Pachyonychia congenita clinical trial using therapeutic self-delivery siRNAs

描述（由申请人提供）：RNA干扰（RNAi）具有彻底改变显性遗传疾病治疗的潜力。小抑制RNA（siRNA）是高度有效和选择性的，表现出显着的单核苷酸特异性。目前正在进行使用siRNA的临床试验，用于包括皮肤在内的许多适应症。在1期资金的支持下，对先天性甲肥厚（PC）进行了一项小型1例患者1b期临床试验。这种皮肤异常是由编码角蛋白6、16和17的基因突变引起的，包括单核苷酸变化。PC患者的主要抱怨是发生在足部压力点上或附近的使人衰弱的疼痛性老茧和起泡。通过病灶内注射siRNA（TD 101）靶向足底上的这些限定区域进行局部siRNA治疗，结果令人鼓舞。不幸的是，与注射到病变中相关的疼痛（需要口服止痛药和局部神经阻滞以允许治疗）阻止了这种给药模式的广泛使用。这一观察结果导致了大量的努力来确定患者友好的（即，很少或没有疼痛）递送选项。这项临床试验是siRNA在皮肤中的首次人类应用，也是第一个靶向突变基因的siRNA。 在本提案的第2阶段，我们延长了第1阶段和第1b阶段临床试验的进展。我们已经发现，未修饰的或稳定的siRNA不容易被皮肤角质形成细胞吸收，但修饰的所谓的“自递送”（sd）siRNA容易被皮肤角质形成细胞吸收。将在小鼠和人类皮肤模型中对sd-siRNA进行额外优化，然后合成GMP材料用于小鼠和兔毒理学研究和临床试验，其中siRNA将通过可溶解微针阵列或局部GeneCream制剂（均在TransDerm开发和生产）递送。GeneCream和微针阵列都被设计为有效地递送sd-siRNA，几乎没有疼痛或没有疼痛（微针突起被设计为仅穿透到非神经支配的表皮）。虽然PC是一种罕见的疾病，但这种疾病的性质使其成为第一次人体siRNA皮肤临床试验的理想原型皮肤疾病（在有限的，定义的区域中表达的定义突变），我们完全期望学到的教训将很容易推广到其他皮肤疾病。 公共卫生相关性：尽管发现了导致许多遗传性皮肤病的潜在突变，但很少有有效的治疗方法出现。siRNA可以靶向特定疾病相关突变的发现预示着一种新的治疗模式的出现。我们已经确定了一种有效的和选择性的siRNA，其靶向角蛋白6a基因中的单核苷酸突变，该突变导致罕见的先天性厚甲皮肤病并阻断其表达。这种siRNA（FDA称为TD 101）已在1b期临床试验中成功测试。尽管有疗效的迹象，但给药途径（大体积皮内注射）的疼痛性质要求我们开发一种对患者更友好的给药系统。该提案的第2阶段允许临床前研究，从而能够使用加载有TD 101 siRNA的自递送版本的微针进行第二次临床试验。应用siRNA技术治疗PC的经验教训应该很容易推广到其他主要皮肤疾病的主机，并可能导致突变或过度表达的致病基因的其他疾病。

DESCRIPTION (provided by applicant): RNA interference (RNAi) has the potential to revolutionize treatment of dominant genetic disorders. Small inhibitory RNAs (siRNAs) are highly potent and selective, demonstrating remarkable single-nucleotide specificity. Clinical trials using siRNAs are currently underway for a number of indications including skin. Facilitated by Phase 1 funding, a small one patient phase 1b clinical trial was undertaken for pachyonychia congenita (PC). This ultrarare skin disorder is caused by mutations, including single nucleotide changes, in the genes encoding keratins 6, 16, and 17. The major complaint of PC patients is the debilitating painful callusing and blistering which occurs on or near the pressure points of the feet. These defined regions on the soles of the feet were targeted for local siRNA treatment by intralesional injection of siRNA (TD101) with encouraging results. Unfortunately, the pain associated with injections into lesions (oral pain medication and regional nerve blocks were required to allow treatment), prevents widespread use of this mode of administration. This observation has led to intense efforts to identify patient-friendly (i.e., little or no pain) delivery options. This clinical trial was the first human use of siRNA in skin and also the first siRNA to target a mutated gene. In Phase 2 of this proposal, we extend the progress of Phase 1 and the Phase 1b clinical trial. We have found that unmodified or stabilized siRNAs are not taken up readily by skin keratinocytes but that modified, so-called "self-delivery" (sd) siRNAs are. Additional optimization of the sd-siRNA will be undertaken in mouse and human skin models followed by synthesis of GMP material for mouse and rabbit toxicology studies and clinical trials in which the siRNA will be delivered by dissolvable microneedle arrays or a topical GeneCream" formulation, both developed and manufactured at TransDerm. GeneCream and microneedle arrays are both designed to effectively deliver sd-siRNA with little or no pain (microneedle protrusions are designed to only penetrate to the non-innervated epidermis). Although PC is a rare disease, the nature of the disorder makes it an ideal prototype skin disorder (defined mutations with expression in limited, defined areas) for first-in-man siRNA skin clinical trials, and we fully expect the lessons learned will be readily generalized to other skin disorders. PUBLIC HEALTH RELEVANCE: Despite the discovery of the underlying mutations responsible for many inherited skin diseases, few effective treatments have emerged. The discovery that siRNAs can be developed to target specific disease-related mutations portends the advent of a new treatment paradigm. We have identified a potent and selective siRNA that targets a single nucleotide mutation in the keratin 6a gene that is responsible for the rare skin disorder pachyonychia congenita and blocks its expression. This siRNA (known to the FDA as TD101) was successfully tested in a Phase 1b clinical trial. Despite signs of efficacy, the painful nature of the administration route (intradermal injection of large volumes) required that we develop a more patient-friendly delivery system. Phase 2 of this proposal allows preclinical studies enabling a second clinical trial using microneedles loaded with a self-delivery version of TD101 siRNA. The lessons learned in applying siRNA technology for treatment of PC should be readily generalizable to a host of other dominant skin disorders and likely to other disorders resulting from mutated or overexpressed disease-causing genes.