T cells transduced with CRISPR replacement show a markedly stronger response to blood cancer lines than with standard transduction techniques. (A) Four-hour cytotoxicity of transduced CD8⁺ cells, as well as parental γδ20 T-cell clone, against an untreated (empty symbols) or zoledronate-pretreated (filled symbols) γδ20 donor-autologous B-LCL. Representative data are shown from 3 donors tested in 2 experiments carried out in duplicate. (B) TNFα secretion by transduced CD8⁺ (top) or CD4⁺ (bottom) T cells after overnight coincubation with a panel of established blood cancer lines of diverse lymphoid and myeloid origin, or patient-derived B-ALL cells. Cancer cells were preincubated with zoledronate for 24 hours before coincubation with T cells. TNFα secretion was normalized by subtracting TNFα produced by T cells alone, and by cancer cells alone. No specific TNFα secretion by T cells was observed in absence of zoledronate pretreatment. Representative data are shown from 3 donors and 2 experiments carried out in duplicate.