Chemotherapy-induced peripheral neuropathy (CIPN), characterized by pain and numbness in hands and feet, is a common side effect of cancer treatment. In most patients, symptoms of CIPN subside after treatment completion. However, in a substantial subgroup, CIPN persists long into survivorship. Impairment in pain resolution pathways may explain persistent CIPN. We investigated the contribution of T cells and endogenous interleukin (IL)-10 to resolution of CIPN. Paclitaxel-induced mechanical allodynia was prolonged in T-cell-deficient (Rag1^−/−) mice compared with wild-type (WT) mice. There were no differences between WT and Rag1^−/−mice in severity of paclitaxel-induced mechanical allodynia. Adoptive transfer of either CD3⁺or CD8⁺, but not CD4⁺, T cells to Rag1^−/−mice normalized resolution of CIPN. Paclitaxel treatment increased the number of T cells in lumbar dorsal root ganglia (DRG), where CD8⁺T cells were the major subset. Inhibition of endogenous IL-10 signaling by intrathecal injection of anti-IL-10 to WT mice or Rag1^−/−mice reconstituted with CD8⁺T cells delayed recovery from paclitaxel-induced mechanical allodynia. Recovery was also delayed in IL-10 knock-out mice. Conversely, administration of exogenous IL-10 attenuated paclitaxel-induced allodynia.In vitro, IL-10 suppressed abnormal paclitaxel-induced spontaneous discharges in DRG neurons. Paclitaxel increased DRG IL-10 receptor expression and this effect requires CD8⁺T cells. In conclusion, we identified a novel mechanism for resolution of CIPN that requires CD8⁺T cells and endogenous IL-10. We propose that CD8⁺T cells increase DRG IL-10 receptor expression and that IL-10 suppresses the abnormal paclitaxel-induced spontaneous discharges by DRG neurons to promote recovery from CIPN.SIGNIFICANCE STATEMENTChemotherapy-induced peripheral neuropathy persists after completion of cancer treatment in a significant subset of patients, whereas others recover. Persistent neuropathy after completion of cancer treatment severely affects quality of life. We propose that understanding how neuropathy resolves will identify novel avenues for treatment. We identified a novel and critical role for CD8⁺T cells and for endogenous IL-10 in recovery from paclitaxel-induced neuropathy in mice. Enhancing the capacity of CD8⁺T cells to promote resolution or increasing IL-10 signaling are promising targets for novel interventions. Clinically, peripheral blood CD8⁺T-cell function and/or the capacity of individuals to produce IL-10 may represent biomarkers of risk for developing persistent peripheral neuropathy after completion of cancer treatment.

In humans, depression is often associated with low-grade inflammation, activation of the tryptophan/kynurenine pathway, and mild lymphopenia. Preclinical research confirms that inflammation induces depression-like behavior through activation of the tryptophan/kynurenine pathway. However, the mechanisms governing recovery from depression are unknown. Understanding the pathways leading to resolution of depression will likely lead to identification of novel targets for treatment. We investigated the contribution of T lymphocytes to the resolution of lipopolysaccharide-induced depression-like behavior. Duration of depression-like behavior was markedly prolonged in mice without mature T or B lymphocytes (Rag1^−/− mice). This prolonged depression-like behavior was associated with persistent upregulation of the tryptophan-metabolizing enzyme indoleamine-2,3-dioxygenase (Ido)1 in the prefrontal cortex (PFC). Reconstitution of Rag1^−/− mice with T lymphocytes normalized resolution of depression-like behavior and expression of Ido1 in the PFC. During resolution of inflammation-induced depression-like behavior, T lymphocytes accumulated in the meninges and were required for induction of interleukin (IL)-10 in the meninges and the PFC. Inhibition of IL-10 signaling by nasal administration of neutralizing anti–IL-10 antibody to WT mice led to persistent upregulation of Ido1 in the PFC and prolonged depression-like behavior. Conversely, nasal administration of recombinant IL-10 in Rag1^−/− mice normalized Ido1 expression and resolution of depression-like behavior. In conclusion, the present data show for the first time that resolution of inflammation-induced depression is an active process requiring T lymphocytes acting via an IL-10–dependent pathway to decrease Ido1 expression in the brain. We propose that targeting the T lymphocyte/IL-10 resolution pathway could represent a novel approach to promote recovery from major depressive disorder.

Nerve injury-induced hyperactivity of primary sensory neurons in the dorsal root ganglion (DRG) contributes to chronic pain development, but the underlying epigenetic mechanisms remain poorly understood. Here we determined genome-wide changes in DNA methylation in the nervous system in neuropathic pain. Spinal nerve ligation (SNL), but not paclitaxel treatment, in male Sprague Dawley rats induced a consistent low-level hypomethylation in the CpG sites in the DRG during the acute and chronic phases of neuropathic pain. DNA methylation remodeling in the DRG occurred early after SNL and persisted for at least 3 weeks. SNL caused DNA methylation changes at 8% of CpG sites with prevailing hypomethylation outside of CpG islands, in introns, intergenic regions, and repetitive sequences. In contrast, SNL caused more gains of methylation in the spinal cord and prefrontal cortex. The DNA methylation changes in the injured DRGs recapitulated developmental reprogramming at the neonatal stage. Methylation reprogramming was correlated with increased gene expression variability. A diet deficient in methyl donors induced hypomethylation and pain hypersensitivity. Intrathecal administration of the DNA methyltransferase inhibitor RG108 caused long-lasting pain hypersensitivity. DNA methylation reprogramming in the DRG thus contributes to nerve injury-induced chronic pain. Restoring DNA methylation may represent a new therapeutic approach to treat neuropathic pain. SIGNIFICANCE STATEMENT Epigenetic mechanisms are critically involved in the transition from acute to chronic pain after nerve injury. However, genome-wide changes in DNA methylation in the nervous system and their roles in neuropathic pain development remain unclear. Here we used digital restriction enzyme analysis of methylation to quantitatively determine genome-wide DNA methylation changes caused by nerve injury. We showed that nerve injury caused DNA methylation changes at 8% of CpG sites with prevailing hypomethylation outside of CpG islands in the dorsal root ganglion. Reducing DNA methylation induced pain hypersensitivity, whereas increasing DNA methylation attenuated neuropathic pain. These findings extend our understanding of the epigenetic mechanism of chronic neuropathic pain and suggest new strategies to treat nerve injury-induced chronic pain.

AMP-activated protein kinase (AMPK) is an energy-sensing kinase that has emerged as a novel therapeutic target for pain due to its ability to inhibit mechanistic target of rapamycin (mTOR) and mitogen activated protein kinase (MAPK) signaling, two signaling pathways that are linked to pain promotion after injury as well as the development of hyperalgesic priming. MAPK and mTOR signaling are also implicated in chemotherapy induced peripheral neuropathy (CIPN). We conducted a series of experiments to gain further insight into how AMPK activators might best be used to treat pain in both sexes in the setting of CIPN from paclitaxel. We also assessed whether hyperalgesic priming emerges from paclitaxel treatment and if this can be prevented by AMPK targeting. AMPK can be pharmacologically activated indirectly through regulation of upstream kinases like liver kinase B1 (LKB1) or directly using positive allosteric modulators. We used the indirect AMPK activators metformin and narciclasine, both of which have been shown to reduce pain in preclinical models but with much different potencies and different efficacies depending on the sex of the animal. We used the direct AMPK activator MK8722 because it is the most potent and specific such activator described to date. Here, the AMPK activators were used in 2 different treatment paradigms. First the drugs were given concurrently with paclitaxel to test whether they prevent mechanical hypersensitivity. Second the AMPK activators were given after the completion of paclitaxel treatment to test whether they reverse established mechanical hypersensitivity. Consistent with our previously published findings with metformin, narciclasine (1 mg/kg) produced an anti-hyperalgesic effect, preventing paclitaxel-induced neuropathy in outbred mice of both sexes. In contrast to metformin, narciclasine also reversed mechanical hypersensitivity in established CIPN. Both metformin (200 mg/kg) and narciclasine prevented the development of hyperalgesic priming induced by paclitaxel treatment. MK8722 (30 mg/kg) had no effect on mechanical hypersensitivity caused by paclitaxel in either the prevention or reversal treatment paradigms. However, MK8722 did attenuate hyperalgesic priming in male and female mice. We conclude that paclitaxel induces robust hyperalgesic priming that is prevented by AMPK targeting and that narciclasine is a particularly attractive candidate for further development as a CIPN treatment.

Transmembrane Ca _v 2.2 (N-type) voltage-gated calcium channels are genetically and pharmacologically validated, clinically relevant pain targets. Clinical block of Ca _v 2.2 (e.g., with Prialt/Ziconotide) or indirect modulation [e.g., with gabapentinoids such as Gabapentin (GBP)] mitigates chronic pain but is encumbered by side effects and abuse liability. The cytosolic auxiliary subunit collapsin response mediator protein 2 (CRMP2) targets Ca _v 2.2 to the sensory neuron membrane and regulates their function via an intrinsically disordered motif. A CRMP2-derived peptide (CBD3) uncouples the Ca _v 2.2–CRMP2 interaction to inhibit calcium influx, transmitter release, and pain. We developed and applied a molecular dynamics approach to identify the A ₁ R ₂ dipeptide in CBD3 as the anchoring Ca _v 2.2 motif and designed pharmacophore models to screen 27 million compounds on the open-access server ZincPharmer. Of 200 curated hits, 77 compounds were assessed using depolarization‐evoked calcium influx in rat dorsal root ganglion neurons. Nine small molecules were tested electrophysiologically, while one (CBD3063) was also evaluated biochemically and behaviorally. CBD3063 uncoupled Ca _v 2.2 from CRMP2, reduced membrane Ca _v 2.2 expression and Ca ²⁺ currents, decreased neurotransmission, reduced fiber photometry-based calcium responses in response to mechanical stimulation, and reversed neuropathic and inflammatory pain across sexes in two different species without changes in sensory, sedative, depressive, and cognitive behaviors. CBD3063 is a selective, first-in-class, CRMP2-based peptidomimetic small molecule, which allosterically regulates Ca _v 2.2 to achieve analgesia and pain relief without negative side effect profiles. In summary, CBD3063 could potentially be a more effective alternative to GBP for pain relief.