确定抗癌药物的作用机制是新药开发的重要一步。在这项研究中，我们建立了一个由 68 个癌基因（MANO panel）组成的综合筛选平台，涵盖 243 个遗传变异，以识别药物疗效的预测标记。使用针对 EGFR、BRAF 和 MAP2K1 的药物进行了验证，证实了该功能筛选组的实用性。对 BRCA2 敲除 DLD1 细胞系 (DLD1-KO) 的筛选显示，表达 SMO 和 GLI1 的细胞对奥拉帕尼具有耐药性。基因集富集分析确定了与 DNA 损伤修复相关的基因，这些基因在过度表达 SMO 和 GLI1 的细胞中富集。 GLI1 表达显着上调与同源重组修复 (HR) 相关的基因（例如 FANC 家族和 BRCA1/2）的表达，这表明 PARP 抑制剂耐药。尽管并非所有核苷酸切除修复 (NER) 途径的代表性基因均上调，但 GLI1 增强了 NER 活性。 GLI1抑制剂在体外和体内均能有效对抗过度表达GLI1的DLD1-KO细胞。此外，奥拉帕尼和GLI1抑制剂的联合治疗对DLD1-KO表现出协同作用，表明GLI1抑制剂针对DNA损伤修复缺陷的癌症的临床应用可能。该平台能够识别与药物敏感性相关的生物标志物，是药物开发的有用工具。© 2024。作者。

Identifying the mechanisms of action of anticancer drugs is an important step in the development of new drugs. In this study, we established a comprehensive screening platform consisting of 68 oncogenes (MANO panel), encompassing 243 genetic variants, to identify predictive markers for drug efficacy. Validation was performed using drugs that targeted EGFR, BRAF, and MAP2K1, which confirmed the utility of this functional screening panel. Screening of a BRCA2-knockout DLD1 cell line (DLD1-KO) revealed that cells expressing SMO and GLI1 were resistant to olaparib. Gene set enrichment analysis identified genes associated with DNA damage repair that were enriched in cells overexpressing SMO and GLI1. The expression of genes associated with homologous recombination repair (HR), such as the FANC family and BRCA1/2, was significantly upregulated by GLI1 expression, which is indicative of PARP inhibitor resistance. Although not all representative genes of the nucleotide excision repair (NER) pathway were upregulated, NER activity was enhanced by GLI1. The GLI1 inhibitor was effective against DLD1-KO cells overexpressing GLI1 both in vitro and in vivo. Furthermore, the combination therapy of olaparib and GLI1 inhibitor exhibited a synergistic effect on DLD1-KO, suggesting the possible clinical application of GLI1 inhibitor targeting cancer with defective DNA damage repair. This platform enables the identification of biomarkers associated with drug sensitivity, and is a useful tool for drug development.© 2024. The Author(s).