A Novel Pharmacologic Inhibitor of the NLRP3 Inflammasome Limits Myocardial Injury After Ischemia–Reperfusion in the Mouse

Abstract

Background The formation of the NLRP3 inflammasome in the heart during AMI amplifies the inflammatory response and mediates further damage. Glyburide has NLRP3-inhibitory activity in vitro, but requires very high doses in vivo, associated with hypoglycemia. The aim of this study was to measure the effects on the NLRP3 inflammasome of 16673-34-0, an intermediate substrate free of the cyclohexylurea moiety, involved in insulin release. Methods and Results We synthetized 16673-34-0 (5-Chloro-2-methoxy-N-[2-(4-sulfamoylphenyl)ethyl]benzamide) that displayed no effect on glucose metabolism. HL-1 cardiomyocytes were treated with LPS+ATP to induce the formation of the NLRP3 inflammasome measured as increased caspase-1 activity and cell death, and 16673-34-0 prevented such effects. 16673-34-0 was well tolerated with no effects on the glucose levels in vivo. Treatment with 16673-34-0 in a model of AMI due to ischemia+reperfusion significantly inhibited the activity of inflammasome (caspase-1) in the heart by 90% (P<0.01) and reduced infarct size, measured at pathology (by >40%, P<0.01) and with troponin I levels (by >70%, P<0.01). Conclusions The small molecule 16673-34-0, an intermediate substrate in the glyburide synthesis free of the cyclohexylurea moiety, inhibits the formation of the NLRP3 inflammasome in cardiomyocytes and limits the infarct size following myocardial ischemia/reperfusion in the mouse, without affecting glucose metabolism.

Molecular Regulation of Cell Fate in Cerebral Ischemia: Role of the Inflammasome and Connected Pathways

“…40 This is in agreement with the observation that inflammasome activation of cardiac fibroblasts is essential for myocardial ischemia/reperfusion injury: 99 inhibition, silencing, or genetic deletion of NLRP3 in the mouse limited the infarct size in experimental acute myocardial infarction. [100][101][102] Intracerebroventricular injection of different pro-caspase-1 inhibitors was shown to protect against ischemic injury up to 7 days after induction of cerebral ischemia in rodents 31,103,104 and NLPR1 inhibition by the use of a blocking antibody demonstrated protective effects in a thromboembolic model 24 hours after induction of cerebral ischemia. 3 Accordingly, NLRP3 deficiency protects against experimental stroke: NLPR3-knockout mice have reduced infarct volumes, and there was reduced neuronal death in vitro, when co-incubated with OGD (oxygen glucose deprivation)-treated microglial cells with reduced NLRP3 expression.…”

Inflammatory and fibrotic responses of cardiac fibroblasts to myocardial damage associated molecular patterns (DAMPs)

“…This resulted in a marked decrease in cell damage (troponin I levels) and infarct size after 24 h [136] and reduced LV dysfunction after 7 days [137]. Additionally, the NLRP3 inhibitor reduced LV dysfunction and remodelling in mice 1 week after permanent LAD ligation without affecting infarct scar size [137].…”

“…Interestingly, a novel small molecule inhibitor of NLRP3 inflammasome formation has been developed (16673-34-0) that can reduce caspase-1 activity in the heart by >90% in mice subjected to experimental MI with reperfusion [136]. This resulted in a marked decrease in cell damage (troponin I levels) and infarct size after 24 h [136] and reduced LV dysfunction after 7 days [137].…”

“…Other attractive DAMP receptor targets include the inflammasome receptor NLRP3. A novel pharmacological NLRP3 inhibitor improved cardiac function in mouse models of ischaemic and non-ischaemic myocardial injury [136,137].…”

Reply to Letter Regarding Article, “NLRP3 Inflammasome as a Therapeutic Target in Myocardial Infarction”

“…Moreover, we congratulate them for developing a novel pharmacological inhibitor of NLRP3 inflammasomes. According to their report, 2) they synthesized 16673-34-0 (5-chloro-2-methoxy-N-[2-(4-sulfamoylphenyl)-ethyl]-benzamide), an intermediate in the synthesis of glyburide, that is free of the cyclohexylurea moiety involved in insulin release, and found that this compound almost completely inhibited ATP-induced IL-1β secretion in J774 macrophages and ATP-induced caspase-1 activation and cell death in HL-1 cardiomyocytes in vitro. This NLRP3 inhibitor also reduced caspase-1 activation and infarct size in a murine model of myocardial ischemia-reperfusion (I/R) injury in vivo.…”