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CN-121991246-A - Double-targeting chimeric antigen receptor specifically binding CD70 and BCMA and application thereof

CN121991246ACN 121991246 ACN121991246 ACN 121991246ACN-121991246-A

Abstract

The present invention relates to a dual targeting chimeric antigen receptor targeting CD70 and BCMA and uses thereof, which has the ability to specifically bind to CD70 and BCMA. The immune effector cell prepared based on the double-targeting chimeric antigen receptor has the advantages of greatly reducing the probability of tumor cells escaping treatment by losing a single antigen, improving the targeting treatment effect, greatly shortening the length of an inserted fragment by using a CD70 nano antibody sequence, reducing the development difficulty of a process end, having excellent proliferation capability, being beneficial to prolonging the in vivo duration and playing a more remarkable anti-tumor role. The present invention provides a therapeutic or ameliorating approach for diseases associated with BCMA-CD70 expression.

Inventors

  • GU MINGYUAN
  • WANG HAIYING
  • XIONG QINGHUI

Assignees

  • 上海恒润达生生物科技股份有限公司

Dates

Publication Date
20260508
Application Date
20241108

Claims (10)

  1. 1. A dual targeting chimeric antigen receptor that targets CD70 and BCMA, wherein said dual targeting chimeric antigen receptor comprises an anti-CD 70 VHH antibody, and an anti-BCMA antibody or antigen binding fragment thereof.
  2. 2. The dual targeting chimeric antigen receptor according to claim 1, wherein the anti-CD 70VHH antibody comprises HCDR1-3 as shown in SEQ ID NO. 11-13, wherein the anti-BCMA antibody or antigen binding fragment thereof comprises HCDR1-3 as shown in SEQ ID NO. 15-17 and LCDR1-3 as shown in SEQ ID NO. 18-20, preferably wherein the anti-CD 70VHH antibody has the amino acid sequence as shown in SEQ ID NO. 14 and wherein the anti-BCMA antibody has the amino acid sequence as shown in SEQ ID NO. 21.
  3. 3. The dual targeted chimeric antigen receptor according to claim 1, comprising a first CAR comprising an extracellular antigen binding domain, a hinge region, a transmembrane domain, an intracellular co-stimulatory domain targeting CD70 and a second CAR comprising an extracellular antigen binding domain, a hinge region, a transmembrane domain, an intracellular co-stimulatory domain and an intracellular signaling domain targeting BCMA, preferably comprising a self-cleaving peptide between the first CAR and the second CAR, more preferably selected from the group consisting of P2A, T2A, E a and F2A.
  4. 4. The dual targeting chimeric antigen receptor according to claim 3, wherein, The hinge region is selected from the group consisting of a CD 8a hinge region, a CD8 hinge region, a CD28 hinge region, an IgD hinge region, an IgG1 Fc CH2CH3 hinge region, and an IgG4 Fc CH2CH3 hinge region; The transmembrane domain is selected from the group consisting of CD3 epsilon, CD4, CD5, CD8 alpha, CD9, CD16, CD22, CD28, CD33, CD37, CD45, CD64, CD80, CD86, CD134, CD137, CD154, the alpha, beta or zeta chain of a T cell receptor; the intracellular co-stimulatory domain is derived from at least one of the 4-1BB、CARD11、CD2、CD7、CD27、CD28、CD30、CD40、CD54、CD83、OX40、CD137、CD134、CD150、CD152、CD223、CD270、PD-L2、PD-L1、CD278、DAP10、LAT、NKD2C、SLP76、TRIM、FcεRIγ、MyD88 and 41BBL intracellular domains; The intracellular signaling domain is derived from at least one of the CD3 zeta, fcrgamma (FCER 1G), fcrbeta (fcepsilon Rib), CD3 gamma, CD3 delta, CD3 epsilon, CD5, CD22, CD79a, CD79b, and CD66d intracellular domains.
  5. 5. The dual targeting chimeric antigen receptor according to claim 1, wherein the dual targeting chimeric antigen receptor has an amino acid sequence as set forth in any one of SEQ ID NOs 1 to 5.
  6. 6. A nucleic acid molecule having a sequence selected from any one of: (1) A coding sequence for a dual targeting chimeric antigen receptor according to any one of claims 1 to 5; (2) The complement of (1).
  7. 7. A nucleic acid construct comprising the nucleic acid molecule of claim 6, preferably the nucleic acid construct is a cloning vector, an expression vector or an integration vector.
  8. 8. A host cell selected from the group consisting of: (1) Expressing and/or secreting a dual targeting chimeric antigen receptor according to any of claims 1 to 5; (2) Comprising the nucleic acid molecule of claim 6, and/or (3) Comprising the nucleic acid construct of claim 7, Preferably, the host cell is an immune effector cell, more preferably a T cell.
  9. 9. A pharmaceutical composition comprising the dual-targeting chimeric antigen receptor of any one of claims 1-5, the nucleic acid molecule of claim 6, the nucleic acid construct of claim 7, or the host cell of claim 8, and a pharmaceutically acceptable adjuvant.
  10. 10. Use of a chimeric antigen receptor according to any one of claims 1 to 5, a nucleic acid molecule according to claim 6, a nucleic acid construct according to claim 7 or a host cell according to claim 8 for the preparation of an activated immune cell or for the preparation of a medicament for the prevention or treatment of a disease or condition associated with CD70 or BCMA expression, Preferably, the disease or condition is selected from one or more of multiple myeloma, acute myelogenous leukemia, non-hodgkin's lymphoma, mantle cell lymphoma, diffuse large cell lymphoma, follicular lymphoma, renal cell carcinoma, pancreatic carcinoma, breast carcinoma, glioblastoma.

Description

Double-targeting chimeric antigen receptor specifically binding CD70 and BCMA and application thereof Technical Field The invention relates to the technical field of biological immunotherapy, in particular to a double-targeting chimeric antigen receptor specifically combined with CD70 and BCMA and application thereof. Background BCMA, also known as TNFRSF17, is a B cell surface molecule belonging to the tumor necrosis factor receptor (tumor necrosis factor receptor, TNFR) family, which is a type I transmembrane receptor. BCMA is mainly expressed on the surface of mature B cells and plasma cells, and can be combined with two ligands of B cell activating factors BAFF or APRIL respectively, thereby playing an important role in the maturation and autoimmune reaction of B cells. BCMA is the receptor most selectively expressed on multiple myeloma cells, and its expression level gradually increases with the differentiation of B cells, and also gradually increases in the course of multiple myeloma. CD70 is a type II transmembrane protein belonging to the tumor necrosis factor family, and is normally expressed predominantly in activated T cells, B cells and mature dendritic cells. Activating the receptor CD27 can promote the activation, proliferation and differentiation of T cells and B cells and regulate immune response. Under pathological conditions, CD70 can be expressed in solid tumors and blood tumor cells, wherein 42% -63% of tumor samples of patients with multiple myeloma are CD70 positive. Analysis of recurrent primary multiple myeloma samples confirmed that 19 cd138+ plasma cells expressed CD70 in 28 samples. The antigen densities were also similar in 5 patients with simultaneous BCMA and CD70 measurements. CD70 is significantly upregulated in high risk multiple myeloma patients, suggesting that it may be a potential immunotherapeutic target for the treatment of MM, worth further preclinical and clinical studies. High recurrence rate is a difficulty and focus of multiple myeloma treatment, although BCMA expression levels are similar at different stages of multiple myeloma (from untreated to recurrent), and are effective therapeutic targets throughout the disease, patients receiving targeted BCMA CAR-T treatment are still at increased risk of recurrence as treatment time increases, although higher complete remission and objective response rates can be obtained. Because of tumor heterogeneity and antigen escape problems caused by BCMA shedding, CAR-T products targeting multiple tumor antigens can effectively reduce relapse caused by BCMA loss. CD70 has high expression ratio in high-risk multiple bone marrow tumor patients, and the expression level is equivalent to BCMA, and is taken as a target independent of BCMA, so that the prospect of combined use of the CD70 and the BCMA is interesting. In addition, the BAFF/APRIL/BCMA functional axis and the CD70/CD27 functional axis play an important role in lupus erythematosus, rheumatoid arthritis and other autoimmune diseases through different mechanisms of action. Thus, the targeted BCMA/CD70 CAR-T cell product can be used not only for treating multiple myeloma patients who have previously been relapsed after treatment with BCMA-related products, but also for reducing tumor recurrence due to single antigen loss in high risk multiple myeloma patients with BCMA positive expression. In addition, BCMA/CD70 CAR-T has the potential to treat autoimmune diseases, further increasing its market value. Chimeric antigen Receptor T cells (CHIMERIC ANTIGEN Receptor T cells, CAR-T) are a novel immunotherapeutic approach against tumor cell surface specific antigens. VHH antibodies, also known as nanobodies, are naturally deleted light chain antibodies found in alpaca peripheral blood, which comprise only one heavy chain variable region (VHH) and two conventional CH2 and CH3 regions, the smallest unit known to bind the antigen of interest. Compared with the traditional artificial modified scFv, the method has the advantages of small molecular weight, easy expression, strong specificity, high affinity, weak immunogenicity to human, short development period and the like. Combining the advantages of immunotherapy and VHH antibodies, a highly effective CAR-T therapy can be developed. There is currently no report on dual targeting chimeric antigen receptor specifically binding human BCMA and CD70 according to the present application. Disclosure of Invention The present invention provides a dual targeting chimeric antigen receptor targeting CD70 and BCMA comprising an anti-CD 70 VHH antibody, and an anti-BCMA antibody or antigen binding fragment thereof. In one or more embodiments, the anti-CD 70 VHH antibody comprises HCDR1-3 as shown in SEQ ID NO. 11-13, the anti-BCMA antibody or antigen binding fragment thereof comprises HCDR1-3 as shown in SEQ ID NO. 15-17 and LCDR1-3 as shown in SEQ ID NO. 18-20, preferably the anti-CD 70 VHH antibody has the amino acid sequence shown in SEQ ID NO.