Integrated genetic screening identifies FEN1 as a selective vulnerability and mediator of temozolomide resistance in glioblastoma stem cells

Glioblastoma stem cells (GSCs) are key drivers of tumor maintenance, therapeutic resistance, and relapse, but targeting them remains clinically elusive due to their overlap with normal neural stem cells (NSCs) and a lack of actionable vulnerabilities. To identify selective vulnerabilities in GSCs, we performed genome-wide CRISPR-Cas9 loss-of-function screening across patient-derived GSC models under standard-of-care treatment conditions.

We identified flap endonuclease 1 (FEN1), a key enzyme in DNA replication and base excision repair, as an essential gene for GSC survival, with enhanced dependency in the context of temozolomide (TMZ) treatment. Genetic knockdown of FEN1 impaired GSC proliferation and self-renewal and extended survival in a patient-derived xenograft model. Pharmacologic inhibition of FEN1 using a small-molecule inhibitor revealed a selective cytotoxic effect in highly aggressive and recurrent GBM models, while sparing NSCs.