Smooth Muscle Myosin PROTACs as a Novel Treatment for Asthma

项目总结/摘要 在2007年NEJM的一篇社论中，我们提出，消耗气道平滑肌的肌球蛋白-收缩性肌球蛋白- 负责肌细胞收缩的tor蛋白-将是预防急性哮喘的有效策略 攻击，但没有药物策略来实现这一点。然而，一种聪明的新化合物， 蛋白水解靶向嵌合体（PROTAC）已经开发出来，可以实现这一目标。这些小， 双功能分子通过刺激靶蛋白的泛素化来催化靶蛋白的蛋白酶体降解。 每个PROTAC含有一个与靶蛋白结合的部分，并通过接头连接至靶蛋白。 结合E3连接酶的第二"诱饵"部分;当两侧都结合时，使E3连接酶靠近第二"诱饵"部分。 靶向蛋白，它泛素化，因此目标降解。该过程是催化性的，因为 PROTAC在靶降解后释放，可用于另一个靶结合循环，E3连接酶再结合。 cruiting，and target目标ubiquitination泛素化.在这里，我们建议测试新的想法，一个PROTAC降解 气道平滑肌细胞内的平滑肌肌球蛋白可作为一种新的治疗方法， 哮喘平滑肌肌球蛋白具有较长的半衰期和缓慢的周转率，因此似乎非常适合于 PROTAC降解。消耗气道肌细胞的肌球蛋白会严重损害收缩， 靶向平滑肌肌球蛋白的PROTACs（SMM-PROTACs）应防止早期急性气道狭窄。 有哮喘的人。在这个R21提案中，我们将进行旨在证明 SMM-PROTAC可以通过执行四个特定目标来抑制支气管收缩： 1)合成并表征一小系列原型SMM-PROTAC 2)证明SMM-PROTAC降解人气道平滑肌中的平滑肌肌球蛋白， 但不降解心脏、骨骼或非肌肉肌球蛋白 3)证明SMM-PROTAC抑制人肺切片中的气道收缩 4)证明SMM-PROTAC被递送到气道钝乙酰甲胆碱（MCh）诱导的Brone- 小鼠睾丸收缩 证明SMM-PROTAC可通过选择性刺激 平滑肌肌球蛋白将证明其作为新型哮喘治疗的进一步临床前开发是合理的。这 这一发现也将证明评价SMM-PROTAC是否抑制其他疾病的进展是合理的， 平滑肌收缩起关键作用。

PROJECT SUMMARY/ABSTRACT In a 2007 editorial in NEJM, we proposed that depleting airway smooth muscle of myosin – the contractile mo- tor protein responsible for myocyte contraction – would be an effective strategy for preventing acute asthma attacks, but no drug strategy to accomplish this was available. However, a clever new class of compounds, Proteolysis Targeting Chimeras (PROTACs), has been developed that could achieve this goal. These small, bifunctional molecules catalyze proteasome degradation of a targeted protein by stimulating its ubiquitination. Each PROTAC contains one moiety that binds to the targeted protein, and is connected through a linker to a second “bait” moiety that binds an E3 ligase; when both sides are bound, the E3 ligase is brought close to the targeted protein, which it ubiquitinates and so targets for degradation. The process is catalytic because the PROTAC is released upon target degradation and is available for another cycle of target binding, E3 ligase re- cruitment, and target ubiquitination. Here, we propose to test the new idea that a PROTAC to degrade smooth muscle myosin within airway smooth muscle cells could be used as a new treatment for asthma. Smooth muscle myosin has a long half-life and slow turnover rate and so seems ideally suited for degradation by PROTACs. Depleting airway myocytes of myosin would drastically impair contraction, and PROTACs targeting smooth muscle myosin (SMM-PROTACs) should prevent acute airway constriction in eve- ryone with asthma. In this R21 proposal, we will pursue proof-of-principle experiments aimed at demonstrating SMM-PROTACs can inhibit bronchoconstriction, through the execution of four specific aims: 1) Synthesize and characterize a small series of prototype SMM-PROTACs 2) Demonstrate that SMM-PROTACs degrade smooth muscle myosin in human airway smooth muscle, but do not degrade cardiac, skeletal, or non-muscle myosins 3) Demonstrate that SMM-PROTACs inhibit constriction of airways in human lung slices 4) Demonstrate that SMM-PROTACs delivered into the airways blunt methacholine (MCh)-induced bron- choconstriction in mice Demonstration that SMM-PROTACs can inhibit bronchoconstriction by selectively stimulating degradation of smooth muscle myosin would justify their further preclinical development as a novel asthma treatment. This finding would also justify evaluation of whether SMM-PROTACs inhibit progression of other diseases in which smooth muscle contraction plays a key role.