SCCA1 distal promoter for gene therapy of cervical intraepithelial neoplsia

宫颈上皮内瘤变（CIN）在这10-20年间有所增加，特别是在15-30岁的年轻人中，尽管晚期宫颈癌在这10-20年间有所减少。人们强烈认为宫颈癌是由人乳头瘤病毒（HPV）感染引起的，90%的人乳头瘤病毒患者会感染宫颈癌。目前治疗CIN的方法有光动力疗法（PDT）、激光治疗、角膜化治疗，但这些治疗方法在治疗效果和无菌性方面还存在一些问题。因此，应该建立一种新的方法来治疗年轻时的CIN，以满足不育的需要。SCCA1基因最初是从宫颈癌组织中克隆出来的，在宫颈癌组织和宫颈正常鳞状组织中均有表达。在本研究中，我们克隆并测序了SCCA1启动子，并对SCCA1启动子的活性进行了表征。为了测定组织特异性启动子活性，我们建立了轻度发育不良、中度发育不良、重度发育不良和CIS细胞系。侵袭性宫颈癌中500 bp上游SCCA1转录起始位点近端启动子被激活，CIN细胞系中3.7 kb上游SCCA1转录起始位点远端启动子被激活。将该远端启动子导入腺病毒中表达p53，制备腺病毒- scca1 -3.7-p53，以确定该载体对侵袭性宫颈癌和CIN细胞株的生长抑制作用。腺病毒- scca1 -3.7-p53可诱导强烈的细胞凋亡变化，尤其是在CIN中，而在浸润性宫颈癌中没有；此外，腺病毒- scca1 -3.7-p53具有强烈的生长抑制作用，特别是在CIN中，而在浸润性宫颈癌中没有。这些结果提示，远端启动子诱导的腺病毒载体（adenovirus - 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.