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Abstract
Introduction/Background Cervical cancer is a prevalent gynecological malignancy in China, with limited treatment options for advanced stages. Despite platinum-based chemotherapy, patients with advanced cervical cancer face a median survival of only 16.8 months, emphasizing the need for alternative therapies. PIK3CA mutations are frequently observed in cervical cancer, leading to the constitutive activation of the PI3K-AKT pathway. Although PI3K inhibitors, like alpelisib (BYL719), show promise, challenges persist, including efficacy prediction and limited single-drug effectiveness.
Methodology Utilizing CRISPR/Cas9 technology, we employed a negative selection strategy to identify efficacy biomarkers for PI3Kα inhibitors in cervical cancer. We knocked down KMT2D and established stable cell lines, assessing responses to BYL719 through CCK8 assays, IC50 calculations, apoptosis, and cell cycle analyses. To elucidate the mechanism, CUT&Tag combined with RNA-seq analysis was performed, exploring the impact of KMT2D on ITGB6 expression and downstream FAK Y397 phosphorylation. In vivo verification involved subcutaneous tumorigenic administration experiments in nude mice with KMT2D knockdown cell lines.
Results KMT2D silencing increased BYL719 sensitivity, apoptosis, and cell cycle arrest in vitro. Overexpression of KMT2D conferred resistance. Mechanistically, KMT2D regulated ITGB6 expression, impacting FAK Y397 phosphorylation and the PI3K/AKT pathway. In vivo, KMT2D knockdown enhanced BYL719 efficacy. To address resistance, we induced PI3Kα inhibitor-resistant cell lines, revealing upregulated phosphorylation-modification-related proteins enriched in the Focal adhesion pathway. Combining BYL719 with the FAK inhibitor Defactinib reversed resistance.
Conclusion Genome-wide CRISPR/Cas9 screening identified KMT2D as an efficacy biomarker for BYL719 in cervical cancer. Silencing KMT2D enhanced BYL719 sensitivity by regulating ITGB6 and FAK Y397 phosphorylation. Additionally, the combination of BYL719 and Defactinib reversed PI3Kα inhibitor resistance. This study provides insights for clinical trials, emphasizing the potential of KMT2D as a predictive marker for PI3Kα inhibitors in cervical cancer treatment.
Disclosures There are no financial conflicts of interest to disclose.
