Affinity and Efficacy Studies of Tetrahydrocannabinolic Acid A at Cannabinoid Receptor Types One and Two
Affinity and Efficacy Studies of Tetrahydrocannabinolic Acid A at Cannabinoid Receptor Types One and Two
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DOI：
10.1089/can.2016.0032
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发表时间：
2017-05-01
影响因子：
3.8
通讯作者：
Glass, Michelle
中科院分区：
文献类型：
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作者：
McPartland, John M.;MacDonald, Christa;Glass, Michelle
简介：大麻生物合成Delta（9）-tetrahydrocannabinolic acid（THCA-A），其脱羧成Delta（9）-tetrahydrocannabinol（THC）。人们对THCA-A的治疗用途越来越感兴趣，但其临床应用可能会受到不稳定性的阻碍。THCA-A缺乏大麻拟似作用;我们假设它对大麻素受体1（CB 1）几乎没有结合亲和力。材料和方法：用高效液相色谱法（HPLC）测试经认证的参考标准的纯度。使用转染的HEK细胞和[H-3] CP 55，940，在竞争结合试验中测量THCA-A和THC对人（h）CB 1和hCB（2）的结合亲和力。使用生物发光共振能量转移（BRET）生物传感器在环腺苷单磷酸酶（cAMP）测定中测量对hCB（1）和hCB（2）的功效。THCA-A在hCB（1）和hCB（2）上显示出小但可测量的结合，相当于分别约为3.1 μ M和12.5 μ M的Ki值。THC对hCB（1）的亲和力高62倍，对hCB（2）的亲和力高125倍。在功效测试中，THCA-A（10 μ M）轻微抑制毛喉素刺激的hCB中的cAMP（1），提示弱激动剂活性，并且对hCB没有可测量的功效（2）.讨论：在我们的THCA-A认证标准品中存在THC与脱羧动力学一致（本文综述的文献），这表明THC的污染几乎是不可避免的。10 μ M时的THCA-A结合接近200 nM时的THC结合。因此，我们怀疑我们的一些THCA-A结合曲线是人为的-从它不可避免的脱羧成THC-和THCA-A的结合亲和力甚至比我们的估计值更弱。我们得出结论，THCA-A对CB 1或CB 2几乎没有亲和力或功效。
Introduction: Cannabis biosynthesizes Delta(9)-tetrahydrocannabinolic acid (THCA-A), which decarboxylates into Delta(9)-tetrahydrocannabinol (THC). There is growing interest in the therapeutic use of THCA-A, but its clinical application may be hampered by instability. THCA-A lacks cannabimimetic effects; we hypothesize that it has little binding affinity at cannabinoid receptor 1 (CB1).Materials and Methods: Purity of certified reference standards were tested with high performance liquid chromatography (HPLC). Binding affinity of THCA-A and THC at human (h) CB1 and hCB(2) was measured in competition binding assays, using transfected HEK cells and [H-3]CP55,940. Efficacy at hCB(1) and hCB(2) was measured in a cyclic adenosine monophosphase (cAMP) assay, using a Bioluminescence Resonance Energy Transfer (BRET) biosensor.Results: The THCA-A reagent contained 2% THC. THCA-A displayed small but measurable binding at both hCB(1) and hCB(2), equating to approximate K-i values of 3.1 mu M and 12.5 mu M, respectively. THC showed 62-fold greater affinity at hCB(1) and 125-fold greater affinity at hCB(2). In efficacy tests, THCA-A (10 mu M) slightly inhibited forskolin-stimulated cAMP at hCB(1), suggestive of weak agonist activity, and no measurable efficacy at hCB(2).Discussion: The presence of THC in our THCA-A certified standard agrees with decarboxylation kinetics (literature reviewed herein), which indicate contamination with THC is nearly unavoidable. THCA-A binding at 10 mu M approximated THC binding at 200nM. We therefore suspect some of our THCA-A binding curve was artifact-from its inevitable decarboxylation into THC-and the binding affinity of THCA-A is even weaker than our estimated values. We conclude that THCA-A has little affinity or efficacy at CB1 or CB2.