基于BRD4新靶点实现分子靶向及131I协同治疗碘难治性分化型甲状腺癌的实验研究

Radioactive iodine refractory differentiated thyroid cancer（RAIR-DTC）is the most important reason that affects the disease free survival, disease specific mortality and life quality of patients. The pathogenesis of RAIR-DTC is still unclear. The abnormal activation of some signaling pathways that can down regulate the expression of NIS and TSHR, and then inhibit the ability of iodine uptake in thyroid cancer cells may be one of the important mechanisms. Molecular targeted therapy for abnormal expression genes is considered to be an important method for the treatment of RAIR-DTC, and some therapeutic effects have been achieved in clinic. But the response rate is still unsatisfactory. Therefore, the key to improve the prognosis of patients is to explore the pathogenesis of RAIR-DTC and find new molecular targets. Our previous study found that BRD4 is over-expression in differentiated thyroid carcinoma (DTC). And JQ-1, one of BRD4 inhibitors, which has obvious anti-tumor effect, can improve the iodine uptake function and the expression level of NIS gene in DTC cells and nude mice model. Therefore, we intend to further clarify the role of BRD4 in tumor proliferation, iodine metabolism and signaling pathways, to study the effect of JQ-1 (BRD4 inhibitor) on tumor inhibition and 131I uptake, to explore the mechanism of synergistic therapy of JQ-1 and 131I in RAIR-DTC. We try to provide a new way of studying the pathogenesis and to explore a new molecular target of RAIR-DTC.

碘难治性分化型甲状腺癌（RAIR-DTC）的发病机制还未明了，如多种信号通路的异常激活使NIS、TSHR等基因表达下调，抑制癌细胞摄碘能力、促进癌细胞的恶性增殖是可能的重要机制之一。分子靶向治疗是治疗RAIR-DTC的重要方法，但反应率仍不尽人意。探究RAIR-DTC的发病机制并寻找新的分子靶点是亟需解决的关键。我们前期研究发现BET蛋白家族中的BRD4在分化型甲状腺癌中高表达，BRD4的抑制剂JQ-1具有明显的抑瘤作用，并可提高DTC癌细胞及裸鼠模型肿瘤组织的摄碘功能及NIS的表达水平。因此，本研究拟通过BRD4基因沉默进一步阐明其在RAIR-DTC肿瘤增殖、碘代谢的分子机制及相关信号通路中的作用，研究BRD4抑制剂JQ-1对RAIR-DTC的抑瘤作用和提高131I摄取率的作用，探索JQ-1与131I协同治疗机制，为研究RAIR-DTC的发病机制、探索新的靶标提供一种新的思路与方法。

本研究以BET蛋白家族中的BRD4基因为突破点，聚焦于碘难治甲状腺癌的治疗。通过构建稳转BRD4-shRNA-8505C和NC-shRNA-8505C细胞系，进行细胞增殖、凋亡、周期、迁移等体外实验和荷瘤小鼠体内实验，并通过western blot探寻其疗效机制。结果表明，沉默BRD4基因可抑制未分化甲状腺癌细胞的增殖、侵袭及迁移能力，促进未分化甲状腺癌细胞系的再分化。BET抑制剂JQ1可抑制未分化甲状腺癌细胞系（8505C和CAL-62）的增殖、侵袭及迁移能力，促进细胞凋亡，改变了细胞周期，并增加甲状腺分化相关蛋白（TTF-1和PAX8）和碘代谢相关蛋白（NIS和TPO）的表达，促进细胞再分化，使得细胞摄碘能力增加。另外，动物实验显示JQ1和131I联合治疗可以明显抑制肿瘤的生长，JQ1可提高未分化甲状腺癌细胞和肿瘤组织的摄碘功能及NIS的表达水平。沉默BRD4基因或使用JQ1均可通过抑制PI3K/AKT信号通路促进NIS的表达。我们的研究在一定程度上揭示了碘难治甲状腺癌的发病机制，也探索了新的靶标BRD4在碘难治甲状腺癌治疗中的潜力。除此之外，我们从生物信息学、中药网络药理学、临床实践出发，围绕甲状腺癌进行了一系列科学研究，为临床中甲状腺癌的治疗提供更多新的思路与方法。