Functional Characterization Of Novel Photo-Switchable Neuromuscular Blockers

Nicotinic acetylcholine receptors (nAChR) are ligand-gated ion channels found in the nervous system and skeletal muscles. They are composed by combinations of α, β, γ, δ, and ε subunits, resulting in distinct receptors with diverse ion selectivity, physiological function and pharmacology. Drugs targeting nAChRs are commonly used in the treatment of neurological disorders and as muscle relaxants. Using azobenzene as basic molecular skeleton, we have designed a series of novel drugs that differentially effect nAChR activity as a function of their isomeric state; isomerization was reversibly induced by light irradiation. Two compounds (CH168 and CH376) behaved as potent ligands with a clear selectivity (up to 60-fold) for the muscular nAChRs (Ki = 35-42 nM), compared to neuronal α7 and α4β2 nAChRs. The trans-isomer of both compounds displayed higher activity against nAChR. CH168 behaved as an inhibitor of muscular nAChR expressed in Xenopus oocytes. At 10 µM, CH168 completely inhibited the activation by acetylcholine (ACh, 50 µM). Cis-isomerization of CH168 by irradiation with UV light (LED, peak at 340 nm) reverted about 20% of inhibition at the same concentration. Our observations suggest that the cis-isomer of CH168 remained bound to nAChR after isomerization, but it was less effective in inhibiting activation by ACh than the trans-isomer. The trans-isomer of CH376 behaved as an agonist, activating nAChR at concentrations of 500 nM and below. In contrast, the cis-isomer of CH376 was unable to render the channel open. Intriguingly, CH168 and CH376 are structurally similar, differing in a single carbon atom in their structure. Thus, we have identified a key element in the structure of these compounds that will be exploited to develop analogues that we can use as either inhibitors or agonists of muscular nAChR. Funding: MINECO (SAF2015-64948-C2-1-R), CSIC (PIE-201580E109), Nutting Foundation Grant (CAVG).