SRF/MYOCD: new targets in Alzheimer's neurovasculature

描述（由申请人提供）： 身体中没有一个器官像大脑那样依赖于持续的血液供应。如果脑血流（CBF）中断，脑功能在几秒钟内停止，其细胞成分的不可逆损伤在几分钟内加剧。在老年患者中，越来越多的证据表明，血管危险因素和降低的CBF在阿尔茨海默病（AD）的发展中起着重要作用，甚至可能引发痴呆症的发作。我们在AD患者软脑膜脑动脉的血管平滑肌细胞（VSMC）中发现了两种过度丰富的相互作用的转录因子，它们对CBF起着关键性的调节作用。血清反应因子（SRF）和心肌素（MYOCD）是与VSMC功能相关的收缩基因程序的有效诱导剂。因此，AD-VSMC显示出几种收缩蛋白的表达升高，这些收缩蛋白在这些细胞中产生过度收缩状态。基于这些新的发现，我们假设，升高的SRF/MYOCD活性的VSMC的小脑动脉导致一个过度收缩的表型，这有助于脑血流量减少和神经血管解偶联在AD。在这个I期申请中，我们奋进进一步验证这些发现，并优化SRF/MYOCD依赖性基因转录的高通量筛选测定。为此，我们已经划定了三个里程碑，我们有信心，我们可以实现作为一个关键的基础，第二阶段的申请，我们同时提交了本快速通道建议。里程碑1将评估MYOCD在年龄匹配的对照VSMC中引发AD-VSMC过度收缩表型的能力。里程碑2将确定AD-VSMC过度收缩表型是否可以用RNAi至SRF来标准化。在里程碑3中，我们将优化最近开发的用于人类细胞的大规模荧光素酶测定，为II期研究中描述的研究做准备。这两个相互作用的转录因子可能是已知的AD脑灌注不足和认知下降的基础，这一发现代表了治疗干预的新范式。此处要解决的里程碑将为该快速通道申请的第II阶段中描述的目标提供重要平台，其中将筛选化合物的多样化小分子库，以寻找特异性破坏SRF/MYOCD相互作用的有机化学物质，从而使VSMC过度收缩正常化，并通过扩展使AD中的脑灌注不足和CBF失调正常化。

DESCRIPTION (provided by applicant): There is no organ in the body as dependent on a continuous supply of blood as the brain. If cerebral blood flow (CBF) is interrupted, brain function ceases within seconds and irreversible damage to its cellular constituents ensues within minutes. In elderly patients, increasing evidence suggests that vascular risk factors and lowered CBF have a major role in the development of Alzheimer's disease (AD) and possibly even trigger the onset of dementia. We have discovered an over-abundance of two interactive transcription factors in vascular smooth muscle cells (VSMC) from pial cerebral arteries of patients with AD which critically regulate CBF. The factors, serum response factor (SRF) and myocardin (MYOCD), are potent inducers of a contractile gene program linked to VSMC function. Consequently, AD-VSMC display elevated expression of several contractile proteins that effect a hypercontractile state in these cells. Based on these novel findings, we hypothesize that elevated SRF/MYOCD activity in VSMC of small cerebral arteries leads to a hypercontractile phenotype, which contributes to CBF reductions and neurovascular uncoupling in AD. In this Phase I application, we endeavor to further validate these findings and optimize a high-throughput screening assay for SRF/MYOCD-dependent gene transcription. To this end, we have delineated three milestones we are confident we can achieve as a critical foundation for a Phase II application that we have simultaneously submitted in this Fast Track proposal. Milestone 1 will assess the ability of MYOCD to elicit an AD-VSMC hypercontractile phenotype in age-matched control VSMC. Milestone 2 will ascertain whether the AD-VSMC hypercontractile phenotype can be normalized with an RNAi to SRF. In Milestone 3, we will optimize a recently developed large-scale luciferase assay for human cells in preparation for studies described in Phase II. The finding that two interactive transcription factors may underlie the known brain hypoperfusion and cognitive decline in AD represents a new paradigm for therapeutic interventions. The milestones to be addressed here will provide a vital platform for aims delineated in Phase II of this Fast Track Application in which a diverse small molecule library of compounds is to be screened for organic chemicals that specifically disrupt the SRF/MYOCD interaction thereby normalizing VSMC hypercontraction and, by extension, brain hypoperfusion and dysregulated CBF in AD.