Synthesis and in vitro and in vivo screening of fused and tethered heterocyclic peptidomimetics for the discovery of new analgesics with decreased side effects

融合和束缚杂环肽模拟物的合成以及体外和体内筛选，以发现副作用减少的新型镇痛药

• 批准号: 10297832
• 负责人: Jay P. McLaughlin
• 金额: $ 29.71万
• 依托单位: FLORIDA INTERNATIONAL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-11 至 2024-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10297832
• 关键词: Absence of pain sensation; Acetic Acids; Address; Affinity; Agonist; Amino Acids; Analgesics; Area; BCL2L11 gene; Behavior assessment; Binding; Binding Proteins; Biological; Biological Assay; Biological Availability; Blood; Blood - brain barrier anatomy; Brain; Clinical; Constipation; Cyclic AMP; Data; Development; Dose; Drug Kinetics; Evaluation; Exhibits; Failure; Generations; Goals; Harvest; Heterocyclic Compounds; Imidazolidines; Imines; In Vitro; Lead; Lead levels; Libraries; Liver; Mass Spectrum Analysis; Measurement; Mediating; Metabolic; Molecular Probes; Morphine; Motor Activity; Mus; Naloxone; Opiate Addiction; Opioid; Opioid Analgesics; Opioid Antagonist; Opioid Receptor; Opioid agonist; Oral; Oral Administration; Outcome; Pain; Parents; Peptides; Peripheral; Permeability; Physiological; Piperazines; Plasma Proteins; Property; Research; Rewards; Sedation procedure; Series; Side; Signal Transduction; Suggestion; System; Tail; Testing; Time; Ventilatory Depression; Vertebral column; Water; Withdrawal; abuse liability; antagonist; aqueous; base; beta-arrestin; computational chemistry; conditioned place preference; design; gastrointestinal epithelium; imidazolone; improved; in vivo; innovation; interest; intravenous administration; kappa opioid receptors; liquid chromatography mass spectrometry; novel; peptidomimetics; radioligand; receptor; recruit; respiratory; scaffold; screening; side effect; small molecule; therapeutic opioid

We hypothesize that the development of peripherally-restricted, mixed-activity opioid receptor agonists will produce robust analgesia while exhibiting reduced side effects, including a lack of respiratory depression or reinforcing properties. In preliminary studies, the in vitro screening of novel bis-imidazolidines and fused heterocyclic lead compounds BIM-22 and FDC-14 identified mixed-opioid receptor agonist activity. After peripheral (i.p.) administration to mice, both compounds demonstrated antinociception equivalent to morphine in the 55oC warm-water tail withdrawal (tail-flick) assay, but FDC-14 was 25 times more potent than morphine in the acetic acid writhing test, with an antinociceptive potency ratio of 72.7, suggestive of peripherally- restricted activity. Confirming this, LC-MS/MS studies detected BIM-22 and FDC-14 in harvested mouse blood, but not brain, after i.v. administration. This proposal addresses two research areas of particular interest for this FOA: i) The development of new and innovative molecular probes for receptors and new strategies for innovative peptidomimetic design, and ii) The identification of structurally diverse, orally active, metabolically stable peripherally-restricted opioid agonists. We propose six interacting aims: In Aim 1, we propose the computationally guided synthesis of heterocyclic peptidomimetics: bis-imidazolidin-2-imines, piperazines, bis- piperazine, bis-imidazolone and fused heterocyclic libraries. In Aim 2, we will screen all compounds in competition radioligand binding assays to determine affinity for μ- (MOR), δ- (DOR), and κ- (KOR) opioid receptors, and in functional assays to identify dual δ/κ or dual δ/µ agonists. In Aim 3, ten selected compounds will be evaluated in vitro for pharmacokinetic properties and screened in vivo for antinociception using the mouse tail-flick assay. Lead compounds identified from this aim will guide 2nd generation SAR studies to enhance peripherally-selective activity. In Aim 4: we will perform full in vivo antinociceptive characterization of lead agonist compounds. The most stable, active 4 agonists not crossing the BBB in Aims 2+3 will be evaluated after i.p. administration with mouse tail-flick and acetic-acid writhing assays for efficacy, duration of action, and opioid receptor selectivity. In Aim 5: we will characterize the bioavailability in mice of the selected 4 agonists following oral administration and confirm their inability to penetrate the blood brain barrier. In Aim 6: The two most potent bioavailable novel agonists identified in Aims 1-5 will be examined for liabilities, specifically antinociceptive tolerance, respiratory and hyperlocomotor effects in the CLAMS physiological and behavioral assessment system, sedation and disruption of coordinated locomotor activity in the rotorod assay, and assessment for rewarding or aversive effects in the conditioned place preference (CPP) assay. Effects on GI transit will also be evaluated to assess effects on constipation. In summary, we expect to generate novel peripherally-restricted, mixed-activity peptidomimetic opioid receptor agonists as both probes and analgesics with reduced side effects, thereby significantly impacting analgesic development.

我们假设，外周受限的混合活性阿片受体激动剂的开发将 产生强劲的止痛，同时表现出较少的副作用，包括缺乏呼吸抑制或 增强性能。在初步研究中，新的双咪唑类化合物和融合化合物的体外筛选 杂环先导化合物BIM-22和FDC-14鉴定了混合阿片受体激动剂活性。之后 外围设备(IP)在小鼠身上，这两种化合物都表现出与吗啡相当的抗伤害作用 在55oC温水甩尾(甩尾)实验中，FDC-14的效力是吗啡的25倍 在醋酸扭体实验中，其抗伤害性效价比为72.7，提示外周- 受限制的活动。为了证实这一点，LC-MS/MS研究在采集的小鼠血液中检测到BIM-22和FDC-14， 但不是大脑，在静脉注射之后。行政管理。这项建议涉及两个特别感兴趣的研究领域 FOA：i)用于受体的新的和创新的分子探针的开发和用于 创新的多肽模拟设计，以及ii)鉴定具有结构多样性、口腔活性、代谢活性的 稳定的外周受限阿片类激动剂。我们提出了六个互动目标：在目标1中，我们提出了 计算导引合成杂环肽类药物：双咪唑烷-2-亚胺，哌嗪，双- 哌嗪，双咪唑酮和融合杂环文库。在目标2中，我们将筛选出 竞争放射配基结合分析测定μ-(MOR)、δ-(DOR)和κ-(KOR)阿片的亲和力 受体，并在功能分析中识别双δ/κ或双δ/µ激动剂。在目标3中，选择了10种化合物 将在体外评估药代动力学特性，并在体内筛选抗伤害效应 小鼠甩尾试验。根据这一目标确定的先导化合物将指导第二代SAR研究 增强外周选择性活动。在目标4中：我们将在体内进行完整的抗伤害性表征 先导激动剂化合物。在AIMS 2+3中，最稳定、最有效的4种激动剂不会越过血脑屏障 在I.P.之后进行评估。小鼠甩尾法和醋酸扭体法给药的疗效、持续时间 作用和阿片受体选择性。在目标5中：我们将表征选定的4种药物在小鼠体内的生物利用度 口服后的激动剂，并确认它们无法穿透血脑屏障。在目标6中： 将检查AIMS 1-5中确定的两种最有效的生物可用新型激动剂的可靠性， 特定的抗伤害性耐受，呼吸和超运动效应在文蛤的生理和 行为评估系统，旋转试验中协调运动活动的镇静和干扰， 以及在条件性位置偏爱(CPP)分析中评估奖励或厌恶效应。对……的影响 还将评估胃肠道转运，以评估对便秘的影响。总而言之，我们希望产生新的 外周受限的混合活性模拟肽类阿片受体激动剂作为探针和镇痛剂 副作用减少，从而显著影响止痛药的发展。

项目成果

Bis-Cyclic Guanidine Heterocyclic Peptidomimetics as Opioid Ligands with Mixed μ-, κ- and δ-Opioid Receptor Interactions: A Potential Approach to Novel Analgesics.

• DOI: 10.3390/ijms23179623
• 发表时间: 2022-08-25
• 期刊: International journal of molecular sciences
• 影响因子: 5.6