MicroRNA to decrease vascular CaV1.2 in hypertension

说明（由申请人提供）：有许多短效药物可用于治疗高血压。然而，大多数患者不能坚持每日多药降压方案，副作用频繁。因此，只有少数高血压患者的血压恢复正常，只有长期的、有针对性的、副作用较小的降压治疗才能从根本上影响西方世界高血压的流行水平。本研究旨在开发一种新的、长期的高血压治疗方法，通过使用含有增强型平滑肌特异性启动子（EnSM221）的腺相关病毒（AAV）载体，传递外源性microRNA，靶向血管l型（CaV1.2）钙通道。值得注意的是，CaV1.2通道已经成为临床用于降血压的钙通道阻断药物的靶点。然而，我们的策略将使用aav介导的CaV1.2通道microRNA的递送来实现钙通道表达的长期和血管特异性敲低。我们的目标包括：1)证明CaV1.2通道microRNA能够在高血压小鼠中至少8周内无严重副作用地降低血压，2)在体内和体外证实在肠系膜小动脉中CaV1.2通道的血管收缩功能受到抑制，3)证明血管CaV1.2通道表达，全细胞Ca2+电流，肠膜动脉细胞K+通道表达和静息膜电位的恢复与AAV向血管传递CaV1.2通道microRNA的降压作用相关。本研究结果将为利用靶向AAV和microRNA技术治疗短期治疗难以控制的全身性高血压等慢性血管异常奠定基础。

DESCRIPTION (provided by applicant): There are many short-acting drugs available to treat hypertension. However, most patients fail to adhere to a daily, multi-drug antihypertensive regimen with frequent side effects. Thus, normal blood pressure is restored in only a minority of hypertensive patients, and only long-term, targeted antihypertensive therapies with fewer side effects will fundamentally impact the epidemic levels of hypertension in the Western world. The studies in this proposal seek to develop a novel, long-term therapy for hypertension by using an adeno-associated viral (AAV) vector containing an enhanced smooth muscle specific promoter (EnSM221) to deliver exogenous microRNA directed against vascular L-type (CaV1.2) calcium channels. Notably, CaV1.2 channels already are the target of clinical calcium channel blocking drugs used to lower blood pressure. However, our strategy will use AAV-mediated delivery of CaV1.2 channel microRNA to enable long- term and vascular-specific knockdown of calcium channel expression. Our aims include: 1) to demonstrate the ability of CaV1.2 channel microRNA to reduce blood pressure for at least 8 weeks without serious side effects in hypertensive mice, 2) confirm in vivo and in vitro that the vasoconstrictor function of CaV1.2 channels is suppressed in small mesenteric arteries, and 3) demonstrate that decreases in vascular CaV1.2 channel expression, whole-cell Ca2+ current, and restoration of K+ channel expression and resting membrane potential in mesenteric arterial cells correlate with the antihypertensive effect of CaV1.2 channel microRNA delivered by AAV to the vasculature. The results of this study will help to establish a foundation for using targeted AAV and microRNA technology to treat systemic hypertension and other chronic vascular abnormalities that are poorly controlled by short-term therapies. PUBLIC HEALTH RELEVANCE: High blood pressure known as hypertension afflicts over 60 million Americans and can lead to even more debilitating conditions. While there are numerous short-acting medications (including calcium channel blockers) to treat hypertension, most patients do not take their medications faithfully and consequently less than one third of patients have their blood pressure adequately controlled. We have developed a small RNA-based therapeutic (microRNA) which decreases expression of a calcium channel found in blood vessels that is also the site of action of all antihypertensive calcium channel blockers. When incorporated into a safe adeno associated virus, our therapeutic can continuously produce more microRNA over a period of months. In addition our therapeutic is coupled to a DNA sequence that will selectively generate the desired microRNA only in vascular smooth muscle cells that make up the wall of blood vessels. This is likely to reduce side effects encountered with many orally administered antihypertensive medications, which spread throughout the body. A longer lasting therapy with fewer side effects may be extremely beneficial for the >40 million Americans whose hypertension is not properly managed.