Adenovirus type 5 EIA gene therapy for ovarian carcinoma

卵巢透明细胞癌（CCC）对铂类化疗耐药与预后不良有关，迫切需要有效的治疗方法。HER 2/neu在CCC中比在其他组织学类型的上皮性卵巢癌中更常上调。本研究的目的是评估抗HER 2抗体tanstuzumab或人腺病毒5型E1 A治疗卵巢CCC的可能性。我们用transtuzumab或E1 A处理了10个CCC细胞系，并评估了细胞活力、增殖和集落形成以及HER 2和野生型p53蛋白和这些信号通路下游分子的表达。通过蛋白质印迹法在所有10个细胞系中检测到不同水平的HER 2蛋白，通过免疫组织化学染色在5个CCC细胞系中检测到不同水平的HER 2蛋白：仅在一个细胞系（RMG-I）中检测到HER 2基因扩增（通过荧光原位杂交）。曲妥珠单抗在测试的四种CCC细胞系（RMG-1、SKOV-2、OVTOKO和OVSAYO）中均不抑制增殖。然而，无论HER 2表达水平如何，用E1 A转染（与对照载体相比）减少了所有10个CCC细胞系中的集落形成。用编码E1 A的腺病毒载体感染RMG-1和SMOV-2细胞导致显著（P<0.05）增殖抑制和细胞死亡增加;这种效应需要p53（而不是p73）蛋白的稳定，并且与Bax的上调和caspase 9的切割有关。其他机制，如p53非依赖性细胞凋亡，也可能参与E1 A介导的CCC细胞死亡。最后，用E1 A治疗延长CCC异种移植模型中的存活（P<0.001）。E1 A基因治疗，由于其稳定野生型p53的能力，是值得探索的治疗模式与卵巢CCC的妇女，通常表达野生型p53。

Resistance of ovarian clear cell carcinoma (CCC) to platinum-based chemotherapy is associated with poor prognosis, and an effective treatment for advanced disease is urgently needed. HER2/neu is up-regulated more often in CCC than in other histologic types of epithelial ovarian cancer. The purpose of this study was to assess possible treatment for ovarian CCC with the anti-HER2 antibody tanstuzumab or human adenovirus type 5 E1A. We treated 10 CCC cell lines with transtuzumab or E1A and assessed cell viability, proliferation, and colony formation and the expression of HER2 and wild-type p53 proteins and molecules downstream of those signaling pathways. HER2 protein was detected at various levels in all 10 cell lines by Western blotting and in 5 CCC cell lines by immunohistochemical staining: HER2 gene amplification was detected (by fluorescence in situ hybridization) in only one cell line (RMG-I). Trastuzumab did not inhibit proliferation in any of the four CCC cell lines tested (RMG-I, SKOV-2, OVTOKO, and OVSAYO). However, transfection with E1A (as compared with control vectors) reduced colony formation in all 10 CCC cell lines regardless of HER2 expression level. Infection of RMG-I and SMOV-2 cells with an adenoviral vector encoding E1A led to significant (P<0.05) suppression of proliferation and enhancement of cell death; this effect required stabilization of p53 (but not p73) protein and was associated with the up-regulation of Bax and the cleavage of caspase9. Other mechanisms, such as p53-independent apoptosis, may also be involved in E1A-mediated cell death in CCC. Finally, treatment with E1A prolonged survival in a CCC xenograft model (P<0.001). E1A gene therapy, because of its ability to stabilize wild-type p53, is worth exploring as a treatment modality for women with ovarian CCC, which typically expresses wild-type p53.