A3 adenosine receptor agonist prevents the development of paclitaxel-induced neuropathic pain by modulating spinal glial-restricted redox-dependent signaling pathways

Abstract

Chemotherapy-induced peripheral neuropathy accompanied by chronic neuropathic pain is the major dose-limiting toxicity of several anticancer agents including the taxane paclitaxel (Taxol). A critical mechanism underlying paclitaxel-induced neuropathic pain is the increased production of peroxynitrite in spinal cord generated in response to activation of the superoxide-generating enzyme, NADPH oxidase. Peroxynitrite in turn contributes to the development of neuropathic pain by modulating several redox-dependent events in spinal cord. We recently reported that activation of the Gi/Gq-coupled A3 adenosine receptor (A3AR) with selective A3AR agonists (ie, IB-MECA) blocked the development of chemotherapy induced-neuropathic pain evoked by distinct agents, including paclitaxel, without interfering with anticancer effects. The mechanism or mechanisms of action underlying these beneficial effects has yet to be explored. We now demonstrate that IB-MECA attenuates the development of paclitaxel-induced neuropathic pain by inhibiting the activation of spinal NADPH oxidase and two downstream redox-dependent systems. The first relies on inhibition of the redox-sensitive transcription factor (NFκB) and mitogen activated protein kinases (ERK and p38) resulting in decreased production of neuroexcitatory/proinflammatory cytokines (TNF-α, IL-1β) and increased formation of the neuroprotective/anti-inflammatory IL-10. The second involves inhibition of redox-mediated posttranslational tyrosine nitration and modification (inactivation) of glia-restricted proteins known to play key roles in regulating synaptic glutamate homeostasis: the glutamate transporter GLT-1 and glutamine synthetase. Our results unravel a mechanistic link into biomolecular signaling pathways employed by A3AR activation in neuropathic pain while providing the foundation to consider use of A3AR agonists as therapeutic agents in patients with chemotherapy-induced peripheral neuropathy.

Keywords: A3; Adenosine; Chemotherapy-induced peripheral neuropathy; Neuroinflammation; Neuropathic pain; Paclitaxel; Spinal cord.

Conflict of interest statement

The authors claim no conflicts of interest.

Copyright © 2014 International Association for the Study of Pain. All rights reserved.

Figures

Fig. 1. IB-MECA prevents paclitaxel-induced mechano-hypersensitivity and spinal NADPH oxidase activity

As compared to baseline on D16, administration of paclitaxel (P, black bars) but not vehicle (V, open bars) leads to the development of mechano-allodynia (A) and mechano–hyperalgesia (B). Spinal cords from paclitaxel-treated animals displayed higher levels of NADPH oxidase activity than their vehicle-treated counterparts (C). These paclitaxel-induced events were blocked by daily administration of the A3AR agonist IB-MECA (D0–15, 0.1 mg/kg/d; dark gray bars, A–C). The selective A3AR antagonist MRS1523 (2 mg/kg/d, D0–15, dark gray hatched bars) prevented IB-MECA’s beneficial actions (A–C). MRS1523 (white hatched bars) administered on its own to paclitaxel animals had no effect on PWTs (A,B). Results are expressed as mean ± SD for n=5–6 rats and analyzed by one-way ANOVA with Dunnett’s comparisons. *P<0.05 vs. Vehicle; †P<0.05 vs. Paclitaxel

Fig. 2. IB-MECA prevents activation of NFκB and MAPKs in the spinal cord during paclitaxel-induced neuropathic pain

When compared to vehicle (V, open bars), administration of paclitaxel (P, black bars) increased IκBα degradation (A), cytosolic phosphorylation of NFκB p65 (B), nuclear translocation of NFκB p65 (C), and phosphorylation of ERK1/2 (D) and p38 (E). These paclitaxel-induced events were blocked by daily administration of IB-MECA (D0–15, 0.1 mg/kg/d; gray bars, A–E). Representative blots are shown. Results are expressed as mean ± SD for n=5–6 rats and analyzed by one-way ANOVA with Dunnett’s comparisons. *P<0.05 vs. Vehicle; †P<0.05 vs. Paclitaxel

Fig. 3. IB-MECA decreases paclitaxel-induced spinal formation of pro-inflammatory TNF-α and IL-1β and increases the anti-inflammatory IL-10

On D16, IB-MECA (0.1 mg/kg/d; D0–15; dark gray bars) attenuated paclitaxel-induced (P, black bars) elevations in TNF-α and IL-1β expression (A) and increased the levels of the anti-inflammatory cytokine, IL-10 (B) as compared to vehicle (V, open bars). The A3AR antagonist MRS1523 blocked IB-MECA’s effects (dark gray hatched bars) but had no effect on vehicle-treated (light gray bars) animals (A,B). Results are expressed as mean ± SD for n=6 and analyzed by one-way ANOVA with Dunnett’s post hoc comparisons. *P<0.05 vs. Vehicle; †P<0.05 vs. Paclitaxel

Fig. 4. IB-MECA prevents post-translational nitration of GLT-1 & GS in the spinal cord during paclitaxel-induced neuropathic pain

Compared with vehicle (V, open bars), paclitaxel (P, black bars) led to significant nitration of the glutamate transporter GLT-1 (A) and GS (B) in spinal cord tissues, events attenuated in rats treated with IB-MECA (0.1 mg/kg/d, D0–15; gray bars). Representative blots are shown. Results are expressed as mean ± SD for n=5 rats and analyzed by one-way ANOVA with Dunnett’s comparisons. *P<0.05 vs. Vehicle; †P<0.05 vs. Paclitaxel

Fig. 5. Proposed schematic representation of mechanisms underlying IB-MECA’s beneficial actions on CIPN

Chemotherapy (paclitaxel)-induced neuropathic pain is associated with increased NADPH oxidase activity within the spinal cord contributing to enhanced peroxynitrite (PN) production. Due to its ability to post-translationally modify protein function, peroxynitrite could be a driver behind many of the spinal neuropathological changes underlying CIPN including 1) the nitration/inactivation of glutamate transporter 1 (GLT-1) and glutamine synthetase (GS) and 2) the activation of redox-dependent signaling pathways (NFκB and MAPK) leading to a surge in glial-associated pro-inflammatory cytokine production (TNF-α and IL-1β). Treatment with the selective A3AR agonist, IB-MECA, not only inhibits paclitaxel-induced pain and the associated spinal events, it also increases the formation of the neuroprotective/anti-inflammatory cytokine, IL-10.