Targeting NLRP3 with ofirnoflast, alone or in combination with semaglutide, ameliorates inflammatory anemia associated with obesity-induced inflammation in rodent models

Abstract

Abstract Background: Inflammatory anemia is a common manifestation of chronic disease that arises when sustained cytokine-mediated inflammation leads to the immune system's sequestration of iron and suppression of erythropoiesis. While this response may serve as a defense against pathogens, persistent inflammatory states can lead to impaired red blood cell production and iron metabolism. Obesity is increasingly recognized as a chronic inflammatory condition, with expanded adipose tissue driving systemic inflammation and altering the bone marrow microenvironment. Increased marrow adiposity disrupts hematopoiesis and promotes anemia of inflammation. The NLRP3 inflammasome is a key sensor of metabolic stress and a central mediator of obesity-induced inflammatory signaling in adipose tissue and the bone marrow. Ofirnoflast (HT-6184) is a selective, orally available NLRP3 inflammasome inhibitor that targets the Nek7 scaffold protein. This study evaluated the therapeutic potential of Ofirnoflast, alone or in combination with the GLP-1 receptor agonist semaglutide, in restoring hematologic function in diet-induced obese mice. Objective: To determine whether pharmacologic inhibition of the NLRP3 inflammasome with Ofirnoflast, alone or in combination with the GLP-1 agonist semaglutide, can prevent or reverse obesity-associated inflammatory anemia in a preclinical diet-induced obese mouse model. Methods: Male C57BL/6 mice were fed a high-fat diet (HFD) for 16 weeks to induce obesity and randomized into four groups: HFD control, semaglutide (0.03 mg/kg), Ofirnoflast (5 mg/kg), and Ofirnoflast + semaglutide. Age-matched lean mice served as healthy controls. Treatment was administered once daily for 5 days. Serum and adipose tissue were collected at endpoint for proteomic profiling (mass spectrometry of >900 serum proteins) and immunohistochemical (IHC) analysis of inflammatory markers. Results: Mice fed a HFD developed features of inflammatory anemia. When compared to lean controls (median hemoglobin read count: 68.7K), obese control mice exhibited dramatically reduced hemoglobin expression (median: 172 reads). Ofirnoflast treatment increased hemoglobin levels to a median of 83.3K reads, suggesting restoration of erythropoiesis. Erythropoietin levels were also elevated in Ofirnoflast-treated mice (1267 reads) relative to obese controls (967 reads). IHC analysis of adipose tissue revealed decreased ASC speck formation (a marker of NLRP3 activation), reduced macrophage infiltration, and diminished crown-like structures in Ofirnoflast-treated animals, consistent with a suppression of adipose tissue inflammation. Conclusion: Obesity-associated anemia of inflammation is driven in part by NLRP3-mediated inflammatory signaling. Pharmacologic inhibition of the NLRP3 inflammasome with Ofirnoflast reduced adipose inflammation and restored key hematologic parameters, including hemoglobin and erythropoietin. These findings support further evaluation of Ofirnoflast as a novel therapeutic for obesity-related anemia and chronic inflammation.