siRNA Inhibition of Keloid Fibrosis in Fibrin Matrix Skin Equivalent Mouse Models

描述(申请人提供)：瘢痕疙瘩是一种旺盛的肿瘤样皮肤疤痕，在10%至20%的深色皮肤种族，即非洲人、西班牙人和亚洲人后裔的皮肤受伤后不可预测地发生。目前还没有有效的治疗方法来预防或控制瘢痕疙瘩病变的生长和扩大，这对受影响个人的心理社会和整体健康状况是严重的。该提案的目标是通过建立基因靶向和纤维蛋白基质皮肤相当的瘢痕疙瘩纤维化小鼠模型，并评估RNA干扰治疗方法，来弥合瘢痕疙瘩研究中的根本障碍。该计划是基于Tuan博士的长期研究努力和Tuan博士和Benya博士合作(多个/共PI)在瘢痕疙瘩发病机制方面的最新进展。目前尚无可用于常规检测或准确反映人体损伤的瘢痕疙瘩纤维化动物模型。此外，为了开发治疗瘢痕疙瘩的基于知识的疗法及其临床应用，候选基因必须在动物模型中进行活体验证，并在此类模型中证明对靶向治疗有反应。在这项申请中提出了三个具体目标来解决这些障碍。特定目标的完成将提供一种强大的、基因靶向的、瘢痕疙瘩来源的皮肤等效动物模型，该模型基于模拟临时伤口基质的三维纤维蛋白凝胶。此外，我们的靶基因PAI-1在瘢痕疙瘩成纤维细胞中过表达，并已被确认为在体外正常和瘢痕疙瘩成纤维细胞中胶原堆积的原因(发表在2008年11月的Am J Pathol上)，将在体内被证实为瘢痕疙瘩纤维化的重要介质，同时展示了RNA干扰治疗方法的有效性。最后，在模型中对正常人成纤维细胞系进行基因操作以模拟瘢痕疙瘩纤维化，将在疗效评估中提供瘢痕疙瘩成纤维细胞的替代品，并绕过用于常规检测的有限的瘢痕疙瘩组织供应。我们还期望这些结果和工具将提高瘢痕疙瘩发病机制研究的发现率，此外，还将为其他研究人员提供一个易于修改的平台，以验证与瘢痕疙瘩纤维化相关的靶基因，并测试其他新的治疗方法。

DESCRIPTION (provided by applicant): Keloids are exuberant, tumor-like skin scars that occur unpredictably after injury to the skin in 10 to 20 % of dark skin races, i.e., people of African, Hispanic, and Asian descents. Currently there is no effective treatment to prevent or control the growth and expansion of keloid lesions, which are grievous to the psycho-social and overall health state of the affected individual. The goal of the proposal is to bridge the fundamental obstacles in keloid research by creating gene-targeted and fibrin matrix skin equivalent mouse models of keloid fibrosis, and assessing a RNA interference therapeutic approach. The plan is based on the long-term research effort of Dr. Tuan and the recent progress in keloid pathogenesis from the collaborative effort of Drs. Tuan and Benya (Multiple/Co-PIs). No animal models of keloid fibrosis currently exist that are applicable to routine testing or accurately reflect the human lesion. Moreover, for the development of knowledge-based therapies and their clinical application for keloids, candidate genes must be validated in vivo in animal models and shown to be responsive to targeted therapeutic treatments in such models. Three specific aims are proposed in this application to address the obstacles. Completion of the specific aims will provide a robust, gene-targeted, keloid-derived, skin equivalent animal model based on a three-dimensional fibrin gel mimic of provisional wound matrix. In addition, our target gene PAI-1, which is overexpressed by keloid fibroblasts and has been identified as causative in collagen accumulation in normal and keloid fibroblasts in vitro (published in Am J Pathol., Nov 2008), will have been validated in vivo as an important mediator of keloid fibrosis, while simultaneously demonstrating efficacy of an RNA interference therapeutic approach. Finally, the genetic manipulation of a normal human fibroblast cell line to mimic keloid fibrosis in the model will provide a substitute for keloid fibroblasts in efficacy evaluation and circumvent the limited supply of keloid tissues for routine testing. We also expect that these results and tools will enhance the rate of discovery research in keloid pathogenesis, furthermore, provide a platform, easily modified, for other investigators to validate target genes of interest in keloid fibrosis and test additional novel therapeutic approaches.