CD8 immunomagnetic selection and interleukin-15 (IL-15) facilitate the isolation of human CD8+ cytotoxic T-lymphocytes (CTL) genetically engineered to express an anti-CD20 chimeric T-cell receptor (cTCR)

2542 Background. Primary human CD8+ CTL can be genetically modified to express a CD20-specific cTCR. We electroporated human peripheral blood mononuclear cells (PBMC) with an anti-CD20 cTCR expressing plasmid and cultured them with interleukin-2 (IL-2). G418-resistant transfectants were cloned and screened by flow cytometry for surface cTCR expression. To optimize the yield of cTCR-expressing clones for a planned phase I clinical trial treating patients with relapsed follicular non-Hodgkin's lymphoma (FL), we compared the utility of CD8 pre-selection and IL-15 with our standard culture method. Methods. CD8+ PBMC were selected by CliniMACS™ anti-CD8 immunomagnetic beads (Miltenyi Biotec) at 99% purity, electroporated with the cTCR plasmid and cultured with IL-2 or IL-15. Clones were phenotyped by flow cytometry. Cytotoxicity was assessed in vitro by a 51Cr release assay and in SCID mice by delay of hind limb paralysis (HLP) induced by a CD20+ Burkitt's lymphoma cell line (Ramos). Results. All IL-2 and IL-1...