Title : Isolation of a novel N-methyltransferase from psychedelic cane toad (Rhinella murina) deployable in bio-based production platforms for psychiatric drug discovery
Abstract:
The skin and paratoid gland secretions of many toads in the Bufonidae family contain several natural products with importance to traditional medicine. Some of these compounds, such the indole amines 5-hydroxy-N,N-dimethyltryptamine (bufotenine) and 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT) are psychedelics with therapeutic potential for the treatment of psychiatric mood disorders. The availability of novel biosynthetic enzymes acting on tryptamine derivatives would facilitate synthetic biology efforts toward biocatalytic and microbial-based production of novel, psychedelic-inspired pharmaceuticals. We mined the transcriptome of Cane toad (Rhinella murina) to isolate a novel N-methyltransferase (RmNMT) homologue distantly related (40-45% amino acid sequence identity) to mammalian indolethylamine N-methyltransferase (INMT) and nicotinamide N-methyltransferase (NNMT). Steady-state enzyme kinetic characterization showed that RmNMT coverts tryptamine and serotonin to N,N-dimethyltryptamine (DMT) and bufotenine, respectively, in a two-step reaction. RmNMT also catalyzes the robust N-methylation of a wide variety of other tryptamine derivatives, although indolethylamines substituted at 7- and 2-positions were poor substrates or were not accepted, respectively. Batch-fed in vivo conversions in Escherichia coli cultures followed by extraction chemistry yielded a pairwise collection of natural and novel primary and N-methylated indole amines in sufficient quantity and purity for pharmacological assessment. Compounds were assayed for 5-HT2A and 5-HT1A receptor-binding activities to investigate the impact of N-methylation on differentially substituted tryptamine derivatives with previously known or potential effects on psychiatric mood. Ki values of N,N-dimethylated tryptamines were approximately 2-5 fold higher than their N-desmethyl counterparts suggesting reduced binding affinity to 5-HT1A. However, timed exposure to MAO (monamine oxidase)-rich human liver microsomes (HLMs) revealed reduced the stability of most N-desmethyl tryptamines, in support of a broad protective function of N-methylation in vivo. Results revealed key structure-activity relationship features informing future drug design efforts.
What will audience learn from your presentation?
- Psychedelic drugs are expected to be represent novel treatments for mood psychiatric disorders
- Existing psychedelics can be improved or built-upon to inspire a new collection of pharmaceuticals available to patients recalcitrant to current treatments
- We use synthetic biology to build psychedelic-inspired drugs; here, we present the example of using natural sources (Toad) to find & develop new tools useful for drug development
- Audience will observe an expansive pipeline to develop psychedelic-inspired drugs, from gene mining in Toad to synthetic biology to pharmaceutical testing and ultimately in vivo models
