HuR介导的转录后调控在青蒿素类药物抗非小细胞肺癌转移中的作用和机制研究

The anti-malarial drug artemisinin has demonstrated potent anti-cancer activity. Our previous studies also indicate that artemisinin inhibits tumor-associated lymphangiogenesis and lymph node metastasis, and protects tumor-harboring mice from death. However, the molecular mechanisms affecting the anti-metastastic activity of artemisinin remain unclear. HuR (human antigen R), a RNA-binding protein, is one of the most important molecules regulating post-transcriptional process, which has been shown to regulate the mRNA stability of several targets and thus promote the invasion and metastasis of many tumors including lung cancer. However, whether the post-transcriptional machinery mediated by HuR participates in the anti-metastasis activity of artemisinin in lung cancer has not been determined. We thus hypothesize that HuR may be a key molecule contributing to the anti-metastasis effects of artemisinin in lung cancer. In this study, we aim to systematically investigate the in vitro and in vivo activity of artemisinin in lung cancer by analyzing the invasion and metastasis of lung cancer cells treated with artemisinin and its analogs and with knockdown or overexpression of HuR, explore the potential molecular mechanisms involved in regulation of HuR functions by artemisinin, and to identify key molecules regulated by HuR that contribute to the effects of artemisinin in lung cancer. Our study may reveal the roles and mechanisms of HuR-mediated post-transcriptional machinery in artemisinin-induced inhibition of invasion and metastasis of lung cancer. Furthermore, our study may provide evidence and explanations for the expanded clinical indications of the anti-cancer agent artemisinin.

近年发现抗疟疾药物青蒿素也具有抗肿瘤活性；我们前期实验证实青蒿素能抑制肿瘤转移，对移植瘤小鼠有保护作用，但青蒿素抗肿瘤的相关分子机制不清。我们还发现转录后调控分子HuR表达定位影响肺癌等实体瘤预后，它通过调控多种侵袭、转移相关分子的mRNA稳定性和蛋白表达参与肿瘤转移，并促进肿瘤细胞迁移。但是，HuR介导的转录后机制是否参与青蒿素抗肿瘤转移未有相关报道。因此，我们假设：HuR转录后调节侵袭、转移分子的表达是青蒿素类药物抗肿瘤转移的关键机制。本课题拟在前期阳性预实验结果的基础上，通过迁移、侵袭等药效学实验、动物实验系统性探讨青蒿素类药物对肺癌细胞生物学的影响；通过分子生物学实验分析青蒿素类药物调节HuR功能的机制，进而明确青蒿素类药物抗肿瘤转移的关键靶分子。本研究旨在探讨转录后调节机制参与青蒿素抗肿瘤作用的分子机制，为增加青蒿素临床适应证、作为备选的治疗肺癌的药物提供可靠的实验依据。

近年发现抗疟疾药物青蒿素也具有抗肿瘤活性。我们前期实验证实青蒿素能抑制肿瘤转移，对移植瘤小鼠有保护作用，但青蒿素抗肿瘤的相关分子机制不清。我们还发现转录后调控分子HuR表达定位影响肺癌等实体瘤预后，它通过调控多种侵袭、转移相关分子的mRNA稳定性和蛋白表达参与肿瘤转移，并促进肿瘤细胞迁移。但是，HuR介导的转录后机制是否参与青蒿素抗肿瘤转移尚不清楚。因此，我们假设HuR转录后调节侵袭、转移分子的表达是青蒿素类药物抗肿瘤转移的关键机制。本研究中，我们发现青蒿素衍生物青蒿琥酯对EGFR非敏感突变细胞A549的增殖、迁移、侵袭具有抑制作用，其机制涉及MMP-9表达下调；通过体外功能实验证实HuR是MMP-9的上游分子，其通过细胞核-细胞浆转运、转录后机制调控其表达，抑制肿瘤生长和转移。我们还发现青蒿琥酯对EGFR突变细胞H1975的迁移、侵袭具有明显抑制作用，其机制涉及CXCR4表达下调。通过功能实验证实HuR是CXCR4的上游分子，HuR通过细胞核-细胞浆转运、转录后机制调控CXCR4表达，抑制肿瘤转移。我们证实了促凋亡蛋白Bim表达与EGFR敏感突变NSCLC EGFR-TKI原发耐药和肿瘤进展密切相关。进一步发现Bim是HuR的靶基因，HuR通过转录后机制调控了Bim表达。在体外实验中，青蒿琥酯对EGFR突变细胞H1975的增殖具有抑制作用，能诱导细胞凋亡，其机制涉及P-IGF-1R及其下游PI3K/AKT通路抑制，及增加的Bim表达；由于HuR是Bim的上游分子，HuR通过细胞核-细胞浆转运、转录后机制调控Bim表达，逆转肿瘤耐药和疾病进展。本研究提示HuR介导的转录后调节机制参与青蒿素抗肿瘤作用的分子机制，这为增加青蒿素临床适应证、作为备选的治疗肺癌的药物提供了可靠实验依据。