Serotonin 2A receptor binding, functional activity, and in vitro metabolism of two trideuteromethoxy 2C-B isotopologues

The aim of this study was to investigate the effects of deuterium substitution on the in vitro pharmacology of the psychedelic drug 2-(4-bromo-2,5-dimethoxyphenyl)ethan-1-amine (2C-B). Two deuterated isotopologues of 2C-B, namely 2-(4-bromo-5-methoxy-2-(methoxy-d3)phenyl)ethan-1-amine (2CB-2OCD₃) and 2-(4-bromo-2-methoxy-5-(methoxy-d3)phenyl)ethan-1-amine (2CB-5OCD₃), were synthesized and tested in cell lines expressing human 5-HT_2A receptors for potential isotope effects on 5-HT_2A receptor binding and functional activity, and for metabolism by human liver microsomes (HLM). The 2C-B isotopologues and 2C-B exhibited picomolar-to-nanomolar potency in binding to [¹²⁵I]DOI-labelled 5-HT_2A receptors and were equally efficacious to 2C-B in stimulating 5-HT_2A-mediated intracellular Ca⁺⁺ release and β-arrestin 2 engagement. No appreciable metabolism was observed for 2C-B or the deuterated analogues by HLM over the time course of the experiment. Deuteration of the ring methoxys of 2C-B appeared to increase affinity at [¹²⁵I]DOI-labelled 5-HT_2A receptors, likely due to dampened internal vibrational frequencies and rotational motions arising from primary and secondary kinetic isotope effects. Deuteration did not affect the ability of the drugs to stimulate intracellular calcium release or to recruit β-arrestin 2, with all compounds displaying similar low nanomolar potency and efficacy in these assays.