PPARγ2-PTEN信号轴调制人前列腺癌变细胞程序性坏死(necroptosis) 的分子机制及其化学预防应用

Loss of PTEN tumor suppressor gene is closely related to the occurrence, development and treatment of prostate cancer. Our series of experiments found that dysregulation of PPARγ-PTEN signal axis led to mouse and human prostate carcinogenesis with active autophagy. Overexpression of PPARγ2 cDNA induced necroptosis/necrosis in human advanced prostate cancer with stromal inflammatory cell accumulation, leading to cancer growth inhibition. The key scientific problems to be solved are further proposed: To clarify the outcome relationships among autophagy, autophagic cell death (ACE) and necroptosis in normal and benign human prostate epithelial cells and their molecular regulation mechanisms. Next, we propose a scientific hypothesis: Modulation of necroptosis in human prostate tumor-initiating cells (TICs) via precise regulation of PPARγ2-PTEN signal axis may serve as an important approach of cancer interference and chemoprevention. We plan to use human normal and benign prostatic epithelial cell lines NHPrE1 and BHPrE1, their 3D-organoid culture and tissue recombination-xenografting (TRX) translational mouse models developed in our laboratory for establishment models of loss of PTEN human prostate carcinogenesis in vitro and in vivo. We will use them for screening a combination of bioactive natural compound Pseudolaric Acid B, PAB and modulators of necroptosis for precise regulation of PPARγ2-PTEN signal axis (a target) to modulate necroptosis in human prostate tumor-initiating cells. It will become an important approach to chemical prevention of prostate carcinogenesis. To elucidate the molecular mechanism by which PPARγ2-PTEN signal axis modulates necroptosis in human prostate tumor-initiating cells for inhibition of carcinogenesis can be used as a new idea to rationally formulate a long-term cancer preventive strategy.

PTEN肿瘤抑制基因失活与前列腺癌发生密切相关。我们在系列实验中发现PPARγ-PTEN信号轴失调导致前列腺上皮细胞癌变，与病理性自噬发生相关；PPARγ2高表达诱导人进展期前列腺癌细胞程序性坏死（necroptosis)和间质炎症细胞聚集，抑制肿瘤生长。提出科学问题：人前列腺上皮细胞癌变过程中自噬、自噬性坏死与程序性坏死的转归关系及其调控机制。并提出科学假说：通过PPARγ2-PTEN信号轴调制人前列腺癌干细胞程序性坏死，进行肿瘤干预和化学预防。我们拟应用本实验室创建的人正常和良性前列腺上皮细胞系NHPrE1和BHPrE1及其微器官培养和组织重组-移植瘤TRX小鼠等模型，构建PTEN失活人前列腺癌变-小鼠移植瘤模型，通过筛选天然药物活性组分土荆皮乙酸PAB与程序性坏死调制剂组合，精准调控PPARγ2-PTEN信号轴调制癌干细胞程序性坏死来抑制肿瘤发生,为制定肿瘤长期化学预防策略提供思路。

近年来，随着人口老龄化及生活习惯的改变，前列腺癌在中国的发病率逐年上升。我们在系列研究中发现PPARγ2-PTEN信号轴失调在前列腺癌的发生发展中发挥重要作用，本项目系统的研究了PPARγ2-PTEN信号轴在人前列腺癌干细胞及癌细胞中的生物学功能及调控机制。.首先，利用人正常前列腺上皮细胞NHPrE1成功构建人前列腺干细胞样细胞NHPrE1-ALDH1A1+/CD133+，PTEN丢失人前列腺癌干细胞样细胞NHPrE1-ALDH1A1+/CD133+/KOPTEN以及PPARγ2高表达人前列腺癌干细胞样细胞NHPrE1-ALDH1A1+/CD133+/KOPTEN/PPARγ2+，发现PTEN失活促进人前列腺干细胞样细胞NHPrE1-ALDH1A1+/CD133+的增殖，且迁移和侵袭能力增强。PPARγ2-PTEN轴功能性重建可显著抑制人前列腺癌干细胞样细胞NHPrE1-ALDH1A1+/CD133+/KOPTEN的增殖，诱导细胞周期阻滞、细胞凋亡、自噬及程序性坏死的发生，并干预了干性相关通路、癌相关转录失调、细胞凋亡和坏死等信号通路。同样，在人前列腺癌细胞LNCaP及PC3（PC3是前期工作，本项目对其进行补充和完善）中重建PPARγ2-PTEN信号轴表达及功能活性，可以在体内、外对人前列腺癌细胞产生生长抑制，并发生细胞周期阻滞和诱导凋亡的发生。进一步对前期通过生物活性小分子化合物库筛选，得到的PPARγ2特异性激活剂土荆皮乙酸（PAB）进行功能分析，发现其可以功能性重建PPARγ2-PTEN信号轴，并抑制人前列腺癌细胞增殖、侵袭和诱导细胞凋亡，提示其具备作为有效的小分子药物对肿瘤干预和化学预防的潜力。最后,本项目探讨了PPARγ2-PTEN信号轴传导过程中的分子调控机制，提出PPARγ2-AKAP12-PTEN/AKT信号传导模式，其调控机制为PPARγ2通过诱导miR-200b-3p上升表达，抑制DNMT3A/3B活性，从而促进AKAP12启动子区去甲基化，增强了PPARγ2与AKAP12启动子区的结合，促进AKAP12的转录，并通过PTEN/AKT信号通路行使下游功能。PPARγ2-AKAP12-PTEN/AKT信号轴的发现可以为前列腺癌转化医学研究及临床实验提供新的靶点，PAB有望应用于前列腺癌的化学预防及临床治疗。