Distinct Mechanism of Cysteine Oxidation-Dependent Activation and Cold Sensitization of Human Transient Receptor Potential Ankyrin 1 Channel by High and Low Oxaliplatin

Takahito Miyake, Saki Nakamura, Zhao Meng, Satoshi Hamano, Keisuke Inoue, Tomohiro Numata, Nobuaki Takahashi, Kazuki Nagayasu, Hisashi Shirakawa, Yasuo Mori, Takayuki Nakagawa, Shuji Kaneko

Oxaliplatin, a third-generation platinum-based chemotherapeutic agent, displays unique acute peripheral neuropathy triggered or enhanced by cold, and accumulating evidence suggests that transient receptor potential ankyrin 1 (TRPA1) is responsible. TRPA1 is activated by oxaliplatin via a glutathione-sensitive mechanism. However, oxaliplatin interrupts hydroxylation of a proline residue located in the N-terminal region of TRPA1 via inhibition of prolyl hydroxylase (PHD), which causes sensitization of TRPA1 to reactive oxygen species (ROS). Furthermore, PHD inhibition endows cold-insensitive human TRPA1 (hTRPA1) with ROS-dependent cold sensitivity. Since cysteine oxidation and proline hydroxylation regulate its activity, their association with oxaliplatin-induced TRPA1 activation and acquirement of cold sensitivity were investigated in the present study. A high concentration of oxaliplatin (1 mM) induced outward-rectifier whole-cell currents and increased the intracellular Ca(2+) concentration in hTRPA1-expressing HEK293 cells, but did not increase the probability of hTRPA1 channel opening in the inside-out configuration. Oxaliplatin also induced the rapid generation of hydrogen peroxide, and the resultant Ca(2+) influx was prevented in the presence of glutathione and in cysteine-mutated hTRPA1 (Cys641Ser)-expressing cells, whereas proline-mutated hTRPA1 (Pro394Ala)-expressing cells showed similar whole-cell currents and Ca(2+) influx. By contrast, a lower concentration of oxaliplatin (100 μM) did not increase the intracellular Ca(2+) concentration but did confer cold sensitivity on hTRPA1-expressing cells, and this was inhibited by PHD2 co-overexpression. Cold sensitivity was abolished by the mitochondria-targeting ROS scavenger mitoTEMPO and was minimal in cysteine-mutated hTRPA1 (Cys641Ser or Cys665Ser)-expressing cells. Thus, high oxaliplatin evokes ROS-mediated cysteine oxidation-dependent hTRPA1 activation independent of PHD activity, while a lower concentration induces cold-induced cysteine oxidation-dependent opening of hTRPA1 via PHD inhibition.