脂质代谢调控基因SRB1和ABCA1在前列腺癌恶性进展中的作用和机制研究

SRB1 (scavenger receptor class B member 1) functions as a receptor for high-density lipoprotein facilitating the uptake of cholesteryl esters, and ABCA1 (ATP-binding cassette transporter member 1 of human transporter sub-family ABCA) also known as the cholesterol efflux regulatory protein (CERP) is a major regulator of cellular cholesterol and phospholipid homeostasis. A hallmark of malignant tumors is disorder of regulatory networks of cellular energy metabolism. Among a variety of malignant tumors including breast cancer and prostate cancer, lipid metabolism was found activated. Previous researches in tissue samples of radical prostatectomy showed that levels of poly-unsaturated fatty acids cholesterol ester are significantly increased in tumor tissues compared with adjacent benign prostate tissues. Furthermore, the ratios of levels in tumor tissues compared with adjacent benign prostate tissues are consistent with degree of invasion and malignancy. Clinical evaluation of malignancy of prostate cancer is related to the content of cholesterol ester, but variations of a single gene cannot directly associate with degree of malignancy. SRB1 and ABCA1 play an important role in this metabolic disorder. Based on previous studies and literatures, the project will focus on the expression of genes related to lipid metabolisms and relationship with progression of prostate cancer. Through simulating tumor microenvironment of high cholesterol, we plan to detect expression of genes and proteins which may contribute to accumulation of cholesterol ester. We plan to analyze the possible mechanism of epithelial-mesenchymal transition by altering tumor microenvironment and identify the key genes involved in microenvironment changes. The project also aim to investigate the possible inhibition of tumor invasion and metastasis by reducing accumulation of cholesteryl ester. Through above plans, we hope to provide new evident involving the pathogenesis and progression of prostate cancer, molecular typing, and find an alternative tool to individualized therapy.

恶性肿瘤的重要标志是细胞能量代谢调控网络紊乱，多种肿瘤包括前列腺癌乳腺癌早期均发现脂质代谢的激活。我们在前期对前列腺癌根治手术组织标本的研究发现，不饱和脂肪酸胆固醇酯在癌组织中的含量明显升高，且其含量与前列腺癌的恶性程度，侵袭进展趋势一致。进一步研究发现前列腺癌的临床恶性进展与胆固醇酯的含量密切相关，但单一基因的变化无法直接与恶性程度相关联。SRB1和ABCA1作为调控细胞胆固醇代谢平衡的重要分子，在这一代谢紊乱中起到重要作用。本项目将主要研究其在前列腺癌中的表达及与前列腺癌进展的关系；模拟高胆固醇酯蓄积肿瘤微环境，检测引起胆固醇酯蓄积相关基因和蛋白表达水平，确定参与前列腺癌恶性进展的关键基因和蛋白表达变化；探讨抑制胆固醇酯蓄积降低前列腺癌侵袭转移可能。从而可能在前列腺癌发病机制，分子分型方面提供新的证据，为个体化治疗提供新的可行途径。

恶性肿瘤的一大重要特征是细胞能量代谢调控网络紊乱，前列腺癌细胞中脂质代谢异常激活。我们采用代谢组学的技术对76例前列腺癌患者和19例良性前列腺增生患者的171个前列腺组织样本进行了检测，发现不饱和脂肪酸胆固醇酯在前列腺癌组织中含量明显升高，而且其含量与前列腺癌的恶性程度侵袭进展去势一致。联合代谢组学和转录组学数据整合分析，发现不饱和脂肪酸合成过程中的SRB1/SRB1和ABCA1显著上调，相应调节的miRNA表达下调。对于胆固醇酯蓄积密切相关的胆固醇摄取相关酶类，SRB1显著上调，其相关miRNA（miR-125a-5p/455-3p/455-5p）显著下调。在前列腺癌细胞系中检测SRB1的表达水平，发现其在肿瘤细胞系22RV1中表达上调。构建稳定干扰SRB1的22RV1细胞系，发现细胞的增殖能力和克隆形成能力减弱，其总胆固醇和胆固醇酯水平均发生下调，提示胆固醇摄入水平下降。在92例前列腺癌组织样本的组织芯片中进行了SRB1的免疫组化检测，我们发现SRB1在肿瘤组织中高表达，而在癌旁正常组织中表达水平较低；进一步分析患者临床资料发现，在高Gleason评分患者人群中其SRB1的表达水平较高，而在低Gleason评分患者人群其SRB1的表达水平较低。而在针对SRB1（NM_005505）序列信息以及前列腺癌发生进展相关的miRNA库，应用miRDB和TargetScan在线数据库进行序列比对，发现miR-125a-5 p/455-3p/455-5p与SRB1的3’UTR可能发生结合。转染miRNA mimics和inhibitors进入细胞，发现肿瘤细胞系的增殖能力和克隆形成能力发生相应变化，SRB1的mRNA表达水平也发生相应下调和上调。构建双荧光素酶报告载体及相应结合位点突变的载体，明确miR-125a-5 p/455-3p/455-5p与SRB1的3’UTR发生直接结合，发挥促进SRB1降解的作用。通过研究我们发现前列腺癌肿瘤细胞中miRNA通过调节SRB1的表达水平，影响胆固醇酯的蓄积，进而影响肿瘤细胞的增殖和克隆形成能力。