anti-TNFα Antibody Infliximab Research Articles

TNFα is a key trigger of inflammation in diet-induced non-obese MASLD in mice

Tumor necrosis factor alpha (TNFα) is thought to be a critical factor in the development of metabolic dysfunction-associated steatotic liver disease (MASLD). Here, we determined the effects of a treatment with the anti-TNFα antibody infliximab and a genetic deletion of TNFα, respectively, in the development of non-obese diet-induced early metabolic dysfunction-associated steatohepatitis (MASH) in mice. The treatment with infliximab improved markers of liver damage in mice with pre-existing early MASH. In TNFα−/− mice, the development of early signs of MASH and insulin resistance was significantly attenuated compared to wild-type animals. While mRNA expression of proinflammatory cytokines like interleukin 1β (Il1b) and interleukin 6 (Il6) were significantly lower in livers of MASH-diet-fed TNFα−/− mice compared to wild-type mice with early MASH, markers of intestinal barrier function were similarly impaired in both MASH-diet-fed groups compared to controls. Our data suggest that TNFα is a key regulator of hepatic inflammation and insulin resistance associated with the development of early non-obese MASH.

Development and Optimization of Therapeutic Analogues of Anti-TNFα Antibody Infliximab

Development and Optimization of Therapeutic Analogues of Anti-TNFα Antibody Infliximab

Vedolizumab is associated with changes in innate rather than adaptive immunity in patients with inflammatory bowel disease

ObjectiveVedolizumab, a monoclonal antibody directed against the integrin heterodimer α4β7, is approved for the treatment of Crohn’s disease and ulcerative colitis. The efficacy of vedolizumab has been suggested to result...

Dextran sulphate sodium colitis in C57BL/6J mice is alleviated by Lactococcus lactis and worsened by the neutralization of Tumor necrosis Factor α

TNFα has a well-established role in inflammatory bowel disease that affects the gastrointestinal tract and is usually manifested as Crohn's disease or ulcerative colitis. We have compared Lactococcus lactis NZ9000 displaying TNFα-binding affibody with control Lactococcus lactis and with anti-TNFα antibody infliximab for the treatment of mice with dextran sulphate sodium (DSS)-induced colitis. L. lactis NZ9000 alleviated the colitis severity one week after colitis induction with DSS, more effectively when administered in preventive fashion prior to, during and after DSS administration. TNFα-binding L. lactis was less effective than control L. lactis, particularly when TNFα-binding L. lactis was administered in preventive fashion. Similarly, an apparently detrimental effect of TNFα neutralization was observed in mice that were intraperitoneally administered anti-TNFα monoclonal antibody infliximab prior to colitis induction. The highest concentrations of tissue TNFα were observed in groups without DSS colitis that were treated either with TNFα-binding L. lactis or infliximab. To conclude, we have confirmed that L. lactis exerts a protective effect on DSS-induced colitis in mice. Contrary to expectations, but in line with some reports, the neutralization of TNFα aggravated disease symptoms in the acute phase of colitis and increased TNFα concentration in colon tissue of healthy mice. Nevertheless, we have demonstrated that oral administration of bacteria with surface displayed TNFα-binding affibody can interfere significantly with TNFα signaling and mimic the infliximab response in the given animal model of colitis.

Infliximab improves endothelial dysfunction in a mouse model of antiphospholipid syndrome: Role of reduced oxidative stress

Antiphospholipid syndrome (APS), induces endothelial dysfunction, oxidative stress and systemic inflammation that may be mediated by TNFα. Thus, we investigated the possible protective effect of the anti-TNFα antibody infliximab (5μg/g) on endothelial function in a mouse APS model (induced by injection of purified human anti-β2GP1-IgG).Seven days after anti-β2GPI-IgG injection, we observed an increase in plasma sVCAM-1 and sE-selectin levels and in aortic mRNA expression of VCAM-1 and E-selectin. This was associated with a decreased endothelium-dependent relaxation of isolated mesenteric arteries to acetylcholine, together with decreased mesenteric eNOS mRNA expression and increased eNOS uncoupling, accompanied by increased iNOS and gp91phox mRNA and increased left ventricular GSH/GSSH ratio. Infliximab significantly improved the NO-mediated relaxing responses to acetylcholine, and induced a decrease in iNOS and gp91phox mRNA expression. The õpro-adhesive and pro-coagulant phenotypes induced by the anti-β2GP1-IgG were also reversed.This study provides the first evidence that TNFα antagonism improves endothelial dysfunction in APS and suggests that endothelial dysfunction is mediated by TNFα and oxidative stress. Therefore, infliximab may be of special relevance in clinical practice.

Cutaneous Adverse Reactions during Anti-Tnf Alpha Treatment for Inflammatory Bowel Diseases: The Experience of the Dermatology Clinic of Cagliari

Background Anti-tumour necrosis factor alpha are a well-documented class of disease-modifying therapy for inflammatory bowel diseases, including Crohn's disease and ulcerative colitis. The monoclonal anti-TNFα antibody Infliximab and Adalimumab are the current approved drugs in Italy, showing high efficacy, but also variable clinically significant adverse effects. Method From 2012, an observational prospective study was activated at the Dermatology Clinic of Cagliari University, to screen all patients with inflammatory bowel diseases treated with Infliximab and Adalimumab for cutaneous adverse reactions and skin cancers. A basal visit, complete of melanocytic lesions digital image recording was performed in all patients referred from the Gastroenterology Unit of the same University Hospital. Subsequent visits were performed when new lesions or modification of the skin appearance occurred. All cutaneous adverse reactions associated with anti-TNF treatment were reported to the Italian Pharmacovigilance network. Results Ninety-one patients were included in the study: 58 (31 women and 27 men) patients affected by Crohn's disease, (the age range was between 16 and 69 years), and 33 patients with ulcerative colitis (15 women and 17 men; age range between 21 and 68 years). Proportion of patients treated with infliximab (52%) was similar to adalimumab (48%). Cutaneous adverse reactions were observed in 38 patients (42%): 20 were taking Infliximab (52%) and 18 Adalimumab (48%). Adverse reactions observed by frequency included: infections (32%), eczematous dermatitis (23%), infusion systemic reactions and at the injection site (5.7%), psoriasis paradoxical reaction (5.7%), followed by a mixture of different cutaneous reactions, including urticaria, photosensitivity, chronic lupus erythematosus, lichenoid eruption, alopecia areata, hypertrichosis. Benign skin tumors eruption (15%) and basal cell carcinoma (2.8%) occurrence were also noted. Patient with dysplastic melanocytic nevi (10%) at screening were regularly followed with digital dermoscopy, but no one showed changes in the study period. Conclusion Our sample of patients had a high rate of skin adverse reactions (42%), but considering severity, no case required definitive discontinuation of therapy. Dermatologist support might be critical to optimize biological agent management in inflammatory bowel diseases, performing a careful basal screening, strict monitoring of adverse reactions and prompt intervention, as well as educating the patient towards skin cancer prevention, considering the long-term drug exposure.

Effect of N-acetylcysteine combined with infliximab on toxic epidermal necrolysis. A proof-of-concept study

IntroductionThe pathophysiology of toxic epidermal necrolysis (TEN) is thought to be related to a drug-induced oxidative stress combined with TNFα overexpression by keratinocytes. None of the current treatments for TEN including systemic corticosteroids, cyclosporine and intravenous administration of immunoglobulins has proven superior over supportive care only. MethodsA total of 10 TEN patients were enrolled to be treated at admission in burn units with the antioxidant N-acetylcysteine [NAC, 150mg/kg in a 20-h intravenous (IV) administration], or the combination of the same IV NAC perfusion with the anti-TNFα antibody infliximab (Remicade®), administered at a 5mg/kg dosage as a single 2-h IV administration. TEN was confirmed by a skin biopsy taken from a bullous lesion. At entry in the trial and 48h later, the illness auxiliary score (IAS) of clinical severity was determined and the extent in altered skin area (erythema and blisters) was assessed as a relative body area. Skin biopsies of both clinically uninvolved and erythematous areas were collected and immunohistochemistry was performed for assessing the density of inflammatory cells (CD8+ T cells, CD68+ macrophages) and keratinocytes enriched in intracellular calcium (Ca++) identified by the Mac387 anti-calprotectin antibody. ResultsNo unexpected drug-induced adverse event was noticed. After 48h of both treatment modalities, improvements were not observed in the extent of skin involvement and in IAS. Immunohistopathology showed the absence of reduction in the amount of intraepidermal inflammatory cells. An increased intracellular Ca++ load in clinically uninvolved keratinocytes and in erythematous epidermis was noticed. This latter finding suggested the progression in the way of the apoptotic process. On burn unit discharge, the survival in each modality of treatment was not improved compared to the expected outcomes determined from the IAS at admission. ConclusionsIn this proof-to-concept attempt, NAC treatment or its combination with infliximab did not appear to reverse the evolving TEN process.

TNFα suppresses IFNγ-induced MHC class II expression on retinal pigmented epithelial cells cultures

One major consequence of retinal pigment epithelium (RPE) cell activation during autoimmune uveitis is the induction of MHC II molecules expression at their surface. IFNγ is regarded as the main cytokine involved in this induction. As TNFα plays a central role in autoimmune uveitis, we investigated its effects on IFNγ-mediated MHC II induction on RPE cells. Retinal pigment epithelium cells (ARPE-19) were stimulated with IFNγ, TNFα and the anti-TNFα antibody infliximab. The expression of MHCII and ICAM-1 was analysed by flow cytometry. The activation and expression of IRF-1 and STAT-1, two proteins involved in IFNγ-signalling pathway, were analysed by WB. Class II transactivator (CIITA) expression was monitored by qRT-PCR and immunoprecipitation. TNFα inhibits IFNγ-induced MHC II expression on ARPE cells in a dose-dependent manner. Infliximab completely reverses the inhibitory effect of TNFα. We did not observe an inhibitory effect of TNFα on the expression of ICAM-1 induced by IFNγ. Similarly, IFNγ-induced STAT1 phosphorylation and IRF1 expression were not affected by TNFα. On the contrary, we found that TNFα suppresses IFNγ-induced CIITA mRNA accumulation and protein expression. TNFα inhibits IFNγ-induced MHC II expression in RPE cells. This inhibitory effect was reversed by infliximab and was not because of a global inhibition of IFNγ -mediated RPE cell activation but rather to a specific down-regulation of CIITA expression. Those findings are consistent with the role of TNFα in the resolution of inflammation and might help to elucidate the complex development of autoimmune uveitis.

Resveratrol Ameliorates TNFα-Mediated Suppression of Erythroid Colony Formation in Human CD34+ Cells Via Cellular Pathways Involving Downregulation of NF-κB.

Abstract 2507Poster Board II-484Although the etiology of anemia remains unexplained in one third of elderly subjects, increasing evidence implicates chronic inflammation in the pathogenesis of anemia in the elderly population. We hypothesize that in addition to classical anemia of inflammation (AI) mediated by hepcidin, a significant proportion of unexplained anemia involves the overexpression of proinflammatory cytokines like TNFα that promote direct suppression of erythroid progenitors to a degree more prominent than the effects of hepcidin, resulting in the development of unexplained anemia marked by absence of the expected biochemical profile of AI. A better understanding of the pathophysiology of anemia subtypes is necessary if we are to develop effective targeted, oral therapies for patients. In the current study, we aimed to evaluate whether resveratrol, a flavanol found in high concentration in red wine that appears to possess potent anti-inflammatory properties, ameliorates TNFα-mediated erythroid colony suppression in human CD34+ cells. Human peripheral blood CD34+ cells from 3 healthy adults were first cultured in methylcellulose medium in the presence of IL-3 (10ng/ml), EPO (1U/ml), SCF (50ng/ml) and increasing doses of TNFα (0-100ng/ml) and erythroid burst-forming units (BFU-E) measured on day 14. Consistent with prior experiments, the percentage of BFU-E colonies was reduced by 60% at day 14 in the presence of 25 and 100ng/ml TNFα (p < 0.01, t-test), which was reversed to baseline with the addition of the neutralizing anti-TNFα antibody infliximab (p < 0.01, t-test). In order to determine whether the addition of resveratrol moderated TNFα-mediated erythroid colony suppression, human CD34+ cells were initially pre-incubated with increasing doses of resveratrol (0-50μM) for 72 hours and cultured in methylcellulose medium containing IL-3 (10ng/ml), EPO (1U/ml), and SCF (50ng/ml) in the presence or absence of 25ng/ml TNFα and varying concentrations of resveratrol (0-50μM) and BFU-E measured on day 14. We noted 50% recovery in BFU-E colony numbers at day 14 in CD34+ cells grown in the presence of 10μM resveratrol and TNFα in comparison with TNFα alone (p < 0.02, t-test). To better characterize TNFα-mediated pathways influenced by resveratrol, we analyzed in parallel CD34+ cells pre-incubated for 72 hours with 10μM resveratrol and grown in serum-free liquid culture containing similar concentrations of IL-3, SCF, and EPO in the presence or absence of 25ng/ml of TNFα and 10μM resveratrol. Total RNA and protein were isolated at multiple timepoints after the addition of TNFα on culture day 4 and TNFα-mediated gene and protein expression as well as hematopoietic microRNAs miR155 and miR451 expression analyzed by real-time RT-PCR and/or Western blot. We found that TNFα-treated CD34+ cells exhibited a 5-fold increased expression of miR155 without change in miR451 expression at 12 hours post TNFα treatment compared with control cells. However, CD34+ cells cultured in the presence of 10μM resveratrol and TNFα revealed downregulation of miR155 expression to baseline levels without change in miR451 expression (p < 0.01, t-test). Resveratrol treatment also resulted in 2-fold increased expression of GATA-1 without an appreciable change in GATA-2 (p < 0.01). Western analysis of whole cell lysates using antibodies recognizing known TNFα-associated signaling pathways revealed robust increases in phospho-p38MAPK, phospho-JNK, and phospho-ERK 1/2 in response to TNFα stimulation compared with control, with no inhibition of these signaling pathways noted in response to resveratrol treatment. We next examined TNFα-mediated NF-κB signaling in the presence or absence of resveratrol by Western analysis and found that the addition of resveratrol resulted in a 50% reduction in phospho-NF-κB levels in response to TNFα, with total NF-κB levels unchanged in all conditions. These data indicate that resveratrol significantly ameliorates TNFα-mediated erythroid colony suppression in human CD34+ cells via novel mechanisms that incorporate downregulation of both miR155 and NF-κB-associated signaling pathways, providing important evidence that resveratrol may be a potentially effective agent in the treatment of TNFα-mediated anemia and inflammation. Disclosures:No relevant conflicts of interest to declare.

Nociceptive and inflammatory responses induced by formalin in the orofacial region of rats: Effect of anti-TNFα strategies

This study evaluated the effects of different anti-TNFα strategies on the nociceptive and inflammatory responses triggered by formalin in the rat orofacial region. Formalin injection (2.5%) into the right upper lip caused a nociceptive response that was biphasic, with the first phase observed between 0 and 3 min and the second phase between 12 and 30 min. Plasma extravasation induced by formalin was time-related and reached the peak at 360 min. The monoclonal antibody anti-TNFα (25 and 50 pg/lip) significantly inhibited the second phase of formalin-induced nociceptive behavior, while the first phase remained unaltered. The systemic treatment with the chimeric anti-TNFα antibody infliximab also caused a significant inhibition of the second phase. Interestingly, the local administration of infliximab (50 pg/lip) produced a significant reduction of both phases of formalin-induced nociception. In addition, the systemic pretreatment with the preferential inhibitor of TNFα synthesis thalidomide (25 and 50 mg/kg, p.o) promoted a marked reduction of the first and second phases of formalin-evoked nociception. The local administration of the monoclonal antibody anti-TNFα (25 and 50 pg/lip) or infliximab (50 pg/lip) markedly reduced the plasma extravasation induced by formalin. Otherwise, formalin-elicited plasma extravasation was not significantly affected by the systemic administration of either infliximab (1 mg/kg; s.c) or thalidomide (50 mg/kg, p.o). Present data suggest that blocking TNFα effects, through different pharmacological tools, could represent a good alternative to control orofacial inflammatory pain that is refractory to other drugs.

Macrophage β3 Integrin Suppresses Hyperlipidemia-Induced Inflammation by Modulating TNFα Expression

High-fat, cholesterol-containing diets contribute to hyperlipidemia. Both high-fat diets and hyperlipidemia are associated with chronic inflammatory diseases like atherosclerosis. Integrins, heterodimeric mediators of inflammatory cell recruitment, are not generally thought to be affected by diet. However, high-fat feeding promotes inflammation, atherosclerosis, and death in hyperlipidemic mice with beta3 integrin deficiency, and treatment of humans from Western populations with oral beta3 integrin inhibitors increases mortality. The mechanisms responsible for these beta3 integrin-associated events are unknown. Here we show that diet-induced death in beta3 integrin-deficient mice is a TNFalpha-dependent process mediated by bone marrow-derived cells. In 2 different hyperlipidemic models, apoE-null and LDL receptor-null mice, beta3-replete animals transplanted with beta3-deficient marrow died with Western-type high-fat feeding whereas beta3-deficient animals transplanted with beta3-replete marrow were rescued from diet-induced death. Transplantation with beta3-deficient marrow also increased atherosclerosis. TNFalpha [corrected] expression was increased in beta3-deficient macrophages and normalized by either retroviral or adenoviral reconstitution of beta3 integrin expression. Treatment with the anti-TNFalpha antibody infliximab rescued beta3 integrin-deficient mice from Western diet-induced death, directly implicating TNFalpha in the pathophysiology triggered by diet-induced hyperlipidemia. These findings suggest that macrophage beta3 integrin, acting through TNFalpha, suppresses inflammation caused by hyperlipidemia attributable to high-fat feeding.