Abstract 3111: PI3K-dependent GAB1/Erk phosphorylation renders head and neck squamous cell carcinoma sensitive to PI3Kα inhibitors

Head and neck cancer is the seventh most common cancer in the world, of which about 90% are head and neck squamous cell carcinoma (HNSCC). Frequent alterations of PIK3CA in HNSCC suggest that PI3Kα is a promising target for HNSCC treatment. CYH33 is a novel PI3Kα-selective inhibitor discovered by our group, which is currently undergoing a phase I clinical trial (NCT03544905) for the treatment of advanced solid tumors, including HNSCC. However, there is an urgent need to elucidate its mechanism of action and improve its efficacy against HNSCC. In this study, we found CYH33 displayed promising but variable therapeutic activity against HNSCC. Inhibition of PI3K/Akt pathway by CYH33 was not sufficient for its activity against HNSCC. Tandem-Mass-Tag (TMT) proteomics and phosphoproteomics were performed to reveal comprehensive regulation of kinome by CYH33. Among them, phosphorylation of MAPK1/Erk at activation sites (T185/Y187) were significantly attenuated by CYH33 in sensitive HNSCC cells. CYH33 significantly suppressed phosphorylation of Erk in sensitive but not resistant cells. Further inhibition of Erk phosphorylation enhanced the activity of CYH33 against HNSCC. Signaling network of differentially phosphorylated proteins illustrated that phosphorylation at multiple sites (Y659, Y406, S651, S648) of the adaptor protein GAB1, which integrates the signaling from PI3K and receptor tyrosine kinases, were significantly downregulated upon CYH33 treatment. CYH33 dose-dependently suppressed the phosphorylation of GAB1 at Y659 in sensitive HNSCC cells, whereas it had little effect in resistant cells. We found that CYH33 attenuated the membrane localization and phosphorylation of GAB1, resulting in reduced Erk phosphorylation and ultimately inhibiting the proliferation of sensitive cells. Meanwhile, activation of EGFR induced GAB1 phosphorylation independent of PI3K in HNSCC. Concurrent inhibition of EGFR synergistically potentiated the activity of CYH33 against HNSCC.In conclusion, we found that PI3Kα-selective inhibitor CYH33 displayed potent activity against HNSCC, which was associated with inhibition of Akt signaling as well as attenuation of PI3K-dependent GAB1/Erk phosphorylation. Simultaneous inhibition of GAB1 phosphorylation independent of PI3K potentiated the anti-HNSCC activity of CYH33. These findings revealed the insight mechanism of CYH33 against HNSCC and provided rational combination regimen for HNSCC. Citation Format: Xu Zhang, Jiao Xu, Xuan Wang, Lan Xu, Xi Zhang, Yi Wang, Shujuan Jiang, Yixiang Zhang, Jian Ding, Chen Qing, Linghua Meng. PI3K-dependent GAB1/Erk phosphorylation renders head and neck squamous cell carcinoma sensitive to PI3Kα inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3111.