Verhees Femke, Demers Imke, Legemaate Dion, Jacobs Robin, Hoeben Ann, Kremer Bernd, Speel Ernst-Jan
Department of Otorhinolaryngology, Head and Neck Surgery, GROW Research Institute for Oncology and Reproduction, Maastricht University Medical Center, 6229HX Maastricht, The Netherlands.
Department of Pathology, GROW Research Institute for Oncology and Reproduction, Maastricht University Medical Center, 6229HX Maastricht, The Netherlands.
Int J Oncol. 2025 Feb;66(2). doi: 10.3892/ijo.2025.5719. Epub 2025 Jan 10.
Human papillomavirus (HPV)‑positive and -negative head and neck squamous cell carcinoma (HNSCC) are often associated with activation of the phosphatidylinositol 3‑kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway due to mutations or amplifications in , loss of or activation of receptor tyrosine kinases. In HPV‑negative tumors, (encoding p16 protein) inactivation or (encoding Cyclin D1 protein) amplification frequently results in sustained cyclin‑dependent kinase (CDK) 4/6 activation. The present study aimed to investigate the efficacy of the CDK4/6 inhibitors (CDKi) palbociclib and ribociclib, and the PI3K/Akt/mTOR pathway inhibitors (PI3Ki) gedatolisib, buparlisib and alpelisib, in suppressing cell viability of HPV‑positive and ‑negative HNSCC cell lines. Inhibitor efficacy was assessed using MTT assay and western blotting analysis. Cell cycle analysis was performed using flow cytometry and apoptosis was assessed using annexin V staining. Metabolic changes in terms of glycolysis and oxidative metabolism were measured by Seahorse XF96 extracellular Flux analysis. The results of the present study showed that both HPV‑positive and ‑negative HNSCC cell lines were sensitive to PI3Ki. In general, PI3Ki decreased PI3K/Akt/mTOR pathway activity, resulting in apoptosis, and decreased oxidative and glycolytic metabolism. The CDKi were particularly effective in blocking HPV‑negative cell line viability, showing decreased retinoblastoma expression and G1‑phase cell cycle arrest, whereas apoptosis was not induced. Thus, PI3Ki and CDKi efficiently inhibited their respective pathways and HNSCC cell viability , with the latter occurring only in HPV‑negative cell lines. Whereas PI3Ki induced apoptosis and attenuated cellular metabolism, CDKi led to cell cycle arrest. Further research should be performed to elucidate whether (a combination of) these inhibitors may be effective therapeutic agents for patients with HNSCC.
人乳头瘤病毒（HPV）阳性和阴性头颈部鳞状细胞癌（HNSCC）常因PI3K基因的突变或扩增、PTEN缺失或受体酪氨酸激酶激活而与磷脂酰肌醇3激酶（PI3K）/Akt/雷帕霉素哺乳动物靶蛋白（mTOR）信号通路的激活相关。在HPV阴性肿瘤中，CDKN2A（编码p16蛋白）失活或CCND1（编码细胞周期蛋白D1）扩增常导致细胞周期蛋白依赖性激酶（CDK）4/6持续激活。本研究旨在探讨CDK4/6抑制剂帕博西尼和瑞博西尼，以及PI3K/Akt/mTOR信号通路抑制剂吉地替尼、布帕利西布和阿培利西布对抑制HPV阳性和阴性HNSCC细胞系细胞活力的疗效。使用MTT法和蛋白质免疫印迹分析评估抑制剂疗效。采用流式细胞术进行细胞周期分析，使用膜联蛋白V染色评估细胞凋亡。通过Seahorse XF96细胞外通量分析测量糖酵解和氧化代谢方面的代谢变化。本研究结果表明，HPV阳性和阴性HNSCC细胞系均对PI3K抑制剂敏感。一般来说，PI3K抑制剂降低PI3K/Akt/mTOR信号通路活性，导致细胞凋亡，并降低氧化代谢和糖酵解代谢。CDK4/6抑制剂在阻断HPV阴性细胞系活力方面特别有效，表现为视网膜母细胞瘤蛋白表达降低和G1期细胞周期阻滞，而未诱导细胞凋亡。因此，PI3K抑制剂和CDK4/6抑制剂可有效抑制各自的信号通路和HNSCC细胞活力，后者仅发生在HPV阴性细胞系中。PI3K抑制剂诱导细胞凋亡并减弱细胞代谢，而CDK4/6抑制剂导致细胞周期阻滞。应进一步开展研究以阐明这些抑制剂（或其组合）是否可能成为HNSCC患者的有效治疗药物。
