HOMOLOGOUS RECOMBINATION IN SKIN CELLS FOR GENE THERAPY

描述：(改编自申请人的摘要)-皮肤细胞是 将新基因引入人类的最有潜力的工具之一 用于局部疾病或全身疾病的基因治疗 代谢性疾病。到目前为止的工作已经确定了 用于表达和传递的表皮角质形成细胞和成纤维细胞 局部和系统的新基因，但技术和安全问题 对当前采用的引入新技术的方法提出质疑 基因进入这些细胞。这项研究的目标是证明 利用同源重组技术将新基因插入到 表皮角质形成细胞和成纤维细胞中的特异性基因座。这 方法应提供可预测、持久且相对安全的 新基因的表达。这个项目的具体目的是：i) 在β-肌动蛋白中插入一个标记基因，细菌酶lacZ。 培养的人角质形成细胞和成纤维细胞的位置或进入 角质形成细胞角蛋白5基因座和II)筛选靶向克隆 并评估LacZ在体外和体内表达的持续时间。 体内，将靶细胞移植到无瘤小鼠体内。

DESCRIPTION: (Adapted from the applicant's abstract) - Skin cells are among the best potential vehicles for introducing new genes into humans for the purpose of gene therapy for local disease or for systemic metabolic disease. Work to date has established the usefulness of epidermal keratinocytes and fibroblasts for expressing and delivering new genes locally and systemically, but technical and safety issues raise questions about currently employed methods used to introduce new genes into those cells. The goal of this research is to demonstrate the feasibility of using homologous recombination to insert new genes into specific gene loci in epidermal keratinocytes and fibroblasts. This method should provide predictable, durable, and relatively safe expression of new genes. The specific aims of this project are I) to insert a marker gene, the bacterial enzyme lacZ, into the beta-actin locus of cultured human keratinocytes and fibroblasts or into the keratin 5 locus of keratinocytes and II) to select clones of targeted cells and to evaluate the duration of lacZ expression in vitro and in vivo, following transplantation of the targeted cells into athymic mice.