Longitudinal analysis of lentiviral and retroviral chimeric antigen receptors' integration sites reveals distinct clonal evolutionary patterns.

Abstract

T-cell malignancies following chimeric antigen receptor (CAR) therapies are partly related to insertional mutagenesis, but the longitudinal evolution of CAR-integration sites (IS) remains understudied. We performed an IS analysis in blood from three tisagenlecleucel (lentiviral), one axicabtagene-ciloleucel and one brexucabtagene-autoleucel (gammaretrovirals) patient at peak expansion and 1-year follow-up. All were complete responders. Peak expansion IS patterns were vector dependent: lentiviral CAR integrated mostly in introns and gammaretrovirals in intergenic regions, closer to transcription start sites. At 1-year post-infusion, lentiviral CAR showed no major clonal proliferation. Gammaretroviral CARs had divergent outcomes: no detectable CAR (axicabtagene-ciloleucel) or low-level oligoclonal persistence (brexucabtagene-autoleucel). Whether this latter evolution is at risk of further CAR malignancies needs further investigations.