Inhibitors of the ENT4 Adenosine Transporter for Cardioprotection

用于心脏保护的 ENT4 腺苷转运蛋白抑制剂

• 批准号: 7907749
• 负责人: John K Buolamwini
• 金额: $ 18.5万
• 依托单位: UNIVERSITY OF TENNESSEE HEALTH SCI CTR
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-14 至 2012-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7907749
• 关键词: 5' -Nucleotidase; Adenosine; Affect; Affinity; American; Biochemical; Biochemistry; Biological Assay; Brain; Cardiac; Cardiac Surgery procedures; Cardiotonic Agents; Cause of Death; Cell Line; Cell membrane; Cellular biology; Chemosensitization; Cholesterol; Clinical; Clinical Trials; Combinatorial Synthesis; Data; Dependence; Development; Dopamine; Dose; Drops; Drug Delivery Systems; Family; Family suidae; Figs - dietary; Flavones; Goals; Grant; Heart; Heart Diseases; Hepatotoxicity; Human; Inhibitory Concentration 50; Injury; Intestines; Intracellular Membranes; Investigation; Ischemia; Kidney; Lead; Left; Life; Light; Literature; Liver; Mammalian Cell; Maps; Medical; Membrane Transport Proteins; Methods; Modeling; Morbidity - disease rate; Muscle Cells; Myocardial; Myocardial Infarction; Myocardial Ischemia; Nucleoside Transporter; Nucleosides; Patients; Pharmaceutical Preparations; Physiological; Physiology; Property; Rattus; Regimen; Regulation; Relative (related person); Reperfusion Injury; Reperfusion Therapy; Research; Risk; Role; Running; Series; Serotonin; Stroke; Structure-Activity Relationship; System; Testing; Tissues; Toxic effect; Ventricular; Work; adenosine transporter; analog; cost; design; effective therapy; extracellular; flavone; high risk; improved; inhibitor/antagonist; insight; interdisciplinary approach; interest; interstitial; member; monoamine; mortality; novel; pharmacophore; preconditioning; prevent; public health relevance; small molecule; synergism; time use; tool; transport inhibitor; uptake

DESCRIPTION (provided by applicant): The advances in treatment notwithstanding, heart disease remains the number one cause of death in the Nation, claiming the lives of nearly one million Americans each year, and running up medical costs estimated at $286.5 billion per year. Ischemia-reperfusion injury is the major cause of complications of cardiac surgery; morbidity and mortality, but current cardioprotective options are very limited and suboptimal in high-risk patients. This leaves a dire need for effective treatments. Approaches to augment the cardioprotective actions of the physiological nucleoside adenosine (Ado) continue to hold promise and remain to be exploited for novel cardioprotective therapies. This project will pursue the identification of high-affinity specific inhibitors of a recently characterized novel pH-dependent cardiac adenosine transporter, ENT4. ENT4 selectively transports adenosine relative to other physiological nucleosides, making it an intriguing transporter to study with respect to adenosine modulation in myocardial ischemia and reperfusion. Currently, there are no known potent or specific inhibitors of ENT4 Ado transport. Thus, the identification of potent specific inhibitors is of high priority, not only for cardiac studies but for studying adenosine regulation by ENT4 in other tissues as well. They will facilitate studies of its role in cardiac ischemia-reperfusion, and show whether or not ENT4 could be a cardioprotection drug target for combination regimens with other cardioprotective agents like ENT1 nucleoside transport inhibitors. In line with our pursuit of adenosine uptake inhibitors as cardioprotective agents for the treatment of ischemia- reperfusion injury, we have cloned and stably expressed the human ENT4 transporter (hENT4) in a nucleoside transporter deficient porcine PK15 (PK15NTD) cell line. We subsequently used this expression system to screen, and identify small molecules inhibitors of ENT4 Ado transport with IC50s values down to submicromolar level, and selectivity up to 78-fold relative to one or the other human plasma membrane equilibrative nucleoside transporters, hENT1 or hENT2. Thus we aim to: 1) optimize the potency and selectivity of these lead compounds through structure- activity relationship (SAR), pharmacophore mapping and 3D-QSAR studies, and to 2) explore the cardioprotective effects of ENT4 inhibitors in an isolated rat heart global ischemia model of myocardial infarction. The new ENT4 inhibitors will be compared with our recently discovered cardioprotective ENT1 inhibitors (Zhu et al., Am. J. Physiol. Heart Circ. Physiol. 2007, 292, H2921-2926). We will explore the possibility that ENT4 inhibitors can be additive or synergistic in combination with ENT1 inhibitors for cardioprotection. A multidisciplinary approach combining parallel combinatorial synthesis, chemoinformatics/3D-QSAR, biochemistry, physiology and cell biology methods will be applied to achieve the aims of the project. The results will provide insights on the role of ENT4 in cardiac ischemia, and its inhibitors in cardioprotection, and provide potential lead compounds for the development of new cardioprotective drugs and/or use as much needed novel research tools to probe ENT4 physiology. The adenosine potentiation approach to cardioprotection could not be more timely, in light of recent clinical results showing that statins, which increase interstitial Ado levels by activating ecto-5'-nucleotidase, can reduce heart attack and stroke risks in people with normal cholesterol levels (Ridker et al., N. Engl. J. Med. 2008; AHA, 2008); and the demonstration of synergism between low dose statin and Ado uptake inhibitor for cardioprotection, when used in combination (Ye et al., 2007). PUBLIC HEALTH RELEVANCE: In this proposal, we seek to study a new class of adenosine uptake inhibitors as cardioprotective agents. We will investigate structure-activity relationships (SARs) and test the cardioprotective properties of the compounds in an isolated rat heart ischemia model.

描述（由申请人提供）：尽管治疗方法取得了进步，但心脏病仍然是美国第一大死因，每年夺去近百万美国人的生命，并且每年增加的医疗费用估计为 2,865 亿美元。缺血再灌注损伤是心脏手术并发症的主要原因；发病率和死亡率，但目前的心脏保护选择非常有限，并且对于高危患者来说不是最佳的。这迫切需要有效的治疗方法。增强生理核苷腺苷 (Ado) 心脏保护作用的方法仍然有希望，并且仍有待开发用于新型心脏保护疗法。该项目将致力于鉴定最近表征的新型 pH 依赖性心脏腺苷转运蛋白 ENT4 的高亲和力特异性抑制剂。相对于其他生理核苷，ENT4 选择性转运腺苷，使其成为研究心肌缺血和再灌注中腺苷调节的有趣转运蛋白。目前，尚无已知的 ENT4 Ado 转运的有效或特异性抑制剂。因此，鉴定有效的特异性抑制剂具有高度优先性，不仅对于心脏研究，而且对于研究 ENT4 在其他组织中的腺苷调节也是如此。他们将促进研究其在心脏缺血再灌注中的作用，并表明 ENT4 是否可以成为与其他心脏保护药物（如 ENT1 核苷转运抑制剂）联合治疗的心脏保护药物靶点。根据我们对腺苷摄取抑制剂作为治疗缺血再灌注损伤的心脏保护剂的追求，我们在核苷转运蛋白缺陷的猪 PK15 (PK15NTD) 细胞系中克隆并稳定表达了人 ENT4 转运蛋白 (hENT4)。随后，我们使用该表达系统筛选并鉴定了 ENT4 Ado 转运的小分子抑制剂，其 IC50 值低至亚微摩尔水平，相对于一种或另一种人质膜平衡核苷转运蛋白 hENT1 或 hENT2，选择性高达 78 倍。因此，我们的目标是：1）通过构效关系（SAR）、药效团图谱和3D-QSAR研究优化这些先导化合物的效力和选择性，2）探索ENT4抑制剂在离体大鼠心脏心肌梗死整体缺血模型中的心脏保护作用。新的 ENT4 抑制剂将与我们最近发现的心脏保护性 ENT1 抑制剂进行比较（Zhu et al., Am. J. Physiol. Heart Circ. Physiol. 2007, 292, H2921-2926）。我们将探索 ENT4 抑制剂与 ENT1 抑制剂联合使用以叠加或协同作用进行心脏保护的可能性。将采用结合平行组合合成、化学信息学/3D-QSAR、生物化学、生理学和细胞生物学方法的多学科方法来实现该项目的目标。这些结果将提供有关 ENT4 在心脏缺血中的作用及其抑制剂在心脏保护中的作用的见解，并为开发新的心脏保护药物和/或使用急需的新颖研究工具来探索 ENT4 生理学提供潜在的先导化合物。鉴于最近的临床结果表明，他汀类药物通过激活 5'-核酸外切酶来增加间质 Ado 水平，可以降低胆固醇水平正常的人心脏病发作和中风的风险，因此用腺苷增强心脏保护方法来得非常及时（Ridker 等人，N. Engl. J. Med. 2008；AHA，2008）；并证明了低剂量他汀类药物和 Ado 摄取抑制剂联合使用时对心脏保护的协同作用（Ye 等，2007）。公共健康相关性：在本提案中，我们寻求研究一类新型腺苷摄取抑制剂作为心脏保护剂。我们将研究构效关系（SAR）并在离体大鼠心脏缺血模型中测试化合物的心脏保护特性。

项目成果

Dipyridamole analogs as pharmacological inhibitors of equilibrative nucleoside transporters. Identification of novel potent and selective inhibitors of the adenosine transporter function of human equilibrative nucleoside transporter 4 (hENT4).

双嘧达莫类似物作为平衡核苷转运蛋白的药理学抑制剂。

• DOI: 10.1016/j.bcp.2013.08.063
• 发表时间: 2013
• 期刊: Biochemical pharmacology
• 影响因子: 5.8
• 作者: Wang,Chunmei;Lin,Wenwei;Playa,Hilaire;Sun,Shan;Cameron,Keyuna;Buolamwini,JohnK
• 通讯作者: Buolamwini,JohnK