The project is divided into three major aims: First, a T cell line will be modified using genome editing in order to generate a novel “plug-and-(dis)play” cell line. This cell line will be engineered to allow for facile TCR reprogramming and for functional screening of candidate TCRs (Aim 1). Next, tumor-infiltrating T cells directly from human cancer patients will be isolated and single-cell sequenced to determine their TCR sequences (Aim 2). Candidate tumor-specific TCRs will then be integrated into the cell line developed in Aim 1 and screened at high-throughput to discover TCRs that are specific to tumor cells (Aim 3).
The development of high-throughput single-cell sorting and sequencing technologies now allow for the identification of individual tumor-reactive TCRs. Accordingly, a number of recent studies have exploited these technologies to demonstrate the feasibility of personalized TCR gene therapy. An important bottleneck in the TCR identification process is the use of primary T cells for screening and characterization purposes. This can become largely impractical, as either freshly isolated or thawed T cell samples are required for screening steps. Thus, the development of a robust TCR screening platform would be extremely beneficial for TCR gene therapy applications. More specifically, the ability to transfer TCR repertoires into a T cell line for facile screening would be highly desirable.
