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. 2021 Nov 16;9(11):e003213. doi: 10.1136/jitc-2021-003213

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© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY. Published by BMJ.

This is an open access article distributed in accordance with the Creative Commons Attribution 4.0 Unported (CC BY 4.0) license, which permits others to copy, redistribute, remix, transform and build upon this work for any purpose, provided the original work is properly cited, a link to the licence is given, and indication of whether changes were made. See https://creativecommons.org/licenses/by/4.0/.

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Clonal expansion in opT cells and identification of TCR. (A) Pie-charts showing representation of a given TCR beta-chain as a percentage of the all the independent TCRs detected. (B) Frequencies and CDR3 beta-chain sequences for the top five organoid-selected TCR (OSR) from each patient. (C) TCR enriched in Pt3 opT and organoid-depleted TCR (ODR1) used for generation of recombinant TCR. (D) A schematic of chimeric TCR that comprised of human Vα and Vβ chains and mouse constant α and β chains and co-expressed with human CD8 gene in SKW-3 cells. (E) Expression of T cell activation marker (CD69) in TCR-expressing SKW-3 cells exposed to autologous (Pt3) tumor organoids. N.S., not significant. **, p<0.01. P value calculated using two-tailed, unpaired t-test. (F) Expression of CD69 in OSR1 expressing SKW-3 cells exposed to allogenic (Pt10 and Pt38) tumor organoids. N.S., not significant. (G) CD8 + T cells from a donor PBMC were transduced with the chimeric TCR (OSR6) and flow sorted by mouse-TCR chimera (mTCR-FITC) expression. (H) OSR6 expressing CD8 + T cells were exposed to autologous (Pt10) or allogenic (Pt38) organoids and media analyzed for IFN-γ by ELISA. ***, p<0.001. P value calculated using two-tailed, unpaired t-test. (I) OSR6 expressing CD8 + T cells were exposed to autologous (Pt10) organoids with either Pan-MHC Class I antibody (W6/32) or Pan-MHC Class II antibody (Tu39) and media analyzed for IFN-γ by ELISA. N.S., not significant. **, p<0.01. P value calculated using two-tailed, unpaired t-test. IFN, interferon; opT, organoid-primed T; PBMCs, peripheral blood mononuclear cells; Pt, patient; TCR, T cell receptor.