Anti-FGFR2b antibody-drug conjugate and use thereof

Abstract

Disclosed in the present invention are an anti-FGFR2b antibody-drug conjugate and the use thereof in the prevention and / or treatment of tumor diseases. The anti-FGFR2b antibody-drug conjugate of the present invention can effectively inhibit the growth of tumor tissue, and has a low toxicity and good safety.

Description

Anti-FGFR2b antibody-drug conjugates and their applications Technical Field

[0001] The present invention relates to an anti-FGFR2b antibody-drug conjugate, and also relates to the use of the anti-FGFR2b antibody-drug conjugate in preventing and / or treating tumor diseases, including but not limited to the use in preventing and / or treating tumor diseases.

[0002] Cross-references

[0003] This application claims priority to and the benefits of international application number "PCT / CN2024 / 085216" filed on April 1, 2024, and the entire contents of the above PCT application are incorporated herein by reference.

[0004] Sequence Listing

[0005] The present invention also includes a sequence listing submitted electronically along with the present application, which is titled “sequence listing anti-FGFR2b”, and the contents of the sequence listing are incorporated herein by reference. Background Art

[0006] According to global cancer statistics in 2020, the global incidence and deaths of lung cancer were 2.2 million and 820,000 respectively, accounting for 11.4% of the total cancer cases. Its incidence rate ranked second among cancers, but its mortality rate ranked first. [1] In China, the number of lung cancer cases and deaths in 2022 was approximately 870,000 and 7.76 million respectively, ranking first among cancers in both incidence and mortality. 2 ]. Non-small cell lung cancer accounts for approximately 85% of lung cancer cases. According to histological classification, it can be divided into ~40% adenocarcinoma, ~30% squamous cell carcinoma, ~10% large cell carcinoma and other rare types of primary lung cancer. Squamous non-small cell lung cancer (sqNSCLC) has very unique driver gene mutation types that are different from lung adenocarcinoma, including 20% ​​and 9% gene amplification of FGFR1 and PDGFRA, respectively, and the gene mutation rates of PIK3CA, PTEN, EGFRvIII, DDR2, and FGFR2 are 14%, 18%, 5%, 4% and 3%, respectively. These mutation types are very low or relatively rare in adenocarcinoma. [3,4,5] For systemic treatment of advanced or metastatic sqNSCLC, priority is given to targeted therapy for driver gene mutations, such as EGFRmut, BRAF V600E, Metexon14, and KRAS G12C. However, <10% of patients with sqNSCLC have driver gene mutations, while >90% of patients receive PD-1 or PD-L1 monotherapy or combination chemotherapy as first-line standard treatment. [6,7]However, for patients with first-line drug resistance, especially those with PD-L1+ expression, there is currently no effective second- or third-line standard treatment. The 5-year overall survival rate for all stages of squamous cell lung cancer is only less than 15%. Therefore, effective targeted therapies for sqNSCLC are urgently needed to address the huge unmet clinical need.

[0007] Gastric cancer is one of the most common cancers in the world, with 1.09 million cases and 770,000 deaths worldwide, ranking fourth in both incidence and mortality. [1] . Among them, the incidence of gastric cancer is higher in East Asia, Eastern Europe and South America, and lower in North America and Africa. Although the prognosis of gastric cancer patients has been significantly improved due to the development of gastric cancer surgical techniques and perioperative treatment, the 5-year overall survival rate of gastric cancer at all stages is only 10-15%. Targeted therapy targeting HER2 has brought new hope to patients with advanced recurrence and metastasis of gastric cancer, but only 10%-15% of patients have HER2 positive expression, and the beneficiary population is limited. For the HER2-negative patient population, the first-line standard treatment is chemotherapy, or PD-1 targeted therapy is applicable to the PD-L1-positive population. However, the applicable PD-L1-positive patient population only accounts for about 30%-40% of patients with advanced gastric cancer. And for HER2-negative patients who are resistant to the first-line treatment, there is no more effective targeted drug in the second line. [8,9] Therefore, there is an urgent need to develop targeted drugs that can achieve clinical breakthroughs in gastric cancer.

[0008] FGFR2b is a highly promising innovative target for solid tumor immunotherapy. It is overexpressed in a variety of solid tumors, such as squamous non-small cell lung cancer, gastric cancer, gastroesophageal junction cancer, triple-negative breast cancer, ovarian cancer, endometrial cancer, cervical cancer, colorectal cancer, intrahepatic bile duct carcinoma, and pancreatic cancer. Antibody-drug conjugates targeting FGFR2b address the lack of targeted therapy for solid tumor indications such as sqNSCLC, gastric cancer, and gastroesophageal junction cancer, as well as the significant unmet clinical need. Summary of the Invention

[0009] The inventors of the present application prepared an anti-FGFR2b antibody-drug conjugate through extensive experiments and creative work, and confirmed that it has good biological activity, thereby completing the present invention.

[0010] To this end, in the first aspect of the present invention, the present invention provides an antibody drug conjugate, a pharmaceutically acceptable salt, solvate or solvate of the salt thereof, wherein the antibody drug conjugate has the structure shown in Formula I,

[0011] Ab-(LD)p

[0012] Formula I

[0013] in:

[0014] Ab is an anti-FGFR2b antibody, the anti-FGFR2b antibody comprises a heavy chain and a light chain, the heavy chain variable complementary determining region 1 (CDR1) comprises a sequence selected from SEQ ID NO: 59, 65, 71, 77, 83, 88, 93, 103, 110, 119, 135, 143, 146, 150 or 152 or a mutant thereof, the heavy chain variable complementary determining region 2 (CDR2) comprises a sequence selected from SEQ ID NO: 60, 66, 72, 78, 84, 94, 111, 118, 120, 124, 141 or 153 or a mutant thereof, and the heavy chain variable complementary determining region 3 (CDR3) comprises a sequence selected from SEQ ID NO: NO: 61, 67, 73, 79, 85, 89, 95, 101, 104, 112, 116, 121, 125, 128, 131, 136, 138, 142, 144, 147, 151, 154 or 158 or a mutant thereof; the light chain variable complementary determining region 1 (CDR1) comprises a sequence selected from SEQ ID NO: 62, 68, 74, 80, 86, 90, 96, 99, 105, 108, 113, 122, 126, 129, 132, 148, 149 or 155 or a mutant thereof, the light chain variable complementary determining region 2 (CDR2) comprises a sequence selected from SEQ ID NO: NO: 63, 69, 75, 81, 91, 97, 106, 114, 133, 139 or 156, or a mutant thereof, and the light chain variable complementarity determining region 3 (CDR3) comprises a sequence selected from SEQ ID NO: 64, 70, 76, 82, 87, 92, 98, 100, 102, 107, 109, 115, 117, 123, 127, 130, 134, 137, 140, 145, 157 or 159, or a mutant thereof;

[0015] D is a cytotoxic agent;

[0016] L is a linker used to connect the anti-FGFR2b antibody and the cytotoxic agent;

[0017] p is 2.0-8.0 (e.g., 2.0-8.0, 2.0-7.0, 2.0-6.0, 2.0-5.0, 2.0-4.0, 3.0-8.0, 3.0-7.0, 3.0-6.0, 3.0-5.0, or 3.0-4.0, or for example, 2.0, 3.0, 4.0, 5.0, 6.0, 7.0, or 8.0.

[0018] The antibody-drug conjugate of the present invention has excellent tumor cell growth inhibition activity both in vivo and in vitro and has good application prospects. The antibody-drug conjugate of the present invention is composed of an anti-FGFR2b antibody and a derivative of dolastatin toxin, MMAE, connected by an MC-vc-PAB linker. Its anti-tumor mechanism of action is that the anti-FGFR2b antibody recognizes tumor surface antigens to form an antigen-ADC complex, which is internalized into the cell to form endosomes. After maturation, the late endosomes fuse with lysosomes. Due to the abundant proteolytic enzymes and acidic environment in the lysosomes, the linker further releases the drug MMAE. Free MMAE binds to microtubules and inhibits their polymerization, thereby blocking cellular functions involved in microtubules, including mitosis, and inhibiting tumor cell proliferation and death. In addition, some free MMAE crosses the cell membrane and kills adjacent tumor cells. This process is independent of whether the tumor cells express the antigen, i.e., a bystander effect.

[0019] It should be noted that the "antibody drug conjugate" is a composition containing ADC molecules with the same or different conjugated toxin to antibody molar ratios (DAR values). Specifically, the present invention provides compositions comprising multiple ADC molecules. In some cases, the multiple ADCs in the composition each contain the same number of drug molecules. In other cases, the multiple ADCs in the composition each contain different numbers of drug molecules.

[0020] The drug-antibody ratio (DAR) refers to the number of drug molecules conjugated to the antibody (e.g., p in Formula I). ​​The number of drug molecules contained in the antibody-drug conjugates of the present invention (e.g., p in Formula I) is generally an integer. When the number of drug molecules contained in the antibody-drug conjugates of the present invention (e.g., p in Formula I) is a fraction, the fraction refers to the average number of drug molecules conjugated to each antibody in a composition comprising multiple ADC molecules.

[0021] The drug-to-antibody ratio (DAR) can be verified by conventional means, such as mass spectrometry, ELISA assays, HIC-HPLC, etc. The quantitative distribution of the ADC in terms of p can also be determined. In some cases, separation, purification, and verification of homogeneous ADCs with a certain p value from ADCs with other drug loadings can be achieved by means such as reverse phase HPLC or electrophoresis.

[0022] In some embodiments, the heavy chain variable complementary determining region 1 (CDR1) of the anti-FGFR2b antibody comprises a sequence selected from SEQ ID NO: 59, 65, 71, 77, 83, 88, 93, 103, 110, 119, 135, 143, 146, 150, or 152, or a mutant thereof, the heavy chain variable complementary determining region 2 (CDR2) comprises a sequence selected from SEQ ID NO: 60, 66, 72, 78, 84, 94, 111, 118, 120, 124, 141, or 153, or a mutant thereof, and the heavy chain variable complementary determining region 3 (CDR3) comprises a sequence selected from SEQ ID NO: 61, 62, 64, 65, 6 ... The sequence shown in NO: 61, 67, 73, 79, 85, 89, 95, 101, 104, 112, 116, 121, 125, 128, 131, 136, 138, 142, 144, 147, 151, 154 or 158, or a mutant thereof.

[0023] In some embodiments, the heavy chain variable complementary determining region 1 (CDR1), heavy chain variable complementary determining region 2 (CDR2), and heavy chain variable complementary determining region 3 (CDR3) of the anti-FGFR2b antibody are selected from the sequence combinations shown in Table 1 below:

[0024] Table 1:

[0025] In some embodiments, the anti-FGFR2b antibody comprises a light chain variable complementary determining region 1 (CDR1) selected from the group consisting of SEQ ID NO: 62, 68, 74, 80, 86, 90, 96, 99, 105, 108, 113, 122, 126, 129, 132, 148, 149 or 155, or a mutant thereof, a light chain variable complementary determining region 2 (CDR2) selected from the group consisting of SEQ ID NO: 63, 69, 75, 81, 91, 97, 106, 114, 133, 139 or 156, or a mutant thereof, and a light chain variable complementary determining region 3 (CDR3) selected from the group consisting of SEQ ID NO: The sequence shown in NO: 64, 70, 76, 82, 87, 92, 98, 100, 102, 107, 109, 115, 117, 123, 127, 130, 134, 137, 140, 145, 157 or 159, or a mutant thereof.

[0026] In some embodiments, the light chain variable complementarity determining region 1 (CDR1), variable complementarity determining region 2 (CDR2), and variable complementarity determining region 3 (CDR3) of the anti-FGFR2b antibody are selected from the sequence combinations shown in Table 2 below:

[0027] Table 2:

[0028] In some embodiments, the anti-FGFR2b antibody comprises a heavy chain variable complementary determining region 1 (CDR1) selected from the sequence shown in SEQ ID NO: 59, 65, 71, 77, 83, 88, 93, 103, 110, 119, 135, 143, 146, 150 or 152 or a mutant thereof, a heavy chain variable complementary determining region 2 (CDR2) selected from the sequence shown in SEQ ID NO: 60, 66, 72, 78, 84, 94, 111, 118, 120, 124, 141 or 153 or a mutant thereof, and a heavy chain variable complementary determining region 3 (CDR3) selected from the sequence shown in SEQ ID NO: NO: 61, 67, 73, 79, 85, 89, 95, 101, 104, 112, 116, 121, 125, 128, 131, 136, 138, 142, 144, 147, 151, 154 or 158 or a mutant thereof; the light chain variable complementary determining region 1 (CDR1) of the anti-FGFR2b antibody comprises a sequence selected from SEQ ID NO: 62, 68, 74, 80, 86, 90, 96, 99, 105, 108, 113, 122, 126, 129, 132, 148, 149 or 155 or a mutant thereof, and the light chain variable complementary determining region 2 (CDR2) comprises a sequence selected from SEQ ID NO: NO: 63, 69, 75, 81, 91, 97, 106, 114, 133, 139 or 156 or a mutant thereof, and the light chain variable complementary determining region 3 (CDR3) comprises a sequence selected from SEQ ID NO: 64, 70, 76, 82, 87, 92, 98, 100, 102, 107, 109, 115, 117, 123, 127, 130, 134, 137, 140, 145, 157 or 159 or a mutant thereof.

[0029] In some embodiments, the heavy chain variable complementary determining region 1 (CDR1), heavy chain variable complementary determining region 2 (CDR2), and heavy chain variable complementary determining region 3 (CDR3) of the anti-FGFR2b antibody are selected from the sequence combinations shown in Table 1; the light chain variable complementary determining region 1 (CDR1), variable complementary determining region 2 (CDR2), and variable complementary determining region 3 (CDR3) of the anti-FGFR2b antibody are selected from the sequence combinations shown in Table 2.

[0030] In some embodiments, the heavy chain variable region framework region 1 (FR1) of the anti-FGFR2b antibody comprises a sequence selected from SEQ ID NO: 160, 168, 175, 188, 194, 204, 210, 224, 233, 284, 259, 265, 268, 271, 274, 281, 295 or 305 or a mutant thereof, the heavy chain variable region framework region 2 (FR2) comprises a sequence selected from SEQ ID NO: 161, 169, 176, 183, 205, 221, 225, 237, 245, 251, 261, 266, 275, 282 or 296 or a mutant thereof, and the heavy chain variable region framework region 3 (FR3) comprises a sequence selected from SEQ ID NO: NO: 162, 170, 177, 184, 189, 195, 200, 206, 211, 216, 226, 231, 234, 239, 242, 246, 252, 254, 260, 262, 267, 272, 276, 297 or 306, or a mutant thereof, and the heavy chain variable region framework region 4 (FR4) comprises a sequence selected from SEQ ID NO: 163, 178, 190, 196, 212, 217, 227, 255, 269 and 298, or a mutant thereof.

[0031] In some embodiments, the light chain variable framework region FR1 of the anti-FGFR2b antibody comprises a sequence selected from SEQ ID NO: 164, 171, 179, 185, 191, 197, 201, 207, 213, 218, 222, 228, 232, 247, 256, 263, 270, 277, 283, 299 or 303, or a mutant thereof; the light chain variable framework region FR2 comprises a sequence selected from SEQ ID NO: 165, 172, 180, 186, 192, 198, 202, 208, 214, 219, 223, 229, 235, 240, 243, 248, 257, 264, 278, 300 or 304, or a mutant thereof; and the light chain variable framework region FR3 comprises a sequence selected from SEQ ID NO: NO: 166, 173, 181, 187, 193, 199, 203, 209, 220, 230, 236, 238, 241, 244, 249, 253, 258, 273, 279, 301, 316 and 317 or a mutant thereof, and the light chain variable framework region FR4 comprises a sequence selected from SEQ ID NO: 167, 174, 182, 215, 250, 280 or 302 or a mutant thereof.

[0032] In some embodiments, the anti-FGFR2 heavy chain variable framework region FR1, heavy chain variable framework region FR2, heavy chain variable framework region FR3 and heavy chain variable framework region FR4 are selected from the sequence combinations shown in Table 3 below:

[0033] Table 3:

[0034] In some embodiments, the heavy chain variable region framework region 1 (FR1) of the anti-FGFR2b antibody comprises a sequence selected from SEQ ID NO: 160, 168, 175, 188, 194, 204, 210, 224, 233, 284, 259, 265, 268, 271, 274, 281, 295 or 305 or a mutant thereof, the heavy chain variable region framework region 2 (FR2) comprises a sequence selected from SEQ ID NO: 161, 169, 176, 183, 205, 221, 225, 237, 245, 251, 261, 266, 275, 282 or 296 or a mutant thereof, and the heavy chain variable region framework region 3 (FR3) comprises a sequence selected from SEQ ID NO: NO: 162, 170, 177, 184, 189, 195, 200, 206, 211, 216, 226, 231, 234, 239, 242, 246, 252, 254, 260, 262, 267, 272, 276, 297 or 306, or a mutant thereof; the heavy chain variable region framework region 4 (FR4) comprises a sequence selected from the group consisting of SEQ ID NO: 163, 178, 190, 196, 212, 217, 227, 255, 269 and 298, or a mutant thereof; the light chain variable framework region FR1 of the anti-FGFR2b antibody comprises a sequence selected from the group consisting of SEQ ID NO: NO: 164, 171, 179, 185, 191, 197, 201, 207, 213, 218, 222, 228, 232, 247, 256, 263, 270, 277, 283, 299 or 303 or a mutant thereof, the light chain variable framework region FR2 comprises a sequence selected from SEQ ID NO: 165, 172, 180, 186, 192, 198, 202, 208, 214, 219, 223, 229, 235, 240, 243, 248, 257, 264, 278, 300 or 304 or a mutant thereof, and the light chain variable framework region FR3 comprises a sequence selected from SEQ ID NO: 1 NO: 166, 173, 181, 187, 193, 199, 203, 209, 220, 230, 236, 238, 241, 244, 249, 253, 258, 273, 279, 301, 316 and 317 or a mutant thereof, and the light chain variable framework region FR4 comprises a sequence selected from SEQ ID NO: 167, 174, 182, 215, 250, 280 or 302 or a mutant thereof.

[0035] In some embodiments, the anti-FGFR2 light chain variable framework region FR1, light chain variable framework region FR2, light chain variable framework region FR3 and light chain variable framework region FR4 are selected from the sequence combinations shown in Table 4 below:

[0036] Table 4:

[0037] In some embodiments, the heavy chain FR1, FR2, FR3, and FR4 of the anti-FGFR2b antibody are selected from the sequence combinations shown in Table 3; the light chain variable regions FR1, FR2, FR3, and FR4 of the anti-FGFR2b antibody are selected from the sequence combinations shown in Table 4.

[0038] In some embodiments, the complementarity determining regions and framework regions of the heavy chain variable region of the antibody are arranged in the following order from the N-terminus to the C-terminus of the sequence: heavy chain framework region 1-heavy chain complementarity determining region 1-heavy chain framework region 2-heavy chain complementarity determining region 2-heavy chain framework region 3-heavy chain complementarity determining region 3-heavy chain framework region 4.

[0039] In some embodiments, the heavy chain of the antibody comprises three constant regions: constant region 1, constant region 2, and constant region 3, consisting of SEQ ID NO: 307, SEQ ID NO: 309, and SEQ ID NO: 310, or consisting of SEQ ID NO: 307, SEQ ID NO: 312, and SEQ ID NO: 313.

[0040] In some embodiments, the heavy chain variable region of the anti-FGFR2b antibody is selected from SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 285, 287, 290, 291 or a homologous sequence thereof, wherein the homologous sequence is a sequence identical to SEQ ID NO: NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 285, 287, 290 or 291 has at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence homology.

[0041] In some embodiments, the heavy chain variable region of the anti-FGFR2b antibody is selected from the sequence shown in SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 285, 287, 290 or 291.

[0042] In some embodiments, the complementarity determining regions and framework regions of the light chain variable region of the antibody are arranged in the following order from the N-terminus to the C-terminus of the sequence: light chain framework region 1-light chain complementarity determining region 1-light chain framework region 2-light chain complementarity determining region 2-light chain framework region 3-light chain complementarity determining region 3-light chain framework region 4.

[0043] In some embodiments, the antibody further comprises a light chain constant region of SEQ ID NO:311.

[0044] In some embodiments, the light chain variable region of the antibody comprises SEQ ID Nos: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 46, 48, 50, 54, 56, 58, 286, 288, 289, 314, 315 or a homologous sequence thereof, wherein the homologous sequence is a sequence identical to SEQ ID Nos: Nos: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 46, 48, 50, 54, 56, 58, 286, 288, 289, 314 or 315 have at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence homology.

[0045] In some embodiments, the light chain variable region of the antibody comprises the sequence shown in SEQ ID Nos: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 46, 48, 50, 54, 56, 58, 286, 288, 289, 314 or 315.

[0046] In some embodiments, the heavy chain variable region and the light chain variable region of the anti-FGFR2b antibody comprise a combination shown in Table 5 below, or comprise a sequence that is at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% homologous to the following sequences, or a combination thereof:

[0047] Table 5:

[0048] In some embodiments, the heavy chain variable region and light chain variable region of the anti-FGFR2b antibody comprise a combination shown in Table 5.

[0049] In some embodiments, the sequences of the heavy chain variable region and the light chain variable region of the anti-FGFR2b antibody are SEQ ID NOs. 285 and 286, respectively.

[0050] In some embodiments, the heavy chain constant region of the anti-FGFR2b antibody is selected from human IgG (such as IgG1, IgG2, IgG3 or IgG4), IgM, IgA, IgD, IgA constant region or mutants of the above constant regions, preferably human IgG1;

[0051] The light chain constant region of the anti-FGFR2b antibody is selected from a human lambda constant region, a kappa constant region or mutants of the above constant regions, preferably a human kappa constant region.

[0052] In some embodiments, the amino acid sequence of the heavy chain of the anti-FGFR2b antibody comprises the sequence shown in SEQ ID NO: 292, or a sequence that is greater than 70%, e.g., greater than 75%, 80%, 85%, 90%, 95%, or 99% identical to the sequence shown in SEQ ID NO: 292;

[0053] The amino acid sequence of the light chain of the anti-FGFR2b antibody comprises the sequence shown in SEQ ID NO: 293, or comprises a sequence that is greater than 70%, such as greater than 75%, 80%, 85%, 90%, 95% or 99% identical to the sequence shown in SEQ ID NO: 293.

[0054] In some embodiments, p is 3.0-5.0.

[0055] In some embodiments, p is 3.5-4.5.

[0056] In some embodiments, p is 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, or 4.5.

[0057] In some embodiments, p is 4.0.

[0058] In some embodiments, the cytotoxic agent is selected from irinotecan, irinotecan derivatives, SN-38, Dxd, gemcitabine, Monomethyl auristatin E (MMAE), Monomethyl auristatin F (MMAF), maytansinoids (e.g., Maytansine DM1, Maytansine DM4), calicheamicin, MGBA (e.g., duocarmycin), doxorubicin, ricin, diphtheria toxin and other toxins, I131, interleukins, radionuclides, tumor necrosis factor, chemokines and nanoparticles.

[0059] In some embodiments, the cytotoxic agent is MMAE.

[0060] The structure of MMAE is:

[0061] In some embodiments, the linker is selected from 6-maleimidocaproyl (MC), maleimidopropionyl (MP), N-succinimidyl 4-(2-pyridylthio) pentanoate (SPP), 4-(N-maleimidomethyl)-cyclohexane-1-carboxyl (MCC), N-succinimidyl (4-iodo-acetyl) aminobenzoate (SIAB), and 6-maleimidocaproyl-valine-citrulline-p-aminobenzyloxycarbonyl (MC-vc-PAB).

[0062] In some embodiments, the linker is 6-maleimidocaproyl-valine-citrulline-p-aminobenzyloxycarbonyl (MC-vc-PAB).

[0063] In some embodiments, the LD described in Formula I is MC-vc-PAB-MMAE, which has the following structure:

[0064] In some embodiments,

[0065] Ab includes:

[0066] (a) heavy chain variable complementarity determining region 1 (CDR1), CDR2, CDR3, and light chain variable complementarity determining region 1 (CDR1), CDR2, CDR3, wherein the sequence of the heavy chain variable region CDR1 is set forth in SEQ ID NO: 59, the sequence of the heavy chain variable complementarity determining region 2 (CDR2) is set forth in SEQ ID NO: 60, and the sequence of the heavy chain variable complementarity determining region 3 (CDR3) is set forth in SEQ ID NO: 61, the sequence of the light chain variable complementarity determining region 1 (CDR1) is set forth in SEQ ID NO: 62, the sequence of the light chain variable complementarity determining region 2 (CDR2) is set forth in SEQ ID NO: 63, and the sequence of the light chain variable complementarity determining region 3 (CDR3) is set forth in SEQ ID NO: 64;

[0067] (b) a heavy chain variable region and a light chain variable region, wherein the sequence of the heavy chain variable region is shown in SEQ ID NO: 285, and the sequence of the light chain variable region is shown in SEQ ID NO: 286; and / or

[0068] (c) a heavy chain and a light chain, wherein the sequence of the heavy chain is shown in SEQ ID NO: 292 and the sequence of the light chain is shown in SEQ ID NO: 293;

[0069] L is MC-vc-PAB; and

[0070] D is MMAE.

[0071] In the second aspect of the present invention, the present invention provides a composition comprising the aforementioned antibody-drug conjugate, a pharmaceutically acceptable salt, solvate or solvate of the salt thereof.

[0072] In some embodiments, the composition further comprises a known chemotherapy drug for treating tumors, such as adriamycin, cyclophosphamide, taxanes (such as paclitaxel (Taxol), docetaxel (Taxotere), capecitabine (Xeloda), gemcitabine (Gemzar), vinorelbine (Navelbine), tamoxifen, aromatase inhibitors (Arimidex, Fulon, Aromasin), 5-FU plus folinic acid, irinotecan (camptosar), oxaliplatin, cisplatin, carboplatin, estramustine, mitoxantrone (Novantrone), prednisone, vincristine (Oncovin), doxorubicin, prednisone, etc., or a combination thereof.

[0073] In some embodiments, the composition further comprises known immunotherapeutic drugs for treating tumors, such as PD-1 monoclonal antibodies (e.g., pembrolizumab, nivolumab), PD-L1 monoclonal antibodies (e.g., atezolizumab), TIGIT monoclonal antibodies, VEGF-A monoclonal antibodies (e.g., bevacizumab), VEGFR2 monoclonal antibodies (e.g., ramucirumab, apatinib), HER2 targeted drugs (e.g., trastuzumab, DS-8201), etc., or combinations thereof.

[0074] In some embodiments, the composition further comprises an immunosuppressant selected from the group consisting of: (1) glucocorticoids, such as cortisone and prednisone; (2) microbial metabolites, such as cyclosporine and tacrolimus; (3) antimetabolites, such as azathioprine and 6-mercaptopurine; (4) polyclonal and monoclonal anti-inflammatory drugs. Anti-lymphocyte antibodies, such as anti-lymphocyte globulin and OKT3; (5) Alkylating agents, such as cyclophosphamide. Specifically, the immunosuppressants include methylprednisolone, prednisone, azathioprine, prograft, nipa, suleburell, cyclosporine, tacrolimus, rapamycin, mycophenolate mofetil, mizoribine, cyclophosphamide, fingolimod, etc.

[0075] In some embodiments, the composition further comprises a pharmaceutically acceptable carrier, diluent, or excipient.

[0076] In the third aspect of the present invention, the present invention provides the aforementioned antibody-drug conjugate, its pharmaceutically acceptable salt, solvate or solvate of the salt, or the use of the aforementioned composition in the preparation of a medicament for preventing and / or treating diseases associated with FGFR2b.

[0077] In some embodiments, the disease associated with FGFR2b is squamous non-small cell lung cancer, gastric cancer, gastroesophageal cancer, triple-negative breast cancer, ovarian cancer, endometrial cancer, cervical cancer, colorectal cancer, intrahepatic bile duct cancer and / or pancreatic cancer.

[0078] In some embodiments, the disease associated with FGFR2b is squamous non-small cell lung cancer, gastric cancer and / or gastroesophageal junction cancer.

[0079] In the fourth aspect of the present invention, the present invention provides a method for preventing and / or treating a disease associated with FGFR2b, comprising: administering to a subject in need thereof a preventive and / or therapeutically effective amount of the aforementioned antibody-drug conjugate, a pharmaceutically acceptable salt, solvate or solvate of the salt, or the aforementioned composition.

[0080] In some embodiments, the disease associated with FGFR2b is squamous non-small cell lung cancer, gastric cancer, gastroesophageal cancer, triple-negative breast cancer, ovarian cancer, endometrial cancer, cervical cancer, colorectal cancer, intrahepatic bile duct cancer and / or pancreatic cancer.

[0081] In some embodiments, the disease associated with FGFR2b is squamous non-small cell lung cancer, gastric cancer and / or gastroesophageal junction cancer.

[0082] In the fifth aspect of the present invention, the present invention provides the aforementioned antibody-drug conjugate, its pharmaceutically acceptable salt, solvate or solvate of the salt, or the aforementioned composition, for use in preventing and / or treating diseases associated with FGFR2b.

[0083] In some embodiments, the disease associated with FGFR2b is squamous non-small cell lung cancer, gastric cancer, gastroesophageal cancer, triple-negative breast cancer, ovarian cancer, endometrial cancer, cervical cancer, colorectal cancer, intrahepatic bile duct cancer and / or pancreatic cancer, etc.

[0084] In some embodiments, the disease associated with FGFR2b is squamous non-small cell lung cancer, gastric cancer and / or gastroesophageal junction cancer.

[0085] The sequences involved in the present invention are shown in Table A.

[0086] Definition of terms

[0087] Unless otherwise indicated, scientific and technical terms used herein have the meanings commonly understood by those skilled in the art. Furthermore, terms and laboratory procedures related to protein and nucleic acid chemistry, molecular biology, cell and tissue culture, microbiology, and immunology used herein are those widely used in the respective fields and are standard procedures. To facilitate a better understanding of the present invention, definitions and explanations of relevant terms are provided below.

[0088] As used herein, the terms "FGFR2IIIb" and "FGFR2b" are used interchangeably and refer to the subtype IIIb splice form of FGFR2. Exemplary FGFR2b sequences include Homo sapiens (human) FGFR2b protein (e.g., precursor sequence with a signal peptide, Genbank number: NP 075259.4); Mus musculus FGFR2b protein (e.g., full sequence, Genbank number: NP_963895.2). The term "anti-FGFR2b antibody" refers to an antibody that can specifically bind to FGFR2b. In some embodiments, the anti-FGFR2b antibodies provided herein specifically bind only to FGFR2b, but do not have detectable binding affinity to FGFR1b, FGFR1c, FGFR2c, FGFR3b, FGFR3c, or FGFR4.

[0089] The carcinogenic mechanism of FGFR2b in squamous non-small cell lung cancer includes, first, oncogene mutations, accounting for ~3%-5%, involving mutations in the extracellular functional domain II (Domain II) and domain III (Domain III), which lead to FGFR2b dimerization and kinase domain transphosphorylation, thereby recruiting linker proteins and activating downstream signaling pathways, as well as mutations in the kinase domain K660N and K660E, which lead to constitutive self-activation of the kinase domain that is independent of ligand and dimerization [10,11,12]. In addition, overexpression of FGFR2b protein increases its dimerization and ligand-independent signaling pathway activation.

[0013] Ligands FGF1, 7, 10, and 22 bind to and activate FGFR2b with high affinity. These ligands are overexpressed in sqNSCLC in an autocrine or paracrine manner in tumor fibroblasts, promoting tumor proliferation, metastasis, and progression.

[0014] In addition, FGFR2b is overexpressed in a variety of solid tumors, including squamous cell lung carcinoma, gastric cancer, gastroesophageal cancer, triple-negative breast cancer, ovarian cancer, endometrial cancer, cervical cancer, colorectal cancer, intrahepatic bile duct cancer, and pancreatic cancer.

[0090] The term FGFR2b-related disease refers to a disease in which FGFR2b expression in tissue cells differs from (e.g., exceeds) normal levels. For example, the level of FGFR2b in a tissue cell compared to the level of FGFR2b in a reference or control (i.e., normal tissue cell) indicates the presence of an FGFR2b-related disease in the subject (particularly a human) from which the tissue cell is derived.

[0091] In the present invention, unless otherwise stated, any numerical range should be understood to include any value or any sub-range within the range.

[0092] In the present invention, the term "antibody" refers to an immunoglobulin molecule typically composed of two pairs of identical polypeptide chains, each pair having a "light" (L) chain and a "heavy" (H) chain. Antibody light chains can be divided into two types: κ and λ. Heavy chains can be divided into five types: μ, δ, γ, α, or ε. Depending on the heavy chain, antibodies can be divided into five types: IgM, IgD, IgG, IgA, and IgE. Within light and heavy chains, the variable and constant regions are connected by a "J" region of approximately 12 or more amino acids, and the heavy chain also contains a "D" region of approximately 3 or more amino acids. Each heavy chain is composed of a heavy chain variable region (VH) and a heavy chain constant region (CH). The heavy chain constant region is composed of three domains (CH1, CH2, and CH3). Each light chain is composed of a light chain variable region (VL) and a light chain constant region (CL). The light chain constant region is composed of one domain, CL. The constant region of an antibody mediates the binding of immunoglobulins to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the complement system component C1q. The VH and VL regions can be further subdivided into highly variable regions called complementarity-determining regions (CDRs), interspersed with more conserved regions called framework regions (FRs). Each heavy chain variable region (VH) and light chain variable region (VL) consists of three CDRs and four FRs arranged from amino terminus to carboxyl terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions (VH and VL) of each heavy chain / light chain pair form the antibody binding site. The assignment of amino acids to regions or domains follows the definitions of Kabat Sequences of Proteins of Immunological Interest (National Institutes of Health, Bethesda, Md. (1987 and 1991)), or Chothia & Lesk (1987) J. Mol. Biol. 196:901-917; Chothia et al. (1989) Nature 342:878-883.

[0093] In the present invention, algorithms used to determine percent sequence identity (homology) and sequence similarity are, for example, BLAST and BLAST 2.0 algorithms, which are described in Altschul et al. (1977) Nucl. Acid. Res. 25:3389-3402 and Altschul et al. (1990) J. Mol. Biol. 215:403-410, respectively. BLAST and BLAST 2.0 can be used to determine percent identity of the amino acid sequences of the present invention, using, for example, the parameters described in the literature or the default parameters. Software for performing BLAST analyses is publicly available through the National Center for Biotechnology Information.

[0094] In the present invention, the amino acid sequence having at least 70% sequence identity with an amino acid sequence includes polypeptide sequences that are substantially identical to the amino acid sequence, for example, those sequences that contain at least 70% sequence identity, preferably at least 75%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity when compared to the polypeptide sequence of the present invention using the methods described herein (e.g., BLAST analysis using standard parameters).

[0095] In the present invention, a variant of an amino acid sequence refers to a sequence having greater than 70% identity to the amino acid sequence, such as greater than 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%, such as a sequence having 3, 2, or 1 amino acid substitutions, deletions, or additions. Preferably, the number of amino acids substituted, added, or deleted does not exceed 3. More preferably, the number of amino acids substituted, added, or deleted does not exceed 2. Most preferably, the number of amino acids substituted, added, or deleted does not exceed 1.

[0096] "Substitution" variants are variants in which at least one amino acid residue in the native sequence has been removed and a different amino acid has been inserted in the same position. The substitutions can be single, in which only one amino acid is substituted in the molecule, or multiple, in which two or more amino acids are substituted in the same molecule. Multiple substitutions can be located at consecutive sites. Likewise, one amino acid can be substituted by multiple residues, in which case such variants include both substitutions and insertions. "Insertion" (or "addition") variants are variants in which one or more amino acids are inserted adjacent to an amino acid at a specific position in a native sequence. Adjacent to an amino acid means linked to the α-carboxyl or α-amino functional group of the amino acid. "Deletion" variants are variants in which one or more amino acids in the native amino acid sequence have been removed. Typically, deletion variants have one or two amino acids deleted in a specific region of the molecule.

[0097] In certain embodiments, less than the theoretical maximum number of drug moieties are coupled to the antibody in the coupling reaction. Generally speaking, antibodies do not contain many free and reactive cysteine ​​thiol groups to which drug moieties can be attached; in fact, most cysteine ​​thiol groups in antibodies exist as disulfide bridges. In certain embodiments, antibodies can be reduced under partial or complete reducing conditions with a reducing agent such as dithiothreitol (DTT) or tricarbonylethylphosphine (TCEP) to generate reactive cysteine ​​thiol groups.

[0098] In some embodiments, the pharmaceutically acceptable salt is an inorganic acid salt or an organic acid salt, wherein the inorganic acid salt is hydrochloride, hydrobromide, hydroiodide, nitrate, bicarbonate and carbonate, sulfate or phosphate, and the organic acid salt is formate, acetate, propionate, benzoate, maleate, fumarate, succinate, tartrate, citrate, ascorbate, α-ketoglutarate, α-glycerophosphate, alkyl sulfonate or aryl sulfonate; preferably, the alkyl sulfonate is methyl sulfonate or ethyl sulfonate; and the aryl sulfonate is benzene sulfonate or p-toluene sulfonate.

[0099] Pharmaceutically acceptable salts can be obtained using standard procedures well known in the art, for example, by reacting a sufficient amount of a basic compound with a suitable acid affording a pharmaceutically acceptable anion.

[0100] As used herein, unless otherwise indicated, the term "prodrug" refers to a derivative that can be hydrolyzed, oxidized, or otherwise reacted under biological conditions (in vitro or in vivo) to provide a compound of the present invention. Prodrugs become active compounds only after this reaction under biological conditions, or they are active in their unreactive form. Prodrugs can generally be prepared using known methods, such as those described in Burger's Medicinal Chemistry and Drug Discovery (1995) 172-178, 949-982 (Manfred E. Wolff, 5th edition).

[0101] In the present invention, solvates refer to forms of the antibody-drug conjugates of the present invention that form complexes in solid or liquid form by coordination with solvent molecules. Hydrates are a specific form of solvates that have coordinated water molecules. In the present invention, hydrates are preferred solvates.

[0102] Methods for preparing various pharmaceutical compositions containing a certain amount of active ingredient are known or will be apparent to those skilled in the art based on the present disclosure. As described in REMINGTON'S PHARMACEUTICAL SCIENCES, Martin, EW, ed., Mack Publishing Company, 19th ed. (1995), the method for preparing the pharmaceutical composition includes incorporating suitable pharmaceutical excipients, carriers, diluents, etc., which are non-toxic to the cells or mammals exposed thereto at the dosages and concentrations employed.

[0103] The pharmaceutical compositions of the present invention may comprise a pH buffered aqueous solution. Alternatively, they may comprise a buffer such as phosphate, citrate, and other organic acids; an antioxidant including ascorbic acid; a low molecular weight (less than about 10 residues) polypeptide; a protein such as serum albumin, gelatin, or immunoglobulin; a hydrophilic polymer such as polyvinylpyrrolidone; an amino acid such as glycine, glutamine, asparagine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, sucrose, trehalose, or dextrin; a chelating agent such as ethylenediaminetetraacetic acid (EDTA); a sugar alcohol such as mannitol or sorbitol; a salt-forming counterion such as sodium; and / or a nonionic surfactant such as Tween™ and polyethylene glycol (PEG).

[0104] The pharmaceutical preparations of the present invention are manufactured by known methods, including conventional mixing, dissolving or lyophilizing methods. The compounds of the present invention can be prepared into pharmaceutical compositions and administered to the patient in various ways suitable for the selected mode of administration, for example, orally or parenterally (by intravenous, intramuscular, topical or subcutaneous routes).

[0105] Therefore, the compounds of the present invention can be systemically administered, for example, orally, in combination with a pharmaceutically acceptable carrier (such as an inert diluent or an assimilable edible carrier). They can be enclosed in hard or soft shell gelatin capsules and can be compressed into tablets. For oral therapeutic administration, the active compound can be combined with one or more excipients and used in the form of swallowable tablets, buccal tablets, lozenges, capsules, elixirs, suspensions, syrups or wafers. Such compositions and preparations should contain at least 0.1% active compound. The proportions of such compositions and preparations can, of course, vary and can account for about 1% to about 99% of the weight of a given unit dosage form. In such therapeutically useful compositions, the amount of active compound is such that an effective dosage level can be achieved.

[0106] Tablets, lozenges, pills, capsules, etc. may also contain: binders such as gum tragacanth, gum arabic, corn starch, or gelatin; excipients such as dicalcium phosphate; disintegrants such as corn starch, potato starch, or alginic acid; lubricants such as magnesium stearate; and sweeteners such as sucrose, fructose, lactose, or aspartame; or flavorings such as mint, wintergreen oil, or cherry flavor. When the unit dosage form is a capsule, it may contain, in addition to the above types of materials, a liquid carrier such as a vegetable oil or polyethylene glycol. Various other materials may be present as a coating or otherwise modify the physical form of the solid unit dosage form. For example, tablets, pills, or capsules may be coated with gelatin, wax, shellac, or sugar. Syrups or elixirs may contain the active compound, sucrose or fructose as a sweetener, methylparaben or propylparaben as a preservative, a dye, and a flavoring (such as cherry flavor or orange flavor). Of course, any material used in preparing any unit dosage form should be pharmaceutically acceptable and substantially non-toxic in the amounts employed. Additionally, the active compound can be incorporated into sustained-release preparations and devices.

[0107] The active compound can also be administered intravenously or intraperitoneally by infusion or injection. Aqueous solutions of the active compound or its salts can be prepared, optionally mixed with a nontoxic surfactant. Dispersions in glycerol, liquid polyethylene glycol, triacetin, and mixtures thereof, and oils can also be prepared. Under ordinary storage and use conditions, these formulations contain a preservative to prevent microbial growth.

[0108] Pharmaceutical dosage forms suitable for injection or infusion can include sterile aqueous solutions or dispersants or sterile powders containing active ingredients (optionally encapsulated in liposomes) suitable for instant preparations of sterile injectable or infusible solutions or dispersants. In all cases, the final dosage form must be sterile, liquid and stable under production and storage conditions. The liquid carrier can be a solvent or liquid dispersion medium, including, for example, water, ethanol, polyols (e.g., glycerol, propylene glycol, liquid polyethylene glycol, etc.), vegetable oils, non-toxic glycerides and suitable mixtures thereof. Suitable fluidity can be maintained, for example, by the formation of liposomes, by maintaining the required particle size in the case of dispersants, or by the use of surfactants. The effect of preventing microorganisms can be produced by various antibacterial and antifungal agents (such as parabens, chlorobutanol, phenol, sorbic acid, thimerosal, etc.). In many cases, it is preferred to include isotonic agents, such as sugars, buffers or sodium chloride. Prolonged absorption of injectable compositions can be produced by using compositions that delay absorption (e.g., aluminum monostearate and gelatin).

[0109] Sterile injectable solutions are prepared by combining the active compound in the required amount in an appropriate solvent with various of the other ingredients enumerated above, as required, followed by filtered sterilization. In the case of sterile powders for the preparation of sterile injectable solutions, the preferred methods of preparation are vacuum drying and freeze drying techniques, which yield a powder of the active ingredient plus any additional desired ingredient present in a previously sterile-filtered solution.

[0110] Useful solid carriers include pulverized solids (such as talc, clay, microcrystalline cellulose, silicon dioxide, aluminum oxide, etc.). Useful liquid carriers include water, ethanol or ethylene glycol or water-ethanol / ethylene glycol mixtures, in which the compounds of the present invention can be dissolved or dispersed in an effective amount, optionally with the aid of non-toxic surfactants. Adjuvants (such as flavors) and additional antimicrobial agents can be added to optimize the properties for a given use.

[0111] Thickeners (such as synthetic polymers, fatty acids, fatty acid salts and esters, fatty alcohols, modified celluloses or modified inorganic materials) can also be used with liquid carriers to form spreadable pastes, gels, ointments, soaps, etc., for application directly to the user's skin.

[0112] The above-mentioned formulations can be presented in unit dosage form, which is a physically discrete unit containing a unit dose suitable for administration to humans and other mammals. The unit dosage form can be a capsule or tablet, or a plurality of capsules or tablets. Depending on the specific treatment involved, the amount of the active ingredient in a unit dose can be varied or adjusted from about 0.1 to about 1000 mg or more.

[0113] In addition, it also includes the application of various new drug dosage forms such as emulsion liposomes, microspheres and nanospheres, such as drugs prepared using particulate dispersion systems including polymeric micelles, nanoemulsions, submicroemulsions, microcapsules, microspheres, liposomes and lipid vesicles (niosomes) (also known as non-ionic surfactant vesicles).

[0114] As used herein, the term "treat" generally refers to obtaining a desired pharmacological and / or physiological effect. The effect may be prophylactic, in terms of completely or partially preventing a disease or its symptoms; and / or therapeutic, in terms of partially or completely stabilizing or curing a disease and / or causing side effects due to the disease. As used herein, "treat" encompasses any treatment of a disease in a patient, including: (a) preventing the onset of a disease or symptom in a patient who is susceptible to the disease or symptom but has not yet been diagnosed with the disease; (b) suppressing the symptoms of a disease, i.e., arresting its development; or (c) relieving the symptoms of a disease, i.e., causing regression of the disease or symptom.

[0115] In the present invention, "subject" refers to a vertebrate. In certain embodiments, the vertebrate refers to a mammal. Mammals include, but are not limited to, livestock (such as cattle), pets (such as cats, dogs, and horses), primates, mice, and rats.

[0116] In certain embodiments, the mammal is a human.

[0117] In the present invention, an "effective amount" refers to an amount that is effective at the necessary dosage and time to achieve the desired therapeutic or preventive effect. The "therapeutically effective amount" of the substance / molecule of the present invention may vary according to factors such as the disease state, age, sex and weight of the individual and the ability of the substance / molecule to elicit the desired response in the individual. A therapeutically effective amount also encompasses an amount in which the therapeutically beneficial effects of the substance / molecule outweigh any toxic or deleterious consequences. A "prophylactically effective amount" refers to an amount that is effective at the necessary dosage and time to achieve the desired preventive effect. Typically, but not necessarily, a prophylactic dose is used in subjects before the onset of disease or in the early stages of the disease, so the prophylactic effective amount will be lower than the therapeutically effective amount. In the case of cancer, a therapeutically effective amount of a drug can reduce the number of cancer cells; reduce tumor size; inhibit (i.e., slow down to a certain extent, preferably stop) cancer cell infiltration into surrounding organs; inhibit (i.e., slow down to a certain extent, preferably stop) tumor metastasis; inhibit tumor growth to a certain extent; and / or alleviate one or more symptoms associated with cancer to a certain extent.

[0118] In the present invention, the 20 conventional amino acids and their abbreviations follow conventional usage. See Immunology-A Synthesis (2nd edition, ES Golub and DR Gren, Eds., Sinauer Associates, Sunderl et al., Mass. (1991)), which is incorporated herein by reference.

[0119] The term "chimeric antibody" refers to antibodies in which the variable region sequences are derived from one species and the constant region sequences are derived from another species, such as antibodies in which the variable region sequences are derived from a mouse antibody and the constant region sequences are derived from a human antibody.

[0120] "Humanized" antibodies refer to non-human (e.g., mouse) antibody forms that are chimeric immunoglobulins, immunoglobulin chains, or fragments thereof (e.g., Fv, Fab, Fab', F(ab')2, or other antigen-binding subsequences of antibodies) that contain minimal sequence derived from non-human immunoglobulin. Preferably, humanized antibodies are human immunoglobulins (recipient antibody) in which residues from a complementarity determining region (CDR) of the recipient antibody are replaced by residues from a CDR of a non-human species (donor antibody) such as mouse, rat, or rabbit having the desired specificity, affinity, and capacity.

[0121] In addition, in humanization, it is also possible to mutate the amino acid residues in the CDR1, CDR2 and / or CDR3 regions of VH and / or VL to thereby improve one or more binding properties (e.g., affinity) of the antibody. For example, PCR-mediated mutations can be used to introduce mutations, and their impact on antibody binding or other functional properties can be evaluated using in vitro or in vivo tests as described herein. Typically, conservative mutations are introduced. Such mutations can be amino acid substitutions, additions, or deletions. In addition, the mutations in the CDRs are typically no more than one or two. Therefore, the humanized antibodies of the present invention also encompass antibodies comprising one or two amino acid mutations in the CDRs. BRIEF DESCRIPTION OF THE DRAWINGS

[0122] FIG1 shows the light chain ( FIG1A ) and heavy chain ( FIG1B ) sequences of the anti-FGFR2b humanized monoclonal antibody humAb1;

[0123] FIG2 shows the binding of antibody humAb1, antibody drug conjugate ADC-1, and ADC control to human, monkey, and mouse FGFR2b, respectively, as measured by biolayer interferometry (BLI) (OCET), and the calculated binding affinities;

[0124] FIG3 shows the ability of humAb1, ADC-1, and ADC control to bind to surface antigens of tumor cells SNU16 ( FIG3A ), KATO III ( FIG3B ), and MFM-223 ( FIG3C ) at different concentrations analyzed by flow cytometry;

[0125] Figure 4 shows the molecular structure of the tubulin-binding drug MMAE;

[0126] Figure 5 shows the molecular structure of the chemical linker-drug, i.e., LD, MC-vc-PAB-MMAE;

[0127] FIG6 is the molecular structure of humAb1-vc-MMAE, an anti-FGFR2 antibody-drug conjugate according to an embodiment of the present invention;

[0128] FIG7 is a hydrophobic interaction chromatogram (HIC) of an antibody-drug conjugate according to an embodiment of the present invention;

[0129] FIG8 shows the cell growth inhibition effects of humAb1, ADC-1, IgG isotype control, and MMAE on HEK293, and gastric cancer cells SNU16 and KATO III, respectively, according to an embodiment of the present invention;

[0130] FIG9 shows the cell growth inhibition effect of ADC-1 and MMAE, respectively, on engineered lung squamous cell carcinoma cells H1703-FGFR2b and H2170-FGFR2b that overexpress FGFR2b, according to an embodiment of the present invention;

[0131] FIG10 is an antibody-dependent cell-mediated cytotoxicity (ADCC) effect of the antibody-drug conjugates implemented in the present invention;

[0132] FIG11 shows the tumor inhibition effect of the antibody-drug conjugate of the present invention on SNU16 gastric cancer-bearing nude mice, including changes in tumor volume over treatment time ( FIG11A ) and body weight over time ( FIG11B );

[0133] FIG12 shows the tumor inhibition effect of the antibody drug conjugate of the present invention on nude mice bearing H1703-FGFR2b lung squamous cell carcinoma, including changes in tumor volume ( FIG12A ) and body weight ( FIG12B ) over the days after treatment. DETAILED DESCRIPTION

[0134] The embodiments of the present invention will be described in detail below with reference to the examples, but it will be understood by those skilled in the art that the following examples are merely illustrative of the present invention and should not be construed as limiting the scope of the invention. Where specific conditions are not specified in the examples, the methods were performed according to conventional conditions or the conditions recommended by the manufacturer. Where the manufacturers of the reagents or instruments are not specified, they are all conventional products that can be obtained commercially.

[0135] The present invention will be further explained below with reference to specific examples, but these examples are not intended to limit the scope of the present invention.

[0136] Example 1 Preparation of humanized anti-FGFR2b monoclonal antibody

[0137] Balb / c mice, CD1 mice, or SJL mice were immunized subcutaneously with human FGFR2b-Fc (Cat. 16485-H02H, Beijing Sino-Biotech Co., Ltd.) in Freund's adjuvant, SAS, or CpG with an initial dose of 50 μg per mouse and then 25 μg per mouse every three weeks. Serum titers of anti-human FGFR2b-Fc were determined by ELISA. The final booster immunization cycle was performed by intravenous injection for three consecutive days at 4 μg, 3 μg, and 3 μg, respectively. Four days later, mouse spleen cells were extracted and fused with mouse myeloma cells. Seven days after fusion, the hybridoma culture supernatant was first screened for FGFR2b-His binding by ELISA. Hybridomas that passed the primary screening underwent secondary screening, including binding to KATO III cells by FACS. Hybridomas that passed the secondary screening were screened and subcloned to obtain mouse monoclonal antibodies.

[0138] The heavy chain and light chain variable region (VH, VL) sequences of mAb1 were determined using standard RACE technology. Total RNA was extracted from the selected hybridoma cell line. SMART RACE cDNA amplification kit (Clontech) was then used to generate the full-length first-strand cDNA containing the 5' end by PCR amplification according to the specification. The PCR product was isolated and purified, and then cloned and sequenced.

[0139] The chimeric antibody mAb1 was then generated by transplanting the VH and VL sequences of mouse Ab1 into a human Fc. Subsequently, a library was constructed using computer simulations to perform backmutation on key framework regions. Through library screening, humanized amino acids at framework region sites that significantly impact antibody affinity were backmutated to the original mouse framework amino acid residues (using Kabat numbering). The humanized anti-FGFR2b antibody was named humAb1. The amino acid sequences of the humAb1 light chain (Figure 1A) and heavy chain (Figure 1B) are shown in Figure 1.

[0140] Example 2 Binding KD of Anti-FGFR2b Antibody Drug Conjugate to FGFR2b Antigen

[0141] The binding of mAb1, ADC-1 and ADC control to human, cynomolgus monkey and mouse FGFR2b antigens was determined by biolayer interferometry (BLI) (OCET). Briefly, the Protein A biosensor was first pre-wetted with PBST (0.05% Tween 20) for 10 minutes. The sample was diluted with PBST-BSA (0.05% Tween, 0.1% BSA) and injected at a continuous concentration (0nM, 1.56nM, 3.12nM, 6.25nM, 12.5nM, 25nM, 50nM), and the surface regeneration of the Protein A biosensor was included in each run cycle. The binding constant and dissociation constant were calculated using OCET RED384 evaluation software (version 1.0). As shown in Figure 2, ADC-1 specifically binds to human FRFR2b with strong binding affinity (nanomolar), and cross-reacts with both cynomolgus monkey and mouse FGFR2b. ADC-1 exhibits similar binding affinity to humAb1 for FGFR2b.

[0142] Example 3: Binding of anti-FGFR2b antibody-drug conjugates to FGFR2b-overexpressing tumor cells

[0143] Gastric cancer cells KATO III (ATCC, HTB-103 TM) were cultured in IMDM (ATCC, 30-2005) medium (Iscove's modified Dulbecco's medium) containing 20% ​​FBS (Gibco, 10091-148) and 1% penicillin-streptomycin (Gibco, 15140-122). SNU16 (ATCC, CRL-5974 TM ) were cultured in RPMI1640 (VivaCell, C3010-0500) medium containing 10% FBS and 1% penicillin-streptomycin, and breast cancer cells MFM-223 (Cobioer, CBP60388) were cultured in EMEM (ATCC, 30-2003) medium (Eagle's minimum essential medium) containing 10% FBS and 1% penicillin-streptomycin. The cells were cultured in a 37°C, 5% CO2 incubator. When the cells reached the logarithmic growth phase, they were digested with 0.25% Trypsin-EDTA (Gibco, 25200-072) and resuspended in PBS for counting. 1E5 cells / 95 μL / well were plated in a 96-well plate (Corning, 3799). The test sample was diluted with PBS to 200 μg / ml and a 4-fold serial dilution was performed. 5 μL of the serially diluted test sample was added to each well and incubated at 4°C for 1 h (1 h represents 1 hour, which can also be represented by 1 hr; similarly, 2 h represents 2 hours, which can also be represented by 2 hrs or 2 hrs; 3 h represents 3 hours, which can also be represented by 3 hrs or 3 hrs). Wash twice with PBS, add PE Goat Anti-Human IgG Fc (PE / phycoerythrin labeled goat anti-human IgG Fc) secondary antibody (abcam, ab98596), incubate at 4℃ for 30min (30min means 30 minutes, similar to the following, such as 4min means 4 minutes). After washing twice with PBS, analyze the cells using a flow cytometer (BD FACSLyric TM Flow cytometry was used to analyze the mean fluorescence intensity (MFI). Graphpad Prism software was used to perform four-parameter curve fitting analysis on the data to obtain the half-effect concentration (EC50) value. The results are shown in Figure 3.

[0144] ADC-1 is humAbl coupled to MC-vc-PAB-MMAE, while the ADC control is an IgG1 isotype control antibody coupled to the same linker, the toxin MC-vc-PAB-MMAE. ADC-1 exhibited similar EC50s as humAbl for binding to tumor cells SNU16 (Figure 3A), KATO III (Figure 3B), and MFM-223 (Figure 3C).

[0145] Example 4 Preparation of Antibody Drug Conjugates

[0146] 1. Purification of monoclonal antibody mAb1

[0147] The monoclonal antibody mAb1 was captured from CHO cell culture medium using Protein A. The purity of mAb1 was >95% as determined by SDS-PAGE and SEC analysis. The resulting antibody protein was dialyzed into a buffer using a 30 kDa membrane, concentrated, and calibrated using a UV spectrophotometer for subsequent coupling reactions.

[0148] 2. Conjugation of monoclonal antibody mAb1 with drug molecules

[0149] Prepare the reducing agent in a buffer solution (20 mM Tris-HCl, pH 6.0) as follows: 1-20 mmol / L TCEP (Tris-2-carboxyethyl-phosphine) (SIGMA, C4706) stock solution. The reducing agent is added within a specific concentration range depending on the desired coupling rate. Mix with a specific concentration of monoclonal antibody (e.g., 5-10 mg / ml) in a specific ratio to achieve a final molar ratio of TCEP to antibody of 1-5:1. Stir the reaction at 15°C for 1-2 hours.

[0150] After TCEP reduction, the antibody was conjugated to LD. A 5 mM concentration of the drug MC-vc-PAB-MMAE (MCE, HY-15575) was dissolved in 20% DMSO (dimethyl sulfoxide, SIGMA, 276855). The drug was added at a molar ratio of 1 to 10:1 and stirred at 25°C for 1 to 2 hours.

[0151] The sample was dialyzed for 10-15 times the volume to remove free drug residues. The sample purity was tested by SDS-PAGE electrophoresis and SEC-HPLC. The coupling status, i.e., DAR value, was tested by HIC-HPLC.

[0152] Example 5 Anti-tumor cell proliferation activity of antibody drug conjugates

[0153] FGFR2b-overexpressing lung squamous cell carcinoma cell line NCI-H2170 (ATCC, CRL-5928) TM )、NCI-H1703(ATCC,CRL-5889 TM) were cultured in RPMI1640 medium supplemented with 10% FBS, 1% penicillin-streptomycin, and 1 μg / ml puromycin (InvivoGen, ant-pr-01). KATO III cells, NCI-H2170 cells overexpressing FGFR2IIIb, and NCI-H1703 cells in logarithmic growth phase were resuspended and seeded into 96-well plates (WHB, WHB-96-01) at a density of 1000 cells / 100 μL / well, 1000 cells / 100 μL / well, and 1000 cells / 100 μL / well, respectively. The cells were cultured overnight at 37°C in a 5% CO2 incubator. SNU16 cells were seeded into 96-well plates at a density of 3000 cells / 100 μL / well on the day of the experiment. The sample was diluted to 21 μM with PBS and serially diluted three-fold for a total of nine concentrations, with 5 μL of sample added to each well of a 96-well plate. A negative control group contained no cells but an equal volume of culture medium (blank); a positive control group contained cells but no test article but 5 μL of PBS (control). The 96-well plate was incubated at 37°C, 5% CO₂ for 5 days. Cell-Titer Glo reagent (Promega, G7573) was added, and after shaking for 10 minutes, chemiluminescence was measured using a microplate reader (TECAN, INSTRUMENT SPARK). The percentage inhibition (% Inhibition) was calculated, and the data were analyzed by four-parameter curve fitting using Graphpad Prism software to obtain the IC₅₀ value. % Inhibition = 1-100% * (Signal - Blank) / (Control - Blank).

[0154] As shown in Figure 8, ADC-1 demonstrated potent, target-specific, and expression-level-dependent growth inhibition against SNU16 and KAT OIII tumor cells, but no specific growth inhibition against HER293 cells. IgG isotype control and humAb1 showed no specific growth inhibition against the corresponding cells; however, the small molecule drug MMAE exhibited similar, nonselective cytotoxicity against these three cell lines.

[0155] As shown in FIG9 , ADC-1 exhibited target-specific and stronger cell growth inhibitory activity against lung squamous cell carcinoma cells H2170-FGFR2b and H1703-FGFR2b than the small molecule drug MMAE.

[0156] Case Study 6: ADCC Effect of Antibody-Drug Conjugates

[0157] NCI-H1703 cells overexpressing FGFR2b were used as target cells. Cells were resuspended and plated at a density of 2E4 cells / 95 μL per well in a 96-well plate (WHB, WHB-96-01). The plates were incubated overnight at 37°C, 5% CO2. Jurkat-Luc NFAT-CD16a-V158 effector cells were resuspended and plated at a density of 1E5 cells / 50 μL per well in the same 96-well plate. The sample was diluted to 150 μg / mL in PBS and serially diluted 5-fold for a total of eight concentrations. 5 μL of sample was added to each well of the 96-well plate. The cells were incubated at 37°C, 5% CO2 for 6 hours. Nano-Glo reagent (Promega, N1120) was added and shaken for 3 minutes (3 min represents 3 minutes, other methods are similar). Chemiluminescence was measured using a microplate reader. The data were analyzed by four-parameter curve fitting using Graphpad Prism software to obtain the EC50 value.

[0158] As shown in FIG10 , ADC-1 showed dose-dependent ADCC functional activity similar to humAbl.

[0159] Example 7 Anti-tumor Effect of Antibody Drug Conjugates on SNU16 Gastric Cancer in Nude Mice

[0160] Human SNU16 gastric cancer nude mouse CDX modeling process: 5x10 6 SNU16 cells were suspended in PBS and injected subcutaneously into the right dorsal region of 6-8 week old BALB / c female nude mice. 3 The mice were randomly divided into 3 groups, each containing 8 mice. The tumor volume was kept uniform across the groups, taking into account the body weight. The groups included the vehicle group, the 10 mg / kg ADC-1 group, and the 10 mg / kg humAb1 group. The average tumor volume in each group was 209 mm. 3 The solvent and ADC-1 were administered three times through the tail vein on D0, D7, and D14, respectively. The humAb1 group received six doses of the drug twice weekly through the tail vein on D0, D4, D7, D11, D14, and D18. ADC-1 refers to the anti-FGFR2b antibody humAb1 conjugated to MC-vc-PAB-MMAE-.

[0161] Data analysis: During the experiment, tumor volume was measured twice a week. The calculation formula of tumor volume (TV) is: TV = l × w 2 / 2. Wherein l and w represent the measured length and width of the tumor, respectively. The relative tumor volume (RTV) is calculated based on the measurement results, RTV = Vf / V0. Wherein V0 is the tumor volume measured at the time of group administration (i.e. Day0, i.e. day 0, may also be represented by Day0, D0 or day0. Similarly, Day11 represents the 11th day after grouping, represented by Day11, D11 or day11, and other Day14, Day46, D14, D46, etc. have the same meaning), and Vf is the tumor volume measured on the last day. Relative tumor proliferation rate T / C (%) = (RTV of the drug administration group / RTV of the solvent group) × 100%. Tumor growth inhibition rate TGI% = (average tumor volume of the solvent group - average tumor volume of the drug administration group) / average tumor volume of the solvent group × 100%.

[0162] The experimental results are shown in Figure 11. Complete tumor regression was observed in 2 / 8 (2 out of 8 nude mice; 6 / 8, 0 / 8, 7 / 8, and so on) of the ADC-1 group on Day 11, Day 14, and Day 46, respectively. In the humAb1 group, 0 / 8 of the tumors had regressed. On Day 46, the tumor growth inhibition (TGI) percentages (%TGI) for the ADC-1 and humAb1 groups compared to the vehicle group were 109% and 35%, respectively. These results demonstrate that ADC-1 exhibits significant tumor suppression, whereas humAb1 exhibits no significant anti-tumor activity (Figure 11A). Both ADC-1 and humAb1 were well tolerated by tumor-bearing mice (Figure 11B).

[0163] Example 8 Antibody Drug Conjugate Inhibitory Effect on H1703 Lung Cancer Overexpressing FGFR2b in Nude Mice CDX Modeling of H1703 Lung Squamous Carcinoma Overexpressing Human FGFR2b in Nude Mice: 5x106 cells were suspended in PBS and injected subcutaneously into the right dorsal region of 6-8 week old BALB / c female nude mice. When the tumor volume reached 90-250mm 3 The mice were randomly divided into groups of 7 mice each, ensuring uniform tumor volume and weight between groups. There were 5 groups in total, namely the solvent group, the chemotherapy group (2.5mg / kg cisplatin + 10mg / kg paclitaxel), and the ADC-1 group with doses of 1mg / kg, 3mg / kg, and 10mg / kg, respectively. After grouping, the average tumor volume of each group was 153mm 3 The vehicle group and different doses of ADC-1 were administered twice via tail vein bolus on D0 and D7, respectively; the chemotherapy group received six intraperitoneal doses on D0, D4, D7, D11, D14, and D18. ADC-1 refers to the anti-FGFR2b antibody humAb1 conjugated to an antibody-drug conjugate (ADC) containing MC-vc-PAB-MMAE.

[0164] Data analysis: During the experiment, tumor volume was measured twice a week. The calculation formula of tumor volume (TV) is: TV = l × w 2 / 2. Where l and w represent the measured length and width of the tumor, respectively. Relative tumor volume (RTV) was calculated based on the measurement results: RTV = Vf / V0. V0 is the tumor volume measured at the time of group dosing (i.e., Day 0), and Vf is the tumor volume measured on the last day. Relative tumor growth rate (T / C) (%) = (RTV of the treatment group / RTV of the solvent group) × 100%. Tumor growth inhibition rate (TGI%) = (average tumor volume of the solvent group - average tumor volume of the treatment group) / average tumor volume of the solvent group × 100%.

[0165] The experimental results are shown in Figure 12. On Day 21, 1 / 7 tumors in the chemotherapy group completely regressed and 1 / 7 partially regressed. In the ADC-11 mg / kg and 3 mg / kg dose groups, 2 / 7 and 7 / 7 tumors partially regressed, respectively; while in the 10 mg / kg dose group, 2 / 7 tumors completely regressed and the other 5 / 7 tumors partially regressed. On Day 21, the tumor growth inhibition rate TGI% of the chemotherapy group and the ADC-1 groups with low to high doses compared with the solvent group was 54%, 99%, 110%, and 113%, respectively. The experimental results show that compared with chemotherapy drugs, the antibody drug conjugate ADC-1 has significant in vivo efficacy in transplanted lung squamous cell carcinoma tumors (Figure 12A), and tumor-bearing mice tolerate the ADC-1 drug well even at high doses (Figure 12B).

[0166] Table A: Comparison table of amino acid sequence numbers and amino acid sequences

[0167] References:

[0168] 1.Sung H,Ferlay J,Siegel RL,etc.CA Cancer J Clin.2021May;71(3):209-249.

[0169] 2. Qin Na, Ma Hongxia, Jin Guangfu, et al. Chinese Medical Journal, 2023, 103(14): 1068-1073.

[0170] 3.Larissa A Pikor 1,Varune R Ramnarine,Stephen Lam,etc.Lung Cancer 2013 Nov;82(2):179-89.

[0171] 4. Perez-Moreno P, Brambilla E, Thomas R, Lung Cancer Manag, etc. Clinical Cancer Research. May 1 2012; 18(9): 2443-2451

[0172] 5.Cancer Genome Atlas Research Network.Comprehensive genomic characterization of squamous cell lung cancer.Nature.Sep 27 2012;489(7417):519-525.

[0173] 6. NCCN Clinical Practice Guidelines (2023.V3): Non-Small Cell Lung Cancer

[0174] 7.2023 CSCO Non-Small Cell Lung Cancer Diagnosis and Treatment Guidelines

[0175] 8. NCCN Clinical Practice Guidelines (2023): Gastric Cancer

[0176] 9.2023 CSCO Gastric Cancer Diagnosis and Treatment Guidelines

[0177] 10.Xiuqin Zhang,Omar A Ibrahimi,Shaun K Olsen,etc.J Biol Chem.2006 Jun 9;281(23):15694-700.

[0178] 11. Genshi Zhao, Wei-Ying Li, Daohong Chen, etc. Mol Cancer Ther. 2011 Nov;10(11):2200-10.

[0179] 12.Rachel G Liao,Joonil Jung,Jeremy Tchaicha,etc.Cancer Res.2013 Aug 15;73(16):5195-205.

[0180] 13.Irina S.Babina,Nicholas C.Turner.Nature Reviews Cancer volume 17,pages318–332(2017)

[0181] 14.Carmen Behrens,Healther Y.Lin,J.Jack Lee,etc.Clin Cancer Res.2008Oct 1;14(19):6014–6022.

Claims

1. An antibody-drug conjugate, and a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, wherein the antibody-drug conjugate has a structure as shown in Formula I: Ab-(LD)p Formula I in: Ab is an anti-FGFR2b antibody, comprising a heavy chain and a light chain, wherein the heavy chain variable complementary determining region 1 (CDR1) comprises a sequence selected from SEQ ID NO: 59, 65, 71, 77, 83, 88, 93, 103, 110, 119, 135, 143, 146, 150 or 152 or a mutant thereof, the heavy chain variable complementary determining region 2 (CDR2) comprises a sequence selected from SEQ ID NO: 60, 66, 72, 78, 84, 94, 111, 118, 120, 124, 141 or 153 or a mutant thereof, and the heavy chain variable complementary determining region 3 (CDR3) comprises a sequence selected from SEQ ID NO: NO: 61, 67, 73, 79, 85, 89, 95, 101, 104, 112, 116, 121, 125, 128, 131, 136, 138, 142, 144, 147, 151, 154 or 158 or a mutant thereof; the light chain variable complementary determining region 1 (CDR1) comprises a sequence selected from SEQ ID NO: 62, 68, 74, 80, 86, 90, 96, 99, 105, 108, 113, 122, 126, 129, 132, 148, 149 or 155 or a mutant thereof, the light chain variable complementary determining region 2 (CDR2) comprises a sequence selected from SEQ ID NO: NO: 63, 69, 75, 81, 91, 97, 106, 114, 133, 139 or 156, or a mutant thereof, and the light chain variable complementarity determining region 3 (CDR3) comprises a sequence selected from SEQ ID NO: 64, 70, 76, 82, 87, 92, 98, 100, 102, 107, 109, 115, 117, 123, 127, 130, 134, 137, 140, 145, 157 or 159, or a mutant thereof; D is a cytotoxic agent; L is a linker used to connect the anti-FGFR2b antibody and the cytotoxic agent; p is 2.0-8.

0.

2. The antibody-drug conjugate according to claim 1, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The anti-FGFR2b antibody has a heavy chain variable complementary determining region 1 (CDR1) comprising a sequence selected from SEQ ID NO: 59, 65, 71, 77, 83, 88, 93, 103, 110, 119, 135, 143, 146, 150 or 152 or a mutant thereof, a heavy chain variable complementary determining region 2 (CDR2) comprising a sequence selected from SEQ ID NO: 60, 66, 72, 78, 84, 94, 111, 118, 120, 124, 141 or 153 or a mutant thereof, and a heavy chain variable complementary determining region 3 (CDR3) comprising a sequence selected from SEQ ID NO: The sequence shown in NO: 61, 67, 73, 79, 85, 89, 95, 101, 104, 112, 116, 121, 125, 128, 131, 136, 138, 142, 144, 147, 151, 154 or 158, or a mutant thereof.

3. The antibody-drug conjugate according to claim 1 or 2, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The heavy chain variable complementary determining region 1 (CDR1), heavy chain variable complementary determining region 2 (CDR2), and heavy chain variable complementary determining region 3 (CDR3) of the anti-FGFR2b antibody are selected from the following sequence combinations:

4. The antibody drug conjugate according to any one of claims 1 to 3, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The anti-FGFR2b antibody has a light chain variable complementary determining region 1 (CDR1) comprising a sequence selected from SEQ ID NO: 62, 68, 74, 80, 86, 90, 96, 99, 105, 108, 113, 122, 126, 129, 132, 148, 149 or 155 or a mutant thereof, a light chain variable complementary determining region 2 (CDR2) comprising a sequence selected from SEQ ID NO: 63, 69, 75, 81, 91, 97, 106, 114, 133, 139 or 156 or a mutant thereof, and a light chain variable complementary determining region 3 (CDR3) comprising a sequence selected from SEQ ID NO: The sequence shown in NO: 64, 70, 76, 82, 87, 92, 98, 100, 102, 107, 109, 115, 117, 123, 127, 130, 134, 137, 140, 145, 157 or 159, or a mutant thereof.

5. The antibody drug conjugate according to any one of claims 1 to 4, or a pharmaceutically acceptable salt, solvate or solvate of the salt thereof, characterized in that: The light chain variable complementary determining region 1 (CDR1), variable complementary determining region 2 (CDR2) and variable complementary determining region 3 (CDR3) of the anti-FGFR2b antibody are selected from the following sequence combinations:

6. The antibody drug conjugate according to any one of claims 1 to 5, or a pharmaceutically acceptable salt, solvate or solvate of the salt thereof, characterized in that: The anti-FGFR2b antibody has a heavy chain variable region framework region 1 (FR1) comprising a sequence selected from SEQ ID NO: 160, 168, 175, 188, 194, 204, 210, 224, 233, 284, 259, 265, 268, 271, 274, 281, 295 or 305 or a mutant thereof, a heavy chain variable region framework region 2 (FR2) comprising a sequence selected from SEQ ID NO: 161, 169, 176, 183, 205, 221, 225, 237, 245, 251, 261, 266, 275, 282 or 296 or a mutant thereof, and a heavy chain variable region framework region 3 (FR3) comprising a sequence selected from SEQ ID NO: NO: 162, 170, 177, 184, 189, 195, 200, 206, 211, 216, 226, 231, 234, 239, 242, 246, 252, 254, 260, 262, 267, 272, 276, 297 or 306, or a mutant thereof, and the heavy chain variable region framework region 4 (FR4) comprises a sequence selected from SEQ ID NO: 163, 178, 190, 196, 212, 217, 227, 255, 269 and 298, or a mutant thereof.

7. The antibody drug conjugate according to any one of claims 1 to 6, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The anti-FGFR2b antibody has a light chain variable framework region 1 (FR1) comprising a sequence selected from SEQ ID NO: 164, 171, 179, 185, 191, 197, 201, 207, 213, 218, 222, 228, 232, 247, 256, 263, 270, 277, 283, 299 or 303 or a mutant thereof, a light chain variable framework region 2 (FR2) comprising a sequence selected from SEQ ID NO: 165, 172, 180, 186, 192, 198, 202, 208, 214, 219, 223, 229, 235, 240, 243, 248, 257, 264, 278, 300 or 304 or a mutant thereof, and a light chain variable framework region 3 (FR3) comprising a sequence selected from SEQ ID NO: NO: 166, 173, 181, 187, 193, 199, 203, 209, 220, 230, 236, 238, 241, 244, 249, 253, 258, 273, 279, 301, 316 and 317 or a mutant thereof, and the light chain variable framework region 4 (FR4) comprises a sequence selected from SEQ ID NO: 167, 174, 182, 215, 250, 280 or 302 or a mutant thereof.

8. The antibody drug conjugate according to any one of claims 1 to 7, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The anti-FGFR2 heavy chain variable framework region 1 (FR1), heavy chain variable framework region 2 (FR2), heavy chain variable framework region 3 (FR3) and heavy chain variable framework region 4 (FR4) are selected from the following sequence combinations:

9. The antibody drug conjugate according to any one of claims 1 to 8, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The anti-FGFR2 light chain variable framework region 1 (FR1), light chain variable framework region 2 (FR2), light chain variable framework region 3 (FR3) and light chain variable framework region 4 (FR4) are selected from the following sequence combinations:

10. The antibody drug conjugate according to any one of claims 1 to 9, or a pharmaceutically acceptable salt, solvate or solvate of the salt thereof, characterized in that: The complementary determining regions and framework regions of the heavy chain variable region of the antibody are arranged in the following order from the N-terminus to the C-terminus of the sequence: heavy chain framework region 1-heavy chain complementary determining region 1-heavy chain framework region 2-heavy chain complementary determining region 2-heavy chain framework region 3-heavy chain complementary determining region 3-heavy chain framework region 4.

11. The antibody drug conjugate according to any one of claims 1 to 10, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The heavy chain of the antibody comprises three constant regions: constant region 1, constant region 2 and constant region 3, which are composed of SEQ ID NO.307, SEQ ID NO.309 and SEQ ID NO.310, or composed of SEQ ID NO:307, SEQ ID NO.312 and SEQ ID NO.

313.

12. The antibody drug conjugate according to any one of claims 1 to 11, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The heavy chain variable region of the anti-FGFR2b antibody is selected from SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 285, 287, 290, 291 or a homologous sequence thereof, wherein the homologous sequence is the same as SEQ ID NO: NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 285, 287, 290 or 291 has at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence homology.

13. The antibody drug conjugate according to any one of claims 1 to 12, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The heavy chain variable region of the anti-FGFR2b antibody is selected from the sequence shown in SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 285, 287, 290 or 291.

14. The antibody-drug conjugate according to any one of claims 1 to 13, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The complementary determining regions and framework regions of the light chain variable region of the antibody are arranged in the following order from the N-terminus to the C-terminus of the sequence: light chain framework region 1-light chain complementary determining region 1-light chain framework region 2-light chain complementary determining region 2-light chain framework region 3-light chain complementary determining region 3-light chain framework region 4.

15. The antibody drug conjugate according to any one of claims 1 to 14, or a pharmaceutically acceptable salt, solvate or solvate of the salt thereof, characterized in that: The antibody further comprises a light chain constant region of SEQ ID NO:

311.

16. The antibody drug conjugate according to any one of claims 1 to 15, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The light chain variable region of the antibody comprises SEQ ID Nos: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 46, 48, 50, 54, 56, 58, 286, 288, 289, 314, 315 or a homologous sequence thereof, wherein the homologous sequence is the same as SEQ ID Nos: Nos: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 46, 48, 50, 54, 56, 58, 286, 288, 289, 314 or 315 have at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence homology.

17. The antibody drug conjugate according to any one of claims 1 to 16, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The light chain variable region of the antibody comprises the sequence shown in SEQ ID Nos: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 46, 48, 50, 54, 56, 58, 286, 288, 289, 314 or 315.

18. The antibody drug conjugate according to any one of claims 1 to 17, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The heavy chain variable region and light chain variable region of the anti-FGFR2b antibody comprise the following combination, or comprise a sequence that is at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% homologous to the following sequence, or a combination thereof:

19. The antibody drug conjugate according to any one of claims 1 to 18, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The heavy chain variable region and light chain variable region of the anti-FGFR2b antibody are respectively the following combinations:

20. The antibody drug conjugate according to any one of claims 1 to 19, or a pharmaceutically acceptable salt, solvate or solvate of the salt thereof, characterized in that: The sequences of the heavy chain variable region and light chain variable region of the anti-FGFR2b antibody are SEQ ID NOs. 285 and 286, respectively.

21. The antibody drug conjugate according to any one of claims 1 to 20, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The heavy chain constant region of the anti-FGFR2b antibody is selected from human IgG, IgM, IgA, IgD, IgA constant regions or mutants of the above constant regions.

22. The antibody drug conjugate according to any one of claims 1 to 21, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The human IgG is IgG1, IgG2, IgG3 or IgG4.

23. The antibody drug conjugate according to any one of claims 1 to 22, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The humanized IgG is IgG1.

24. The antibody drug conjugate according to any one of claims 1 to 23, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The light chain constant region of the anti-FGFR2b antibody is selected from a human lambda constant region, a kappa constant region or a mutant of the above constant region.

25. The antibody drug conjugate according to any one of claims 1 to 24, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The light chain constant region of the anti-FGFR2b antibody is selected from a human kappa constant region.

26. The antibody drug conjugate according to any one of claims 1 to 25, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The amino acid sequence of the heavy chain of the anti-FGFR2b antibody comprises the sequence shown in SEQ ID NO: 292, or comprises a sequence that is greater than 70%, 75%, 80%, 85%, 90%, 95% or 99% identical to SEQ ID NO:

292.

27. The antibody drug conjugate according to any one of claims 1 to 26, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The amino acid sequence of the heavy chain of the anti-FGFR2b antibody comprises the sequence shown in SEQ ID NO:

292.

28. The antibody drug conjugate according to any one of claims 1 to 27, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The amino acid sequence of the light chain of the anti-FGFR2b antibody comprises the sequence shown in SEQ ID NO: 293, or comprises a sequence that is greater than 70%, 75%, 80%, 85%, 90%, 95% or 99% identical to SEQ ID NO:

293.

29. The antibody drug conjugate according to any one of claims 1 to 28, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The amino acid sequence of the light chain of the anti-FGFR2b antibody comprises the sequence shown in SEQ ID NO:

293.

30. The antibody drug conjugate according to any one of claims 1 to 29, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: p is 2.0-7.

0.

31. The antibody drug conjugate according to any one of claims 1 to 30, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: 3.0-6.0。 32. The antibody drug conjugate according to any one of claims 1 to 31, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: p is 3.0-4.

0.

33. The antibody drug conjugate according to any one of claims 1 to 32, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: p is 3.0, 3.4, 3.8 or 4.

0.

34. The antibody drug conjugate according to any one of claims 1 to 33, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: p is 4.

0.

35. The antibody drug conjugate according to any one of claims 1 to 34, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The cytotoxic agent is selected from irinotecan, irinotecan derivatives, Dxd, SN-38, gemcitabine, Monomethyl auristatin E (MMAE), Monomethyl auristatin F (MMAF), maytansinoids (e.g., Maytansine DM1, Maytansine DM 4), calicheamicin, MGBA (e.g., duocarmycin), doxorubicin, ricin, diphtheria toxin and other toxins, I131, interleukins, nuclides, tumor necrosis factors, chemokines or nanoparticles.

36. The antibody drug conjugate according to any one of claims 1 to 35, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The cytotoxic agent is Monomethyl auristatin E (MMAE) or Monomethyl auristatin F (MMAF).

37. The antibody drug conjugate according to any one of claims 1 to 36, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The linker is selected from 6-maleimidocaproyl (MC), maleimidopropionyl (MP), N-succinimidyl 4-(2-pyridylthio) pentanoate (SPP), 4-(N-maleimidomethyl)-cyclohexane-1-formyl (MCC), N-succinimidyl (4-iodo-acetyl) aminobenzoate (SIAB) or 6-maleimidocaproyl-valine-citrulline-p-aminobenzyloxycarbonyl (MC-vc-PAB).

38. The antibody drug conjugate according to any one of claims 1 to 37, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The linker is 6-maleimidocaproyl-valine-citrulline-p-aminobenzyloxycarbonyl (MC-vc-PAB).

39. The antibody drug conjugate according to any one of claims 1 to 37, or a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof, characterized in that: The antibodies include: 1) heavy chain variable complementary determining regions CDR1, CDR2, and CDR3, and light chain variable complementary determining regions CDR1, CDR2, and CDR3; wherein the sequence of the heavy chain variable complementary determining region 1 (CDR1) is set forth in SEQ ID NO: 59, the sequence of the heavy chain variable complementary determining region 2 (CDR2) is set forth in SEQ ID NO: 60, and the sequence of the heavy chain variable complementary determining region 3 (CDR3) is set forth in SEQ ID NO: 61; the sequence of the light chain variable complementary determining region 1 (CDR1) is set forth in SEQ ID NO: 62, the sequence of the light chain variable complementary determining region 2 (CDR2) is set forth in SEQ ID NO: 63, and the sequence of the light chain variable complementary determining region 3 (CDR3) is set forth in SEQ ID NO: 64; 2) a heavy chain variable framework region and a light chain variable framework region; wherein the sequence of the heavy chain variable framework region is as shown in SEQ ID NO: 285, and the sequence of the light chain variable framework region is as shown in SEQ ID NO: 286; and / or 3) a heavy chain and a light chain; wherein the sequence of the heavy chain is shown in SEQ ID NO: 292, and the sequence of the light chain is shown in SEQ ID NO: 293; The linker L is MC-vc-PAB; The cytotoxic agent D is MMAE.

40. A pharmaceutical composition comprising at least one of the antibody-drug conjugates of any one of claims 1-39, a pharmaceutically acceptable salt, solvate, or solvate of the salt thereof; optionally further comprising at least one of known chemotherapeutic drugs, immunotherapeutic drugs, and immunosuppressants for treating tumors; or optionally further comprising at least one pharmaceutically acceptable carrier, diluent, or excipient.

41. Use of the antibody-drug conjugate according to any one of claims 1 to 39, a pharmaceutically acceptable salt, solvate or solvate of the salt thereof, or the pharmaceutical composition according to claim 40 in the preparation of a medicament for preventing and / or treating a disease associated with FGFR2b.

42. The method for preventing and / or treating a disease associated with FGFR2b according to claim 41, characterized in that: The diseases associated with FGFR2b are squamous non-small cell lung cancer, gastric cancer, gastroesophageal junction cancer, triple-negative breast cancer, ovarian cancer, endometrial cancer, cervical cancer, colorectal cancer, intrahepatic bile duct cancer and / or pancreatic cancer.

43. The method for preventing and / or treating a disease associated with FGFR2b according to claim 41 or 42, characterized in that: The disease associated with FGFR2b is squamous non-small cell lung cancer, gastric cancer and / or gastroesophageal junction cancer.

44. A method for preventing and / or treating a disease associated with FGFR2b, comprising: administering to a subject in need thereof a preventively and / or therapeutically effective amount of at least one of the antibody-drug conjugates according to any one of claims 1 to 39, a pharmaceutically acceptable salt, solvate or solvate of the salt thereof, or the pharmaceutical composition according to claim 40.

45. The method for preventing and / or treating a disease associated with FGFR2b according to claim 44, characterized in that: The diseases associated with FGFR2b include squamous non-small cell lung cancer, gastric cancer, gastroesophageal junction cancer, triple-negative breast cancer, ovarian cancer, endometrial cancer, cervical cancer, colorectal cancer, intrahepatic bile duct cancer and / or pancreatic cancer.

46. ​​The method for preventing and / or treating a disease associated with FGFR2b according to claim 44 or 45, characterized in that: The disease associated with FGFR2b is squamous non-small cell lung cancer, gastric cancer and / or gastroesophageal junction cancer.

47. Use of the antibody drug conjugate according to any one of claims 1 to 39, its pharmaceutically acceptable salt, solvate or solvate of the salt, or the pharmaceutical composition according to claim 40 for preventing and / or treating diseases associated with FGFR2b.

48. The method for preventing and / or treating diseases associated with FGFR2b according to claim 47, characterized in that: The diseases associated with FGFR2b include squamous non-small cell lung cancer, gastric cancer, gastroesophageal junction cancer, triple-negative breast cancer, ovarian cancer, endometrial cancer, cervical cancer, colorectal cancer, intrahepatic bile duct cancer and / or pancreatic cancer.

49. The method for preventing and / or treating a disease associated with FGFR2b according to claim 47 or 48, wherein: The disease associated with FGFR2b is squamous non-small cell lung cancer, gastric cancer and / or gastroesophageal junction cancer.

Images