Hu Hao, Chen Wuqiang, Zhang Shuo, Xue Yuzheng, He Youzhao, Gu Yuanlong
Hepatobiliary and Pancreatic Surgery, Affiliated Hospital of Jiangnan University, 585 Xingyuan Rd, Liangxi District, Wuxi, 214041, China.
School of Medicine, Jiangnan University, Wuxi 214122, China.
J Cancer. 2021 Jul 25;12(18):5622-5632. doi: 10.7150/jca.58824. eCollection 2021.
Although we previously revealed that DNA-dependent protein kinase catalytic subunit (DNA-PKcs) is overexpressed in pancreatic ductal adenocarcinoma (PDAC) and important for gemcitabine resistance, the role of DNA-PKcs in the progression and metastasis of PDAC remain unclear. To date, the upstream signaling events stimulating DNA-PKcs overexpression in PDAC are still not well characterized. Expression of DNA-PKcs was measured by western blot. The levels of miRNA-101 and lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) were detected by real-time PCR. Cell viability was determined by CCK-8. Cell migration and cell invasion were measured by transwell assay. The regulatory relationship between NEAT1 and miR-101 was determined by a luciferase assay. DNA-PKcs expression was significantly elevated in human PDAC tissues and cells. DNA-PKcs overexpression was correlated with TNM stage and lymph node metastasis. Higher expression of DNA-PKcs was closely correlated with patients of worse overall survival (OS). DNA-PKcs knockdown suppresses malignant behaviors of PDAC cells. Further study showed that miRNA-101 level was decreased in PDAC tissues and cells, which could be responsible for DNA-PKcs overexpression and DNA-PKcs mediated oncogenic actions in PDAC cells. Moreover, NEAT1 functions as an oncogene influencing cell proliferation, migration and invasion in part by serving as a competing endogenous RNA (ceRNAs) modulating miR-101 expression, leading to up-regulation of DNA-PKcs. These findings suggest that NEAT1/miR-101-dependent up-regulation of DNA-PKcs promotes the malignant behaviors of PDAC cells. The NEAT1/miR-101/DNA-PKcs axis may serve as a viable prognostic marker and therapeutic target for PDAC.
尽管我们之前揭示了DNA依赖性蛋白激酶催化亚基（DNA-PKcs）在胰腺导管腺癌（PDAC）中过表达且对吉西他滨耐药很重要，但DNA-PKcs在PDAC进展和转移中的作用仍不清楚。迄今为止，刺激PDAC中DNA-PKcs过表达的上游信号事件仍未得到很好的表征。通过蛋白质印迹法检测DNA-PKcs的表达。通过实时PCR检测miRNA-101和长链非编码RNA核旁斑组装转录本1（NEAT1）的水平。通过CCK-8法测定细胞活力。通过Transwell实验测量细胞迁移和细胞侵袭。通过荧光素酶实验确定NEAT1与miR-101之间的调控关系。DNA-PKcs在人PDAC组织和细胞中表达显著升高。DNA-PKcs过表达与TNM分期和淋巴结转移相关。DNA-PKcs的高表达与总生存期（OS）较差的患者密切相关。敲低DNA-PKcs可抑制PDAC细胞的恶性行为。进一步研究表明，PDAC组织和细胞中miRNA-101水平降低，这可能是PDAC细胞中DNA-PKcs过表达和DNA-PKcs介导的致癌作用的原因。此外，NEAT1作为一种癌基因，部分通过作为调节miR-101表达的竞争性内源RNA（ceRNA）影响细胞增殖、迁移和侵袭，导致DNA-PKcs上调。这些发现表明，NEAT1/miR-101依赖性上调DNA-PKcs促进了PDAC细胞的恶性行为。NEAT1/miR-101/DNA-PKcs轴可能作为PDAC可行的预后标志物和治疗靶点。
