lnc-RI正调控LIG4表达增强结直肠癌放射抵抗性及机制研究

Colorectal cancer preoperative radiotherapy is a routine treatment of advanced colorectal cancer. Research on colorectal cancer radiation resistance prediction and intervention is an important. Lnc-RI is a long non-coding RNA induced by radiation, which was previously identified by the applicant. Previous studies have shown that lnc-RI may regulate the NHEJ repair process of DNA double-strand breaks by modulating the expression of LIG4. Further study showed that lnc-RI showed high expression in some colorectal cancer specimens and found that colorectal cancer cell line HCT-116 lnc-RI high expression levels had higher radiation resistance than lnc-RI low expression cells. It is hypothesized that the high expression of lnc-RI enhances its radiation resistance by positively regulating LIG4 and promoting NHEJ repair of DSBs in postoperative colorectal cancer cells. Three studies were conducted: 1) confirmed that lnc-RI enhanced the radiosensitivity of colorectal cancer cells by promoting DSB NHEJ repair; 2) clarified the role of LIG4 in lnc-RI regulation of radiation resistance and lnc-RI regulation of LIG4； 3) combined with clinical tumor case analysis the correlation between lnc-RI expression levels and colorectal cancer radiation resistance.

术前放射治疗是局限性晚期结直肠癌的常规治疗手段，结直肠癌放射抵抗性研究具有重要的临床价值。LncRNA作为一类新的功能分子，代表新的方向领域。我们前期研究发现敲低lnc-RI（本实验室筛选获得的放射诱导表达lncRNA）水平，抑制LIG4（NHEJ修复关键分子）表达的同时增加DNA双链断裂，降低细胞的放射抵抗性。进一步研究发现，lnc-RI在部分结直肠癌标本呈现高表达。结合DSB修复水平是影响细胞放射抗性关键因素的文献报道，提出如下科学设想： lnc-RI通过正调控LIG4，促进结直肠癌细胞DSBs的NHEJ修复，增强其放射抵抗性。拟开展三个方面研究：1）证实lnc-RI通过促进DSB的NHEJ修复增强结直肠癌细胞放射抵抗性；2）明确LIG4在lnc-RI调节放射抵抗性中的作用及lnc-RI调节LIG4的机制；3）结合临床肿瘤病例分析lnc-RI表达水平与结直肠癌放射抵抗性的相关性。

本项目探索长链非编码RNA分子lnc-RI对结直肠癌细胞放射敏感性的调控作用和分子机制，旨在寻找解决结直肠癌放射抗性的干预靶点。主要从下面三个方面开展研究：1）证实lnc-RI通过促进DSB的NHEJ修复增强结直肠癌细胞放射抵抗性；2）明确LIG4在lnc-RI调节放射抵抗性中的作用及lnc-RI调节LIG4的机制；3）结合临床肿瘤病例分析lnc-RI表达水平与结直肠癌放射抵抗性的相关性。主要研究结果包括：（1）下调lnc-RI表达抑制结直肠癌细胞在体内外的生长，促进其凋亡，诱导了内源性DSBs的增加并引起细胞周期阻滞。下调lnc-RI表达抑制了NHEJ修复效率且明显抑制了其关键蛋白LIG4的表达，lnc-RI通过竞争结合miR-4727-5p调控LIG4的表达进而参与调控DNA损伤修复。下调lnc-RI表达增强了结直肠癌细胞的放射敏感性。（2）MiR-1587 mimics 抑制了结直肠癌细胞的生长并促进其凋亡，伴随着DSBs的增加并引起了细胞周期阻滞。MiR-1587通过靶向结合LIG4 mRNA 3’UTR区调控LIG4的表达，miR-1587 mimics增强了结直肠癌细胞的放射敏感性。本研究提示lnc-RI可能是结直肠癌潜在的放射治疗靶点，并且发现MiR-1587通过靶向结合LIG4 mRNA 3’UTR区参与调控结直肠癌细胞DNA损伤修复与放射敏感性。