Inhibition of IRF5 cellular activity with cell-penetrating peptides that target homodimerization

Abstract

The transcription factor interferon regulatory factor 5 (IRF5) plays essential roles in pathogen-induced immunity downstream of Toll-, nucleotide-binding oligomerization domain–, and retinoic acid–inducible gene I–like receptors and is an autoimmune susceptibility gene. Normally, inactive in the cytoplasm, upon stimulation, IRF5 undergoes posttranslational modification(s), homodimerization, and nuclear translocation, where dimers mediate proinflammatory gene transcription. Here, we report the rational design of cell-penetrating peptides (CPPs) that disrupt IRF5 homodimerization. Biochemical and imaging analysis shows that IRF5-CPPs are cell permeable, noncytotoxic, and directly bind to endogenous IRF5. IRF5-CPPs were selective and afforded cell type– and species-specific inhibition. In plasmacytoid dendritic cells, inhibition of IRF5-mediated interferon-α production corresponded to a dose-dependent reduction in nuclear phosphorylated IRF5 [p(Ser462)IRF5], with no effect on pIRF5 levels. These data support that IRF5-CPPs function downstream of phosphorylation. Together, data support the utility of IRF5-CPPs as novel tools to probe IRF5 activation and function in disease.

Inhibition of IRF5 hyperactivation protects from lupus onset and severity

“…Last, given the heterogeneity of SLE and the differential kinetics and cell-type specificity of IRF5 activation in NZB/W F1 and MRL/lpr mice, it is unlikely that there is a single trigger or pathway leading to IRF5 activation. Indeed, we recently reported that the kinetics of Ser462 IRF5 phosphorylation and nuclear translocation were distinct depending on the stimulus (39). Unfortunately, this phospho-antibody does not detect endog-disease and contributed to increased survival; significant reductions in proteinuria occurred at 30 weeks (Figure 7, C-G, and Supplemental Figure 10A).…”

“…Identification of IRF5 as a global risk factor for autoimmune and inflammatory diseases (5,11,20,(36)(37)(38), coupled with its increased activation in the blood of patients with SLE, indicates that IRF5 is an attractive target for therapeutic inhibition. While C-terminal phosphorylation and dimerization represent steps amenable to inhibition (39), neither has been definitively shown to be an absolute requirement for nuclear translocation (35). An alternate approach to inhibiting IRF5 stems from the finding that either N-or C-terminal regions of IRFs can act as dominant-negative (DN) mutants to block transactivation ability (2,29,(40)(41)(42)(43)(44).…”

“…Altogether, these data show that IRF5 represents a particularly valuable, dual-function therapeutic target to treat autoimmune and inflammatory diseases. To date, this is the first report to our knowledge of a selective IRF5 inhibitor that directly binds to IRF5 to inhibit nuclear translocation and has in vivo clinical efficacy in murine models of lupus (39,83,84).…”

“…Healthy donor PBMCs were preincubated with an inhibitor and stimulated with SLE sera for 2 hours, and then IRF5 activation was assessed (19). N5-1 induced a significant reduction 39). Unfortunately, these inhibitors were not stable for in vivo analysis, nor did they similarly inhibit both human and murine IRF5.…”

Anti-SARS-CoV-2 IgG from severely ill COVID-19 patients promotes macrophage hyper-inflammatory responses

“…In addition to fostamatinib, also other drugs that target key molecules in FcγR signaling could be efficacious to counteract anti-Spike IgG-induced inflammation in COVID-19 patients. For example, the Sykdependent FcγR signaling pathway critically depends on the transcription factor IRF5 (15,28), which can be targeted using cell penetrating peptides (40). Furthermore, FcγR stimulation is known to induce metabolic reprogramming of human macrophages (28), which is also observed in COVID-19 patients (41), and therefore may provide additional targets for therapy.…”