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TCR for identifying human papilloma virus HPV16-E7 antigen

A HPV16-E7, human papilloma virus technology, applied in the direction of virus, antiviral agent, virus/phage, etc., can solve problems such as little effect

Active Publication Date: 2019-10-22
深圳市因诺转化医学研究院 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although HPV vaccination can effectively prevent cancers caused by HPV infection, for cancer patients already infected with HPV, especially advanced cancer patients, conventional treatments such as surgery, radiotherapy, and chemotherapy have little effect on the remission of the patient's condition

Method used

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  • TCR for identifying human papilloma virus HPV16-E7 antigen
  • TCR for identifying human papilloma virus HPV16-E7 antigen
  • TCR for identifying human papilloma virus HPV16-E7 antigen

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Example 1: HLA-A2-HPV16-E7 11-19 Cloning and sequencing of specific T cells

[0047] Peripheral blood mononuclear cells (PBMC) derived from HLA-A2 genotype and HPV-positive cervical cancer patients were stimulated in vitro by chemically synthesized short peptide YMLDLQPET (Sangon Bioengineering (Shanghai) Co., Ltd.). After 2 rounds of polypeptide stimulation, polyclonal T cells (1×10 5 ) and 1×10 5 T2 cells loaded with YMLDLQPET short peptide (target cells) or non-target short peptides (control cells) were co-cultured overnight at 37°C. The next day, the release of cytokine IFN-r in the culture supernatant was detected. Positive polyclonal T cells (target cell OD 450 - control cell OD 450 >1.0) for limited dilution cloning, 14 days later, ELISA was used to detect the reactivity of T cells and target cells in each well after limited dilution. Pick positive monoclonal T cells for rapid expansion. After 14 days of rapid amplification, mouse anti-human CD8 antibody (...

Embodiment 2

[0069] Example 2: Preparation of HLA-A2-HPV16-E7 11-19 Specific TCR Gene Modified T Cells

[0070] The target TCR was cloned into the retroviral vector pMSGV1 (addgene) to construct the pMSGV1-G8 TCR vector. The virus packaging cell line 293GP cells were transfected with pMSGV1-G8 TCR and pVSV-G plasmids to prepare retrovirus and use the virus supernatant to transduce T cells.

[0071] The transfection operation was as follows: 293GP cells were seeded into 6-well plates (6×10 5 On the first day, pMSGV1-G8TCR and pVSV-G plasmids were transfected into 293GP cells (2 μg pMSGV1-G8TCR and 1.4 μg pVSV-G / well), and on the same day, anti-human CD3 antibody (OKT3) was used to activate the PBMC; on the third day, collect the culture medium containing the virus supernatant, and add fresh culture medium (DMEN containing 10% fetal bovine serum) to the 293GP cells; use the collected virus supernatant to centrifugally transfect activated T cells On the 4th day, adopt the same method to tr...

Embodiment 3

[0072] Example 3: HLA-A2-HPV16-E7 11-19 In vitro functional verification of specific TCR gene-modified T cells

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Abstract

The invention relates to the technical field of T cell antigen receptors, in particular to TCR capable of specifically identifying and combining with a human papilloma virus antigen HPV16-E7, a nucleotide sequence encoding the TCR or nucleic acid molecules of a complementary sequence of the nucleotide sequence, a carrier containing the nucleic acid molecules, cells transducing the nucleic acid molecules or carrier, a drug composition containing the TCR, the nucleic acid molecules, the carrier or the cells which are taken as active ingredients and application of the TCR, nucleic acid molecules,carrier, cells and drug composition which are used for preparing drugs for treating tumors or virus infection respectively.

Description

technical field [0001] The present invention relates to the technical field of T cell antigen receptors, in particular to a TCR capable of specifically recognizing and binding to human papillomavirus antigen HPV16-E7, and a nucleic acid molecule comprising a nucleotide sequence encoding the TCR or its complementary sequence, and A vector containing said nucleic acid molecule, and a cell transduced with said nucleic acid molecule or said vector, and a pharmaceutical composition comprising said TCR, nucleic acid molecule, vector or cell as an active ingredient, and said TCR, nucleic acid molecule, Uses of vectors, cells, pharmaceutical compositions. Background technique [0002] Human papillomavirus (HPV) is a small double-stranded DNA virus. According to the difference of pathogenicity of HPV, HPV can be divided into high-risk type and low-risk type. Studies have confirmed that the persistent infection of high-risk HPV (including HPV-16, 18, 31, 33 and 45) is the cause of v...

Claims

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Application Information

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IPC IPC(8): C07K14/725C12N15/867C12N15/12C12N5/10A61K35/17A61P35/00A61P31/20
CPCC07K14/7051C12N5/0636A61K35/17A61P35/00A61P31/20C12N2740/10043
Inventor 王明军陈磊董莲花
Owner 深圳市因诺转化医学研究院
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