SELECTIVE MANIPULATION OF GENE EXPRESSION IN SKIN

一个合作项目将使用单链合成DNA 与靶向的DNA调节因子形成三链体结构的寡核苷酸 区域（TFO）来测试它们可以被用作 皮肤细胞功能障碍的基因特异性疗法。皮肤是理想的模型 基因操作，因为它可以很容易地评估和生长在 体外牛皮癣和皮肤癌将成为选择性 操纵在第一个项目中，合成寡核苷酸类似物 将被设计为靶向表皮生长的启动子结构域， 因子受体基因（EGF-R），并测试其选择性 抑制EGF-R在A431表皮癌细胞系中的表达， EGF-R。由于银屑病过度表达EGF-R以及TGF-α， 然后，我们将研究寡核苷酸的能力， 体外效应，它们下调银屑病患者EGF-R能力 病变在第二个项目中，我们将尝试操纵 谷胱甘肽-S-转移酶pi，GST-pi的表达， 在黑色素瘤和鳞状细胞癌中过表达， 导致多药耐药性。将测试TFO的容量 选择性抑制GST-π首先在黑色素瘤细胞系，随后， 转移性黑色素瘤至植入裸鼠的皮肤。在这两种模型中，皮肤 将在应用寡核苷酸后除去， 通过斑点杂交、北方印迹和原位杂交检测mRNA表达 杂交方法这些研究可能会导致一类新的 治疗药物，并提供深入了解银屑病的发病机制 和癌症的抗药性。

A collaborative project will use single stranded synthetic DNA oligonucleotides which form triplex structures with targeted DNA regulatory regions (TFOs) to test the hypothesis that they may be used as gene-specific therapy for skin cell dysfunction. The skin is an ideal model for genetic manipulation because it can be readily evaluated and grown in vitro. Psoriasis and skin cancer will be targeted for selective manipulation. In the first project, synthetic oligonucleotide analogues will be designed to target the promoter domain of the epidermal growth factor receptor gene (EGF-R) and tested for their capacity to selectively repress EGF-R in the A431 epidermal carcinoma line which overexpressed EGF-R. Since psoriasis overexpresses EGF-R as well as TGF-alpha which binds to that receptor, we will then study the ability of oligonucleotides with in vitro effect, for their capacity to downregulate EGF-R in psoriatic lesions. In the second project, we will attempt to manipulate the expression of Glutathione-S-transferase pi, GST-pi, an enzyme which is overexpressed in melanoma and squamous cell carcinoma and which may contribute to multidrug resistance. TFOs will be tested for their capacity to selectively repress GST-pi first in melanoma lines and subsequently, in metastatic melanoma to skin implanted into nude mice. In both models, skin will be removed following application of the oligonucleotides and assayed for mRNA expression by dot hybridization, Northern blotting, and in situ hybridization. These studies may lead to the development of a new class of therapeutic agents and provide insight into the pathogenesis of psoriasis and drug resistance in cancer.