Computation-assisted discovery of bioactive minor cannabinoids from hemp

该项目旨在对大麻中微量大麻素的潜在治疗效果进行机制研究。我们建议将人工智能整合到我们的工作流程中，以加速生物活性次要大麻素的发现。我们专注于慢性疼痛作为非精神活性大麻素的治疗益处。大量文献表明，大麻中的萜酚类物质可能具有治疗疼痛和其他由低度慢性炎症引起的疾病的潜力。识别源自天然产物 (NP) 的潜在疗法的传统经典方法是费力且耗时的生物测定引导分馏。正如我们的可行性研究所示，我们用于发现生物活性纳米颗粒的人工智能辅助方法加速了发现流程。我们预计我们将成功识别出以前未在临床相关生物测定中研究过的生物活性大麻素。我们的具体目标是： 具体目标 1：预测和验证 CBD 耗尽的大麻提取物中单个大麻素在与伤害感受相关的细胞培养模型中的影响。 Columbia Basin Bioscience 运营着一家工业规模的 CBD 制造工厂，利用富含大麻素的大麻提取物对 CBD 进行重结晶。我们将使用各个重结晶阶段的上清液（“母液”）作为我们在计算辅助发现生物活性物质中的次要大麻素来源。我们将使用稳定过表达重组人 TRPV1 的 HEK-293 细胞作为与伤害感受相关的细胞培养模型来进行钙流测定。我们还将测试 FAAH/COX-2 抑制活性，因为针对内源性大麻素和内香草素系统的双重作用化合物正在成为慢性疼痛管理的新型治疗选择。我们将通过在相同的测定中测试单独的纯大麻素来验证最有效的活性预测因子。我们还将从我们的大麻素标准存储库中确定 17 种次要大麻素的生物活性，其中可能包括也可能不包括我们发现管道中的生物活性预测因子。 具体目标 2：预测和验证生物活性大麻素之间的协同/相互作用效应。我们的人工智能发现管道能够根据含有生物活性大麻素的粗馏分中纳米颗粒绝对浓度的测量来检测协同、相加或拮抗效应，我们将为其建立浓度效应曲线。我们将量化一部分正宗大麻素之间的药理学相互作用，其中包括我们标准库中的 17 种次要大麻素，这些大麻素在我们的检测中显示出生物活性，或者据报道可以减弱 TRPV1 相关的疼痛信号传导。我们预计所提出的人工智能辅助发现方法将产生大麻素候选物，减弱伤害性 TRPV1-FAAH/COX-1 信号通路，以便在临床前疼痛模型中进行进一步的机制测试。从 CBD 行业的废品中开发出次要大麻素作为止痛剂，不仅有利于慢性疼痛患者，而且有利于可持续的农产品加工业。

This project aims to conduct mechanistic research of potential therapeutic benefits of minor cannabinoids in hemp. We propose to integrate Artificial Intelligence in our workflow to accelerate the discovery of bioactive minor cannabinoids. We focus on chronic pain as our therapeutic benefit of non-psychoactive cannabinoids. A large body of literature suggests that terpenophenolics from hemp may have potential in the treatment of pain and other conditions that are driven by low-grade chronic inflammation. The conventional, classic approach to identify potential therapeutics derived from natural products (NPs) is laborious and time-consuming bioassay-guided fractionation. Our AI-assisted approach for the discovery of bioactive NPs accelerates the discovery pipeline, as our feasibility studies show. We expect that we will succeed in identifying bioactive cannabinoids that have previously not been studied in clinically relevant bioassays. Our Specific Aims are: Specific Aim 1: To predict and validate the effects of individual cannabinoids in CBD-depleted hemp extracts in cell culture models relevant to nociception. Columbia Basin Bioscience operates an industrial scale CBD manufacturing facility which makes use of recrystallization of CBD from cannabinoid-enriched hemp extracts. We will use the supernatants of various recrystallization stages (“mother liquors”) as our source of minor cannabinoids in our computation-assisted discovery of bioactives. We will perform calcium flux assays using HEK-293 cells stably over-expressing recombinant human TRPV1 as a cell culture model relevant to nociception. We will also test for FAAH/COX-2 inhibitory activity as dual acting compounds targeting the endocannabinoid and endovanilloid systems are emerging as a novel treatment option for chronic pain management. We will validate the most potent predictors of activity by testing individual, pure cannabinoids in the same assays. We will also determine the bioactivity of 17 minor cannabinoids from our repository of cannabinoid standards, which may or may not include predictors of bioactivity from our discovery pipeline. Specific Aim 2: To predict and validate synergistic/interactive effects between bioactive cannabinoids. Our AI-discovery pipeline has the ability to detect synergistic, additive, or antagonistic effects based on measurements of absolute concentrations of NPs in crude fractions containing bioactive cannabinoids for which we will establish concentration-effect curves. We will quantify the pharmacological interactions among a subset of authentic cannabinoids including 17 minor cannabinoids from our repository of standards which show bioactivity in our assays or which have been reported to attenuate TRPV1-related pain signaling. We expect that the proposed AI-assisted discovery approach will generate cannabinoid candidates that attenuate nociceptive TRPV1-FAAH/COX-1 signaling pathways for further mechanistic testing in preclinical models of pain sensation. The development of minor cannabinoids as pain mitigators from waste products of the CBD-industry will benefit not only chronic pain patients but also a sustainable agro-industry.