Targeted protein stabilization using Protein Rescue Targeting Chimeras (PRESTACs)

以泛素途径为靶点是治疗各种破坏性疾病的潜在选择，包括 癌症和神经退行性变。针对嵌合体的异双功能蛋白水解酶的最新研究进展， (PROTACs)，它将E3连接酶招募到感兴趣的靶标，以泛素化和降解靶标，是一种 前景看好的治疗途径。PROTAC已经被开发用于降解广泛的蛋白质家族，并且 新的目标正在不断地被调查。在细胞中，连接酶对泛素的结合可被去连接酶逆转。 共轭蛋白水解酶(DUBS)，这两类酶的动态调节蛋白质周转， 隔间和其他过程。一些细胞病理是由密钥的过度降解引起的 蛋白质，例如，肿瘤抑制因子，病理状态可以通过减弱降解来纠正， 从而节省了关键蛋白质。从形式上讲，DUB可以被一种异双功能分子招募来 使目标蛋白去泛素化并防止其降解，因此假设 被称为针对嵌合体的蛋白质救援(PRESTAC)的异双功能分子可以通过以下方式产生 将配基与靶配基化学连接，并对其生理功能进行评估。普罗根拉有 鉴定和表征了与USP7结合并激活其DUB功能的新型小分子USP7配体。 这些分子，连同感兴趣靶标的配体，将被用于设计和合成前STAC， 这将代表DUB活动与疾病相关靶点的首次联系，并提供了一种新的工具 扩展基于UPS的疗法。作为概念的证明，双功能分子将被合成来招募 USP7以稳定核受体PPAR，已知有几种激动剂。噻唑烷二酮类 针对PPAR的TZD通过诱导降低胰岛素抵抗的基因来治疗2型糖尿病。配基结合 TZD到PPAR不仅调控转录，而且导致PPAR泛素化和蛋白酶体 退化。因此，稳定PPAR将延长其激活时间，并增强这些药物的作用。 PRESTAC将使用Progenra的USP7激活剂和罗格列酮设计，罗格列酮是一种与TZD相互作用的药物 并且对PPAR的影响也得到了很好的表征。PRESTAC候选人将接受生化评估， 生物物理上，以及在NIH3T3细胞中形成三元复合体(USP7-PRESTAC-PPAR)的能力，以 稳定PPAR，并影响PPAR激活的下游靶点(例如，PPAR报告分析， 脂联素水平)。

Targeting the ubiquitin pathway is a potential option for treating a variety of devastating diseases, including cancers and neurodegeneration. Recent focus on heterobifunctional proteolysis targeting chimeras, (PROTACs), which recruit E3 ligases to targets of interest for ubiquitination and degradation of the target, is a promising therapeutic avenue. PROTACs have been developed to degrade a wide range of protein families, and new targets are continually being investigated. In cells, ubiquitin conjugation by ligases is reversed by de- conjugating proteases (DUBs), and the dynamics of these two enzyme classes regulates protein turnover, compartmentation, and other processes. Some cell pathologies are driven by excessive degradation of a key protein, for example, a tumor suppressor, and the pathological state can be corrected by attenuating degradation, thereby sparing the critical protein. Formally, DUBs could be recruited by a heterobifunctional molecule to deubiquitinate target proteins and prevent their degradation, and it is therefore hypothesized that heterobifunctional molecules called Protein Rescue Targeting Chimeras (PRESTACs) can be generated by chemically linking a DUB ligand to a target ligand and evaluated for their physiological function. Progenra has identified and characterized novel small molecule USP7 ligands that bind USP7 and activate its DUB function. These molecules, along with ligands for targets of interest, will be used to design and synthesize PRESTACs, which will represent the first tethering of DUB activity to disease-relevant targets and provide a novel tool for expansion of UPS-based therapies. As proof of concept, bifunctional molecules will be synthesized to recruit the DUB USP7 to stabilize the nuclear receptor PPAR, for which several agonists are known. Thiazolidinediones (TZDs) targeting PPAR treat type 2 diabetes by inducing genes that decrease insulin resistance. Ligand binding of TZDs to PPAR not only regulates transcription, but also leads to PPAR ubiquitination and proteasomal degradation. Thus, stabilizing PPAR will prolong its activation and augment the effects of these drugs. PRESTACs will be designed using Progenra’s USP7 activators and rosiglitazone, a TZD whose interaction with and effects on PPAR are well-characterized. PRESTAC candidates will be evaluated biochemically, biophysically, and in NIH 3T3 cells for their ability to form ternary complexes (USP7-PRESTAC-PPAR), to stabilize PPAR, and to affect downstream targets of PPAR activation (for example, PPAR reporter assays, levels of adiponectin).