APPROACHES TO CORRECT DISEASES WITH GENETICALLY MODIFIED CELLS OF HUMAN SKIN

对于许多遗传疾病，异常基因和正常基因一样， 已被定义，克隆，并在体外表达。 分子生物学的这些进展 遗传学使我们有可能纠正遗传疾病， 通过体细胞（体细胞）将非缺陷基因导入患病宿主 基因疗法）。 这种方法似乎特别适合于 后天畸形和异常遗传性皮肤病 形态发生，例如，Darier病，着色性干皮病，表皮病 大疱，以及那些以缺乏 血浆蛋白，例如，血友病、糖尿病等。一个中心问题， 现在处于生物医学界的前沿，专注于如何最好地 将这些无缺陷的基因导入患病个体。 是我们 通过基因修饰皮肤细胞的矫正， 风险相对较低，应用广泛，因此具有相当大的 临床应用 我们相信， 方法的适用性可以回答临床前移植 遗传修饰的人皮肤等同物或其组分， 无胸腺啮齿类动物，并通过实验分析这种无胸腺啮齿类动物的选定参数， 皮肤作为时间的函数。 具体目标： 1.临床前基因移植技术的发展 改造皮肤给人类 a.将现有的人类皮肤表皮移植到无胸腺 啮齿类动物的表皮是由转基因的 角质形成细胞 B.产生一种基因修饰的皮肤等同物， 可移植的遗传修饰的成纤维细胞或角质形成细胞 无胸腺啮齿动物身上 C.产生浸渍有基因修饰的真皮基质， 成纤维细胞，可以放置在啮齿动物皮下， 皮内注射。 2.为了确定含有这些物质的皮肤的长期功能， 转基因成分;特别是剂量的能力 反应，转染基因的产生和稳定性，以及 转染对皮肤固有结构和功能的影响。 3.以确定是否隐性X连锁鱼鳞病，可以纠正 用类固醇基因转染的角质形成细胞或成纤维细胞 硫酸酯酶，并移植到无胸腺啮齿动物。 这些实验 旨在定义这项技术的潜力和局限性。

For many genetic disorders, the abnormal, as well as the normal gene have been defined, cloned, and expressed in vitro. These advances in molecular genetics have made it possible to correct genetic disease by bringing the non-defective gene into the afflicted host via somatic cells (somatic cell gene therapy). This approach appears to be particularly suitable to heritable skin diseases of acquired malformation and abnormal morphogenesis, e.g., Darier's disease, xeroderma pigmentosa, epidermolysis bullosa, and those disorders which are characterized by a deficiency of a plasma protein, e.g., hemophilia, diabetes mellitus, etc. A central issue, now at the forefront of the biomedical community, focuses on how best to introduce these non-defective genes into afflicted individuals. It is our thesis that correction by genetically modifying cells of the skin has relatively low risk, has broad application and thus has considerable clinical utility. We are persuaded that the feasibility, limitations and methods for applicability can be answered preclinically by transplanting genetically modified human skin equivalents or components thereof to athymic rodents and by experimentally analyzing selected parameters of such skin as a function of time. Specific aims: 1. Development of the preclinical technology for transplanting genetically modified skin to humans. a. Replacing the existing epidermis of human skin grafted onto athymic rodents with an epidermis generated from genetically modified keratinocytes. b. Generating a genetically modified skin-equivalent carrying normal or genetically modified fibroblasts or keratinocytes which can be transplanted onto athymic rodents. c. Generating a dermal matrix impregnated with genetically modified fibroblasts that can be placed subcutaneously in rodents, and possibly intradermally in humans. 2. To determine the long-term functionality of skin containing these genetically modified components; specifically the capacity for a dose response, the production by and stability of the transfected gene, and the effect of transfection on structure and function inherent to skin. 3. To determine whether recessive X-linked ichthyosis, can be corrected with keratinocytes or fibroblasts transfected with the gene for steroid sulfatase, and transplanted to athymic rodents. These experiments are designed to define the potential and the limitations of this technology.