Abstract
Transfer of tumour antigen-specific receptors to T cells requires efficient delivery and integration of transgenes, and currently most clinical studies are using gamma retroviral or lentiviral systems. Whilst important proof-of-principle data has been generated for both chimeric antigen receptors and αβ T cell receptors, the current platforms are costly, time-consuming and relatively inflexible. Alternative, more cost-effective, Sleeping Beauty transposon-based plasmid systems could offer a pathway to accelerated clinical testing of a more diverse repertoire of recombinant high affinity T cell receptors. Nucleofection of hyperactive SB100X transposase-mediated stable transposition of an optimised murine-human chimeric T cell receptor specific for Wilm’s tumour antigen from a Sleeping Beauty transposon plasmid. Whilst transfer efficiency was lower than that mediated by lentiviral transduction, cells could be readily enriched and expanded, and mediated effective target cells lysis in vitro and in vivo. Integration sites of transposed TCR genes in primary T cells were almost randomly distributed, contrasting the predilection of lentiviral vectors for transcriptionally active sites. The results support exploitation of the Sleeping Beauty plasmid based system as a flexible and adaptable platform for accelerated, early-phase assessment of T cell receptor gene therapies.
Highlights
Gamma retroviral-mediated transfer of recombinant receptors to redirect T cell immunity against tumour and viral antigens is being extensively investigated in early phase clinical trials
Isolated human peripheral blood lymphocytes (PBL) from healthy HLA-A2 donors were stimulated for 48 hours with anti-CD3/CD28 microbeads and human IL-2 exposed to a single round of lentivirus infection or nucleofected with transposon/transposase DNA plasmids
Flow cytometry following staining with anti-Vb2.1 antibody identified primary T cells expressing cell surface Wilms’ Tumour antigen 1 (WT1)-TCR and revealed comparable gene transfer
Summary
Gamma retroviral-mediated transfer of recombinant receptors to redirect T cell immunity against tumour and viral antigens is being extensively investigated in early phase clinical trials. HIV-1 derived lentiviral vector based T cell redirection with a CAR against CD19 was reported [6], and these vectors are thought to improve efficiency of gene transfer and yield more functional T cells after ex-vivo transduction. Both retroviral [7] and lentiviral [8] modification of T cells expressing recombinant T cell receptors against the Wilm’s tumour (WT1) antigen has been described and Phase 1 testing of an enhanced HLA-A2/WT1 peptide specific TCR has been initiated. Transduction efficiency, integration sites and redirection of human primary T cells against WT1 antigen was assessed and compared to lentiviral transduction procedures
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