Department of Urology, Zhuji People's Hospital, Zhuji, China.
Am J Mens Health. 2024 Sep-Oct;18(5):15579883241273305. doi: 10.1177/15579883241273305.
While cisplatin remains a frontline treatment for bladder cancer (BCa), the onset of resistance greatly hampers its effectiveness. RAC3 is closely linked to chemoresistance in cancer cells, but its specific role in cisplatin resistance within BCa is still elusive. RAC3 expression in BCa was analyzed using bioinformatics and quantitative polymerase chain reaction (qPCR). The gene set enrichment analysis (GSEA) identified RAC3-enriched pathways and the correlation between RAC3 and fatty acid synthase (FASN), a gene involved in fatty acid synthesis. Potential upstream transcription factors of RAC3 were predicted and their interaction with RAC3 was confirmed via dual-luciferase and chromatin immunoprecipitation (ChIP) assays. T24/DDP, a cisplatin-resistant BCa cell line, was established to probe into the regulatory role of RAC3 in cisplatin resistance. Cell proliferation was evaluated by colony formation and the IC values after cisplatin treatment were determined using cell counting kit-8 (CCK-8). The levels of free fatty acids and triglycerides (TGs), as well as the expression of DGAT2 and FASN proteins, were measured to gauge the extent of fatty acid synthesis in cells. Elevated expression of RAC3 was observed in BCa and the cisplatin-resistant BCa cells (T24/DDP). The knockdown of RAC3 within T24/DDP cells was demonstrated to counteract cisplatin resistance. Subsequent analyses identified RAC3 as being notably enriched in the fatty acid synthesis pathway, with Kruppel-like factor 1 (KLF1) emerging as a key upstream regulator. The overexpression of RAC3 was correlated with increased cisplatin resistance in T24/DDP cells, an effect that was mitigated by the addition of the FASN inhibitor, Orlistat. Furthermore, the downregulation of KLF1 suppressed RAC3 expression, disrupted fatty acid synthesis, and attenuated cisplatin resistance in T24/DDP cells. Conversely, the co-overexpression of RAC3 counteracted the effects conferred by KLF1 knockdown. Our study has validated that KLF1 activates RAC3 to mediate fatty acid synthesis and promote cisplatin resistance in BCa, suggesting the KLF1/RAC3 axis as a potential target for combating cisplatin-resistant BCa.
顺铂仍然是膀胱癌（BCa）的一线治疗药物，但耐药性的出现极大地影响了其疗效。RAC3 与癌细胞的化疗耐药性密切相关，但它在 BCa 中对顺铂耐药性的具体作用仍不清楚。使用生物信息学和定量聚合酶链反应（qPCR）分析 BCa 中的 RAC3 表达。基因集富集分析（GSEA）鉴定了 RAC3 富集的途径以及 RAC3 与参与脂肪酸合成的脂肪酸合酶（FASN）之间的相关性。预测了 RAC3 的潜在上游转录因子，并通过双荧光素酶和染色质免疫沉淀（ChIP）实验证实了它们与 RAC3 的相互作用。建立了顺铂耐药的 BCa 细胞系 T24/DDP，以探讨 RAC3 在顺铂耐药中的调节作用。通过集落形成评估细胞增殖，并用细胞计数试剂盒-8（CCK-8）测定顺铂处理后的 IC 值。测量细胞中游离脂肪酸和甘油三酯（TGs）的水平以及 DGAT2 和 FASN 蛋白的表达，以评估脂肪酸合成的程度。在 BCa 和顺铂耐药的 BCa 细胞（T24/DDP）中观察到 RAC3 的表达升高。在 T24/DDP 细胞中敲低 RAC3 被证明可以抵抗顺铂耐药性。随后的分析表明，RAC3 在脂肪酸合成途径中明显富集，而 Kruppel 样因子 1（KLF1）是关键的上游调节剂。RAC3 的过表达与 T24/DDP 细胞中顺铂耐药性的增加相关，而 FASN 抑制剂奥利司他的加入减轻了这种作用。此外，KLF1 的下调抑制了 RAC3 的表达，破坏了脂肪酸的合成，并减轻了 T24/DDP 细胞中的顺铂耐药性。相反，RAC3 的共过表达抵消了 KLF1 敲低的作用。我们的研究验证了 KLF1 通过激活 RAC3 介导脂肪酸合成并促进 BCa 中的顺铂耐药性，表明 KLF1/RAC3 轴可能是治疗顺铂耐药性 BCa 的潜在靶点。
