缺氧时HIF-1α转录激活自噬蛋白Beclin 1促进鼻咽癌转移机制的研究

We had proved that inhibition of Aur-A/Snail/MAPK signaling could partly suppress HIF-1α induced distant metastasis for human nasopharyngeal carcinoma. We supposed that HIF-1α induced metastasis might be raced by varied pathways for nasopharyngeal carcinoma. We further found that HIF-1α level was closely correlated with autophagic key regulator Beclin 1, and with tumor metastasis for nasopharyngeal as well as other cancer types. Moreover, three Hypoxia Responsive Elements (HREs), the specific gene region to be transcriptional combined by HIF-1α, were detected in Beclin 1 enhancer area in nasopharyngeal cancer CNE-1 cells. We therefore raised that HIF-1α induced metastasis might also be mediated by transcriptional upregulating Beclin 1, and further activating autophagy for nasopharyngeal carcinoma. It has been known that the stabilization of autophagic Beclin 1/PI3K complex might be ascribed to their interaction with MAPK signaling. In this study, we will characterize the underlying mechanism between HIF-1α and Beclin 1 in promoting metastasis for nasopharyngeal carcinoma. Furthermore, the crosstalk between Aur-A/Snail/MAPK and HIF-1α/Beclin 1/autophagy signaling in regulating metastasis will be elucidated. Importantly, the malignant phenotype of HIF-1α/Beclin 1/autophagy in regulation of metastasis would be further confirmed in clinical nasopharyngeal carcinoma tissues. In conclusion, we will address the transcriptional mechanism between HIF-1α and Beclin 1, and characterize their crosstalk with Aur-A/Snail/MAPK signaling, highlighting the essential role of hypoxia induced autophagy in promoting metastasis for nasopharyngeal carcinoma.

青年基金项目证实抑制Aur-A/Snail/MAPK通路仅能部分阻断缺氧时HIF-1α促发的鼻咽癌转移，那么HIF-1α促发的转移是否存在其他途径？我们发现在鼻咽癌等肿瘤中HIF-1α与自噬关键蛋白Beclin 1的表达呈正相关，且促进转移；深入研究发现Beclin 1基因增强子区含有HIF-1α启动转录需结合的特异序列-缺氧反应元件HRE。由此我们提出了HIF-1α促进鼻咽癌转移的新途径：HIF-1α转录结合Beclin 1-HRE，诱导自噬促进转移。已知MAPK可稳定Beclin 1-自噬复合体。本课题拟在三维转移模型中探讨HIF-1α转录Beclin 1、调控自噬及转移的机制；研究Aur-A/Snail通路调控MAPK，稳定Beclin 1-自噬复合体的机制；结合临床资料验证，最终获得HIF-1α转录Beclin 1、串话Aur-A/Snail/MAPK通路、调控自噬及转移的可靠证据

由于肿瘤组织增殖迅速且肿瘤血管发育畸形，导致肿瘤内部处于缺氧环境。我们前期的研究证实，缺氧环境诱导的HIF-1α表达升高及其通路激活是导致肿瘤转移和放疗抵抗的重要原因。在本项目的工作中，我们进一步研究了HIF-1α与自噬通路、DNA损伤修复通路的串话，并针对其分子机制和临床意义做了深入研究。我们的研究证实：Beclin 1、ULK1、ATG10等自噬相关分子抑制胃癌肿瘤复发和远端转移，综合多个分子的模型可有效预测胃癌病人的预后情况（Am J Transl Res，2016）。此外，Beclin 1表达水平与肿瘤转移和预后的负相关也在胆管癌中得到验证（Plos one，2013）。通过基因组范围的CRISPER-CAS9基因敲除病毒库，我们筛选发现NONO是HIF-1α诱导肿瘤放疗抵抗的重要下游分子。进一步研究发现，NONO通过促进EGFR的蛋白表达，并在细胞核内与EGFR蛋白结合，促进下游DNA损伤修复和放疗抵抗（未发表）。在临床水平，项目负责人根据临床工作经验和需求，在鼻咽癌的临床反射治疗中，尝试了插植三维适形放疗方案。我们发现，该方案能够有效提高局部早期复发鼻咽癌患者的治疗效果，重要的是周围正常组织放射损伤小，放射防护效果理想（Plos one，2014）。此外，在肿瘤放疗抵抗和肿瘤转移的机制研究中，使用三维细胞培养模型能够更好的模拟肿瘤或正常组织结构，与传统二维细胞培养模型相比具有巨大优势。项目负责人探索构建了新型的三维细胞培养方案，并获得了2项发明专利（项目负责人为第一发明人）。. 在本项目的资助下，项目负责人的科研工作获得了良性发展，获得了多个人才项目支持，其中包括2014年度广东省自然杰出青年科学基金、中山大学附属第六医院“五个五”人才专项，并获得获得联合国下属国际遗传工程和生物技术中心组织（International Centre for Genetic Engineering and Bio-technology，ICGEB）的科研基金资助。目前，后续各项研究正在按计划开展中。