背景:癌症干细胞样细胞是各种癌症治疗耐药和转移的关键屏障,包括乳腺癌,然而,潜在的机制仍然难以捉摸。通过全基因组lncRNA表达谱,我们发现LINC00115在化疗耐药的乳腺癌干细胞样细胞(BCSC)中强烈上调.
方法:进行LncRNA微阵列分析以记录耐紫杉醇(PTX)的MDA-MB-231BCSC(ALDH)和非BCSC(ALDH-)中lncRNA的丰度变化。进行RNA下拉和RNA免疫沉淀(RIP)测定以确定LINC00115的结合蛋白。在TNBC转移性淋巴结组织中检查了LINC00115途径的临床意义。LINC00115的生物学功能通过功能增益和功能丧失研究进行了研究。通过RNA测序探索分子机制,质谱,和CRISPR/Cas9敲除系统。通过异种移植动物模型检查LINC00115的治疗潜力。
结果:LINC00115作为支架lncRNA连接SETDB1和PLK3,导致耐药BCSC中K106和K200处PLK3的SETDB1甲基化增强。PLK3甲基化降低HIF1α的PLK3磷酸化,从而增加HIF1α稳定性。HIF1α,反过来,上调ALKBH5以减少LINC00115的m6A修饰,导致YTHDF2依赖性m6A修饰的RNA的降解减弱,并增强LINC00115的稳定性。因此,在三阴性乳腺癌中,这种正反馈循环激发了BCSC表型,并增强了化疗耐药和转移.SETDB1抑制剂TTD-IN与LINC00115ASO在异种移植动物模型中使PTX抗性细胞对化疗的反应敏感。LINC00115、甲基化PLK3、SETDB1和HIF1α的相关表达是临床三阴性乳腺癌的预后。
结论:我们的发现揭示了LINC00115作为BCSC的关键调节因子,并强调了靶向LINC00115和SETDB1作为化疗耐药乳腺癌的潜在治疗策略。
Cancer stem-like cell is a key barrier for therapeutic resistance and metastasis in various cancers, including breast cancer, yet the underlying mechanisms are still elusive. Through a genome-wide lncRNA expression profiling, we identified that LINC00115 is robustly upregulated in chemoresistant breast cancer stem-like cells (BCSCs).
LncRNA microarray assay was performed to document abundance changes of lncRNAs in paclitaxel (PTX)-resistant MDA-MB-231 BCSC (ALDH+) and non-BCSC (ALDH-). RNA pull-down and RNA immunoprecipitation (RIP) assays were performed to determine the binding proteins of LINC00115. The clinical significance of the LINC00115 pathway was examined in TNBC metastatic lymph node tissues. The biological function of LINC00115 was investigated through gain- and loss-of-function studies. The molecular mechanism was explored through RNA sequencing, mass spectrometry, and the CRISPR/Cas9-knockout system. The therapeutic potential of LINC00115 was examined through xenograft animal models.
LINC00115 functions as a scaffold lncRNA to link SETDB1 and PLK3, leading to enhanced SETDB1 methylation of PLK3 at both K106 and K200 in drug-resistant BCSC. PLK3 methylation decreases PLK3 phosphorylation of HIF1α and thereby increases HIF1α stability. HIF1α, in turn, upregulates ALKBH5 to reduce m6A modification of LINC00115, resulting in attenuated degradation of YTHDF2-dependent m6A-modified RNA and enhanced LINC00115 stability. Thus, this positive feedback loop provokes BCSC phenotypes and enhances chemoresistance and metastasis in triple-negative breast cancer. SETDB1 inhibitor TTD-IN with LINC00115 ASO sensitizes PTX-resistant cell response to chemotherapy in a xenograft animal model. Correlative expression of LINC00115, methylation PLK3, SETDB1, and HIF1α are prognostic for clinical triple-negative breast cancers.
Our findings uncover LINC00115 as a critical regulator of BCSC and highlight targeting LINC00115 and SETDB1 as a potential therapeutic strategy for chemotherapeutic resistant breast cancer.