Hsp90 Inhibition in the Spinal Cord: Novel Strategies and Targets for Opioid Treatment

Abstract

Pain is defined as an unpleasant sensory and affective experience typically associated with tissue damage. As such, it is often prevented, avoided, or attenuated when possible. Acute pain signals organisms to avoid noxious stimuli, but chronic pain is maladaptive and may still be experienced long after the harmful stimuli are no longer present. In the treatment of both acute and chronic pain, opioids are often and widely prescribed, as they can be highly potent and efficacious. Opioids inhibit neuronal signaling that transmits nociceptive information from peripheral nerve endings to the central nervous system (CNS). Importantly, opioids also produce a wide range of side effects, including constipation, reward, tolerance, addiction, respiratory depression, and death. Considering the increase in the prevalence of opioid-related deaths and opioid use disorder diagnoses in the U.S. today, seeking safer alternatives or co-therapeutics to treat acute and chronic pain is a worthwhile venture. Recent studies from our lab and others have shown that non-selectively inhibiting heat shock protein 90 (Hsp90) in the spinal cord augments the potency of systemically administered opioids in mouse models of acute and chronic pain. Because varied isoforms of Hsp90 are expressed differentially in diverse tissues, the resultant effects of nonselective Hsp90 inhibition vary by route of administration and the compartments and cell types in which they act. Nonspecific inhibition of Hsp90 in the brain blocks opioid-induced antinociception but, when delivered intrathecally, systemic opioid potency is increased. This led our lab to believe that the actions of specific Hsp90 isoforms play different roles in modulating opioid signaling cascades in a context-specific manner. Thus, with the treatment of isoform-specific small molecule Hsp90 inhibitors, isoforms expressed specifically in the spinal cord, but not the brain, could be targeted and subsequent enhancement of opioid antinociception occurred even when they were delivered systemically. These isoform-specific small molecule inhibitors of Hsp90 show high affinity, potency, solubility, and distribution, and when co-administered with morphine, a reduced or unchanged side-effect profile compared to morphine alone. The mechanisms by which these outcomes occur are incompletely understood. However, much light has been shed on the mechanisms involved in this phenomenon. Using a proteomic approach we found that, 24hrs after spinal administration of the nonselective Hsp90 inhibitor, sodium-and-chloride dependent GABA reuptake transporter 2 (GAT-2) expression was upregulated. Upon further analysis, the pharmacological blockade of this protein attenuated the enhancement of opioid analgesia from Hsp90 inhibition in the spinal cord. To target GAT-2 more specifically, GAT-2 was knocked out via CRISPR/Cas9 in the spinal cord. After which, again the effect of Hsp90 inhibition on morphine was absent. Further, to distinguish between GABA-A and GABA-B mediated mechanisms, GABA agonists and antagonists were administered intrathecally and showed sex-specific differences in the role of GABAergic transmission in influencing Hsp90-modulated opioid antinociception. These data suggest that, in part, that GABA signaling in the spinal cord exerts a mediating role in the enhancement of opioid analgesia resulting from Hsp90 inhibition. The details of the underlying circuitry engaged in this mechanism are explicated in further detail later in this paper. In summary, isoform-selective Hsp90 inhibitors have been revealed to be promising co-therapeutics to offset or abolish the harmful side effects of commonly prescribed opioids, and the mechanism by which they produce these effects is likely by disabling a tonically inhibitory GABAergic projection that inhibits opioidergic interneurons in the dorsal horn of the spinal cord.