C2230, a preferential use- and state-dependent CaV2.2 channel blocker, mitigates pain behaviors across multiple pain models
Cheng Tang, … , Olga A. Korczeniewska, Rajesh Khanna
Cheng Tang, … , Olga A. Korczeniewska, Rajesh Khanna
Published December 10, 2024
Citation Information: J Clin Invest. 2025;135(4):e177429. https://doi.org/10.1172/JCI177429.
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Research Article Neuroscience

C2230, a preferential use- and state-dependent CaV2.2 channel blocker, mitigates pain behaviors across multiple pain models

Abstract

Antagonists — such as Ziconotide and Gabapentin — of the CaV2.2 (N-type) calcium channels are used clinically as analgesics for chronic pain. However, their use is limited by narrow therapeutic windows, difficult dosing routes (Ziconotide), misuse, and overdoses (Gabapentin), as well as a litany of adverse effects. Expansion of novel pain therapeutics may emerge from mechanism-based interrogation of CaV2.2. Here, we report the identification of C2230, an aryloxy-hydroxypropylamine, as a CaV2.2 blocker. C2230 trapped and stabilized inactivated CaV2.2 in a slow-recovering state and accelerated the open-state inactivation of the channel, conferring an advantageous use-dependent inhibition profile. C2230 inhibited CaV2.2 during high-frequency stimulation, while sparing other voltage-gated ion channels. C2230 inhibited CaV2.2 in dorsal root and trigeminal ganglia neurons from rats, marmosets, and humans in a G-protein-coupled-receptor–independent manner. Further, C2230 reduced evoked excitatory postsynaptic currents and excitatory neurotransmitter release in the spinal cord, leading to relief of neuropathic, orofacial, and osteoarthritic pain-like behaviors via 3 different routes of administration. C2230 also decreased fiber photometry-based calcium responses in the parabrachial nucleus, mitigated aversive behavioral responses to mechanical stimuli after neuropathic injury, and preserved protective pain responses, all without affecting motor or cardiovascular function. Finally, site-directed mutation analysis demonstrated that C2230 binds differently than other known CaV2.2 blockers, making it a promising lead compound for analgesic development.

Authors

Cheng Tang, Kimberly Gomez, Yan Chen, Heather N. Allen, Sara Hestehave, Erick J. Rodríguez-Palma, Santiago Loya-Lopez, Aida Calderon-Rivera, Paz Duran, Tyler S. Nelson, Siva Rama Raju Kanumuri, Bijal Shah, Nihar R. Panigrahi, Samantha Perez-Miller, Morgan K. Schackmuth, Shivani Ruparel, Amol Patwardhan, Theodore J. Price, Paramjit S. Arora, Ravindra K. Sharma, Abhisheak Sharma, Jie Yu, Olga A. Korczeniewska, Rajesh Khanna

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Figure 4

C2230 inhibits total calcium currents in rat and marmoset trigeminal sensory neurons.

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C2230 inhibits total calcium currents in rat and marmoset trigeminal sen...
(A) Representative traces of total calcium currents from rat trigeminal ganglia (TG) neurons treated with 0.1% DMSO (control; blue), 20 μM C2230 (orange), 500 nM ω-conotoxin-GVIA (ω-Ctx-GVIA; burgundy), C2230 + ω-Ctx-GVIA (green), or ω-Ctx-GVIA + C2230 (dark purple). (B) Summary of total ICa2+ density-voltage relationship. (C) Bar graphs of peak total ICa2+ density from rat TGs -treated as indicated. P values as indicated, Kruskal-Wallis test followed by Dunn’s post hoc test, n = 9–13 cells per condition from 3 independent experiments. (D) Time-course of CaV2.2 current inhibition by sequential perfusion of C2230 and ω-Ctx-GVIA. Inset: Bar graph illustrating the normalized current (Norm I) of each condition at the indicated time points. P values as indicated, 1-way ANOVA followed by Tukey multiple comparison test, n = 3–5 cells per condition from 2 independent experiments. (E) Time-course of CaV2.2 currents inhibition by sequential perfusion of ω-Ctx-GVIA and C2230 perfusion. Inset: Bar graph illustrating the normalized current (Norm I) at the time points as indicated. P values as indicated, 1-way ANOVA followed by Tukey multiple comparison test, n = 5 cells per condition from 2 independent experiments. (F) Representative traces of total calcium currents from marmoset TGs incubated with 0.1% DMSO (control; blue circles), 20 μM C2230 (orange squares), or 500 nM ω-Ctx-GVIA (burgundy hexagons). (G) Summary of total ICa2+ density-voltage relationship. (H) Bar graphs of peak total ICa2+ density from marmoset TGs pretreated as indicated. P values as indicated, Kruskal-Wallis test followed by Dunn’s post hoc test, n = 10–13 cells per condition from 1 independent experiment. Error bars indicate mean ± SEM. See Supplemental Table 3 for full statistical analysis.