Molecular Basis for Allosteric Inhibition of Acid-Sensing Ion Channel 1a by Ibuprofen

Abstract

A growing body of evidence links certain aspects of nonsteroidal anti-inflammatory drug (NSAID) pharmacology with acid-sensing ion channels (ASICs), a small family of excitatory neurotransmitter receptors implicated in pain and neuroinflammation. The molecular basis of NSAID inhibition of ASICs has remained unknown, hindering the exploration of this line of therapy. Here, we characterized the mechanism of inhibition, explored the molecular determinants of sensitivity, and sought to establish informative structure-activity relationships, using electrophysiology, site-directed mutagenesis, and voltage-clamp fluorometry. Our results show that ibuprofen is an allosteric inhibitor of ASIC1a, which binds to a crucial site in the agonist transduction pathway and causes conformational changes that oppose channel activation. Ibuprofen inhibits several ASIC subtypes, but certain ibuprofen derivatives show some selectivity for ASIC1a over ASIC2a and vice versa. These results thus define the NSAID/ASIC interaction and pave the way for small-molecule drug design targeting pain and inflammation.

Divalent cation and chloride ion sites of chicken acid sensing ion channel 1a elucidated by x-ray crystallography

“…Placement of a Cl - ion at this high pH binding site off of Lys 77 and Lys 422 partially occludes the narrow extracellular fenestrations that provide access for Na + ions to the channel pore ( Fig 5D and 5E ) and that undergo a dramatic expansion upon channel activation[ 23 ]. Furthermore, a recent study demonstrated that Lys 422, positioned less than 3 Å from the high pH Cl - site, is important for inhibition of ASIC1a by ibuprofen[ 35 ], indicating that this region of the channel is critical for proton-dependent gating and serves as a binding site for channel modulators. Moreover, while the biological significance of a high pH Cl - site on the resting channel remains unknown, the absence of a bound Cl - at the canonical low pH site in crystal structures of the resting channel at high pH demonstrates how Cl - ions, in addition to protons and divalent cations, play an important role in the pH-dependent function of ASICs.…”

Divalent cation and chloride ion sites of chicken acid sensing ion channel 1a elucidated by x-ray crystallography

“…Placement of a Cl − ion at this high pH binding site off of Lys 77 and Lys 422 partially occludes the narrow extracellular fenestrations that provide access for Na+ ions to the channel pore (Figure 5D-E) and that undergo a dramatic expansion upon channel activation(23). Furthermore, a recent study demonstrated that Lys 422, positioned less than 3 Å from the high pH Cl − site, is important for inhibition of ASIC1a by ibuprofen(35), indicating that this region of the channel is critical for proton-dependent gating and serves as a binding site for channel modulators. Moreover, while the biological significance of a high pH Cl − site on the resting channel remains unknown, the absence of a bound Cl − at the canonical low pH site in crystal structures of the resting channel at high pH demonstrates how Cl − ions, in addition to protons and divalent cations, play an important role in the pH-dependent function of ASICs.…”

Synaptic signals mediated by protons and acid‐sensing ion channels

“…Ibuprofen and flurbiprofen, for example, inhibit ASIC1a containing channels with an IC50 value of 350 µM. The proposed mechanism of action of the nonsteroidal anti-inflammatory drugs is that of allosteric inhibition of the channels by slowing the recovery after inactivation (Dorofeeva, Barygin, Staruschenko, Bolshakov, & Magazanik, 2008;Lynagh, Romero-Rojo, Lund, & Pless, 2017). Other drugs like Aspirin and salicylate inhibit ASIC3 containing channels (Voilley, 2004).…”