Identification of genetic vulnerabilities in brain tumors using CRISPR-Cas9 screening

Despite considerable gains in understanding the genetic architecture of brain tumours, little progress has been made in improving survival for patients with Glioblastoma (GBM) which remains a universally lethal cancer. Standard of care treatment for GBM includes chemotherapy with the oral alkylating agent temozolomide (TMZ), which increases survival by just 3 months on average with many patients deriving no benefit. To identify new therapeutic strategies, a comprehensive picture of genetic vulnerabilities and mechanisms of chemotherapeutic resistance across this heterogenous cancer is needed. To this end, we have performed series of genome-wide CRISPR-Cas9 fitness/essentiality screens in eight patient-derived GBM-stem cell (GSCs) lines with accompanying chemogenomic screens uncovering mechanisms of TMZ sensitivity and resistance. Our results uncover GBM-specific core and context-specific vulnerabilities underlying multiple processes such as the GSC state, cellular signaling and proteostasis. Small molecule inhibitors targeting several of these fitness genes lead to robust inhibition of GSC proliferation highlighting that the screens uncover new actionable therapeutic targets. Genome-wide chemogenomic TMZ resistance screens confirmed mutations in mismatch repair pathway genes as the principal source of acquired TMZ resistance. TMZ hypersensitivity screens with sublethal TMZ doses identified multiple loss of function mutations that increase TMZ sensitivity including many members of the Fanconi Anemia/Interstrand Crosslink Repair pathways, the MCM8-9 helicase complex and the previously uncharacterized stress-responsive RNA binding protein ZC3H7A. Our results comprise one of the largest series of CRISPR-Cas9 screens in solid tumour primary cell lines and unveil new strategies for treating this high fatality cancer.