获得性耐药是限制索拉非尼在肝癌治疗中应用的主要原因。本研究旨在探讨一种新型长链非编码RNA(lncRNA)的作用和机制,lnc-TSI,肝癌的索拉非尼耐药。lnc-TSI与miR-4726-5p的相互作用,使用生物信息学工具预测miR-4726-5p和KCNMA1。lnc-TSI/miR-4726-5p/KCNMA1轴分子在临床样品和细胞系中的表达,以及索拉非尼耐药肝癌细胞系,使用qRT-PCR或蛋白质印迹测定。lnc-TSI的表达式,miR-4726-5p,通过质粒转染或慢病毒感染在HepG2和Huh7细胞中操作KCNMA1。CCK-8,流式细胞术,和Tunel测定法用于确定该轴在HCC索拉非尼耐药性中的作用。使用索拉非尼抗性HepG2和Huh7细胞建立异种移植模型,随后进行体内索拉非尼处理以确认体外发现。Lnc-TSI和KCNMA1在HCC临床样本和细胞系中表达显著下调,特别是在索拉非尼抗性中,而mi-4726-5p呈现相反的表达模式。Lnc-TSI与miR-4726-5p相互作用,Lnc-TSI通过在HCC细胞中形成miR-4726-5p作为ceRNA。lnc-TSI和KCNMA1的过表达促进索拉非尼处理的HCC细胞的凋亡和降低的细胞活力,从而减轻索拉非尼的耐药性。miR-4726-5p模拟物逆转KCNMA1介导的索拉非尼增敏作用,而lnc-TSI的额外过表达逆转了miR-4726-5p的作用。体内分析还显示,ln-TSI的过表达通过增加KCNMA1表达和降低miR-4726-5p表达来减少接种索拉非尼抗性HCC细胞的小鼠中的索拉非尼抗性。lnc-TSI/miR-4726-5p/KCNMA1轴在调节HCC对索拉非尼的耐药性中起关键作用,并可能作为临床治疗肝癌索拉非尼耐药的治疗靶点。
Acquired drug resistance is a main reason for limiting the application of sorafenib in HCC treatment. This study aimed to explore the role and mechanisms of a novel long non-coding RNA (lncRNA), lnc-TSI, in sorafenib resistance of HCC. The interaction between lnc-TSI and miR-4726-5p, and miR-4726-5p and
KCNMA1 were predicted using bioinformatic tools. Expression of the molecules in the lnc-TSI/miR-4726-5p/
KCNMA1 axis in clinical samples and cell lines, as well as the sorafenib resistant HCC cell lines, was determined using qRT-PCR or western blotting. Expressions of lnc-TSI, miR-4726-5p, and
KCNMA1 were manipulated in HepG2 and Huh7 cells through plasmid transfection or lentivirus infection. The CCK-8, flow cytometry, and Tunel assays were employed to determine the role of this axis on sorafenib resistance of HCC. A xenograft model was established using sorafenib-resistant HepG2 and Huh7 cells followed by in vivo sorafenib treatments to confirm the in vitro findings. Lnc-TSI and
KCNMA1 expressions were significantly downregulated in HCC clinical samples and cell lines, especially in sorafenib resistance ones, while mi-4726-5p presented a reversed expression pattern. Lnc-TSI interacted with miR-4726-5p, and Lnc-TSI acts as a ceRNA via sponging miR-4726-5p in HCC cells. Overexpression of lnc-TSI and
KCNMA1 promoted apoptosis and decreased cell viability of sorafenib-treated HCC cells, thus alleviated sorafenib resistance. miR-4726-5p mimic reversed the KCNMA1-mediated sorafenib sensitivity-promoting effect, while additional overexpression of lnc-TSI reversed the effect of miR-4726-5p. In vivo analysis also showed that overexpression of ln-TSI diminished sorafenib resistance in mice inoculated with sorafenib-resistant HCC cells via increasing KCNMA1 expression and decreasing miR-4726-5p expression. The lnc-TSI/miR-4726-5p/KCNMA1 axis plays a critical role in regulating the resistance of HCC to sorafenib, and might serve as a therapeutic target to manage sorafenib resistance of HCC in clinic.