Activation of APOBEC3B and Induced APOBEC3 Mutations in Self-Inactivating Lentiviral-Based CAR T Cell Production Decreases the Efficiency of CAR T Cell Production

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Presentation Time: 01:45 PM - 02:00 PM
Abstract ID: 4781 - B

Presenting Author:
Jack Swanson , Graduate Student at Mayo Clin. Grad. Sch. of Biomed. Sci., Mayo Clin., Minnesota

Abstract:

Apolipoprotein B mRNA editing enzyme, catalytic polypeptides (APOBECs) are a family of cytidine deaminase enzymes well described in anti-retroviral defense which convert cytidines to uracils (C-U), leading to C-T mutations in DNA. We investigated whether activation of APOBEC3B during the process of making chimeric antigen receptor (CAR) T cells via lentiviral vector affects the overall mutational integrity of resulting CAR T cells. To investigate this question, we quantified APOBEC3B expression and APOBEC signature mutations both post transfection in 293T viral producer cells and post transduction in human T cells. We observed abundant APOBEC3B activation and APOBEC3 signature mutations after transfection of plasmids to make lentivirus, but minimal APOBEC3B activation with a few additional APOBEC3 signature mutations after transduction. To test the functional relevance of APOBEC3B activity, we overexpressed APOBEC3B in 293T cells, and produced lentivirus from the 293T cells overexpressing APOBEC3B. The CAR T cells transduced with lentivirus from the 293T cells overexpressing APOBEC3B exhibited reduced killing ability compared to CAR T cells from 293T cells with no transgene. Taken together, these data suggest that APOBEC3B activation during the process of CAR T cell production induces mutations and reduces CAR T cell killing, and that selective ablation of APOBEC enzymes in GMP-grade lentiviral producer cells may enhance the overall quality of CAR T cells for clinical use.