Background/Purpose: NLRP3-activating mutations cause cryopyrinopathies of which NOMID is the most severe phenotype. NLRP3 assembles a protein complex, responsible for the maturation of IL-1β and IL-18. NOMID-associated inflammation, not bone phenotype is resolved by IL-1 biologics, but these drugs require parenteral injection and are costly. Engineered mice harboring a mutation found in NOMID patients exhibit several features of the human disease, including excessive IL-1β and IL-18 secretion and low bone mass. Here, we tested the hypothesis that the p38 MAPK/MK2 (p38) axis is central in the biosynthesis of pro-IL-1β and NLRP3, and as a result, inhibition of this pathway may provide efficacy in NOMID mice. We used our proprietary p38/MK2 pathway inhibitor, CDD-450 that is predicted to be safer and more efficacious than global p38 inhibitors.

Methods: CDD-450 selectivity for the p38/MK2 complex was evaluated by comparing p38 phosphorylation of MK2 vs other substrates including PRAK. CDD-450 efficacy was assessed in vitro and in vivo. For in vivo studies, floxed mice expressing NLRP3 carrying D301N substitution, ortholog of human D303N, were crossed with Cre-ER for postnatal conditional activation of NLRP3 upon exposure to tamoxifen, injected for 3 consecutive days over 2 weeks to generate NOMID mice. Mice were fed with normal or 1000 ppm CDD-450-formulated chow beginning 3 days before Tam exposure. This formulation yielded steady-state serum CDD-450 levels, which in pilot studies suppressed LPS-induced cytokine production by >80%. The studies were carried for up to 7 weeks.

Results: CDD-450 inhibited p38 phosphorylation of MK2 with low nanomolar potency and was >700-800x less potent at inhibiting p38 phosphorylation of PRAK and ATF2. NOMID mice produced excessive amounts of IL-1β; approximately 35% of these mice died during the course of the study. However, mice who survived, exhibited severe body weight loss and developed neutrophilia, thrombocytosis, lymphopenia, anemia, and splenomegaly. Remarkably, all CDD-450-treated mice survived and exhibited reduced weight loss as a result of abrogated or at least attenuated disease severity. NOMID mice also exhibited osteopenia, coincident with increased numbers of osteoclasts, responses that were inhibited >95% by CDD-450. Furthermore, in vitro osteoclast differentiation of mouse bone marrow-derived macrophages and IL-1β production by these cells were also blocked by CDD-450. From a translational prospective, human peripheral blood mononuclear cells (PBMC) were isolated from a cryopyrin-associated periodic syndrome (CAPS) patient, in which the disease is triggered upon exposure to low temperatures. CAPS PBMC, but not WT PBMC produced large amounts of IL-1β at 32^oC, an event that was inhibited by CDD-450 in a dose-dependent manner. In contrast, WT PBMC cultured at 37^oC produced IL-1β only when they were exposed to LPS, a response that was also reduced by CDD-450.

Conclusion: CDD-450 is a selective inhibitor of the p38/MK2 axis. Importantly, this compound exhibits sustained efficacy in NOMID mice and human cell systems. These results support ongoing efforts for clinical development of this compound for the treatment of inflammatory disorders.

ACR Meeting Abstracts - https://acrabstracts.org/abstract/nomid-associated-complications-in-mice-are-prevented-by-cdd-450-a-small-molecule-inhibitor-of-the-mitogen-activated-protein-kinase-activated-protein-kinase-2-mk2-pathway/