Extended Data Fig. 7: Potential resistant mechanisms of death receptor activation mediated senolysis.
From: cFLIP suppression and DR5 activation sensitize senescent cancer cells to senolysis
a, CRISPR-based resistance screen was performed on cFLIP null SK-Hep1 cells to identify genes whose inhibition can result in resistance to alisertib mediated senolysis. The cFLIP null SK-Hep1 cells were infected with genome-wide Brunello gRNA collection lentivirus and cultured with 0.5 µM alisertib and 1 µg/ml doxycycline for 14 days. The control arm is without alisertib. Deep-sequencing was used to determine changes in library representation. b, Top hits were selected by the fold enrichment upon senolysis (N = 3 independently viral infected cell cultures). c, Real-time PCR of Bid in cells A549 and SK-Hep1 with shRNA against Bid. d, Cell viability accessed by colony formation assay on Bid knockdown senescent A549 and SK-Hep1 cells treated with conatumumab. e, Real-time PCR of Caspase 8 in cells A549 with shRNA against Caspase 8. f, Cell viability on Caspase 8 knockdown senescent A549 cells treated with conatumumab. g, Cell viability on FADD suppressed senescent A549 cells treated with Lz-TRAIL. h, Real-time PCR of FADD in cells A549 with shRNA against FADD. i, Dose-response curve in 1 µM alisertib-induced senescent PLC/PRF/5 cells treated with conatumumab, the result represents at least 3 times with similar results. j, Real-time PCR of FADD in cells A549 infected with FADD overexpression vector. k, Cell viability on FADD overexpressed senescent A549 cells treated with Lz-TRAIL. Error bars in this figure panel c-h, j, k represent as mean ± standard deviations, N = 3 independent experiments, statistical significance was calculated by two-tailed t-test.
