Cytokine receptor clustering in sensory neurons with an engineered cytokine fusion protein triggers unique pain resolution pathways

Abstract

Significance Interactions between the immune system and nervous system regulate chronic pain. Cytokines are well-known immune-regulatory molecules that have activities beyond regulation of the immune system, including regulation of pain pathways. We identified that fusion of regulatory cytokines interleukin (IL)-4 and IL-10 into the fusion protein IL4–10 FP provides unexpected properties to inhibit pain by clustering the respective receptor chains of both cytokines in sensory neurons. IL4–10 FP promotes unique signaling pathways and gene-expression profiles that resolve chronic inflammatory pain. Thus IL4–10 FP is an immune biologic that effectively inhibits pain by activating pain-resolution pathways in sensory neurons through clustering of cytokine receptors. This unique effect discriminates IL4–10 FP from a combination therapy with individual regulatory cytokines. New therapeutic approaches to resolve persistent pain are highly needed. We tested the hypothesis that manipulation of cytokine receptors on sensory neurons by clustering regulatory cytokine receptor pairs with a fusion protein of interleukin (IL)-4 and IL-10 (IL4–10 FP) would redirect signaling pathways to optimally boost pain-resolution pathways. We demonstrate that a population of mouse sensory neurons express both receptors for the regulatory cytokines IL-4 and IL-10. This population increases during persistent inflammatory pain. Triggering these receptors with IL4–10 FP has unheralded biological effects, because it resolves inflammatory pain in both male and female mice. Knockdown of both IL4 and IL10 receptors in sensory neurons in vivo ablated the IL4–10 FP-mediated inhibition of inflammatory pain. Knockdown of either one of the receptors prevented the analgesic gain-of-function of IL4–10 FP. In vitro, IL4–10 FP inhibited inflammatory mediator-induced neuronal sensitization more effectively than the combination of cytokines, confirming its superior activity. The IL4–10 FP, contrary to the combination of IL-4 and IL-10, promoted clustering of IL-4 and IL-10 receptors in sensory neurons, leading to unique signaling, that is exemplified by activation of shifts in the cellular kinome and transcriptome. Interrogation of the potentially involved signal pathways led us to identify JAK1 as a key downstream signaling element that mediates the superior analgesic effects of IL4–10 FP. Thus, IL4–10 FP constitutes an immune-biologic that clusters regulatory cytokine receptors in sensory neurons to transduce unique signaling pathways required for full resolution of persistent inflammatory pain.