Mentor Name: Ruby Sims

Cell based immunotherapies in the form of T Cells genetically modified to express chimeric antigen receptors (CAR) are providing additional treatment options for relapsed and/or refractory childhood cancers. A current hurdle to the wider implementation of these therapies is that the manufacturing of CAR T cells requires costly and viral vector-based ex vivo modification. Lipid nanoparticle (LNP) technologies have shown significant promise in the non-viral engineering of CAR T cells in vivo, yet their broader clinical translation for this application is currently limited by their nonspecific delivery. My role in this project focuses on designing and optimizing an LNP platform for the targeted engineering of human primary T Cells. Utilizing a LNP platform optimized to transfect primary human T cells by my mentor Dr Ruby Sims, we will utilize microfluidic synthesis of LNPs, incorporating an additional click chemistry enabled pegylated-lipid. The surface of the LNPs will then be decorated with aCD3 monoclonal antibodies utilizing a click chemistry conjugation strategy and size exclusion chromatography (SEC) enabled clean-up of unbound antibodies. Our preliminary results suggest that aCD3-mofidied LNPs and optimized, non-targeting LNPs show comparable transfection of activated, primary human T cells. However, the addition of CD3 targeting antibodies enables the transfection of resting T cells, not observed for LNPs absent of aCD3. This project will probe the mechanism by which these aCD3-modified LNPs enable resting T cell transfection, investigating the LNP mediated activation in an effort to increase the efficacy of this system and ensure that our LNP platform performs across different donors.