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CN-121992039-A - Method for enhancing CAR-T cell function

CN121992039ACN 121992039 ACN121992039 ACN 121992039ACN-121992039-A

Abstract

The invention discloses a method for enhancing the function of CAR-T cells. In particular, the use of an Id2 protein to enhance CAR-T cell activation, proliferation, killing and tumor growth inhibition, cytokine secretion, persistence and/or anti-tumor effects is disclosed. Also disclosed is a method of enhancing CAR-T cell function comprising increasing the level and/or activity of an Id2 protein in a CAR-T cell. According to the invention, id2 is fused on the CAR gene to construct the CAR-T cell over-expressed by Id2, experiments prove that the Id2 protein can enhance the persistence, activation and proliferation capacity, killing capacity, cytokine secretion capacity and the like of the CAR-T cell, and the killing capacity of the CAR-T cell to tumor cells can be obviously enhanced in the CAR-T cell of both human and mice, so that the anti-tumor effect is improved, and the CAR-T cell has stronger tumor treatment effect.

Inventors

  • ZHANG FUPING
  • SHI YIXUAN
  • GAO PING
  • LI CHAO
  • KANG YUCHEN

Assignees

  • 中国科学院微生物研究所

Dates

Publication Date
20260508
Application Date
20241101

Claims (10)

  1. Use of the id2 protein in any one of the following: A1 For enhancing CAR-T cell activation and/or proliferation capacity or for the preparation of a product for enhancing CAR-T cell activation and/or proliferation capacity; A2 Use in enhancing the killing capacity of CAR-T cells or in the preparation of a product for enhancing the killing capacity of CAR-T cells; A3 Use in promoting secretion of cytokines by CAR-T cells or in the preparation of a product for promoting secretion of cytokines by CAR-T cells; A4 Use in improving CAR-T cell persistence or in the manufacture of a product for improving CAR-T cell persistence; A5 Use in increasing the ability of CAR-T cells to inhibit tumor growth or in the manufacture of a product for increasing the ability of CAR-T cells to inhibit tumor growth; a6 To improve the anti-tumor effect of the CAR-T cells or to prepare a product for improving the anti-tumor effect of the CAR-T cells.
  2. 2. A fusion protein comprising an Id2 protein and a chimeric antigen receptor.
  3. 3. The fusion protein of claim 2, wherein the chimeric antigen receptor targets CD19.
  4. 4. Use of a biomaterial, characterized in that the use is any one of the following: b1 For enhancing CAR-T cell activation and/or proliferation capacity or for the preparation of a product for enhancing CAR-T cell activation and/or proliferation capacity; b2 Use in enhancing the killing capacity of CAR-T cells or in the preparation of a product for enhancing the killing capacity of CAR-T cells; b3 Use in promoting secretion of cytokines by CAR-T cells or in the preparation of a product for promoting secretion of cytokines by CAR-T cells; b4 Use in improving CAR-T cell persistence or in the manufacture of a product for improving CAR-T cell persistence; B5 Use in increasing the ability of CAR-T cells to inhibit tumor growth or in the manufacture of a product for increasing the ability of CAR-T cells to inhibit tumor growth; b6 The application of the anti-tumor effect of the CAR-T cells is improved or the product for improving the anti-tumor effect of the CAR-T cells is prepared; The biological material is any one of the following: c1 A nucleic acid molecule encoding the Id2 protein of claim 1; c2 A nucleic acid molecule encoding the fusion protein of claim 2 or 3; C3 An expression cassette comprising the nucleic acid molecule of C1) or C2); c4 A recombinant vector comprising the nucleic acid molecule of C1) or C2); c5 A recombinant microorganism comprising a nucleic acid molecule according to C1) or C2); C6 A recombinant host cell comprising a nucleic acid molecule according to C1) or C2).
  5. 5. A method of improving or enhancing CAR-T cell function, comprising increasing the level and/or activity of an Id2 protein in a CAR-T cell.
  6. 6. The method of claim 5, wherein said increasing the level and/or activity of the Id2 protein in the CAR-T cell is achieved by increasing the expression level of a gene encoding the Id2 protein in the CAR-T cell.
  7. 7. The method of claim 5 or 6, wherein increasing the expression of the gene encoding the Id2 protein in the CAR-T cell is achieved by introducing the gene encoding the Id2 protein into the T cell after fusion with the CAR gene.
  8. 8. A modified T cell comprising the fusion protein of claim 2 or 3.
  9. 9. A method of making the modified T cell of claim 8, comprising introducing a nucleic acid molecule encoding the fusion protein of claim 2 or 3 into a T cell for expression, thereby obtaining the modified T cell.
  10. 10. Use of the Id2 protein of claim 1, the fusion protein of claim 2 or 3, or the biomaterial of claim 4 in the preparation of CAR-T cells.

Description

Method for enhancing CAR-T cell function Technical Field The invention belongs to the technical field of cellular immunotherapy, and particularly relates to a method for enhancing the function of CAR-T cells. Background Chimeric antigen receptor-T cell (CHIMERIC ANTIGENS receptor-T cell, CAR-T) and T cell receptor-engineered T cell (ENGINEERED T CELL receptor-T cell, TCR-T) therapies have achieved surprising success in tumor therapy as representative therapies for cellular immunotherapy. Three scientists in Gross G, 1989, for the first time, proposed the concept of CAR, indicating that conferring gene sequences expressing specific antibodies to cytotoxic T Cells (CTL) could allow T cells to achieve antigen-specific, non-MHC restricted activation and enhancement of effects. T cells expressing the CAR can recognize and bind to tumor antigens, thereby attacking the tumor cells. The CAR-T cell therapy has shown good targeting, killing and persistence on hematological tumor in clinical test, provides a new solution for immune cell therapy, and has great development potential and application prospect. CARs are engineered synthetic receptors, typically consisting of an extracellular domain, a transmembrane domain, and an intracellular domain. The extracellular domain is mainly composed of a single-chain variable fragment (scFv) responsible for recognizing and binding to an antigen and a Linker (Linker) that serves to link VH and VL. Since the extracellular domain of CAR-T is a universal antibody sequence, CAR-expressing T cells are not MHC-restricted, thus exhibiting stronger T cell activation and anti-tumor responses. The transmembrane domain is responsible for linking the extracellular domain and the intracellular domain of the CAR molecule, thereby transducing ligand recognition signals into the cell. The intracellular domain is composed primarily of costimulatory and signaling domains, the costimulatory domain providing primarily the second signal for CAR-T cell activation, the most common costimulatory domains being CD28 and 4-1BB. The signal transduction domain is mainly CD3 zeta. After the CAR recognizes the tumor antigen, tyrosine of ITAM motif of CD3 zeta is phosphorylated, ZAP70 is recruited, and then kinase activation cascade reaction is started, finally, the CAR-T cell is thoroughly activated, various transcription factors are activated, and proliferation and differentiation of the T cell and expression of an activation gene are promoted. Like effector T cells, CAR-T cells can directly mediate apoptosis of tumor cells via the perforin/granzyme pathway after activation. Over thirty years of development, CAR-T cells can be divided into 4 stages of development, no co-stimulatory signal CAR-T (generation 1), single co-stimulatory signal CAR-T (generation 2), dual/multi co-stimulatory signal CAR-T (generation 3), and precise CAR-T (generation 4), to date. Since 2017 FDA approved North and Ji Lide CAR-T cell products were marketed, several CAR-T products have been marketed worldwide at present, all for use in the treatment of hematological malignancies. The injection comprises CAR-T, carvykti which is first obtained in China, namely the Sidamascene injection, and is mainly used for treating multiple myeloma. Nevertheless, CAR-T immunotherapy is still imperfect, and there are many problems in clinical treatment, such as limited ability of CAR-T to infiltrate solid tumors, short survival time, poor effector capacity, etc. in inhibitory tumor microenvironment, many research teams are exploring innovative therapies to meet clinical needs. Therefore, the novel CAR-T cells and the therapies thereof are intensively researched and developed, the structure of the CAR is improved and perfected, the problems of insufficient proliferation capability, impaired effector function and the like of the CAR-T cells in a tumor microenvironment are solved, the proliferation capability and the anti-tumor capability of the CAR-T cells are enhanced, and the novel CAR-T cells have important significance and wide application value for improving the clinical curative effect, can benefit more patients, and are beneficial to promoting the commercialization process of the CAR-T cells. Disclosure of Invention The technical problem to be solved by the invention is how to improve or enhance the function of CAR-T cells. The technical problems to be solved are not limited to the described technical subject matter, and other technical subject matter not mentioned herein will be clearly understood by those skilled in the art from the following description. To solve the above technical problem, the present invention provides first the use of an Id2 protein in any of the following: A1 For enhancing CAR-T cell activation and/or proliferation capacity or for the preparation of a product for enhancing CAR-T cell activation and/or proliferation capacity; A2 Use in enhancing the killing capacity of CAR-T cells or in the preparation of a product for en