靶向抑制BTAK基因对卵巢癌细胞化疗敏感性的影响及机制研究

The mortality of ovarian cancer is leading the first place of female gynaecological malignances because of its biological behavior of dissemination and metastasis as well as lack of sensitivity during chemotherapy. The mechanisms of chemosensitivity are not only manifold, but involute. Generally, inhibiting proliferation and/or inducing apoptosis is thought as the common final way of most drugs. We previously reported that the expression of BTAK was frequently up-regulated in human ovarian cancer tissues, and there was close correlation between the expression of BTAK and p53. So it might be a potential target for gene therapy. Some researches have found that BTAK expression was associated with poor prognosis and resistance to chemotherapy. In our study, we select BTAK mRNA as one target, try to down-regulate the expression of BTAK in ovarian cancer cell lines by RNA interference. Then we can investigate the effect of targeting BTAK suppression on proliferation, apotosis and cell cycle changes in vitro. Next chemosensitivity against paclitaxel can be investigated by MTT assay and cell determination in vitro and the status of tumor formation in vivo. Our study will lay the experimental foundation for ovarian cancer therapy targeting on BTAK, and also may provide a new clue to the combination of chemotherapy and gene therapy for ovarian cancer patients.

卵巢癌死亡率位居女性生殖道恶性肿瘤之首，主要原因是卵巢癌易于转移播散，且治疗中容易出现耐药性。其耐药机制多重复杂，但一般认为抑增殖和/或促凋亡是大多数化疗药物作用的最终途径。我们的前期工作证实BTAK基因在卵巢癌中表达升高，且与p53高表达密切相关，可能是潜在的治疗靶点；并且研究表明BTAK基因可能与卵巢癌的预后不良及化疗耐药有关。因此，本研究拟通过RNA干扰特异性下调BTAK基因在卵巢癌细胞的表达，分别从细胞水平及动物水平深入探讨靶向抑制BTAK基因对卵巢癌细胞增殖、凋亡、细胞周期的影响以及对紫杉醇化疗敏感性的影响，为探索以BTAK基因为干预靶点的卵巢癌治疗奠定研究基础，同时为卵巢癌的基因治疗联合化疗提供一种新思路。

根据项目研究计划，对BTAK蛋白在卵巢细胞系及石蜡标本组织芯片中的表达水平进行了检测，结果发现携带BRCA杂合性突变的卵巢和卵巢癌细胞中BTAK的表达显著高于正常卵巢细胞，交界性及恶性肿瘤中的表达亦高于良性肿瘤及正常卵巢组织，但BTAK表达水平与肿瘤恶性程度及组织学类型未见显著相关性，提示BTAK基因可能在卵巢癌的早期阶段发挥作用，其表达水平升高可能是早期卵巢癌发生的信号。本项目成功构建了重组逆转录病毒表达载体并转染SKOV-3和OVCA432细胞，从而建立了稳定抑制BTAK基因的卵巢癌细胞株，结果发现BTAK基因靶向抑制后，其蛋白表达显著下调、细胞生长速度明显变慢，且在一定程度上能增强肿瘤细胞对紫杉醇的药物敏感性。另外鸡胚绒毛尿囊膜(CAM)模型和裸鼠体内成瘤实验均显示，靶向抑制BTAK能够显著抑制卵巢癌细胞诱导肿瘤形成，免疫组化染色结果表明，BTAK低表达的肿瘤组织中新生血管的数量及ki-67的表达也降低，其促肿瘤增殖及促血管生成的能力亦受到显著抑制。综合以上结果，推测靶向抑制BTAK基因可以抑制肿瘤增殖及血管新生，在一定程度上协同增强肿瘤细胞对紫杉醇的药物敏感性，有望成为临床治疗卵巢癌的一个潜在的作用靶点。本项目研究结果目前发表SCI论文1篇，国内核心期刊1篇，协助培养硕士研究生1名。