前列腺癌细胞通过表达NanogP8来获得永生性及去势耐受性的分子机制研究

Prostate cancer (PCa) comprises a spectrum of phenotypically heterogeneous cells and may harbor stem cell-like cells commonly referred to as prostate cancer stem cells (PCSCs). Although the PCSC concept is still in debate and the true molecular nature and clinical significance of PCSCs remain to be fully elucidated, it is undeniable that malignant tumors are immortal at the population level. One of the key unanswered biological questions of therapeutic implication is how PCa cells in general and PCSCs in particular are regulated at the molecular level with respect to their self-perpetuating (immortal) tumorigenic potential, i.e., self-renewal properties. Remarkably, malignant cells share a similar gene expression profile with embryonic stem cells, which express several core regulators of self-renewal and pluripotency including Nanog (also called Nanog1), a homeodomain transcription factor located on chromosome 12. We have observed that human PCa and other cancer cells do not express Nanog1 but rather express a retrotransposed Nanog1 homolog, called NanogP8, located on chromosome 15. Knocking down NanogP8 significantly inhibits the PCSC properties in vitro and tumor regeneration of PCa cells. In contrast, NanogP8 overexpression is sufficient to confer PCSC properties, enhances tumor regeneration, and significantly promotes development of castration-resistant PCa (CRPC). Our recent whole-genome ChIP-Seq experiment has uncovered distinct NanogP8 DNA-binding profiles. Furthermore, newly generated NanogP8 transgenic animals develop hyperplastic lesions in the target organs. These observations, taken together, lead to our overarching hypotheses that 1) NanogP8 plays a causal role in the development of CRPC; 2) NanogP8 promotes tumor development by regulating PCSC self-renewal via unique signaling pathways; and 3) NanogP8 possesses protumorigenic activity. The current project tests these three hypotheses, mainly in PCa, with the following three specific aims...1) To test the hypothesis that NanogP8 is causally involved in the development of CRPC;.2) To elucidate the mechanisms underlying NanogP8-regulated PCSC self-renewal; and.3) To explore the protumorigenic roles of NanogP8 using novel transgenic animal models...Accomplishment of the goals proposed herein should greatly advance our understanding towards how NanogP8 regulates the immortality of PCa cells and contribute to PCSC self-renewal and CRPC formation. It will also facilitate development of mechanism-based therapeutics that specifically targets PCSCs.

前列腺癌(prostate cancer, PCa)细胞，特别是癌干细胞(prostate cancer stem cell, PCSC)通过何种机制调控自我更新以维系其永生性并产生去势耐受性，是抗癌领域亟待解决的基础问题。类似胚胎干细胞中的Nanog1，癌细胞可表达一种与其同源的转座子NanogP8。本实验室通过敲除与过表达实验，观测到NanogP8在调控PCa细胞致瘤性及去势耐受过程中发挥重要作用。基于此，拟构建报道基因，利用ChIP-Seq及转基因模型，1)论证NanogP8可引发去势耐受形成；2)揭示NanogP8调控PCSC自我更新的机制；3)论证NanogP8原发致瘤活性。此三目标紧密围绕NanogP8的基因特征、蛋白调控及细胞效应三个方面，以期解析NanogP8调控癌细胞永生性及去势耐受性的机制，为研发以NanogP8为靶点、针对PCSC的靶向抗癌手段提供转化医学平台。

肿瘤细胞如何获得永生性，并不被人们所了解。我们最近研究证实，癌细胞可以篡夺与胚胎干细胞自我更新相关的关键因子，来维系它们的永生特性。胚胎干细胞可以表达一种Nanog1蛋白，其位于12号染色体，与细胞的自我更新有关。与之相反，肿瘤细胞优先表达一种与Nanog1同源的转座子，称为NanogP8，其位于15号染色体，与细胞的致瘤潜力相关。在本项目中，我们进一步阐明了NanogP8在前列腺癌细胞中的分子功能及其机制。为了深入理解人类前列腺癌细胞如何调控自我更新、增殖和启动肿瘤发生等一系列问题，我们对参与调节胚胎干细胞的“干性”和肿瘤细胞永生性的大约30个因子进行了无偏倚的筛查，其结果提示NanogP8分子在调控细胞“干性”方面发挥了关键作用。我们的研究证实，人类前列腺癌和一些其他癌细胞并不表达Nanog1，而是表达与Nanog1具有同源结构域的转座子，称为NanogP8，位于15号染色体上。体外敲除NanogP8能够显著抑制肿瘤干细胞的活性，有效遏制前列腺癌、乳腺癌及结肠癌细胞的再生与增殖。相反地，NanogP8的过表达可以维持肿瘤干细胞的活性，促进肿瘤再生，并显著推进去势抗性前列腺癌的发生发展。启动子示踪研究表明，表达Nanog的前列腺癌细胞具有肿瘤干细胞特性。我们的研究揭示了NanogP8 DNA特异性结合谱。在新建立的NanogP8转基因动物中，其靶器官发生了增生性病变。综上，本项目研究论证了：1) NanogP8具有启动肿瘤发生的特性; 2) NanogP8通过独特的信号转导通路调控肿瘤干细胞自我更新，从而促进肿瘤发展；3) NanogP8能够引发去势耐受性前列腺癌的形成。应用基因启动子示踪系统、时间轴视频显微监测技术、全基因组 ChIP-Seq测序以及NanogP8转基因动物模型，本项目研究任务已如期实施。本项目的研究工作推进了人们对于前列腺癌细胞及癌干细胞生物行为的认知水平，有望加速研发能够特异性针对肿瘤干细胞的靶向抗癌手段。