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US-12624119-B2 - Isolated antigen-binding protein and application thereof

US12624119B2US 12624119 B2US12624119 B2US 12624119B2US-12624119-B2

Abstract

An isolated antigen-binding protein, having one or more of the following properties: 1) capable of binding to human and monkey-derived GITR proteins at a K D value of 7×10 −12 or below, wherein the K D value is measured by BLI method; 2) capable of stimulating immune cell proliferation; 3) capable of stimulating immune cells to secrete IFN-γ, wherein the secretion is measured in T cell viability assay; 4) capable of inhibiting tumor growth and/or tumor cell proliferation; 5) capable of activating GITR signaling pathway; 6) capable of inhibiting the binding of GITR to GITRL.

Inventors

  • Xin Zhang
  • Erxia SHI
  • Ting Xu
  • Hui Ma
  • Yangyang YUAN
  • Shanshan NING
  • Shilong FU
  • Xiaolong PAN
  • Liyao ZHOU
  • Meng Zhao

Assignees

  • SHIHUIDA PHARMACEUTICAL GROUP (JILIN) CO., LTD.

Dates

Publication Date
20260512
Application Date
20200716
Priority Date
20190717

Claims (17)

  1. 1 . An isolated antigen-binding protein, having one or more of the following properties: 1) Capable of binding to human and monkey-derived GITR proteins at a K D value of 7×10 −12 M or below, wherein the K D value is measured by Bio-Layer Interferometry (BLI) method; 2) capable of stimulating immune cell proliferation; 3) capable of stimulating immune cells to secrete IFN-γ, wherein the secretion is measured in T cell viability assay; 4) capable of inhibiting tumor growth and/or tumor cell proliferation; 5) capable of activating GITR signaling pathway; 6) capable of inhibiting the binding of GITR to GITRL, wherein VH of the isolated antigen-binding protein comprises HCDR3 as set forth in SEQ ID NO: 4, HCDR1 as set forth in SEQ ID NO: 2, and HCDR2 as set forth in SEQ ID NO: 3, and wherein VL of the isolated antigen-binding protein comprises LCDR1 as set forth in SEQ ID NO: 10, LCDR2 as set forth in SEQ ID NO: 11, and LCDR3 as set forth in SEQ ID NO: 12.
  2. 2 . The isolated antigen-binding protein according to claim 1 , wherein the VL comprises framework regions L-FR1, L-FR2, L-FR3, and L-FR4, wherein a C-terminus of the L-FR1 is directly or indirectly linked to an N-terminus of the LCDR1, wherein the L-FR1 comprises an amino acid sequence as shown in any one of SEQ ID NOs: 13, 30, 34.
  3. 3 . The isolated antigen-binding protein according to claim 2 , wherein the L-FR2 is located between the LCDR1 and the LCDR2, and the L-FR2 comprises an amino acid sequence as shown in any one of SEQ ID NOs: 14, 31.
  4. 4 . The isolated antigen-binding protein according to claim 2 , wherein the L-FR3 is located between the LCDR2 and the LCDR3, and the L-FR3 comprises an amino acid sequence as shown in any one of SEQ ID NOs: 15, 32.
  5. 5 . The isolated antigen-binding protein according to claim 2 , wherein an N-terminus of the L-FR4 is linked to a C-terminus of the LCDR3, and the L-FR4 comprises an amino acid sequence as shown in any one of SEQ ID NOs: 16, 33.
  6. 6 . The isolated antigen-binding protein according to claim 1 , wherein the VL comprises an amino acid sequence as shown in any one of SEQ ID NOs: 9, 20-22.
  7. 7 . The isolated antigen-binding protein according to claim 1 , comprising an antibody light chain constant region, and the antibody light chain constant region comprises a human Igκ constant region.
  8. 8 . The isolated antigen-binding protein according to claim 1 , comprising an antibody light chain LC, and the LC comprises an amino acid sequence as shown in any one of SEQ ID NOs: 64, 46-48.
  9. 9 . The isolated antigen-binding protein according to claim 1 , wherein the VH comprises framework regions H-FR1, H-FR2, H-FR3, and H-FR4, a C-terminus of the H-FR1 is directly or indirectly linked to an N-terminus of the HCDR1, and wherein the H-FR1 comprises an amino acid sequence as shown in any one of SEQ ID NOs: 5, 25, 29.
  10. 10 . The isolated antigen-binding protein according to claim 9 , wherein the H-FR2 is located between the HCDR1 and the HCDR2, and the H-FR2 comprises an amino acid sequence as shown in any one of SEQ ID NOs: 6, 26.
  11. 11 . The isolated antigen-binding protein according to claim 9 , wherein the H-FR3 is located between the HCDR2 and the HCDR3, and the H-FR3 comprises an amino acid sequence as shown in any one of SEQ ID NOs: 7, 27.
  12. 12 . The isolated antigen-binding protein according to claim 9 , wherein an N-terminus of the H-FR4 is linked to a C-terminus of the HCDR3, and the H-FR4 comprises an amino acid sequence as shown in any one of SEQ ID NOs: 8, 28.
  13. 13 . The isolated antigen-binding protein according to claim 1 , wherein the VH comprises an amino acid sequence as shown in any one of SEQ ID NOs: 1, 17-19.
  14. 14 . The isolated antigen-binding protein according to claim 1 , comprising an antibody heavy chain constant region, and the antibody heavy chain constant region is derived from a human IgG1 heavy chain constant region.
  15. 15 . The isolated antigen-binding protein according to claim 1 , comprising an antibody heavy chain HC, and the HC comprises an amino acid sequence as shown in any one of SEQ ID NOs: 63, 43-45.
  16. 16 . A pharmaceutical composition, comprising the isolated antigen-binding protein according to claim 1 , as well as optionally a pharmaceutically acceptable adjuvant.
  17. 17 . A method for activating GITR, comprising administering the isolated antigen-binding protein according to claim 1 .

Description

CROSS-REFERENCE TO RELATED APPLICATIONS This application is a National Stage Application and claims priority under 35 U.S.C. § 371 to Patent Cooperation Treaty application PCT/CN2020/102472, filed Jul. 16, 2020, which claims the benefit of Chinese application CN 201910646093.8 filed Jul. 17, 2019. Priority is claimed to these applications and the disclosure of these prior applications is considered part of the disclosure of this application and to the extent allowed the entire contents of the aforementioned applications are incorporated herein. SEQUENCE LISTING This application incorporates by reference in its entirety the Sequence Listing entitled “2022 Jan. 13_262790-502844_ST25.txt”, is 95,099 bytes in size and was created on Jan. 13, 2022, and filed electronically herewith. FIELD OF THE INVENTION The present application relates to the field of biomedicine, and particularly to an isolated antigen-binding protein and an application thereof. BACKGROUND OF THE INVENTION GITR is a member of the TNF receptor family. GITR is a type I transmembrane protein with a molecular weight of about 26 kDa, which has 14-28% homology with other molecules in the TNF receptor family (Gurney A L, et al, 1999). GITR is constitutively expressed on Treg at a high level, but expressed on unactivated CD4+ T cells, CD8+ T cells, NK and NKT cells at lower levels. When T cells are activated, GITR is highly expressed in CD4+ T, CD8+ T and NK cells (S. Ronchetti, et al., 2004). The natural ligand of GITR, GITRL, is expressed on APC. After GITRL binds to GITR, it provides costimulatory signals, regulates antigen-specific T cell responses, and strengthens cellular and humoral immunity (Selvakumar Sukumar, et al., 2017). SUMMARY OF THE INVENTION The present application provides an isolated antigen-binding protein, having one or more of the following properties: 1) capable of binding to human and monkey-derived GITR proteins at a KD value of 7×10−12 M or below, wherein the KD value is measured by BLI method; 2) capable of stimulating immune cell proliferation; 3) capable of stimulating immune cells to secrete IFN-7, wherein the secretion is measured in T cell viability assay; 4) capable of inhibiting tumor growth and/or tumor cell proliferation; 5) capable of activating GITR signaling pathway; 6) capable of inhibiting the binding of GITR to GITRL. In some embodiments, the isolated antigen-binding protein of the present application comprises at least one CDR in VH whose amino acid sequence is shown in SEQ ID NO: 23. In some embodiments, the isolated antigen-binding protein of the present application comprises HCDR3 in VH whose amino acid sequence is shown in SEQ ID NO: 23. In some embodiments, the isolated antigen-binding protein of the present application comprises HCDR2 in VH whose amino acid sequence is shown in SEQ ID NO: 23. In some embodiments, the isolated antigen-binding protein of the present application comprises HCDR1 in VH whose amino acid sequence is shown in SEQ ID NO: 23. In some embodiments, the isolated antigen-binding protein of the present application comprises at least one CDR in VL whose amino acid sequence is shown in SEQ ID NO: 24. In some embodiments, the isolated antigen-binding protein of the present application comprises LCDR1 in VL whose amino acid sequence is shown in SEQ ID NO: 24. In some embodiments, the isolated antigen-binding protein of the present application comprises LCDR2 in VL whose amino acid sequence is shown in SEQ ID NO: 24. In some embodiments, the isolated antigen-binding protein of the present application comprises LCDR3 in VL whose amino acid sequence is shown in SEQ ID NO: 24. In some embodiments, the HCDR3 comprises an amino acid sequence as shown in SEQ ID NO: 4. In some embodiments, the HCDR1 comprises an amino acid sequence as shown in SEQ ID NO: 2. In some embodiments, the HCDR2 comprises an amino acid sequence as shown in SEQ ID NO: 3. In some embodiments, the LCDR1 comprises an amino acid sequence as shown in SEQ ID NO: 10. In some embodiments, the LCDR2 comprises an amino acid sequence as shown in SEQ ID NO: 11. In some embodiments, the LCDR3 comprises an amino acid sequence as shown in SEQ ID NO: 12. In some embodiments, the isolated antigen-binding protein of the present application comprises an antibody or an antigen-binding fragment thereof. In some embodiments, the antigen-binding fragment comprises Fab, Fab′, F(ab)2, Fv fragment, F(ab′)2, scFv, di-scFv and/or dAb. In some embodiments, the antibody is a humanized antibody. In some embodiments, the isolated antigen-binding protein of the present application competes with a reference antibody for binding to GITR proteins, wherein the reference antibody comprises HCDR1-3 in VH whose amino acid sequence is shown in SEQ ID NO: 23 as well as LCDR1-3 in VL whose amino acid sequence is shown in SEQ ID NO: 24. In some embodiments, the HCDR1-3 of the reference antibody include amino acid sequences as shown in SEQ ID NO: 2, SEQ ID NO: 3 and