187 - Use of Elutriated Lymphocytes for Manufacturing CD19-Chimeric Antigen Receptor T Cells Improves the Quantity of Transduced T Cells in the Final Product

Abstract

Clinical trials of CD19-Chimeric Antigen Receptor (CAR) T cells manufactured from autologous peripheral blood mononuclear cell (PBMC) concentrates for the treatment of B cell lymphoma and leukemia have been promising. Yields of CD19-CAR T cells have been variable and we have shown that this variability is due in part to the contamination of the PBMCs used as starting cellular material with monocytes and granulocytes. We investigated the use of PBMC concentrates enriched for lymphocytes using counter-flow elutriation for CD19-CAR T cell manufacturing. Elutriation allows for the rapid, closed system separation of lymphocytes from monocytes and granulocytes. CD19-CAR T cells were manufactured from 26 patients using autologous PBMCs, anti-CD3/CD28 magnetic beads to promote T cell expansion, two days of retroviral transduction and 7 days of culture. For 6 products lymphocyte enrichment was performed using anti-CD3/CD28 beads, for 15 plastic adherence and anti-CD3/CD28 bead selection and for 5 elutriation. One product manufactured with anti-CD3/CD28 bead selected T cells did not contain enough cells for one clinical dose (1 × 106 transduced cells per kg). Among the other 25 products all were rich in T cells (>93%) and had high transduction efficiency (32% to 90%). The quantity of transduced T cells in the final product was greatest for those manufactured with elutriated lymphocytes (581 ± 212 × 106 cells/kg) and was the least for products manufactured with anti-CD3/CD28 bead selected cells (253 ± 227 × 106 cells/kg; p < 0.04). Products manufactured from plastic adherence and anti-CD3/CD28 selected cells had an intermediated quantity of transduced T cells (393 ± 176 × 106 cells/kg). In conclusion, enrichment of PBMCs for lymphocytes using elutriation increases the quantity of transduced cells produced. These results provide further evidence that CAR T cell expansion is inhibited by monocytes and granulocytes.