Hsa_circ_0089466调控GPR183致ERK通路激活对喉鳞癌干细胞干性维持及化疗抵抗的作用与机制研究

Chemotherapy resistance of laryngeal squamous cell carcinoma (LSCC) is a clinical bottleneck that impedes its comprehensive treatment to improve the prognosis level. CD133 and CD44 are the recognized surface markers of cancer stem cells, and the CD133 and CD44 dual-positive cancer stem cells shows significant resistance to chemotherapy, however, the upstream mechanism of this phenotype is unclear. The CD133+CD44+ double-positive LSCC cells were successfully enriched with injection of cisplatin into xenografts continuously for three generations. Whole transcriptomic sequencing revealed that GPR183 and circ_0089466 were co-upregulated in the CD133+CD44+ LSCC cells, and circ_0089466 regulated GPR183 mRNA and protein levels. Knockdown of GPR183 inhibits invasion and migration of LSCC cells. Moreover, GPR183 knockdown inhibits the expression of stem cell markers, while enhances the sensitivity to cisplatin in the CD133+CD44+ LSCC cells. It has been proved that MTA1 activates ERK pathway to regulate stemness and chemotherapy resistance of cancer cells. Our previous study demonstrated that GPR183 binds to MTA1 directly in LSCC cells. Therefore, this project intends to elucidate the underlying mechanisms of GPR183 regulates stemness and chemoresistance of LSCC stem cells via binding to MTA1 and activating the ERK signaling pathway. Furthermore, the mechanism of circ_0089466-mediated upregulation of GPR183 will be explored in this study. The potential values of circ_0089466/GPR183 for LSCC therapy will be evaluated through animal experiments and clinical samples analysis.Results of this project will provide new targets for improving sensitivity of LSCC chemotherapy.

化疗抵抗是阻碍喉鳞癌预后水平提升的重要临床瓶颈问题。CD133+CD44+双阳性肿瘤干细胞化疗抵抗能力显著，但上游调控机制不清。课题组连续对三代裸鼠移植瘤注射顺铂驯化并成功富集CD133+CD44+双阳性喉鳞癌干细胞。测序发现GPR183显著上调，与之共上调的circ_0089466可影响其转录与翻译水平。敲降GPR183显著抑制喉鳞癌细胞迁移侵袭能力，增强顺铂敏感性，同时CD133+CD44+双阳性喉鳞癌干细胞干性基因表达下调。文献报道MTA1可激活ERK通路调控肿瘤干细胞干性及化疗抵抗，而我们证实GPR183可与MTA1直接结合。本项目首先从体外实验解析circ_0089466表观调节GPR183对MTA1激活ERK通路调控喉鳞癌干细胞干性及化疗抵抗性的机制，其次通过动物实验和临床病例分析circ_0089466/GPR183的喉鳞癌临床治疗应用价值，为该病化疗增敏提供新的靶向策略。

恶性增殖、化疗抵抗是阻碍喉鳞癌预后水平提升的重要临床瓶颈问题。G蛋白偶联受体在细胞增殖、迁移、分化及稳态维持等发面发挥有重要生物学功能，参与多种肿瘤的发生发展。其中GPR183在喉鳞癌组织中高表达，不但调控喉鳞癌细胞的恶性增殖、迁移和侵袭能力，而且能够促进喉鳞癌细胞的干性及化疗抵抗。另外体内实验也表明敲降GPR183能够抑制喉鳞癌细胞的成瘤能力。同时本研究明确了circ_0089466在喉鳞癌中可能通过靶向GPR183调控喉鳞癌的发生发展。本研究还揭示miR-515-5p抑制喉鳞癌细胞的表型能力，并通过体内实验进一步揭示miR-515-5p对体内成瘤能力的影响。分析circ_0089466、miR-515-5p及GPR183等标记基因作为喉鳞癌分子分型标志物的潜能，可能为喉鳞癌的综合治疗方案提供有益的参考。