NIFID

Abstract

<h3>Abstract</h3> Adoptive transfer of T cells expressing a transgenic T cell receptor (TCR) has the potential to revolutionize immunotherapy of infectious diseases and cancer. However, the generation of defined TCR-transgenic T cell medicinal products with predictable <i>in vivo</i> function still poses a major challenge and limits broader and more successful application of this ‘living drug’. Here, by studying 51 different TCRs, we show that conventional genetic engineering by viral transduction leads to variable TCR expression and T cell product functionality as a result of interference with the endogenous TCR, variable transgene copy numbers, and un-targeted transgene integration. In contrast, CRISPR/Cas9-mediated TCR gene editing enables defined, targeted TCR insertion with concomitant knock-out of the endogenous receptor. Thereby, T cell products display more homogenous and physiological TCR expression which results in increased functionality and – importantly – less variable <i>in vivo</i> T cell responses in comparison to conventionally generated T cells. Hence, targeted TCR gene editing increases the predictability of TCR-transgenic T cell product function, which represents a crucial aspect for clinical application in adoptive T cell immunotherapy. <h3>BRIEF SUMMARY</h3> Safer, more functional and predictable transgenic T cell products for immunotherapy can be generated via CRISPR/Cas9-mediated targeted TCR engineering