Targeting Inflammasome Activation Research Articles

Induction of the Inflammasome Pathway by Tyrosine Kinase Inhibitors Provides an Actionable Therapeutic Target for Hepatocellular Carcinoma.

During the last decade, tyrosine kinase inhibitors (TKIs) sorafenib and regorafenib have been standard systemic treatments for advanced hepatocellular carcinoma (HCC). Previous data associated sorafenib with inflammasome activation. However, the role of the inflammasome in sorafenib and regorafenib signaling has not been described in liver cancer patients. For this purpose, we analyzed inflammasome-related transcriptomic changes in a murine HCC model. Our data confirmed inflammasome activation after both TKI treatments, sharing a similar pattern of increased gene expression. According to human database results, transcriptional increase of inflammasome genes is associated with poorer prognosis for male liver cancer patients, suggesting a sex-dependent role for inflammasome activation in HCC therapy. In biopsies of HCC and its surrounding tissue, we detected durable increases in the inflammasome activation pattern after sorafenib or regorafenib treatment in male patients. Further supporting its involvement in sorafenib action, inflammasome inhibition (MCC950) enhanced sorafenib anticancer activity in experimental HCC models, while no direct in vitro effect was observed in HCC cell lines. Moreover, activated human THP-1 macrophages released IL-1β after sorafenib administration, while 3D Hep3B spheres displayed increased tumor growth after IL-1β addition, pointing to the liver microenvironment as a key player in inflammasome action. In summary, our results unveil the inflammasome pathway as an actionable target in sorafenib or regorafenib therapy and associate an inflammasome signature in HCC and surrounding tissue with TKI administration. Therefore, targeting inflammasome activation, principally in male patients, could help to overcome sorafenib or regorafenib resistance and enhance the efficacy of TKI treatments in HCC.

Targeting Inflammasome Activation in Viral Infection: A Therapeutic Solution?

Inflammasome activation is exclusively involved in sensing activation of innate immunity and inflammatory response during viral infection. Accumulating evidence suggests that the manipulation of inflammasome assembly or its interaction with viral proteins are critical factors in viral pathogenesis. Results from pilot clinical trials show encouraging results of NLRP3 inflammasome suppression in reducing mortality and morbidity in SARS-CoV-2-infected patients. In this article, we summarize the up-to-date understanding of inflammasomes, including NLRP3, AIM2, NLRP1, NLRP6, and NLRC4 in various viral infections, with particular focus on RNA viruses such as SARS-CoV-2, HIV, IAV, and Zika virus and DNA viruses such as herpes simplex virus 1. We also discuss the current achievement of the mechanisms involved in viral infection-induced inflammatory response, host defense, and possible therapeutic solutions.

The anti-caspase 1 inhibitor VX-765 reduces immune activation, CD4+ T cell depletion, viral load, and total HIV-1 DNA in HIV-1 infected humanized mice.

HIV-1 infection results in the activation of inflammasome that may facilitate viral spread and establishment of viral reservoirs. We evaluated the effects of the caspase-1 inhibitor VX-765 on HIV-1 infection in humanized NSG mice engrafted with human CD34+ hematopoietic stem cells. Expression of caspase-1, NLRP3, and IL-1β was increased in lymph nodes and bone marrow between day 1 and 3 after HIV-1 infection (mean fold change (FC) of 2.08, 3.23, and 6.05, p<0.001, respectively). IFI16 and AIM2 expression peaked at day 24 and coincides with increased IL-18 levels (6.89 vs 83.19 pg/ml, p=0.004), increased viral load and CD4+ T cells loss in blood (p<0.005 and p<0.0001, for the spleen respectively). Treatment with VX-765 significantly reduced TNF-α at day 11 (0.47 vs 2.2 pg/ml, p=0.045), IL-18 at day 22 (7.8 vs 23.2 pg/ml, p=0.04), CD4+ T cells (44.3% vs 36,7%, p=0.01), viral load (4.26 vs 4.89 log 10 copies/ml, p=0.027), and total HIV-1 DNA in the spleen (1 054 vs 2 889 copies /106 cells, p=0.029). We demonstrated that targeting inflammasome activation early after infection may represent a therapeutic strategy towards HIV cure to prevent CD4+ T cell depletion and reduce immune activation, viral load, and the HIV-1 reservoir formation.

Inflammasomes in Human Immunodeficiency Virus Type 1 Infection.

Innate immune responses are the host's first line of defense against human immunodeficiency virus type 1 (HIV-1) infection, with pattern recognition receptors detecting viral specific pathogen-associated molecular patterns and initiating antiviral responses. In response to HIV-1 nucleic acids or proteins, some pattern recognition receptors have the ability to assemble a large multiprotein complex called the inflammasome, which triggers pro-inflammatory cytokine release and a form of lytic programmed cell death called pyroptosis. Here, we review our current understanding of the mechanism of the inflammasome in sensing HIV-1 infection. Furthermore, we discuss the contribution of inflammasome activation in HIV-1 pathogenesis as well as potential strategies of targeting inflammasome activation for the treatment of HIV-1 infection.

Targeting Inflammasome Activation in COVID-19: Delivery of RNA Interference-Based Therapeutic Molecules.

Mortality and morbidity associated with COVID-19 continue to be significantly high worldwide, owing to the absence of effective treatment strategies. The emergence of different variants of SARS-CoV-2 is also a considerable source of concern and has led to challenges in the development of better prevention and treatment strategies, including vaccines. Immune dysregulation due to pro-inflammatory mediators has worsened the situation in COVID-19 patients. Inflammasomes play a critical role in modulating pro-inflammatory cytokines in the pathogenesis of COVID-19 and their activation is associated with poor clinical outcomes. Numerous preclinical and clinical trials for COVID-19 treatment using different approaches are currently underway. Targeting different inflammasomes to reduce the cytokine storm, and its associated complications, in COVID-19 patients is a new area of research. Non-coding RNAs, targeting inflammasome activation, may serve as an effective treatment strategy. However, the efficacy of these therapeutic agents is highly dependent on the delivery system. MicroRNAs and long non-coding RNAs, in conjunction with an efficient delivery vehicle, present a potential strategy for regulating NLRP3 activity through various RNA interference (RNAi) mechanisms. In this regard, the use of nanomaterials and other vehicle types for the delivery of RNAi-based therapeutic molecules for COVID-19 may serve as a novel approach for enhancing drug efficacy. The present review briefly summarizes immune dysregulation and its consequences, the roles of different non-coding RNAs in regulating the NLRP3 inflammasome, distinct types of vectors for their delivery, and potential therapeutic targets of microRNA for treatment of COVID-19.

724 Evaluation of an anti-human IL-1β antibody in monosodium urate crystals-induced peritonitis model in hIL-1β HuGEMM™ mice

BackgroundGout is a chronic inflammatory disease featuring the deposition of monosodium urate (MSU) crystals in the synovial fluid of patients, followed by NLRP3 (NOD-, LRR- and pyrin domain-containing protein 3) inflammasome activation and bioactive IL-1β release, which recruits neutrophils to the local inflammation sites. Blocking IL-1β function is becoming a a potent therapeutic approach for gout and gouty arthritis. Conventional MSU-induced peritonitis in C57BL/6 mice provides a simple and rapid evaluation of therapeutics targeting inflammasome activation. However, this murine model has limitations when it comes to the evaluation of human-specific antibodies, for example, anti-human IL-1β (anti-hIL-1β) monoclonal antibodies (mAb). Thus, a murine model to assess the efficacy of anti-hIL-1β mAb is needed. We have developed a hIL-1β knock-in mouse model (hIL-1β HuGEMM™), which is able to facilitate the pre-clinical evaluation of drugs targeting specific human biological molecules especially when mouse ortholog is not available. Therefore, an MSU crystals induced peritonitis model using hIL-1β HuGEMM™ mice provides a robust model to evaluate therapies targeting hIL-1β.MethodsMSU crystals were injected intraperitoneally into human IL-1β (hIL-1β) knock-in mice, where the coding sequence of mouse IL-1β was replaced by hIL-1β. Prior to MSU crystal administration, mice received treatment of either vehicle or anti-hIL-1β antibody. Six hours facilitate post MSU crystal injection, serum and lavage flushed with PBS were collected. Subsequently, cytokine protein levels in the serum were determined by MSD, and the population of polymorphonuclear leukocytes (PMNs) (live CD11b+ Ly-6GHi cells) in the lavage was analysed by flow cytometry.ResultsThe vehicle treatment group showed a dramatic increase in hIL-1β secretion and PMN leukocytes, in comparison to the group that did not receive MSU, which suggests a successful induction of acute inflammatory response in the peritoneal cavity. In contrast, mice that received a single administration of anti-hIL-1β antibody 24 hours prior to MSU injection exhibited a significantly lower level of hIL-1β when compared to the vehicle treatment group, which implies that the anti-hIL-1β mAb efficaciously neutralized hIL-1β secretion. In addition, TNF-α and IL-6, two further cytokines downstream of IL-1β, were significantly reduced in the anti-hIL-1β mAb treatment group. However, the PMN leukocyte infiltration in the anti-hIL-1β mAb treatment group did not change in comparison to the vehicle group.ConclusionsIn this study, an MSU crystals-induced peritonitis model was successfully established in hIL-1β HuGEMM mice, which has the potential to evaluate immune therapeutics with anti-hIL-1β blockades.

Flavonoids: Nutraceuticals for Rheumatic Diseases via Targeting of Inflammasome Activation.

Inflammation, an innate immune response that prevents cellular damage caused by pathogens, consists of two successive mechanisms, namely priming and triggering. While priming is an inflammation-preparation step, triggering is an inflammation-activation step, and the central feature of triggering is the activation of inflammasomes and intracellular inflammatory protein complexes. Flavonoids are natural phenolic compounds predominantly present in plants, fruits, and vegetables and are known to possess strong anti-inflammatory activities. The anti-inflammatory activity of flavonoids has long been demonstrated, with the main focus on the priming mechanisms, while increasing numbers of recent studies have redirected the research focus on the triggering step, and studies have reported that flavonoids inhibit inflammatory responses and diseases by targeting inflammasome activation. Rheumatic diseases are systemic inflammatory and autoimmune diseases that primarily affect joints and connective tissues, and they are associated with numerous deleterious effects. Here, we discuss the emerging literature on the ameliorative role of flavonoids targeting inflammasome activation in inflammatory rheumatic diseases.

IC100: a novel anti-ASC monoclonal antibody improves functional outcomes in an animal model of multiple sclerosis

BackgroundThe inflammasome adaptor apoptosis-associated speck-like protein containing a CARD (ASC) is involved in immune signaling by bridging the interactions between inflammasome sensors and caspase-1. Strong experimental evidence has shown that ASC−/− mice are protected from disease progression in animal models of multiple sclerosis (MS), suggesting that targeting inflammasome activation via ASC inhibition may be a promising therapeutic strategy in MS. Thus, the goal of our study is to test the efficacy of IC100, a novel humanized antibody targeting ASC, in preventing and/or suppressing disease in the experimental autoimmune encephalomyelitis (EAE) model of MS.MethodsWe employed the EAE model of MS where disease was induced by immunization of C57BL/6 mice with myelin oligodendrocyte glycoprotein peptide 35–55 (MOG35–55). Mice were treated with vehicle or increasing doses of IC100 (10, 30, and 45 mg/kg) and clinical disease course was evaluated up to 35 days post EAE induction. Immune cell infiltration into the spinal cord and microglia responses were assessed.ResultsWe show that IC100 treatment reduced the severity of EAE when compared to vehicle-treated controls. At a dose of 30 mg/kg, IC100 significantly reduced the number of CD4+ and CD8+ T cells and CD11b+MHCII+ activated myeloid cells entering the spinal cord from the periphery, and reduced the number of total and activated microglia.ConclusionsThese data indicate that IC100 suppresses the immune-inflammatory response that drives EAE development and progression, thereby identifying ASC as a promising target for the treatment of MS as well as other neurological diseases with a neuroinflammatory component.

Autophagy Protects Against Developing Increased Lung Permeability and Hypoxemia by Down Regulating Inflammasome Activity and IL-1β in LPS Plus Mechanical Ventilation-Induced Acute Lung Injury

Targeting inflammasome activation to modulate interleukin (IL)-1β is a promising treatment strategy against acute respiratory distress syndrome and ventilator-induced lung injury (VILI). Autophagy is a key regulator of inflammasome activation in macrophages. Here, we investigated the role of autophagy in the development of acute lung injury (ALI) induced by lipopolysaccharide (LPS) and mechanical ventilation (MV). Two hours before starting MV, 0.2 mg/kg LPS was administered to mice intratracheally. Mice were then placed on high-volume MV (30 ml/kg with 3 cmH2O positive end-expiratory pressure for 2.5 h without additional oxygen application). Mice with myeloid-specific deletion of the autophagic protein ATG16L1 (Atg16l1fl/fl LysMCre) suffered severe hypoxemia (adjusted p < 0.05) and increased lung permeability (p < 0.05, albumin level in bronchoalveolar lavage fluid) with significantly higher IL-1β release into alveolar space (p < 0.05). Induction of autophagy by fasting-induced starvation led to improved arterial oxygenation (adjusted p < 0.0001) and lung permeability (p < 0.05), as well as significantly suppressed IL-1β production (p < 0.01). Intratracheal treatment with anti-mouse IL-1β monoclonal antibody (mAb; 2.5 mg/kg) significantly improved arterial oxygenation (adjusted p < 0.01) as well as lung permeability (p < 0.05). On the other hand, deletion of IL-1α gene or use of anti-mouse IL-1α mAb (2.5 mg/kg) provided no significant protection, suggesting that the LPS and MV-induced ALI is primarily dependent on IL-1β, but independent of IL-1α. These observations suggest that autophagy has a protective role in controlling inflammasome activation and production of IL-1β, which plays a critical role in developing hypoxemia and increased lung permeability in LPS plus MV-induced acute lung injury.

The role of microglial inflammasome activation in pyroptotic cell death following penetrating traumatic brain injury

BackgroundTraumatic brain injury remains a significant cause of death and disability in the USA. Currently, there are no effective therapies to mitigate disability except for surgical interventions necessitating a need for continued research into uncovering novel therapeutic targets. In a recent study, we used a rodent model of penetrating traumatic brain injury known as penetrating ballistic-like brain injury (PBBI) to examine the role of innate immunity in post-traumatic secondary injury mechanisms. We previously reported that the inflammasome, a multiprotein complex composed of apoptosis-associated speck-like protein containing card and caspase-1, plays a role in secondary cell death mechanisms after PBBI, including inflammatory cell death (pyroptosis).MethodsIn the current study, we used flow cytometry analysis to evaluate activated microglia and CD11b-positive leukocytes after PBBI and assessed inflammasome activation and pyroptosis of specific cellular populations. Sprague-Dawley male rats underwent PBBI or sham-operated procedures and ipsilateral cortical regions processed for flow cytometry and cellular analysis. Flow cytometry results were compared using one-way ANOVA followed by Tukey’s multiple comparisons.ResultsAt 48 h following PBBI, there was an increase in activated microglia and infiltrating leukocytes compared to sham controls that were associated with increased caspase-1 activity. Using a florescent probe to identify caspase-1 activity and a fluorescent assay to determine cell viability, evidence for pyroptosis in CD11b+ cells was also determined. Finally, while post-traumatic treatment with an anti-ASC antibody had no effect on the number of activated microglia and infiltrating leukocytes, antibody treatment decreased caspase-1 activity in both resident microglia and infiltrating leukocytes and reduced pyroptotic CD11b+ cell death.ConclusionsThese results provide evidence for inflammasome activation in microglia and infiltrating leukocytes after penetrating traumatic brain injury and a role for pyroptotic cell death in the pathophysiology. In addition to inhibiting neuronal cell death, therapeutic treatments targeting inflammasome activation may also provide beneficial effects by reducing the potentially detrimental consequences of activated microglia and infiltrating CD11b+ leukocytes following penetrating traumatic brain injury.

Targeting the inflammasome as host directed therapy for Mycobacterium tuberculosis and HIV co-infection

Abstract Mycobacterium tuberculosis (Mtb) infection continues to pose a significant public health threat with 1.3 million deaths occurring annually. People with HIV are among the most susceptible to Mtb infection due to the microbial synergism occurring during co-infection which promotes inappropriate inflammation as a potential driver of lung pathology. However, the immunomodulatory mechanisms driving pulmonary inflammation and pathogenesis are not fully understood. Our preliminary studies suggest that synergistic inflammasome activation within macrophages, the cellular host of both Mtb and HIV, leads to a caspase-1 dependent increase in pro-inflammatory cytokines. Using a humanized mouse model of co-infection developed in our laboratory, we have evaluated the in vivo inhibition of caspase-1 using the pro-drug VX-765 to limit mature IL-1β secretion and therefore reduce pulmonary inflammation. Treatment with VX-765 in humanized and WT (BALB/C) mice resulted in a decrease of lung inflammatory cytokine and chemokine production. Mycobacterial burden in the lung and sites of dissemination were unaffected by VX-765 treatment, suggesting inhibition of caspase-1 activity did not suppress protective immune responses. By using monocyte-derived macrophages (MDM) for an in vitro co-infection assay, we observed reduced inflammatory responses to co-infection following caspase-1 inhibition. These preliminary studies suggest a potential role of VX-765 as a host directed therapeutic targeting inflammasome activation in co-infected patients. With host directed therapy emerging as a possible new line of “anti-AIDS” therapy, the impact of these drugs on Mtb infection and the course of treatment is of great clinical importance.

Therapeutically Targeting the Inflammasome Product in a Chimeric Model of Endometriosis-Related Surgical Adhesions.

Development of adhesions commonly occurs in association with surgery for endometriosis. Even in the absence of surgery, women with endometriosis appear to be at an enhanced risk of developing adhesions. In the current study, we utilized a chimeric mouse model of experimental endometriosis in order to examine the role of inflammasome activation in the development of postsurgical adhesions. Mice were randomized to receive peritoneal injections of human endometrial tissue fragments or endometrial tissue conditioned media (CM) from women with or without endometriosis 16 hours after ovariectomy and placement of an estradiol-releasing silastic capsule. A subset of mice receiving CM was also treated with interleukin (IL) 1 receptor antagonist (IL-1ra). Our studies demonstrate that peritoneal injection of endometrial tissue fragments near the time of surgery resulted in extensive adhesive disease regardless of tissue origin. However, adhesion scores were significantly higher in mice receiving CM from tissues acquired from patients with endometriosis compared to control tissue CM ( P = .0001). Cytokine bead array analysis of endometrial CM revealed enhanced expression of IL-1β from patients with endometriosis compared to controls ( P < .01). Finally, the ability of human tissue CM to promote adhesive disease was dramatically reduced in mice cotreated with IL-1ra ( P < .0001). Our data implicate enhanced expression of IL-1β in women with endometriosis as a potential causal factor in their increased susceptibility of developing postsurgical adhesions. Thus, targeting inflammasome activation may be an effective strategy for the prevention of surgical adhesions in patients with endometriosis.

The NLRP3 and Pyrin Inflammasomes: Implications in the Pathophysiology of Autoinflammatory Diseases.

Inflammasomes are multiprotein complexes that critically control different aspects of innate and adaptive immunity. Among them we could highlight the release of pro-inflammatory cytokines that induce and maintain the inflammatory response. Usually, inflammasomes result from oligomerization of a nucleotide-binding domain-like receptor (NLR) after sensing different pathogenic or endogenous sterile dangerous signals; however, other proteins such as absent in melanoma 2, retinoic acid-inducible gene I, or pyrin could also form inflammasome platforms. Inflammasome oligomerization leads to caspase-1 activation and the processing and release of the pro-inflammatory cytokines, such as interleukin (IL)-1β and IL-18. Mutations in different inflammasomes are causative for multiple periodic hereditary syndromes or autoinflammatory diseases, characterized by acute systemic inflammatory flares not associated with infections, tumors, or autoimmunity. This review focuses on germline mutations that have been described in cryopyrin-associated periodic syndrome (CAPS) for NLRP3 or in familial Mediterranean fever (FMF) and pyrin-associated autoinflammation with neutrophilic dermatosis (PAAND) for MEFV. Besides the implication of inflammasomes in autoinflammatory syndromes, these molecular platforms are involved in the pathophysiology of different illnesses, including chronic inflammatory diseases, degenerative processes, fibrosis, or metabolic diseases. Therefore, drug development targeting inflammasome activation is a promising field in expansion.

Chenodeoxycholic acid activates NLRP3 inflammasome and contributes to cholestatic liver fibrosis.

Accumulation of hydrophobic bile acids in the liver contributes to cholestatic liver injury. Inflammation induced by excessive bile acids is believed to play a crucial role, however, the mechanisms of bile acids triggered inflammatory response remain unclear. Recent studies have highlighted the effect of NLRP3 inflammasome in mediating liver inflammation and fibrosis. In this study, we for the first time showed that chenodeoxycholic acid (CDCA), the major hydrophobic primary bile acid involved in cholestatic liver injury, could dose-dependently induce NLRP3 inflammasome activation and secretion of pro-inflammatory cytokine-IL-1β in macrophages by promoting ROS production and K+ efflux. Mechanistically, CDCA triggered ROS formation in part through TGR5/EGFR downstream signaling, including protein kinase B, extracellular regulated protein kinases and c-Jun N-terminal kinase pathways. Meanwhile, CDCA also induced ATP release from macrophages which subsequently causes K+ efflux via P2X7 receptor. Furthermore, in vivo inhibition of NLRP3 inflammasome with caspase-1 inhibitor dramatically decreased mature IL-1β level of liver tissue and ameliorated liver fibrosis in bile duct ligation (BDL) mouse model. In conclusion, excessive CDCA may represent an endogenous danger signal to activate NLRP3 inflammasome and initiate liver inflammation during cholestasis. Our finding offers a mechanistic basis to ameliorate cholestatic liver fibrosis by targeting inflammasome activation.

Multiple inflammasome complexes are activated in autistic spectrum disorders

BackgroundInflammasomes are multimeric protein platforms involved in the regulation of inflammatory responses whose activity results in the production of proinflammatory cytokines. Because neuroinflammation is observed in autistic spectrum disorders (ASD), a neurologic condition of childhood resulting in a complex behavioural impairment, we analyzed the inflammasomes activity in ASD. Additionally we verified whether alterations of the gastrointestinal (GI) barriers might play a role in inflammasomes activation. MethodsThe activity of the inflammasomes, the concentration of the inflammasomes-derived proinflammatory cytokines interleukin (IL)-1β and IL-18, and serum parameters of GI damage were analyzed in 25 ASD children, 23 healthy siblings (HS) and 30 unrelated age-matched healthy controls (HC). ResultsA significant upregulation of the AIM2 and the NLRP3 inflammasomes and an increased production of IL-1β and IL-18 that was associated with a consistent reduction of IL-33, an anti inflammation cytokine were observed in ASD alone. Notably, in a possible immune-mediated attempt to dampen inflammation, IL-37, a suppressor of innate inflammatory responses, was significantly augmented in these same children. Finally, intestinal fatty acid binding protein (IFABP), an index of altered GI permeability, was significantly increased in serum of ASD and HS. ConclusionsThese results show that the inflammasomes are activated in ASD and shed light on the molecular mechanisms responsible for ASD-associated neuroinflammation. The observation that GI alterations could be present as well in ASD offers a possible link between such alterations and neuroinflammation. Therapeutic strategies targeting inflammasome activation could be useful in ASD.

The NLRP3 and NLRP1 inflammasomes are activated in Alzheimer's disease.

BackgroundInterleukin-1 beta (IL-1β) and its key regulator, the inflammasome, are suspected to play a role in the neuroinflammation observed in Alzheimer’s disease (AD); no conclusive data are nevertheless available in AD patients.ResultsmRNA for inflammasome components (NLRP1, NLRP3, PYCARD, caspase 1, 5 and 8) and downstream effectors (IL-1β, IL-18) was up-regulated in severe and MILD AD. Monocytes co-expressing NLRP3 with caspase 1 or caspase 8 were significantly increased in severe AD alone, whereas those co-expressing NLRP1 and NLRP3 with PYCARD were augmented in both severe and MILD AD. Activation of the NLRP1 and NLRP3 inflammasomes in AD was confirmed by confocal microscopy proteins co-localization and by the significantly higher amounts of the pro-inflammatory cytokines IL-1β and IL-18 being produced by monocytes. In MCI, the expression of NLRP3, but not the one of PYCARD or caspase 1 was increased, indicating that functional inflammasomes are not assembled in these individuals: this was confirmed by lack of co-localization and of proinflammatory cytokines production.ConclusionsThe activation of at least two different inflammasome complexes explains AD-associated neuroinflammation. Strategies targeting inflammasome activation could be useful in the therapy of AD.

Abstract 3173: The NLRC4 inflammasome promotes breast cancer progression in diet-induced obese mice

Abstract Obesity is not only a known risk factor for the development of ER+ breast cancer in postmenopausal woman, but is also associated with a poor clinical outcome across all subtypes of breast cancer. Obesity is also one of the only known risk factors for the development of invasive ductal carcinoma (IDC) versus ductal carcinoma in-situ (DCIS; non-invasive). A number of hypotheses have been proposed to explain the association between obesity and breast cancer progression. One such hypothesis is that the chronic inflammation in adipose tissue associated with obesity, especially adipose tissue in the mammary glands, creates a microenvironment that promotes breast cancer progression by acting as a reservoir of pro-inflammatory cytokines (including interleukin IL-1β and IL-6). One source of pro-inflammatory cytokines in adipose tissue is activated inflammasomes in infiltrating macrophages. Inflammasomes are multiprotein complexes whose main function is the activation of IL-1β and IL-18 during inflammation via Caspase 1-mediated proteolytic cleavage. Chronic inflammation, NLRP3 inflammasome activation and IL-1β are thought to be a major cause of obesity-associated insulin resistance and are involved in many human diseases including type-2 diabetes and cancer. However, the mechanism linking obesity and breast cancer progression remains unknown. We hypothesize that obesity-induced inflammasome activation in mammary adipose tissue results in active IL-1β which in turn promotes breast cancer progression. Using a syngeneic orthotopic transplant model of breast cancer in C57Bl/6 mice (Py8119 cells and E0771 cells) we have shown that diet-induced obese (DIO) mice (mice fed a high fat diet) have increased tumor growth and metastasis compared to control mice (mice fed normal chow). Tumors from DIO mice have increased macrophage infiltration and caspase-1 activation compared to tumors from control mice. Additional studies have shown that treating DIO mice with an IL1R1 (the receptor for IL-1) blocking antibody reduces tumor growth to that observed in control mice. Furthermore, NLRC4 deficient DIO mice had reduced tumor growth (compared to wild-type DIO) mice, while there was no decrease in tumor growth in NLRP3 deficient DIO mice, indicating that NLRC4 inflammasome activation promotes breast cancer growth in obese mice. Finally, tumors from DIO mice had increased vascularization (as evidenced by an increase in CD31+ cells) compared to control mice, which was decreased in NLRC4 deficient mice or mice treated with the anti-IL1R1 antibody. This data indicates that obesity induced NLRC4 inflammasome activation in infiltrating macrophages in the mammary gland promotes breast cancer progression possibly through increased angiogenesis in the tumor microenvironment. Thus, targeting inflammasome activation or the IL-1/IL1R1 axis represents a rational target for the treatment of breast cancer in obese patients. Citation Format: Ryan Kolb, Nicholas Borcherding, Yinghong Liu, Fang Yuan, Qing Xie, Fayyaz Sutterwala, Weizhou Zhang. The NLRC4 inflammasome promotes breast cancer progression in diet-induced obese mice. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3173. doi:10.1158/1538-7445.AM2015-3173

Abstract 4098: Inflammasome activation in obesity-associated breast cancer progression

Abstract The purpose of this research is to determine the role of inflammasome-mediated production of interleukin(IL)-1β in obesity-associated breast cancer progression. Previous studies have shown that obesity is a risk factor for both developing breast cancer and breast cancer progression. Obesity is also associated with the development of invasive ductal carcinoma (IDC) versus ductal carcinoma in-situ (DCIS; non-invasive). A number of hypotheses have been proposed to explain the association between obesity and breast cancer progression. It has been hypothesized that the chronic inflammation in adipose tissue associated with obesity, especially adipose tissue in the mammary glands, creates a microenvironment that promotes breast cancer progression by acting as a reservoir of pro-inflammatory cytokines (including IL-1β and IL-6). One source of pro-inflammatory cytokines in adipose tissue is activated inflammasomes in infiltrating macrophages. Inflammasomes are multiprotein complexes whose main function is the activation of IL-1β and IL-18 during inflammation via Caspase 1 (Casp1)-mediated proteolytic cleavage. Chronic inflammation, Nlrp3 inflammasome activation and IL-1β are thought to be a major cause of obesity-associated insulin resistance and are involved in many human diseases including type-2 diabetes and cancer. However, the mechanism linking obesity and breast cancer progression remains unknown. We hypothesize that obesity-induced inflammasome activation in mammary adipose tissue results in active IL-1β which in turn promotes breast cancer progression. To determine the role of inflammasome activation in obesity-associated breast cancer progression, we will use inflammasome-deficient mice or neutralizing antibodies to inhibit IL-1 signaling in a syngeneic orthotopic transplant model of Py8119 cells in obese mice (mice fed a high fat diet). Our results show that Py8119 tumors have increased tumor growth and metastasis in obese mice compared to lean mice (mice fed normal chow). Furthermore, tumors from obese mice had increased expression of IL-1β and its receptor IL1 receptor 1 (IL1R1) as well as increased Casp1. Tumors from obese mice also had increase in M1 (which can secrete IL-1β) and M2 (which can be recruited in response to IL-1β signaling) tumor-infiltrating macrophages. Finally, inhibition of IL-1β signaling using an IL1R1 neutralizing antibody or Casp1 deficiency results in reduced tumor growth in obese mice. In conclusion, our results suggest that obesity-induced inflammasome activation promotes breast cancer progression in obese mice. Targeting inflammasome activation or neutralizing the IL-1/IL1R axis represents a rational therapy to treat obese patients with invasive breast cancer. Citation Format: Ryan Kolb, Yinghong Liu, Qing Xie, Nicholas Borcherding, Wei Li, Weizhou Zhang. Inflammasome activation in obesity-associated breast cancer progression. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4098. doi:10.1158/1538-7445.AM2014-4098

Inflammasomes: Caspase-1-Activating Platforms with Critical Roles in Host Defense

Activation of the inflammatory cysteine protease caspase-1 in inflammasome complexes plays a critical role in the host response to microbial infections. Inflammasome activation induces inflammation through secretion of the pro-inflammatory cytokines interleukin (IL)-1β and IL-18 and through extracellular release of the alarmin high mobility group box 1. Moreover, caspase-1 activation by inflammasomes counters bacterial replication and induces pyroptosis, a specialized cell death program that removes infected immune cells as part of the host defense system. It is thus not surprising that bacterial and viral pathogens evolved virulence factors targeting inflammasome activation and activity. Here, we provide an overview of the distinct inflammasome complexes that are activated in a pathogen-specific manner and discuss the diverse strategies employed by viruses and bacteria to modulate inflammasome function.