Pot1端粒保护机制在正常及肿瘤细胞间的差异及其在肿瘤特异性治疗中的应用研究

Telomere dysfunction and telomerase reactivation has been implicated in human cancer pathogenesis. Also, lots of compelling studies have demonstrated that targeting telomeres would be a novel potential approach towards the end of cancer elimination. However, the therapeutic targets capable of discriminatingly destroying cancer cellular telomere structure without imparing the normal cellular telomere function have not been identified yet. We have got convincing data indicating that the structural and functional relationship underlying the telomeric protein Pot1 to protect telomeres in human normal cells has been widely changed in human telomerase active cancer cells, thereby providing an approach to solve the major challenge facing by the current telomere-based targeted cancer therapies. As this novel finding may have important implications in both studies of cancer pathogenesis and cancer therapy, we will focus our major interests of this proposed project on the following questions: what details in the structural-functional relationship for Pot1 performing the same telomere protection are different between the human normal and cancer cells; How the Pot1 having the normal structural-functional relationship changed functions in cancer cells for protecting the cancer cellular telomeres and what other unknown aberrant mechanisms may involve in this cancerous Pot1 telomere protection pathway. By answering these these questions, we will get to the specifial aim for this project to discover a potential approach towards specifically targeting the aberrant telomere protection by Pot1 and evaluate the potential significances for this approach in cancer specific therapy.

端粒异常以及端粒酶重新活化已经被证明与人类肿瘤发生有普遍性的内在联系。大量研究也已经证明靶向端粒可以成为根治肿瘤的一条希望途径。然而能够实现特异性靶向肿瘤异常端粒结构而不影响正常细胞端粒功能的治疗性靶标至今仍未被发现。我们在最近研究中已获得确凿证据表明端粒蛋白Pot1在发挥端粒保护效应中涉及到的结构-功能关系及相应分子相互作用在正常人体细胞和端粒酶阳性肿瘤细胞间存在实质性差异，有可能为解决上述问题打开一个突破口。基于这一新发现在肿瘤研究中潜在的重大理论和现实意义，本课题将重点研究Pot1保护端粒的结构功能关系在肿瘤和正常细胞间差异性的具体体现；肿瘤细胞中结构功能关系改变的Pot1蛋白发挥端粒保护生物学效应的功能状态及相应异常的功能维持机制。通过对这些问题的阐明，揭示出靶向抑制肿瘤异常的Pot1端粒保护机制的可行性分子途径及其在肿瘤特异性治疗上的潜在意义。

细胞癌变中获得端粒酶端粒DNA长度稳定功能是大多数人肿瘤发生的必要条件,是肿瘤细胞获得永生化生长能力的基础。但大量证据表明肿瘤发育中在获得端粒酶端粒稳定功能之前已经历了极度端粒缩短,造成肿瘤端粒保护Shelterin复合体一种特殊的功能缺陷,表现为肿瘤端粒发生异常高频的病理性融合。但肿瘤端粒shelterin结构上发生怎样的改变并不清楚。端粒融合涉及shelterin对端粒末端单链DNA的保护不足。POT1是shelterin中唯一的端粒单链DNA结合因子，同时也是shelterin中端粒单链DNA的专业性保护因子。本项目研究POT1端粒单链DNA结合保护功能在正常和肿瘤细胞端粒shelterin结构中潜在的差异性及在肿瘤细胞靶向特异性杀伤中的应用。取得以下重大发现。我们的试验数据证明，由TPP1募集的POT1在正常细胞端粒shelterin结构中表现为正统的端粒单链DNA结合保护因子，组成型结后定位在端粒单链DNA上，并完全依赖其自身的单恋DNA结合活性发挥保护作用。但同样由TPP1募集的POT1在端粒酶阳性肿瘤细胞端粒结构中丧失与端粒单链DNA稳定结合能力，与其自身的单链DNA结合活性无关，同时证明了POT1在肿瘤端粒shelterin结构中获得一种在正常端粒shelterin结构中无法工作的不依赖其单链DNA结合能力的端粒单链DNA保护功能。在以上发现基础上，我们进一步证明靶向性打断POT1和TPP1在shelterin结构中的蛋白-蛋白连接不影响正常细胞端粒shelterin的完整保护功能，但专一性地去除肿瘤端粒shelterin中的POT1，肿瘤端粒DNA损伤反应活化，具有完全杀死各种组织来源的端粒酶阳性肿瘤细胞，而对不同组织来源的端粒酶阴性和阳性的正常细胞的生存和长期生长没有影响。本研究揭示了人正常细胞固有的shelterin POT1端粒单链DNA结合定位的结构构象在端粒酶阳性肿瘤中普遍发生了改变，使肿瘤端粒shelterin结构固有性存在POT1端粒单链DNA结合缺陷，提出了端粒shelterin结构重塑与肿瘤发生密切相关的新理论，同时建立了一个靶向端粒shelterin结构的肿瘤细胞特异性杀伤的新方法，提供了一条潜在的肿瘤异常端粒shelterin结构靶向性治疗新途径，具有重大科学意义。有关本项目研究发现的论文一投稿。