Structure Activity Relationship Studies of Psychostimulant Synthetic Cathinones Reveal Nanomolar Antagonist Potency of α‐Pyrrolidinohexiophenone (α‐PHP) at Human Muscarinic M2Receptors

Abstract

Synthetic cathinones (SCs), psychostimulant drugs of abuse, are β‐keto analogs of amphetamine that primarily target monoamine transporters as inhibitors or substrates. Certain pyrrolidine‐containing SCs, such as α‐Pyrrolidinohexiophenone (α‐PHP) and methylenedioxypyrovalerone (MDPV), are potent monoamine transporter inhibitors that appear to produce distinct toxic, autonomic—e.g., tachycardia and arrythmia—and neuropsychiatric—e.g., delirium and cognitive impairments—adverse events compared to N‐alkylated SCs and amphetamines, which are monoamine transporter substrates. Although the apparent differences in toxicity could be caused by unique pharmacodynamic effects at monoamine transporters, we hypothesized there is another mechanism. We focused on muscarinic receptors (MRs), as numerous MR antagonists are well‐characterized deliriants that also cause tachycardia. We conducted pharmacological assays assessing human M1, M2, M3, M4, and M5R activity of 10 pyrrolidine‐containing SCs, systematically evaluating the effect of the α‐carbon chain length of methylenedioxy‐containing and unsubstituted α‐propiophenone analogues. [3H]Scopolamine competition binding assays revealed that increasing the α‐carbon chain length of unsubstituted, but not methylenedioxy derivatives led to a stepwise enhancement in affinity (Ki) at all MR subtypes, with highest affinities typically observed at M2Rs. Relative to all other SCs tested, α‐PHP had the highest affinity at M2Rs, with a calculated pKi of 6.28 (Ki= 520 nM). We next assessed the M2R functional effects of α‐PHP using M2R—cAMP and M2R—β‐arrestin recruitment assays; α‐PHP behaved as an M2R competitive antagonist in both assays. α‐PHP antagonized oxotremorine‐elicited cAMP inhibition with an apparent pKb of 7.29 (Kb= 50 nM) and antagonized oxotremorine‐elicited β‐arrestin recruitment with an apparent pKb of 6.22 (Kb= 602 nM). Toxicology studies report that serum concentrations of commonly abused pyrrolidine‐containing SCs, including α‐PHP, can range from 713 to 2,619 nM. Thus, α‐PHP’s potency at M2Rs is physiologically relevant and likely contributes to its toxicity. Given the dominant function of M2Rs to slow heart rate, α‐PHP may cause tachycardia, in part, by inhibition of cardiac M2Rs. This inhibitory effect on the parasympathetic nervous system, in combination with sympathomimetic effects caused by monoamine transporter inhibition could explain the serious adverse cardiovascular events associated with certain pyrrolidine‐containing SCs.Support or Funding InformationFunding: NIDA R21DA040907