Unveiled cold sensitivity of TRPA1 by the prolyl hydroxylation inhibition-induced sensitization to ROS in oxaliplatin-induced acute peripheral neuropathy

[Speaker] Takayuki Nakagawa:1
[Co-author] Takahito Miyake:2, Saki Nakamura:2, Meng Zhao:2, Kanako So:2, Hisashi Shirakawa:2, Yasuo Mori:3, Shuji Kaneko:2
1:Department of Clinical Pharmacology and Therapeutics, Kyoto University Hospital, Japan, 2:Department of Molecular Pharmacology, Graduate School of Pharmaceutical Sciences, Kyoto University, Japan, 3:Department of Synthetic Chemistry and Biological Chemistry, Graduate School of Enginnering, Kyoto University, Japan

Background: Chemotherapy-induced peripheral neuropathy (CIPN) is a common adverse effect of several chemotherapeutic agents. Recent accumulating evidence suggests that some transient receptor potential (TRP) channels are responsible for CIPN. Oxaliplatin, a platinum chemotherapeutic agent, induces a peculiar cold-triggered CIPN in almost all patients within hours after its infusion. We previously reported that oxaliplatin elicits rapid-onset cold hypersensitivity via TRP ankyrin 1 (TRPA1) in mice (Mol Pain, 2012). In this study, we further explored the molecular mechanisms underlying TRPA1 in oxaliplatin-induced acute cold hypersensitivity.

Methods: Whole-cell patch-clamp recordings or Ca2+ imaging experiments were performed in primary cultured DRG neurons or HEK293 cells expressing hTRPA1 or its mutants. C57BL6/J male mice (6-8 weeks) were injected with oxaliplatin (5 mg/kg, i.p.). Cold sensitivity was scored in the cold-plate test.

Results: In hTRPA1-expressing cells, treatment with oxaliplatin (100 μM) or its metabolite analog, dimethyl oxalate (DMO), for 2 h enhanced H2O2-evoked TRPA1 activation, suggesting TRPA1 sensitization to reactive oxygen species (ROS). Although hTRPA1 showed little responses to cold, oxaliplatin or DMO pretreatment induced cold-evoked hTRPA1 response, which was mediated through ROS-mediated oxidation. TRPA1 is activated by ROS through oxidative modification of cysteine residues, but also by relief from prolyl hydroxylase (PHD)-mediated hydroxylation of a proline residue (P394). Thus, we determined whether the oxaliplatin-induced hTRPA1 sensitization to cold/ROS is PHD inhibition-dependent. We found that oxaliplatin or DMO has an ability to inhibit PHD activity, and the TRPA1 sensitization to cold or ROS was inhibited in a hTRPA1 proline mutant resistant to PHDs (P394A) or by overexpressing PHD2. Furthermore, a PHD inhibitor or P394A mutation mimicked the cold-evoked responses. Finally, we confirmed our in vitro findings could fit an in vivo mechanism of TRPA1 in mice.

Conclusions: These results suggest that oxaliplatin or its metabolite inhibits the PHD-mediated prolyl hydroxylation of TRPA1 and enables the channel to sense cold-evoked mitochondria-derived ROS, which endows TRPA1 with indirect cold sensitivity in oxaliplatin-induced acute peripheral neuropathy.
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