Numerous studies demonstrate that neuroinflammation is a key player in the progression of Alzheimer’s disease (AD). Interleukin (IL)-1β is a main inducer of inflammation and therefore a prime target ...for therapeutic options. The inactive IL-1β precursor requires processing by the the nucleotide-binding oligomerization domain-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome into a mature and active form. Studies have shown that IL-1β is up-regulated in brains of patients with AD, and that genetic inactivation of the NLRP3 inflammasome improves behavioral tests and synaptic plasticity phenotypes in a murine model of the disease. In the present study, we analyzed the effect of pharmacological inhibition of the NLRP3 inflammasome using dapansutrile (OLT1177), an oral NLRP3-specific inhibitor that is safe in humans. Six-month-old WT and APP/PS1 mice were fed with standard mouse chow or OLT1177-enriched chow for 3 mo. The Morris water maze test revealed an impaired learning and memory ability of 9-mo-old APP/PS1 mice (P = 0.001), which was completely rescued by OLT1177 fed to mice (P = 0.008 to untreated APP/PS1). Furthermore, our findings revealed that 3 mo of OLT1177 diet can rescue synaptic plasticity in this mouse model of AD (P = 0.007 to untreated APP/PS1). In addition, microglia were less activated (P = 0.07) and the number of plaques was reduced in the cortex (P = 0.03) following NLRP3 inhibition with OLT1177 administration. We also observed an OLT1177 dose-dependent normalization of plasma metabolic markers of AD to those of WT mice. This study suggests the therapeutic potential of treating neuroinflammation with an oral inhibitor of the NLRP3 inflammasome.
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
IL-1β and IL-18 are pro-inflammatory cytokines that are linked to inflammation. Activation of the NOD-like receptor protein 3 (NLRP3) inflammasome is involved in the maturation and secretion of IL-1β ...and IL-18 and, thus, plays a key role in the pathogenesis of many inflammatory conditions, including multiple sclerosis (MS). OLT1177™ (Dapansutrile) is a newly developed drug that is safe in humans and inhibits specifically the NLRP3 inflammasome. In the present study, we investigated whether OLT1177 exerts therapeutic effects in experimental autoimmune encephalomyelitis (EAE), a mouse model of MS. We found that EAE mice fed an OLT1177-enriched diet prophylactically were significantly protected against functional deficits and demyelination in the spinal cord. We also demonstrated that prophylactic oral administration of OLT1177 led to marked reduction (~2- to 3-fold) in the protein levels of IL-1β and IL-18, as well as, IL-6 and TNFα, in the spinal cord of EAE mice. Moreover, prophylactic oral administration of OLT1177 significantly attenuated the infiltration of CD4 T cells and macrophages in the spinal cord. We also demonstrated that oral administration of OLT1177, starting at disease onset, resulted in significant amelioration of the clinical signs of EAE. Overall, these first data suggest that OLT1177 could have clinical benefit for the treatment of MS in humans.
Activation of the NLRP3 inflammasome induces maturation of IL-1β and IL-18, both validated targets for treating acute and chronic inflammatory diseases. Here, we demonstrate that OLT1177, an orally ...active β-sulfonyl nitrile molecule, inhibits activation of the NLRP3 inflammasome. In vitro, nanomolar concentrations of OLT1177 reduced IL-1β and IL-18 release following canonical and noncanonical NLRP3 inflammasome activation. The molecule showed no effect on the NLRC4 and AIM2 inflammasomes, suggesting specificity for NLRP3. In LPS-stimulated human blood-derived macrophages, OLT1177 decreased IL-1β levels by 60% and IL-18 by 70% at concentrations 100-fold lower in vitro than plasma concentrations safely reached in humans. OLT1177 also reduced IL-1β release and caspase-1 activity in freshly obtained human blood neutrophils. In monocytes isolated from patients with cryopyrin-associated periodic syndrome (CAPS), OLT1177 inhibited LPS-induced IL-1β release by 84% and 36%. Immunoprecipitation and FRET analysis demonstrated that OLT1177 prevented NLRP3-ASC, as well as NLRP3-caspase-1 interaction, thus inhibiting NLRP3 inflammasome oligomerization. In a cell-free assay, OLT1177 reduced ATPase activity of recombinant NLRP3, suggesting direct targeting of NLRP3. Mechanistically, OLT1177 did not affect potassium efflux, gene expression, or synthesis of the IL-1β precursor. Steady-state levels of phosphorylated NF-κB and IkB kinase were significantly lowered in spleen cells from OLT1177-treated mice. We observed reduced IL-1β content in tissue homogenates, limited oxidative stress, and increased muscle oxidative metabolism in OLT1177-treated mice challenged with LPS. Healthy humans receiving 1,000 mg of OLT1177 daily for 8 d exhibited neither adverse effects nor biochemical or hematological changes.
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
Spinal cord injury (SCI) leads to irreversible functional deficits due to the disruption of axons and the death of neurons and glial cells. The inflammatory response that occurs in the injured spinal ...cord results in tissue degeneration; thus, targeting inflammation after acute SCI is expected to ameliorate histopathological evidence indicative of damage and, consequently, reduce functional disabilities. Interleukin 1 beta (IL-1β) and interleukin 18 (IL-18) are pro-inflammatory cytokines members of the IL-1 family that initiate and propagate inflammation. Here, we report that protein levels of IL-1β and IL-18 were increased in spinal cord parenchyma after SCI, but with different expression profiles. Whereas levels of IL-1β were rapidly increased reaching peak levels at 12 h after the injury, levels of IL-18 did not increase until 7 days after the injury. Since activation of the NLRP3 inflammasome is required for the processing and release of IL-1β and IL-18, we intraperitoneally administered OLT1177, a selective inhibitor of the NLRP3 inflammasome, to reduce the contribution of these cytokines to SCI. At a dose of 200 mg/kg, OLT1177 protected against neurological deficits and histological evidence of damage. OLT1177 also reduced the levels of IL-1β in the spinal cord after contusion injury and diminished the accumulation of neutrophils and macrophages at later time points. These data suggest that targeting the NLRP3 inflammasome with OLT1177 could be a novel therapeutic strategy to arrest neuroinflammation and reduce functional impairments after acute SCI in humans.
•NLRP3 inflammasome is activated in the spinal cord after contusion injury•OLT1177 inhibits activation of the NLRP3 inflammasome in the injured spinal cord•OLT1177 reduces the accumulation of neutrophils and macrophages in the lesioned spinal cord•OLT1177 minimizes locomotor deficits and myelin loss after spinal cord injury
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Abstract
Background
The symptoms of asthma arise on the basis of a chronic inflammation of the airways. The NLRP3 inflammasome plays a central role in regulating inflammation e.g. by activating ...interleukin (IL) 1 cytokines, which in turn promote allergic inflammation. This study aims to evaluate the therapeutic potential of the specific NLRP3 inhibitor OLT1177® on experimental allergic asthma (EAA) in mice.
Methods
The impact of OLT1177® on NLRP3 expression, caspase-1 activation, and IL-1β release were assessed in vivo and in allergen-restimulated mononuclear cells (MNC) from ovalbumin (OVA)-sensitized mice. The effect of intra-peritoneal OLT1177®-treatment on OVA- and house dust mite (HDM) EAA and on poly(I:C)-triggered exacerbation of EAA were assessed. Furthermore, the effect of oropharyngeal aspiration of OLT1177® and oral OLT1177® via an enriched diet were also evaluated in mice with OVA-induced EAA.
Results
OLT1177® reduced NLRP3 expression, caspase-1 activation, and levels of activated IL-1β in vitro and in mice with OVA-induced EAA. Intra-peritoneal treatment with OLT1177® significantly diminished allergic airway inflammation, mucus hyperproduction, and airway hyperresponsiveness (AHR) in mice with OVA- and HDM-induced EAA as well as in mice with poly(I:C)-triggered exacerbation of EAA. While oropharyngeal aspiration of OLT1177® had no effect on EAA, oral OLT1177® via feeding an enriched diet markedly reduced all pathophysiologic hallmarks of EAA in mice.
Conclusion
Intra-peritoneal and oral delivery of OLT1177® is effective to inhibit the NLRP3 inflammasome in vivo and to treat EAA in mice.
This study was mainly supported by the German Center for Lung Research (DZL). The study was also supported by Olatec Therapeutics LLC and the Interleukin Foundation.
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