Anti-hGDF-15 antibody for use in treating cancer in patients in combination with cancer antigen-targeted drug conjugates that induce cancer cell stress.

Abstract

The present invention relates to the use of an anti-hGDF-15 antibody for treating a patient's cancer in combination with a cancer antigen-targeting drug conjugate, such as an antibody-drug conjugate that induces cancer cell stress.

Description

Technology Field

[0001] The present invention relates to the use of an anti-hGDF-15 antibody for treating cancer in patients in combination with a cancer antigen-targeting drug conjugate, such as an antibody-drug conjugate that induces cancer cell stress. Additionally, the present invention relates to a combination therapy for cancer treatment further comprising an immune checkpoint blocker. Furthermore, the present invention provides a combination product, a kit, and a pharmaceutical composition comprising an anti-hGDF-15 antibody and an antibody-drug conjugate. Background Technology

[0002] Antibody-drug conjugates (ADCs) have brought significant advancements to the field of targeted cancer therapy by combining the specificity of monoclonal antibodies with the potent cytotoxicity of small molecule drugs. This therapeutic approach leverages the ability of antibodies to selectively bind to antigens primarily expressed on the surface of cancer cells to deliver attached cytotoxic drugs directly to the tumor site. The targeted delivery of cytotoxic drugs reduces the systemic toxicity commonly associated with traditional chemotherapy by minimizing impact on healthy cells.

[0003] An ADC typically consists of three components: a monoclonal antibody specific to a tumor-associated antigen, a cytotoxic drug (payload), and a linker that stably attaches the drug to the antibody. When the ADC-antibody complex binds to the target antigen, it is internalized by cancer cells, causing the cytotoxic drug to be released and inducing cellular stress or apoptosis in the cancer cells.

[0004] Recent observations indicate that ADCs can stimulate a potent anti-cancer immune response by inducing cancer cell stress or apoptosis, in addition to direct cytotoxic effects. For example, ADCs have been shown to induce a form of apoptosis known as immunogenic cell death (ICD) in cancer cells (Cao et al., 2017; Rios-Doria et al., 2017; Bauzon et al., 2019; D'Amico et al., 2019; Boshuizen et al., 2021; Devra Olson et al., 2022).

[0005] ICDs are characterized particularly by the release of various danger signals and cancer antigens, which can enhance the mobilization and activation of immune cells. The exposure of caleticulin on the cell surface and the release of ATP and HMGB1 are phenomena involved in this process, acting as 'eat me' signals to dendritic cells and other antigen-presenting cells. These immune cells process and present cancer antigens, effectively triggering a secondary immune response against cancer (Kroemer et al., 2022).

[0006] Despite the therapeutic potential of targeted cancer therapy, many patients still die from cancer, and the need to improve cancer therapy using ADCs remains in the field of technology. Prior art literature

[0007] Bauzon, M. et al. (2019) 'Maytansine-bearing antibody-drug conjugates induce in vitro hallmarks of immunogenic cell death selectively in antigen-positive target cells', OncoImmunology, 8(4), p. e1565859.Boshuizen, J. et al. (2021) 'Cooperative Targeting of Immunotherapy-Resistant Melanoma and Lung Cancer by an AXL-Targeting Antibody-Drug Conjugate and Immune Checkpoint Blockade', Cancer Research, 81(7), pp. 1775-1787.D'Amico, L. et al. (2019) 'A novel anti-HER2 anthracycline-based antibody-drug conjugate induces adaptive anti-tumor immunity and potentiates PD-1 blockade in breast cancer', Journal for ImmunoTherapy of Cancer, 7(1), p. 16.Devra Olson et al. (2022) '1187 Enfortumab vedotin induces immunogenic cell death, elicits antitumor immune memory, and shows enhanced preclinical activity in combination with immune checkpoint inhibitors', Journal for ImmunoTherapy of Cancer, 10(Suppl 2), p. A1231.Olson, D. et al.(2022) '1187 Enfortumab vedotin induces immunogenic cell death, elicits antitumor immune memory, and shows enhanced preclinical activity in combination with immune checkpoint inhibitors', in Regular and Young Investigator Award Abstracts. SITC 37th Annual Meeting (SITC 2022) Abstracts, BMJ Publishing Group Ltd, pp. A1229-A1229.Rios-Doria, J. et al. (2017) 'Antibody-Drug Conjugates Bearing Pyrrolobenzodiazepine or Tubulysin Payloads Are Immunomodulatory and Synergize with Multiple Immunotherapies', Cancer Research, 77(10), pp. 2686-2698.Cao, A.T. et al. (2017) 'Abstract 5588: Brentuximab vedotin-driven immunogenic cell death enhances antitumor immune responses, and is potentiated by PD1 inhibition in vivo', Cancer Research, 77(13_Supplement), p. 5588.Kroemer, G. et al. (2022) 'Immunogenic cell stress and death', Nature Immunology, 23(4), pp. 487-500.Schuberth-Wagner, C. et al.(2023) Immuno-suppressive role of tumour-derived GDF-15 on myeloid cells;; Annals of Oncology, Volume 34, S190Zhou Z, Li W, Song Y, Wang L, Zhang K, Yang J, Zhang W, Su H, Zhang Y. Growth differentiation factor-15 suppresses maturation and function of dendritic cells and inhibits tumor-specific immune response. PLoS One. 2013 Nov 13;8(11):e78618.Haake M, Haack B, Schafer T, Harter PN, Mattavelli G, Eiring P, Vashist N, Wedekink F, Genssler S, Fischer B, Dahlhoff J, Mokhtari F, Kuzkina A, Welters MJP, Benz TM, Sorger L, Thiemann V, Almanzar G, Selle M, Thein K, Spath J, Gonzalez MC, Reitinger C, Ipsen-Escobedo A, Wistuba-Hamprecht K, Eichler K, Filipski K, Zeiner PS, Beschorner R, Goedemans R, Gogolla FH, Hackl H, Rooswinkel RW, Thiem A, Roche PR, Joshi H, Puhringer D, Wockel A, Diessner JE, Rudiger M, Leo E, Cheng PF, Levesque MP, Goebeler M, Sauer M, Nimmerjahn F, Schuberth-Wagner C, von Felten S, Mittelbronn M, Mehling M, Beilhack A, van der Burg SH, Riedel A, Weide B, Dummer R, Wischhusen J. Tumor-derived GDF-15 blocks LFA-1 dependent T cell recruitment and suppresses responses to anti-PD-1 treatment. Nat Commun. 2023 Jul 20;14(1):4253.Collins, Denis M., et al. "Acquired resistance to antibody-drug conjugates." Cancers 11.3 (2019): 394.Tiligada, E. "Chemotherapy: induction of stress responses." Endocrine-related cancer 13.Supplement_1 (2006): S115-S124.

[0008] The inventors considered that they could address unmet clinical needs in the field of cancer therapy by investigating the role of growth differentiation factor-15 (GDF-15), a stress-induced cytokine, in cancer targeted therapy.

[0009] Unexpectedly, the inventors discovered that cancer antigen-targeting drug conjugates, such as ADCs, induce GDF-15 production in cancer cells. Specifically, the inventors discovered that increasing the concentrations of trastuzumab deruxtecan (anti-HER2) and sacituzumab govitecan (anti-TROP2) as ADCs induces GDF-15 production, accompanied by the induction of apoptosis, as evidenced by a decrease in cell viability. Similarly, the inventors discovered that GDF-15 is also induced by monomethylauristatin E (MMAE), a microtubule inhibitor that inhibits mitotic cell division and ultimately induces apoptosis.

[0010] GDF-15 is known to inhibit several key processes in anticancer immunity. It impairs the activation and maturation of antigen-presenting cells (APCs), including macrophages and dendritic cells, and interferes with the effective sensitization and subsequent expansion of T cells by DCs (Schuberth-Wagner, C., et al., 2023; Zhou, Zhizhong, et al. 2023). Beyond these effects, GDF-15 significantly reduces T cell adhesion to endothelial cells, thereby hindering tumor invasion and extravasation, which are essential for direct cancer attack (Haake, Markus, et al. 2023).

[0011] In addition, it is known that the overexpression of GDF-15 contributes to an immunosuppressive environment, helping cancer cells evade immune surveillance and response.

[0012] In short, the experimental evidence presented in this application supports that antibodies conjugated with cancer antigen-targeting drug conjugates, such as topoisomerase I inhibitors or microtubule inhibitors, induce cancer cell stress and GDF-15 production.

[0013] Although not intended to be a theoretical conclusion, the production of GDF-15 appears to be a stress response in cancer cells resulting from treatment with ADCs. Furthermore, the presence of GDF-15 in the tumor microenvironment is expected to reduce the efficacy of cancer therapies using antigen-targeted drug conjugates, such as ADCs, by counteracting the immune-stimulating activities of cancer cell stress or apoptosis. This reduction in efficacy applies particularly to the synergistic activity of antigen-targeted drug conjugates with immunotherapies, such as immune checkpoint blockades.

[0014] Accordingly, based on the findings of the inventors, GDF-15 appears to reduce the therapeutic efficacy of antigen-targeted drug conjugates and contribute to the development of therapeutic resistance.

[0015] Clearly, the inventors demonstrate that neutralizing GDF-15 improves the anti-tumor efficacy of ADC therapy, particularly achieving tumor regression, enhanced immune infiltration, and increased activation of T cells and macrophages. Furthermore, the experimental data provided in this application demonstrates that GDF-15 reduces the therapeutic activity of combination therapy using ADCs and checkpoint blockades / inhibitors, and that neutralizing GDF-15 enhances the efficacy of combination therapy using ADCs and immune checkpoint inhibitors / blockades.

[0016] Therefore, neutralization of GDF-15 is expected to improve synergistic activity with immunotherapies, such as ADC activation and immune checkpoint blockade, by counteracting GDF-15-mediated therapeutic resistance and the inhibition of ADC-induced immune cell activation.

[0017] In light of the foregoing, the present invention relates to an anti-hGDF-15 antibody or its antigen-binding portion (i.e., its hGDF-15 binding portion) for use in the treatment of cancer in patients in combination with a cancer antigen-targeting drug conjugate that induces cancer cell stress. The anti-hGDF-15 antibody and its antigen-binding portion (i.e., its hGDF-15 binding portion) are expected to act synergistically with the cancer antigen-targeting drug conjugate therapy and enhance an adaptive anticancer immune response, thereby improving therapeutic efficacy.

[0018] Accordingly, the present invention has the potential to establish long-term immune surveillance to potentially prevent cancer recurrence and improve patient outcomes in various types of cancer.

[0019] Accordingly, the present invention provides the following preferred embodiments:

[0020] 1. An anti-hGDF-15 antibody or its antigen-binding portion for use in a method of treating cancer in human patients in combination with a cancer antigen-targeting drug conjugate that induces cancer cell stress.

[0021] 2. An anti-hGDF-15 antibody or its antigen-binding portion for use according to Item 1, in which cancer cell stress is immunogenic.

[0022] 3. An anti-hGDF-15 antibody or its antigen-binding portion for use according to item 1 or 2, in which cancer cell stress is associated with the induction of hGDF-15 expression in a patient.

[0023] 4. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 1 to 3, wherein the cancer antigen-targeting drug conjugate induces apoptosis of cancer cells.

[0024] 5. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 1 to 4, wherein cancer cell death is immunogenic cell death (ICD).

[0025] 6. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 1 to 5, in which immunogenic cell death (ICD) is associated with the induction of hGDF-15 expression in a patient.

[0026] 7. An anti-hGDF-15 antibody or an antigen-binding portion thereof for use according to any one of items 1 to 6, wherein the anti-hGDF-15 antibody or its antigen-binding portion is a neutralizing anti-hGDF-15 antibody or its antigen-binding portion.

[0027] 8. An anti-hGDF-15 antibody or an antigen-binding portion thereof for use according to any one of items 1 to 7, wherein the anti-hGDF-15 antibody or its antigen-binding portion comprises a heavy chain variable domain comprising a CDR1 region represented by the amino acid sequence presented in SEQ ID NO. 1, a CDR2 region represented by the amino acid sequence presented in SEQ ID NO. 2, and a CDR3 region represented by the amino acid sequence presented in SEQ ID NO. 3, and a light chain variable domain comprising a CDR1 region represented by the amino acid sequence presented in SEQ ID NO. 4, a CDR2 region represented by the amino acid sequence ser-ala-ser, and a CDR3 region represented by the amino acid sequence presented in SEQ ID NO. 5.

[0028] 9. An anti-hGDF-15 antibody or an antigen-binding portion thereof for use according to any one of items 1 to 8, the anti-hGDF-15 antibody or the antigen-binding portion thereof comprising a heavy chain variable domain having at least 90% identity, preferably at least 95% identity, more preferably at least 98% identity, with the amino acid sequence represented by SEQ ID NO. 6 or the amino acid sequence presented in SEQ ID NO. 6, and a light chain variable domain having at least 90% identity, preferably at least 95% identity, more preferably at least 98% identity, with the amino acid sequence represented by SEQ ID NO. 7 or the amino acid sequence presented in SEQ ID NO. 7.

[0029] 10. An anti-hGDF-15 antibody or an antigen-binding portion thereof for use according to any one of items 1 to 9, comprising a heavy chain having an amino acid sequence having at least 90% identity, preferably at least 95% identity, more preferably at least 98% identity, with respect to the amino acid sequence represented by SEQ ID NO. 8 or the amino acid sequence presented in SEQ ID NO. 8, and a light chain having an amino acid sequence having at least 90% identity, preferably at least 95% identity, more preferably at least 98% identity, with respect to the amino acid sequence represented by SEQ ID NO. 9 or the amino acid sequence presented in SEQ ID NO. 9.

[0030] 11. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 1 to 10, wherein the anti-hGDF-15 antibody or its antigen-binding portion comprises a heavy chain having an amino acid sequence represented by SEQ ID NO. 8 and a light chain having an amino acid sequence represented by SEQ ID NO. 9.

[0031] 12. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 1 to 10, wherein the anti-hGDF-15 antibody or its antigen-binding portion competes with the antibody defined in item 11 for specific binding to hGDF-15.

[0032] 13. An anti-hGDF-15 antibody or an antigen-binding portion thereof for use according to any one of items 1 to 12, wherein the anti-hGDF-15 antibody or the antigen-binding portion thereof binds to a stereotypical or discontinuous epitope on hGDF-15 composed of the amino acid sequences of SEQ ID NO. 12 and SEQ ID NO. 13.

[0033] 14. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 1 to 7, wherein the anti-hGDF-15 antibody or its antigen-binding portion is visugromab, ponsegromab, or rilologrotug.

[0034] 15. An anti-hGDF-15 antibody or an antigen-binding portion thereof for use according to any one of items 1 to 14, wherein the anti-hGDF-15 antibody or the antigen-binding portion thereof is bisugromab.

[0035] 16. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 1 to 15, comprising a cancer antigen-targeting drug conjugate comprising a cancer cell stress-inducing drug linked to a cancer antigen-targeting portion.

[0036] 17. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 16, wherein the drug is an anticancer agent.

[0037] 18. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 16 or 17, wherein the drug is a chemotherapy drug.

[0038] 19. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with any one of items 16 to 18, wherein the drug is a cytotoxic drug.

[0039] 20. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 19, wherein the cytotoxic drug is selected from the group consisting of microtubule inhibitors, topoisomerase I or II inhibitors, DNA-damaging agents, protein synthesis inhibitors, RNA polymerase III inhibitors, transcription inhibitors, apoptosis inducers, NAMPT inhibitors, proteasome inhibitors, kinase inhibitors, PROTACs, NIR-PIT drugs, and immune-activating substances.

[0040] 21. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with item 19 or 20, wherein the cytotoxic drug is a microtubule inhibitor or a topoisomerase I or II inhibitor.

[0041] 22. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 19 to 21, wherein the cytotoxic drug is a microtubule inhibitor.

[0042] 23. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 20 to 22, wherein the microtubule inhibitor is selected from the group consisting of monomethylauristatin E (MMAE), monomethylauristatin F (MMAF), methansin (DM-1), auristatin F-HPA, auristatin-0101, DM21, DM4, metansinoid, eribulin, and SC209.

[0043] 24. An anti-hGDF-15 antibody or its antigen-binding portion for use according to item 20 or 21, wherein the topoisomerase I inhibitor is selected from the group consisting of Dxd, SN-38, irinotecan, topotecan, rubitecan, exatecan, belotecan, MLN576, and camptothecin.

[0044] 25. An anti-hGDF-15 antibody or its antigen-binding portion for use according to Item 20 or 21, wherein the topoisomerase II inhibitor is selected from the group consisting of etoposide, idarubicin, mitoxantrone, PNU-159682, daunorubicin, teniposide, epirubicin, and doxorubicin.

[0045] 26. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 20, wherein the DNA damaging agent is selected from the group consisting of caliceamycin, SG3199 / PBD dimer, PBD, and indolinobenzodiazepine (IGN).

[0046] 27. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 20, wherein the protein synthesis inhibitor is selected from the group consisting of PE38, geldanamycin, tylanstatin A, and carmapicin B analogs.

[0047] 28. An anti-hGDF-15 antibody or its antigen-binding portion for use according to Item 20, wherein the RNA polymerase III inhibitor is selected from the group consisting of α-amanitine, beta-amanitine, phalloidin, trichothecene T-2, velucarin A and loridin A.

[0048] 29. An anti-hGDF-15 antibody or its antigen-binding portion for use according to Item 20, wherein the transcription inhibitor is selected from the group consisting of triptolide, ST7464AA1, vorinobstat, and dashinostat.

[0049] 30. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 20, wherein the apoptosis inducer is a BCL-XL inhibitor such as clezutoclax.

[0050] 31. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 20, wherein the NAMPT inhibitor is an FK-866 analog.

[0051] 32. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 20, wherein the proteasome inhibitor is selected from the group consisting of bortezomib, carfilzomib, ixazomib, and carmapicin B analogs.

[0052] 33. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 20, wherein the kinase inhibitor is selected from the group consisting of genistein, neolympostine, dasatinib, and staurosporine.

[0053] 34. An anti-hGDF-15 antibody or its antigen-binding portion for use according to Item 20, wherein the PROTAC is selected from the group consisting of BET / BRD degraders (GNE-987, MZ1 analogs, BRD4 / VHL, BRD4 / CRBN), ERa degraders (ERa / XIAP, ERa / VHL), TGFbR2 degraders (TGFbR2 / VHL), BRM degraders (BRL / VHL), and GSTP1 degraders (SMOL006).

[0054] 35. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 20, wherein the NIR-PIT drug is selected from water-soluble phthalocyanine derivatives such as the silicon phthalocyanine derivative IR700 (IRDye700DX).

[0055] 36. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 20, wherein the immune-activating substance is selected from the group consisting of STING agonists, TLR7 and / or TLR8 agonists and zuvotolimod.

[0056] 37. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 16 to 19, wherein the drug inducing cancer cell stress is selected from the group consisting of MMAE, MMAF / auristatin-F, Dxd, DM-1, SN-38, camptothecin, and DM-4.

[0057] 38. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 37, wherein the drug inducing cancer cell stress is MMAE.

[0058] 39. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 16 to 38, wherein the drug is linked to a cancer antigen-targeting portion through a cleavable linker.

[0059] 40. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 16 to 38, wherein the drug is linked to the cancer antigen-targeting portion via a non-cleavable linker.

[0060] 41. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 16 to 40, wherein the cancer antigen-targeting portion of a cancer antigen-targeting drug conjugate binds to cancer cells.

[0061] 42. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 16 to 41, wherein the cancer antigen-targeting portion is a ligand, peptide, or antibody.

[0062] 43. An anti-hGDF-15 antibody for use in accordance with Item 42, wherein the cancer antigen-targeting portion is an antibody, or its antigen-binding portion.

[0063] 44. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 41 to 43, wherein the cancer antigen-targeting portion binds to a target antigen on a cancer cell.

[0064] 45. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 44, where the target antigen is not hGDF-15.

[0065] 46. ​​Target antigens are Nectin-4, HER2, Trop-2, TF (CD142), CD30, CD22, CD79b, BCMA, CD19, FRα, CD33, LIV-1, HER3, CD25, NaPi2b, B7-H4, c-Met, B7-H3, B7-H4, PTK7, ADAM9, CEACAM5, 5T4, ALK, AXL, GRP20, CDH6, TA-MUC1, KAAG1, DLK1, DLL3, SLAMF7, CA125, C4.4A / LYPD3, CDH3, CDH6, CAIX, CD20, CD26 / DPP4, CD37, CD38, CD138, CD46, ICAM4 / CD54, CD56 / NCAM1, CD70, CD73, CD74, CD205, With CD248, C-KIT, CLDN6, CLDN18.2, CLL-1, RET, CRIPTO, DLK-1, DLL3, EGFR, CD105, ENPP3, EPCAM, EPHA2, FAP, FGFR2 / CD332, FLT3, GDNF / GFRA1, GPC2, GPNMB, Guanylyl Cyclase (GCC), IGF-1R, ITGAV, Sialyl-di-Lewis, LGR5, LIV1A, LRRC15, MSLN, STEAP1, PSMA, TMEFF2, NOTCH3, PTK7, SLC44A4, SLC46A3, SLITRK6, TIM-1, LY6E, Cadherin, PD-L1, CD228, FOLR1, CTLA4, GPR20, HGFR, CD123, PSMA, ROR1, and ETBR An anti-hGDF-15 antibody or its antigen-binding portion for use according to item 44 or 45, selected from the group formed.

[0066] 47. An anti-hGDF-15 antibody or its antigen-binding portion for use according to Item 46, wherein the target antigen is selected from Nectin-4, HER2, Trop-2, TF (CD142), CD30, CD22, CD79b, BCMA, CD19, FRα and CD33.

[0067] 48. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 1 to 47, wherein the cancer antigen-targeting drug conjugate is an antibody-drug conjugate (ADC).

[0068] 49. ADCs include Enfortumab vedotin, trastuzumab deruxtecan, trastuzumab emtansine, sacituzumab govitecan, tisotumab vedotin, Brentuximab vedotin, Inotuzumab ozogamicin, Moxetumomab pasudotox, Polatuzumab vedotin, Belantamab mafodotin, Loncastuximab tesirine, and Mirvetuximab sorabtansine-gynx. soravtansine-gynx), Gemtuzumab ozogamicin, disitamab vedotin, ladiratuzumab vedotin, datopotamab deruxtecan, patritumab deruxtecan, camidanlumab tesirine, upifitamab rilsodotin, XMT-1592, XMT-1660, XMT-2056, telisotuzumab vedotin, ABBV-400, mirzotamab clezutoclax, cofetuzumab felidotin pelidotin), MORAb-202 (parletuzumab), STRO-002, IMGN632, IMGC936, ASP-1929, isacituzumab govitecan,An anti-hGDF-15 antibody for use according to Item 48, selected from the group consisting of SKB264 and tusamitamab ravtansine,

[0069] 50. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 49, wherein the ADC is selected from the group consisting of enfortumab vedotin, trastuzumab deruxtecan, trastuzumab emtansine, datopotamab deruxtecan and sacituzumab govitecan.

[0070] 51. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 1 to 50, wherein the cancer is a solid tumor.

[0071] 52. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 51, wherein the cancer is selected from the group consisting of urothelial carcinoma (UC), non-small cell lung cancer (NSCLC), pancreatic cancer, head and neck cancer, breast cancer, colorectal cancer, anal cancer, gastric cancer, liver cancer, biliary tract cancer, ovarian cancer, prostate cancer, gastric cancer, esophageal cancer, kidney cancer, thyroid cancer, endometrial cancer, cervical cancer, testicular cancer, melanoma, and skin cancer.

[0072] 53. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with item 51 or 52, wherein the cancer is urothelial carcinoma and the drug of the cancer antigen-targeting drug conjugate is a microtubule inhibitor.

[0073] 54. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 53, wherein the microtubule inhibitor is MMAE.

[0074] 55. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with item 53 or 54, wherein the cancer antigen-targeting drug conjugate is enfortumab vedotin.

[0075] 56. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with item 51 or 52, wherein the cancer is breast cancer and the drug of the cancer antigen-targeting drug conjugate is a topoisomerase I inhibitor.

[0076] 57. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 56, wherein the topoisomerase I inhibitor is Dxd.

[0077] 58. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with item 56 or 57, wherein the cancer antigen-targeting drug conjugate is trastuzumab deruxtecan.

[0078] 59. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with item 51 or 52, wherein the cancer is breast cancer and the drug of the cancer antigen-targeting drug conjugate is a topoisomerase I inhibitor.

[0079] 60. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 59, wherein the topoisomerase I inhibitor is SN-38.

[0080] 61. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with item 59 or 60, wherein the cancer antigen-targeting drug conjugate is sacituzumab govitecan.

[0081] 62. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with item 51 or 52, wherein the cancer is NSCLC and the drug of the cancer antigen-targeting drug conjugate is a topoisomerase I inhibitor.

[0082] 63. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with Item 62, wherein the topoisomerase I inhibitor is Dxd.

[0083] 64. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with item 62 or 63, wherein the cancer antigen-targeting drug conjugate is trastuzumab deruxtecan.

[0084] 65. An anti-hGDF-15 antibody or an antigen-binding portion thereof for use in a method of treating cancer in human patients, wherein the anti-hGDF-15 antibody or the antigen-binding portion thereof is administered in combination with at least one antibody-drug conjugate.

[0085] 66. An anti-hGDF-15 antibody or an antigen-binding portion thereof for use in accordance with Item 65, such as as defined in any one of Items 7 to 15.

[0086] 67. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with item 65 or 66, as defined in any one of items 48 to 50.

[0087] 68. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with any one of items 65 to 67, wherein the antibody-drug conjugate induces cancer cell stress.

[0088] 69. An anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with any one of items 65 to 68, such as the cancer defined in item 51 or 52.

[0089] 70. Anti-hGDF-15 antibody or its antigen-binding portion for use in accordance with any one of items 65 to 69, as defined in any one of items 53 to 64 for cancer and drugs.

[0090] 71. An anti-hGDF-15 antibody or its antigen-binding portion and an antibody-drug conjugate (ADC) for use according to any one of items 65 to 70, wherein the anti-hGDF-15 antibody or its antigen-binding portion and the antibody-drug conjugate (ADC) are administered in combination with an immune checkpoint blocker (ICB), optionally, the ICB is selected from the list of PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors and LAG-3 inhibitors, preferably a PD-1 inhibitor or a PD-L1 inhibitor.

[0091] 72. An anti-hGDF-15 antibody or its antigen-binding portion for use according to Item 71, wherein the ICB is selected from the group consisting of nivolumab, atezolizumab, pembrolizumab, avelumab, cemiplimab, durvalumab, dostalimab, retipanlimab, toripalimab, syntilimab, tislellizumab, camrelizumab, valstilimab, envapolimab, pakmilimab, cosivelimab and spartalizumab, preferably nivolumab or pembrolizumab.

[0092] 73. An antibody-drug conjugate (ADC) for use in a method of treating cancer in human patients, wherein the antibody-drug conjugate is administered in combination with an anti-hGDF-15 antibody or its antigen-binding portion.

[0093] 74. An antibody-drug conjugate (ADC) for use according to Item 73, wherein the anti-hGDF-15 antibody or its antigen-binding portion is as defined in any one of Items 7 through 15.

[0094] 75. An antibody-drug conjugate (ADC) for use according to item 73 or 74, as defined in any one of items 48 to 50.

[0095] 76. An antibody-drug conjugate (ADC) for use according to any one of items 73 to 75, wherein the antibody-drug conjugate induces cancer cell stress.

[0096] 77. An antibody-drug conjugate (ADC) for use according to any one of items 73 through 76, such as cancer as defined in item 51 or 52.

[0097] 78. Antibody-drug conjugates (ADCs) for use according to any one of items 73 through 77, wherein the cancer and the drug are as defined in any one of items 53 through 64.

[0098] 79. An antibody-drug conjugate (ADC) for use according to any one of items 73 to 78, wherein the antibody-drug conjugate (ADC) and an anti-hGDF-15 antibody or its antigen-binding portion is administered in combination with an immune checkpoint blocker (ICB), said ICB is selected from the list of PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors and LAG-3 inhibitors, preferably a PD-1 inhibitor or a PD-L1 inhibitor.

[0099] 80. An antibody-drug conjugate (ADC) for use according to item 79, wherein the ICB is selected from the group consisting of nivolumab, atezolizumab, pembrolizumab, avelumab, cemiplimab, durvalumab, dostalimab, retipanlimab, toripalimab, syntilimab, tislellizumab, camrelizumab, valstilimab, envapolimab, pakmilimab, cocibelimab and spartalizumab, preferably nivolumab or pembrolizumab.

[0100] 81. An anti-hGDF-15 antibody or a combination product consisting of its antigen-binding portion and an antibody-drug conjugate for use in a method of treating cancer in human patients.

[0101] 82. A combination product for use according to Item 81, wherein the anti-hGDF-15 antibody or its antigen-binding portion is as defined in any one of Items 7 to 15.

[0102] 83. An antibody-drug conjugate combination product for use according to item 81 or 82, as defined in any one of items 48 to 50.

[0103] 84. A combination product for use according to any one of items 81 to 83, wherein the antibody-drug conjugate induces cancer cell stress.

[0104] 85. A combination product for use according to any one of items 81 through 84, such as as defined in item 51 or 52.

[0105] 86. A combination product for use according to any one of items 81 through 85, wherein the cancer and the drug are as defined in any one of items 53 through 64.

[0106] 87. A combination product for use according to any one of items 81 to 86, wherein an antibody-drug conjugate (ADC) and an anti-hGDF-15 antibody or its antigen-binding portion is administered in combination with an immune checkpoint blocker (ICB), optionally, the ICB is selected from the list of PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors and LAG-3 inhibitors, preferably a PD-1 inhibitor or a PD-L1 inhibitor.

[0107] 88. A combination product for use according to item 87, wherein the ICB is selected from the group consisting of nivolumab, atezolizumab, pembrolizumab, avelumab, cemiplimab, durvalumab, dostalimab, retipanlimab, toripalimab, syntilimab, tislellizumab, camrelizumab, valstilimab, envapolimab, pakmilimab, cocibelimab, and spartalizumab, preferably nivolumab or pembrolizumab.

[0108] 89. A kit comprising an anti-hGDF-15 antibody or its antigen-binding portion and an antibody-drug conjugate.

[0109] 90. A kit according to item 89 in which the anti-hGDF-15 antibody or its antigen-binding portion is as defined in any one of items 7 to 15.

[0110] 91. A kit according to item 89 or 90 in which an antibody-drug conjugate is defined in any one of items 48 to 50.

[0111] 92. A kit according to items 89 to 91 in which an antibody-drug conjugate induces cancer cell stress.

[0112] 93. A kit according to items 89 to 92, further comprising an immune checkpoint blocker (ICB), optionally, wherein the ICB is selected from the list of PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors and LAG-3 inhibitors, preferably a PD-1 inhibitor or a PD-L1 inhibitor.

[0113] 94. A kit according to item 93 in which the ICB is selected from the group consisting of nivolumab, atezolizumab, pembrolizumab, avelumab, cemiplimab, durvalumab, dostalimab, retipanlimab, toripalimab, syntilimab, tislellizumab, camrelizumab, valstilimab, envapolimab, pakmilimab, cocibelimab, and spartalizumab, preferably nivolumab or pembrolizumab.

[0114] 95. A pharmaceutical composition comprising a combination of an anti-hGDF-15 antibody and an antibody-drug conjugate.

[0115] 96. A pharmaceutical composition according to Item 95 in which an anti-hGDF-15 antibody or its antigen-binding portion is as defined in any one of Items 7 to 15.

[0116] 97. A pharmaceutical composition according to item 95 or 96, wherein the antibody-drug conjugate is as defined in any one of items 48 to 50.

[0117] 98. A pharmaceutical composition according to items 95 to 97 in which an antibody-drug conjugate induces cancer cell stress.

[0118] 99. A pharmaceutical composition according to any one of items 95 to 98, further comprising an immune checkpoint blocker (ICB), optionally, wherein the ICB is selected from the list of PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors and LAG-3 inhibitors, preferably a PD-1 inhibitor or a PD-L1 inhibitor.

[0119] 100. A pharmaceutical composition according to item 99 in which the ICB is selected from the group consisting of nivolumab, atezolizumab, pembrolizumab, avelumab, cemiplimab, durvalumab, dostalimab, retipanlimab, toripalimab, syntilimab, tislellizumab, camrelizumab, valstilimab, envapolimab, pakmilimab, cocibelimab, and spartalizumab, preferably nivolumab or pembrolizumab.

[0120] 101. Use of an anti-hGDF-15 antibody or its antigen-binding portion in combination with a cancer antigen-targeting drug conjugate in the manufacture of a drug for the treatment of cancer in human patients, optionally wherein the cancer antigen-targeting drug conjugate is an antibody-drug conjugate (ADC).

[0121] 102. Use of an anti-hGDF-15 antibody or its antigen-binding portion in the manufacture of a drug for the treatment of cancer in human patients, wherein the drug further comprises a cancer antigen-targeting drug conjugate, and optionally, the cancer antigen-targeting drug conjugate is an antibody-drug conjugate (ADC).

[0122] 103. Use of an antibody-drug conjugate (ADC) in the manufacture of a drug for the treatment of cancer in human patients, wherein the drug further comprises an anti-hGDF-15 antibody or an antigen-binding portion thereof.

[0123] 104. Any of items 101 to 103, wherein an anti-hGDF-15 antibody or its antigen-binding portion and an antibody-drug conjugate (ADC) are administered in combination with an immune checkpoint blocker (ICB), optionally, the ICB is selected from the list of PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors and LAG-3 inhibitors, preferably a PD-1 inhibitor or a PD-L1 inhibitor, and additionally optionally, the ICB is selected from the group consisting of nivolumab, atezolizumab, pembrolizumab, avelumab, cemiplimab, durvalumab, dostalimab, retipanlimab, toripalimab, syntilimab, tislellizumab, camrelizumab, valstilimab, envapolimab, pakmilab, cocibelimab and spartalizumab, preferably nivolumab or pembrolizumab. Usage according to one item.

[0124] 105. An anti-hGDF-15 antibody or its antigen-binding portion as defined in any one of items 7 to 15, or a use according to any one of items 101 to 104.

[0125] 106. Use of an antibody-drug conjugate according to any one of items 101 to 105, as defined in any one of items 48 to 50.

[0126] 107. Use according to any one of items 101 to 106 in which a cancer antigen-targeting drug conjugate or antibody-drug conjugate induces cancer cell stress.

[0127] 108. An anti-hGDF-15 antibody or its antigen-binding portion for use according to any one of items 1 to 72, wherein the anti-hGDF-15 antibody or its antigen-binding portion and the antigen-targeting drug conjugate or antibody-drug conjugate are administered at the same or different times; an antibody-drug conjugate (ADC) for use according to any one of items 73 to 80; a combination product for use according to any one of items 81 to 88; or a use according to any one of items 101 to 107. Brief explanation of the drawing

[0128] Fig. 1: Induction of apoptosis by increasing concentration of sacituzumab govitecan As shown in the figure, the cell viability of MCF-7 or BxPC-3 cancer cell lines was quantitatively evaluated upon treatment with sacituzumab or sacituzumab govitecan. BxPC-3 alone and MCF-7 alone controls are indicated by bars in the upper left of the figure. Fig. 2: In vitro induction of GDF-15 release by sacituzumab govitecan GDF-15 induction by sacituzumab or sacituzumab govitecan in cancer cell lines (MCF-7 or BxPC-3) was evaluated by ELISA assay. The untreated control group is indicated on the left side of the figure (concentration="0"). Fig. 3: Induction of apoptosis by increasing concentration of SN-38 When treated with SN-38, the cell viability of cancer cell lines (SK-MEL-5, MCF-7, BxPC-3, PC-3) was quantitatively evaluated. Fig. 4: Induction of in vitro GDF-15 release by SN-38 GDF-15 induction by SN-38 in cancer cell lines (SK-MEL-5, MCF-7, BxPC-3, PC-3) was evaluated by ELISA analysis. Fig. 5: Induction of apoptosis by increased concentration of camptothecin The cell viability of cancer cell lines (SK-MEL-5, MCF-7, BxPC-3, PC-3) was quantitatively evaluated upon treatment with camptothecin. The control groups—MCF-7 alone, PC-3 alone, BxPC-3 alone, and SK-MEL-5 alone—are shown as bars in the upper left of the figure. Fig. 6: In vitro induction of GDF-15 release by camptothecin GDF-15 induction by camptothecin in cancer cell lines (SK-MEL-5, MCF-7, BxPC-3, PC-3) was evaluated by ELISA analysis. Fig. 7: Induction of apoptosis by increasing concentration of trastuzumab deruxtecan The cell viability of cancer cell lines (SK-MEL-5, MCF-7, BxPC-3, PC-3) was quantitatively evaluated upon treatment with trastuzumab or trastuzumab deruxtecan. PC-3, BxPC-3, MCF-7, and the SK-MEL-5 control group are represented by bars on the left side of the figure. Fig. 8: In vitro induction of GDF-15 release by trastuzumab deruxtecan GDF-15 induction by trastuzumab or trastuzumab deruxtecan in cancer cell lines (SK-MEL-5, MCF-7, BxPC-3, PC-3) was evaluated by ELISA analysis. The untreated control group is shown on the left side of each panel in the figure (concentration="0"). Fig. 9: Induction of apoptosis by increasing concentration of Dxd Cell viability was quantitatively evaluated in cancer cell lines (SK-MEL-5, MCF-7, BxPC-3, PC-3) upon Dxd treatment. The control groups—MCF-7 alone, PC-3 alone, BxPC-3 alone, and SK-MEL-5 alone—are represented by bars in the upper left of the figure. Fig. 10: Induction of in vitro GDF-15 release by Dxd GDF-15 induction by Dxd in cancer cell lines (SK-MEL-5, MCF-7, BxPC-3, PC-3) was evaluated by ELISA analysis. Fig. 11: Induction of apoptosis by increasing concentration of MMAE Cell viability was quantitatively evaluated for cancer cell lines (Panc02, MCF-7, BxPC-3, PC-3) treated with MMAE. Panc02 alone, MCF-7 alone, PC-3 alone, and BxPC-3 alone control groups are shown as bars in the upper left of the figure. Fig. 12: Induction of in vitro GDF-15 release by MMAE GDF-15 induction by MMAE in cancer cell lines (Panc02, MCF-7, BxPC-3, PC-3) was evaluated by ELISA analysis. Fig. 13: Induction of apoptosis by increasing concentrations of MMAE (monomethylauristatin E) & enfortumab vedotin Cell viability was quantitatively evaluated for cancer cell lines (RT-4, TUHR4TKB, OS-RC-2, ZR-75-30) treated with MMAE & enfortumab vedotin. The values ​​presented were calculated as the percentage (%) of living cells compared to the untreated control group. The calculated IC50 values ​​are shown in the legend. Fig. 14: In vitro induction of GDF-15 release by MMAE (monomethylauristatin E) & enfortumab vedotin GDF-15 induction by MMAE & enfortumab vedotin at approximately IC50 concentrations of each compound in cancer cell lines (RT-4, TUHR4TKB, OS-RC-2, ZR-75-30) was evaluated by ELISA analysis. Fig. 15: Induction of apoptosis by increasing concentrations of DM-1 (mertansine) & trastuzumab emtansine Cell viability was quantitatively evaluated in cancer cell lines (OS-RC-2, ZR-75-30) treated with DM-1 and trastuzumab emtansine. The values ​​presented were calculated as the percentage (%) of living cells relative to the untreated control group. The calculated IC50 values ​​are shown in the legend. Fig. 16: In vitro induction of GDF-15 release by DM-1 (mertansine) & trastuzumab emtansine GDF-15 induction by DM-1 & trastuzumab emtansine at approximately IC50 concentrations of each compound in cancer cell lines (OS-RC-2, ZR-75-30) was evaluated by ELISA analysis. Fig. 17: Induction of apoptosis by increasing concentrations of DxD & trastuzumab deruxtecan Cell viability was quantitatively evaluated in cancer cell lines (Caov-3, MCF-7, RT-4, SW-780, TUHR4TKB, NCI-N87, SW-837, HepG2, JHH-1, NCI-H2122) upon treatment with DxD & trastuzumab deruxtecan. The values ​​presented were calculated as the percentage (%) of living cells relative to the untreated control group. The calculated IC50 values ​​are shown in the legend. Fig. 18: In vitro induction of GDF-15 release by DxD and trastuzumab deruxtecan GDF-15 induction by DxD & trastuzumab deruxtecan at approximately IC50 concentrations of each compound in cancer cell lines (Caov-3, MCF-7, RT-4, SW-780, TUHR4TKB, NCI-N87, SW-837, HepG2, JHH-1, NCI-H2122) was evaluated by ELISA analysis. Fig. 19: Induction of apoptosis by increasing concentrations of SN-38 & sacituzumab govitecan Cell viability was quantitatively evaluated in cancer cell lines (Caov-3, MCF-7, RT-4, TUHR4TKB, SW-780, OVCAR-3, NCI-N87, SW-837, HT-1376, NCI-H747, NCI-H2122) upon treatment with SN-38 & sacituzumab govitecan. The values ​​presented were calculated as the percentage (%) of living cells relative to the untreated control group. The calculated IC50 values ​​are shown in the legend. Fig. 20: In vitro induction of GDF-15 release by SN-38 & sacituzumab govitecan GDF-15 induction by SN-38 & sacituzumab govitecan at approximately IC50 concentrations of each compound in cancer cell lines (Caov-3, MCF-7, RT-4, TUHR4TKB, SW-780, OVCAR-3, NCI-N87, SW-837, HT-1376, NCI-H747, NCI-H2122) was evaluated by ELISA analysis. Fig. 21: Induction of apoptosis by increasing concentrations of caliceamicin & SG3199 Cell viability was quantitatively evaluated in cancer cell lines (AMO-1, JHH-1, OS-RC-2, HT-1376, MCF-7) upon treatment with caliceamicin and SG3199. The values ​​presented were calculated as the percentage (%) of living cells relative to the untreated control group. The calculated IC50 values ​​are shown in the legend. Fig. 22: In vitro induction of GDF-15 release by caliceamicin & SG3199 GDF-15 induction by caliceamicin & SG3199 at approximately IC50 concentrations of each compound in cancer cell lines (AMO-1, JHH-1, OS-RC-2, HT-1376, MCF-7) was evaluated by ELISA analysis. Fig. 23: Induction of apoptosis in RT-4 cells by increasing concentrations of sacituzumab, SN-38 & sacituzumab govitecan Cell viability was quantitatively evaluated in RT-4 cancer cell line upon treatment with any one of sacituzumab, SN-38, or sacituzumab govitecan. The values ​​presented were calculated as the percentage (%) of living cells compared to the untreated control group. Fig. 24: Induction of ICD (immunogenic apoptosis) in RT-4 cells by sacituzumab govitecan Quantification of ICD-associated DAMPs (risk-associated molecular patterns) indicated by eATP assessed by bioluminescence assay, HMGB1 assessed by ELISA assay, and ecto-caleticulin potential assessed by flow cytometry after treatment of cancer cell line RT-4 with sacituzumab govitecan. Values ​​shown are standardized fold changes relative to the untreated control at all time points. Fig. 25: Induction of apoptosis in RT-4 cells by increasing concentrations of enfortumab, MMAE & enfortumab vedotin Cell viability was quantitatively evaluated in cancer cell line RT-4 upon treatment with any one of enfortumab, MMAE, or enfortumab vedotin. The values ​​presented were calculated as the percentage (%) of living cells compared to the untreated control group. Fig. 26: Induction of ICD (immunogenic apoptosis) in RT-4 cells by enfortumab vedotin Quantification of eATP assessed by bioluminescence assay, HMGB1 assessed by ELISA assay, and ICD-associated DAMPs indicated by ecto-caleticulin potential assessed by flow cytometry after treatment of cancer cell line RT-4 with enfortumab vedotin. Values ​​shown are standardized fold changes relative to the untreated control group for all time points. Fig. 27: Induction of apoptosis in RT-4 cells by increasing concentrations of trastuzumab, DxD & trastuzumab deruxtecan Cell viability in RT-4 cancer cell line was quantified upon treatment with any one of trastuzumab, DxD, or trastuzumab deruxtecan. The values ​​presented were calculated as the percentage (%) of living cells relative to the untreated control group. Fig. 28: Induction of ICD (immunogenic apoptosis) in RT-4 cells by trastuzumab deruxtecan Quantification of eATP assessed by bioluminescence assay, HMGB1 assessed by ELISA assay, and ICD-associated DAMPs indicated by ecto-caleticulin potential assessed by flow cytometry after treatment of cancer cell line RT-4 with trastuzumab deruxtecan. Values ​​shown are standardized fold changes relative to the untreated control group for all time points. Fig. 29: Induction of apoptosis in RT-4 cells by increasing concentrations of trastuzumab, DM-1 & trastuzumab emtansine Cell viability was quantitatively evaluated for cancer cell line RT-4 upon treatment with any one of trastuzumab, DM-1, or trastuzumab emtansine. The values ​​presented were calculated as the percentage (%) of living cells relative to the untreated control group. Fig. 30: Induction of ICD (immunogenic apoptosis) in RT-4 cells by trastuzumab emtansine Quantification of eATP assessed by bioluminescence assay, HMGB1 assessed by ELISA assay, and ICD-associated DAMPs indicated by ecto-caleticulin potential assessed by flow cytometry after treatment of cancer cell line RT-4 with trastuzumab emtansine. Values ​​shown are standardized fold changes relative to the untreated control group for all time points. Fig. 31: Induction of human versus murine GDF-15 protein expression by sacituzumab govitecan in a xenograft tumor model Human versus mouse GDF-15 induction in serum collected after treatment with sacituzumab govitecan in MCF-7 tumor-bearing mice was measured by ELISA analysis. Tumors from animals treated as a vehicle control were used as a control. Fig. 32: Induction of human GDF-15 and IP-10 (ICD marker) gene expression within tumors by enfortumab vedotin in a xenograft tumor model Induction of human GDF-15 mRNA and IP-10 / CXCL10 (ICD markers) evaluated by RT-qPCR (calculated by the ΔΔCT method) in tumors finally collected from HT-1376 tumor-bearing mice treated with enfortumab vedotin. Tumors from animals treated as vehicle controls were used as controls. Fig. 33: Induction of human versus mouse GDF-15 protein expression by enfortumab vedotin in a xenograft tumor model Human versus mouse GDF-15 induction in serum collected after treatment with enfortumab vedotin in HT-1376 tumor-bearing mice was measured by ELISA analysis. Tumors from animals treated as vehicle controls were used as controls. Fig. 34: Reduction in tumor volume by anti-GDF-15 antibody (aGDF-15) in combination with enfortumab vedotin in a synaptic tumor model Tumor volumes of MC38 / hNectin4 transplanted animals treated with any one of aGDF-15, enfortumab vedotin, or a combination of aGDF-15 and enfortumab vedotin were evaluated by caliper measurement. Untreated animals were used as controls. Average tumor volumes are shown over time. Fig. 35: Tumor volume of individual animals treated with aGDF-15, enfortumab vedotin & aGDF-15 + enfortumab vedotin Tumor volumes of MC38 / hNectin4 transplanted animals treated with aGDF-15, enfortumab vedotin, or a combination of aGDF-15 and enfortumab vedotin were evaluated by caliper measurement. Untreated animals were used as a control group. The average tumor volume over time for individual animals is presented by treatment group. Fig. 36: Tumor growth inhibition by aGDF-15 in combination with enfortumab vedotin in a synaptic tumor model Tumor growth inhibition was evaluated in MC38 / hNectin4 transplanted animals treated with any one of aGDF-15, enfortumab vedotin, or a combination of aGDF-15 and enfortumab vedotin. Tumor growth inhibition was calculated by dividing the tumor volume of individual mice on day 25 by the average control tumor volume on the same day. Fig. 37: Induction of intratumoral immune cell infiltration and activation by aGDF-15 in combination with enfortumab vedotin in a synonymic tumor model Immunocytes from the tumors finally collected from MC38 / hNectin4 transplanted animals treated with any one of aGDF-15, enfortumab vedotin, or a combination of aGDF-15 and enfortumab vedotin were evaluated by flow cytometry. Changes in immune infiltration and antigen-presenting cell (APC) activation induced by various treatments were quantified. Fig. 38: Induction of CD4 T cell and CD8 T activation in PBMCs by aGDF-15 in combination with enfortumab vedotin in a synaptic tumor model Immune cells were evaluated by flow cytometry in PBMCs collected at the median time point (day 8 after the start of treatment) in MC38 / hNectin4 transplanted animals treated with any one of aGDF-15, enfortumab vedotin, or a combination of aGDF-15 and enfortumab vedotin. Changes in CD4 T cell & CD8 T cell activation caused by various treatments were quantified. Fig. 39: Induction of CD4 T cell & CD8 T proliferation in PBMCs by aGDF-15 in combination with enfortumab vedotin and anti-PD1 antibody (aPD1) in a synonymic tumor model Immune cells were evaluated by flow cytometry in PBMCs collected at the median time point (day 8 after the start of treatment) in MC38 / hNectin4 transplanted animals treated with any combination of aGDF-15, enfortumab vedotin, aPD1, or aGDF-15, enfortumab vedotin, and aPD1. Changes in CD4 T cell and CD8 T cell proliferation caused by various treatments were quantified. Statistical significance in degrees is indicated by an asterisk: *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, and ****P ≤ 0.0001. P-values ​​were calculated using one-way ANOVA in GraphPad Prism, and significance thresholds were determined by the software. Specific details for implementing the invention

[0129] Definition and General Description

[0130] Unless specifically defined in this specification, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art of the present invention.

[0131] All methods and materials similar or equivalent to those described herein may be used to practice or test the invention, and suitable methods and materials are described herein.

[0132] All references mentioned in this specification are incorporated by reference in their entirety for all purposes. Patent applications are referred to in this specification by application and / or publication numbers. Non-patent literature mentioned in this specification may be cited by reference in its entirety or in its abbreviated form (e.g., Cao et al., 2017), followed by a full reference after mention in the “References” section and / or text.

[0133] The present invention provides an anti-hGDF-15 antibody or its antigen-binding portion for use in cancer treatment in patients in combination with a cancer antigen-targeting drug conjugate that induces cancer cell stress.

[0134] Sequence alignment of the sequence according to the present invention is performed using the BLAST algorithm (see Altschul et al. (1990) "Basic local alignment search tool." Journal of Molecular Biology 215. p. 403-410; Altschul et al.: (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 25:3389-3402). Preferably, the following parameters are applied: max target sequence 10; word size 3; BLOSUM 62 matrix; gap cost: presence 11, extension 1; conditional configuration score matrix adjustment. Accordingly, when used in relation to the sequence, terms such as "identity" or "identical" refer to the identity value obtained using the BLAST algorithm.

[0135] As used herein, the term “antibody” refers to any functional antibody capable of specifically binding to a target antigen, as generally described in Chapter 7 of Paul, WE (Ed.): Fundamental Immunology 2nd Ed. Raven Press, Ltd., New York 1989, incorporated herein by reference. Without particular limitation, the term “antibody” encompasses antibodies derived from any suitable species of origin, including chickens and mice, goats, non-human primates, and mammals such as humans. Preferably, the antibody is a humanized antibody. The antibody is preferably a monoclonal antibody that can be prepared by methods well known in the art. The term “antibody” encompasses IgG-1, -2, -3, or -4, IgE, IgA, IgM, or IgD isotype antibodies. The term “antibody” encompasses monomeric antibodies (e.g., IgD, IgE, IgG) or oligomeric antibodies (e.g., IgA or IgM). The term "antibody" also encompasses isolated antibodies and genetically engineered antibodies, such as chimeric antibodies, without specific limitation.

[0136] The nomenclature for antibody domains follows terms known in the art. Each antibody monomer comprises two heavy chains and two light chains, as is generally known in the art. Of these, each heavy chain and light chain comprises a variability domain important for antigen binding (designated VH for the heavy chains and VL for the light chains). These heavy and light chain variability domains comprise the regions FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4 (FR, framework region; CDR, complementarity determining region also known as the supervariability region) (in order from N-terminus to C-terminus). The identification and assignment of the aforementioned antibody regions in antibody sequences are generally based on Kabat et al. (Sequences of proteins of immunological interest, US Dept. of Health and Human Services, Public Health Service, National Institutes of Health, Bethesda, Md. 1983), or Chothia et al. This can be performed in accordance with (Conformations of immunoglobulin hypervariable regions. Nature. 1989 Dec 21-28;342 (6252):877-83) or using the IMGT / V-QUEST software described in Giudicelli et al. (IMGT / V-QUEST, an integrated software program for immunoglobulin and T cell receptor VJ and VDJ rearrangement analysis. Nucleic Acids Res. 2004 Jul 1;32 (Web Server issue):W435-40), which is incorporated herein by reference. Preferably, the antibody regions mentioned above are identified and assigned using the IMGT / V-QUEST software.

[0137] “Monoclonal antibodies” are antibodies derived from a population of antibodies that are essentially homogeneous, and the antibodies are substantially identical in sequence (i.e., identical except for a small proportion of antibodies with naturally occurring sequence variations, such as amino acid modifications at the N-terminus and C-terminus). Unlike polyclonal antibodies, which contain a mixture of various antibodies targeting multiple epitopes, monoclonal antibodies target the same epitope and are therefore highly specific. The term “monoclonal antibody” includes (non-limitingly) antibodies obtained from a population of monoclonal cells derived from a single cell clone, such as antibodies produced by the hybridoma method described in Kohler and Milstein (Nature, 1975 Aug 7;256 (5517):495-7) or Harlow and Lane (“Antibodies: A Laboratory Manual,” Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York 1988), which are incorporated herein by reference. Monoclonal antibodies may also be obtained by other suitable methods, including phage display techniques as described in Clackson et al. (Nature. 1991 Aug 15;352 (6336):624-8) or Marks et al. (J Mol Biol. 1991 Dec 5;222 (3):581-97), which are incorporated herein by reference. Monoclonal antibodies may be antibodies optimized for antigen-binding properties, such as reduced Kd values, optimized binding, and dissociation kinetics, through methods known in the art. For example, Kd values ​​may be optimized through display methods, including phage display, to construct affinity-mature monoclonal antibodies. The term "monoclonal antibody" is not limited to antibody sequences derived from a specific origin species or a single origin species. Thus, the meaning of the term "monoclonal antibody" encompasses chimeric monoclonal antibodies, such as humanized monoclonal antibodies.The anti-hGDF-15 antibody according to the present invention is preferably a monoclonal antibody.

[0138] A "humanized antibody" is an antibody containing a human sequence and a small non-human sequence that confers binding specificity to a target antigen (e.g., human GDF-15). Typically, a humanized antibody is constructed by replacing the hypervariable region sequence of a human acceptor antibody with the hypervariable region sequence of a non-human donor antibody (e.g., mouse, rabbit, or rat donor antibody) that binds to the target antigen (e.g., human GDF-15, also referred to as "hGDF-15"). In some cases, the framework region sequence of the acceptor antibody may also be replaced with the corresponding sequence of the donor antibody. In addition to the sequences derived from the acceptor and donor antibodies, the "humanized antibody" may or may not contain other (additional or substituted) residues or sequences. These other residues or sequences may serve to further improve antibody properties, such as binding properties (e.g., reduction of the Kd value) and / or immunogenic properties (e.g., reduction of antigenicity in humans). Non-limiting examples of methods for generating humanized antibodies are known in the art, for example, Riechmann et al. (Nature. 1988 Mar 24; 332 (6162):323-7) or Jones et al. (Nature. 1986 May 29-Jun 4; 321 (6069):522-5), which are incorporated herein by reference.

[0139] The term "human antibody" refers to antibodies containing human variable and invariant domain sequences. This definition encompasses antibodies having human sequences in which a single amino acid substitution or modification exists that may serve to further enhance antibody properties, such as binding properties (e.g., reduction of Kd value) and / or immunogenic properties (e.g., reduction of antigenicity in humans). The term "human antibody" excludes humanized antibodies in which a portion of a non-human sequence confers binding specificity to a target antigen.

[0140] As used herein, the “antigen-binding portion” of an antibody refers to a portion of the antibody that retains the ability for the corresponding antibody to specifically bind to an antigen (e.g., hGDF-15, PD-1, or PD-L1). This ability may be determined, for example, by determining the ability of the antigen-binding portion to compete with the antibody for specific binding to the antigen through methods known in the art. The antigen-binding portion may comprise one or more fragments of the antibody. Without particular limitation, the antigen-binding portion may be constructed through any suitable method known in the art, including recombinant DNA methods and preparation by chemical or enzymatic fragmentation of the antibody. The antigen-binding portion may be a Fab fragment, an F(ab') fragment, an F(ab')2 fragment, a single-strand antibody (scFv), a single-domain antibody, a diabody, or any other portion(s) of the antibody that retain the ability to specifically bind to the antigen.

[0141] As used herein, the terms “bind” or “bind” refer to specific binding to a target antigen (e.g., human GDF-15). Preferably, the Kd value is less than 100 nM, more preferably less than 50 nM, even more preferably less than 10 nM, even more preferably less than 5 nM, and most preferably less than 2 nM.

[0142] As used herein, an antibody or its antigen-binding portion that "competes" with the second (reference) anti-hGDF-15 antibody (e.g., an anti-hGDF-15 antibody having a heavy chain having an amino acid sequence represented by SEQ ID NO. 8 and a light chain having an amino acid sequence represented by SEQ ID NO. 9) means that the "competing" (first) antibody or its antigen-binding portion can reduce the binding of the 10 nM second antibody reference solution to human or recombinant human GDF-15 by at least 50%. Generally, "competes" means that the concentration of the (first) antibody or its antigen-binding portion required to reduce the binding of the 10 nM second antibody reference solution to human or recombinant human GDF-15 by at least 50% is less than 1000 nM, preferably less than 100 nM, more preferably less than 10 nM. Binding is measured by surface plasmon resonance measurements or enzyme-linked immunosorbent assays (ELISA).

[0143] As used herein, the term “epitope” refers to a small portion of an antigen that forms a binding site for an antibody.

[0144] As used herein, the term “single-domain antibody” (also referred to as “Nanobody™”) is an antibody fragment consisting of a single monomeric variable antibody domain. The structure and method of preparing single-domain antibodies are known in the art, for example, in Holt LJ et al. (“Domain antibodies: proteins for therapy.” Trends Biotechnol. 2003 Nov;21 (11):484-90), Saerens D et al. (“Single-domain antibodies as building blocks for novel therapeutics.” Curr Opin Pharmacol. 2008 Oct;8 (5):600-8. Epub 2008 Aug 22.) and Arbabi Ghahroudi M et al. (“Selection and identification of single domain antibody fragments from camel heavy-chain antibodies.” FEBS Lett. 1997 Sep 15;414 (3):521-6), which are incorporated herein by reference.

[0145] According to the present invention, each instance in which the terms “comprising,” “comprising,” and “comprising” are mentioned may optionally be replaced with the terms “consisting of,” “consisting of,” and “consisting of,” respectively.

[0146] The anti-hGDF-15 antibody or its antigen-binding portion to be used according to the present invention

[0147] The "anti-hGDF-15 antibody or its antigen-binding portion" according to the present invention is an antibody or its antigen-binding portion capable of specifically binding to human GDF-15 (hGDF-15), e.g., recombinant mature hGDF-15. Examples of such antibodies are described in the preferred embodiments and claims.

[0148] The anti-hGDF-15 antibody or its antigen-binding portion may be a neutralizing antibody or its antigen-binding portion. In this regard, the term “neutralization” is understood not to mean complete neutralization, but may mean any detectable neutralization, e.g., at least 50%, at least 80%, or at least 90% neutralization. To determine whether the antibody or its antigen-binding portion is a neutralizing anti-hGDF-15 antibody or its antigen-binding portion, an analysis based on a model system for measuring T cell attachment to human umbilical vein endothelial cells (HUVECs), as described in Example 3 of WO2017 / 055613, may be used, the said literature is incorporated herein by reference.

[0149] In this regard, T cells were pre-incubated with 100 ng / ml GDF-15 for 1 hour, or pre-incubated with 100 ng / ml GDF-15 that had been pre-incubated with 10 µg / ml antibody for 1 hour, and then tested briefly as described below.

[0150] T cell flow / adhesion experiment (at HUVEC):

[0151] Day 1:

[0152] - μ-slide VI 0.4 (ibidi GmbH, Germany) was coated with fibronectin (100 μg / mL): 30 μl per loading port. This was incubated at 37°C for 1 hour (or use pre-coated slides).

[0153] - After removing fibronectin through aspiration, rinse with HUVEC medium.

[0154] - HUVECs were trypsin-treated from a 6-well plate (Count: 2x10 5 / ㎖ (Total 2 ㎖))

[0155] - Wash this to make 1x10 6 Diluted to cells / mL

[0156] - 30 µl of HUVEC was applied to the loading port of μ-slide VI and verified under a microscope.

[0157] - μ-slide VI was covered with a lid and incubated at 37°C and 5% CO2.

[0158] Day 2:

[0159] - HUVECs were activated with TNFα (10 ng / mL) and IFNγ (10 ng / mL) in channels 2-5 (see table below): all media were removed from the channels and replaced with cytokine-containing preheated media.

[0160] Day 3:

[0161] - T cells were isolated (negative isolation of pan T cells).

[0162] - T cells in well 1 (1x10 6 Pre-incubated for 1 hour with or without GDF-15 (100 ng / mL) in cells / mL.

[0163] - HUVECs were pre-incubated with GDF-15 (100 ng / mL) in channels 4 and 5 for 1 hour: all medium was removed by suction from the loading port, and both loading ports were filled with preheated medium containing GDF-15.

[0164] - Preheated the stage-top incubator next to the microscope and coupled it with a gas mixture (5% CO2, 16% O2, 79% N2).

[0165] - 3 x 50 ml syringes were prepared:

[0166] ■ T cells (1x10 6 Cells / mL): 1 mL

[0167] ■ T cell GDF15 (1x10 6 Cells / mL): 1 mL

[0168] ■ Badge

[0169] - Connected Syringe 1 to Channel 1 (see table below) and allowed flow (0.5 dyn / cm²) 2 : 0.38 ml / min = 22.8 ml / h).

[0170] - T cells were flowed for 3 minutes, and during that time, 10 regions of interest were pre-designated on the microscope.

[0171] - Each region of interest was videotaped for 5 seconds.

[0172] - The remaining channels were evaluated similarly to channel 1 (fh) using T cell samples specified in the table below.

[0173] Channel # Endothelial cells T cells in motion note 1 Non-irritating HUVEC T cells [Negative Control Group] 2 Stimulated HUVEC T cells [Positive Control Group] 3 Stimulated HUVEC T cell GDF-15 4 Stimulated HUVECGDF-15 T cells 5 Stimulated HUVECGDF-15 T cell GDF-15

[0174] Alternative setting (according to Haake et al. 2023, which is incorporated herein by reference):

[0175] HUVECs (Lonza, Catalog #: CC-2517, used up to 5 passages) were cultured on chamber slides for 2–3 days prior to overnight activation with TNF-α (1000 U / mL) and IFNβ (500 U / mL) or IFNγ (500 U / mL). Healthy donor PBMCs obtained from soft layers no more than 24 hours prior to the experiment were purified using the MACS® Cell Isolation Kit (Miltenyi Biotec) to obtain pan CD8+ T cells (Human CD8+ T Cell Isolation Kit #130-096-495). Purified T cells were stored in 1% BSA at 4°C and acclimatized to 37°C 1 hour prior to the experiment. Jurkat cells were used instead of primary T cells. Physiological flow (0.5 Dyns / cm²) was measured through HUVEC culture slides mounted on a heated microscope chamber (37°C) using a calibrated pump. 2A ) was constructed. Then, chemokines (1 μM CXCL12α or 0.5 μM CXCL9 + 0.5 μM CXCL10) and GDF-15 (100 ng / mL) or a vehicle control (100 mM acetic acid) were perfused onto the activated HUVEC monolayer for 5 minutes and then incubated for 15 minutes (Step 1). Afterward, wash buffer was pumped onto the HUVEC for 10 minutes to remove any unbound CXCL chemokines or GDF-15. Leukocyte suspensions were pretreated for 20 minutes with rhGDF-15 (100 ng / mL), rhGDF-15 (100 ng / mL) + anti-hGDF-15 antibody (20 µg / mL), or blocking anti-LFA-1 antibody TS1 / 18 (20 µg / mL) (# MA1810, ThermoFisher Scientific) (positive control), or vehicle control, then centrifuged and resuspended in wash medium. Subsequently, leukocytes were perfused onto HUVECs for 6 minutes (Step 2), followed by flushing wash buffer onto the cells for 50 minutes under a pressure of 0.1 Pa (Step 3). During Steps 2 and 3, captured leukocytes were imaged every 30 seconds using a phase-contrast microscope. The generated short video sequences allowed for the analysis of individual leukocytes over a wide area. Unit field (0.19 mm²) 2 Total number of attachment events per / mm 2 It was represented as follows. T cells attached to the surface of HUVECs showed a phase-white / gray appearance, while migrating (mobilized) T cells showed a phase-black appearance. The number of cells captured at each time point was equal to the total number of cells attached to both sides of the HUVEC layer. Migration events (phase black) were expressed as the percentage of total T cells (phase gray + black) captured from the flow per unit field. The effect of GDF-15 on T cell adhesion was confirmed using pan T cells or additional T cell subgroups in the same experimental setup. The indicated time points (30 min or 15 min) are specified. 3 to 4 donors were used per condition.

[0176] Non-limiting examples of anti-hGDF-15 antibodies are described in US 2020 / 0055930 (see, for example, Tables 2 and 5), WO2021197171A1, WO2017189724A1, WO2020039321A2, WO2023018803 A1 (see, for example, Table 2), WO2022189936A1 and WO 2014 / 100689 A1, which are incorporated herein by reference.

[0177] As stated herein, the terms "CTL-002", "H1L5", and bisugromab are used synonyms. These terms refer to antibodies having the heavy chain amino acid sequence of SEQ ID NO. 8 and the light chain amino acid sequence of SEQ ID NO. 9. All references to the amino acid sequences of the antibodies of the present invention are intended to include post-translational modifications of these sequences occurring in mammalian cells, such as CHO cells, including but not limited to N-glycosylation, O-glycosylation, deamidation, Asp isomerization / fragmentation, pyro-glutamate formation, C-terminal lysine removal, and Met / Trp oxidation.

[0178] A preferred anti-GDF-15 antibody useful for the present invention may be any one selected from the group consisting of bisugromab, ponsegromab, relogrotug (i.e., AV-380) and an antibody having a heavy chain variable region formed by SEQ ID NO. 19 and a light chain variable region formed by SEQ ID NO. 20 (e.g., AZD8853), and preferably is bisugromab.

[0179] The heavy chain of ponsegromab consists of the following amino acids:

[0180] QVQLVQSGAEVKKPGSSVKVSCKASGYTFSSYNIDWVRQAPGQGLEWMGGINPIFGTAFYNQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREAITTVGAMDHWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEAAGAPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG (서열번호 15).

[0181] 폰세그로맙의 경쇄는 다음 아미노산으로 구성된다:

[0182] EIVLTQSPATLSLSPGERATLSCRTSQSVHNYLAWYQQKPGQAPRLLIYDASTRADGIPARFSGSGSGTDFTLTISSLEPEDFAVYYCQQFWSWPWTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (서열번호 16).

[0183] 릴로그로투그의 중쇄는 다음 아미노산으로 구성된다:

[0184] QVQLVQSGAEVKKPGSSVKVSCKASGYTFSDYNMDWVRQAPGQGLEWMGQINPNNGLIFFNQKFKGRVTLTADKSTSTAYMELSSLRSEDTAVYYCAREAITTVGAMDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (서열번호 17)

[0185] 릴로그로투그의 경쇄는 다음 아미노산으로 구성된다:

[0186] DIQMTQSPSSLSASVGDRVTITCRTSENLHNYLAWYQQKPGKAPKLLIYDAKTLADGVPSRFSGSGSGTDYTLTISSLQPEDFATYYCQHFWSSPYTFGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (서열번호 18)

[0187] AZD8853의 중쇄 가변 영역은 다음 아미노산으로 구성된다:

[0188] QVQLVQSGSELKKPGASVKVSCKASGYTFTDYNMDWIRQSPGKGLEWIGDINPNQGGTFYNQKFKDRATLTVDKSTSTAYMELRSLRSDDTAVYYCAREEKLYFGLMDYWGQGTTVTVSS (서열번호 19)

[0189] The light chain variable region of AZD8853 consists of the following amino acids:

[0190] DIQMTQSPSSLSASVGDRVTITCRASENIYSYLAWYQQKPGKAPKLLVYNAKTLAEGVPSRFSGSGSGIDFTLTISSLQPEDFATYYCQHQYGSPPTFGQGTKLEIK (SEQ ID NO: 20)

[0191] An analysis for determining whether the first anti-hGDF-15 antibody competes with the second (referenced) anti-hGDF-15 antibody for specific binding to hGDF-15 is known in the art.

[0192] Example of analysis to determine whether the first anti-hGDF-15 antibody competes with the second anti-hGDF-15 antibody for specific binding to hGDF-15:

[0193] To determine whether two distinct anti-GDF-15 antibodies can bind to a single GDF-15 dimer, a sandwich ELISA is performed. Accordingly, the anti-GDF-15 capture antibody (i.e., the second (reference) antibody) is immobilized on the bottom of a 96-well plate (50 µl at 2 µg / mL, RT, overnight). It is coated with an isomorphic control antibody as a negative control. The next day, the coating solution is removed, and the plate is washed three times with 150 µl of PBS 0.05% Tween. For the washing procedure, 150 µl of PBS 0.05% Tween is added to the wells, the plate is shaken in a sink, and the plate is patted on a paper towel to remove the wash solution. To block, 150 µl of PBS 1% BSA is added to the wells and incubated at room temperature for 1–2 hours. Next, rhGDF-15 is applied at a specified concentration (50 µl / well diluted in PBS 1% BSA) for 1.5 to 2 hours at RT. After removing the GDF-15 solution and washing three times with 150 µl of PBS 0.05% Tween, 50 µl of the specified detection antibody (i.e., primary anti-hGDF-15 antibody or control antibody) is added to each well, diluted to a final concentration of 1 µg / mL in PBS 1% BSA. After incubating at RT for 1 hour and washing three times with PBS 0.05% Tween, a species-specific HRP-labeled secondary antibody is added and left for 1 hour to bind to the Fc portion of the detection antibody (where: anti-human HRP / anti-chicken HRP 1:5000 in PBS 1% BSA). Wash the plate three times with PBS 0.05% Tween, then prepare the TMB substrate (50 µl of TMB diluted 1:100 in 0.1M pH 6 sodium acetate, H2O21:5000) and pipette 50 µl into each well. After an incubation period of 5-15 minutes, when a luscious blue color is observed, add an equal volume of stop solution (1M H2SO4) and measure the OD at 450 nm in Tecan Sunrise.

[0194] hGDF-15

[0195] hGDF-15 is, for example, ELECSYS, summarized by Wollert et al. (Wollert KC, Kempf T, Giannitsis E, et al. An Automated Assay for Growth Differentiation Factor 15. J Appl Lab Med An AACC Publ. 2018;1 (5):510-521. doi:10.1373 / jalm.2016.022376). ® As in the GDF15 assay (Roche Diagnostics), measurements can be performed non-limitingly through ELISAs including R&D systems Quantikine ELISA, immunoradiographic counting, luminex™ sandwich assay and electrochemiluminescence sandwich assay, the above literature is incorporated herein by reference.

[0196] Alternatively, serum hGDF-15 levels can be determined by known electrochemiluminescence immunoassays using antibodies against GDF-15. For example, Roche Elecsys ® The technique can be used for such electrochemiluminescence immunoassays.

[0197] All mentioned analyses capture and quantify GDF-15 based on the immunosandwich principle using monoclonal or polyclonal antibodies. Depending on the reagents used and their combinations, free GDF-15 or total GDF-15 (free GDF-15 and GDF-15 conjugated with CTL-002) is measured.

[0198] hGDF-15 expression in a patient may be, for example, the patient's hGDF-15 serum level. Accordingly, according to the present invention, cancer cell stress or immunogenic apoptosis (ICD) associated with the induction of hGDF-15 expression in a patient, induced by the cancer antigen-targeting drug conjugate of the present invention, can be detected by measuring hGDF-15 serum levels before and after the initiation of treatment using the cancer antigen-targeting drug conjugate, and an increase in hGDF-15 serum levels indicates the presence of hGDF-15 expression induction in the patient.

[0199] Cancer antigen-targeting drug conjugates that induce cancer cell stress

[0200] According to the present invention, the term “cancer antigen-targeting drug conjugate inducing cancer cell stress” is not particularly limited and relates, in particular, to a conjugate of a drug and a cancer antigen target portion that specifically delivers a drug into a cancer cell, internalizes it, and subsequently releases the drug that induces cell stress in the cancer cell from within.

[0201] Those skilled in the art recognize methods to assess whether cancer antigen-targeted drug conjugates induce cancer cell stress (e.g., by reference to Collins, Denis M., et al. 2019, and Tiligada, E. 2006, incorporated herein by reference). As a non-limiting example, cancer cell stress may be assessed by determining hGDF-15 induction as further described herein and illustrated in the experimental section. Likewise, those skilled in the art also recognize that cancer antigen-targeted drug conjugates can induce cancer cell death, such as immunogenic apoptosis (ICD), in cancer cells.

[0202] In the context of the cancer antigen-targeting drug conjugate used according to the present invention, the words "payload," "cargo," and "drug" are used interchangeably and are intended to have the same meaning.

[0203] Non-limiting examples of drugs used in the cancer antigen-targeting drug conjugate of the present invention are selected from the group consisting of microtubule inhibitors, topoisomerase I or II inhibitors, DNA damaging agents, protein synthesis inhibitors, and immune-activating substances.

[0204] In particular, cancer targeted therapies using ADCs are known to induce cancer cell stress or apoptosis, followed by stimulating a potent anti-cancer immune response. A well-characterized pathway induced by ADCs is called immunogenic apoptosis (ICD) of cancer cells (see, e.g., Cao et al., 2017; Rios-Doria et al., 2017; Bauzon et al., 2019; D'Amico et al., 2019; Boshuizen et al., 2021; Devra Olson et al., 2022, which are incorporated herein by reference). ICD is associated with the release of various danger signals and cancer antigens that can enhance the recruitment and activation of immune cells.

[0205] Immunogenic cell death (ICD) is a process generally known in the art in which a drug induces apoptosis in cells, such as cancer cells, by stimulating the immune system (see, for example, Kroemer et al. 2022, which is incorporated herein by reference).

[0206] For example, exposure to caleticulin on the cell surface and the release of ATP and HMGB1 are known to be associated with ICD. Subsequently, treated and presented cancer antigens effectively trigger an immune response against cancer (Kroemer et al., 2022).

[0207] The inventors surprisingly demonstrated that cancer cell lines produce GDF-15 in response to ADC exposure, contributing to an immunosuppressive environment.

[0208] Accordingly, the inventors determined that a cancer antigen-targeting drug conjugate that induces cancer cell stress would achieve a suboptimal therapeutic outcome by limiting the immune response against cancer due to immunosuppression by cancer cells.

[0209] According to the present invention, cancer treatment using a combination of a cancer antigen-targeting drug conjugate and an anti-hGDF-15 antibody or its antigen-binding portion enables reactivation, repair, reinforcement, or enhancement of an adaptive cancer immune response.

[0210] For example, when used for medical purposes or in the manufacture of pharmaceuticals, the antigen-targeting drug conjugate and the anti-hGDF-15 antibody (or its antigen-binding portion) may be used simultaneously or at different times. It is obvious to those skilled in the art that when two substances, such as the anti-hGDF-15 antibody and the antigen-targeting drug conjugate, are used "in combination," they are not necessarily administered or manufactured simultaneously, but may be administered or manufactured at different times.

[0211] ADC to be used according to the present invention

[0212] Antibody-drug conjugates (ADCs) are a type of anticancer agent that enables highly cancer-selective treatment by combining selectivity for target proteins with potent cytotoxic drugs, as described herein. ADCs are chemoimmunotherapy agents and potent targeted chemotherapy agents.

[0213] In the context of antibody-drug conjugates (ADCs) used according to the present invention, the words "payload," "cargo," and "drug" are used interchangeably and are intended to have the same meaning.

[0214] The cytotoxic drug used in the context of the present invention is not particularly limited and ideally induces the production of hGDF-15 in cancer cells.

[0215] Methods for evaluating GDF-15 levels are commonly known in the art, and exemplary ELISA-based methods are described above. Furthermore, screening methods for evaluating whether a given cytotoxic drug induces GDF-15 production in cancer cell lines are part of routine practice in the art.

[0216] In addition, the cytotoxic drug ideally possesses at least one of the following characteristics: high efficacy, high cytotoxic activity (half-maximum inhibitory concentration (IC50) value of less than nanomolar in vitro), high stability in systemic circulation, sufficient solubility of the antibody in an aqueous environment and biochemical properties that allow conjugation to the antibody or its antigen-binding fragment, low immunogenicity, small molecular weight, and a long half-life.

[0217] For example, when used for medical purposes or to manufacture pharmaceuticals, the antibody-drug conjugate and the anti-hGDF-15 antibody (or its antigen-binding portion) may be used simultaneously or at different times. It is obvious to those skilled in the art that if the anti-hGDF-15 antibody and the antibody-drug conjugate are used "in combination," they are not necessarily administered or manufactured simultaneously, but may be administered or manufactured at different times.

[0218] Exemplary classes of cytotoxic drugs used in approved ADCs are microtubule inhibitors or DNA damage agents, as shown in Table 1 below.

[0219] An exemplary ADC is shown below Table 1 and 2 Summarized in.

[0220] Table 1 List of approved ADCs:

[0221] Drug conjugate name Brand name Payload (drug) Drug Class / Mechanism of Action (MOA) Target antigen Enfortumab Vedotin Padcev Monomethylauristatin E (MMAE) microtubule inhibitors Nectin-4 Trastuzumab deruxtecan Enhertu Deruxtecan (Dxd) Topoisomerase I inhibitor HER2 Trastuzumab emtansine Kadcyla Mertansin (DM-1) microtubule inhibitors HER2 Sacituzumab govitecan Trodelvy SN-38 Topoisomerase I inhibitor Trop-2 Tisowumab Vedotin Tivdak Monomethylauristatin E (MMAE) microtubule inhibitors TF (CD142) Brentuximab Vedotin Adcetris Monomethylauristatin E (MMAE) microtubule inhibitors CD30 Inotuzumab ozogamicin Besponsa Kalikeamisin DNA cutting CD22 Mosectumomab Pasodotox Lumoxiti PE38 Inhibition of protein synthesis CD22 Folatuzumab Vedotin Polivy Monomethylauristatin E (MMAE) microtubule inhibitors CD79b Belantamab Mapodotin Blenrep Monomethylauristatin F (MMAF) microtubule inhibitors BCMA Roncastuzumab Tesirin Zynlonta SG3199 / PBD dimer DNA cutting CD19 Mirvetousimab sorabtansin-gynx Elahere DM4 microtubule inhibitors FRα Gemtuzoumab ozogamicin Mylotarg Kalikeamisin DNA cutting CD33

[0222] Table 2 List of related ADCs:

[0223] Drug conjugate name Payload (drug) Drug Class / MOA Target antigen Dicitamab Vedotin Monomethylauristatin E (MMAE) microtubule inhibitors HER2 Radiratuzumab Vedotin Monomethylauristatin E (MMAE) microtubule inhibitors LIV-1 Datopotamab deruxtecan Dxd Topoisomerase I inhibitor Trop-2 Patritumab deruxtecan Dxd Topoisomerase I inhibitor HER3 Kamidanlumab Tesirin PBD DNA cross-link CD25 Upipitamab rilsodotin Auristatin F-HPA microtubule inhibitors NaPi2b XMT-1592 Auristatin F-HPA microtubule inhibitors NaPi2b XMT-1660 Auristatin F-HPA microtubule inhibitors B7-H4 XMT-2056 Exclusive Sting Agent Immune activation HER2 Telisotuzumab Vedotin Monomethylauristatin E (MMAE) microtubule inhibitors c-Met ABBV-400 Topoisomerase I inhibitor c-Met Mirzotamab Clezutoclax BCL-XL inhibitor B7-H3 Cofetuzumab felidotin Auristatin-0101 microtubule inhibitors PTK7 MORAb-202 (parletuzumab) Eribulin microtubule inhibitors FRα STRO-002 SC209 microtubule inhibitors FRα IMGN632 Indolinobenzodiazepine (IGN) DNA alkylation CD123 IMGC936 DM21 microtubule inhibitors ADAM9 Tusamitamab Labtancin Meitansinoid microtubule inhibitors CEACAM5

[0224] Target antigen to be used according to the present invention

[0225] Suitable target antigens (cancer antigens) that enable cancer antigen-targeting drug conjugates to selectively target cancer cells are known in the art, and those skilled in the art recognize screening methods for selecting additional cancer antigens that are suitable targets for cancer antigen-targeting drug conjugates, such as ADCs.

[0226] According to the present invention, the cancer antigen-targeting portion of the conjugate is not limited to antibodies that bind to target antigens on cancer cells, but also includes targeting portions based on ligands or peptides. Regardless of the exact nature of the cancer antigen-targeting portion, the conjugate of the present invention is delivered to the cancer cell through specific interactions between the conjugate and the cancer cell.

[0227] An exemplary target antigen recognized by the cancer antigen-targeting drug conjugate of the present invention must be expressed exclusively or primarily in cancer cells to reduce or prevent off-target toxicity. Furthermore, binding to the target antigen should ideally lead to the internalization of the conjugate within the cancer cells. Additionally, for the antigen to be recognized by the conjugate, it is ideal for the antigen to be located on the cell surface rather than inside the cell. Finally, since circulating soluble antigens can cause undesirable binding of the conjugate outside the cancer site, the antigen must not be secreted or released outside the cancer cells.

[0228] Non-restrictive examples for the target antigen may be selected from any one of the following: Nectin-4, HER2, Trop-2, TF (CD142), CD30, CD22, CD22, CD79b, BCMA, CD19, FRα, CD33, LIV-1, HER3, CD25, NaPi2b, B7-H4, c-Met, B7-H3, PTK7, ADAM9, CEACAM5, 5T4, ALK, AXL, GRP20, CDH6, TA-MUC1, KAAG1, DLK1, DLL3, SLAMF7, CA125, C4.4A / LYPD3, CDH3, CDH6, CAIX, CD20, CD26 / DPP4, CD37, CD38, CD138, CD46, ICAM4 / CD54, CD56 / NCAM1, CD70, CD73, CD74, CD205, CD248, C-KIT, CLDN6, CLDN18.2, CLL-1, RET, CRIPTO, DLK-1, DLL3, EGFR, CD105, ENPP3, EPCAM, EPHA2, FAP, FGFR2 / CD332, FLT3, GDNF / GFRA1, GPC2, GPNMB, guanylyl cyclase (GCC), IGF-1R, ITGAV, Sialyl-di-Lewis, LGR5, LIV1A, LRRC15, MSLN, STEAP1, PSMA, TMEFF2, NOTCH3, PTK7, SLC44A4, SLC46A3, SLITRK6, TIM-1, LY6E and ETBR.

[0229] Cancer and cancer antigen-targeting drug conjugates

[0230] In one embodiment according to all other embodiments of the present invention, cancer is a “solid tumor.” A “solid tumor” is a cancer that forms one or more solid tumors. Solid tumors that form such solid tumors are generally known in the art. The term “solid tumor” encompasses both the primary tumor formed by cancer and possible secondary tumors also known as metastases. A “solid tumor” includes all non-hematological cancers and, without limitation, colorectal cancer, gastric cancer, anal cancer, gastric cancer, esophageal cancer, renal cancer, thyroid cancer, endometrial cancer, testicular cancer, melanoma, skin cancer, bladder cancer, non-small cell lung cancer, head and neck squamous cell carcinoma, hepatocellular carcinoma, liver cancer, biliary tract cancer, prostate adenocarcinoma, pancreatic cancer, uterine cancer, urothelial carcinoma, cervical cancer, ovarian cancer, thyroid cancer, cutaneous squamous cell carcinoma, mesothelioma, cancer of unknown primary site (CUP), and breast carcinoma.

[0231] In one embodiment according to all other embodiments of the present invention, the cancer is selected from the group consisting of urothelial carcinoma (UC), non-small cell lung cancer (NSCLC), pancreatic cancer, head and neck cancer, breast cancer, colorectal cancer, anal cancer, gastric cancer, liver cancer, biliary tract cancer, ovarian cancer, prostate cancer, gastric cancer, esophageal cancer, kidney cancer, thyroid cancer, endometrial cancer, cervical cancer, testicular cancer, mesothelioma, cancer of unknown primary site, melanoma, and skin cancer.

[0232] Likewise, the cancer antigen-targeting drug conjugate according to all other embodiments of the present invention is preferably an ADC selected from the group consisting of enfortumab vedotin, trastuzumab deruxtecan, trastuzumab emtansine, datopotamab deruxtecan, and sacituzumab govitecan.

[0233] Immuno-checkpoint blocker used according to the present invention

[0234] Cancer cells possess genomic mutations that produce cancer cell antigens specific to cancer cells and distinct from the antigens of non-cancerous cells. Therefore, an uninhibited, intact immune system must recognize these cancer cell antigens to trigger an immune response against them. However, most cancers have developed mechanisms of immune tolerance toward these antigens. One type of mechanism by which cancer cells achieve this immune tolerance is the use of immune checkpoints. As used herein, the term "immune checkpoint" generally refers to an immunological mechanism capable of inhibiting an immune response. More specifically, an immune checkpoint is a mechanism characterized by the inhibition of an immune response by molecules (or groups of immune system molecules) of the immune system by inhibiting the activation of immune system cells. These immune system molecules (or groups of molecules) that inhibit an immune response by inhibiting the activation of immune system cells are also known as checkpoint molecule(s).

[0235] As used herein, the term “immune checkpoint blocker” (ICB) is a molecule capable of blocking immune checkpoints. Although it is understood that the hGDF-15 inhibitor used according to the present invention affects the immune system, including effects on CD8+ T cells, as used herein, the term “immune checkpoint blocker” does not refer to the hGDF-15 inhibitor but refers to a molecule different from the hGDF-15 inhibitor.

[0236] The terms "immune checkpoint blocker" (ICB) and "immune checkpoint inhibitor" (ICI) may be used interchangeably.

[0237] The most common immune checkpoint blockers known to date are inhibitors of immune checkpoint molecules, such as human PD-1 inhibitors and human PD-L1 inhibitors. Additional immune checkpoint blockers are inhibitors of anti-LAG-3, anti-B7H3, anti-TIM3, anti-VISTA, anti-TIGIT, anti-KIR, anti-CD27, anti-CD137, anti-CD40, anti-OX40, anti-GITR, anti-ICOS, anti-SIGLEC7, anti-CTLA4, anti-CD276, as well as IDO. Thus, as used according to the present invention, a preferred form of immune checkpoint blocker is an inhibitor of an immune checkpoint molecule. Alternatively, the immune checkpoint blocker may be an activator of a co-stimulatory signal that neutralizes the immune checkpoint.

[0238] Methods for measuring the efficacy of immune checkpoint blockers include in vitro binding assays, primary T cell-based cytokine release assays, and in vivo model systems. Furthermore, Promega has developed a commercially available bioluminescent reporter system for PD-1 / PD-L1, which is mentioned, for example, below: Mei Cong, Ph.D. et al.: Advertorial: Novel Bioassay to Assess PD-1 / PD-L1 Therapeutic Antibodies in Development for Immunotherapy Bioluminescent Reporter-Based PD-1 / PD-L1 Blockade Bioassay. (http: / / www.genengnews.com / gen-articles / advertorial-novel-bioassay-to-assess-pd-1-pd-l1-therapeutic-antibodies-in-development-for-immun / 5511 / ).

[0239] Desirable immune checkpoint blockers are human PD-1 inhibitors and human PD-L1 inhibitors.

[0240] As used herein, the term “human PD-1 inhibitor” may be any molecule capable of specifically inhibiting the function of human PD-1. Non-limiting examples of such molecules are antibodies capable of binding to human PD-1 and DARPin (Designed Ankyrin Repeat Proteins) capable of binding to human PD-1. Preferably, the PD-1 inhibitor to be used according to the present invention is an antibody capable of binding to human PD-1, more preferably a monoclonal antibody capable of binding to human PD-1. Most preferably, the monoclonal antibody capable of binding to human PD-1 is selected from the group consisting of nivolumab, pembrolizumab, cemiflimab, pidilizumab, AMP-224, dostarilab, retipanlimab, toripalimab, syntilimab, tisrelizumab, camrelizumab, valstilimab, and spartalizumab, and more preferably nivolumab or pembrolizumab.

[0241] As used herein, the term "human PD-L1 inhibitor" may be any molecule capable of specifically inhibiting the function of human PD-L1. Non-limiting examples of such molecules are antibodies capable of binding to human PD-L1 and DARPin (Designed Ankyrin Repeat Proteins) capable of binding to human PD-L1. Preferably, the human PD-L1 inhibitor to be used according to the present invention is an antibody capable of binding to human PD-L1, more preferably a monoclonal antibody capable of binding to human PD-L1. Most preferably, the monoclonal antibody capable of binding to human PD-L1 is selected from the group consisting of atezolizumab, avelumab, durvalumab, BMS-936559, MPDL3280A, MEDI4736, MSB0010718C, enbapolimab, pakmilimab, and cosivelimab.

[0242] composition

[0243] It is understood that the antibody according to the present invention may be administered in the form of a pharmaceutical composition. Such pharmaceutical composition is prepared in a manner that allows for appropriate storage and administration by using pharmaceutically acceptable components, such as, for example, a carrier, an excipient, or a stabilizer.

[0244] These pharmaceutically acceptable components are not toxic at the levels used when administering the pharmaceutical composition to human patients. The pharmaceutically acceptable components added to the pharmaceutical composition may vary depending on the route of administration.

[0245] Generally, pharmaceutically acceptable ingredients used in connection with the present invention are used in accordance with knowledge available in the art, e.g., Remington's Pharmaceutical Sciences, Ed. AR Gennaro, 20th edition, 2000, Williams & Wilkins, PA, USA.

[0246] The present invention is illustrated by the following non-limiting embodiments.

[0247] Examples

[0248] In vitro cytotoxicity and GDF-15 induction analysis:

[0249] hGDF-15 induction was examined in the following exemplary cancer cell lines: BxPC-3, Panc02 (pancreatic cancer), MCF-7 (breast cancer), PC-3 (prostate cancer), and SK-MEL-5 (skin cancer).

[0250] Detach adhered tumor cells from culture dishes using trypsin-EDTA solution, count them, and place 0.3–1 x 10⁶ cells in each culture medium. 5 It was adjusted to a cell / mL density. 100 µL of this cell suspension was placed in a flat-bottomed 96-well plate at a density of 0.3–1 x 10⁶ per well. 4 Cells were inoculated into wells. Cells were incubated at 37°C and 5% CO2 for 24 hours to allow attachment.

[0251] ADCs, non-conjugated antibodies, or ADC payload drugs were dissolved according to the manufacturer's recommendations and prepared as a series of 4-fold concentrated serial dilutions in culture medium appropriate for each cancer cell line. 50 µl of each drug dilution was added to the inoculated tumor cells, and the culture medium was filled to a volume of 200 µl per well. The assay plates were then transferred to an incubator and incubated for 3 to 5 days at 37°C and 5% CO2.

[0252] After the incubation period ended, 100 µl of the culture supernatant was retrieved, loaded into a new round-bottom 96-well plate, and frozen at -80°C. The tumor cells remaining in the analysis plate were used to perform a CellTiter-Glo viability assay (Promega) according to the manufacturer's instructions. The concentration of GDF-15 in the frozen cell culture supernatant was determined using a human (Cat# DY957) or mouse (Cat# DY6385) GDF-15 DuoSet ELISA kit (R&D Systems) according to the manufacturer's protocol.

[0253] Results Summary

[0254] The inventors have demonstrated that hGDF-15 production is induced by ADCs (see Figs. 2 and 8) and non-conjugated drugs (payloads) (see Figs. 4, 6, 10 and 12) in various cancer cell lines.

[0255] Specifically, hGDF-15 production was induced with increasing concentrations of trastuzumab deruxtecan (anti-HER2, see Fig. 2) and sacituzumab govitecan (anti-TROP2, see Fig. 8) as ADCs, as well as their non-conjugated toxin drugs (payloads) Dxd (see Fig. 10) and SN-38 (see Fig. 4), respectively, simultaneously with the induction of apoptosis as evidenced by reduced cell viability (see Fig. 1, 3, 7, and 9).

[0256] Likewise, the inventors also discovered that hGDF-15 is induced by the microtubule inhibitor monomethylauristatin E (MMAE) (see Fig. 12) and the topoisomerase I inhibitor camptothecin (see Fig. 6), which also occurred simultaneously with the induction of apoptosis, as evidenced by a decrease in cell viability (see Figs. 11 and 5).

[0257] Dxd, SN-38, and camptothecin inhibit topoisomerase I, which causes DNA damage and subsequent apoptotic cell death, whereas MMAE is a microtubule inhibitor that inhibits mitotic cell division and ultimately induces apoptosis.

[0258] Based on experimental data, the induction of hGDF-15 production in response to ADC treatment is expected to reduce the therapeutic activity of ADCs and synergy with immunotherapies, such as immune checkpoint blockade, by counteracting the immunostimulatory activity of ADC-induced ICDs.

[0259] Accordingly, hGDF-15 is expected to mediate the development of suboptimal anticancer responses and treatment resistance.

[0260] In short, anti-hGDF-15 antibodies capable of neutralizing the function of hGDF-15, or their antigen-binding portions, are expected to act synergistically with tumor-targeted therapy regimens and enhance adaptive anticancer immune responses.

[0261] In vitro cytotoxicity and hGDF-15 induction analysis

[0262] hGDF-15 induction was investigated in the following exemplary cell lines belonging to different tumor entities: lung cancer (NCI-H2122), bladder cancer (RT-4, HT-1376, SW-780), breast cancer (MCF-7, ZR-75-30), ovarian cancer (Caov-3, OVCAR-3), liver cancer (HepG2, JHH-1), gastric cancer (NCI-N87), colorectal cancer (SW-837, NCI-H747), renal cancer (TUHR4TKB, OS-RC-2), and hematological cancer (AMO-1).

[0263] Detach adhered tumor cells from culture dishes using trypsin-EDTA solution, count them, and place 0.4-1 x 10⁶ in each culture medium 5 Adjusted to a cell / mL density. 100 µL of cell suspension was placed in a flat-bottomed 96-well plate at a density of 0.4–1 x 10⁶ per well. 4 Cells were inoculated into wells. Cells were incubated at 37°C and 5% CO2 for 24 hours to allow attachment.

[0264] ADCs, non-conjugated antibodies, or ADC payloads were lysed according to the manufacturer's recommendations and prepared as a 4-fold concentrated 9-step serial dilution series in the appropriate culture medium for each cancer cell line. 10 µl of each drug dilution was added to the inoculated tumor cells, and the culture medium was filled to a volume of 200 µl per well. The assay plates were then transferred to an incubator and incubated for 3 to 5 days at 37°C and 5% CO2.

[0265] After the incubation period ended, 100 µl of the culture supernatant was retrieved, loaded into a new round-bottom 96-well plate, and frozen at -80°C. The tumor cells remaining in the analysis plate were used to perform a CellTiter-Glo viability assay (Promega #G7572) according to the manufacturer's instructions. Using the frozen cell culture supernatant, the concentration of hGDF-15 was determined at the IC50 of each drug using the human (Cat# DY957) DuoSet ELISA kit (R&D Systems) according to the manufacturer's protocol.

[0266] Results Summary

[0267] The inventors demonstrated that hGDF-15 production is induced by ADCs and their non-conjugated drugs (payloads) in a number of tumor entity-derived cell lines (see Figs. 14, 16, 18, 20, and 22).

[0268] Specifically, hGDF-15 production was induced by enfortumab vedotin (see Fig. 14), trastuzumab emtansine (see Fig. 16), trastuzumab deruxtecan (see Fig. 18), and sacituzumab govitecan (see Fig. 20). Induction of apoptosis, as evidenced by reduced cell viability, was also observed in all tested ADCs (see Figs. 13, 15, 17, 19).

[0269] Likewise, the inventors also confirmed that hGDF-15 is induced by non-conjugated payloads of approved ADCs (e.g., see MMAE in Fig. 14, DM-1 in Fig. 16, DxD in Fig. 18, and SN-38 in Fig. 20) or ADCs currently in clinical trials (e.g., see Caliceamicin and SG3199 in Fig. 22). Induction of apoptosis, as evidenced by reduced cell viability, was also observed in all payloads examined (see Figs. 13, 15, 17, 19, and 21).

[0270] MMAE and DM-1 are microtubule inhibitors that inhibit mitotic cell division, ultimately leading to apoptotic cell death. DxD and SN-38 are topoisomerase 1 inhibitory payloads that induce DNA damage, ultimately leading to apoptotic cell death. Chalicemycin and the PBD dimer S3199 are also DNA damage-inducing payloads that cause apoptotic cell death.

[0271] Based on experimental data, the induction of hGDF-15 production in response to ADC treatment is expected to reduce the therapeutic activity of ADCs and synergy with immunotherapies, such as immune checkpoint blockade, by counteracting the immunostimulatory activity of ADCs.

[0272] Therefore, hGDF-15 is expected to mediate the development of suboptimal anticancer responses and treatment resistance.

[0273] Experimental data support the idea that anti-hGDF-15 antibodies capable of neutralizing the function of hGDF-15, or their antigen-binding portions, can act synergistically with tumor-targeted therapy regimens to enhance adaptive anticancer immune responses.

[0274] Major subclasses of ADCs induce a hallmark of immunogenic cell death in vitro:

[0275] Using the RT-4 cell line (bladder cancer), we confirmed ADC / payload-induced killing and the induction of extracellular ATP, HMGB1, and ecto-calreticulin, which are ICD hallmarks.

[0276] Detach attached RT-4 cells from culture dishes using trypsin-EDTA solution, count them, and 1.4 x 10⁶ in each culture medium 5 The density was adjusted to cells / mL. 500 µL of this cell suspension was placed in a flat-bottomed 24-well plate at a volume of 7 x 10⁴ per well. 4 Cells were inoculated into wells. Cells were incubated at 37°C and 5% CO2 for 24 hours to allow attachment.

[0277] ADCs, non-conjugated antibodies, or ADC payloads were lysed according to the manufacturer's recommendations and prepared at a 3-fold concentration in appropriate culture medium, and added to inoculated tumor cells. The assay plates were incubated at 37°C and 5% CO2 for 24, 48, or 72 hours.

[0278] After 72 hours, CellTiter-Glo viability assay (Promega #G7572) was performed using tumor cells from one analysis plate according to the manufacturer's instructions. Another plate was used for ICD evaluation, and the supernatant was collected at various time points to detect ICD markers. eATP was analyzed by RealTime Glo™ (Promega, #GA5010) luminescence reading, and HMGB1 was analyzed using HMGB1 Express ELISA (IBL, #30164033) according to the manufacturer's instructions. After collecting the supernatant, the remaining cells were carefully detached and viability stained (Zombie Violet™ Fixable Viability Kit, Biolegend, #423113). Additionally, the same cells were stained to detect the translocation of caleticulin to the surface by performing a flow cytometry staining protocol using an anti-caleticulin antibody according to the manufacturer's (aCalr-PE, Abcam, #ab209577) instructions. Samples were acquired using a MACSQuant16 (Miltenyi Biotec) flow cytometry device. The geometric mean of PE of viable / dead negative singlet cells was standardized against the untreated control sample.

[0279] Results Summary

[0280] Upregulation of hGDF-15 was observed after treating RT-4 cell lines (bladder cancer) with ADCs containing different payloads, namely microtubule inhibitors or topoisomerase inhibitors (see Figs. 14, 18, and 20). The inventors confirmed the upregulation of known ICD hallmarks in RT-4 cells simultaneously with ADC-induced apoptosis (see Figs. 13, 25, 27, and 29) (see Figs. 24, 26, 28, and 30).

[0281] Specifically, it was confirmed that topoisomerase inhibitors as payloads (see sacituzumab govitecan in Fig. 23 and trastuzumab deruxtecan in Fig. 27) and microtubule inhibitors as payloads (see enfortumab vedotin in Fig. 25 and trastuzumab emtansine in Fig. 29) can induce apoptosis in RT-4 cells, whereas backbone antibodies did not. This apoptosis was accompanied by an increase in eATP, HMGB1, and ecto-caleticulin (see sacituzumab govitecan in Fig. 24, enfortumab vedotin in Fig. 26, trastuzumab deruxtecan in Fig. 28, and trastuzumab emtansine in Fig. 30).

[0282] eATP, HMGB1, and ecto-caleticulin are risk-associated molecular patterns (DAMPs) that form the signature characteristics of immunogenic cell death. These DAMPs subsequently activate antigen-presenting cells, further triggering adaptive immunity against the antigen, and thus contribute to the efficacy of ADCs.

[0283] Based on experimental data, the ability of ADCs to induce ICD hallmarks is expected to contribute to efficacy and antitumor activity. However, the simultaneous increase in hGDF-15 may counteract the immune-enhancing activity of ADC-induced ICDs.

[0284] ADC-induced GDF-15 release is observed in tumors rather than systemic sources.

[0285] To detect human-derived GDF-15 compared to murine-derived GDF-15, human tumors were transplanted into immunocompromised mice using an in vivo study involving a tumor xenograft model.

[0286] To this end, 1 x 10⁶ MCF-7 human breast cancer cells in Matrigel were injected into a total of 45 8-week-old female NMRI nu / nu mice. 7Dogs were inoculated subcutaneously. On day 15 after inoculation, when the tumor volume reached approximately 200 mm³, mice were immediately randomized into study groups of nine. This day was designated as Day 0, and the mice were treated once. Specifically, the mice were intravenously injected with either the vehicle control or the 5 mg / kg antibody-drug conjugate sacituzumab govitecan. Blood samples for serum preparation were collected 24 hours after treatment and again 2 days later. To minimize animal burden, blood was collected from only three mice per group at each time point. Subsequently, serum GDF-15 levels were evaluated using muline GDF-15 ELISA (R&D systems, Cat# DY6385) and human GDF-15 ELISA (R&D systems, Cat# DY957) to distinguish between tumor-origin hGDF-15 and mGDF-15 from surrounding tissues.

[0287] A different xenograft model was used, and 2 x 10⁶ HT-1376 human bladder cancer cells in Matrigel were injected into a total of 54 8-9 week old female NOG mice. 6 Dogs were subcutaneously inoculated into the lateral side of the right hind leg. Thirty-two days after inoculation, when the tumor volume reached approximately 300 mm³, 18 mice were immediately randomized into each study group. This day was designated as Day 0, and the mice were treated once. The mice were intravenously injected with enfortumab vedotin as a vehicle control or a 5 mg / kg antibody-drug conjugate. Blood collection for serum preparation was performed after Day 13. Subsequently, serum GDF-15 levels were evaluated using muline GDF-15 ELISA (R&D systems, Cat# DY6385) and human GDF-15 ELISA (R&D systems, Cat# DY957) to distinguish between tumor-origin hGDF-15 and mGDF-15 from surrounding tissues.

[0288] In addition, animal tumors were excised at the end of the study (Day 13) for further RNA analysis. RT-PCR evaluation was performed on the tumor samples, and the rapid-frozen HT1376 tumors were transferred to an RNA stabilization solution for dissection. Tissue slices were isolated using a rotor-stator device while maintaining an RNAse inhibition environment. RNA was isolated according to the manufacturer's instructions (RNeasy mini kit, Quiagen, #74104). RNA yield and purity were determined using a Nanodrop device. RNA concentrations were matched across all samples, and genomic DNA digestion and reverse transcription were performed according to the manufacturer's instructions (SS IV VILO Master mix, Thermo Fisher Scientific, #11766050). Cycle thresholds were obtained on a Quantstudio 6 flex device using TaqMan FAST master mix (Thermo Fisher Scientific, #4444557) and respective probes (Thermo Fisher Scientific; Gapdh: Hs02786624_g1, #4331182; Gdf15: Hs00171132_m1, #4331182; Cxcl10: Hs00171042_m1, #4331182). Relative gene expression of Gdf15 and IP-10 / CXCL10 was compared to Gapdh using the ΔΔCT method, and the treatment groups were compared to the geometric mean of the vehicle control group for the corresponding target gene and reference gene.

[0289] Results Summary

[0290] The inventors demonstrated that an ADC can target and release hGDF-15 into the serum of mice implanted with tumors of human origin (see Figs. 31-33).

[0291] Specifically, immunocompromised mice transplanted with human MCF-7 cells and then treated with sacituzumab govitecan showed significantly increased levels of human GDF-15 detected in serum collected from the animals at 24 and 48 hours after treatment, but no murine GDF-15 was detected (see Fig. 31).

[0292] Similarly, the inventors confirmed that treatment with enfortumab vedotin in another xenograft mouse model using HT-1376 human tumors increased intratumoral GDF-15 induction as evaluated by RT-PCR (see Fig. 32). This increase in hGDF-15 gene expression could also be interpreted as the detection of higher levels of human GDF-15 protein in the serum of these mice on day 13 (see Fig. 33). No such increase was observed in murine GDF-15 levels (see Fig. 33).

[0293] The inventors also confirmed that GDF-15 expression is accompanied by an increase in ICD-related markers. Specifically, after treatment with enfortumab vedotin, an increase in GDF-15 gene expression occurred simultaneously with an increase in IP-10 / CXCL10 expression at the mRNA level (see Fig. 32). CXCL10 is a chemokine associated with ICD and was used as an ICD hallmark.

[0294] Experimental data support the fact that ADCs carrying topoisomerase 1 inhibitors or microtubule inhibitors as payloads can induce hGDF-15 release from targeted tumors in examined xenograft models. The ADCs induced GDF-15 within the tumor, and simultaneous ICD induction was achieved. The induction of hGDF-15 production in response to ADC treatment is expected to counteract the immunostimulatory activity of ADC-induced ICDs, thereby reducing the therapeutic activity of ADC treatment and synergy with immunotherapies such as immune checkpoint blockade.

[0295] Accordingly, hGDF-15 is expected to mediate the development of suboptimal anticancer responses and treatment resistance.

[0296] Therefore, experimental evidence supports the fact that anti-hGDF-15 antibodies are expected to act synergistically with tumor-targeted therapy regimens and enhance adaptive anti-cancer immune responses.

[0297] The combination of ADC and GDF-15 blockade enhances the in vivo anti-tumor immune response.

[0298] In an in vivo study involving a winter murine model, the efficacy of GDF-15 neutralization combined with ADC treatment was evaluated.

[0299] To this end, MC38, a colorectal cancer cell line overexpressing human Nectin-4, was used for transplantation. A total of 32 female C57BL / 6 mice aged 6–8 weeks were transplanted with 1 x 10⁶ MC38 / hNectin4 tumor cells in 0.1 ml PBS. 6 Dogs were subcutaneously inoculated into the lateral side of the right hind leg. On day 13 following inoculation, the tumors reached an average size of approximately 100 mm³. Subsequently, mice were randomly assigned to four study groups of eight mice each, and treatment was initiated. The treatment start date was designated as Day 0. The mice served as a control group, receiving either no treatment, intravenous injection of 3 mg / kg enfortumab vedotin once a week, or intraperitoneal injection of 10 mg / kg anti-hGDF-15 antibody twice a week. The fourth group received a combination treatment consisting of intravenous injection of 3 mg / kg enfortumab vedotin once a week and intraperitoneal injection of 10 mg / kg anti-hGDF-15 antibody twice a week. Mice were monitored regularly after tumor inoculation, and body weight was measured twice a week. Tumor size was assessed twice a week using calipers, and tumor volume was calculated in mm³. Blood was collected from animals on day 8 after the start of treatment to perform flow cytometry for PBMC. On day 26 after the start of treatment, animals were sacrificed and tumors were analyzed by flow cytometry.

[0300] To perform flow cytometry evaluation after resection, the tumor was washed in an RPMI1640 and any excess tissue was removed. Subsequently, the tumor was weighed to count the absolute number of cells and cut into small pieces. Cells were isolated from the tumor using a Muline Tumor Dissociation Kit (Miltenyi Biotec, Cat#130-096-730) and a gentleMACS tissue separator (Miltenyi) according to the manufacturer's instructions. After obtaining single-cell suspensions, Fc receptors were blocked (Fc block, BD Biosciences, #553142) to prevent non-specific antibody binding, and then surface staining was performed using a fluorescence-conjugated antibody cocktail (aCD45, Biolegend #103149; aCD80, BioLegend #104726; aCD4 BD #612761; aCD8 eBiosciences #61-0081-82 a41BB, BioLegend #106106; aPD1, BD #744546; aCD39, BD #567295). For blood samples, erythrocyte lysis was performed. Subsequently, cells were fixed and permeated using a commercially available kit (eBioscience, #00-5523-00) to enable flow cytometry staining. For tumor samples, the absolute cell count was determined by adding counting beads. Subsequently, data was acquired via flow cytometry and analyzed using Kaluza software. The absolute cell count was calculated using a unified standard equation that combines the number of cells and beads, volume, and bead concentration.

[0301] Results Summary

[0302] The inventors demonstrated that a neutralizing anti-GDF-15 antibody improves the anti-tumor efficacy of ADC therapy. Specifically, tumor regression, enhanced immune infiltration, and increased activation of T cells and macrophages were achieved when the anti-GDF-15 antibody was combined with an ADC (see Figs. 34-38).

[0303] Precisely, using the MC38 / hNectin-4 muline model, the combination of enfortumab vedotin and the anti-GDF-15 antibody achieved tumor growth inhibition (see Figs. 34 and 36). This tumor regression was accompanied by increased intratumoral immune infiltration, as confirmed by an increase in intratumoral %CD45+ cell levels resulting from the combination of GDF-15 neutralization using the anti-GDF-15 antibody and enfortumab vedotin (see Fig. 37). Furthermore, the combination of the ADC and the anti-GDF-15 antibody also achieved intratumoral APC activation, as supported by an increase in CD80-expressing macrophages (see Fig. 37).

[0304] In addition, the inventors demonstrated that the combination of GDF-15 neutralization and ADC treatment further activates CD4 T cells in PBMCs, as evidenced by an increase in 41BB+ CD4 T cells (see Fig. 38). Likewise, in the combination group (ADC, anti-GDF-15 antibody), more CD8 T cells were detected, and more CD8 T cells expressing CD39+ and PD1+ were detected (see Fig. 38).

[0305] Experimental data support the fact that GDF-15 reduces the therapeutic activity of ADC therapy and that GDF-15 neutralization enhances the anti-tumor efficacy of ADCs. Therefore, it is expected that GDF-15 neutralization will improve synergy with immunotherapies, such as ADC activation and immune checkpoint blockade, by counteracting GDF-15-mediated therapeutic resistance and suppressing ADC-induced immune cell activation.

[0306] In short, anti-GDF-15 antibodies capable of neutralizing the function of GDF-15, or their antigen-binding portions, are expected to act synergistically with tumor-targeted therapy regimens and enhance adaptive anticancer immune responses.

[0307] GDF-15 blockade enhances the efficacy of combined ADCs and immune checkpoint blockades (ICBs) / immune checkpoint inhibitors (ICIs) in vivo.

[0308] In an in vivo study involving a winter murine model, the efficacy of combining GDF-15 neutralization with an ADC and an anti-PD1 antibody (aPD1) was investigated.

[0309] MC38, a colorectal cancer cell line overexpressing human Nectin-4, was used for transplantation. A total of 64 female C57BL / 6 mice aged 6–8 weeks were transplanted with 1 x 10⁶ MC38 / hNectin4 tumor cells in 0.1 ml PBS. 6 Dogs were subcutaneously inoculated into the side of the right hind leg. On day 13 after inoculation, the tumors reached an average size of approximately 100 mm³. Subsequently, mice were randomly assigned to four study groups of eight mice each to begin treatment. The treatment start date was designated as day 0. Mice were either left untreated as a control group, or received intraperitoneal injections of 10 mg / kg anti-GDF-15 antibody twice weekly, 10 mg / kg anti-PD-1 antibody twice weekly, or 3 mg / kg enfortumab vedotin once weekly. In the combination groups, a combination of intravenous injection of 3 mg / kg enfortumab vedotin once weekly and intraperitoneal injections of 10 mg / kg anti-GDF-15 antibody or 10 mg / kg anti-PD-1 antibody twice weekly was administered. The other control group was administered a combination of intraperitoneal injections of 10 mg / kg anti-GDF-15 antibody and 10 mg / kg anti-PD-1 antibody twice a week. The triple combination group included all three of these compounds. Mice were monitored regularly after tumor inoculation, and body weight was measured twice a week. Blood was collected from the animals on day 8 after the start of treatment for flow cytometry of PBMCs.

[0310] Erythrocyte lysis was performed for flow cytometry of blood. Subsequently, cells were fixed and permeated to enable flow cytometry staining using a commercially available kit (eBioscience, #00-5523-00). Nuclear staining was performed using a fluorescence-conjugated antibody cocktail (aCD45, Biolegend #103149; aCD4 BD #612761; aCD8 eBiosciences #61-0081-82; Ki67 eBioscience, #69-5698-82; CD44 Biolegend #103006; CD62L Biolegend #104440). Afterward, data were acquired via flow cytometry and analyzed using Kaluza software.

[0311] In another synaptic murine model, MC38 cancer cells overexpressing human Trop-2 were used for transplantation. A total of 64 6–8-week-old female C57BL / 6-hTrop2 mice expressing the human Trop2 epitope were transplanted with 1 x 10⁶ MC38 / hTrop2 tumor cells in 0.1 ml PBS. 6Dogs were subcutaneously inoculated into the side of the right hind leg. On day 7 after inoculation, the tumors reached an average size of approximately 100 mm³. Subsequently, mice were randomly assigned to four study groups of eight mice each to begin treatment. The treatment start date was designated as Day 0. Mice were either left untreated as a control group, or received intraperitoneal injections of 10 mg / kg anti-GDF-15 antibody twice a week, 0.3 mg / kg anti-PD-1 antibody once a week, or 10 mg / kg sacituzumab govitecan once a week. The combination groups were administered a combination of intravenous injection of 10 mg / kg sacituzumab govitecan once a week and intraperitoneal injections of 10 mg / kg anti-GDF-15 antibody or 0.3 mg / kg anti-PD-1 antibody twice a week. Another control group was administered a combination of intraperitoneal injections of 10 mg / kg anti-GDF-15 antibody and 0.3 mg / kg anti-PD-1 antibody twice weekly. The triple combination group included all three compounds. After tumor inoculation, mice were monitored regularly, and body weight was measured twice weekly. Tumor size was assessed twice weekly using a caliper, and tumor volume was calculated in mm³. Kaplan-Meier analysis was performed to indicate animal viability following various treatments.

[0312] Results Summary

[0313] The inventors confirmed that the therapeutic efficacy of an ADC is improved when combined with an immune checkpoint block (ICB) / inhibition (ICI) upon GDF-15 neutralization. When an anti-GDF-15 antibody was combined with an ADC and an ICI, enhanced proliferation of circulating T cells was achieved (see Fig. 39). Using the MC38 / hNectin-4 muline model, cell proliferation of CD4 and CD8 central memory T cells was increased in the combination of enfortumab vedotin, aPD1, and aGDF-15 (see Fig. 39). Ki67 is a classic proliferation marker, and higher levels of Ki67 expression were observed in CD4 and CD8 T cells upon GDF-15 neutralization in addition to combination therapy with enfortumab vedotin and aPD1.

[0314] The inventors also experimentally demonstrated in an MC38 / hTrop-2 mouse model that a triple combination consisting of an anti-GDF-15 antibody, an ADC, namely sacituzumab govitecan, and an anti-PD1 antibody, improves therapeutic efficacy compared to an ADC combined with either an anti-GDF-15 antibody or an anti-PD1 antibody, as evidenced by improved animal viability in the triple combination group.

[0315] Therefore, experimental data demonstrate that GDF-15 reduces the therapeutic activity of combination therapies using ADCs and anti-PD1 antibodies, and that the efficacy of combination therapies using ADCs and immune checkpoint inhibition / blockade is enhanced upon GDF-15 neutralization. Accordingly, the neutralization of GDF-15 is also expected to improve synergy with immunotherapies, such as ADC activation and immune checkpoint blockade, by counteracting GDF-15-mediated therapeutic resistance and the suppression of GDF-15-mediated ADC-induced immunostimulatory activity.

[0316] In short, anti-GDF-15 antibodies capable of neutralizing the function of GDF-15, or their antigen-binding portions, are expected to act synergistically with tumor-targeted therapy regimens to enhance adaptive anticancer immune responses.

[0317] order

[0318] SEQ ID NO. 1 (Peptide sequence of the heavy chain CDR1 region of the monoclonal anti-human GDF-15 antibody):

[0319] GFSLSTSGMG

[0320] SEQ ID NO. 2 (Peptide sequence of the heavy chain CDR2 region of the monoclonal anti-human GDF-15 antibody):

[0321] IYWDDDK

[0322] SEQ ID NO. 3 (Peptide sequence of the heavy chain CDR3 region of the monoclonal anti-human GDF-15 antibody):

[0323] ARSSYGAMDY

[0324] SEQ ID NO. 4 (Peptide sequence of the light chain CDR1 region of the monoclonal anti-human GDF-15 antibody):

[0325] QNVGTN

[0326] Light chain CDR2 region peptide sequence of monoclonal anti-human GDF-15 antibody:

[0327] SAS

[0328] SEQ ID NO. 5 (Peptide sequence of the light chain CDR3 region of the monoclonal anti-human GDF-15 antibody):

[0329] QQYNNFPYT

[0330] Sequence No. 6 (Heavy chain variability domain of monoclonal anti-human GDF-15 antibody):

[0331] QITLKESGPTLVKPTQTLTLTCTFSGFSLTSGMGVSWIRQPPGKGLEWLAHIYWDDDKRYNPTLKSRLTITKDPSKNQVVLTMTNMDPVDTATYYGAMDYWGQGTLVTVSSASTKGP

[0332] Sequence No. 7 (Light chain variability domain of monoclonal anti-human GDF-15 antibody):

[0333] DIVLTQSPSFLSASVGDRVTITCKASQNVGTNVAWFQQKPGKSPKALIYSASYRYSGVPDRFTGSGSGTEFTLTISSLQPEDFAAYFCQQYNNFPYTFGGGTKLEIKRT

[0334] SEQ ID NO. 8 (Heavy chain of monoclonal anti-human GDF-15 antibody CTL-002 lacking leader peptide sequence):

[0335] QITLKESGPTLVKPTQTLTLTCTFSGFSLTSGMGVSWIRQPPGKGLEWLAHIYWDDDKRYNPTLKSRLTITKDPSKNQVVLTMTNMDPVDTATYYCARSSYGAMDYWGQG TLVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKT ISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLG

[0336] SEQ ID NO. 9 (Light chain of monoclonal anti-human GDF-15 antibody CTL-002 lacking leader peptide sequence):

[0337] DIVLTQSPSFLSASVGDRVTITCKASQNVGTNVAWFQQKPGKSPKALIYSASYRYSGVPDRFTGSGSGTEFTLTISSLQPEDFAAYFCQQYNNFPYTFGGGTKLEIK RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0338] SEQ ID NO. 10 (Heavy chain variability domain of anti-human GDF-15 antibody H1L5):

[0339] QITLKESGPTLVKPTQTLTLTCTFSGFSLTSGMGVSWIRQPPGKGLEWLAHIYWDDDKRYNPTLKSRLTITKDPSKNQVVLTMTNMDPVDTATYYCARSSYGAMDYWGQG TLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDK THTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK TISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG

[0340] SEQ ID NO. 11 (Light chain variability domain of anti-human GDF-15 antibody H1L5):

[0341] DIVLTQSPSFLSASVGDRVTITCKASQNVGTNVAWFQQKPGKSPKALIYSASYRYSGVPDRFTGSGSGTEFTLTISSLQPEDFAAYFCQQYNNFPYTFGGGTKLEIK RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0342] SEQ ID NO. 12 (GDF-15 peptide containing a portion of the GDF-15 epitope binding to B1-23):

[0343] EVQVTMCIGACPSQFR

[0344] SEQ ID NO. 13 (GDF-15 peptide containing a portion of the GDF-15 epitope binding to B1-23):

[0345] TDTGVSLQTYDDLLAKDCHCI

[0346] Sequence No. 14 (Recombinant mature hGDF-15 protein):

[0347] GSARNGDHCPLGPGRCCRLHTVRASLEDLGWADWVLSPREVQVTMCIGACPSQFRAANMHAQIKTSLHRLKPDTVPAPCCVPASYNPMVLIQKTDTGVSLQTYDDLLAKDCHCI

[0348] Sequence No. 15 (heavy chain of ponsegromab):

[0349] QVQLVQSGAEVKKPGSSVKVSCKASGYTFSSYNIDWVRQAPGQGLEWMGGINPIFGTAFYNQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCAREAITTVGAMDHWGQ GTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCD KTHTCPPCPAPEAAGAPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIE KTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG

[0350] Sequence No. 16 (Light chain of ponsegromab):

[0351] EIVLTQSPATLSLSPGERATLSCRTSQSVHNYLAWYQQKPGQAPRLLIYDASTRADGIPARFSGSGSGTDFTLTISSLEPEDFAVYYCQQFWSWPWTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0352] 서열번호 17 (릴로그로투그의 중쇄):

[0353] QVQLVQSGAEVKKPGSSVKVSCKASGYTFSDYNMDWVRQAPGQGLEWMGQINPNNGLIFFNQKFKGRVTLTADKSTSTAYMELSSLRSEDTAVYYCAREAITTVGAMDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

[0354] 서열번호 18 (릴로그로투그의 경쇄):

[0355] DIQMTQSPSSLSASVGDRVTITCRTSENLHNYLAWYQQKPGKAPKLLIYDAKTLADGVPSRFSGSGSGTDYTLTISSLQPEDFATYYCQHFWSSPYTFGQGTKLEIK RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0356] Sequence No. 19 (heavy chain variable region of AZD8853):

[0357] QVQLVQSGSELKKPGASVKVSCKASGYTFTDYNMDWIRQSPGKGLEWIGDINPNQGGTFYNQKFKDRATLTVDKSTSTAYMELRSLRSDDTAVYYCAREEKLYFGLLMDYWGQGTTVTVSS

[0358] Sequence No. 20 (Light chain variable region of AZD8853):

[0359] DIQMTQSPSSLSASVGDRVTITCRASENIYSYLAWYQQKPGKAPKLLVYNAKTLAEGVPSRFSGSGSGIDFTLTISSLQPEDFATYYCQHQYGSPPTFGQGTKLEIK Industrial applicability

[0360] The anti-hGDF-15 antibody or its antigen-binding portion used for cancer treatment in human patients can be industrially manufactured and marketed as a product for the specified methods and uses in accordance with the stated pharmaceutical manufacturing standards. Thus, the present invention is industrially available.

Claims

Claim 1 An anti-hGDF-15 antibody or its antigen-binding portion for use in a method of treating cancer in human patients in combination with a cancer antigen-targeting drug conjugate that induces cancer cell stress. Claim 2 In claim 1, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the cancer cell stress is immunogenic. Claim 3 In claim 1 or 2, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the cancer cell stress is associated with the induction of hGDF-15 expression in the patient. Claim 4 In any one of claims 1 to 3, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the cancer antigen-targeting drug conjugate induces apoptosis of cancer cells. Claim 5 In any one of claims 1 to 4, an anti-hGDF-15 antibody or its antigen-binding portion, wherein the death of the cancer cells is immunogenic cell death (ICD). Claim 6 In any one of claims 1 to 5, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the immunogenic cell death (ICD) is associated with the induction of hGDF-15 expression in the patient. Claim 7 In any one of claims 1 to 6, the anti-hGDF-15 antibody or its antigen-binding portion is a neutralizing anti-hGDF-15 antibody or its antigen-binding portion. Claim 8 An anti-hGDF-15 antibody or its antigen-binding portion according to any one of claims 1 to 7, wherein the anti-hGDF-15 antibody or its antigen-binding portion comprises a heavy chain variable domain comprising a CDR1 region represented by the amino acid sequence presented in SEQ ID NO. 1, a CDR2 region represented by the amino acid sequence presented in SEQ ID NO. 2, and a CDR3 region represented by the amino acid sequence presented in SEQ ID NO. 3, and a light chain variable domain comprising a CDR1 region represented by the amino acid sequence presented in SEQ ID NO. 4, a CDR2 region represented by the amino acid sequence ser-ala-ser, and a CDR3 region represented by the amino acid sequence presented in SEQ ID NO.

5. Claim 9 An anti-hGDF-15 antibody or its antigen-binding portion according to any one of claims 1 to 8, wherein the anti-hGDF-15 antibody or its antigen-binding portion comprises a heavy chain variable domain having at least 90% identity, preferably at least 95% identity, more preferably at least 98% identity, with an amino acid sequence represented by SEQ ID NO. 6 or an amino acid sequence presented in SEQ ID NO. 6, and a light chain variable domain having at least 90% identity, preferably at least 95% identity, more preferably at least 98% identity, with an amino acid sequence represented by SEQ ID NO. 7 or an amino acid sequence presented in SEQ ID NO.

7. Claim 10 An anti-hGDF-15 antibody or its antigen-binding portion according to any one of claims 1 to 9, wherein the anti-hGDF-15 antibody or its antigen-binding portion comprises a heavy chain having at least 90% identity, preferably at least 95% identity, more preferably at least 98% identity, with the amino acid sequence represented by SEQ ID NO. 8 or the amino acid sequence presented in SEQ ID NO. 8, and a light chain having at least 90% identity, preferably at least 95% identity, more preferably at least 98% identity, with the amino acid sequence represented by SEQ ID NO. 9 or the amino acid sequence presented in SEQ ID NO.

9. Claim 11 In any one of claims 1 to 10, the anti-hGDF-15 antibody or its antigen-binding portion having a heavy chain comprising an amino acid sequence represented by SEQ ID NO. 8 and a light chain comprising an amino acid sequence represented by SEQ ID NO.

9. Claim 12 An anti-hGDF-15 antibody or its antigen-binding portion, wherein, in any one of claims 1 to 10, the anti-hGDF-15 antibody or its antigen-binding portion competes with the antibody defined in claim 11 for specific binding to hGDF-15. Claim 13 An anti-hGDF-15 antibody or its antigen-binding portion according to any one of claims 1 to 12, wherein the anti-hGDF-15 antibody or its antigen-binding portion binds to a stereotypical or discontinuous epitope on hGDF-15 consisting of the amino acid sequences of SEQ ID NO. 12 and SEQ ID NO.

13. Claim 14 An anti-hGDF-15 antibody or its antigen-binding portion according to any one of claims 1 to 7, wherein the anti-hGDF-15 antibody or its antigen-binding portion is visugromab, ponsegromab, or rilologrotug, or is an antibody having a heavy chain variable region formed of SEQ ID NO. 19 and a light chain variable region formed of SEQ ID NO.

20. Claim 15 In any one of claims 1 to 14, the anti-hGDF-15 antibody or the antigen-binding portion thereof is bisugromab. Claim 16 In any one of claims 1 to 15, the anti-hGDF-15 antibody or its antigen-binding portion comprising a cancer cell stress-inducing drug connected to the cancer antigen-targeting portion of the cancer antigen-targeting drug conjugate. Claim 17 In paragraph 16, the above drug is an anti-hGDF-15 antibody or its antigen-binding portion, which is an anticancer agent. Claim 18 In paragraph 16 or 17, the above drug is an anti-hGDF-15 antibody or its antigen-binding portion, which is a chemotherapy drug. Claim 19 In any one of claims 16 to 18, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the drug is a cytotoxic drug. Claim 20 In claim 19, the cytotoxic drug is an anti-hGDF-15 antibody or its antigen-binding portion, selected from the group consisting of microtubule inhibitors, topoisomerase I or II inhibitors, DNA-damaging agents, protein synthesis inhibitors, RNA polymerase III inhibitors, transcription inhibitors, apoptosis inducers, NAMPT inhibitors, proteasome inhibitors, kinase inhibitors, PROTAC, NIR-PIT drugs, and immune-activating substances. Claim 21 In claim 19 or 20, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the cytotoxic drug is a microtubule inhibitor or a topoisomerase I or II inhibitor. Claim 22 In any one of claims 19 to 21, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the cytotoxic drug is a microtubule inhibitor. Claim 23 In any one of claims 20 to 22, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the microtubule inhibitor is selected from the group consisting of monomethylauristatin E (MMAE), monomethylauristatin F (MMAF), methansin (DM-1), auristatin F-HPA, auristatin-0101, DM21, DM4, metansinoid, eribulin, and SC209. Claim 24 In claim 20 or 21, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the topoisomerase I inhibitor is selected from the group consisting of Dxd, SN-38, irinotecan, topotecan, rubitecan, exatecan, belotecan, MLN576, and camptothecin. Claim 25 In claim 20 or 21, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the topoisomerase II inhibitor is selected from the group consisting of etoposide, idarubicin, mitoxantrone, PNU-159682, daunorubicin, teniposide, epirubicin, and doxorubicin. Claim 26 In paragraph 20, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the DNA damaging agent is selected from the group consisting of caliceamycin, SG3199 / PBD dimer, PBD, and indolinobenzodiazepine (IGN). Claim 27 In claim 20, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the protein synthesis inhibitor is selected from the group consisting of PE38, geldanamycin, tylanstatin A, and carmapicin B analogs. Claim 28 In claim 20, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the RNA polymerase III inhibitor is selected from the group consisting of α-amanitine, beta-amanitine, phalloidin, trichothecene T-2, velucarin A, and loridin A. Claim 29 In paragraph 20, the above transcription inhibitor is an anti-hGDF-15 antibody or its antigen-binding portion selected from the group consisting of triptolide, ST7464AA1, vorinostat, and dashinostat. Claim 30 In paragraph 20, the above-mentioned apoptosis inducer is an anti-hGDF-15 antibody or its antigen-binding portion, which is a BCL-XL inhibitor such as clezutoclax. Claim 31 In paragraph 20, the above NAMPT inhibitor is an anti-hGDF-15 antibody or its antigen-binding portion, which is an FK-866 analog. Claim 32 In claim 20, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the proteasome inhibitor is selected from the group consisting of bortezomib, carfilzomib, ixazomib, and carmapicin B analogs. Claim 33 In paragraph 20, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the kinase inhibitor is selected from the group consisting of genistein, neolympostine, dasatinib, and staurosporine. Claim 34 In claim 20, the above PROTAC is an anti-hGDF-15 antibody or its antigen-binding portion, wherein the PROTAC is selected from the group consisting of BET / BRD degrading agent (GNE-987, MZ1 analog, BRD4 / VHL, BRD4 / CRBN), ERa degrading agent (ERa / XIAP, ERa / VHL), TGFbR2 degrading agent (TGFbR2 / VHL), BRM degrading agent (BRL / VHL), and GSTP1 degrading agent (SMOL006). Claim 35 In paragraph 20, the above NIR-PIT drug is an anti-hGDF-15 antibody or its antigen-binding portion selected from water-soluble phthalocyanine derivatives such as silicon phthalocyanine derivative IR700 (IRDye700DX). Claim 36 In paragraph 20, the immune-activating substance is an anti-hGDF-15 antibody or its antigen-binding portion, selected from the group consisting of STING agonists, TLR7 and / or TLR8 agonists and botolimod. Claim 37 In any one of claims 16 to 19, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the drug causing cancer cell stress is selected from the group consisting of MMAE, MMAF / auristatin-F, Dxd, DM-1, SN-38, camptothecin, and DM-4. Claim 38 In paragraph 37, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the drug causing the cancer cell stress is MMAE. Claim 39 In any one of claims 16 to 38, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the drug is connected to the cancer antigen-targeting portion through a cleavable linker. Claim 40 In any one of claims 16 to 38, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the drug is connected to the cancer antigen-targeting portion via a non-cleavable linker. Claim 41 In any one of claims 16 to 40, an anti-hGDF-15 antibody or its antigen-binding portion, wherein the cancer antigen-targeting portion of the cancer antigen-targeting drug conjugate binds to a cancer cell. Claim 42 An anti-hGDF-15 antibody or its antigen-binding portion, wherein the cancer antigen-targeting portion is a ligand, peptide, or antibody in any one of claims 16 to 41. Claim 43 In paragraph 42, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the cancer antigen-targeting portion is an antibody. Claim 44 In any one of claims 41 to 43, an anti-hGDF-15 antibody or its antigen-binding portion, wherein the cancer antigen-targeting portion binds to a target antigen on a cancer cell. Claim 45 In paragraph 44, the target antigen is not hGDF-15, but an anti-hGDF-15 antibody or its antigen-binding portion. Claim 46 In claim 44 or 45, the target antigen is Nectin-4, HER2, Trop-2, TF (CD142), CD30, CD22, CD79b, BCMA, CD19, FRα, CD33, LIV-1, HER3, CD25, NaPi2b, B7-H4, c-Met, B7-H3, B7-H4, PTK7, ADAM9, CEACAM5, 5T4, ALK, AXL, GRP20, CDH6, TA-MUC1, KAAG1, DLK1, DLL3, SLAMF7, CA125, C4.4A / LYPD3, CDH3, CDH6, CAIX, CD20, CD26 / DPP4, CD37, CD38, CD138, CD46, ICAM4 / CD54, CD56 / NCAM1, CD70, CD73, CD74, CD205, CD248, C-KIT, CLDN6, CLDN18.2, CLL-1, RET, CRIPTO, DLK-1, DLL3, EGFR, CD105, ENPP3, EPCAM, EPHA2, FAP, FGFR2 / CD332, FLT3, GDNF / GFRA1, GPC2, GPNMB, guanylyl cyclase (GCC), IGF-1R, ITGAV, Sialyl-di-Lewis, LGR5, LIV1A, LRRC15, MSLN, STEAP1, PSMA, TMEFF2, NOTCH3, PTK7, SLC44A4, SLC46A3, SLITRK6, TIM-1, LY6E, Cadherin, PD-L1, CD228, FOLR1, CTLA4, GPR20, HGFR, CD123, An anti-hGDF-15 antibody or its antigen-binding portion selected from the group consisting of PSMA, ROR1, and ETBR. Claim 47 In claim 46, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the target antigen is selected from Nectin-4, HER2, Trop-2, TF (CD142), CD30, CD22, CD79b, BCMA, CD19, FRα and CD33. Claim 48 In any one of claims 1 to 47, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the cancer antigen-targeting drug conjugate is an antibody-drug conjugate (ADC). Claim 49 In claim 48, the said ADC is Enfortumab vedotin, trastuzumab deruxtecan, trastuzumab emtansine, sacituzumab govitecan, tisotumab vedotin, Brentuximab vedotin, Inotuzumab ozogamicin, moxetumomab pasudotox, polatuzumab vedotin, belantamab mafodotin, loncastuximab tesirine, mirvetuzimab sorabtansine-gynx (Mirvetuximab soravtansine-gynx), Gemtuzumab ozogamicin, disitamab vedotin, radiratuzumab vedotin, datopotamab deruxtecan, patritumab deruxtecan, camidanlumab tesirine, upifitamab rilsodotin, XMT-1592, XMT-1660, XMT-2056, telisotuzumab vedotin, ABBV-400, mirzotamab clezutoclax, cofetuzumab felidotin (cofetuzumab pelidotin), MORAb-202 (parletuzumab), STRO-002, IMGN632, IMGC936, ASP-1929, isacituzumab govitecan,An anti-hGDF-15 antibody or its antigen-binding portion selected from the group consisting of SKB264 and tusamitamab ravtansine. Claim 50 In claim 49, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the ADC is selected from the group consisting of enfortumab vedotin, trastuzumab deruxtecan, trastuzumab emtansine, datopotamab deruxtecan and sacituzumab govitecan. Claim 51 In any one of claims 1 to 50, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the cancer is a solid tumor. Claim 52 In claim 51, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the cancer is selected from the group consisting of urothelial carcinoma (UC), non-small cell lung cancer (NSCLC), pancreatic cancer, head and neck cancer, breast cancer, colorectal cancer, anal cancer, gastric cancer, liver cancer, biliary cancer, ovarian cancer, prostate cancer, gastric cancer, esophageal cancer, kidney cancer, thyroid cancer, endometrial cancer, cervical cancer, testicular cancer, melanoma, and skin cancer. Claim 53 An anti-hGDF-15 antibody or its antigen-binding portion, wherein the cancer is urothelial carcinoma and the drug of the cancer antigen-targeting drug conjugate is a microtubule inhibitor. Claim 54 In paragraph 53, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the microtubule inhibitor is MMAE. Claim 55 In paragraph 53 or 54, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the cancer antigen-targeting drug conjugate is enfortumab vedotin. Claim 56 In claim 51 or 52, the cancer is breast cancer, and the drug of the cancer antigen-targeting drug conjugate is a topoisomerase I inhibitor, an anti-hGDF-15 antibody or its antigen-binding portion. Claim 57 In paragraph 56, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the topoisomerase I inhibitor is Dxd. Claim 58 In paragraph 56 or 57, the cancer antigen-targeting drug conjugate is trastuzumab deruxtecan, an anti-hGDF-15 antibody, or its antigen-binding portion. Claim 59 In claim 51 or 52, the cancer is breast cancer, and the drug of the cancer antigen-targeting drug conjugate is a topoisomerase I inhibitor, an anti-hGDF-15 antibody or its antigen-binding portion. Claim 60 In paragraph 59, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the topoisomerase I inhibitor is SN-38. Claim 61 In paragraph 59 or 60, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the cancer antigen-targeting drug conjugate is sacituzumab govitecan. Claim 62 An anti-hGDF-15 antibody or its antigen-binding portion, wherein the cancer is NSCLC and the drug of the cancer antigen-targeting drug conjugate is a topoisomerase I inhibitor. Claim 63 In paragraph 62, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the topoisomerase I inhibitor is Dxd. Claim 64 In paragraph 62 or 63, the cancer antigen-targeting drug conjugate is trastuzumab deruxtecan, an anti-hGDF-15 antibody, or its antigen-binding portion. Claim 65 An anti-hGDF-15 antibody or its antigen-binding portion for use in a method of treating cancer in human patients, wherein said anti-hGDF-15 antibody or its antigen-binding portion is administered in combination with one or more antibody-drug conjugates. Claim 66 In paragraph 65, the anti-hGDF-15 antibody or its antigen-binding portion is as defined in any one of paragraphs 7 to 15. Claim 67 In paragraph 65 or 66, the antibody-drug conjugate is an anti-hGDF-15 antibody or its antigen-binding portion as defined in any one of paragraphs 48 to 50. Claim 68 In any one of claims 65 to 67, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the antibody-drug conjugate induces cancer cell stress. Claim 69 In any one of paragraphs 65 to 68, the cancer is an anti-hGDF-15 antibody or its antigen-binding portion as defined in paragraph 51 or 52. Claim 70 In any one of paragraphs 65 to 69, the cancer and the drug are an anti-hGDF-15 antibody or its antigen-binding portion as defined in any one of paragraphs 53 to 64. Claim 71 In any one of claims 65 to 70, the anti-hGDF-15 antibody or its antigen-binding portion and antibody-drug conjugate (ADC) are administered in combination with an immune checkpoint blocker (ICB), optionally, the ICB is selected from a list of PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors and LAG-3 inhibitors, preferably a PD-1 inhibitor or a PD-L1 inhibitor, the anti-hGDF-15 antibody or its antigen-binding portion. Claim 72 In claim 71, the anti-hGDF-15 antibody or its antigen-binding portion, wherein the ICB is selected from the group consisting of nivolumab, atezolizumab, pembrolizumab, avelumab, cemiplimab, durvalumab, dostalimab, retipanlimab, toripalimab, syntilimab, tislellizumab, camrelizumab, valstilimab, envapolimab, pakmilimab, cosivelimab, and spartalizumab, preferably nivolumab or pembrolizumab. Claim 73 An antibody-drug conjugate (ADC) for use in a method of treating cancer in human patients, wherein the antibody-drug conjugate is administered in combination with an anti-hGDF-15 antibody or its antigen-binding portion. Claim 74 In paragraph 73, the antibody-drug conjugate (ADC) wherein the anti-hGDF-15 antibody or its antigen-binding portion is as defined in any one of paragraphs 7 to 15. Claim 75 In paragraph 73 or 74, the antibody-drug conjugate is an antibody-drug conjugate (ADC) as defined in any one of paragraphs 48 to 50. Claim 76 An antibody-drug conjugate (ADC) according to any one of claims 73 to 75, wherein the antibody-drug conjugate induces cancer cell stress. Claim 77 In any one of paragraphs 73 to 76, the cancer is an antibody-drug conjugate (ADC) as defined in paragraph 51 or 52. Claim 78 In any one of paragraphs 73 to 77, the cancer and the drug are antibody-drug conjugates (ADCs) as defined in any one of paragraphs 53 to 64. Claim 79 An antibody-drug conjugate (ADC) according to any one of claims 73 to 78, wherein the antibody-drug conjugate (ADC) and the anti-hGDF-15 antibody or its antigen-binding portion is administered in combination with an immune checkpoint blocker (ICB), and wherein the ICB is selected from a list consisting of PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors and LAG-3 inhibitors, and preferably a PD-1 inhibitor or a PD-L1 inhibitor. Claim 80 An antibody-drug conjugate (ADC) according to claim 79, wherein the ICB is selected from the group consisting of nivolumab, atezolizumab, pembrolizumab, avelumab, cemiplimab, durvalumab, dostalimab, retipanlimab, toripalimab, syntilimab, tislellizumab, camrelizumab, valstilimab, envapolimab, pakmilimab, cosivelimab, and spartalizumab, preferably nivolumab or pembrolizumab. Claim 81 A combination product composed of an anti-hGDF-15 antibody or its antigen-binding portion and an antibody-drug conjugate for use in cancer treatment methods for human patients. Claim 82 A combination product according to claim 81, wherein the anti-hGDF-15 antibody or its antigen-binding portion is as defined in any one of claims 7 to 15. Claim 83 A combination product according to claim 81 or 82, wherein the antibody-drug conjugate is as defined in any one of claims 48 to 50. Claim 84 A combination product according to any one of claims 81 to 83, wherein the antibody-drug conjugate induces cancer cell stress. Claim 85 A combination product according to any one of paragraphs 81 to 84, wherein the arm is as defined in paragraph 51 or 52. Claim 86 A combination product in any one of paragraphs 81 to 85, wherein the cancer and the drug are as defined in any one of paragraphs 53 to 64. Claim 87 A combination product according to any one of claims 81 to 86, wherein the antibody-drug conjugate (ADC) and the anti-hGDF-15 antibody or its antigen-binding portion is administered in combination with an immune checkpoint blocker (ICB), optionally, the ICB is selected from a list of PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors and LAG-3 inhibitors, preferably a PD-1 inhibitor or a PD-L1 inhibitor. Claim 88 A combination product according to claim 87, wherein the ICB is selected from the group consisting of nivolumab, atezolizumab, pembrolizumab, avelumab, cemiplimab, durvalumab, dostalimab, retipanlimab, toripalimab, syntilimab, tislellizumab, camrelizumab, valstilimab, envapolimab, pakmilimab, cosivelimab, and spartalizumab, preferably nivolumab or pembrolizumab. Claim 89 A kit comprising an anti-hGDF-15 antibody or its antigen-binding portion and an antibody-drug conjugate. Claim 90 In claim 89, the above anti-hGDF-15 antibody or its antigen-binding portion is as defined in any one of claims 7 to 15, a kit. Claim 91 A kit according to claim 89 or 90, wherein the antibody-drug conjugate is as defined in any one of claims 48 to 50. Claim 92 A kit according to any one of claims 89 to 91, wherein the antibody-drug conjugate induces cancer cell stress. Claim 93 A kit according to any one of claims 89 to 92, further comprising an immune checkpoint blocker (ICB), optionally wherein the ICB is selected from a list of PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors and LAG-3 inhibitors, preferably a PD-1 inhibitor or a PD-L1 inhibitor. Claim 94 A kit according to claim 93, wherein the ICB is selected from the group consisting of nivolumab, atezolizumab, pembrolizumab, avelumab, cemiplimab, durvalumab, dostalimab, retipanlimab, toripalimab, syntilimab, tislellizumab, camrelizumab, valstilimab, envapolimab, pakmilimab, cocibelimab, and spartalizumab, preferably nivolumab or pembrolizumab. Claim 95 A pharmaceutical composition comprising a combination of an anti-hGDF-15 antibody and an antibody-drug conjugate. Claim 96 A pharmaceutical composition according to claim 95, wherein the anti-hGDF-15 antibody or its antigen-binding portion is as defined in any one of claims 7 to 15. Claim 97 A pharmaceutical composition according to claim 95 or 96, wherein the antibody-drug conjugate is as defined in any one of claims 48 to 50. Claim 98 A pharmaceutical composition according to any one of claims 95 to 97, wherein the antibody-drug conjugate induces cancer cell stress. Claim 99 A pharmaceutical composition according to any one of claims 95 to 98, further comprising an immune checkpoint blocker (ICB), optionally wherein the ICB is selected from a list of PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors and LAG-3 inhibitors, preferably a PD-1 inhibitor or a PD-L1 inhibitor. Claim 100 A pharmaceutical composition according to claim 99, wherein the ICB is selected from the group consisting of nivolumab, atezolizumab, pembrolizumab, avelumab, cemiplimab, durvalumab, dostalimab, retipanlimab, toripalimab, syntilimab, tislellizumab, camrelizumab, valstilimab, envapolimab, pakmilimab, cocibelimab, and spartalizumab, preferably nivolumab or pembrolizumab. Claim 101 Use of an anti-hGDF-15 antibody or its antigen-binding portion combined with a cancer antigen-targeting drug conjugate in the manufacture of a drug for the treatment of cancer in human patients, optionally wherein the cancer antigen-targeting drug conjugate is an antibody-drug conjugate (ADC). Claim 102 Use of an anti-hGDF-15 antibody or its antigen-binding portion in the manufacture of a drug for the treatment of cancer in human patients, wherein said drug further comprises a cancer antigen-targeting drug conjugate, and optionally, said cancer antigen-targeting drug conjugate is an antibody-drug conjugate (ADC). Claim 103 Use of an antibody-drug conjugate (ADC) in the manufacture of a drug for the treatment of cancer in human patients, wherein the drug further comprises an anti-hGDF-15 antibody or an antigen-binding portion thereof. Claim 104 In any one of claims 101 to 103, the anti-hGDF-15 antibody or its antigen-binding portion and antibody-drug conjugate (ADC) are administered in combination with an immune checkpoint blocker (ICB), optionally, said ICB is selected from a list consisting of PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors and LAG-3 inhibitors, preferably a PD-1 inhibitor or a PD-L1 inhibitor, and additionally optionally, said ICB is selected from the group consisting of nivolumab, atezolizumab, pembrolizumab, avelumab, cemiplimab, durvalumab, dostalimab, retipanlimab, toripalimab, syntilimab, tislellizumab, camrelizumab, valstilimab, envapolimab, pakmilimab, cocibelimab and spartalizumab, preferably nivolumab or Pembrolizumab, use. Claim 105 A use in any one of claims 101 to 104, wherein the anti-hGDF-15 antibody or its antigen-binding portion is as defined in any one of claims 7 to 15. Claim 106 In any one of claims 101 to 105, the use of the antibody-drug conjugate as defined in any one of claims 48 to 50. Claim 107 In any one of claims 101 to 106, the use wherein the cancer antigen-targeting drug conjugate or antibody-drug conjugate induces cancer cell stress. Claim 108 An anti-hGDF-15 antibody or its antigen-binding portion, antibody-drug conjugate (ADC), combination product or use according to any one of claims 1 to 72, 73 to 80, 81 to 88, or 101 to 107, wherein the anti-hGDF-15 antibody or its antigen-binding portion and the antigen-targeting drug conjugate or antibody-drug conjugate are administered at the same or different times.

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