DJ-1在胃癌多药耐药中的作用及其分子机制研究

Multidrug resistance (MDR) is usually one of the main causes for failure of chemotherapy against malignant tumors, including gastric cancer. At present, the molecular mechanisms underlying MDR of gastric cancer have not been fully elucidated, indicating that other unknown molecules or pathways may be involved in the development of MDR of gastric cancer. DJ-1 is a novel mitogen-dependent oncogene product. Our previous work found that the levels of DJ-1 expression were significantly higher in multidrug resistant gastric cancer cell lines than in its parental cells. Additionally, we found that DJ-1 knockdown significantly increased the sensitivity of multidrug resistant gastric cancer cell lines to multiple chemotherapy drubs. These results indicated that DJ-1 might be closely related to multidrug resistance in gastric cancer cells. However, whether DJ-1 is directly involved in the development of MDR of gastric cancer remains to be further clarified. Therefore, the current investigation will be undertaken to elucidate the role of DJ-1 in gastric cancer MDR in multi-levels by gene transfection, RNA interference, MTT assay, histoculture drug response assay (HDRA), and nude tumorigenesis model. Moreover, this study will further explore the molecular regulation mechanisms by which DJ-1 promotes gastric cancer MDR from the perspective of PI3K/Akt-Nrf2-ARE signaling pathways and the change of MRP1, GST, NQO1, UGT, and HO-1 expression. This study will provide novel target and new strategies for reversing the MDR of gastric cancer.

多药耐药是胃癌化疗失败的主要原因，其分子机制仍未阐明。DJ-l蛋白是一种丝裂原依赖性癌基因产物。我们前期研究发现DJ-1在多药耐药胃癌细胞株中表达明显高于其敏感亲本胃癌细胞株，同时发现下调DJ-1表达能显著提高多药耐药胃癌细胞对化疗药物的敏感性，提示DJ-1可能与胃癌多药耐药密切相关，但DJ-1是否直接参与胃癌多药耐药及其相应调控机制有待于进一步明确，为此，本研究拟采用基因转染、RNA干扰、MTT、组织培养药物敏感实验以及裸鼠体内成瘤等实验，进一步从体外、体内、组织水平多途径、多层次验证DJ-1在胃癌多药耐药中的作用。同时，从PI3K/Akt-Nrf2-ARE信号通路及其调控的MRP1、GST、NQO1、UGT及HO-1等多药耐药相关蛋白表达改变的角度进一步探讨DJ-1参与胃癌多药耐药的分子机制。该研究的顺利完成将为阐明胃癌多药耐药本质提供新思路和新方向，为逆转多药耐药提供新靶点和新基础。

胃癌是我国发病率和死亡率较高的恶性肿瘤之一。多药耐药（MDR）的发生是导致胃癌化疗失败的主要原因。因此，深入探索胃癌MDR发生的分子机制，确立胃癌MDR防治靶点对临床上逆转胃癌MDR、提高胃癌疗效具有重要意义。DJ-l蛋白是一种丝裂原依赖性癌基因产物，与肿瘤密切相关。本研究探讨了DJ-1在胃癌MDR中的作用及其分子机制。结果发现，与亲本的SGC7901胃癌细胞相比，多药耐药的SGC7901/VCR胃癌细胞中DJ-1表达上调，化疗药物的IC50值、克隆形成率及阿霉素泵出率均明显增加，而G0/G1比值、细胞凋亡率，细胞移植瘤化疗后的生长抑制率均显著降低。有趣的是，上述效应在DJ-1沉默的SGC7901/VCR/shDJ-1胃癌细胞中被逆转，而在DJ-1过表达的SGC7901/DJ-1胃癌细胞中被重现。这些结果表明DJ-1参与了胃癌MDR的发生。在探讨DJ-1促胃癌MDR的分子机制中，我们发现，与SGC7901胃癌细胞相比，SGC7901/VCR胃癌细胞中DJ-1与PTEN蛋白相互作用增加，PTEN酶活性下降，Akt磷酸化增加，此外，Nrf2磷酸化及其入核增加，同时，P-gp、Bcl-2、GST、NQO1及HO-1表达上调。同样，上述效应在SGC7901/VCR/shDJ-1胃癌细胞中被逆转，而在SGC7901/DJ-1胃癌细胞中被重现。更为重要的是，我们进一步发现，SGC7901/VCR及SGC7901/DJ-1胃癌细胞经PI3K抑制剂LY294002预处理或pLVX-shNrf2慢病毒感染后，Nrf2核转位、P-gp, Bcl-2, GST, NQO1及HO-1表达以及细胞多药耐药性均明显降低；而对DJ-1表达，DJ-1与PTEN相互作用及PTEN酶活性均无明显影响。此外，上述细胞经LY294002预处理后Akt磷酸化及Nrf2磷酸化明显被抑制；而pLVX-shNrf2慢病毒感染后仅抑制Nrf2磷酸化但对Akt磷酸化无影响。这些结果在胃癌细胞裸鼠移植瘤模型上也得以证实。总之，上述结果表明DJ-1促胃癌MDR的分子机制可能与其负调控PTEN活性间接激活PI3K/Akt/Nrf2信号通路，进而上调P-gp, Bcl-2, GST, NQO1及HO-1的表达有关。本研究为逆转胃癌MDR提供了新靶点和新的干预环节。本研究按计划全面完成，发表学术论文8篇，并培养硕士研究生3名。