Investigating immunophenotypic and transcriptional heterogeneity as biomarkers of pain centralization in rheumatoid arthritis

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Over half of patients with rheumatoid arthritis (RA) report pain despite treatment with strong disease-modifying antirheumatic drugs (DMARDs). Our research group was one of the first to show that this pain is associated with increased pain sensitivity in areas distant from inflamed joints, suggestive of abnormalities in the way the brain and spinal cord regulate pain. The pain associated with this process is termed “centralized pain” and is frequently treated with opioid analgesics. However, opioids are minimally effective and often misused. The development of non-opioid analgesics has been hindered by our limited understanding of how the centralized pain phenotype relates to changes at the cellular level. In the research setting, centralized pain is often assessed using quantitative sensory testing (QST). However, QST is time-consuming, requires significant assessor training, and is prone to measurement error. There is a critical unmet need to develop quantifiable measurements of the altered cellular state that distinguish patients with centralized pain. It is imperative to address this need to achieve our long-term goal of developing safe and efficacious non-opioid pain analgesics for patients with rheumatic diseases. The objective of this high-risk, high-reward proposal is to generate cutting-edge insights into the relationship between peripheral blood mononuclear cells (PBMCs) and centralized pain in patients with RA. Our proposal emphasizes an unbiased, high-throughput approach, using multi-parameter flow cytometry and single-cell RNA sequencing (scRNA-seq) to identify characteristics of PBMCs that are associated with centralized pain after DMARD therapy. Our central hypothesis is that differences in the composition and gene expression of circulating immune cells, particularly monocytes, will characterize RA patients with a centralized pain phenotype vs. those without a centralized pain phenotype. In Aim 1, we will determine the association between the immunophenotypic profile of PBMCs and centralized pain. We will enroll 50 RA patients with minimal joint inflammation but varying levels of pain after DMARD treatment. These patients will undergo QST to test whether centralized pain, assessed by pressure pain thresholds at the trapezius muscle, is associated with: a) the proportion of each cell type in circulation, and b) continuous measures of activation status for each cell type. In Aim 2, we will identify differences in the transcriptional heterogeneity of circulating PBMCs between patients with a centralized pain phenotype vs. those without a centralized pain phenotype. We will perform scRNA-seq on a subset of patients from Aim 1 with the highest (N = 10) and lowest (N = 10) levels of centralized pain. We will investigate differences in transcriptional subpopulations and cell-type-specific gene expression between the two groups. Our proposal has high impact potential because, if successful, it will deliver novel insights into the role of circulating PBMCs in altered pain regulation in patients with RA, the prototypical systemic inflammatory condition. Data from this proposal will be used to inform an R01 application to identify a reproducible peripheral blood biomarker for centralized pain in patients with RA.