SCCA1 distal promoter for gene therapy of cervical intraepithelial neoplsia

宫颈上皮内瘤变(CIN)在这10-20年间呈上升趋势，特别是在15-30岁的年轻人群中，尽管晚期宫颈癌在这10-20年间有所减少。有强烈的证据表明，宫颈癌是由人乳头瘤病毒(HPV)感染引起的，90%的HPV患者会感染宫颈癌。目前治疗CIN的方法有光动力疗法(PDT)、激光疗法、角化疗法，但这些疗法在疗效和灭菌方面仍存在一些问题。因此，对于需要治疗不孕症的青壮年CIN应建立新的治疗方法。SCCA1基因最初是从宫颈癌组织中克隆出来的，在宫颈癌组织和正常宫颈鳞状组织中表达。本研究对SCCA1启动子进行了克隆和测序，并对其启动子活性进行了鉴定。为了确定组织特异性启动子的活性，我们建立了轻度不典型增生、中度不典型增生、重度不典型增生和CIS的细胞系。在浸润性宫颈癌中，SCCA1上游500kb转录起始点的启动子近端被激活；在CIN细胞系中，SCCA1上游转录起始点的3.7kb启动子远端被激活。将此远端启动子导入腺病毒表达P53，构建重组腺病毒SCCA1-3.7-P53，观察其对宫颈癌细胞系和CIN细胞系的生长抑制作用。腺病毒SCCA1-3.7-P53对宫颈上皮内瘤变(CIN)有明显的细胞凋亡抑制作用，但对宫颈浸润性癌无明显影响，对宫颈上皮内瘤变(CIN)有明显的抑制作用，但对宫颈浸润性癌无明显抑制作用。这些结果表明，远端启动子导入腺病毒载体(腺病毒-SCCA1-3.7-P53)具有治疗CIN的潜力。

Cervical intraepithelial neoplasia (CIN) has been increased for these 10-20 years especially in the young ages with 15-30 years, although advanced cervical cancer has been decreased for these 10-20 years. It has been strongly suggested that cervical cancer is developed by the infection of human papillomaivurs (HPV) and is infected in 90% of patients with HPV. CIN is treated by photodynamic therapy (PDT), laser treatment, cornization, but these treatment still nave some problems concerning treatment effect and steriligy. Therefore, a new method should be established for the treatment of CIN in young ages to be needed for sterility. SCCA1 gene was originally cloned from cervical cancer tissue and was expressed in cervical cancer tissue and normal squamous tissue in the cervix. In this study, we cloned and sequenced SCCA1 promoter, and characterized the promoter activity of SCCA1 promoter. To use for the determination of tissue specific promoter activity, we established cell lines of mild dysplasia, moderate dysplasia, severe dyplasia, and CIS. The proximal promoter of 500-bp upstream SCCA1 transcriptional start site was activated in invasive cervical cancer, and the distal promoter of 3.7-kb upstream SCCA1 transcriptional start site was activated in CIN cell lines. This distal promoter was introduced into adenoviru to express p53 to develop Adenovirus-SCCA1-3.7-p53 in order to determine growth inhibitory effect of this vector in invasive cervical cancer and CIN cell lines. Adenovirus-SCCA1-3.7-p53 induced strong apoptic changes especially in CIN but not in invasive cervical cancer, furthermore, showed strong growth inhibitory effect especially in CIN but not in invasive cervical cancer. From these results, it is suggested that distal promoter indroduced adenovirus vector (Adenovirus-SCCA1-3.7-p53) has the potential to treat CIN.