VLK drives extracellular phosphorylation of EphB2 to govern the EphB2-NMDAR interaction and injury-induced pain.

Phosphorylation of hundreds of protein extracellular domains is mediated by two kinase families, yet the significance of these kinases is underexplored. Here, we find that the presynaptic release of the tyrosine directed-ectokinase, Vertebrate Lonesome Kinase (VLK/Pkdcc), is necessary and sufficient for the direct extracellular interaction between EphB2 and GluN1 at synapses, for phosphorylation of the ectodomain of EphB2, and for injury-induced pain. Pkdcc is an essential gene in the nervous system, and VLK is found in synaptic vesicles, and is released from neurons in a SNARE-dependent fashion. VLK is expressed by nociceptive sensory neurons where presynaptic sensory neuron-specific knockout renders mice impervious to post-surgical pain, without changing proprioception. VLK defines an extracellular mechanism that regulates protein-protein interaction and non-opioid-dependent pain in response to injury.

Epigenomic landscape of the human dorsal root ganglion: sex differences and transcriptional regulation of nociceptive genes.

Gene expression is influenced by chromatin architecture via controlled access of regulatory factors to DNA. To better understand regulation of gene expression in the human dorsal root ganglion (hDRG) we used bulk and spatial transposase-accessible chromatin technology followed by sequencing (ATAC-seq). We detected a total of 3005 differentially accessible chromatin regions (DARs) between sexes using bulk ATAC-seq. DARs in female hDRG mapped mainly to the X chromosome. In males, DARs were found in autosomal genes. We also found differential transcription factor binding motifs within DARs. EGR1/3 and SP1/4 were abundant in females, and JUN, FOS and other AP-1 family members in males. With the aim of dissecting the open chromatin profile in hDRG neurons, we used spatial ATAC-seq. Consistent with our bulk ATAC-seq data, most of the DARs in female hDRG were located in X chromosome genes. Neuron cluster showed higher chromatin accessibility in GABAergic, glutamatergic, and interferon-related genes in females, and in Ca2+-signaling-related genes in males. Sex differences in open chromatin transcription factor binding sites in neuron-proximal barcodes were consistent with the bulk data, having EGR1 transcription factor activity in females and AP-1 family members in males. Accordingly, we showed higher expression of EGR1 in female hDRG compared to male with in-situ hybridization. Our findings point to epigenomic sex differences in the hDRG that likely underlie divergent transcriptional responses that determine mechanistic sex differences in pain.

How to differentiate induced pluripotent stem cells into sensory neurons for disease modelling: a functional assessment.

BACKGROUND: Human induced pluripotent stem cell (iPSC)-derived peripheral sensory neurons present a valuable tool to model human diseases and are a source for applications in drug discovery and regenerative medicine. Clinically, peripheral sensory neuropathies can result in maladies ranging from a complete loss of pain to severe painful neuropathic disorders. Sensory neurons are located in the dorsal root ganglion and are comprised of functionally diverse neuronal types. Low efficiency, reproducibility concerns, variations arising due to genetic factors and time needed to generate functionally mature neuronal populations from iPSCs remain key challenges to study human nociception in vitro. Here, we report a detailed functional characterization of iPSC-derived sensory neurons with an accelerated differentiation protocol ("Anatomic" protocol) compared to the most commonly used small molecule approach ("Chambers" protocol). Anatomic's commercially available RealDRG™ were further characterized for both functional and expression phenotyping of key nociceptor markers. METHODS: Multiple iPSC clones derived from different reprogramming methods, genetics, age, and somatic cell sources were used to generate sensory neurons. Manual patch clamp was used to functionally characterize both control and patient-derived neurons. High throughput techniques were further used to demonstrate that RealDRGs™ derived from the Anatomic protocol are amenable to high throughput technologies for disease modelling. RESULTS: The Anatomic protocol rendered a purer culture without the use of mitomycin C to suppress non-neuronal outgrowth, while Chambers differentiations yielded a mix of cell types. Chambers protocol results in predominantly tonic firing when compared to Anatomic protocol. Patient-derived nociceptors displayed higher frequency firing compared to control subject with both, Chambers and Anatomic differentiation approaches, underlining their potential use for clinical phenotyping as a disease-in-a-dish model. RealDRG™ sensory neurons show heterogeneity of nociceptive markers indicating that the cells may be useful as a humanized model system for translational studies. CONCLUSIONS: We validated the efficiency of two differentiation protocols and their potential application for functional assessment and thus understanding the disease mechanisms from patients suffering from pain disorders. We propose that both differentiation methods can be further exploited for understanding mechanisms and development of novel treatments in pain disorders.

Synovial fibroblast gene expression is associated with sensory nerve growth and pain in rheumatoid arthritis.

It has been presumed that rheumatoid arthritis (RA) joint pain is related to inflammation in the synovium; however, recent studies reveal that pain scores in patients do not correlate with synovial inflammation. We developed a machine-learning approach (graph-based gene expression module identification or GbGMI) to identify an 815-gene expression module associated with pain in synovial biopsy samples from patients with established RA who had limited synovial inflammation at arthroplasty. We then validated this finding in an independent cohort of synovial biopsy samples from patients who had early untreated RA with little inflammation. Single-cell RNA sequencing analyses indicated that most of these 815 genes were most robustly expressed by lining layer synovial fibroblasts. Receptor-ligand interaction analysis predicted cross-talk between human lining layer fibroblasts and human dorsal root ganglion neurons expressing calcitonin gene-related peptide (CGRP+). Both RA synovial fibroblast culture supernatant and netrin-4, which is abundantly expressed by lining fibroblasts and was within the GbGMI-identified pain-associated gene module, increased the branching of pain-sensitive murine CGRP+ dorsal root ganglion neurons in vitro. Imaging of solvent-cleared synovial tissue with little inflammation from humans with RA revealed CGRP+ pain-sensing neurons encasing blood vessels growing into synovial hypertrophic papilla. Together, these findings support a model whereby synovial lining fibroblasts express genes associated with pain that enhance the growth of pain-sensing neurons into regions of synovial hypertrophy in RA.

Rhythmic motor behavior explains individual differences in grammar skills in adults.

A growing body of literature has reported the relationship between music and language, particularly between individual differences in perceptual rhythm skill and grammar competency in children. Here, we investigated whether motoric aspects of rhythm processing-as measured by rhythmic finger tapping tasks-also explain the rhythm-grammar connection in 150 healthy young adults. We found that all expressive rhythm skills (spontaneous, synchronized, and continued tapping) along with rhythm discrimination skill significantly predicted receptive grammar skills on either auditory sentence comprehension or grammaticality well-formedness judgment (e.g., singular/plural, past/present), even after controlling for verbal working memory and music experience. Among these, synchronized tapping and rhythm discrimination explained unique variance of sentence comprehension and grammaticality judgment, respectively, indicating differential associations between different rhythm and grammar skills. Together, we demonstrate that even simple and repetitive motor behavior can account for seemingly high-order grammar skills in the adult population, suggesting that the sensorimotor system continue to support syntactic operations.

Elevations in the Mitochondrial Matrix Protein Cyclophilin D Correlate With Reduced Parvalbumin Expression in the Prefrontal Cortex of Patients With Schizophrenia.

BACKGROUND AND HYPOTHESIS: Cognitive deficits in schizophrenia are linked to dysfunctions of the dorsolateral prefrontal cortex (DLPFC), including alterations in parvalbumin (PV)-expressing interneurons (PVIs). Redox dysregulation and oxidative stress may represent convergence points in the pathology of schizophrenia, causing dysfunction of GABAergic interneurons and loss of PV. Here, we show that the mitochondrial matrix protein cyclophilin D (CypD), a critical initiator of the mitochondrial permeability transition pore (mPTP) and modulator of the intracellular redox state, is altered in PVIs in schizophrenia. STUDY DESIGN: Western blotting was used to measure CypD protein levels in postmortem DLPFC specimens of schizophrenic patients (n = 27) and matched comparison subjects with no known history of psychiatric or neurological disorders (n = 26). In a subset of this cohort, multilabel immunofluorescent confocal microscopy with unbiased stereological sampling methods were used to quantify (1) numbers of PVI across the cortical mantle (20 unaffected comparison, 14 schizophrenia) and (2) PV and CypD protein levels from PVIs in the cortical layers 2-4 (23 unaffected comparison, 18 schizophrenia). STUDY RESULTS: In schizophrenic patients, the overall number of PVIs in the DLPFC was not significantly altered, but in individual PVIs of layers 2-4 PV protein levels decreased along a superficial-to-deep gradient when compared to unaffected comparison subjects. These laminar-specific PVI alterations were reciprocally linked to significant CypD elevations both in PVIs and total DLPFC gray matter. CONCLUSIONS: Our findings support previously reported PVI anomalies in schizophrenia and suggest that CypD-mediated mPTP formation could be a potential contributor to PVI dysfunction in schizophrenia.

Vinorelbine causes a neuropathic pain-like state in mice via STING and MNK1 signaling associated with type I interferon induction.

Type I interferons (IFNs) increase the excitability of dorsal root ganglia (DRGs) neurons via MNK-eIF4E signaling to promote pain sensitization in mice. Activation of stimulator of interferon response cGAMP interactor 1 (STING) signaling is pivotal for type I IFN induction. We hypothesized that vinorelbine, a chemotherapeutic and activator of STING, would cause a neuropathic pain-like state in mice via STING signaling in DRG neurons associated with IFN production. Vinorelbine caused tactile allodynia and grimacing in wild-type (WT) mice and increased p-IRF3, type I IFNs, and p-eIF4E in peripheral nerves. Supporting our hypothesis, vinorelbine failed to induce IRF3-IFNs-MNK-eIF4E in StingGt/Gt mice and, subsequently, failed to cause pain. The vinorelbine-elicited increase of p-eIF4E was not observed in Mknk1-/- (MNK1 knockout) mice in peripheral nerves consistent with the attenuated pro-nociceptive effect of vinorelbine in these mice. Our findings show that activation of STING signaling in the periphery causes a neuropathic pain-like state through type I IFN signaling to DRG nociceptors.

Highly specific σ2R/TMEM97 ligand FEM-1689 alleviates neuropathic pain and inhibits the integrated stress response.

The sigma 2 receptor (σ2R) was described pharmacologically more than three decades ago, but its molecular identity remained obscure until recently when it was identified as transmembrane protein 97 (TMEM97). We and others have shown that σ2R/TMEM97 ligands alleviate mechanical hypersensitivity in mouse neuropathic pain models with a time course wherein maximal antinociceptive effect is approximately 24 h following dosing. We sought to understand this unique antineuropathic pain effect by addressing two key questions: do these σ2R/TMEM97 compounds act selectively via the receptor, and what is their downstream mechanism on nociceptive neurons? Using male and female conventional knockout mice for Tmem97, we find that a σ2R/TMEM97 binding compound, FEM-1689, requires the presence of the gene to produce antinociception in the spared nerve injury model in mice. Using primary mouse dorsal root ganglion neurons, we demonstrate that FEM-1689 inhibits the integrated stress response (ISR) and promotes neurite outgrowth via a σ2R/TMEM97-specific action. We extend the clinical translational value of these findings by showing that FEM-1689 reduces ISR and p-eIF2α levels in human sensory neurons and that it alleviates the pathogenic engagement of ISR by methylglyoxal. We also demonstrate that σ2R/TMEM97 is expressed in human nociceptors and satellite glial cells. These results validate σ2R/TMEM97 as a promising target for further development for the treatment of neuropathic pain.

4E-BP1-dependent translation in nociceptors controls mechanical hypersensitivity via TRIM32/type I interferon signaling.

Activation of the mechanistic target of rapamycin complex 1 (mTORC1) contributes to the development of chronic pain. However, the specific mechanisms by which mTORC1 causes hypersensitivity remain elusive. The eukaryotic initiation factor 4E-binding protein 1 (4E-BP1) is a key mTORC1 downstream effector that represses translation initiation. Here, we show that nociceptor-specific deletion of 4E-BP1, mimicking activation of mTORC1-dependent translation, is sufficient to cause mechanical hypersensitivity. Using translating ribosome affinity purification in nociceptors lacking 4E-BP1, we identified a pronounced translational up-regulation of tripartite motif-containing protein 32 (TRIM32), an E3 ubiquitin ligase that promotes interferon signaling. Down-regulation of TRIM32 in nociceptors or blocking type I interferon signaling reversed the mechanical hypersensitivity in mice lacking 4E-BP1. Furthermore, nociceptor-specific ablation of TRIM32 alleviated mechanical hypersensitivity caused by tissue inflammation. These results show that mTORC1 in nociceptors promotes hypersensitivity via 4E-BP1-dependent up-regulation of TRIM32/interferon signaling and identify TRIM32 as a therapeutic target in inflammatory pain.

RNA isoform expression landscape of the human dorsal root ganglion (DRG) generated from long read sequencing.

Splicing is a post-transcriptional RNA processing mechanism that enhances genomic complexity by creating multiple isoforms from the same gene. Diversity in splicing in the mammalian nervous system is associated with neuronal development, synaptic function and plasticity, and is also associated with diseases of the nervous system ranging from neurodegeneration to chronic pain. We aimed to characterize the isoforms expressed in the human peripheral nervous system, with the goal of creating a resource to identify novel isoforms of functionally relevant genes associated with somatosensation and nociception. We used long read sequencing (LRS) to document isoform expression in the human dorsal root ganglia (hDRG) from 3 organ donors. Isoforms were validated in silico by confirming expression in hDRG short read sequencing (SRS) data from 3 independent organ donors. 19,547 isoforms of protein-coding genes were detected using LRS and validated with SRS and strict expression cutoffs. We identified 763 isoforms with at least one previously undescribed splice-junction. Previously unannotated isoforms of multiple pain-associated genes, including ASIC3, MRGPRX1 and HNRNPK were identified. In the novel isoforms of ASIC3, a region comprising ~35% of the 5'UTR was excised. In contrast, a novel splice-junction was utilized in isoforms of MRGPRX1 to include an additional exon upstream of the start-codon, consequently adding a region to the 5'UTR. Novel isoforms of HNRNPK were identified which utilized previously unannotated splice-sites to both excise exon 14 and include a sequence in the 5' end of exon 13. The insertion and deletion in the coding region was predicted to excise a serine-phosphorylation site favored by cdc2, and replace it with a tyrosine-phosphorylation site potentially phosphorylated by SRC. We also independently confirm a recently reported DRG-specific splicing event in WNK1 that gives insight into how painless peripheral neuropathy occurs when this gene is mutated. Our findings give a clear overview of mRNA isoform diversity in the hDRG obtained using LRS. Using this work as a foundation, an important next step will be to use LRS on hDRG tissues recovered from people with a history of chronic pain. This should enable identification of new drug targets and a better understanding of chronic pain that may involve aberrant splicing events.

How to differentiate induced pluripotent stem cells into sensory neurons for disease modelling: a comparison of two protocols.

Background: Human induced pluripotent stem cell (iPSC)-derived peripheral sensory neurons present a valuable tool to model human diseases and are a source for applications in drug discovery and regenerative medicine. Clinically, peripheral sensory neuropathies can result in maladies ranging from a complete loss of pain to severe painful neuropathic symptoms. Sensory neurons are located in the dorsal root ganglion and are comprised of functionally diverse neuronal types. Low efficiency, reproducibility concerns, variations arising due to genetic factors and time needed to generate functionally mature neuronal populations from iPSCs for disease modelling remain key challenges to study human nociception in vitro. Here, we report a detailed characterization of iPSC-derived sensory neurons with an accelerated differentiation protocol ("Anatomic" protocol) compared to the most commonly used small molecule approach ("Chambers" protocol). Methods: Multiple iPSC clones derived from different reprogramming methods, genetics, age, and somatic cell sources were used to generate sensory neurons. Expression profiling of sensory neurons was performed with Immunocytochemistry and in situ hybridization techniques. Manual patch clamp and high throughput cellular screening systems (Fluorescence imaging plate reader, automated patch clamp and multi-well microelectrode arrays recordings) were applied to functionally characterize the generated sensory neurons. Results: The Anatomic protocol rendered a purer culture without the use of mitomycin C to suppress non-neuronal outgrowth, while Chambers differentiations yielded a mix of cell types. High throughput systems confirmed functional expression of Na+ and K+ ion channels. Multi-well microelectrode recordings display spontaneously active neurons with sensitivity to increased temperature indicating expression of heat sensitive ion channels. Patient-derived nociceptors displayed higher frequency firing compared to control subject with both, Chambers and Anatomic differentiation approaches, underlining their potential use for clinical phenotyping as a disease-in-a-dish model. Conclusions: We validated the efficiency of two differentiation protocols and their potential application for understanding the disease mechanisms from patients suffering from pain disorders. We propose that both differentiation methods can be further exploited for understanding mechanisms and development of novel treatments in pain disorders.

S. aureus drives itch and scratch-induced skin damage through a V8 protease-PAR1 axis.

Itch is an unpleasant sensation that evokes a desire to scratch. The skin barrier is constantly exposed to microbes and their products. However, the role of microbes in itch generation is unknown. Here, we show that Staphylococcus aureus, a bacterial pathogen associated with itchy skin diseases, directly activates pruriceptor sensory neurons to drive itch. Epicutaneous S. aureus exposure causes robust itch and scratch-induced damage. By testing multiple isogenic bacterial mutants for virulence factors, we identify the S. aureus serine protease V8 as a critical mediator in evoking spontaneous itch and alloknesis. V8 cleaves proteinase-activated receptor 1 (PAR1) on mouse and human sensory neurons. Targeting PAR1 through genetic deficiency, small interfering RNA (siRNA) knockdown, or pharmacological blockade decreases itch and skin damage caused by V8 and S. aureus exposure. Thus, we identify a mechanism of action for a pruritogenic bacterial factor and demonstrate the potential of inhibiting V8-PAR1 signaling to treat itch.

Analysis of matrisome expression patterns in murine and human dorsal root ganglia.

The extracellular matrix (ECM) is a dynamic structure of molecules that can be divided into six different categories and are collectively called the matrisome. The ECM plays pivotal roles in physiological processes in many tissues, including the nervous system. Intriguingly, alterations in ECM molecules/pathways are associated with painful human conditions and murine pain models. Nevertheless, mechanistic insight into the interplay of normal or defective ECM and pain is largely lacking. The goal of this study was to integrate bulk, single-cell, and spatial RNA sequencing (RNAseq) datasets to investigate the expression and cellular origin of matrisome genes in male and female murine and human dorsal root ganglia (DRG). Bulk RNAseq showed that about 65% of all matrisome genes were expressed in both murine and human DRG, with proportionally more core matrisome genes (glycoproteins, collagens, and proteoglycans) expressed compared to matrisome-associated genes (ECM-affiliated genes, ECM regulators, and secreted factors). Single cell RNAseq on male murine DRG revealed the cellular origin of matrisome expression. Core matrisome genes, especially collagens, were expressed by fibroblasts whereas matrisome-associated genes were primarily expressed by neurons. Cell-cell communication network analysis with CellChat software predicted an important role for collagen signaling pathways in connecting vascular cell types and nociceptors in murine tissue, which we confirmed by analysis of spatial transcriptomic data from human DRG. RNAscope in situ hybridization and immunohistochemistry demonstrated expression of collagens in fibroblasts surrounding nociceptors in male and female human DRG. Finally, comparing human neuropathic pain samples with non-pain samples also showed differential expression of matrisome genes produced by both fibroblasts and by nociceptors. This study supports the idea that the DRG matrisome may contribute to neuronal signaling in both mouse and human, and that dysregulation of matrisome genes is associated with neuropathic pain.

MNK inhibitor eFT508 (Tomivosertib) suppresses ectopic activity in human dorsal root ganglion neurons from dermatomes with radicular neuropathic pain.

Spontaneous activity in dorsal root ganglion (DRG) neurons is a key driver of neuropathic pain in preclinical models and in patients suffering from this largely untreated disease. While many intracellular signaling mechanisms have been examined in preclinical models that drive this spontaneous activity (SA), none of these have been tested directly on spontaneously active human nociceptors. Using cultured DRG neurons recovered during thoracic vertebrectomy surgeries, we show that inhibition of mitogen activated protein kinase interacting kinase (MNK) with eFT508 (25 nM) reverses SA in human sensory neurons associated with painful dermatomes. MNK inhibition in spontaneously active nociceptors decreased action potential amplitude and produced alterations in the magnitude of afterhyperpolarizing currents suggesting modification of Na+ and K+ channel activity downstream of MNK inhibition. The effects of MNK inhibition on SA took minutes to emerge and were reversible over time with eFT508 washout. MNK inhibition with eFT508 led to a profound loss of eIF4E Serine 209 phosphorylation, a specific target of the kinase, within 2 min of drug treatment, consistent with the rapid action of the drug on SA in electrophysiology experiments. Our results create a compelling case for the future testing of MNK inhibitors in clinical trials for neuropathic pain.

Intrinsic Homeostatic Plasticity in Mouse and Human Sensory Neurons.

In response to changes in activity induced by environmental cues, neurons in the central nervous system undergo homeostatic plasticity to sustain overall network function during abrupt changes in synaptic strengths. Homeostatic plasticity involves changes in synaptic scaling and regulation of intrinsic excitability. Increases in spontaneous firing and excitability of sensory neurons are evident in some forms of chronic pain in animal models and human patients. However, whether mechanisms of homeostatic plasticity are engaged in sensory neurons under normal conditions or altered after chronic pain is unknown. Here, we showed that sustained depolarization induced by 30mM KCl induces a compensatory decrease in the excitability in mouse and human sensory neurons. Moreover, voltage-gated sodium currents are robustly reduced in mouse sensory neurons contributing to the overall decrease in neuronal excitability. Decreased efficacy of these homeostatic mechanisms could potentially contribute to the development of the pathophysiology of chronic pain.

Harmonized cross-species cell atlases of trigeminal and dorsal root ganglia.

Peripheral sensory neurons in the dorsal root ganglion (DRG) and trigeminal ganglion (TG) are specialized to detect and transduce diverse environmental stimuli including touch, temperature, and pain to the central nervous system. Recent advances in single-cell RNA-sequencing (scRNA-seq) have provided new insights into the diversity of sensory ganglia cell types in rodents, non-human primates, and humans, but it remains difficult to compare transcriptomically defined cell types across studies and species. Here, we built cross-species harmonized atlases of DRG and TG cell types that describe 18 neuronal and 11 non-neuronal cell types across 6 species and 19 studies. We then demonstrate the utility of this harmonized reference atlas by using it to annotate newly profiled DRG nuclei/cells from both human and the highly regenerative axolotl. We observe that the transcriptomic profiles of sensory neuron subtypes are broadly similar across vertebrates, but the expression of functionally important neuropeptides and channels can vary notably. The new resources and data presented here can guide future studies in comparative transcriptomics, simplify cell type nomenclature differences across studies, and help prioritize targets for future pain therapy development.

Tiam1 creates a painful link between dendritic spine remodeling and NMDA receptors.

Dendritic spine remodeling in the dorsal horn is associated with many chronic pain models. Li et al. demonstrate that Tiam1 links Rac1-mediated spine changes to NMDA receptor activity to promote behavioral signs of chronic pain in rodents.

Metastatic Infiltration of Nervous Tissue and Periosteal Nerve Sprouting in Multiple Myeloma-Induced Bone Pain in Mice and Human.

Multiple myeloma (MM) is a neoplasia of B plasma cells that often induces bone pain. However, the mechanisms underlying myeloma-induced bone pain (MIBP) are mostly unknown. Using a syngeneic MM mouse model, we show that periosteal nerve sprouting of calcitonin gene-related peptide (CGRP+) and growth associated protein 43 (GAP43+) fibers occurs concurrent to the onset of nociception and its blockade provides transient pain relief. MM patient samples also showed increased periosteal innervation. Mechanistically, we investigated MM induced gene expression changes in the dorsal root ganglia (DRG) innervating the MM-bearing bone of male mice and found alterations in pathways associated with cell cycle, immune response and neuronal signaling. The MM transcriptional signature was consistent with metastatic MM infiltration to the DRG, a never-before described feature of the disease that we further demonstrated histologically. In the DRG, MM cells caused loss of vascularization and neuronal injury, which may contribute to late-stage MIBP. Interestingly, the transcriptional signature of a MM patient was consistent with MM cell infiltration to the DRG. Overall, our results suggest that MM induces a plethora of peripheral nervous system alterations that may contribute to the failure of current analgesics and suggest neuroprotective drugs as appropriate strategies to treat early onset MIBP.SIGNIFICANCE STATEMENT Multiple myeloma (MM) is a painful bone marrow cancer that significantly impairs the quality of life of the patients. Analgesic therapies for myeloma-induced bone pain (MIBP) are limited and often ineffective, and the mechanisms of MIBP remain unknown. In this manuscript, we describe cancer-induced periosteal nerve sprouting in a mouse model of MIBP, where we also encounter metastasis to the dorsal root ganglia (DRG), a never-before described feature of the disease. Concomitant to myeloma infiltration, the lumbar DRGs presented blood vessel damage and transcriptional alterations, which may mediate MIBP. Explorative studies on human tissue support our preclinical findings. Understanding the mechanisms of MIBP is crucial to develop targeted analgesic with better efficacy and fewer side effects for this patient population.

Vinorelbine causes a neuropathic pain-like state in mice via STING and MNK1 signaling associated with type I interferon induction.

Type I interferons (IFNs) increase the excitability of dorsal root ganglion (DRG) neurons via activation of MNK-eIF4E translation signaling to promote pain sensitization in mice. Activation of STING signaling is a key component of type I IFN induction. Manipulation of STING signaling is an active area of investigation in cancer and other therapeutic areas. Vinorelbine is a chemotherapeutic that activates STING and has been shown to cause pain and neuropathy in oncology clinical trials in patients. There are conflicting reports on whether STING signaling promotes or inhibits pain in mice. We hypothesized that vinorelbine would cause a neuropathic pain-like state in mice via STING and signaling pathways in DRG neurons associated with type I IFN induction. Vinorelbine (10 mg/kg, i.v.) induced tactile allodynia and grimacing in WT male and female mice and increased p-IRF3 and type I IFN protein in peripheral nerves. In support of our hypothesis, vinorelbine-mediated pain was absent in male and female StingGt/Gt mice. Vinorelbine also failed to induce IRF3 and type I IFN signaling in these mice. Since type I IFNs engage translational control via MNK1-eIF4E in DRG nociceptors, we assessed vinorelbine-mediated p-eIF4E changes. Vinorelbine increased p-eIF4E in DRG in WT animals but not in StingGt/Gt or Mknk1-/- (MNK1 KO) mice. Consistent with these biochemical findings, vinorelbine had an attenuated pro-nociceptive effect in male and female MNK1 KO mice. Our findings support the conclusion that activation of STING signaling in the peripheral nervous system causes a neuropathic pain-like state that is mediated by type I IFN signaling to DRG nociceptors.