Inhibition of both NOX and TNF-α exerts substantial renoprotective effects in renal ischemia reperfusion injury rat model.

BACKGROUND AND AIMS: Acute kidney injury (AKI) due to renal ischemia-reperfusion injury (RIRI) is associated with high morbidity and mortality, with no renoprotective drug available. Previous research focused on single drug targets, yet this approach has not reached translational success. Given the complexity of this condition, we aimed to identify a disease module and apply a multitarget network pharmacology approach. METHODS: Identification of a disease module with potential drug targets was performed utilizing Disease Module Detection algorithm using NADPH oxidases (NOXs) as seeds. We then assessed the protective effect of a multitarget network pharmacology targeting the identified module in a rat model of RIRI. Rats were divided into five groups; sham, RIRI, and RIRI treated with setanaxib (NOX inhibitor, 10 mg/kg), etanercept (TNF-α inhibitor, 10 mg/kg), and setanaxib and etanercept (5 mg/kg each). Kidney functions, histopathological changes and oxidative stress markers (MDA and reduced GSH) were assessed. Immunohistochemistry of inflammatory (TNF-α, NF-κB) apoptotic (cCasp-3, Bax/Bcl 2), fibrotic (α-SMA) and proteolysis (MMP-9) markers was performed. RESULTS: Our in-silico analysis yielded a disease module with TNF receptor 1 (TNFR1A) as the closest target to both NOX1 and NOX2. Targeting this module by a low-dose combination of setanaxib, and etanercept, resulted in a synergistic effect and ameliorated ischemic AKI in rats. This was evidenced by improved kidney function and reduced expression of inflammatory, apoptotic, proteolytic and fibrotic markers. CONCLUSIONS: Our findings show that applying a multitarget network pharmacology approach allows synergistic renoprotective effect in ischemic AKI and might pave the way towards translational success.

Effects of tumor necrosis factor-alpha inhibitors on lipid profiles in patients with psoriasis: a systematic review and meta-analysis.

Background: There is no consensus on the effect of tumor necrosis factor-alpha (TNF-alpha) inhibitors on lipid profiles in patients with psoriasis. This study aimed to investigate the effects of TNF-alpha inhibitors on lipid profiles (triglycerides, total cholesterol, low-density lipoprotein, or high-density lipoprotein) in patients with psoriasis. Methods: We searched PubMed, Embase, and Cochrane Library databases for articles published before October 17, 2023. Four TNF-alpha inhibitors (infliximab, etanercept, adalimumab, and certolizumab) were included in our study. (PROSPERO ID: CRD42023469703). Results: A total of twenty trials were included. Overall results revealed that TNF-alpha inhibitors elevated high-density lipoprotein levels in patients with psoriasis (WMD = 2.31; 95% CI: 0.96, 3.67; P = 0.001), which was supported by the results of sensitivity analyses excluding the effect of lipid-lowering drugs. Subgroup analyses indicated that high-density lipoprotein levels were significantly increased in the less than or equal to 3 months group (WMD = 2.88; 95% CI: 1.37, 4.4; P < 0.001), the etanercept group (WMD = 3.4; 95% CI = 1.71, 5.09, P < 0.001), and the psoriasis group (WMD = 2.52; 95% CI = 0.57, 4.48, P = 0.011). Triglyceride levels were significantly increased in the 3 to 6-month group (WMD = 4.98; 95% CI = 1.97, 7.99, P = 0.001) and significantly decreased in the 6-month and older group (WMD = -19.84; 95% CI = -23.97, -15.7, P < 0.001). Additionally, Triglyceride levels were significantly increased in the psoriasis group (WMD = 5.22; 95% CI = 2.23, 8.21, P = 0.001). Conclusion: Our results revealed that TNF-alpha inhibitors might temporarily increase high-density lipoprotein levels in patients with psoriasis. However, changes in triglycerides were not consistent among the different durations of treatment, with significant increases after 3 to 6 months of treatment. Future prospective trials with long-term follow-up contribute to confirming and extending our findings. Systematic Review Registration: https://www.crd.york.ac.uk/PROSPERO/, identifier CRD42023469703.

TNF-α blockage by etanercept restores spatial learning and reduces cellular degeneration in the hippocampus during liver cirrhosis.

Hepatic encephalopathy (HE) is one of the most debilitating cerebral complications of liver cirrhosis. The one-year survival of patients with liver cirrhosis and severe encephalopathy is less than 50%. Recent studies have indicated that neuroinflammation is a new player in the pathogenesis of HE, which seems to be involved in the development of cognitive impairment. In this study, we demonstrated neurobehavioral and neuropathological consequences of liver cirrhosis and tested the therapeutic potential of the tumor necrosis factor-α (TNF-α) inhibitor, etanercept. Sixty male adult Wistar albino rats (120-190 g) were allocated into four groups, where groups I and IV served as controls. Thioacetamide (TAA; 300 mg/kg) was intraperitoneally injected twice a week for five months to induce liver cirrhosis in group II (n = 20). Both TAA and etanercept (2 mg/kg) were administered to group III (n = 20). At the end of the experiment, spatial learning was assessed using Morris water maze. TNF-α was detected in both serum and hippocampus. The excised brains were also immunohistochemically stained with glial fibrillary acidic protein (GFAP) to estimate both the number and integrity of hippocampal astrocytes. Ultrastructural changes in the hippocampus were characterized by transmission electron microscopy. The results showed that blocking TNF-α by etanercept was accompanied by a lower TNF-α expression and a higher number of GFAP-positive astrocytes in the hippocampus. Etanercept intervention alleviated the neuronal and glial degenerative changes and impeded the deterioration of spatial learning ability. In conclusion, TNF-α is strongly involved in the development of liver cirrhosis and the associated encephalopathy. TNF-α blockers may be a promising approach for management of hepatic cirrhosis and its cerebral complications.

Autocrine positive feedback of tumor necrosis factor from activated microglia proposed to be of widespread relevance in chronic neurological disease.

Over a decade's experience of post-stroke rehabilitation by administering the specific anti-TNF biological, etanercept, by the novel perispinal route, is consistent with a wide range of chronically diminished neurological function having been caused by persistent excessive cerebral levels of TNF. We propose that this TNF persistence, and cerebral disease chronicity, largely arises from a positive autocrine feedback loop of this cytokine, allowing the persistence of microglial activation caused by the excess TNF that these cells produce. It appears that many of these observations have never been exploited to construct a broad understanding and treatment of certain chronic, yet reversible, neurological illnesses. We propose that this treatment allows these chronically activated microglia to revert to their normal quiescent state, rather than simply neutralizing the direct harmful effects of this cytokine after its release from microglia. Logically, this also applies to the chronic cerebral aspects of various other neurological conditions characterized by activated microglia. These include long COVID, Lyme disease, post-stroke syndromes, traumatic brain injury, chronic traumatic encephalopathy, post-chemotherapy, post-irradiation cerebral dysfunction, cerebral palsy, fetal alcohol syndrome, hepatic encephalopathy, the antinociceptive state of morphine tolerance, and neurogenic pain. In addition, certain psychiatric states, in isolation or as sequelae of infectious diseases such as Lyme disease and long COVID, are candidates for being understood through this approach and treated accordingly. Perispinal etanercept provides the prospect of being able to treat various chronic central nervous system illnesses, whether they are of infectious or non-infectious origin, through reversing excess TNF generation by microglia.

Tocilizumab suppresses NF-kappa B activation via toll-like receptor 9 signaling by reducing cell-free DNA in rheumatoid arthritis.

Endogenous DNA is released into the bloodstream as cell-free DNA (cfDNA) following cell death and is associated with various pathological conditions. However, their association with therapeutic drugs against rheumatoid arthritis (RA) remains unknown. Therefore, we investigated the significance of cfDNA in RA treated with tocilizumab and tumour necrosis factor inhibitor (TNF-I). Biological DMARDs (bDMARDs), including tocilizumab and TNF-I, were administered to 77 and 59 RA patients, respectively. Plasma cfDNA levels were measured at weeks 0, 4, and 12 by quantitative polymerase chain reaction. Disease activity was evaluated at the same time point using DAS28ESR. cfDNA levels from RA synovial cells treated with tocilizumab or etanercept for 24 h were measured. Human toll-like receptor 9 (hTLR9)-expressing HEK293 cells, which release secreted embryonic alkaline phosphatase (SEAP) upon NF-κB activation, were stimulated by cfDNA from RA patients, and subsequently, SEAP levels were determined. NF-κB translocation was evaluated by immunofluorescence staining with or without tocilizumab. The DAS28ESR significantly improved in both bDMARD groups at week 12. However, plasma cfDNA levels significantly decreased in the tocilizumab group at week 12 compared to that in week 0. cfDNA levels correlated with DAS28ESR in biological treatment-naïve patients administered tocilizumab. cfDNA levels in synovial cells were significantly suppressed by tocilizumab treatment and unaltered with etanercept. HEK293 cells released SEAP upon cfDNA stimulation, and the observed NF-κB nuclear translocation was suppressed by tocilizumab. Tocilizumab suppressed inflammation via the TLR9 pathway by decreasing cfDNA levels. Regulation of cfDNA may be a therapeutic target for RA.

TNF-α induced extracellular release of keratinocyte high-mobility group box 1 in Stevens-Johnson syndrome/toxic epidermal necrolysis: Biomarker and putative mechanism of pathogenesis.

Decreased epidermal high-mobility group box 1 (HMGB1) expression is an early marker of epidermal injury in Stevens-Johnson syndrome/toxic epidermal necrolysis (SJS/TEN). Etanercept, an anti-tumor necrosis factor therapeutic, is effective in the treatment of SJS/TEN. The objective was to characterize antitumor necrosis factor-alpha (TNF-α)-mediated HMGB1 keratinocyte/epidermal release and etanercept modulation. HMGB1 release from TNF-α treated (± etanercept), or doxycycline-inducible RIPK3 or Bak-expressing human keratinocyte cells (HaCaTs) was determined by western blot/ELISA. Healthy skin explants were treated with TNF-α or serum (1:10 dilution) from immune checkpoint inhibitor-tolerant, lichenoid dermatitis or SJS/TEN patients ± etanercept. Histological and immunohistochemical analysis of HMGB1 was undertaken. TNF-α induced HMGB1 release in vitro via both necroptosis and apoptosis. Exposure of skin explants to TNF-α or SJS/TEN serum resulted in significant epidermal toxicity/detachment with substantial HMGB1 release which was attenuated by etanercept. Whole-slide image analysis of biopsies demonstrated significantly lower epidermal HMGB1 in pre-blistered SJS/TEN versus control (P < 0.05). Keratinocyte HMGB1 release, predominantly caused by necroptosis, can be attenuated by etanercept. Although TNF-α is a key mediator of epidermal HMGB1 release, other cytokines/cytotoxic proteins also contribute. Skin explant models represent a potential model of SJS/TEN that could be utilized for further mechanistic studies and targeted therapy screening.

Docosahexaenoic acid and etanercept could reduce functional and metabolic alterations during collagen-induced arthritis in rats without any synergistic effect.

AIMS: Rheumatoid arthritis is an autoimmune disease which induces chronic inflammation and increases the risk for sarcopenia and metabolic abnormalities. Nutritional strategies using omega 3 polyunsaturated fatty acids could be proposed to alleviate inflammation and improve the maintenance of lean mass. Independently, pharmacological agents targeting key molecular regulators of the pathology such as TNF alpha could be proposed, but multiple therapies are frequently necessary increasing the risk for toxicity and adverse effects. The aim of the present study was to explore if the combination of an anti-TNF therapy (Etanercept) with dietary supplementation with omega 3 PUFA could prevent pain and metabolic effects of RA. MATERIALS AND METHODS: RA was induced using collagen-induced arthritis (CIA) in rats to explore of supplementation with docosahexaenoic acid, treatment with etanercept or their association could alleviate symptoms of RA (pain, dysmobility), sarcopenia and metabolic alterations. KEY FINDINGS: We observed that Etanercept had major benefits on pain and RA scoring index. However, DHA could reduce the impact on body composition and metabolic alterations. SIGNIFICANCE: This study revealed for the first time that nutritional supplementation with omega 3 fatty acid could reduce some symptoms of rheumatoid arthritis and be an effective preventive treatment in patients who do not need pharmacological therapy, but no sign of synergy with an anti-TNF agent was observed.

Transcriptomic responses of peripheral blood mononuclear cells to cyclosporin and etanercept in a female infant with juvenile generalized pustular psoriasis.

Generalized pustular psoriasis (GPP) is a rare but severe form of psoriasis. An early onset of the diseases is correlated with mutations among IL36RN, CARD14, AP1S3, MPO and SERPINA3 genes. Systemic biological agents including anti-TNF-α, anti-IL-17, anti-IL-12/IL-23, anti-IL1R, anti-IL1ß and anti-IL-36R act as novel treatment methods for GPP. Herein we report a female infant clinically diagnosed with GPP since she was 10-month-old. Results of whole-exome sequencing (WES) and Sanger sequencing revealed a reported heterozygous IL36RN (c.115+6T>C) and another reported heterozygous SERPINA3 frame-shifting variant (c.1247_1248del). Initial cyclosporin treatment for the patient led to a partial remission of the symptoms. However, the patient reached nearly total remission of pustules and erythema after anti-TNF-α inhibitor etanercept treatment. Results of further RNA sequencing (RNA-seq) done on peripheral blood mononuclear cells correlated with the clinical responses, showing that cyclosporin suppressed a portion of the neutrophil-related genes, while most genes associated with neutrophil activation, neutrophil-mediated immunity and degranulation were downregulated by the subsequent etanercept treatment. We report this case to demonstrate WES and RNA-seq in combination could come in handy in reaching a precise diagnosis and in evaluating or even predicting the molecular alterations underlying clinical treatment effectiveness.

The role of proteasome in muscle wasting of experimental arthritis.

BACKGROUND: Rheumatoid arthritis is an autoimmune inflammatory disease that often leads patients to muscle impairment and physical disability. This study aimed to evaluate changes in the activity of proteasome system in skeletal muscles of mice with collagen-induced arthritis (CIA) and treated with etanercept or methotrexate. METHODS: Male DBA1/J mice were divided into four groups (n = 8 each): CIA-Vehicle (treated with saline), CIA-ETN (treated with etanercept, 5.5 mg/kg), CIA-MTX (treated with methotrexate, 35 mg/kg) and CO (healthy control group). Mice were treated two times a week for 6 weeks. Clinical score and hind paw edema were measured. Muscles were weighted after euthanasia and used to quantify proteasome activity, gene (MuRF-1, PMSα4, PSMß5, PMSß6, PSMß7, PSMß8, PSMß9, and PSMß10), and protein (PSMß1, PSMß5, PSMß1i, PSMß5i) expression of proteasome subunits. RESULTS: Both treatments slowed disease development, but only CIA-ETN maintained muscle weight compared to CIA-MTX and CIA-Vehicle groups. Etanercept treatment showed caspase-like activity of 26S proteasome similar to CO group, while CIA-Vehicle and CIA-MTX had higher activity compared to CO group (p: 0.0057). MuRF-1 mRNA expression was decreased after etanercept administration compared to CIA-Vehicle and CO groups (p: 0.002, p: 0.007, respectively). PSMß8 and PSMß9 mRNA levels were increased in CIA-Vehicle and CIA-MTX compared to CO group, while CIA-ETN presented no difference from CO. PMSß6 mRNA expression was higher in CIA-Vehicle and CIA-MTX groups than in CO group. Protein levels of the PSMß5 subunit were increased in CO group compared to CIA-Vehicle; after both etanercept and methotrexate treatments, PSMß5 expression was higher than in CIA-Vehicle group and did not differ from CO group expression (p: 0.0025, p: 0.001, respectively). The inflammation-induced subunit ß1 (LMP2) was enhanced after methotrexate treatment compared to CO group (p: 0.043). CONCLUSIONS: The results of CIA-Vehicle show that arthritis increases muscle proteasome activation by enhanced caspase-like activity of 26S proteasome and increased PSMß8 and PSMß9 mRNA levels. Etanercept treatment was able to maintain the muscle weight and to modulate proteasome so that its activity and gene expression were compared to CO after TNF inhibition. The protein expression of inflammation-induced proteasome subunit was increased in muscle of CIA-MTX group but not following etanercept treatment. Thus, anti-TNF treatment may be an interesting approach to attenuate the arthritis-related muscle wasting.

Tumour necrosis factor blockade after asphyxia in foetal sheep ameliorates cystic white matter injury.

Cystic white matter injury is highly associated with severe neurodevelopmental disability and cerebral palsy in preterm infants, yet its pathogenesis remains poorly understood and there is no established treatment. In the present study, we tested the hypothesis that slowly evolving cystic white matter injury after hypoxia-ischaemia is mediated by programmed necrosis initiated by tumour necrosis factor. Tumour necrosis factor blockade was begun 3 days after hypoxia-ischaemia to target the tertiary phase of injury, when most secondary cell death is thought to be complete. Chronically instrumented preterm foetal sheep (0.7 gestation) received 25 min of hypoxia-ischaemia induced by complete umbilical cord occlusion or sham-umbilical cord occlusion (controls, n = 10), followed by intracerebroventricular infusion of the soluble TNF inhibitor, Etanercept, at 3, 8 and 13 days after umbilical cord occlusion (n = 9) or vehicle (n = 9). Foetal brains were processed for histology at 21 days after umbilical cord occlusion. Umbilical cord occlusion with vehicle was associated with a spectrum of macroscopic white matter degeneration, including white matter atrophy, ventriculomegaly and overt temporal lobe cystic white matter injury. Oligodendrocyte maturational arrest and impaired labelling of myelin proteins, characteristic of diffuse white matter injury, was observed in the parietal lobe and surrounding the cystic lesions in the temporal lobe. Etanercept markedly attenuated cystic white matter injury on the side of the intracerebroventricular infusion, with partial contralateral protection. Further, Etanercept improved oligodendrocyte maturation and labelling of myelin proteins in the temporal and parietal lobes. The present study shows that cystic white matter injury reflects late-onset tertiary cell death mediated by delayed neuroinflammation through the tumour necrosis factor pathway. Delayed tumour necrosis factor blockade markedly attenuated cystic white matter injury and restored oligodendrocyte maturation and deficits in myelin protein expression. These data suggest that delayed tumour necrosis factor blockade may represent a viable therapeutic strategy to reduce the risk of cystic and diffuse white matter injury and potentially cerebral palsy after preterm birth, with a surprisingly wide therapeutic window.

The implication of long non-coding RNA expression profile in rheumatoid arthritis: Correlation with treatment response to tumor necrosis factor inhibitor.

OBJECTIVE: This study aimed to investigate the linkage of long non-coding RNA (lncRNA) expression profile with etanercept response in rheumatoid arthritis (RA) patients. METHODS: Peripheral blood mononuclear cell (PBMC) samples were collected from 80 RA patients prior to etanercept treatment. Samples from eight responders and eight non-responders at week 24 (W24) were proposed to RNA-sequencing, then 10 candidate lncRNAs were sorted and their PBMC expressions were validated by reverse transcription quantitative chain reaction (RT-qPCR) in 80 RA patients. Subsequently, clinical response by lncRNA (CRLnc) prediction model was established. RESULTS: RNA-sequencing identified 254 up-regulated and 265 down-regulated lncRNAs in W24 responders compared with non-responders, which were enriched in immune or joint related pathways such as B-cell receptor signaling, osteoclast differentiation and T-cell receptor signaling pathways, etc. By reverse transcription quantitative chain reaction (RT-qPCR) validation: Two lncRNAs were correlated with W4 response, three lncRNAs were correlated with W12 response, seven lncRNAs were correlated with W24 response. Subsequently, to construct and validate CRLnc prediction model, 80 RA patients were randomly divided into test set (n = 40) and validation set (n = 40). In the test set, lncRNA RP3-466P17.2 (OR = 9.743, P = .028), RP11-20D14.6 (OR = 10.935, P = .007), RP11-844P9.2 (OR = 0.075, P = .022), and TAS2R64P (OR = 0.044, P = .016) independently related to W24 etanercept response; then CRLnc prediction model integrating these four lncRNAs presented a good value in predicting W24 etanercept response (Area Under Curve (AUC): 0.956, 95%CI: 0.896-1.000). However, in the validation set, the CRLnc prediction model only exhibited a certain value in predicting W24 etanercept response (AUC: 0.753, 95%CI: 0.536-0.969). CONCLUSIONS: CRLnc prediction model is potentially a useful tool to instruct etanercept treatment in RA patients.

TNF or EGFR inhibition equally block AKI-to-CKD transition: opportunities for etanercept treatment.

BACKGROUND: Inflammation is a key driver of the transition of acute kidney injury to progressive fibrosis and chronic kidney disease (AKI-to-CKD transition). Blocking a-disintegrin-and-metalloprotease-17 (ADAM17)-dependent ectodomain shedding, in particular of epidermal growth factor receptor (EGFR) ligands and of the type 1 inflammatory cytokine tumor necrosis factor (TNF), reduces pro-inflammatory and pro-fibrotic responses after ischemic AKI or unilateral ureteral obstruction (UUO), a classical fibrosis model. Metalloprotease or EGFR inhibition show significant undesirable side effects in humans. In retrospective studies anti-TNF biologics reduce the incidence and progression of CKD in humans. Whether TNF has a role in AKI-to-CKD transition and how TNF inhibition compares to EGFR inhibition is largely unknown. METHODS: Mice were subjected to bilateral renal ischemia-reperfusion injury or unilateral ureteral obstruction. Kidneys were analyzed by histology, immunohistochemistry, qPCR, western blot, mass cytometry, scRNA sequencing, and cytokine profiling. RESULTS: Here we show that TNF or EGFR inhibition reduce AKI-to-CKD transition and fibrosis equally by about 25%, while combination has no additional effect. EGFR inhibition reduced kidney TNF expression by about 50% largely by reducing accumulation of TNF expressing immune cells in the kidney early after AKI, while TNF inhibition did not affect EGFR activation or immune cell accumulation. Using scRNAseq data we show that TNF is predominantly expressed by immune cells in AKI but not in proximal tubule cells (PTC), and PTC-TNF knockout did not affect AKI-to-CKD transition in UUO. Thus, the anti-inflammatory and anti-fibrotic effects of the anti-TNF biologic etanercept in AKI-to-CKD transition rely on blocking TNF that is released from immune cells recruited or accumulating in response to PTC-EGFR signals. CONCLUSION: Short-term anti-TNF biologics during or after AKI could be helpful in the prevention of AKI-to-CKD transition.

Combination therapy of a TRPV2 agonist with a TNF inhibitor achieves sustained suppression of disease severity and reduced joint damage.

We aimed to compare a transient receptor potential vanilloid 2 (TRPV2) agonist with a TNF inhibitor, and to test the potential of their combination in collagen-induced arthritis (CIA) as a potential future strategy for rheumatoid arthritis (RA). Following the onset of CIA DBA1/j mice were started on treatment with either vehicle, etanercept (8 mg/kg three times a week), the TRPV2 agonist O1821 (20-30 mg/kg/day), or a combination of both. Mice were scored over a 61-day period. Synovial tissues were obtained for RNA sequencing. Mice on monotherapy with either O1821 or etanercept developed milder clinical disease. The O1821 protection was observed at an earlier time-point than in the etanercept group. The combination therapy group achieved a more robust and sustained reduction in disease severity than either monotherapy group. All treatment groups had reduced scores for synovial inflammation, synovial hyperplasia, and erosive changes, compared with controls, with the combination group achieving the most significant protection. RNA sequencing and pathway analyses of synovial tissues identified pathways and processes regulated by the TRPV2 agonist, such as chemotaxis and cytokine receptor signaling, including IL6R. The combination therapy affected additional pathways not seen in the monotherapy groups. In conclusion, the TRPV2 agonist achieved an overall similar reduction in arthritis severity and histology scores as etanercept, but the combination therapy achieved a more sustained disease control and more pronounced reduction in joint damage, suggesting a potential future option for improving disease control in RA. RNA sequencing analyses identified new pathways regulated by TRPV2, and also by the combination treatment.

TNF-α‒Mediated Keratinocyte Expression and Release of Matrix Metalloproteinase 9: Putative Mechanism of Pathogenesis in Stevens‒Johnson Syndrome/Toxic Epidermal Necrolysis.

Stevens‒Johnson syndrome and toxic epidermal necrolysis (SJS/TEN) are severe cutaneous adverse drug reactions characterized by widespread keratinocyte cell death and epidermal detachment. At present, there is little understanding of how the detachment occurs or how it is abrogated by the TNF-α inhibitor etanercept, an effective SJS/TEN treatment. RNA sequencing was used to identify upregulated transcripts in formalin-fixed paraffin-embedded SJS/TEN skin biopsies. Epidermal matrix metalloproteinase 9 (MMP9) expression was assessed by immunohistochemistry in skin biopsies and cultured human skin explants exposed to serum from patients with cutaneous adverse drug reactions. TNF-α‒induced MMP9 expression and activity and its abrogation by etanercept were determined using the HaCaT immortalized keratinocyte cell line. Epidermal MMP9 expression was significantly higher in SJS/TEN skin (70.6%) than in healthy control skin (0%) (P = 0.0098) and nonbullous skin reactions (10.7%) (P = 0.0002). SJS/TEN serum induced significant MMP9 expression and collagenase activity in healthy skin explants, which was reduced by etanercept. Etanercept was also able to negate the TNF-α‒induced MMP9 expression in the HaCaT cell line. Data suggest that elevated epidermal MMP9 expression and collagenase activity are a putative pathogenic mechanism in SJS/TEN, which is limited by etanercept. Modulation of MMP9 expression and activity represents, to our knowledge, a previously unreported therapeutic target for the treatment of SJS/TEN.

A randomized controlled pilot trial of etanercept and alpha-1 antitrypsin to improve autologous islet engraftment.

BACKGROUND: In total pancreatectomy with islet auto-transplantation, successful diabetes outcomes are limited by islet loss from the instant blood mediated inflammatory response. We hypothesized that blockade of the inflammatory response with either etanercept or alpha-1-antitrypsin would improve islet function and insulin independence. METHODS: We randomized 43 participants to receive A1AT (90 mg/kg x 6 doses, n = 13), or etanercept (50 mg then 25 mg x 5 doses, n = 14), or standard care (n = 16), aiming to reduce detrimental effects of innate inflammation on early islet survival. Islet graft function was assessed using mixed meal tolerance testing, intravenous glucose tolerance testing, glucose-potentiated arginine-induced insulin secretion studies, HbA1c, and insulin dose 3 months and 1 year post-TPIAT. RESULTS: We observed the most robust acute insulin response (AIRglu) and acute C-peptide response to glucose (ACRglu) at 3 months after TPIAT in the etanercept-treated group (p ≤ 0.02), but no differences in other efficacy measures. The groups did not differ overall at 1 year but when adjusted by sex, there was a trend towards a sex-specific treatment effect in females (AIRglu p = 0.05, ACRglu p = 0.06), with insulin secretion measures highest in A1AT-treated females. CONCLUSION: Our randomized trial supports a potential role for etanercept in optimizing early islet engraftment but it is unclear whether this benefit is sustained. Further studies are needed to evaluate possible sex-specific responses to either treatment. CLINICAL TRIAL NOTATION: This study was performed under an Investigational New Drug Application (IND #119828) from the Food and Drug Administration and was registered on clinicaltrials.gov (NCT#02713997).

Cytokine modulation by etanercept ameliorates metabolic syndrome and its related complications induced in rats administered a high-fat high-fructose diet.

The aim of the present study was to investigate the effect of etanercept (ETA)-an anti-tumor necrosis factor α (TNF-α) monoclonal antibody-on metabolic disorders such as obesity, hypertension, dyslipidemia, and insulin resistance associated with the metabolic syndrome (MS). MS was induced in rats via high-fat high-fructose (HFHF) administration for 8 weeks. Rats were divided into three groups: negative control, HFHF model, and ETA-treated groups [HFHF + ETA (0.8 mg/kg/twice weekly, subcutaneously) administered in the last 4 weeks]. ETA effectively diminished the prominent features of MS via a significant reduction in the percent body weight gain along with the modulation of adipokine levels, resulting in a significant elevation of serum adiponectin consistent with TNF-α and serum leptin level normalization. Moreover, ETA enhanced dyslipidemia and the elevated blood pressure. ETA managed the prominent features of MS and its associated complications via the downregulation of the hepatic inflammatory pathway that induces nonalcoholic steatohepatitis (NASH)-from the expression of Toll-like receptor 4, nuclear factor kappa B, and TNF-α until that of transforming growth factor-in addition to significant improvements in glucose utilization, insulin sensitivity, and liver function parameter activity and histopathological examination. ETA was effective for the treatment of all prominent features of MS and its associated complications, such as type II diabetes mellitus and NASH.

Etanercept alleviates psoriasis by reducing the Th17/Treg ratio and promoting M2 polarization of macrophages.

INTRODUCTION: This study aimed to investigate the effect of etanercept in psoriasis and its underlying mechanism. METHODS: Female mice were treated with imiquimod (IMQ) to induce psoriasis, and intraperitoneally administered etanercept (0.1-0.4 mg/ml). The RAW264.7 cells were treated with LPS and IFN-γ to polarize to M1, and were treated with IL-13 and IL-4 to polarize to M2. RESULTS: In our study, Etanercept markedly reduced the psoriasis area and severity index scores, and epidermal thickness of mice induced by IMQ. In addition, etanercept reduced the levels of TNF-α and IL-6/12/23, and enhanced the levels of IL-4/10, reduced Th17/Treg ratio and facilitated the polarization of macrophages to M2 in psoriasis model mice. Furthermore, etanercept inhibited the JAK/STAT3 pathway and increased the protein levels of SOCS1 and SOCS3. CONCLUSIONS: In conclusion, our findings indicated that etanercept could inhibit the JAK/STAT3 pathway to reduce Th17/Treg ratio and promote M2 polarization, thereby alleviating psoriasis of mice.

Machine learning predicts response to TNF inhibitors in rheumatoid arthritis: results on the ESPOIR and ABIRISK cohorts.

OBJECTIVES: Around 30% of patients with rheumatoid arthritis (RA) do not respond to tumour necrosis factor inhibitors (TNFi). We aimed to predict patient response to TNFi using machine learning on simple clinical and biological data. METHODS: We used data from the RA ESPOIR cohort to train our models. The endpoints were the EULAR response and the change in Disease Activity Score (DAS28). We compared the performances of multiple models (linear regression, random forest, XGBoost and CatBoost) on the training set and cross-validated them using the area under the receiver operating characteristic curve (AUROC) or the mean squared error. The best model was then evaluated on a replication cohort (ABIRISK). RESULTS: We included 161 patients from ESPOIR and 118 patients from ABIRISK. The key selected features were DAS28, lymphocytes, ALT (aspartate aminotransferase), neutrophils, age, weight, and smoking status. When predicting EULAR response, CatBoost achieved the best performances of the four tested models. It reached an AUROC of 0.72 (0.68-0.73) on the train set (ESPOIR). Better results were obtained on the train set when etanercept and monoclonal antibodies were analysed separately. On the test set (ABIRISK), these models respectively achieved on AUROC of 0.70 (0.57-0.82) and 0.71 (0.55-0.86). Two decision thresholds were tested. The first prioritised a high confidence in identifying responders and yielded a confidence up to 90% for predicting response. The second prioritised a high confidence in identifying inadequate responders and yielded a confidence up to 70% for predicting non-response. The change in DAS28 was predicted with an average error of 1.1 DAS28 points. CONCLUSION: The machine learning models developed allowed predicting patient response to TNFi exclusively using data available in clinical routine.

Discovery of novel targets in a complex regional pain syndrome mouse model by transcriptomics: TNF and JAK-STAT pathways.

Complex Regional Pain Syndrome (CRPS) represents severe chronic pain, hypersensitivity, and inflammation induced by sensory-immune-vascular interactions after a small injury. Since the therapy is unsatisfactory, there is a great need to identify novel drug targets. Unbiased transcriptomic analysis of the dorsal root ganglia (DRG) was performed in a passive transfer-trauma mouse model, and the predicted pathways were confirmed by pharmacological interventions. In the unilateral L3-5 DRGs 125 genes were differentially expressed in response to plantar incision and injecting IgG of CRPS patients. These are related to inflammatory and immune responses, cytokines, chemokines and neuropeptides. Pathway analysis revealed the involvement of Tumor Necrosis Factor (TNF) and Janus kinase (JAK-STAT) signaling. The relevance of these pathways was proven by abolished CRPS IgG-induced hyperalgesia and reduced microglia and astrocyte markers in pain-associated central nervous system regions after treatment with the soluble TNF alpha receptor etanercept or JAK inhibitor tofacitinib. These results provide the first evidence for CRPS-related neuroinflammation and abnormal cytokine signaling at the level of the primary sensory neurons in a translational mouse model and suggest that etanercept and tofacitinib might have drug repositioning potentials for CRPS-related pain.

Effects of Etanercept on TNF-α Inhibition in Rats with Adenine-Induced Chronic Kidney Disease.

Introduction: Chronic kidney disease (CKD) constitutes a chronic inflammatory state associated with an increase in inflammatory mediators and profibrotic molecules such as tumor necrosis factor-α (TNF-α). Etanercept (ETA) is a TNF inhibitor widely used in treatment of autoimmune inflammatory diseases. However, the effects of TNF-α inhibition in the establishment of CKD have not been fully elucidated. We evaluate the effects of TNF inhibition by ETA in adenine- (Ad-) induced CKD in rats. Methods: Rats were divided into three groups: control, renal injury model, and model plus ETA (2 mg/kg, 3 times per week (w); sc). Renal injury was induced by Ad administration (100 mg/kg, daily for 2 or 4 w; orogastric). Serum TNF-α levels and biochemical parameters for renal function were evaluated. Histopathological changes in the kidney were assessed using H&E and Masson's trichrome staining and also immunostaining for tubular cells. Results: Ad administration produced a renal functional decline, tubular atrophy, interstitial inflammation, and fibrosis for 2 w, followed by renal anemia, several renal dysfunctions, tubular atrophy, and fibrosis at 4 w. A significant increase in serum TNF-α levels was observed from 2 w of Ad administration and remained elevated up to 4 w. Treatment with ETA partially reduced kidney damage but was very effective to blocking serum TNF-α. Conclusion: Although inhibition of TNF by ETA was very effective in reducing serum TNF-α, this strategy was partially effective in preventing Ad-induced CKD.