Background/Objectives: HNSCC is a highly aggressive malignancy marked by the dysregulation of the cell cycle. In HPV−HNSCC, mutations in the CDKN2A gene frequently result in the loss of the p16 protein, a key inhibitor of the cyclin D1/CDK4/6 complex. This loss results in unchecked G1/S phase progression. The CDK4/6 inhibitor palbociclib has shown therapeutic potential in HPV−HNSCC by inducing G1 phase arrest and reducing cell viability. In this study, we investigated the molecular mechanisms by which palbociclib affects cell viability in HPV−HNSCC. Methods: Four HPV−HNSCC cell lines were treated with palbociclib, and RNA sequencing was performed to assess changes in gene expression. Cell viability was measured using the MTT assay. To further investigate protein localization, interactions, and function, we used immunofluorescence staining, co-immunoprecipitation, small molecule inhibitors, and siRNA-mediated knockdown. Results: We demonstrate that palbociclib downregulates survivin, a protein that plays dual roles in mitosis and apoptosis, thereby inhibiting cell proliferation. We also found that survivin is overexpressed in HPV−HNSCC. Inhibiting survivin dimerization using the compound LQZ-7i significantly reduces cell viability and promotes its export from the nucleus to the cytoplasm. Additionally, we identified USP1, a deubiquitinase, as both a downstream target of CDK4/6 and a key regulator of survivin stability. Inhibiting USP1 activity or silencing its expression significantly reduces survivin levels. Conclusions: Our findings highlight survivin as a critical mediator of cell proliferation in HPV−HNSCC and suggest that targeting the CDK4/6-USP1-survivin axis may offer a promising therapeutic strategy.
背景/目的：头颈部鳞状细胞癌（HNSCC）是一种高度侵袭性的恶性肿瘤，其特征是细胞周期失调。在HPV阴性HNSCC中，CDKN2A基因的突变常导致p16蛋白缺失，而p16蛋白是细胞周期蛋白D1/CDK4/6复合物的关键抑制剂。这种缺失导致G1/S期进程失控。CDK4/6抑制剂帕博西尼通过诱导G1期阻滞和降低细胞活力，在HPV阴性HNSCC中显示出治疗潜力。本研究旨在探讨帕博西尼影响HPV阴性HNSCC细胞活力的分子机制。方法：用帕博西尼处理四种HPV阴性HNSCC细胞系，并通过RNA测序评估基因表达变化。采用MTT法检测细胞活力。为进一步研究蛋白质定位、相互作用和功能，我们使用了免疫荧光染色、免疫共沉淀、小分子抑制剂和siRNA介导的敲低技术。结果：我们发现帕博西尼能下调survivin蛋白的表达，该蛋白在有丝分裂和凋亡中发挥双重作用，从而抑制细胞增殖。我们还发现survivin在HPV阴性HNSCC中过表达。使用化合物LQZ-7i抑制survivin二聚化可显著降低细胞活力，并促进其从细胞核输出至细胞质。此外，我们发现去泛素化酶USP1既是CDK4/6的下游靶点，也是survivin稳定性的关键调节因子。抑制USP1活性或沉默其表达可显著降低survivin水平。结论：我们的研究结果强调了survivin是HPV阴性HNSCC细胞增殖的关键介质，并提示靶向CDK4/6-USP1-survivin轴可能是一种有前景的治疗策略。
肿瘤(癌症)患者之家