Thioridazine reverses trastuzumab resistance in gastric cancer by inhibiting S-phase kinase associated protein 2-mediated aerobic glycolysis

BACKGROUND Trastuzumab constitutes the fundamental component of initial therapy for patients with advanced human epidermal growth factor receptor 2(HER-2)-positive gastric cancer(GC).However,the efficacy of this treatment is hindered by substantial challenges associated with both primary and acquired drug resistance.While S-phase kinase associated protein 2(Skp2)overexpression has been implicated in the malignant progression of GC,its role in regulating trastuzumab resistance in this context remains uncertain.Despite the numerous studies investigating Skp2 inhibitors among small molecule compounds and natural products,there has been a lack of successful commercialization of drugs specifically targeting Skp2.AIM To discover a Skp2 blocker among currently available medications and develop a therapeutic strategy for HER2-positive GC patients who have experienced progression following trastuzumab-based treatment.METHODS Skp2 exogenous overexpression plasmids and small interfering RNA vectors were utilized to investigate the correlation between Skp2 expression and trastuzumab resistance in GC cells.Q-PCR,western blot,and immunohistochemical analyses were conducted to evaluate the regulatory effect of thioridazine on Skp2 expression.A cell counting kit-8 assay,flow cytometry,a amplex red glucose/glucose oxidase assay kit,and a lactate assay kit were utilized to measure the proliferation,apoptosis,and glycolytic activity of GC cells in vitro.A xenograft model established with human GC in nude mice was used to assess thioridazine's effectiveness in vivo.RESULTS The expression of Skp2 exhibited a negative correlation with the sensitivity of HER2-positive GC cells to trastuzumab.Thioridazine demonstrated the ability to directly bind to Skp2,resulting in a reduction in Skp2 expression at both the transcriptional and translational levels.Moreover,thioridazine effectively inhibited cell proliferation,exhibited antiapoptotic properties,and decreased the glucose uptake rate and lactate production by suppressing Skp2/protein kinase B/mammalian target of rapamycin/glucose transporter type 1 signaling pathways.The combination of thioridazine with either trastuzumab or lapatinib exhibited a more pronounced anticancer effect in vivo,surpassing the efficacy of either monotherapy.CONCLUSION Thioridazine demonstrates promising outcomes in preclinical GC models and offers a novel therapeutic approach for addressing trastuzumab resistance,particularly when used in conjunction with lapatinib.This compound has potential benefits for patients with Skp2-proficient tumors.

曲古菌素A对血管平滑肌细胞p27kip1表达的调控机制研究

目的探讨组蛋白去乙酰化酶抑制剂曲古菌素A(Trichostatin A,TSA)对血管平滑肌细胞(VSMC)的p27kip1表达的影响和调控机制。方法半定量逆转录聚合酶链反应(RT-PCR)检测p27kip1的mRNA水平,蛋白印迹测定p27kip1和S-phase kinase-associated protein-2(skp2)蛋白表达,荧光分光光度法测定20S蛋白酶体活性。结果100 ng/ml TSA不影响VSMC中p27kip1的mRNA水平。100 ng/mlTSA显著抑制血清诱导的p27kip1蛋白下调,并延长p27kip1蛋白的半衰期。100 ng/ mlTSA抑制血清诱导的skp2表达上调,且skp2表达与相应时点p27kip1蛋白呈负相关。100 ng/ml TSA对20S蛋白酶体活性物影响。结论TSA对VSMC的p27kip1表达调控不是在转录水平上,而是通过翻译后机制抑制血清诱导VSMC的p27kip1蛋白降解,其机制可能与TSA抑制泛素连接酶亚单位skp2表达有关。