Artificial T cell adapter molecule-transduced TCR-T cells demonstrated improved proliferation only when transduced in a higher intensity

Abstract An artificial T cell adapter molecule (ATAM) was generated to improve persistence of T cell receptor (TCR) gene-transduced T cell (TCR-T) in a preceding study. ATAMs are gene-modified CD3ζ with the intracellular domain of 4-1BB inserted in the middle of CD3ζ. NY-ESO-1 TCR-T cells transduced with ATAM with two separated virus vector demonstrated superior proliferation upon antigen stimulation. To further develop clinically applicable ATAM-transduced TCR-T cells, we attempted to make single virus-vector to transduce TCR and ATAM simultaneously. Because we failed to observe improved proliferation capacity upon stimulation after 1 virus-vector (1vv) transduction, we compared TCR-T cells transduced with 1vv and 2 virus-vector (2vv) methods to elucidate the reason. In Jurkat-reporter cells, ATAM transduced by 2vv method demonstrated higher intensity than 1vv method, and the ATAM intensity was associated with the increased NF-κB signals upon stimulation. In ATAM-transduced primary T cells, transduced ATAM by 2vv method showed higher intensity and better proliferation. ATAM-transduced TCR-T cells demonstrated improved proliferation only when ATAM was transduced in a higher intensity. To create the simpler transduction method, we need to develop a strategy to make a higher ATAM expression to prove the efficacy of ATAM-transduction in TCR-T therapy.