226 ‘Off-the-shelf’ CD19-CAR-T cells generated from umbilical cord blood display superior anti-tumor fitness and lower pro-inflammatory activity as compared to peripheral blood-derived engineered T cells

BackgroundThe principal goal of this study is to generate ’off-the-shelf’ CD19-CAR-T cells utilizing umbilical cord blood (UCB) as source of T lymphocytes and characterize through multi-omics their phenotype and functional properties.MethodsT cells were isolated from UBCs (N=15) and, upon activation in vitro using CD3 and CD28 mAbs, transduced with lentiviral vectors encoding for CD19-CD28z-or CD19–4-1BBz-CARs. Engineered T cells were also produced from the peripheral blood lymphocytes (PBL; N=5) as reference. The multidimensional phenotype analysis was utilized to assess the differentiation and activation status of the T cells. CD19-CAR-T cells were co-incubated or not with either CD19+ or CD19- target cells to mimic the antigen-mediated engagement of the CARs, and then, multi-omics analyses, including metabolomics, transcriptomics, and in vitro functional assays, (Elispot, Luminex) were performed.ResultsCD19-CAR T cells with early stage of differentiation (CD45RA+) co-expressing either ICOS or BTLA were observed in UCB- vs. PBL-CAR T cells (p