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Methods of isolating t cell receptors having antigenic specificity for a cancer-specific mutation

A specific and antigenic technology, applied in the direction of cancer antigen components, genetically modified cells, receptors/cell surface antigens/cell surface determinants, etc., can solve the problems of difficult identification of TCR and patient isolation

Pending Publication Date: 2017-08-18
UNITED STATES OF AMERICA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For example, TCRs that specifically recognize cancer antigens can be difficult to identify and / or isolate from patients

Method used

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  • Methods of isolating t cell receptors having antigenic specificity for a cancer-specific mutation
  • Methods of isolating t cell receptors having antigenic specificity for a cancer-specific mutation
  • Methods of isolating t cell receptors having antigenic specificity for a cancer-specific mutation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0124] This example demonstrates a method for identifying one or more genes in the nucleic acid of a patient's cancer cells, each gene containing a cancer-specific mutation encoding a mutated amino acid sequence.

[0125]A 43-year-old woman (Patient (Pt.) 3737) with extensively metastatic cholangiocarcinoma who had undergone multiple chemotherapy regimens was enrolled in TIL-based adoptive cell therapy (ACT ) scheme. The clinical characteristics of patient 3737 are shown in Table 1.

[0126] Table 1

[0127]

[0128] *Acquisition sites for TIL generation and whole exome sequencing

[0129] + Functional status: ECOG, Eastern Cooperative Oncology Group

[0130] Lung metastases were excised and used as a source for whole-exome sequencing and generation of therapeutic T cells. Table 2 shows the somatic mutations identified by whole exome sequencing of metastatic pulmonary nodules from patient 3737. Tumor nodules were estimated to be approximately 70% of the tumor by pathol...

Embodiment 2

[0135] This example demonstrates the method of: inducing the patient's autologous APCs to present the mutated amino acid sequence; co-cultivating the patient's autologous T cell population with the autologous APCs presenting the mutated amino acid sequence; and selecting the autologous T cells that: (a ) is co-cultured with autologous APCs presenting the mutated amino acid sequence, and (b) is antigen specific for the mutated amino acid sequence presented in the context of MHC molecules expressed by the patient.

[0136] For each mutation identified in Example 1, a minigene construct was designed encoding the mutated amino acids flanked on each side by 12 amino acids from the endogenous protein. Multiple minigenes were synthesized in tandem to generate tandem minigene (TMG) constructs (Table 3). In Table 3, underlines indicate mutated amino acids and novel sequences encoded by point mutations or nucleotide insertions or deletions. For the splice site donor mutations (HLA-DOA ...

Embodiment 3

[0150] This example demonstrates that autologous open repertoire peripheral blood T cells genetically engineered with the TCR-Vβ22 chain (matched to its α chain) of ERBB2IP-specific CD4+ T cells identified in Example 2, are endowed with the ability to target the mutant Specific reactivity of ERBB2IP peptides.

[0151] The clonality of the mutant ERBB2IP-specific CD4+ T cells identified in Example 2 was characterized by sorting them after antigen-specific activation using OX40 as an activation marker. These cells were then bulk expanded and cloned by limiting dilution. Flow cytometry-based measurements of overall TCR-Vβ indicated that the overall expanded population was greater than 95% Vβ22+, and 10 / 11 clones assessed were pure Vβ22+. TCR sequence analysis revealed identical TCRβV-D-J sequences in 6 / 6 Vβ22+ clones tested (Table 5), suggesting that the majority of ERBB2IP-mutation reactive T cells contained a predominant Vβ22+ T cell clone.

[0152] table 5

[0153]

[01...

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Abstract

Disclosed are methods of isolating a TCR having antigenic specificity for a mutated amino acid sequence encoded by a cancer-specific mutation, the method comprising: identifying one or more genes in the nucleic acid of a cancer cell of a patient, each gene containing a cancer-specific mutation that encodes a mutated amino acid sequence; inducing autologous APCs of the patient to present the mutated amino acid sequence; co-culturing autologous T cells of the patient with the autologous APCs that present the mutated amino acid sequence; selecting the autologous T cells; and isolating a nucleotide sequence that encodes the TCR from the selected autologous T cells, wherein the TCR has antigenic specificity for the mutated amino acid sequence encoded by the cancer-specific mutation. Also disclosed are related methods of preparing a population of cells, populations of cells, TCRs, pharmaceutical compositions, and methods of treating or preventing cancer.

Description

[0001] Incorporation by reference into electronic submissions [0002] Incorporated herein by reference in its entirety is the computer-readable nucleotide / amino acid sequence listing filed concurrently with this application and identifying the following: 29,577 bytes ASCII (text) with filename "718291ST25.TXT" dated 2014 September 15th. Background technique [0003] Adoptive cell therapy (ACT) using cells genetically engineered to express anticancer antigen T-cell receptors (TCRs) can produce positive clinical responses in some cancer patients. However, obstacles to the successful application of TCR-engineered cells to broadly treat cancer and other diseases remain. For example, TCRs that specifically recognize cancer antigens can be difficult to identify and / or isolate from patients. Therefore, improved methods of obtaining cancer-responsive TCRs are needed. [0004] Summary of the invention [0005] One embodiment of the invention provides a method of isolating a TCR or...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/725C12N5/10A61K39/00A61P35/00
CPCA61K39/0011C07K14/7051C12N5/0636C12N5/0638A61K2039/5158A61K2039/5156C12N2510/00C12N2501/505A61P35/00A61P35/02C12N2501/998C12N2502/99C12Q1/6881C12Q2600/156C12Q2600/158G01N33/56977G01N33/6866G01N33/6869G01N2333/4725G01N2333/525G01N2333/535G01N2333/54G01N2333/5406G01N2333/5409G01N2333/5425G01N2333/5428G01N2333/55G01N2333/57G01N2333/70596
Inventor 埃里克·特兰卢勇成保罗·E·罗宾斯史蒂文·A·罗森伯格
Owner UNITED STATES OF AMERICA
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