Small Molecule Positive Allosteric Modulation of TRPV1 Activation by Vanilloids and Acidic pH

Abstract

Transient receptor potential cation channel subfamily V member 1 (TRPV1) is a high-conductance, nonselective cation channel strongly expressed in nociceptive primary afferent neurons of the peripheral nervous system and functions as a multimodal nociceptor gated by temperatures greater than 43°C, protons, and smallmolecule vanilloid ligands such as capsaicin. The ability to respond to heat, low pH, vanilloids, and endovanilloids and altered sensitivity and expression in experimental inflammatory and neuropathic pain models made TRPV1 a major target for the development of novel, nonopioid analgesics and resulted in the discovery of potent antagonists. In human clinical trials, observations of hyperthermia and the potential for thermal damage by suppressing the ability to sense noxious heat suggested that full-scale blockade of TRPV1 function can be counterproductive and subtler pharmacological approaches are necessary. Here we show that the dihydropyridine derivative 4,5-diethyl-3-(2-methoxyethylthio)-2-methyl-6-phenyl-1,4-(Ϯ)-dihydropyridine-3,5-dicarboxylate (MRS1477) behaves as a positive allosteric modulator of both proton and vanilloid activation of TRPV1. Under inflammatorymimetic conditions of low pH (6.0) and protein kinase C phosphorylation, addition of MRS1477 further increased sensitivity of already sensitized TPRV1 toward capsaicin. MRS1477 does not affect inhibition by capsazepine or ruthenium red and remains effective in potentiating activation by pH in the presence of an orthosteric vanilloid antagonist. These results indicate a distinct site on TRPV1 for positive allosteric modulation that may bind endogenous compounds or novel pharmacological agents. Positive modulation of TRPV1 sensitivity suggests that it may be possible to produce a selective analgesia through calcium overload restricted to highly active nociceptive nerve endings at sites of tissue damage and inflammation.

Positive allosteric modulation of TRPV1 as a novel analgesic mechanism

“…We previously demonstrated that MRS1477 exhibits no agonist activity at TRPV1, but potentiates the calcium influx induced by orthosteric vanilloid agonists and TRPV1 stimulation by protons or sensitization by phorbol ester [46,47]. In the previous study, we also observed MRS1477 potentiation of intraperitoneal capsaicin-induced hypothermia [46].…”

“…In the previous study, we also observed MRS1477 potentiation of intraperitoneal capsaicin-induced hypothermia [46]. In the present study, we were able to induce analgesia through functional inactivation of peripheral primary afferent nerve terminals similar to the long-lasting effect of the potent TRPV1 agonist RTX, which served as our positive control [8].…”

“…In chronic pain states, nociceptive nerve endings remain active and do not transition into an inactive state without pharmacological intervention, since the endings have multiple mechanisms to cope with transmembrane calcium flux [62]. Thus, despite a low pH and an endovanilloidenriched environment, the added "push" from a PAM would be necessary to achieve nerve terminal inactivation [46].…”

“…In an effort to combine the ease of systemic administration of antagonists with the potent, long-lasting analgesic qualities of agonists, we are exploring positive allosteric modulation of TRPV1 as a novel analgesic mechanism [46]. An ideal TRPV1 positive allosteric modulator (PAM) would have no intrinsic activity but would enhance receptor activation by an orthosteric agonist and, as a result, would act conditionally to potentiate only TRPV1 on afferent nerves terminating in regions of tissue damage or inflammation where channels are highly active (see Figure 1).…”

“…Here, we present in vivo evidence that it is possible to positively modulate agonist-activated TRPV1-expressing nerve terminals to the point of deafferentation. MRS1477, a small molecule 1,4-dihydropyridine that we have previously identified [47] and characterized [46] in vitro, was used to potentiate capsaicin-activated nerve terminals in the rat hind paw. We examined analgesic effects of terminal inactivation by measuring acute thermal nociception and changes in gene expression markers of peripheral nerve terminal ablation, damage, or axotomy in lumbar dorsal root ganglia.…”