one of claims 1 to 119, 131 to 148 and 176 to 224, a CD86 variant polypeptide according to any one of claims 120 to 130, a LAG3 variant according to any one of claims 149 to 175, an immunoconjugate according to claim 225, a nucleic acid molecule according to claim 226, a vector according to claim 227, and / or a cell according to claim 228, optionally further comprising a pharmaceutically acceptable carrier.

230. 228. A method for producing a fusion polypeptide according to any one of claims 1 to 119, 131 to 148 and 176 to 224, a CD86 variant polypeptide according to any one of claims 120 to 130, and a LAG3 variant according to any one of claims 149 to 175, the method comprising culturing a cell according to claim 228 under conditions allowing expression of said fusion polypeptide.

231. 226. A method for blocking the interaction of PD-L1 protein with PD-1, comprising administering an effective amount of a fusion polypeptide of any one of claims 1-119, 131-148, and 176-224.

232. A method for stimulating antigen-presenting cells and / or promoting T-cell activation, comprising a LAG3 variant according to any one of claims 149 to 175.

233. The method of claim 232, wherein the stimulation of the antigen-presenting cells includes one or more selected from the group consisting of increased expression of costimulatory molecules in the antigen-presenting cells, morphological changes and maturation of the antigen-presenting cells, increased secretion of chemokines from the antigen-presenting cells, and improved phagocytic ability of the antigen-presenting cells.

234. 228. A method of inhibiting the growth and / or proliferation of a tumor or tumor cells, said method comprising administering an effective amount of a fusion polypeptide of any one of claims 1 to 119, 131 to 148 and 176 to 224, a CD86 variant polypeptide of any one of claims 120 to 130, a LAG3 variant of any one of claims 149 to 175, an immunoconjugate of claim 225, a nucleic acid molecule of claim 226, a vector of claim 227, a cell of claim 228, and / or a composition of claim 229.

235. 229. Use of a fusion polypeptide according to any one of claims 1 to 119, 131 to 148 and 176 to 224, a CD86 mutant polypeptide according to any one of claims 120 to 130, a LAG3 mutant according to any one of claims 149 to 175, an immunoconjugate according to claim 225, a nucleic acid molecule according to claim 226, a vector according to claim 227, a cell according to claim 228 and / or a composition according to claim 229 in the manufacture of a medicament, wherein said medicament is used for the prevention, amelioration and / or treatment of a tumor.

236. 236. The use of claim 235, wherein the tumor comprises a solid tumor and / or a hematological tumor.

237. 237. The use of any one of claims 235 to 236, wherein the tumor comprises a tumor selected from the group consisting of colon tumor, breast tumor, lung tumor, gastric tumor, melanoma, head and neck tumor, lymphoma, nasopharyngeal tumor, cervical tumor, esophageal tumor, kidney tumor, cutaneous squamous cell carcinoma, endometrial tumor, liver tumor, bladder tumor, urothelial tumor and skin tumor.

Images