American Association for Cancer Research
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Figure S4 from Rapid Construction of Antitumor T-cell Receptor Vectors from Frozen Tumors for Engineered T-cell Therapy

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posted on 2023-04-03, 22:41 authored by Takemasa Tsuji, Akira Yoneda, Junko Matsuzaki, Anthony Miliotto, Courtney Ryan, Richard C. Koya, Kunle Odunsi

Fig. S4 describes about the recovery of antigen-specific TCR from plasmid mixtures.




Roswell Park Alliance Foundation, Ovarian Cancer Research Fund Alliance

New York State Stem Cell Science



T cells genetically engineered with tumor antigen–specific T-cell receptor (TCR) genes have demonstrated therapeutic potential in patients with solid tumors. In order to achieve broader application, an efficient method to identify TCR genes for an array of tumor antigens and HLA restriction elements is required. Here, we have developed a method to construct a TCR-expression library from specimens, including frozen tumor biopsies, that contain antigen-specific T cells. TCR-expressing cassettes were constructed and cloned in a retroviral plasmid vector within 24 hours by unbiased PCR amplification of TCR α and β chain variable regions assembled with TCR constant regions. The method was validated by constructing TCR-expressing vectors from tumor antigen–specific T-cell clones and functionally assessing TCR gene–transduced T cells. We applied this method to frozen ovarian tumor specimens that were infiltrated by tumor antigen–specific T cells. The tumor-derived TCR libraries were expressed in peripheral T cells from healthy volunteers and screened for tumor antigen–specific TCR pairs with the use of an MHC/peptide tetramer reagent. Tumor antigen–specific TCR-expressing transgenes were recovered from isolated tetramer-positive T cells. Peripheral T cells that were engineered with library-derived TCR gene showed potent therapeutic antitumor effect in a tumor xenograft model. Our method can efficiently and rapidly provide tumor-specific TCR-expressing viral vectors for the manufacture of therapeutic and personalized antitumor T-cell products. Cancer Immunol Res; 6(5); 594–604. ©2018 AACR.

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