PARP inhibitor counteracts Temozolomide Resistance in Glioblastoma Multiforme
BackgroundGlioblastoma multiforme (GBM) is the most common malignant primary brain tumour in adults and is invariably associated with poor prognosis. Resistance to Temozolomide (TMZ), the standard chemotherapeutic agent, remains a major clinical challenge, particularly due to DNA mismatch repair (MMR) deficiencies. The aim of this study was to determine whether combining TMZ with the poly(ADP-ribose) polymerase inhibitor Olaparib (OLA) could overcome TMZ resistance in GBM. MethodsWe conducted in vitro experiments using U251 cell-line, including a TMZ-resistant derivative, and primary GBM cultures derived from patient tumours. A CRISPR/Cas9 knockout screen was employed to identify genes involved in TMZ resistance. Cell viability, proliferation, and morphology were assessed following treatment with TMZ, OLA, or their combination. ResultsThe CRISPR screen identified inactivation of MMR pathway genes as key mediators of TMZ resistance. Co-treatment with OLA and TMZ demonstrated synergistic cytotoxicity in both parental and TMZ-resistant U251 cells, as well as in primary GBM cultures at diagnosis or relapse. Notably, OLA restored sensitivity to TMZ in MMR-deficient contexts and in tumours expressing O6-methylguanine-DNA-methyltransferase (MGMT). The combination treatment induced persistent DNA damage, cell cycle disruption, and cell death. ConclusionsThese findings provide strong preclinical evidence that combining TMZ with OLA can effectively overcome key mechanisms of TMZ resistance in GBM. This approach offers a promising therapeutic strategy warranting further clinical investigation. IMPORTANCE OF THE STUDYTemozolomide (TMZ) resistance remains a major therapeutic obstacle in glioblastoma (GBM), often driven by MMR deficiency or MGMT expression. While poly(ADP-ribose) polymerase (PARP) inhibitors have shown potential in other cancers, their role in overcoming TMZ resistance in GBM has remained unclear. In this study, a CRISPR screen identified MMR deficiency as a key driver of TMZ resistance. We further demonstrate that co-treatment with the PARP inhibitor Olaparib (OLA) restores TMZ sensitivity in both MMR-deficient and MGMT-expressing GBM cells and patient-derived cultures. These findings provide strong preclinical evidence supporting PARP inhibition as a promising therapeutic strategy to overcome chemoresistance in GBM and justify further clinical investigation. KEY POINTSO_LIPARP inhibitor Olaparib restores temozolomide sensitivity in resistant GBM cells C_LIO_LICombination therapy overcomes resistance driven by MMR deficiency or MGMT expression C_LIO_LIDual treatment induces persistent DNA damage and apoptosis in glioblastoma primary cultures C_LI