The Cancer Chemotherapeutic Paclitaxel Increases Human and Rodent Sensory Neuron Responses to TRPV1 by Activation of TLR4.

Peripheral neuropathy is dose limiting in paclitaxel cancer chemotherapy and can result in both acute pain during treatment and chronic persistent pain in cancer survivors. The hypothesis tested was that paclitaxel produces these adverse effects at least in part by sensitizing transient receptor potential vanilloid subtype 1 (TRPV1) through Toll-like receptor 4 (TLR4) signaling. The data show that paclitaxel-induced behavioral hypersensitivity is prevented and reversed by spinal administration of a TRPV1 antagonist. The number of TRPV1(+) neurons is increased in the dorsal root ganglia (DRG) in paclitaxel-treated rats and is colocalized with TLR4 in rat and human DRG neurons. Cotreatment of rats with lipopolysaccharide from the photosynthetic bacterium Rhodobacter sphaeroides (LPS-RS), a TLR4 inhibitor, prevents the increase in numbers of TRPV1(+) neurons by paclitaxel treatment. Perfusion of paclitaxel or the archetypal TLR4 agonist LPS activated both rat DRG and spinal neurons directly and produced acute sensitization of TRPV1 in both groups of cells via a TLR4-mediated mechanism. Paclitaxel and LPS sensitize TRPV1 in HEK293 cells stably expressing human TLR4 and transiently expressing human TRPV1. These physiological effects also are prevented by LPS-RS. Finally, paclitaxel activates and sensitizes TRPV1 responses directly in dissociated human DRG neurons. In summary, TLR4 was activated by paclitaxel and led to sensitization of TRPV1. This mechanism could contribute to paclitaxel-induced acute pain and chronic painful neuropathy. Significance statement: In this original work, it is shown for the first time that paclitaxel activates peripheral sensory and spinal neurons directly and sensitizes these cells to transient receptor potential vanilloid subtype 1 (TRPV1)-mediated capsaicin responses via Toll-like receptor 4 (TLR4) in multiple species. A direct functional interaction between TLR4 and TRPV1 is shown in rat and human dorsal root ganglion neurons, TLR4/TRPV1-coexpressing HEK293 cells, and in both rat and mouse spinal cord slices. Moreover, this is the first study to show that this interaction plays an important role in the generation of behavioral hypersensitivity in paclitaxel-related neuropathy. The key translational implications are that TLR4 and TRPV1 antagonists may be useful in the prevention and treatment of chemotherapy-induced peripheral neuropathy in humans.

MAPK signaling downstream to TLR4 contributes to paclitaxel-induced peripheral neuropathy.

Toll-like receptor 4 (TLR4) has been implicated as a locus for initiation of paclitaxel related chemotherapy induced peripheral neuropathy (CIPN). This project explores the involvement of the immediate down-stream signal molecules in inducing paclitaxel CIPN. Mitogen-activated protein kinases (MAPKs) and nuclear factor-κB (NFκB) were measured in dorsal root ganglia (DRG) and the spinal cord over time using Western blot and immunohistochemistry in a rat model of paclitaxel CIPN. The effects of MAPK inhibitors in preventing and reversing behavioral signs of CIPN were also measured (group sizes 4-9). Extracellular signal related kinase (ERK1/2) and P38 but not c-Jun N terminal kinase (JNK) or PI3K-Akt signaling expression was increased in DRG. Phospho-ERK1/2 staining was co-localized to small CGRP-positive DRG neurons in cell profiles surrounding large DRG neurons consistent with satellite glial cells. The expression of phospho-P38 was co-localized to small IB4-positive and CGRP-positive DRG neurons. The TLR4 antagonist LPS derived from Rhodobacter sphaeroides (LPS-RS) inhibited paclitaxel-induced phosphorylation of ERK1/2 and P38. The MAPK inhibitors PD98059 (MEK1/2), U0126 (MEK1/2) and SB203580 (P38) prevented but did not reverse paclitaxel-induced behavioral hypersensitivity. Paclitaxel treatment resulted in phosphorylation of Inhibitor α of NFκB (IκBα) in DRG resulting in an apparent release of NFκB from the IκBα-NFκB complex as increased expression of nuclear NFκB was also observed. LPS-RS inhibited paclitaxel-induced translocation of NFκB in DRG. No change was observed in spinal NFκB. These results implicate TLR4 signaling via MAP kinases and NFκB in the induction and maintenance of paclitaxel-related CIPN.

Toll-like receptor 4 signaling contributes to Paclitaxel-induced peripheral neuropathy.

UNLABELLED: This paper tests the contribution of the toll-like receptors, TLR4 in particular, in the initiation and maintenance of paclitaxel-related chemotherapy-induced peripheral neuropathy. TLR4 and its immediate downstream signaling molecules-myeloid differentiation primary response gene 88 (MyD88) and toll/interleukin 1 receptor domain-containing adapter-inducing interferon-ß (TRIF)-were found to be increased in the dorsal root ganglion (DRG) using Western blot by day 7 of paclitaxel treatment. The behavioral phenotype, the increase of both TLR4 and MyD88, was blocked by cotreatment with the TLR4 antagonist lipopolysaccharide-Rhodobacter sphaeroides during chemotherapy. A similar, but less robust, behavioral effect was observed using intrathecal treatment of MyD88 homodimerization inhibitory peptide. DRG levels of TLR4 and MyD88 reduced over the next 2 weeks, whereas these levels remained increased in spinal cord through day 21 following chemotherapy. Immunohistochemical analysis revealed TLR4 expression in both calcitonin gene-related peptide-positive and isolectin B4-positive small DRG neurons. MyD88 was only found in calcitonin gene-related peptide-positive neurons, and TRIF was found in both calcitonin gene-related peptide-positive and isolectin B4-positive small DRG neurons as well as in medium- and large-size DRG neurons. In the spinal cord, TLR4 was only found colocalized to astrocytes but not with either microglia or neurons. Intrathecal treatment with the TLR4 antagonist lipopolysaccharide-R. sphaeroides transiently reversed preestablished chemotherapy-induced peripheral neuropathy mechanical hypersensitivity. These results strongly implicate TLR4 signaling in the DRG and the spinal cord in the induction and maintenance of paclitaxel-related chemotherapy-induced peripheral neuropathy. PERSPECTIVE: The toll-like receptor TLR4 and MyD88 signaling pathway could be a new potential therapeutic target in paclitaxel-induced painful neuropathy.