Targeting CYP2J to reduce paclitaxel-induced peripheral neuropathic pain.

Chemotherapy-induced peripheral neuropathic pain (CIPNP) is a severe dose- and therapy-limiting side effect of widely used cytostatics that is particularly difficult to treat. Here, we report increased expression of the cytochrome-P450-epoxygenase CYP2J6 and increased concentrations of its linoleic acid metabolite 9,10-EpOME (9,10-epoxy-12Z-octadecenoic acid) in dorsal root ganglia (DRGs) of paclitaxel-treated mice as a model of CIPNP. The lipid sensitizes TRPV1 ion channels in primary sensory neurons and causes increased frequency of spontaneous excitatory postsynaptic currents in spinal cord nociceptive neurons, increased CGRP release from sciatic nerves and DRGs, and a reduction in mechanical and thermal pain hypersensitivity. In a drug repurposing screen targeting CYP2J2, the human ortholog of murine CYP2J6, we identified telmisartan, a widely used angiotensin II receptor antagonist, as a potent inhibitor. In a translational approach, administration of telmisartan reduces EpOME concentrations in DRGs and in plasma and reverses mechanical hypersensitivity in paclitaxel-treated mice. We therefore suggest inhibition of CYP2J isoforms with telmisartan as a treatment option for paclitaxel-induced neuropathic pain.

N-naphthoyl-beta-naltrexamine (NNTA), a highly selective and potent activator of µ/kappa-opioid heteromers.

Numerous G protein-coupled receptors (GPCRs) have been shown to form heteromeric receptors in cell-based assays. Among the many heteromers reported in the opioid receptor family are µ/κ, κ/δ, and µ/δ. However, the in vivo physiological and behavioral relevance for the proposed heteromers have not yet been established. Here we report a unique example of a ligand, N-naphthoyl-ß-naltrexamine (NNTA) that selectively activates heteromeric µ/κ-opioid receptors in HEK-293 cells and induces potent antinociception in mice. NNTA was an exceptionally potent agonist in cells expressing µ/κ-opioid receptors. Intriguingly, it was found to be a potent antagonist in cells expressing only µ-receptors. In the mouse tail-flick assay, intrathecal (i.t.) NNTA produced antinociception that was ~100-fold greater than by intracerebroventricular (i.c.v.) administration. The κ-antagonist, norBNI, decreased the i.t. potency, and the activity was virtually abolished in µ-opioid receptor knockout mice. No tolerance was induced i.t., but marginal tolerance (3-fold) was observed via the i.c.v. route. Moreover, NNTA produced neither significant physical dependence nor place preference in the ED50 dose range. Taken together, this work provides an important pharmacologic tool for investigating the in vivo functional relevance of heteromeric µ/κ-opioid receptors and suggests an approach to potent analgesics with fewer deleterious side effects.

Bortezomib-induced neuropathy is in part mediated by the sensitization of TRPV1 channels.

TRPV1 is an ion channel that transduces noxious heat and chemical stimuli and is expressed in small fiber primary sensory neurons that represent almost half of skin nerve terminals. Tissue injury and inflammation result in the sensitization of TRPV1 and sustained activation of TRPV1 can lead to cellular toxicity though calcium influx. To identify signals that trigger TRPV1 sensitization after a 24-h exposure, we developed a phenotypic assay in mouse primary sensory neurons and performed an unbiased screen with a compound library of 480 diverse bioactive compounds. Chemotherapeutic agents, calcium ion deregulators and protein synthesis inhibitors were long-acting TRPV1 sensitizers. Amongst the strongest TRPV1 sensitizers were proteasome inhibitors, a class that includes bortezomib, a chemotherapeutic agent that causes small fiber neuropathy in 30-50% of patients. Prolonged exposure of bortezomib produced a TRPV1 sensitization that lasted several days and neurite retraction in vitro and histological and behavioral changes in male mice in vivo. TRPV1 knockout mice were protected from epidermal nerve fiber loss and a loss of sensory discrimination after bortezomib treatment. We conclude that long-term TRPV1 sensitization contributes to the development of bortezomib-induced neuropathy and the consequent loss of sensation, major deficits experienced by patients under this chemotherapeutic agent.

Breaking barriers to novel analgesic drug development.

This corrects the article DOI: 10.1038/nrd.2017.87.

Breaking barriers to novel analgesic drug development.

Acute and chronic pain complaints, although common, are generally poorly served by existing therapies. This unmet clinical need reflects a failure to develop novel classes of analgesics with superior efficacy, diminished adverse effects and a lower abuse liability than those currently available. Reasons for this include the heterogeneity of clinical pain conditions, the complexity and diversity of underlying pathophysiological mechanisms, and the unreliability of some preclinical pain models. However, recent advances in our understanding of the neurobiology of pain are beginning to offer opportunities for developing novel therapeutic strategies and revisiting existing targets, including modulating ion channels, enzymes and G-protein-coupled receptors.

Two to tango: GPCR oligomers and GPCR-TRP channel interactions in nociception.

G protein coupled receptors (GPCRs) represent the largest family of cell surface receptors that are involved in regulating several physiological and behavioral responses in organisms. Indeed, over half of all the approved drugs on the market target GPCRs. Over the past twenty years, several lines of evidence have suggested that GPCRs associate to form oligomeric structures that substantially expand the complexity of signaling processes in vivo. In addition, GPCRs have also been shown to functionally regulate ion channels and help fine-tune neurotransmission. In this review, we will discuss recent advances in both mechanisms, with specific focus on opioid receptors, cannabinoid receptors and transient receptor potential (TRP) calcium channels in nociception. A better understanding of such mechanisms will be imperative in designing analgesics devoid of deleterious side effects and mitigating drug abuse.

An immunocytochemical-derived correlate for evaluating the bridging of heteromeric mu-delta opioid protomers by bivalent ligands.

Bivalent ligands that contain two pharmacophores linked by a spacer are promising tools to investigate the pharmacology of opioid receptor heteromers. Evidence for occupation of neighboring protomers by two phamacophores of a single bivalent ligand (bridging) has relied mainly on pharmacological data. In the present study, we have employed an immunocytochemical correlate to support in vivo biological studies that are consistent with bridging. We show that a bivalent mu agonist/delta antagonist (MDAN-21) that is devoid of tolerance due to possible bridging of mu and delta protomers prevents endocytosis of the heteromeric receptors in HEK-293 cells. Conversely, a bivalent ligand (MDAN-16) with a short spacer or monovalent mu agonist give rise to robust internalization. The data suggest that the immobilization of proximal mu and delta protomers is due to bridging by MDAN-21. The finding that MDAN-21 and its shorter spacer homologue MDAN-16 possess equivalent activity in HEK-293 cells, but produce dramatically divergent internalization of mu-delta heteromer, is relevant to the role of internalization and tolerance.

Clinically employed opioid analgesics produce antinociception via µ-δ opioid receptor heteromers in Rhesus monkeys.

Morphine and related drugs are widely employed as analgesics despite the side effects associated with their use. Although morphine is thought to mediate analgesia through mu opioid receptors, delta opioid receptors have been implicated in mediating some side effects such as tolerance and dependence. Here we present evidence in rhesus monkeys that morphine, fentanyl, and possibly methadone selectively activate mu-delta heteromers to produce antinociception that is potently antagonized by the delta opioid receptor antagonist, naltrindole (NTI). Studies with HEK293 cells expressing mu-delta heteromeric opioid receptors exhibit a similar antagonism profile of receptor activation in the presence of NTI. In mice, morphine was potently inhibited by naltrindole when administered intrathecally, but not intracerebroventricularly, suggesting the possible involvement of mu-delta heteromers in the spinal cord of rodents. Taken together, these results strongly suggest that, in primates, mu-delta heteromers are allosterically coupled and mediate the antinociceptive effects of three clinically employed opioid analgesics that have been traditionally viewed as mu-selective. Given the known involvement of delta receptors in morphine tolerance and dependence, our results implicate mu-delta heteromers in mediating both antinociception and these side effects in primates. These results open the door for further investigation in humans.

The δ opioid receptor agonist SNC80 selectively activates heteromeric µ-δ opioid receptors.

Coexpressed and colocalized µ- and δ-opioid receptors have been established to exist as heteromers in cultured cells and in vivo. However the biological significance of opioid receptor heteromer activation is less clear. To explore this significance, the efficacy of selective activation of opioid receptors by SNC80 was assessed in vitro in cells singly and coexpressing opioid receptors using a chimeric G-protein-mediated calcium fluorescence assay, SNC80 produced a substantially more robust response in cells expressing µ-δ heteromers than in all other cell lines. Intrathecal SNC80 administration in µ- and δ-opioid receptor knockout mice produced diminished antinociceptive activity compared with wild type. The combined in vivo and in vitro results suggest that SNC80 selectively activates µ-δ heteromers to produce maximal antinociception. These data contrast with the current view that SNC80 selectively activates δ-opioid receptor homomers to produce antinociception. Thus, the data suggest that heteromeric µ-δ receptors should be considered as a target when SNC80 is employed as a pharmacological tool in vivo.

Standard opioid agonists activate heteromeric opioid receptors: evidence for morphine and [d-Ala(2)-MePhe(4)-Glyol(5)]enkephalin as selective µ-δ agonists.

Research in the opioid field has relied heavily on the use of standard agonist ligands such as morphine, [d-Ala(2)-MePhe(4)-Glyol(5)]enkephalin (DAMGO), U69593, bremazocine, [d-Pen(2)d-Pen(5)]enkephalin (DPDPE), and deltorphin-II as tools for investigating the three major types of opioid receptors, MOP (µ), KOP (κ), and DOP (δ), that mediate antinociception. The functional selectivity of these ligands has been based on the assumption that opioid receptors exist as homomers. As numerous studies in cultured cells have suggested that opioid receptors can associate both as homomers and heteromers, we have investigated the selectivity of these standard ligands using intracellular calcium release and [(35)S]GTPγS assays in HEK-293 cells that contain singly and coexpressed opioid receptors. The present study reveals that morphine and DAMGO, traditionally classified as µ selective agonists, selectively activate µ-δ heteromeric opioid receptors with greater efficacy than homomeric opioid receptors. Moreover, standard ligands that have been widely employed as κ- and δ-selective agonists display little or no differences in the activation of homomeric and heteromeric opioid receptors. The far-reaching implications of these results are discussed.