Non-canonical NF-κB Pathway Research Articles

BAFF Promotes FLS Activation Through BAFFR-Mediated Non-canonical NF-κB Pathway and the Effects of CP-25.

B cell activating factor (BAFF) has been shown to play a key role in regulating B cell function, but little is known about whether BAFF affects the function of fibroblast-like synoviocyte (FLS), an effector cell of rheumatoid arthritis (RA). CP-25, a new ester derivative of paeoniflorin, could alleviate the arthritis symptoms of collagen-induced arthritis (CIA) mice by inhibiting BAFF-mediated abnormal activation of B cells. In this study, we aimed to understand the mechanism by which BAFF activates FLS and the effect of CP-25 on FLS function. Therefore, the proliferation and migration abilities of FLS and key proteins on the non-canonical NF-κB pathway were examined. The results showed that compared with the FLS of normal rats/OA patients, the expression of BAFF-R, TRAF2, NIK, p-IKKα, P100, and P52 was higher in the FLS of AA rats/RA patients, while the expression of TRAF3 was lower. And, BAFF promotes FLS activation by activating the non-canonical NF-κB signaling pathway. Meanwhile, BAFFR-siRNA inhibited the proliferation of FLS and the activation of non-canonical NF-κB signaling in FLS induced by BAFF. Additionally, CP-25 could inhibit abnormal proliferation and migration of FLS by regulating non-canonical NF-κBsignaling. We concluded that BAFF may act as an important role in facilitating the function of FLS through the BAFFR-mediated non-canonical NF-κB pathway, which would be useful for revealing the pathological mechanism of RA. And CP-25 may become a potential new drug for the treatment of RA, providing a scientific basis for the development of new drugs to treat RA.

Anti-Rheumatoid Arthritis Effects of Dangshen Ejiao Pill Mediated through Nuclear Factor-κB Pathway

Dangshen Ejiao (DE) pill, a traditional Chinese hospital preparation, has been used for the treatment of rheumatoid arthritis (RA) in China for decades with an unknown mechanism. This study aims to investigate the anti-RA and anti-inflammatory effect of DE using a Collagen-Induced Arthritis (CIA) model and explore its underlying mechanism of action. The quality control analysis of DE was performed by the phytochemical determination of 9 main monomers. The anti-RA effect of DE was validated by its reduction of inflammatory responses and CCP level in the CIA model. DE inhibited the production of pro-inflammatory cytokines IL-1β and IL-6 both in vivo and in vitro. Further mechanism study revealed that DE might alleviate inflammatory responses in the treatment of RA by blocking both canonical and noncanonical NF-κB pathways, and thus provided a powerful scientific foundation for its expanded application and approval as an anti-RA drug.

LCL161 enhances expansion and survival of engineered anti-tumor T cells but is restricted by death signaling.

The genesis of SMAC mimetic drugs is founded on the observation that many cancers amplify IAP proteins to facilitate their survival, and therefore removal of these pathways would re-sensitize the cells towards apoptosis. It has become increasingly clear that SMAC mimetics also interface with the immune system in a modulatory manner. Suppression of IAP function by SMAC mimetics activates the non-canonical NF-κB pathway which can augment T cell function, opening the possibility of using SMAC mimetics to enhance immunotherapeutics. We have investigated the SMAC mimetic LCL161, which promotes degradation of cIAP-1 and cIAP-2, as an agent for delivering transient costimulation to engineered BMCA-specific human TAC T cells. In doing so we also sought to understand the cellular and molecular effects of LCL161 on T cell biology. LCL161 activated the non-canonical NF-κB pathway and enhanced antigen-driven TAC T cell proliferation and survival. Transcriptional profiling from TAC T cells treated with LCL161 revealed differential expression of costimulatory and apoptosis-related proteins, namely CD30 and FAIM3. We hypothesized that regulation of these genes by LCL161 may influence the drug's effects on T cells. We reversed the differential expression through genetic engineering and observed impaired costimulation by LCL161, particularly when CD30 was deleted. While LCL161 can provide a costimulatory signal to TAC T cells following exposure to isolated antigen, we did not observe a similar pattern when TAC T cells were stimulated with myeloma cells expressing the target antigen. We questioned whether FasL expression by myeloma cells may antagonize the costimulatory effects of LCL161. Fas-KO TAC T cells displayed superior expansion following antigen stimulation in the presence of LCL161, suggesting a role for Fas-related T cell death in limiting the magnitude of the T cell response to antigen in the presence of LCL161. Our results demonstrate that LCL161 provides costimulation to TAC T cells exposed to antigen alone, however LCL161 did not enhance TAC T cell anti-tumor function when challenged with myeloma cells and may be limited due to sensitization of T cells towards Fas-mediated apoptosis.

Cell-Penetrating Peptide Conjugated Au Nanoclusters Selectively Suppress Refractory Lymphoma Cells via Targeting Both Canonical and Noncanonical NF-κB Signaling Pathways.

Activated B cell-like diffuse large B-cell lymphoma (ABC-DLBCL) is the most aggressive form of DLBCL, with a significantly inferior prognosis due to resistance to the standard R-CHOP immunochemotherapy. Survival of ABC-DLBCL cells addicted to the constitutive activations of both canonical and noncanonical NF-κB signaling makes them attractive therapeutic targets. However, a pharmaceutical approach simultaneously targeting the canonical and noncanonical NF-κB pathway in the ABC-DLBCL cell is still lacking. Peptide-conjugated gold nanoclusters (AuNCs) have emerged unique intrinsic biomedical activities and possess a great potential in cancer theranostics. Here, we demonstrated a Au25 nanocluster conjugated by cell-penetrating peptides that can selectively repress the growth of ABC-DLBCL cells by inducing efficient apoptosis, more efficiently than glutathione (GSH)-conjugated AuNCs. The mechanism study showed that the cell-penetrating peptides enhanced the cellular internalization efficiency of AuNCs, and the selective repression in ABC-DLBCL cells is due to the inhibition of inherent constitutive canonical and noncanonical NF-κB activities by AuNCs. Several NF-κB target genes involved in chemotherapy resistance in ABC-DLBCL cells, including anti-apoptotic Bcl-2 family members and DNA damage repair proteins, were effectively down-regulated by the AuNC. The emerged novel activity of AuNCs in targeting both arms of NF-κB signaling in ABC-DLBCL cells may provide a promising candidate and a new insight into the rational design of peptide-conjugated Au nanomedicine for molecular targeting treatment of refractory lymphomas.

Comparison of the safety and efficacy of fingolimod and tofacitinib in the zebrafish model of colitis.

Oral targeted small molecules, including sphingosine 1 phosphate receptor (S1PR) modulators and tyrosine kinase inhibitors (TKIs), seem to revolutionize the management of inflammatory bowel disease (IBD). To select the most effective treatment, there is an unmet need to comparatively study their mechanism of action, efficacy, and toxicity in the preclinical stage and further translate it into clinical practice. Using 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced adult zebrafish colitis model, LC50 of fingolimod and tofacitinib were determined based on the acute toxicity test to compare aquatic toxicity potential. Subsequently, the efficacy of different concentrations of tofacitinib and fingolimod was compared using flow cytometry, qPCR, and histopathology analyses. TNBS significantly reduced the length of villi, and the number of goblet cells increased the level of TNF-α, MyD88, and NF-κB2, the thickness of villi and necrosis, and induced histopathological changes. All of these parameters were reversed almost dose-dependently with both medications, with the highest concentration of fingolimod being superior to other groups. Additionally, results from qPCR analysis suggested that these medications might suppress canonical and non-canonical NF-κB pathways by targeting toll-like receptors and MyD88. LC50 of tofacitinib and fingolimod was 0.9014 and 0.36mg/L, respectively. Hence, both are in the cory 1 of the Global Harmonization System (GHS) aquatic toxicity and are toxic to adult zebrafish life. Given the better efficacy of fingolimod, it is worth translating the effectiveness and safety of S1PR modulators into IBD patients and comparing them with TKIs in head-to-head studies; albeit, their toxicity should not be overlooked.

Sustained activation of non-canonical NF-κB signalling drives glycolytic reprogramming in doxorubicin-resistant DLBCL.

DLBCL is the most common lymphoma with high tumor heterogeneity. Treatment refractoriness and relapse from R-CHOP therapy in patients remain a clinical problem. Activation of the non-canonical NF-κB pathway is associated with R-CHOP resistance. However, downstream targets of non-canonical NF-κB mediating R-CHOP-induced resistance remains uncharacterized. Here, we identify the common mechanisms underlying both intrinsic and acquired resistance that are induced by doxorubicin, the main cytotoxic component of R-CHOP. We performed global transcriptomic analysis of (1) a panel of resistant versus sensitive and (2) isogenic acquired doxorubicin-resistant DLBCL cell lines following short and chronic exposure to doxorubicin respectively. Doxorubicin-induced stress in resistant cells activates a distinct transcriptional signature that is enriched in metabolic reprogramming and oncogenic signalling. Selective and sustained activation of non-canonical NF-κB signalling in these resistant cells exacerbated their survival by augmenting glycolysis. In response to doxorubicin, p52-RelB complexes transcriptionally activated multiple glycolytic regulators with prognostic significance through increased recruitment at their gene promoters. Targeting p52-RelB and their targets in resistant cells increased doxorubicin sensitivity in vitro and in vivo. Collectively, our study uncovered novel molecular drivers of doxorubicin-induced resistance that are regulated by non-canonical NF-κB pathway. We reveal new avenues of therapeutic targeting for R-CHOP-treated refractory/relapsed DLBCL patients.

CARD11 mutation and HBZ expression induce lymphoproliferative disease and adult T-cell leukemia/lymphoma

Adult T-cell leukemia/lymphoma (ATL) is caused by human T-cell leukemia virus type 1 (HTLV-1). In addition to HTLV-1 bZIP factor (HBZ), a leukemogenic antisense transcript of HTLV-1, abnormalities of genes involved in TCR-NF-κB signaling, such as CARD11, are detected in about 90% of patients. Utilizing mice expressing CD4+ T cell-specific CARD11(E626K) and/or CD4+ T cell-specific HBZ, namely CARD11(E626K)CD4-Cre mice, HBZ transgenic (Tg) mice, and CARD11(E626K)CD4-Cre;HBZ Tg double transgenic mice, we clarify these genes’ pathogenetic effects. CARD11(E626K)CD4-Cre and HBZ Tg mice exhibit lymphocytic invasion to many organs, including the lungs, and double transgenic mice develop lymphoproliferative disease and increase CD4+ T cells in vivo. CARD11(E626K) and HBZ cooperatively activate the non-canonical NF-κB pathway, IRF4 targets, BATF3/IRF4/HBZ transcriptional network, MYC targets, and E2F targets. Most KEGG and HALLMARK gene sets enriched in acute-type ATL are also enriched in double transgenic mice, indicating that these genes cooperatively contribute to ATL development.

Abstract A059: NF-κB inducing kinase (NIK) deletion accelerates the progression of pancreatic cancer

Abstract Pancreatic ductal adenocarcinoma (PDAC) is the most common type of pancreatic cancer and the leading cause of cancer-related deaths. The canonical NF-κB pathway displays critical functions in the development of PDAC. However, the relationship between the non-canonical NF-κB pathway and PDAC is still obscure. Here, we analyzed the role of the non-canonical NF-κB signaling in precancerous lesions and PDAC development by deleting NF-κB inducing kinase (NIK), an essential kinase for the activation of the non-canonical pathway. We used KC mice (Pdx1-cre/LSL-KrasG12D) that develop pancreatic intraepithelial neoplasias (PanINs) already at the age of 8 weeks. To evaluate the role of the non-canonical NF-κB signaling, KC mice were crossed with NIK-floxed mice (NiC) to generate the KNiC (Pdx1-cre/KRASG12D/NIKfl/fl) mouse model. Mice were injected with cerulein to promote the development of pancreatitis and support the development of PanINs. At the age of 8 weeks, mice were sacrificed and analyzed. Furthermore, to analyze the role of the non-canonical pathway in PDAC development, we crossed -the rapidly PDAC developing- KPC mice (Pdx1-cre/LSL-KrasG12D/p53fl/fl) to NiC mice and generated the KPNiC (Pdx1-cre/KRASG12D/NIKfl/fl/p53fl/fl) mouse model. Mice were analyzed at the age of 8 weeks or when they reached their humane endpoint. Analysis of the pancreata revealed that deletion of NIK supported the development of higher-grade precancerous lesions and an increased remodeling area. Importantly, NIK deletion accelerated the establishment of PDAC already by the age of 8 weeks. In addition, the pancreatic/body weight ratio was significantly higher in the KNiC group compared to the KC group (Mean: KC 0.018 vs KNiC 0.030, P<0.05). These results were further verified by Ki67 staining, where we detected more proliferating cells in KNiC pancreata (Mean: KC 66.53/mm2 vs KNiC 205.60/mm2, P<0.05). Moreover, FN1 expression level (Mean: KC 1.00 vs KNiC 2.14, P<0.05) and the number α-SMA+ cells (Mean: KC 6.73% vs KNiC 17.47%, P<0.05) were dramatically elevated in KNiC pancreata, indicating increased desmoplastic reaction after NIK deletion. Interestingly, KNiC pancreata displayed stronger activation of the STAT3 pathway which is associated with PanIN development and progression (Mean: KC 1.00 vs KNiC 1.73, P<0.05). With respect to the KPC and KPNiC mice, NIK deletion strongly accelerated the development of PDAC at the age of 8 weeks (83.3% of KPNiC mice, 25% of KPC mice). Finally, the median survival of KPNiC mice (71 days) was remarkably shorter than the survival of KPC mice (85 days). Our data demonstrate that deletion of NIK in KC mice promoted the progression of precancerous lesions, a stronger fibrotic reaction, and activation of the STAT3 pathway. Further, deletion of NIK in KPC mice accelerated cancer development and shortened the lifespan of the mice. These results indicate that inactivation of the non-canonical NF-κB pathway leads to PDAC deterioration. Analysis on later time points and further examination of the mechanism is ongoing. Citation Format: Ziwei Du, Miltiadis Tsesmelis, Thomas Wirth. NF-κB inducing kinase (NIK) deletion accelerates the progression of pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer; 2022 Sep 13-16; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2022;82(22 Suppl):Abstract nr A059.

Identification of TSC22D3 As a Novel Immune-Related Prognostic Biomarker of Acute Myeloid Leukemia

Acute myeloid leukemia (AML) is the most common type of hematological neoplasms in adults. TSC22 Domain Family Member 3 (TSC22D3) is one of glucocorticoid-induced genes and has an important regulatory role in immunosuppressive and cell proliferation. However, its prognostic value and immune infiltration effect in acute myeloid leukemia are remain elusive. Here, we investigated the association of TSC22D3 with immune infiltrates and its prognostic value in AML patients. We first analyzed 173 patients from the TCGA and 70 healthy controls from GTEx databases and found that TSC22D3 was highly expressed in AML (Figure A)and closely associated with various biological processes and signaling pathways, including stress-activated MAPK cascade and proteoglycans in cancer and cytokine-cytokine receptor interaction (Figure B). In addition, GESA analysis demonstrated that TSC22D3 was closely corelated with several critical signaling pathways including Toll-like receptor 4 (TLR4) pathway, EGFR pathway, NOTCH pathway, TNFR2 non-canonical NF-κB pathway (Figure C), which further highlighted the important role of TSC22D3 in the pathogenesis of AML. Immunogen analysis found positive associations with various immune cells, especially macrophages (Figure D), and TSC22D3 was also significantly associated with PD-1 and CTLA-4 expression in AML (Figure E). Our results suggest that TSC22D3 is closely associated with immune injectors and promotes immune escape in AML patients in the tumor microenvironment. High level of TSC22D3 was related with poor prognosis of AML in age, BM blasts and several gene mutation (IDH1 R132, FLT3) subgroups (Figure F), indicated that TSC22D3 can be identified as a prognostic molecule in these clinical feature subgroups. Subsequently, a calibration nomogram based on the age and genetic risk level of the clinical patient was conducted. There was a favorable consistency between the actual observed and predicted values for 1-, 3-, 5-year OS based on the calibration chart (Figure G). Based on the complementary perspective for respective tumors, the constructed model provided a personalized score for individual patients. Taken together, this study firstly reported that the high expression of TSC22D3 was significantly associated with the poor survival rate and immune infiltration in AML patients, which may promote the tumorigenesis of AML through enhancing the abnormal inflammation and immune escape. In particular, the immunomodulatory effects of TSC22D3 may play a crucial role in predicting the prognosis and have the potential to become a new biomarker of AML. Thus, this study identified that TSC22D3 may be useful as an immunomodulatory agent for AML clinical management and provided a new insight for further investigation on the clinicopathological significance and molecular pathogenesis of AML. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

TRAF3 Loss-of-Function Reveals the Non-Canonical NF-Κb Pathway As a Therapeutic Target in Diffuse Large B-Cell Lymphoma

TRAF3 Loss-of-Function Reveals the Non-Canonical NF-Κb Pathway As a Therapeutic Target in Diffuse Large B-Cell Lymphoma

Non-canonical NF-κB contributes to endothelial pyroptosis and atherogenesis dependent on IRF-1

Cell inflammation and death are closely linked processes contributing to endothelial dysfunction, which plays a critical role in atherogenesis. Activation of the NLRP3 inflammasome causes pyroptosis, the Gasdermin D (GSDMD)-mediated inflammatory cell death. The non-canonical NF-κB pathway has been implicated in inflammation; however, its role in NLRP3 inflammasome-mediated endothelial dysfunction has not been investigated. This study investigated a role for the non-canonical NF-κB pathway in regulating endothelial pyroptosis as it relates to atherogenesis. Immunohistochemistry indicated inflammasome activation in the endothelial cells (EC) of human atherosclerotic arteries. Flow cytometry and Western blot analysis revealed that oxidized low-density lipoprotein (oxLDL) activated the NLRP3 inflammasome, concomitant with the activation of non-canonical NF-κB in primary human aortic EC. Interference of NF-κB inducing kinase (NIK), the key regulator of the non-canonical pathway, significantly attenuated oxLDL- or LPS/ATP-induced NLRP3 inflammasome activation, pyroptosis, IL-1β, and IL-18 secretion. In contrast, overexpression of NIK exacerbated these responses. Chromatin immunoprecipitation revealed that activation of the non-canonical NF-κB pathway upregulated the transcription factor IRF-1 through RelB/p52 binding to its promoter region at -782/-770. In addition to the known target CASP1, RNA sequencing further identified GSDMD as a target gene of IRF-1. IRF-1 but not RelB/p52 interacted with the GSDMD promoter at -526/-515 and the CASP1 promoter at -11/10 to promote the expression and CASP1-mediated activation of GSDMD. Consistent with the observations in cultured endothelium, endothelial-specific deficiency of NIK or IRF-1 attenuated atherosclerosis in high-fat diet-fed Apoe-null mice. These data demonstrate that the non-canonical NF-κB pathway contributes to NLRP3 inflammasome-mediated endothelial pyroptosis and the development of atherosclerosis through GSDMD activation in a manner dependent on IRF-1. Further investigation may facilitate the identification of specific therapeutic targets for atherosclerotic heart diseases.

Norcantharidin liposome emulsion hybrid delivery system enhances PD-1/PD-L1 immunotherapy by agonizing the non-canonical NF-κB pathway

PD-1/L1 checkpoint blockade has gained approval in terms of treating patients suffering from hepatocellular carcinoma (HCC). It should be noted that the PD-1/L1 inhibitor (α-PD-1/L1) has a low overall response rate when used as a single agent. Accordingly, the combination of α-PD-1/L1 and a series of therapies to further increase the response rate has become a major research direction. In our previous study, we developed a novel norcantharidin (NCTD) liposome emulsion hybrid delivery system (NE) with enhanced anticancer activity and reduced toxicity. In this study, NE was combined with α-PD-1/L1 for treating HCC. The combination therapy exhibited an enhanced antitumor activity, which led to the up-regulated expression levels of white blood cells, interleukin 12 (IL-12), interferon γ (IFN-γ), PD-L1, as well as CD8. Furthermore, the combination of NE and α-PD-1 achieved the optimal efficiency. NCTD-based chemotherapy is capable of synergizing with α-PD-1/L1 while enhancing checkpoint immunotherapy. It follows a mechanism that NCTD agonizes the non-canonical NF-κB pathway of dendritic cells for better activating CD8+T cells. Furthermore, NCTD may enhance antitumor immunity due to the leukogenic effect. In brief, new therapeutic regimens were provided for anti-HCC treatment by integrating NE to PD-1/L1 immunotherapy.

SARS-CoV-2 and HIV: Impact on Pulmonary Epithelial Cells.

The SARS-CoV-2 pandemic provides a natural opportunity for the collision of coronavirus disease-2019 (COVID-19) with chronic infections, which place numerous individuals at high risk of severe COVID-19. Infection with Human Immunodeficiency Virus (HIV), a global epidemic, remains a major public health concern. Whether prior HIV+ status exacerbates COVID-19 warrants investigation. Herein, we characterized the impact of SARS-CoV-2 in human bronchial epithelial cells (HBECs) previously exposed to HIV. We optimized the air-liquid interface (ALI) cell culture technique to allow for challenges with HIV at the basolateral cell surface and SARS-CoV-2 spike protein on the apical surface, followed by genetic analyses for cellular stress/toxicity and innate/adaptive immune responses. Our results suggest that the IL-10 pathway was consistently activated in HBECs treated with spike, HIV, or a combination. Recombinant spike protein elicited COVID-19 cytokine storms while HIV activated different signaling pathways. HIV-treated HBECs could no longer activate NF-kB, pro-inflammatory TRAF-6 ubiquitination nor RIP1 signaling. Combinations of HIV and SARS-CoV-2 spike increased gene expression for activation of endoplasmic reticulum-phagosome pathway and downregulated non-canonical NF-kB pathways that are key in functional regulatory T cells and RNA Polymerase II transcription. Our in vitro studies suggest that prior HIV infection may not exacerbate COVID-19. Further in vivo studies are warranted to advance this field.

Mechanisms underlying synergism between circularized tumor necrosis factor-related apoptosis inducing ligand and bortezomib in bortezomib-sensitive or -resistant myeloma cells.

Mechanisms underlying interactions between a novel, clinically relevant circularized tumor necrosis factor-related apoptosis inducing ligand (TRAIL) agonist, circularly permuted TRAIL (CPT) have been examined in multiple myeloma (MM) cells sensitive or resistant to bortezomib (BTZ). Various MM cell lines for example, U266, including those resistant to bortezomib-resistant U266 cells were exposed to low nanomolar concentrations of bortezomib±CPT and apoptosis monitored. Circularly permuted TRAIL and bortezomib synergistically induced apoptosis in both BTZ-naïve and -resistant cells. The regimen up-regulated DR4 receptor internalization in MM cells, known to modulate both NF-κB and extrinsic apoptotic pathways. CPT/BTZ disrupted the non-canonical NF-κB pathway, reflected by tumor necrosis factor (TNF) receptor associated factors 3 (TRAF3) up-regulation, NF-κB inducing kinase down-regulation, diminished p52 and p50 processing, and B-cell lymphoma-extra large (BCL-XL) down-regulation, but failed to inactivate the canonical NF-κB pathway, reflected by unchanged or increased expression of phospho-p65. The regimen also sharply increased extrinsic apoptotic pathway activation. Cells exhibiting TRAF3 knock-down, dominant-negative Fas-associated protein with death domain, knock-down of caspase-8, BCL-2/BCL-XL, or exposure to a caspase-9 inhibitor displayed markedly reduced CPT/BTZ sensitivity. Concordant results were observed in bortezomib-resistant cells. The regimen was also active in the presence of stromal cells and was relatively sparing toward normal CD34+ hematopoietic cells. Finally, ex vivo results revealed synergism in primary MM primary cells, including those BTZ, and the CPT/BTZ regimen significantly decreased tumor growth in a patient-derived MM xenograft model. These results indicate that the CPT/BTZ regimen acts via the non-canonical NF-κB as well as intrinsic/extrinsic apoptotic pathways to induce cell death in MM cells, and may represent an effective strategy in the setting of bortezomib resistance.

RANKL elevation activates the NIK/NF-κB pathway, inducing obesity in ovariectomized mice.

Menopausal women are susceptible to visceral obesity, which increases the risk of metabolic disorders. However, the mechanisms of menopause-induced visceral fat accumulation are not fully understood. Circulating levels of receptor activator of nuclear factor-kappa B (NF-κB) ligand (RANKL) are elevated in an animal model of menopause. RANKL, a multifunctional cytokine, activates the NF-κB pathway, which serves as a pivotal mediator of inflammatory responses. Here, we investigated whether RANKL-induced non-canonical NF-κB pathway activation induces inflammation and lipid accumulation in adipose tissues. RANKL induced Tnfa expression via the non-canonical NF-κB pathway in bone marrow cells. We therefore analyzed aly/aly mice, in which the non-canonical NF-κB pathway is not activated, owing to an inactive form of NF-κB-inducing kinase. A postmenopausal obesity model was generated by ovariectomy and subsequent high-fat and high-sucrose diet feeding. In aly/aly mice with postmenopausal obesity, serum RANKL levels were elevated, and hepatic lipid accumulation and adipocyte hypertrophy were suppressed, resulting in reduced macrophage infiltration and inflammatory cytokine mRNA expression in visceral adipose tissue. Furthermore, aly/aly mice showed protection from glucose intolerance and insulin resistance, which were observed in ovariectomized WT obese mice. These findings indicate that non-canonical NF-κB pathway activation via serum RANKL elevation contributes to postmenopausal obesity.

NF-κB Inducing Kinase Regulates Intestinal Immunity and Homeostasis.

Intestinal immunity and homeostasis are maintained through the regulation of cytokine trafficking, microbiota, necrosis and apoptosis. Intestinal immunity and homeostasis participate in host defenses and inflammatory responses locally or systemically through the gut-organ axis. NF-κB functions as a crucial transcription factor mediating the expression of proteins related to the immune responses. The activation of NF-κB involves two major pathways: canonical and non-canonical. The canonical pathway has been extensively studied and reviewed. Here, we present the current knowledge of NIK, a pivotal mediator of the non-canonical NF-κB pathway and its role in intestinal immunity and homeostasis. This review also discusses the novel role of NIK signaling in the pathogenesis and treatment of inflammatory bowel disease.

CGAS-STING drives the IL-6-dependent survival of chromosomally instable cancers.

Chromosomal instability (CIN) drives cancer cell evolution, metastasis and therapy resistance, and is associated with poor prognosis1. CIN leads to micronuclei that release DNA into the cytoplasm after rupture, which triggers activation of inflammatory signalling mediated by cGAS and STING2,3. These two proteins are considered to be tumour suppressors as they promote apoptosis and immunosurveillance. However, cGAS and STING are rarely inactivated in cancer4, and, although they have been implicated in metastasis5, it is not known why loss-of-function mutations do not arise in primary tumours4. Here we show that inactivation of cGAS-STING signalling selectively impairs the survival of triple-negative breast cancer cells that display CIN. CIN triggers IL-6-STAT3-mediated signalling, which depends on the cGAS-STING pathway and the non-canonical NF-κB pathway. Blockade of IL-6 signalling by tocilizumab, a clinically used drug that targets the IL-6 receptor (IL-6R), selectively impairs the growth of cultured triple-negative breast cancer cells that exhibit CIN. Moreover, outgrowth of chromosomally instable tumours is significantly delayed compared with tumours that do not display CIN. Notably, this targetable vulnerability is conserved across cancer types that express high levels of IL-6 and/or IL-6R in vitro and in vivo. Together, our work demonstrates pro-tumorigenic traits of cGAS-STING signalling and explains why the cGAS-STING pathway is rarely inactivated in primary tumours. Repurposing tocilizumab could be a strategy to treat cancers with CIN that overexpress IL-6R.

Changes Within H3K4me3-Marked Histone Reveal Molecular Background of Neutrophil Functional Plasticity.

Neutrophils are a heterogenous population capable of both antimicrobial functions and suppressor ones, however, no specific pattern of transcription factors controlling this plasticity has been identified. We observed rapid changes in the neutrophil status after stimulation with LPS, pre-activating concentration of TNF-α, or IL-10. Chromatin immunoprecipitation sequencing (ChIP-Seq) analysis of histone H3K4me3 allowed us to identify various transcriptional start sites (TSSs) associated with plasticity and heterogeneity of human neutrophils. Gene Ontology analysis demonstrated great variation within target genes responsible for neutrophil activation, cytokine production, apoptosis, histone remodelling as well as NF-κB transcription factor pathways. These data allowed us to assign specific target genes positioned by H3K4me3-marked histone with a different pattern of gene expression related to NF-κB pathways, apoptosis, and a specific profile of cytokines/chemokines/growth factors realised by neutrophils stimulated by LPS, IL-10, or TNF-α. We discovered IL-10-induced apoptotic neutrophils being transcriptionally active cells capable of switching the profile of cytokines/chemokines/growth factors desired in resolving inflammation via non-canonical NF-κB pathway with simultaneous inhibition of canonical NF-κB pathway. As apoptotic/suppressive neutrophils induced by IL-10 via positioning genes within H3K4me3-marked histone were transcriptionally active, newly described DNA binding sites can be considered as potential targets for immunotherapy.Graphical H3K4me3 histone ChIP-Seq analysis reveals molecular drivers critical for switching neutrophils from their pro- to anti-inflammatory properties.

Role of IKKε in the Metabolic Diseases: Physiology, Pathophysiology, and Pharmacology.

IKKε (inhibitor of nuclear factor kappa-B kinase ε) is a member of the noncanonical NF-κB pathway. It participates in the inflammatory response and innate immunity against bacteria. In recent decades, IKKε has been closely associated with metabolic regulation. Inhibition of the IKKε pathway can improve fat deposition in the liver, reduce subcutaneous fat inflammation, and improve liver gluconeogenesis in obesity. IKKε is expected to be a new therapeutic target for metabolic diseases such as nonalcoholic fatty liver disease, diabetes, and obesity. Herein, we summarize the structural characterization, physiological function, and pathological role of IKKε in metabolic diseases and small molecule inhibitors of IKKε.

Comparison of the Safety and Efficacy of Tofacitinib and Fingolimod in TNBS‐Induced Colitis Model in Adult Zebrafish: The Role of Myd88/NF‐κB/TNF‐α Signaling Pathway

Due to the inadequate response, side effects, parenteral route of administration, and immunogenicity, oral targeted small molecules seem to replace monoclonal antibodies in the future management of ulcerative colitis (UC). In order to pave the way for clinical translations of small molecules in UC, for the first time, we compared the efficacy of fingolimod and tofacitinib in preclinical model of colitis, and uncovered another mechanism of action that these medicines may act through.We implicated adult zebrafish to find out its pros/cons, investigated aquatic toxicity of the mentioned compounds, and designed a procedure to evaluate compound's efficacy in adult zebrafish colitis model; hence, facilitating screening of further agents.Tofacitinib and fingolimod were investigated to be in category 1 of Globally Harmonized System (GHS) aquatic toxicity classification with the LC50 of 0.9014 and 0.36 mg/L, respectively, based on the acute toxicity assessment within 96 h.Considering efficacy assessments, colitis was induced through intrarectal administration of 160 mM TNBS for 48 h. Then, animals were exposed to different concentrations of tofacitinib (0.25, 0.57, 0.72, 0.90 mg/L) and fingolimod (0.23, 0.28, 0.36, 0.45 mg/L) for 96 h. Subsequent analyses revealed that TNBS significantly reduced length of villi, number of goblet cells, level of TNF‐α, MyD88, and NF‐κB2, increased thickness of villi and necrosis, and induced histopathological changes, including intestinal edema, submucosal hemorrhage/hyperemia with the presence of inflammatory cells. All the aforementioned parameters were considerably enhanced almost dose‐dependently with both agents, wherein the effect of 0.45 mg/L fingolimod was superior to other groups. As well, there were no signs of hyperemia/hemorrhage in harvested intestinal samples. Resultantly, we revealed that these medications may suppress canonical and non‐canonical NF‐κB pathways through targeting toll‐like receptors and MyD88.Given the better efficacy and multi‐targeted action of fingolimod, it worth to focus on sphingosine‐1 phosphate receptor modulators in clinical trials of UC and compare them with other classes of medications; albeit, their ecotoxicity aspect should not be overlooked.