Building and Optimizing a Virus-specific T Cell Receptor Library for Targeted Immunotherapy in Viral Infections

Banu, Nasirah; Chia, Adeline; Ho, Zi Zong; Garcia, Alfonso Tan; Paravasivam, Komathi; Grotenbreg, Gijsbert M.; Bertoletti, Antonio; Gehring, Adam J.

Our goal was to harness the potential of TCR gene transfer and develop a virus-specific TCR library, optimized and ready for clinical use. We expanded virus-specific T cells from healthy and resolved donors and cloned 10 virus-specific TCRs to 5 different viruses restricted to 4 HLA class-I molecules commonly found in the general population. To establish a standardized TCR gene cassette we employed previously published methods to optimize TCR expression with minimal modification to the wild type amino acid sequence [13] [14] [15] [16] [17] . Our library of 10 TCRs ideally positioned us to probe the specific effects of simple modifications, such as inverting the orientation of the TCR alpha and beta genes in the expression cassette, which led to a significant increase in TCR expression and cytokine production. We also found that the function (IFN-c production) of engineered T cells could be further augmented with the addition of Toll-like receptor (TLR) ligands to the culture during the transduction procedure; increasing the frequency of IFN-c producing cells up to 70%. The core library of virus-specific TCRs presented here, each one optimized for expression in primary human T cells, could provide a steppingstone to effective treatments for viral infections.