Analysis of a Novel Homeobox gene in CV Development

描述(由申请人提供)：这是一份修订后的(A1)R01竞争性续签申请，最初侧重于我们在胚胎心脏中发现的一种名为Hop的新的同源结构域蛋白的作用。我们发现，Hop功能丧失会导致发育性心脏缺陷，过度表达会导致心肌肥大。HOP可以通过招募I类HDAC在分子水平上作为转录共抑制因子发挥作用。这一发现使我们测试了HDAC抑制剂的效果，我们发现这些药物可以阻止Hop过度表达引起的心肌肥大，也可以阻止其他应激引起的心肌肥厚，包括β-肾上腺素能激动剂和拉伸。因此，我们寻找了解释这些影响的特定的I类HDAC2(S)，并发现在发育中的心脏和成人心脏中有高水平的HDAC2。HDAC2功能丧失导致对Hop诱导的心肌肥大和β-肾上腺素能激动剂诱导的肥厚具有抵抗力的小鼠。我们已经证明，在这些模型中，对肥大的抵抗是由GSK3-β的结构性激活引起的，因为GSK3-β的拮抗剂恢复了HDAC2缺失动物的肥大能力。我们的数据表明，HDAC2通过直接抑制一种名为INPP5F的新型肌醇多聚磷酸酶来影响AKT-GSK3-β途径，该酶具有降解PIP3的功能，从而影响AKT级联反应。这些发现具有直接的临床和翻译意义，因为磷酸酶是治疗心血管疾病的极好的可用药靶点，而且HDAC抑制剂已经在治疗癌症的临床试验中，并且很容易被改造成用于心脏领域。因此，我们将通过以下步骤进行这些观察：1)开发HDAC2的等位基因，以执行组织和时间特异性缺失，并确定HDAC2功能丧失对预先建立的肥厚逆转的影响；2)阐明Hop和HDAC2之间的遗传和生化相互作用；以及3)通过细胞和小鼠的功能获得和丧失方法检测INPP5F的功能。这些实验为开发治疗心肌肥厚和心力衰竭的新的治疗途径和范例提供了强大的前景，并对更广泛的细胞和器官生长调节领域产生了影响。 公共卫生相关性：该项目关注充血性心力衰竭的原因，并寻找新的治疗靶点。在美国，心力衰竭是导致死亡和残疾的主要原因，而且发病率还在上升。我们已经发现，目前正在进行癌症治疗临床试验的药物HDAC抑制剂在动物模型中预防心力衰竭方面具有有益的效果。该项目探索了一种在心脏表达的名为HDAC2的酶是HDAC抑制剂的分子靶点的可能性，以及它通过调节心脏中一种名为INPP5F的新型磷酸酶来发挥作用的可能性。我们希望确定HDAC2和INPP5F是否代表心脏病特定疗法开发的新靶点。

DESCRIPTION (provided by applicant): This is a revised (A1) competitive renewal application for an R01 initially focused on the role of a novel homeodomain protein called Hop that we discovered in the embryonic heart. We have found that Hop loss of function results in a developmental cardiac defect and that over-expression results in cardiac hypertrophy. Hop can function at the molecular level as a transcriptional co-repressor by recruiting class I HDACs. This finding led us to test the effects of HDAC inhibitors and we have found that these agents can block cardiac hypertrophy induced by Hop over-expression and also hypertrophy resulting from other stresses including beta-adrenergic agonists and stretch. Therefore, we have sought the specific class I HDAC(s) that account for these effects, and we have identified high levels of HDAC2 in the developing and adult heart. HDAC2 loss of function results in mice that are resistant to Hop- induced cardiac hypertrophy and to hypertrophy induced by beta-adrenergic agonists. We have shown that resistance to hypertrophy in these models is caused by constitutive activation of GSK3-beta, since antagonists of GSK3-beta restore the ability to hypertrophy on HDAC2 null animals. Our data indicates that HDAC2 affects the AKT-GSK3-beta pathway by directly repressing a novel inositol polyphosphate phosphatase called INPP5F, which functions to degrade PIP3 and thus affects the AKT cascade. These findings have direct clinical and translational implications since phosphatases are excellent drugable targets for the treatment of cardiovascular disease and because HDAC inhibitors are already in clinical trials for the treatment of cancer and could be readily adapted for use in the cardiac arena. Therefore, we will pursue these observations by: 1) Developing a floxed allele of HDAC2 in order to perform tissue and temporal specific deletion and to determine the effects of HDAC2 loss of function on regression of pre-established hypertrophy; 2) Elucidation of the genetic and biochemical interaction between Hop and HDAC2, and; 3) Examination of the function of INPP5F through gain and loss of function approaches in cells and in mice. These experiments offer strong prospects for developing new therapeutic avenues and paradigms for the treatment of cardiac hypertrophy and heart failure, with implications for the more general fields of cellular and organ growth regulation. PUBLIC HEALTH RELEVANCE: This project focuses on the causes of congestive heart failure, and on finding new therapeutic targets. Heart failure is a leading cause of death and disability in the United States, and the incidence is rising. We have found that HDAC inhibitors, which are drugs currently in clinical trials for cancer therapy, have beneficial effects in terms of heart failure prevention in animal models. This project explores the possibility that an enzyme called HDAC2, expressed in the heart, is the molecular target for HDAC inhibitors, and that it functions by regulating a novel phosphatase in the heart called INPP5F. We hope to determine if HDAC2 and INPP5F represent new targets for the development of specific therapies for heart disease.