Abstract
The discovery that paracetamol is metabolized to the potent TRPV1 activator N-(4-hydroxyphenyl)-5Z,8Z,11Z,14Z-eicosatetraenamide (AM404) and that this metabolite contributes to paracetamol’s antinociceptive effect in rodents via activation of TRPV1 in the central nervous system (CNS) has provided a potential strategy for developing novel analgesics. Here we validated this strategy by examining the metabolism and antinociceptive activity of the de-acetylated paracetamol metabolite 4-aminophenol and 4-hydroxy-3-methoxybenzylamine (HMBA), both of which may undergo a fatty acid amide hydrolase (FAAH)-dependent biotransformation to potent TRPV1 activators in the brain. Systemic administration of 4-aminophenol and HMBA led to a dose-dependent formation of AM404 plus N-(4-hydroxyphenyl)-9Z-octadecenamide (HPODA) and arvanil plus olvanil in the mouse brain, respectively. The order of potency of these lipid metabolites as TRPV1 activators was arvanil = olvanil>>AM404> HPODA. Both 4-aminophenol and HMBA displayed antinociceptive activity in various rodent pain tests. The formation of AM404, arvanil and olvanil, but not HPODA, and the antinociceptive effects of 4-aminophenol and HMBA were substantially reduced or disappeared in FAAH null mice. The activity of 4-aminophenol in the mouse formalin, von Frey and tail immersion tests was also lost in TRPV1 null mice. Intracerebroventricular injection of the TRPV1 blocker capsazepine eliminated the antinociceptive effects of 4-aminophenol and HMBA in the mouse formalin test. In the rat, pharmacological inhibition of FAAH, TRPV1, cannabinoid CB1 receptors and spinal 5-HT3 or 5-HT1A receptors, and chemical deletion of bulbospinal serotonergic pathways prevented the antinociceptive action of 4-aminophenol. Thus, the pharmacological profile of 4-aminophenol was identical to that previously reported for paracetamol, supporting our suggestion that this drug metabolite contributes to paracetamol’s analgesic activity via activation of bulbospinal pathways. Our findings demonstrate that it is possible to construct novel antinociceptive drugs based on fatty acid conjugation as a metabolic pathway for the generation of TRPV1 modulators in the CNS.
Highlights
Paracetamol is an effective analgesic, a single oral dose of 1000 mg having an NNT of 3.6 for at least 50% pain reduction over 4–6 hours in patients with postoperative pain [1]
We show that incubation of mouse brain homogenates with another primary amine, HMBA, leads to the formation of arvanil and olvanil (Fig. 1A), which are both recognized as potent TRPV1 activators [9,10,11]
To establish whether fatty acid conjugation of 4-aminophenol and HMBA occurs in living animals, we determined the contents of AM404, HPODA, arvanil and olvanil in the brain 20 min after intraperitoneal injection of 4-aminophenol (30 and 100 mg/kg) and HMBA (100 and 300 mg/kg) in mice
Summary
Paracetamol is an effective analgesic, a single oral dose of 1000 mg having an NNT (numbers needed to treat) of 3.6 for at least 50% pain reduction over 4–6 hours in patients with postoperative pain [1]. Mapping the in vivo metabolism of paracetamol in different organs indicated an extensive de-acetylation to the primary amine 4-aminophenol in the liver [2]. This well-known paracetamol metabolite enters the brain, where it undergoes conjugation with arachidonic acid to yield AM404, a reaction dependent on the enzyme FAAH [2]. The design of molecules undergoing FAAH-dependent biotransformation to TRPV1 active drug metabolites in the CNS represents a plausible strategy for developing novel antinociceptive agents that may be more effective and less toxic than paracetamol
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