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Knockdown siRNA Targeting the Mitochondrial Sodium-Calcium Exchanger-1 Inhibits the Protective Effects of Two Cannabinoids Against Acute Paclitaxel Toxicity

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Abstract

Treatment with cannabidiol (CBD) or KLS-13019 (novel CBD analog), has previously been shown to prevent paclitaxel-induced mechanical allodynia in a mouse model of chemotherapy-induced peripheral neuropathy (CIPN). The mechanism of action for CBD- and KLS-13019-mediated protection now has been explored with dissociated dorsal root ganglion (DRG) cultures using small interfering RNA (siRNA) to the mitochondrial Na+ Ca2+ exchanger-1 (mNCX-1). Treatment with this siRNA produced a 50–55% decrease in the immunoreactive (IR) area for mNCX-1 in neuronal cell bodies and a 72–80% decrease in neuritic IR area as determined with high-content image analysis. After treatment with 100 nM KLS-13019 and siRNA, DRG cultures exhibited a 75 ± 5% decrease in protection from paclitaxel-induced toxicity; whereas siRNA studies with 10 μM CBD produced a 74 ± 3% decrease in protection. Treatment with mNCX-1 siRNA alone did not produce toxicity. The protective action of cannabidiol and KLS-13019 against paclitaxel-induced toxicity during a 5-h test period was significantly attenuated after a 4-day knockdown of mNCX-1 that was not attributable to toxicity. These data indicate that decreases in neuritic mNCX-1 corresponded closely with decreased protection after siRNA treatment. Pharmacological blockade of mNCX-1 with CGP-37157 produced complete inhibition of cannabinoid-mediated protection from paclitaxel in DRG cultures, supporting the observed siRNA effects on mechanism.

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Summary

Based on the high-content analysis of decreases in mNCX-1 immunoreactive area in DRG neurons, the siRNA approach confirmed the importance of this target in producing acute (5 h) protective actions of CBD and KLS-13019. Coupled with the pharmacological studies, the target knockdown observations suggested that with both cannabinoids, the mNCX-1 target was sufficient to mediate the protection from toxicity produced by paclitaxel in sensory neurons. An unexpected finding was the identification of neuritic mitochondria as a target for the protective action of these cannabinoids. A working hypothesis emerging from these studies is that the cellular location of a target may be of equal importance to the identity of that target, thereby completing or helping to define the mechanistic concept.

Funding

These studies were supported by a Grant from the National Institute on Drug Abuse of the National Institutes of Health (R41DA044898).

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Authors and Affiliations

  1. Advanced Neural Dynamics, Inc, Pennsylvania Biotechnology Center, 3805 Old Easton Road, Doylestown, PA, 18902, USA

    Douglas E. Brenneman

  2. Kannalife Sciences, Inc, Pennsylvania Biotechnology Center, Doylestown, PA, 18902, USA

    Douglas E. Brenneman & William A. Kinney

  3. Center for Substance Abuse Research, Department of Pharmacology, Lewis Katz School of Medicine, Temple University, Philadelphia, PA, 19140, USA

    Sara Jane Ward

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  1. Douglas E. Brenneman
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  2. William A. Kinney
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  3. Sara Jane Ward
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Correspondence to Douglas E. Brenneman.

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Brenneman, D.E., Kinney, W.A. & Ward, S.J. Knockdown siRNA Targeting the Mitochondrial Sodium-Calcium Exchanger-1 Inhibits the Protective Effects of Two Cannabinoids Against Acute Paclitaxel Toxicity. J Mol Neurosci 68, 603–619 (2019). https://doi.org/10.1007/s12031-019-01321-z

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