The lipid hydrolase NAAA as a target for non-addictive analgesic medications

安全有效的镇痛药的发现是一个迫切的社会需求和美国国立卫生研究院的愈合倡议的核心。在这里，我们验证了一种假设，即n -酰基乙醇胺酸酰胺酶（NAAA）提供了一种没有滥用潜力的镇痛药物的新靶点。NAAA水解棕榈酰乙醇酰胺（PEA），这是一种脂质信使，通过参与核受体PPAR-α来抑制痛感。最近，国际疼痛研究协会（IASP）对临床前文献进行了荟萃分析，发现了NAAA抑制剂的实质性抗痛觉性，但这种特性背后的细胞底物尚不清楚。我们发现，与野生型幼崽相比，naaa敲除（ko）小鼠(i)对福尔马林的不良反应显著降低；（ii）在慢性收缩损伤（CCI）模型中未出现持续的感觉异常。相反，即使给予低于阈值的福尔马林剂量，Naaa-过表达和para-ko小鼠也表现出强烈的攻击性行为。NAAA -ko小鼠对疼痛的敏感性降低不是由于发育代偿，因为它可以通过给药NAAA抑制剂ARN19702来表现。重要的是，ARN19702在小鼠条件位置偏好（CPP）测试中没有发挥积极的激励作用，这表明它缺乏奖励特性。这些数据表明NAAA是发现非成瘾性镇痛药的一个有希望的靶点。我们将在两个具体目标中检验这一假设：目标1。验证NAAA作为镇痛药物发现的分子靶点。将解决三个问题：(1)NAAA抑制剂直接针对哪些细胞类型？我们将生成细胞特异性NAAA-ko小鼠系，并在福尔马林和CCI模型中评估疼痛相关行为和对NAAA阻断的敏感性。同时，我们将确定病理性疼痛对野生型小鼠背根神经节（DRG）和脊髓（SC）中naaa调节信号的影响。(2)哪些细胞类型介导naaa调控的抗感知信号？我们将通过创建细胞特异性PPAR-α-ko小鼠系，并在福尔马林和CCI测试中评估它们的疼痛相关反应及其对NAAA抑制剂的敏感性，来鉴定参与NAAA依赖性信号传导的PPAR-α-表达细胞。(3) NAAA抑制剂的药效学特征是否与安全有效地用于疼痛治疗相一致？我们将确定ARN19702是否(i)减轻自发性伤害感受，（ii）在反复给药后产生耐受性，以及（iii）表现出奖励和/或成瘾的潜力。目标2。开发改进的NAAA抑制剂并检查其作为镇痛剂的疗效。需要更好的NAAA抑制剂来评估NAAA在疼痛治疗中的可能作用。我们将结合基于计算结构的药物和构效关系研究来发现具有改进的药效学和药代动力学特性、强大的镇痛活性和无成瘾性的新型药物。

The discovery of safe and effective analgesics is an urgent societal need and a centerpiece of NIH’s HEAL initiative. Here, we test the hypothesis that the enzyme N-Acylethanolamine Acid Amidase (NAAA) offers a novel target for analgesic medications devoid of abuse potential. NAAA hydrolyzes palmitoylethanolamide (PEA), a lipid messenger that suppresses nociception by engaging the nuclear receptor PPAR-α. The substantial antinociceptive properties of NAAA inhibitors have been recently recognized by a meta-analysis of the preclinical literature conducted by the International Association for the Study of Pain (IASP), but the cellular substrates underlying such properties are still unknown. We found that Naaa-knock out (ko) mice (i) have markedly reduced nocifensive responses to formalin, compared to wild-type littermates; and (ii) fail to develop persistent sensory abnormalities in the chronic constriction injury (CCI) model. Conversely, Naaa- overexpressing and Ppara-ko mice exhibit robust nocifensive behavior even when given a subthreshold formalin dose. The reduced sensitivity of Naaa-ko mice to pain does not result from developmental compensation, because it can be phenocopied by administration of the NAAA inhibitor ARN19702. Importantly, ARN19702 does not exert positive motivational effects in the mouse conditioned place preference (CPP) test, which is suggestive of a lack of rewarding properties. These data point to NAAA as a promising target for the discovery of non-addictive analgesics. We will test this hypothesis in two specific aims: Aim 1. Validate NAAA as a molecular target for analgesic drug discovery. Three questions will be addressed: (1) What cell types are directly targeted by NAAA inhibitors? We will generate cell-specific NAAA-ko mouse lines and evaluate, in the formalin and CCI models, pain-related behaviors and sensitivity to NAAA blockade. In parallel, we will determine the impact of pathological pain on NAAA-regulated signaling in dorsal root ganglia (DRG) and spinal cord (SC) of wild-type mice. (2) What cell types mediate NAAA-regulated antinociceptive signaling? We will identify PPAR-α-expressing cells involved in NAAA-dependent signaling by creating cell-specific PPAR-α-ko mouse lines and evaluating their pain-related responses and their sensitivity to NAAA inhibitors in the formalin and CCI tests. (3) Is the pharmacodynamic profile of NAAA inhibitors compatible with safe and effective use in pain therapy? We will determine whether ARN19702 (i) alleviates spontaneous nociception, (ii) produces tolerance after repeated administration, and (iii) exhibits rewarding and/or addicting potential. Aim 2. Develop improved NAAA inhibitors and examine their efficacy as analgesic agents. Better NAAA inhibitors are needed to assess NAAA’s possible role in pain therapy. We will combine computational structure-based drug and structure-activity relationship studies to discover novel agents with improved pharmacodynamic and pharmacokinetic properties, robust analgesic activity, and no addictive potential.