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CN-121991219-A - Antibody coupling drug targeting TM4SF20 and preparation method and application thereof

CN121991219ACN 121991219 ACN121991219 ACN 121991219ACN-121991219-A

Abstract

The invention relates to the technical field of antibody coupling medicaments. The invention provides an antibody coupling drug targeting TM4SF20, a preparation method and application thereof, wherein the antibody comprises VLCDR1, VLCDR2 and VLCDR3 with amino acid sequences shown as SEQ ID NO 1-3, and VHCDR1, VHCDR2 and VHCDR3 with amino acid sequences shown as SEQ ID NO 4-6. The invention verifies that the specificity of the kit is high in pancreatic cancer through in vivo and in vitro experiments and clinical samples, is related to poor prognosis of pancreatic cancer, designs a neutralizing antibody alpha-TM 4SF20 aiming at a TM4SF20 target spot, verifies that the alpha-TM 4SF20 can be well internalized into cells through an intracellular experiment, couples the alpha-TM 4SF20 and Dxd into ADC drugs, and verifies the effectiveness and the safety of the ADC drugs through in vivo and in vitro experiments.

Inventors

  • HAO JIHUI
  • CHANG ANTAO
  • LIU XINYUE
  • LIU WEISHUAI
  • WU CHAO

Assignees

  • 天津市肿瘤医院（天津医科大学肿瘤医院）

Dates

Publication Date
20260508
Application Date
20250902

Claims (10)

  1. 1. An antibody targeting TM4SF20 is characterized by comprising VLCDR1, VLCDR2 and VLCDR3 with amino acid sequences shown as SEQ ID NO 1-3, and VHCDR1, VHCDR2 and VHCDR3 with amino acid sequences shown as SEQ ID NO 4-6.
  2. 2. The antibody of claim 1, wherein the nucleotide sequence encoding VLCDR1 is shown in SEQ ID NO. 7, the nucleotide sequence encoding VLCDR2 is shown in SEQ ID NO. 8, the nucleotide sequence encoding VLCDR3 is shown in SEQ ID NO. 9, the nucleotide sequence encoding VHCDR1 is shown in SEQ ID NO. 10, the nucleotide sequence encoding VHCDR2 is shown in SEQ ID NO. 11, and the nucleotide sequence encoding VHCDR3 is shown in SEQ ID NO. 12.
  3. 3. The antibody of claim 1, wherein the amino acid sequence of the light chain of the antibody is shown in SEQ ID NO. 13 and the amino acid sequence of the heavy chain is shown in SEQ ID NO. 14.
  4. 4. The antibody of claim 3, wherein the light chain of the antibody has a nucleotide sequence shown in SEQ ID NO. 15 and the heavy chain has a nucleotide sequence shown in SEQ ID NO. 16.
  5. 5. The use of the antibody of any one of claims 1-4 in the preparation of an antibody-conjugated drug targeting TM4SF 20.
  6. 6. An antibody conjugated drug targeting TM4SF20, characterized in that the antibody conjugated drug consists of an antibody according to any one of claims 1-4, a linker and a cytotoxic agent, said antibody and cytotoxic agent being linked via a linker.
  7. 7. The antibody conjugated drug of claim 6, wherein the linker is a tetrapeptide chain having an amino acid sequence GGFG and the cytotoxic agent is a DX-8951 derivative.
  8. 8. A method of preparing an antibody-conjugated drug according to claim 6 or 7, comprising the steps of: (1) Mixing DPBS, the antibody of any one of claims 1-4 and TCEP, and incubating to obtain a mixed solution 1; (2) Arginine is added into the mixed solution to obtain mixed solution 2; (3) And adding MC-GGFG-DXD solution into the mixed solution 2, adding DMSO, and then incubating to obtain the antibody coupling drug.
  9. 9. The preparation method of claim 8, wherein the molar equivalent ratio of the antibody to TCEP in the step (1) is 1:10-14, the imparting time is 100-140 min, the final concentration of arginine in the step (2) is 450-470 mM, the solvent of the MC-GGFG-DXD solution in the step (3) is DMSO, the molar equivalent ratio of the antibody to MC-GGFG-DXD is 1:14-18, the DMSO accounts for 8-12% of the total volume after the DMSO addition, and the incubation time is 50-70 min.
  10. 10. Use of an antibody-conjugated drug according to claim 6 or 7 for the preparation of a product for the treatment of pancreatic cancer.

Description

Antibody coupling drug targeting TM4SF20 and preparation method and application thereof Technical Field The invention relates to the technical field of antibody coupling medicines, in particular to an antibody coupling medicine targeting TM4SF20, and a preparation method and application thereof. Background Pancreatic cancer is malignancy with mortality ranking global 4, with survival rates of less than 9% in 5 years. Traditional pancreatic cancer treatment means mainly comprise surgical excision, chemical drug treatment (chemotherapy for short) and radioactive treatment (radiotherapy for short). For resectable pancreatic cancer, surgery is the treatment of choice. However, since pancreatic cancer is often associated with lymph node invasion or distant organ metastasis at the beginning of the onset, only less than 20% of patients have surgical treatment opportunities at the time of diagnosis. For critically resectable (borderline resectable, BR) and locally advanced (locally advanced, LA) pancreatic cancer patients, while conventional chemotherapy regimens, based on FOLFIRINOX and AG regimens, can to some extent enhance the surgical resection rate of pancreatic cancer, the overall prognosis of pancreatic cancer patients is not significantly improved. For pancreatic cancer which cannot be resected by surgery, chemotherapy and radiotherapy are the main treatment modes at present, but the comprehensive treatment effect of the pancreatic cancer is not improved obviously because the pancreatic cancer lacks effective targeted treatment drugs, has the characteristics of high fibrosis of the essence of tumor and cold tumor, and the conventional chemotherapy scheme is extremely easy to generate drug resistance and strong chemotherapy side effect in a short period. Immunotherapy has made significant progress in cancer treatment in recent years as a new therapeutic strategy. However, in pancreatic cancer, immunotherapy still faces a series of challenges and bottlenecks. On one hand, the immune microenvironment of pancreatic cancer is abnormal and complex, the tumor evades various immune attack modes, and the current immune treatment method often has the problems of unstable curative effect, large side effect and the like. Currently, in addition to chemotherapy, radiotherapy and immunotherapy, antibody-drug conjugate (ADC) drugs are increasingly developed. The ADC medicine is based on the theory assumption of magic bullets, and can search and develop medicines which only kill pathogens and do not affect normal tissues and cells aiming at the special structural characteristics of the pathogens. Currently, 16 ADC drugs are commercially available worldwide, and the treatment field relates to lymphoma, leukemia, pancreatic cancer, multiple myeloma, pancreatic cancer, head and neck cancer, urothelial cancer and the like. However, in pancreatic cancer, no ADC drugs are currently marketed. Therefore, a target spot with high specificity expression in pancreatic cancer is searched, and an ADC medicine is designed aiming at the target spot, so that a new treatment idea is hopeful to be provided for diagnosis and treatment of pancreatic cancer. Disclosure of Invention The invention aims to provide an antibody coupling drug targeting TM4SF20, a preparation method and application thereof, a specific high-expression protein target point in pancreatic cancer is searched, a specific antibody aiming at the target point is designed, an ADC drug is further synthesized, and the effectiveness and safety of the ADC drug are verified through in-vitro experiments of cells and in-vivo experiments of mice and animals. In order to achieve the above object, the present invention provides the following technical solutions: The invention provides an antibody targeting TM4SF20, which comprises VLCDR1, VLCDR2 and VLCDR3 with amino acid sequences shown as SEQ ID NO. 1-3, and VHCDR1, VHCDR2 and VHCDR3 with amino acid sequences shown as SEQ ID NO. 4-6. Preferably, the nucleotide sequence encoding VLCDR1 is shown as SEQ ID NO. 7, the nucleotide sequence encoding VLCDR2 is shown as SEQ ID NO. 8, the nucleotide sequence encoding VLCDR3 is shown as SEQ ID NO. 9, the nucleotide sequence encoding VHCDR1 is shown as SEQ ID NO. 10, the nucleotide sequence encoding VHCDR2 is shown as SEQ ID NO. 11, and the nucleotide sequence encoding VHCDR3 is shown as SEQ ID NO. 12. Preferably, the amino acid sequence of the light chain of the antibody is shown as SEQ ID NO. 13, and the amino acid sequence of the heavy chain is shown as SEQ ID NO. 14. Preferably, the nucleotide sequence of the light chain of the antibody is shown as SEQ ID NO. 15, and the nucleotide sequence of the heavy chain is shown as SEQ ID NO. 16. The invention also provides application of the antibody in preparation of an antibody coupling drug targeting TM4SF 20. The invention also provides an antibody conjugate drug targeting TM4SF20, which consists of the antibody, a connector and a cytotoxic agent, w