TWEAK/Fn14 signalling driven super-enhancer reprogramming promotes pro-metastatic metabolic rewiring in triple-negative breast cancer.

Triple Negative Breast Cancer (TNBC) is the most aggressive breast cancer subtype suffering from limited targeted treatment options. Following recent reports correlating Fibroblast growth factor-inducible 14 (Fn14) receptor overexpression in Estrogen Receptor (ER)-negative breast cancers with metastatic events, we show that Fn14 is specifically overexpressed in TNBC patients and associated with poor survival. We demonstrate that constitutive Fn14 signalling rewires the transcriptomic and epigenomic landscape of TNBC, leading to enhanced tumour growth and metastasis. We further illustrate that such mechanisms activate TNBC-specific super enhancers (SE) to drive the transcriptional activation of cancer dependency genes via chromatin looping. In particular, we uncover the SE-driven upregulation of Nicotinamide phosphoribosyltransferase (NAMPT), which promotes NAD+ and ATP metabolic reprogramming critical for filopodia formation and metastasis. Collectively, our study details the complex mechanistic link between TWEAK/Fn14 signalling and TNBC metastasis, which reveals several vulnerabilities which could be pursued for the targeted treatment of TNBC patients.

[Relationships between Cxcl12, Tweak, Notch1, and Yap mRNA Expression Levels in Molecular Mechanisms of Liver Fibrogenesis].

Current data on the molecular mechanisms of liver fibrosis and cirrhosis fail to fully explain all stages of their development. Interactions between individual genes and signaling pathways are known to play an important role in their functions. However, data on their relationships are insufficient and often contradictory. For the first time, mRNA expression of Notch1, Notch2, Yap1, Tweak (Tnfsf12), Fn14 (Tnfrsf12a), Ang, Vegfa, Cxcl12 (Sdf), Nos2, and Mmp-9 was studied in detail at several stages of thioacetamide-induced liver fibrosis in Wistar rats. A factor analysis isolated three factors, which combined highly correlated target genes. The first factor included four genes: Cxcl12 (r = 0.829, p < 0.05), Tweak (r = 0.841, p < 0.05), Notch1 (r = 0.848, p < 0.05), and Yap1 (r = 0.921, p < 0.05). The second factor described the correlation between Mmp-9 (r = 0.791, p < 0.05) and Notch2 (r = 0.836, p < 0.05). The third factor included Ang (r = 0.748, p < 0.05) and Vegfa (r = 0.679, p < 0.05). The Nos2 and Fn14 genes were not included in any of the factors. The gene grouping by mRNA expression levels made it possible to assume a pathogenetic relationship between their products in the development of fibrotic changes due to liver toxicity.

Genetically predicted TWEAK mediates the association between lipidome and Keratinocyte Carcinomas.

BACKGROUND: Reports suggest that lipid profiles may be linked to the likelihood of developing skin cancer, yet the exact causal relationship is still unknown. OBJECTIVE: This study aimed to examine the connection between lipidome and skin cancers, as well as investigate any possible mediators. METHODS: A two-sample Mendelian randomization (MR) analysis was conducted on 179 lipidomes and each skin cancer based on a genome-wide association study (GWAS), including melanoma, basal cell carcinoma (BCC), and squamous cell carcinoma (SCC). Then, Bayesian weighted MR was performed to verify the analysis results of two-sample MR. Moreover, a two-step MR was employed to investigate the impact of TNF-like weak inducer of apoptosis (TWEAK)-mediated lipidome on skin cancer rates. RESULTS: MR analysis identified higher genetically predicted phosphatidylcholine (PC) (17:0_18:2) could reduce the risk of skin tumors, including BCC (OR = 0.9149, 95% CI: 0.8667-0.9658), SCC (OR = 0.9343, 95% CI: 0.9087-0.9606) and melanoma (OR = 0.9982, 95% CI: 0.9966-0.9997). The proportion of PC (17:0_18:2) predicted by TWEAK-mediated genetic prediction was 6.6 % in BCC and 7.6% in SCC. The causal relationship between PC (17:0_18:2) and melanoma was not mediated by TWEAK. CONCLUSION: This study identified a negative causal relationship between PC (17:0_18:2) and keratinocyte carcinomas, a small part of which was mediated by TWEAK, and most of the remaining mediating factors are still unclear. Further research on other risk factors is needed in the future.

Proteome-Wide Mendelian Randomization Analysis Identified Potential Drug Targets for Atrial Fibrillation.

Background Finding effective and safe therapeutic drugs for atrial fibrillation (AF) is an important concern for clinicians. Proteome-wide Mendelian randomization analysis provides new ideas for finding potential drug targets. Methods and Results Using a proteome-wide Mendelian randomization approach, we assessed the genetic predictive causality between thousands of proteins and AF risk and found that genetically predicted plasma levels of phosphomevalonate kinase, tumor necrosis factor ligand superfamily member 12, sulfhydryl oxidase 2, interleukin-6 receptor subunit alpha, and low-affinity immunoglobulin gamma Fc region receptor II-b might decrease AF risk, while genetically predicted plasma levels of beta-mannosidase, collagen alpha-1(XV) chain, ANXA4 (annexin A4), COF2 (cofilin-2), and RAB1A (Ras-related protein Rab-1A) might increase AF risk (P<3.4×10-5). By using different Mendelian randomization methods and instrumental variable selection thresholds, we performed sensitivity analyses in 30 scenarios to test the robustness of positive findings. Replication analyses were also performed in independent samples to further avoid false-positive findings. Drugs targeting tumor necrosis factor ligand superfamily member 12, interleukin-6 receptor subunit alpha, low-affinity immunoglobulin gamma Fc region receptor II-b, and annexin A4 are approved or in development. The results of the phenome-wide Mendelian randomization analysis showed that changing the plasma levels of phosphomevalonate kinase, cofilin-2, annexin A4, Ras-related protein Rab-1A, sulfhydryl oxidase 2, and collagen alpha-1(XV) chain did not increase the risk of other diseases while decreasing the risk of AF. Conclusions We found a significant causal association between genetically predicted levels of 10 plasma proteins and AF risk. Four of these proteins have drugs targeting them that are approved or in development, and our results suggest the potential for these drugs to treat AF or cause AF. Sulfhydryl oxidase 2, low-affinity immunoglobulin gamma Fc region receptor II-b, and beta-mannosidase have not been suggested by previous laboratory or epidemiological studies to be associated with AF and may reveal new pathophysiological pathways as well as therapeutic targets for AF.

Relief of Extracellular Matrix Deposition Repression by Downregulation of IRF1-Mediated TWEAK/Fn14 Signaling in Keloids.

Keloids represent a fibrotic disorder characterized by the excessive deposition of extracellular matrix (ECM). However, the mechanisms through which ECM deposition in keloids is regulated remain elusive. In this study, we found that the expression of both TWEAK and its cognate receptor Fn14 was significantly downregulated in keloids and that TWEAK/Fn14 signaling repressed the expression of ECM-related genes in keloid fibroblasts. The IRF1 gene was essential for this repression, and the TWEAK/Fn14 downstream transcription factor p65 directly bound to the promoter of the IRF1 gene and induced its expression. Furthermore, in patients with keloid, the expression of TWEAK and Fn14 was negatively correlated with that of ECM genes and positively correlated with that of IRF1. These observations indicate that relief of TWEAK/Fn14/IRF1-mediated ECM deposition repression contributes to keloid pathogenesis, and the identified mechanism and related molecules provide potential targets for keloid treatment in the future.

Status of TWEAK DNA methylation and mRNA expression in systemic lupus erythematosus.

OBJECTIVE: Draw upon research into the serum concentration, mRNA expression, and DNA methylation of TNF-like weak inducer of apoptosis (TWEAK) in the peripheral blood of systemic lupus erythematosus patients and healthy controls in an attempt to investigate the epigenetics associated with TWEAK in the pathogenesis of systemic lupus erythematosus (SLE). METHODS: A total of 178 SLE patients (SLE group) and 131 sex-age matched healthy controls (HC group) were recruited. Enzyme-linked immunosorbent assays (ELISA) was used to detect serum protein concentration of TWEAK. TWEAK mRNA expression was analyzed by Real-time quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). Methylation levels of the promotor of TWEAK were measured using quantitative DNA methylation analysis on the MassARRAY spectrometry. RESULTS: Serum TWEAK concentrations were not statistically significant in SLE patients and HCs. Nevertheless, serum TWEAK concentrations were significantly lower in patients with renal involvement when compared to those without it. Serum TWEAK concentrations were reduced in clinically active patients (SLEDAI ≥ 10) compared with clinically stable patients (SLEDAI < 10). It was also significantly associated with SLEDAI. Compared with the HC group, the TWEAK mRNA expression in the SLE group was significantly lower. The global DNA methylation levels of TWEAK in the SLE group were observed to be significantly higher than the HC group. SLE patients with renal involvement, and the clinically active patients had higher TWEAK global methylation as well as exhibited variation in certain CpG island methylation. Furthermore, TWEAK methylation negatively correlated with TWEAK mRNA expression. CONCLUSION: This study suggests that TWEAK DNA methylation is a valuable as a focus for epigenetic studies because of it potentially influencing TWEAK gene expression in SLE patients. Aberrant DNA methylation of TWEAK may be involved in the initiation and development of SLE.

Lower Expression of TWEAK is Associated with Poor Survival and Dysregulate TIICs in Lung Adenocarcinoma.

Background: Lung cancer remains the leading cause of cancer death worldwide, and the most subtype is lung adenocarcinoma (LUAD). Tumor-infiltrating immune cells (TIICs) greatly impact the prognosis of LUAD. Tumor necrosis factor-like weak inducer of apoptosis (TWEAK), signal via its receptor fibroblast growth factor-inducible 14 (Fn14), dysregulates immune cell recruitment within tumor environment, thus promoting the progression of autoimmune diseases and cancer. We aimed to explore its role in LUAD. Methods: The expression level of TWEAK was explored in Tumor Immune Estimation Resource 2.0 (TIMER2.0) and Oncomine databases. The Tumor Immune Dysfunction and Exclusion (TIDE) and Lung Cancer Explorer (LCE) databases were applied to evaluate the survival in correlation to TWEAK expression. TIICs were assessed with TIMER2.0 and TIDE datasets. The expression of TWEAK protein was detected in LUAD cell lines and also in tissue samples from LUAD patients via western blotting or combination with immunochemistry. Results: Our results showed that TWEAK was downregulated in LUAD tumors compared to normal tissues in TIMER2.0, Oncomine, cell lines, and clinical specimens. Poor survival was uncovered in lower TWEAK expression of LUAD patients in LCE (meta - HR = 0.84 [95% CI, 0.76-0.92]) and TCGA (Continuous Z = -1.97, p = 0.0486) and GSE13213@PRECOG (Continuous Z = -4.25, p = 2.12e - 5) in TIDE. Multiple tumor-infiltrating immune cells (TIICs) were found closely correlated with TWEAK expression in LUAD, especially hematopoietic stem cell (Rho = 0.505, p = 2.78e - 33), common lymphoid progenitor (Rho = -0.504, p = 3.79e - 33), and myeloid-derived suppressor cells (MDSCs) (Rho = -0.615, p = 1.36e - 52). Conclusion: Lower level of TWEAK was linked with poor survival and aberrant recruitment and phenotype of TIICs in LUAD, which might motivate immune escape and weaken the effects of immunotherapy.

[Methylation status and expression of TWEAK gene promoter region in peripheral blood of patients with rheumatoid arthritis].

OBJECTIVE: To explore the relationship between tumor necrosis factor like weak inducer of apoptosis (TWEAK) gene and the pathogenesis of rheumatoid arthritis (RA) by detecting the DNA methylation level, mRNA expression level and serum protein concentration of TWEAK gene in peripheral blood. METHODS: The MassARRAY method was used to detect the DNA methylation level of the TWEAK gene in the peripheral blood of 112 RA patients and 86 matched healthy volunteers. The real-time quantitative polymerase chain reaction method was used to detect the mRNA expression level of the TWEAK gene in the peripheral blood of the subjects. The enzyme-linked immunosorbent assay method was used to detect the serum TWEAK protein concentration of the subjects. The TWEAK gene DNA methylation level, mRNA expression level and serum protein concentration between the RA group and the healthy control group were compared, and the relationship between it and the degree of disease activity analyzed. RESULTS: The overall DNA methylation level of TWEAK gene and the DNA methylation levels of CpG_11, CpG_17.18.19.20, CpG_40.41.42 site in the RA group were higher than those in the healthy control group (P=0.002, P=0.01, P=0.006, P=0.002, respectively). The DNA methylation level of CpG_55.56 site in the high disease activity group was higher than that in the medium and low disease activity group (P=0.041). The expression level of TWEAK gene mRNA in the peripheral blood of the RA group was lower than that of the healthy control group (P=0.023). The expression level of TWEAK gene mRNA in the high disease activity group was lower than that in the medium and low disease activity group (P=0.035). The serum TWEAK protein concentration of the RA group was not significantly different from that of the healthy control group (P=0.508), but it was positively correlated with the mRNA expression level (r=0.482, P < 0.001). CONCLUSION: The TWEAK gene is closely related to the onset and progression of RA, and its hypermethylation state may be one of the epigenetic mechanisms regulating its low mRNA expression, and it can be used as one of the important indicators for clinical monitoring and evaluation of RA.

Road to Metastasis: The TWEAK Pathway as a Discriminant between Metastasizing and Non-Metastasizing Thick Melanomas.

Cutaneous melanoma (CM) is the most aggressive form of skin cancer, and its worldwide incidence is rapidly increasing. Early stages can be successfully treated by surgery, but once metastasis has occurred, the prognosis is poor. However, some 5-10% of thick (≥2 mm) melanomas do not follow this scenario and run an unpredictable course. Little is known about the factors that contribute to metastasis in some patient with thick melanomas and the lack thereof in thick melanoma patients who never develop metastatic disease. We were therefore interested to study differential gene expression and pathway analysis and compare non-metastatic and metastatic thick melanomas. We found that the TNF-like weak inducer of apoptosis (TWEAK) pathway was upregulated in thick non-metastasizing melanomas. MAP3K14 (NIK1), BIRC2 (cIAP1), RIPK1, CASP7, CASP8, and TNF play an important role in inhibiting proliferation and invasion of tumor cells via the activation of the non-canonical NF-κB signaling pathway. In particular, this pathway sensitizes melanoma cells to TNF-alpha and activates the apoptosis module of the TWEAK pathway in thick non-metastasizing melanomas. Hence, our study suggests a potential role of the TWEAK pathway in inhibiting thick melanoma from metastasis. Exploitation of these genes and the pathway they control may open future therapeutic avenues.

TWEAKing the Hippocampus: The Effects of TWEAK on the Genomic Fabric of the Hippocampus in a Neuropsychiatric Lupus Mouse Model.

Neuropsychiatric manifestations of systemic lupus erythematosus (SLE), specifically cognitive dysfunction and mood disorders, are widely prevalent in SLE patients, and yet poorly understood. TNF-like weak inducer of apoptosis (TWEAK) has previously been implicated in the pathogenesis of neuropsychiatric lupus (NPSLE), and we have recently shown its effects on the transcriptome of the cortex of the lupus-prone mice model MRL/lpr. As the hippocampus is thought to be an important focus of NPSLE processes, we explored the TWEAK-induced transcriptional changes that occur in the hippocampus, and isolated several genes (Dnajc28, Syne2, transthyretin) and pathways (PI3K-AKT, as well as chemokine-signaling and neurotransmission pathways) that are most differentially affected by TWEAK activation. While the functional roles of these genes and pathways within NPSLE need to be further investigated, an interesting link between neuroinflammation and neurodegeneration appears to emerge, which may prove to be a promising novel direction in NPSLE research.

Changes in Cerebrospinal Fluid Balance of TNF and TNF Receptors in Naïve Multiple Sclerosis Patients: Early Involvement in Compartmentalised Intrathecal Inflammation.

An imbalance of TNF signalling in the inflammatory milieu generated by meningeal immune cell infiltrates in the subarachnoid space in multiple sclerosis (MS), and its animal model may lead to increased cortical pathology. In order to explore whether this feature may be present from the early stages of MS and may be associated with the clinical outcome, the protein levels of TNF, sTNF-R1 and sTNF-R2 were assayed in CSF collected from 122 treatment-naïve MS patients and 36 subjects with other neurological conditions at diagnosis. Potential correlations with other CSF cytokines/chemokines and with clinical and imaging parameters at diagnosis (T0) and after 2 years of follow-up (T24) were evaluated. Significantly increased levels of TNF (fold change: 7.739; p < 0.001), sTNF-R1 (fold change: 1.693; p < 0.001) and sTNF-R2 (fold change: 2.189; p < 0.001) were detected in CSF of MS patients compared to the control group at T0. Increased TNF levels in CSF were significantly (p < 0.01) associated with increased EDSS change (r = 0.43), relapses (r = 0.48) and the appearance of white matter lesions (r = 0.49). CSF levels of TNFR1 were associated with cortical lesion volume (r = 0.41) at T0, as well as with new cortical lesions (r = 0.56), whilst no correlation could be found between TNFR2 levels in CSF and clinical or MRI features. Combined correlation and pathway analysis (ingenuity) of the CSF protein pattern associated with TNF expression (encompassing elevated levels of BAFF, IFN-γ, IL-1ß, IL-10, IL-8, IL-16, CCL21, haptoglobin and fibrinogen) showed a particular relationship to the interaction between innate and adaptive immune response. The CSF sTNF-R1-associated pattern (encompassing high levels of CXCL13, TWEAK, LIGHT, IL-35, osteopontin, pentraxin-3, sCD163 and chitinase-3-L1) was mainly related to altered T cell and B cell signalling. Finally, the CSF TNFR2-associated pattern (encompassing high CSF levels of IFN-ß, IFN-λ2, sIL-6Rα) was linked to Th cell differentiation and regulatory cytokine signalling. In conclusion, dysregulation of TNF and TNF-R1/2 pathways associates with specific clinical/MRI profiles and can be identified at a very early stage in MS patients, at the time of diagnosis, contributing to the prediction of the disease outcome.

TNF-α induces endothelial-mesenchymal transition promoting stromal development of pancreatic adenocarcinoma.

Endothelial-mesenchymal transition (EndMT) is an important source of cancer-associated fibroblasts (CAFs), which facilitates tumour progression. PDAC is characterised by abundant CAFs and tumour necrosis factor-α (TNF-α). Here, we show that TNF-α strongly induces human endothelial cells to undergo EndMT. Interestingly, TNF-α strongly downregulates the expression of the endothelial receptor TIE1, and reciprocally TIE1 overexpression partially prevents TNF-α-induced EndMT, suggesting that TNF-α acts, at least partially, through TIE1 regulation in this process. We also show that TNF-α-induced EndMT is reversible. Furthermore, TNF-α treatment of orthotopic mice resulted in an important increase in the stroma, including CAFs. Finally, secretome analysis identified TNFSF12, as a regulator that is also present in PDAC patients. With the aim of restoring normal angiogenesis and better access to drugs, our results support the development of therapies targeting CAFs or inducing the EndMT reversion process in PDAC.

TWEAK Signaling Pathway Blockade Slows Cyst Growth and Disease Progression in Autosomal Dominant Polycystic Kidney Disease.

BACKGROUND: In autosomal dominant polycystic kidney disease (ADPKD), cyst development and enlargement lead to ESKD. Macrophage recruitment and interstitial inflammation promote cyst growth. TWEAK is a TNF superfamily (TNFSF) cytokine that regulates inflammatory responses, cell proliferation, and cell death, and its receptor Fn14 (TNFRSF12a) is expressed in macrophage and nephron epithelia. METHODS: To evaluate the role of the TWEAK signaling pathway in cystic disease, we evaluated Fn14 expression in human and in an orthologous murine model of ADPKD. We also explored the cystic response to TWEAK signaling pathway activation and inhibition by peritoneal injection. RESULTS: Meta-analysis of published animal-model data of cystic disease reveals mRNA upregulation of several components of the TWEAK signaling pathway. We also observed that TWEAK and Fn14 were overexpressed in mouse ADPKD kidney cysts, and TWEAK was significantly high in urine and cystic fluid from patients with ADPKD. TWEAK administration induced cystogenesis and increased cystic growth, worsening the phenotype in a murine ADPKD model. Anti-TWEAK antibodies significantly slowed the progression of ADPKD, preserved renal function, and improved survival. Furthermore, the anti-TWEAK cystogenesis reduction is related to decreased cell proliferation-related MAPK signaling, decreased NF-κB pathway activation, a slight reduction of fibrosis and apoptosis, and an indirect decrease in macrophage recruitment. CONCLUSIONS: This study identifies the TWEAK signaling pathway as a new disease mechanism involved in cystogenesis and cystic growth and may lead to a new therapeutic approach in ADPKD.

Inhibition of fibroblast growth factor-inducible 14 attenuates experimental tubulointerstitial fibrosis and profibrotic factor expression of proximal tubular epithelial cells.

BACKGROUND AND AIM: As a proinflammatory cytokine, tumor necrosis factor-like weak inducer of apoptosis (TWEAK) participates in the progression of renal fibrosis by binding to its receptor, fibroblast growth factor-inducible 14 (Fn14). However, the effect of Fn14 inhibition on tubular epithelial cell-mediated tubulointerstitial fibrosis remains unclear. This study aimed to elucidate the role of TWEAK/Fn14 interaction in the development of experimental tubulointerstitial fibrosis as well as the protective effect of Fn14 knockdown on proximal tubular epithelial cells. METHODS: A murine model of unilateral ureteral obstruction was constructed in both wild-type and Fn14-deficient BALB/c mice, followed by observation of the tubulointerstitial pathologies. RESULTS: Fn14 deficiency ameliorated the pathological changes, including inflammatory cell infiltration and cell proliferation, accompanied by reduced production of profibrotic factors and extracellular matrix deposition. In vitro experiments showed that TWEAK dose-dependently enhanced the expression of collagen I, fibronectin, and α-smooth muscle actin in proximal tubular epithelial cells. Interestingly, TWEAK also upregulated the expression levels of Notch1/Jagged1. Fn14 knockdown and Notch1/Jagged1 inhibition also mitigated the effect of TWEAK on these cells. CONCLUSIONS: In conclusion, TWEAK/Fn14 signals contributed to tubulointerstitial fibrosis by acting on proximal tubular epithelial cells. Fn14 inhibition might be a therapeutic strategy for protecting against renal interstitial fibrosis.

Remodeling of Neurotransmission, Chemokine, and PI3K-AKT Signaling Genomic Fabrics in Neuropsychiatric Systemic Lupus Erythematosus.

Cognitive dysfunction and mood changes are prevalent and especially taxing issues for patients with systemic lupus erythematosus (SLE). Tumor necrosis factor (TNF)-like weak inducer of apoptosis (TWEAK) and its cognate receptor Fn14 have been shown to play an important role in neurocognitive dysfunction in murine lupus. We profiled and compared gene expression in the cortices of MRL/+, MRL/lpr (that manifest lupus-like phenotype) and MRL/lpr-Fn14 knockout (Fn14ko) adult female mice to determine the transcriptomic impact of TWEAK/Fn14 on cortical gene expression in lupus. We found that the TWEAK/Fn14 pathway strongly affects the expression level, variability and coordination of the genomic fabrics responsible for neurotransmission and chemokine signaling. Dysregulation of the Phosphoinositide 3-kinase (PI3K)-AKT pathway in the MRL/lpr lupus strain compared with the MRL/+ control and Fn14ko mice was particularly prominent and, therefore, promising as a potential therapeutic target, although the complexity of the transcriptomic fabric highlights important considerations in in vivo experimental models.

Evaluation of Computationally Designed Peptides against TWEAK, a Cytokine of the Tumour Necrosis Factor Ligand Family.

The tumour necrosis factor-like weak inducer of apoptosis (TWEAK) is a member of the tumour necrosis factor ligand family and has been shown to be overexpressed in tumoral cells together with the fibroblast growth factor-inducible 14 (Fn14) receptor. TWEAK-Fn14 interaction triggers a set of intracellular pathways responsible for tumour cell invasion and migration, as well as proliferation and angiogenesis. Hence, modulation of the TWEAK-Fn14 interaction is an important therapeutic goal. The targeting of protein-protein interactions by external agents, e.g., drugs, remains a substantial challenge. Given their intrinsic features, as well as recent advances that improve their pharmacological profiles, peptides have arisen as promising agents in this regard. Here, we report, by in silico structural design validated by cell-based and in vitro assays, the discovery of four peptides able to target TWEAK. Our results show that, when added to TWEAK-dependent cellular cultures, peptides cause a down-regulation of genes that are part of TWEAK-Fn14 signalling pathway. The direct, physical interaction between the peptides and TWEAK was further elucidated in an in vitro assay which confirmed that the bioactivity shown in cell-based assays was due to the targeting of TWEAK. The results presented here are framed within early pre-clinical drug development and therefore these peptide hits represent a starting point for the development of novel therapeutic agents. Our approach exemplifies the powerful combination of in silico and experimental efforts to quickly identify peptides with desirable traits.

Tumor Necrosis Factor-Like Weak Inducer of Apoptosis (TWEAK) Enhances Activation of STAT3/NLRC4 Inflammasome Signaling Axis through PKCδ in Astrocytes: Implications for Parkinson's Disease.

Astrocytic dysfunction has been implicated in Parkinson's disease (PD) pathogenesis. While the Tumor necrosis factor-like weak inducer of apoptosis (TWEAK)/Fn14 signaling axis is known to play a role in PD-like neuropathology, the molecular mechanisms that govern this process remain poorly understood. Herein, we show that TWEAK levels are elevated in PD serum compared to controls. Moreover, using both U373 human astrocyte cells and primary mouse astrocytes, we demonstrate that TWEAK induces mitochondrial oxidative stress as well as protein kinase C delta (PKCδ) and signal transducer and activator of transcription 3 (STAT3) activation, accompanied by NLRC4 inflammasome activation and upregulation and release of proinflammatory cytokines, including IL-1ß, TNF-α, and IL-18. Mechanistically, TWEAK-induced PKCδ activation enhances the STAT3/NLRC4 signaling pathway and other proinflammatory mediators through a mitochondrial oxidative stress-dependent mechanism. We further show that PKCδ knockdown and mito-apocynin, a mitochondrial antioxidant, suppress TWEAK-induced proinflammatory NLRC4/STAT3 signaling and cellular oxidative stress response. Notably, we validated our in vitro findings in an MPTP mouse model of PD and in mice receiving intrastriatal administration of TWEAK. These results indicate that TWEAK is a key regulator of astroglial reactivity and illustrate a novel mechanism by which mitochondrial oxidative stress may influence dopaminergic neuronal survival in PD.

Tumor Necrosis Factor-Like Weak Inducer of Apoptosis (TWEAK)/Fibroblast Growth Factor-Inducible 14 (Fn14) Axis in Cardiovascular Diseases: Progress and Challenges.

Cardiovascular diseases (CVD) are the leading cause of mortality in Western countries. CVD include several pathologies, such as coronary artery disease, stroke, peripheral artery disease, and aortic aneurysm, among others. All of them are characterized by a pathological vascular remodeling in which inflammation plays a key role. Interaction between different members of the tumor necrosis factor superfamily and their cognate receptors induce several biological actions that may participate in CVD. The cytokine tumor necrosis factor-like weak inducer of apoptosis (TWEAK) and its functional receptor, fibroblast growth factor-inducible 14 (Fn14), are abundantly expressed during pathological cardiovascular remodeling. The TWEAK/Fn14 axis controls a variety of cellular functions, such as proliferation, differentiation, and apoptosis, and has several biological functions, such as inflammation and fibrosis that are linked to CVD. It has been demonstrated that persistent TWEAK/Fn14 activation is involved in both vessel and heart remodeling associated with acute and chronic CVD. In this review, we summarized the role of the TWEAK/Fn14 axis during pathological cardiovascular remodeling, highlighting the cellular components and the signaling pathways that are involved in these processes.

MicroRNA-889 Inhibits Autophagy To Maintain Mycobacterial Survival in Patients with Latent Tuberculosis Infection by Targeting TWEAK.

Autophagy plays an important role in protecting the host against pathogens. Mycobacterium tuberculosis can suppress autophagy and then remain dormant and survive within the host for an extended period, which is responsible for latent tuberculosis infection (LTBI). Here, we explored the role of microRNAs (miRNAs) in LTBI. The miRNA profiles were explored using the next-generation sequencing approach, followed by quantitative reverse transcription-PCR validation. The biological function of candidate miRNA was evaluated using immunoblotting, immunofluorescence techniques, and enzyme-linked immunosorbent assay in an in vitro human TB granuloma model. An increased miR-889 expression was observed in patients with LTBI compared with that in patients without infection. The reporter assay identified tumor necrosis factor (TNF)-like weak inducer of apoptosis (TWEAK) as the target of miR-889. Mycobacterial infection induced TWEAK upregulation in the early phase. TWEAK induced autophagy and promoted mycobacterial autophagosome maturation through activation of AMP-activated protein kinase (AMPK). Upon entry to LTBI status, elevated miR-889 levels were associated with TNF alpha (TNF-α) and granuloma formation/maintenance. MiR-889 inhibited autophagy via posttranscriptional suppression of TWEAK expression to maintain mycobacterial survival in granulomas. Adalimumab (anti-TNF-α monoclonal antibody) treatment reduced levels of both TNF-α and miR-889 and caused granuloma destruction and LTBI reactivation. The circulating miR-889 and TWEAK levels were correlated with LTBI and subsequently associated with anti-TNF-α-related LTBI reactivation in patients. We propose that miR-889 and TWEAK can act as targets that can be manipulated for antimycobacterial therapeutic purposes and act as candidate biomarkers for LTBI and LTBI reactivation, respectively.IMPORTANCE TB remains a leading cause of morbidity and mortality worldwide. Approximately one-quarter of the world's population has latent TB infection. TWEAK is a multiple-function cytokine and may be used as a target for the treatment of rheumatic diseases, cardiovascular diseases, and renal diseases. Here, we demonstrated a novel relationship between TWEAK and activation of the autophagic machinery which promotes antimycobacterial immunity. Additionally, TB infection is highly dynamic and determined by the interaction between the host and mycobacterium. We demonstrated a mechanism of fine-tuned balance between the mycobacterium and host for granuloma formation and/or maintenance in LTBI status. Once patients entered LTBI status, the upregulation of miR-889 was associated with TNF-α levels and granuloma formation to maintain mycobacterial survival. Adalimumab (a TNF-α inhibitor) reduced both TNF-α and miR-889 levels and caused LTBI reactivation and, thus, TWEAK enhancement. MiR-889 and TWEAK may become potential diagnostic biomarkers or therapeutic targets for LTBI and LTBI reactivation, respectively.

Experimental atopic dermatitis is dependent on the TWEAK/Fn14 signaling pathway.

Tumor necrosis factor (TNF)-like weak inducer of apoptosis (TWEAK) acts through its receptor fibroblast growth factor inducible 14 (Fn14), and participates in skin inflammation. Both TWEAK and Fn14 are highly expressed in skin lesions of patients with atopic dermatitis. The purpose of this study was to further explore the effect of Fn14 inhibition on experimental atopic dermatitis. Experimental atopic dermatitis was induced in the wild-type and Fn14 knock-out BALB/c mice. The effect of TWEAK/Fn14 interaction on keratinocytes was studied in an in-vitro model of atopic dermatitis. Fn14 deficiency ameliorates skin lesions in the mice model, accompanied by less infiltration of inflammatory cells and lower local levels of proinflammatory cytokines, including TWEAK, TNF-α and interleukin (IL)-17. Fn14 deficiency also attenuates the up-regulation of TNFR1 in skin lesions of atopic dermatitis. Moreover, topical TWEAK exacerbates skin lesion in the wild-type but not in the Fn14 knock-out mice. In vitro, TWEAK enhances the expressions of IL-17, IL-18 and IFN-γ in keratinocytes under atopic dermatitis-like inflammation. These results suggest that Fn14 deficiency protects mice from experimental atopic dermatitis, involving the attenuation of inflammatory responses and keratinocyte apoptosis. In the context of atopic dermatitis-like inflammation, TWEAK modulates keratinocytes via a TNFR1-mediated pathway.