Human T-bet Governs Innate and Innate-like Adaptive IFN-γ Immunity against Mycobacteria.

Inborn errors of human interferon gamma (IFN-γ) immunity underlie mycobacterial disease. We report a patient with mycobacterial disease due to inherited deficiency of the transcription factor T-bet. The patient has extremely low counts of circulating Mycobacterium-reactive natural killer (NK), invariant NKT (iNKT), mucosal-associated invariant T (MAIT), and Vδ2+ γδ T lymphocytes, and of Mycobacterium-non reactive classic TH1 lymphocytes, with the residual populations of these cells also producing abnormally small amounts of IFN-γ. Other lymphocyte subsets develop normally but produce low levels of IFN-γ, with the exception of CD8+ αß T and non-classic CD4+ αß TH1∗ lymphocytes, which produce IFN-γ normally in response to mycobacterial antigens. Human T-bet deficiency thus underlies mycobacterial disease by preventing the development of innate (NK) and innate-like adaptive lymphocytes (iNKT, MAIT, and Vδ2+ γδ T cells) and IFN-γ production by them, with mycobacterium-specific, IFN-γ-producing, purely adaptive CD8+ αß T, and CD4+ αß TH1∗ cells unable to compensate for this deficit.

Human Adaptive Immunity Rescues an Inborn Error of Innate Immunity.

The molecular basis of the incomplete penetrance of monogenic disorders is unclear. We describe here eight related individuals with autosomal recessive TIRAP deficiency. Life-threatening staphylococcal disease occurred during childhood in the proband, but not in the other seven homozygotes. Responses to all Toll-like receptor 1/2 (TLR1/2), TLR2/6, and TLR4 agonists were impaired in the fibroblasts and leukocytes of all TIRAP-deficient individuals. However, the whole-blood response to the TLR2/6 agonist staphylococcal lipoteichoic acid (LTA) was abolished only in the index case individual, the only family member lacking LTA-specific antibodies (Abs). This defective response was reversed in the patient, but not in interleukin-1 receptor-associated kinase 4 (IRAK-4)-deficient individuals, by anti-LTA monoclonal antibody (mAb). Anti-LTA mAb also rescued the macrophage response in mice lacking TIRAP, but not TLR2 or MyD88. Thus, acquired anti-LTA Abs rescue TLR2-dependent immunity to staphylococcal LTA in individuals with inherited TIRAP deficiency, accounting for incomplete penetrance. Combined TIRAP and anti-LTA Ab deficiencies underlie staphylococcal disease in this patient.

An integrated multi-tissue approach for endometriosis candidate biomarkers: a systematic review.

Biomarker identification could help in deciphering endometriosis pathophysiology in addition to their use in the development of non invasive diagnostic and prognostic approaches, that are essential to greatly improve patient care. Despite extensive efforts, no single potential biomarker or combination has been clinically validated for endometriosis.Many studies have investigated endometriosis-associated biological markers in specific tissues, but an integrative approach across tissues is lacking. The aim of this review is to propose a comprehensive overview of identified biomarkers based on tissue or biological compartment, while taking into account endometriosis phenotypes (superficial, ovarian or deep, or rASRM stages), menstrual cycle phases, treatments and symptoms.We searched PubMed and Embase databases for articles matching the following criteria: 'endometriosis' present in the title and the associated term 'biomarkers' found as Medical Subject Headings (MeSH) terms or in all fields. We restricted to publications in English and on human populations. Relevant articles published between 01 January 2005 (when endometriosis phenotypes start to be described in papers) and 01 September 2022 were critically analysed and discussed.Four hundred forty seven articles on endometriosis biomarkers that included a control group without endometriosis and provided specific information on endometriosis phenotypes are included in this review. Presence of information or adjustment controlling for menstrual cycle phase, symptoms and treatments is highlighted, and the results are further summarized by biological compartment. The 9 biological compartments studied for endometriosis biomarker research are in order of frequency: peripheral blood, eutopic endometrium, peritoneal fluid, ovaries, urine, menstrual blood, saliva, feces and cervical mucus. Adjustments of results on disease phenotypes, cycle phases, treatments and symptoms are present in 70%, 29%, 3% and 6% of selected articles, respectively. A total of 1107 biomarkers were identified in these biological compartments. Of these, 74 were found in several biological compartments by at least two independent research teams and only 4 (TNF-a, MMP-9, TIMP-1 and miR-451) are detected in at least 3 tissues with cohorts of 30 women or more.Integrative analysis is a crucial step to highlight potential pitfalls behind the lack of success in the search for clinically relevant endometriosis biomarkers, and to illuminate the physiopathology of this disease.

PPARγ partial agonist LPSF/GQ-16 prevents dermal and pulmonary fibrosis in HOCl-induced systemic sclerosis (SSc) and modulates cytokine production in PBMC of SSc patients.

Thiazolidinediones (TZD) are synthetic molecules that have a range of biological effects, including antifibrotic and anti-inflammatory, and they may represent a promising therapeutic strategy for systemic sclerosis (SSc). The aim of this study was to investigate the immunomodulatory and antifibrotic properties of LPSF/GQ-16, a TZD derivative, in peripheral blood mononuclear cells (PBMC) from SSc patients and in a murine model of SSc HOCl-induced. The PBMC of 20 SSc patients were stimulated with phytohemagglutinin (PHA) and treated with LPSF/GQ-16 for 48 h, later cytokines in the culture supernatants were quantified by sandwich enzyme-linked immunosorbent assay (ELISA) or cytometric bead array (CBA). Experimental SSc was induced by intradermal injections of hypochlorous acid (HOCl) for 6 weeks. HOCl-induced SSc mice received daily treatment with LPSF/GQ-16 (30 mg/kg) through intraperitoneal injections during the same period. Immunological parameters were evaluated by flow cytometry and ELISA, and dermal and pulmonary fibrosis were evaluated by RT-qPCR, hydroxyproline dosage and histopathological analysis. In PBMC cultures, it was possible to observe that LPSF/GQ-16 modulated the secretion of cytokines IL-2 (p < 0.001), IL-4 (p < 0.001), IL-6 (p < 0.001), IL-17A (p = 0.006), TNF (p < 0.001) and IFN-γ (p < 0.001). In addition, treatment with LPSF/GQ-16 in HOCl-induced SSc mice promoted a significant reduction in dermal thickening (p < 0.001), in the accumulation of collagen in the skin (p < 0.001), down-regulated the expression of fibrosis markers in the skin (Col1a1, α-Sma and Tgfß1, p < 0.001 for all) and lungs (Il4 and Il13, p < 0.001 for both), as well as reduced activation of CD4 + T cells (p < 0.001), B cells (p < 0.001) and M2 macrophages (p < 0.001). In conclusion, LPSF/GQ-16 showed immunomodulatory and antifibrotic properties, demonstrating the therapeutic potential of this molecule for SSc.

High Frequency of Specific Polysaccharide Antibody Deficiency in Adults With Unexplained, Recurrent and/or Severe Infections With Encapsulated Bacteria.

BACKGROUND: Primary immunodeficiencies (PIDs) in adults are mainly revealed by recurrent and/or severe bacterial infections. The objective of this study was to evaluate a systematic research strategy of PIDs in adults with unexplained bacterial infections, with a special focus on specific polysaccharide antibody deficiency (SPAD). METHODS: In this prospective multicenter study, inclusion criteria were recurrent benign upper and lower respiratory tract infections (RTIs) for at least two years (group 1), at least one upper or lower RTI requiring hospitalization (group 2), and/or at least one invasive infection documented with encapsulated bacteria (group 3). Main exclusion criteria were all local and general conditions that could explain infections. If no PID diagnosis was made, response to polysaccharide antigens was assessed using a pneumococcal polysaccharide vaccine. RESULTS: From March 2015 to March 2020, 118 patients were included (37 males, median age of 41 years): 73, 17, and 28 in groups 1, 2, and 3, respectively. Forty-seven PIDs were diagnosed, giving an estimated frequency of 39.8% (95% confidence interval [CI] [30.4, 48.8]). SPAD was the most frequent diagnosis by far (n = 37/47, 78.7%), and was made in 23, 5, and 9 patients from groups 1 to 3, respectively. All SPAD patients received conjugate vaccines and, according to their infectious history, were on surveillance or treated with preventive antibiotics (n = 6) and/or with immunoglobulins replacement therapy (n = 10), the latter being dramatically efficient in all cases. CONCLUSIONS: Considering its high prevalence among adults with unexplained recurrent and/or severe bacterial infections, SPAD should be screened in those patients. CLINICAL TRIALS REGISTRATION: NCT02972281.

Second-trimester amniotic fluid proteins changes in subsequent spontaneous preterm birth.

INTRODUCTION: The global sequence of the pathogenesis of preterm labor remains unclear. This study aimed to compare amniotic fluid concentrations of extracellular matrix-related proteins (procollagen, osteopontin and IL-33), and of cytokines (IL-19, IL-6, IL-20, TNFα, TGFß, and IL-1ß) in asymptomatic women with and without subsequent spontaneous preterm delivery. MATERIAL AND METHODS: We used amniotic fluid samples of singleton pregnancy, collected by amniocentesis between 16 and 20 weeks' gestation, without stigmata of infection (i.e., all amniotic fluid samples were tested with broad-range 16 S rDNA PCR to distinguish samples with evidence of past bacterial infection from sterile ones), during a randomized, double-blind, placebo-controlled trial to perform a nested case-control laboratory study. Cases were women with a spontaneous delivery before 37 weeks of gestation (preterm group). Controls were women who gave birth at or after 39 weeks (full term group). Amniotic fluid concentrations of the extracellular matrix-related proteins and cytokines measured by immunoassays were compared for two study groups. CLINICALTRIALS: gov: NCT00718705. RESULTS: Between July 2008 and July 2011, in 12 maternal-fetal medicine centers in France, 166 women with available PCR-negative amniotic fluid samples were retained for the analysis. Concentrations of procollagen, osteopontin, IL-19, IL-6, IL-20, IL-33, TNFα, TGFß, and IL-1ß were compared between the 37 who gave birth preterm and the 129 women with full-term delivery. Amniotic fluid levels of procollagen, osteopontin, IL-19, IL-33, and TNFα were significantly higher in the preterm than the full-term group. IL-6, IL-20, TGFß, and IL-1ß levels did not differ between the groups. CONCLUSIONS: In amniotic fluid 16 S rDNA PCR negative samples obtained during second-trimester amniocentesis, extracellular matrix-related protein concentrations (procollagen, osteopontin and IL-33), together with IL-19 and TNFα, were observed higher at this time in cases of later spontaneous preterm birth.

Role of thyroid dysimmunity and thyroid hormones in endometriosis.

Endometriosis is characterized by the presence of ectopic endometrial cells outside the uterine cavity. Thyroid autoimmunity has been associated with endometriosis. This work investigated the potential pathophysiological link between endometriosis and thyroid disorders. Transcripts and proteins involved in thyroid metabolism are dysregulated in eutopic and ectopic endometrium of endometriotic patients, leading to resistance of ectopic endometrium to triiodothyronine (T3) action and local accumulation of thyroxine (T4). Thyroid-stimulating hormone (TSH) acts as a proliferative and prooxidative hormone on all endometria of endometriosis patients and controls, whereas T3 and T4 act to specifically increase ectopic endometrial cell proliferation and reactive oxygen species (ROS) production. Mouse studies confirmed the data gained in vitro since endometriotic implants were found to be bigger when thyroid hormones increased. A retrospective analysis of endometriosis patients with or without a thyroid disorder revealed an increased chronic pelvic pain and disease score in endometriotic patients with a thyroid disorder.

A New Manganese Superoxide Dismutase Mimetic Improves Oxaliplatin-Induced Neuropathy and Global Tolerance in Mice.

Reactive oxygen species (ROS) are produced by every aerobic cell during mitochondrial oxidative metabolism as well as in cellular response to xenobiotics, cytokines, and bacterial invasion. Superoxide Dismutases (SOD) are antioxidant proteins that convert superoxide anions (O2•-) to hydrogen peroxide (H2O2) and dioxygen. Using the differential in the level of oxidative stress between normal and cancer cells, SOD mimetics can show an antitumoral effect and prevent oxaliplatin-induced peripheral neuropathy. New Pt(IV) conjugate prodrugs (OxPt-x-Mn1C1A (x = 1, 1-OH, 2)), combining oxaliplatin and a Mn SOD mimic (MnSODm Mn1C1A) with a covalent link, were designed. Their stability in buffer and in the presence of sodium ascorbate was studied. In vitro, their antitumoral activity was assessed by the viability and ROS production of tumor cell lines (CT16, HCT 116, KC) and fibroblasts (primary culture and NIH 3T3). In vivo, a murine model of colorectal cancer was created with subcutaneous injection of CT26 cells in Balb/c mice. Tumor size and volume were measured weekly in four groups: vehicle, oxaliplatin, and oxaliplatin associated with MnSODm Mn1C1A and the bis-conjugate OxPt-2-Mn1C1A. Oxaliplatin-induced peripheral neuropathy (OIPN) was assessed using a Von Frey test reflecting chronic hypoalgesia. Tolerance to treatment was assessed with a clinical score including four items: weight loss, weariness, alopecia, and diarrhea. In vitro, Mn1C1A associated with oxaliplatin and Pt(IV) conjugates treatment induced significantly higher production of H2O2 in all cell lines and showed a significant improvement of the antitumoral efficacy compared to oxaliplatin alone. In vivo, the association of Mn1C1A to oxaliplatin did not decrease its antitumoral activity, while OxPt-2-Mn1C1A had lower antitumoral activity than oxaliplatin alone. Mn1C1A associated with oxaliplatin significantly decreased OIPN and also improved global clinical tolerance of oxaliplatin. A neuroprotective effect was observed, associated with a significantly improved tolerance to oxaliplatin without impairing its antitumoral activity.

Long-term exposure to monoclonal anti-TNF is associated with an increased risk of lymphoma in BAFF-transgenic mice.

The impact of treatment on the risk of lymphoma in patients with rheumatoid arthritis (RA) is unclear. Here, we aimed to assess if the risk of lymphoma differs according to the type of tumor necrosis factor inhibitor (TNFi), comparing monoclonal anti-TNF antibodies to the soluble TNF receptor. We used B cell activating factor belonging to the TNF family (BAFF)-transgenic (Tg) mice as a model of autoimmunity-associated lymphoma. Six-month-old BAFF-Tg mice were treated with TNFi for 12 months. Histological examination of the spleen, assessment of the cellular composition of the spleen by flow cytometry and assessment of B cell clonality were performed at euthanasia. Crude mortality and incidence of lymphoma were significantly higher in mice treated with monoclonal anti-TNF antibodies compared to both controls and mice treated with the soluble TNF receptor, even at a high dose. Flow cytometry analysis revealed decreased splenic macrophage infiltration in mice treated with monoclonal anti-TNF antibodies. Overall, this study demonstrates, for the first time, that a very prolonged treatment with monoclonal anti-TNF antibodies increase the risk of lymphoma in B cell-driven autoimmunity. These data suggest a closer monitoring for lymphoma development in patients suffering from B cell-driven autoimmune disease with long-term exposure to monoclonal anti-TNF antibodies.

TGFß promotes low IL10-producing ILC2 with profibrotic ability involved in skin fibrosis in systemic sclerosis.

OBJECTIVE: Innate lymphoid cells-2 (ILC2) were shown to be involved in the development of lung or hepatic fibrosis. We sought to explore the functional and phenotypic heterogeneity of ILC2 in skin fibrosis within systemic sclerosis (SSc). METHODS: Blood samples and skin biopsies from healthy donor or patients with SSc were analysed by immunostaining techniques. The fibrotic role of sorted ILC2 was studied in vitro on dermal fibroblast and further explored by transcriptomic approach. Finally, the efficacy of a new treatment against fibrosis was assessed with a mouse model of SSc. RESULTS: We found that ILC2 numbers were increased in the skin of patients with SSc and correlated with the extent of skin fibrosis. In SSc skin, KLRG1- ILC2 (natural ILC2) were dominating over KLRG1+ ILC2 (inflammatory ILC2). The cytokine transforming growth factor-ß (TGFß), whose activity is increased in SSc, favoured the expansion of KLRG1- ILC2 simultaneously decreasing their production of interleukin 10 (IL10), which regulates negatively collagen production by dermal fibroblasts. TGFß-stimulated ILC2 also increased myofibroblast differentiation. Thus, human KLRG1- ILC2 had an enhanced profibrotic activity. In a mouse model of SSc, therapeutic intervention-combining pirfenidone with the administration of IL10 was required to reduce the numbers of skin infiltrating ILC2, enhancing their expression of KLRG1 and strongly alleviating skin fibrosis. CONCLUSION: Our results demonstrate a novel role for natural ILC2 and highlight their inter-relationships with TGFß and IL10 in the development of skin fibrosis, thereby opening up new therapeutic approaches in SSc.

Efficacy, safety and immunological profile of combining rituximab with belimumab for adults with persistent or chronic immune thrombocytopenia: results from a prospective phase 2b trial.

B-cell activating factor may be involved in the failure of B-cell depleting therapy with rituximab in immune thrombocytopenia (ITP) by promoting the emergence of splenic long-lived plasma cells. From results obtained in mouse models, we hypothesized that combining rituximab with sequential injections of belimumab could increase the rate of response at one year in patients with persistent or chronic ITP by preventing the emergence of these long-lived plasma cells. The study was a single-center, single arm, prospective phase 2b trial (RITUX-PLUS, NCT03154385) investigating the safety and efficacy of rituximab given at a fixed dose of 1,000 mg, two weeks apart, combined with five infusions of belimumab, 10 mg/kg at week 0 (W0)+2 days, W2+2 days, W4, W8 and W12 for adults with primary persistent or chronic ITP. The primary endpoint was the total number of patients achieving an overall response (complete response + response) at W52 according to a standard definition. In total, 15 non-splenectomized adults, nine (60%) with persistent IPT and six (40%) with chronic ITP, were included. No severe adverse event, infection, or severe hypogammaglobulinemia was observed. Thirteen patients achieved an initial overall response. At W52, 12 (80%) patients achieved an overall response, including ten (66.7%) with complete response. When compared with a cohort of patients receiving rituximab alone, the kinetics of B-cell repopulation appeared similar, but the number of circulating T follicular helper cells was significantly decreased with belimumab combination therapy. Combining rituximab and belimumab seems a promising strategy in ITP, with high efficacy and acceptable safety.

Doctor's aptitude for switching from innovator etanercept to biosimilar etanercept in inflammatory rheumatic diseases: experience from a single French rheumatology tertiary care center.

OBJECTIVES: To describe the switch to biosimilar etanercept (bETN), evaluate factors associated with this switch, and evaluate the efficacy of this switch in a real-life setting METHODS: We included patients, from October 2016 to April 2017, with rheumatoid arthritis (RA) and spondyloarthritis (SpA) who received innovator ETN (iETN) for at least 6 months. After receiving information on biosimilars, all physicians were invited to propose a switch from iETN to bETN. Factors associated with bETN discontinuation were explored by univariate and multivariate analyses. We estimated the proportion of patients still on bETN over time by Kaplan-Meier survival analysis. We assessed serum trough concentrations of iETN and bETN and anti-drug antibodies to ETN. RESULTS: Overall, 183 outpatients were eligible for a potential switch; 94 (51.6%) switched from iETN to bETN. The probability of a switch was greater with an older than younger aged physician (mean [SD] age 50.4 [14.3] with a switch vs 44.8 [11.3] with no switch, p = 0.005) and the physician having a full-time academic position than other position (56.4% with a switch vs 13.5% with no switch, p < 0.001). After a 6-month follow-up, bETN retention rate was 83% (95% CI: 0.76-0.92). The first cause of bETN discontinuation was inefficacy (50%). On multivariate analysis, no factor was independently associated with a bETN switch or discontinuation. Drug trough levels did not significantly differ by discontinuation or continuation of bETN. No patient showed anti-drug antibodies. CONCLUSION: The probability of switching from iETN to bETN was likely related to physician characteristics.

Prevention of esophageal stricture after circumferential endoscopic submucosal dissection using a modified self-assembling peptide.

Circumferential endoscopic resection (ER) of the esophageal mucosa could find its place in the treatment of dysplastic Barrett's esophagus or extensive squamous cell neoplasia. However, the occurrence of esophageal strictures remains a major complication after ER exceeding 75% of the circumference. The aim of this study was to assess the effect of a modified, pH = 2, self-assembling peptide matrix (4[Arg-Ala-Asp-Ala]) (SAP) on the development of esophageal stricture after circumferential ER in a swine model. We performed a circumferential ER in 35 swine under general anesthesia. Five animals were included in the control group, 11 animals received the SAP matrix immediately after the resection, and 11 received the SAP matrix associated to a local steroid immediately after the resection. Follow-up endoscopy and esophagogram were performed before slaughter and necropsy at day 14. Eight treated animals were kept alive until day 28. At day 14, 27% of the animals in the SAP group developed a symptomatic stricture versus 100% in the control group (P = 0.008) and 50% in the SAP-triamcinolone group (P = 0.11). Application of an SAP matrix after circumferential ER in the swine allowed a significant reduction of the incidence of symptomatic stricture at day 14. Adding triamcinolone brought no significant improvement.

Endometriosis phenotypes are associated with specific serum metabolic profiles determined by proton-nuclear magnetic resonance.

RESEARCH QUESTION: What is the correlation between serum metabolic profile and endometriosis phenotype? DESIGN: A pilot study nestled in a prospective cohort study at a university hospital, including 46 patients with painful endometriosis who underwent surgery and 21 controls who did not have macroscopic endometriotic lesions. Endometriosis was strictly classified into two groups of 23 patients each: endometrioma (OMA) and deep infiltrating endometriosis (DIE). Serum samples were collected before surgery for metabolomic profiling based on proton-nuclear magnetic resonance spectroscopy in combination with statistical approaches. Comparative identification of the metabolites in the serum from endometriosis patients and from controls was carried out, including an analysis according to endometriosis phenotype. RESULTS: The serum metabolic profiles of the endometriosis patients revealed significantly lower concentrations of several amino acids compared with the controls, whereas the concentrations of free fatty acids and ketone bodies were significantly higher. The OMA and the DIE phenotypes each had a specific metabolic profile, with higher concentrations of two ketone bodies in the OMA group, and higher concentrations of free fatty acids and lipids in the DIE group. CONCLUSION: Proton-nuclear magnetic resonance-based metabolomics of serum samples were found to have ample potential for identifying metabolic changes associated with endometriosis phenotypes. This information may improve our understanding of the pathogenesis of endometriosis.

The follicular fluid metabolome differs according to the endometriosis phenotype.

RESEARCH QUESTION: Is there a follicular fluid-specific metabolic profile in deep infiltrating endometriosis (DIE) depending on the presence of an associated ovarian endometrioma (OMA) that could lead to the identification of biomarkers for diagnosis and prognosis of the disease? DESIGN: In this prospective cohort study, proton nuclear magnetic resonance (1H-NMR) experiments were carried out on 50 follicular fluid samples from patients presenting with DIE, associated or not associated with an OMA, and 29 follicular fluid samples from patients with infertility caused by a tubal obstruction. RESULTS: Concentrations of glucose, citrate, creatine and amino acids such as tyrosine and alanine were lower in women with DIE than control participants, whereas concentrations of lactate, pyruvate, lipids and ketone bodies were higher. Metabolic analysis revealed enhanced concentrations of glycerol and ketone bodies in patients with OMA, indicative of an activation of lipolysis followed by beta-oxidation. Concentrations of lactate and pyruvate were increased in patients without OMA, whereas the concentration of glucose was decreased, highlighting activation of the anaerobic glycolysis pathway. Differences in concentrations of amino acids such as threonine and glutamine were also statistically relevant in discriminating between the presence or absence of OMA. CONCLUSIONS: Results indicate a mitochondrial dysregulation in endometriosis phenotypes, with a modified balance between anaerobic glycolysis and beta-oxidation in OMA phenotypes that could affect the fertility of women with endometriosis. As the composition of the follicular fluid has been shown to be correlated with oocyte development and outcome of implantation after fertilization, these findings may help explain the high level of infertility in these patients.

Identification of TP53 mutated group using a molecular and immunohistochemical classification of endometrial carcinoma to improve prognostic evaluation for adjuvant treatments.

INTRODUCTION: Molecular classification of endometrial carcinoma has been proposed to predict survival. However, its role in patient management remains to be determined. We aimed to identify whether a molecular and immunohistochemical classification of endometrial carcinoma could improve decision-making for adjuvant therapy. METHODS: All consecutive patients treated for endometrial carcinoma between 2010 and 2017 at Cochin University Hospital were included. Clinical risk of relapse was based on European Society for Medical Oncology-European Society of Gynaecological Oncology-European SocieTy for Radiotherapy & Oncology (ESMO-ESGO-ESTRO) consensus. The clinical event of interest was event-free survival. Formalin-fixed paraffin-embedded tissue samples were processed for histopathological analysis and DNA extraction. The nuclear expression of mismatch repair and TP53 proteins was analyzed by immunohistochemistry. Next-generation sequencing of a panel of 15 genes including TP53 and POLE was performed using Ampliseq panels on Ion Torrent PGM (ThermoFisher). Tumors were allocated into four molecular groups using a sequential method based on next-generation sequencing and immunohistochemistry data: (1) POLE/ultramutated-like; (2) MSI/hypermutated-like (mismatch repair-deficient); (3) TP53-mutated (without POLE mutations or mismatch repair deficiency); (4) not otherwise specified (the remaining tumors). RESULTS: 159 patients were included; 125 tumors were available for molecular characterization and distributed as follows: (1) POLE/ultramutated-like: n=4 (3%); (2) MSI/hypermutated-like: n=35 (30%); (3) TP53-mutated: n=30 (25%); and (4) not otherwise specified: n=49 (42%). Assessing the TP53 status by immunohistochemistry only rather than next-generation sequencing would have misclassified 6 tumors (5%). TP53-mutated tumors were associated with poor prognosis, independently of International Federation of Gynecology and Obstetrics (FIGO) stage and histological grade (Cox-based adjusted hazard ratio (aHR) 5.54, 95% CI 2.30 to 13.4), and independently of clinical risk of relapse (aHR 3.92, 95% CI 1.59 to 9.64). Among patients with FIGO stage I-II tumors, 6 (38%) TP53-mutated tumors had low/intermediate clinical risk of relapse and did not receive adjuvant chemotherapy or radiotherapy. CONCLUSION: Endometrial carcinoma molecular classification identified potentially under-treated patients with poor molecular prognosis despite being at low/intermediate clinical risk of relapse. Consideration of molecular classification in adjuvant therapeutic decisions should be evaluated in prospective trials.

Dimethyl fumarate, a two-edged drug: Current status and future directions.

Dimethyl fumarate (DMF) is a fumaric acid ester registered for the treatment of relapsing-remitting multiple sclerosis (RRMS). It induces protein succination leading to inactivation of cysteine-rich proteins. It was first shown to possess cytoprotective and antioxidant effects in noncancer models, which appeared related to the induction of the nuclear factor erythroid 2 (NF-E2)-related factor 2 (NRF2) pathway. DMF also displays antitumor activity in several cellular and mice models. Recently, we showed that the anticancer mechanism of DMF is dose-dependent and is paradoxically related to the decrease in the nuclear translocation of NRF2. Some other studies performed indicate also the potential role of DMF in cancers, which are dependent on the NRF2 antioxidant and cellular detoxification program, such as KRAS-mutated lung adenocarcinoma. It, however, seems that DMF has multiple biological effects as it has been shown to also inhibit the transcription factor nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), thus blocking downstream targets that may be involved in the development and progression of inflammatory cascades leading to various disease processes, including tumors, lymphomas, diabetic retinopathy, arthritis, and psoriasis. Herein, we present the current status and future directions of the use of DMF in various diseases models with particular emphases on its targeting of specific intracellular signal transduction cascades in cancer; to shed some light on its possible mode of action.

Reduced α-2,6 sialylation regulates cell migration in endometriosis.

STUDY QUESTION: Is endometriosis associated with aberrant sialylation patterns and what is the potential impact of such anomalies on cell migratory properties? SUMMARY ANSWER: The reduced α-2,6 sialylation patterns in the peritoneal fluid of endometriosis-affected women and in stromal and epithelial cells from endometriotic lesions could be associated with enhanced cell migration. WHAT IS KNOWN ALREADY: Endometriosis is considered to be a benign disease although, like cancer, it has the characteristic of being an invasive disease with cells that have an enhanced capacity to migrate. Aberrant sialylation has been reported in various malignancies and it has been linked to tumour invasion and metastasis. STUDY DESIGN, SIZE, DURATION: We conducted a prospective laboratory study in a tertiary-care university hospital. We investigated non-pregnant patients who were <42 years of age (n = 273) when they underwent surgery for a benign gynaecological condition. PARTICIPANTS/MATERIALS, SETTING, METHODS: The study population consisted of 102 women with histologically proven endometriosis and 71 endometriosis-free controls, who underwent a complete surgical exploration of the abdominopelvic cavity. Peritoneal fluids were collected during the surgical procedures, and endometrial and endometriotic biopsies were performed on all of the patients to generate stromal and epithelial primary cell cultures. The expression of α-2,6-sialyltransferase (ST6GALNAC1) was studied in eutopic and ectopic endometria of endometriosis patients and in eutopic endometria of controls by reverse transcription followed by quantitative real-time polymerase chain reaction (RT-qPCR). The α-2,6 sialylation levels were measured by ELISA in the peritoneal fluids of patients and controls and by western-blot in primary endometrial and endometriotic cell cultures using Sambucus nigra agglutinin (SNA), an α-2,6 sialic acid-binding lectin. A transwell migration assay after incubation of the cells with neuraminidase was also performed to evaluate the impact of desialylation on eutopic endometrial stromal cell migration. MAIN RESULTS AND THE ROLE OF CHANCE: ST6GALNAC1 gene expression was significantly lower in endometriotic lesions compared to that in eutopic endometrium of endometriosis-affected patients and healthy endometrium (16-fold for both; P < 0.01). We observed a significant reduction in SNA levels in the peritoneal fluids of endometriosis-affected women compared to control women (median optic density (OD), 0.257; range, 0.215-0.279 versus median OD, 0.278; range 0.238-0.285; P < 0.01), as well as in stromal (mean OD, 705 907; standard error of the mean (SEM), 141 549 versus mean OD, 1.16 × 106; SEM, 107,271; P < 0.05) and epithelial (mean OD, 485 706; SEM, 179 681 versus mean OD, 1.25 × 106; SEM, 232 120; P < 0.05) ectopic endometriotic cells compared to control eutopic cells, indicating reduced α-2,6 sialylation. Finally, in the transwell migration assay, the eutopic endometrial cells of endometriosis patients migrated significantly more into the lower chamber after incubation with neuraminidase, indicating enhanced migration by these cells after desialylation. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: Our control group involved patients operated for benign gynaecological conditions (e.g. tubal infertility, uterine fibroids or ovarian cysts) which may also be associated with altered sialylation patterns. WIDER IMPLICATIONS OF THE FINDINGS: The hyposialylation pattern of endometriotic cells appeared to be associated with enhanced migratory abilities, which might contribute to the establishment of early endometriotic implants. Further research is needed to confirm these findings, as this could lead to new potential therapeutic targets for this complex disorder. STUDY FUNDING AND COMPETING INTEREST(S): No external funding was received and there are no conflicts of interest.

Impact of MPO-ANCA-mediated oxidative imbalance on renal vasculitis.

Glomerulonephritis is a severe complication of microscopic polyangiitis (MPA), a small-vessel vasculitis associated with anti-myeloperoxidase antibodies (MPO-ANCA). We previously showed the pathogenic effects of MPO-ANCA that activate MPO to trigger an oxidative burst mainly through HOCl production, contributing to endothelial injury and lung fibrosis. The aim of this study was to investigate the relationship between MPO-induced oxidative stress, anti-oxidant defenses and renal histological lesions in MPA patients. We therefore analyzed histological data from a prospective cohort of ANCA-associated glomerulonephritis. Serum-mediated HOCl production, advanced oxidation protein products (AOPP), and thiol concentration in sera were determined. From 38 patients included, histological classification noted 50% focal glomerulonephritis, 15.8% crescentic-glomerulonephritis, and 34.2% mixed-glomerulonephritis. MPA patients' sera displayed higher HOCl production by MPO ( P < 0.001), higher AOPP ( P < 0.001) and thiol ( P < 0.01) levels, compared with healthy subjects. The presence of cellular crescents was associated with higher serum-mediated HOCl production ( P = 0.049) and lower thiol levels ( P = 0.022) at disease onset. Higher thiol concentrations were associated with focal glomerulonephritis ( P = 0.042), less interstitial fibrosis ( P = 0.039) and hyalinosis ( P = 0.066). In remission, HOCl production was decreased ( P < 0.01), and thiol concentration remained high ( P = 0.39). Our findings suggest that HOCl production by activated MPO could contribute to the very early stage of glomerulonephritis, whereas thiol may exert a protective effect against the development of renal vasculitis and glomerulosclerosis. This study highlights the importance of oxidative defenses to counteract the process of MPO-ANCA associated glomerulonephritis.

Niclosamide Prevents Systemic Sclerosis in a Reactive Oxygen Species-Induced Mouse Model.

Systemic sclerosis (SSc) is a connective tissue disorder characterized by fibrosis of the skin and inner organs, vasculopathy, and immunological abnormalities. Recent insights on the implication of STAT3, AKT, and Wnt/ß-catenin in fibrosis have prompted us to investigate, in a mouse model of ROS-induced SSc, the effects of niclosamide, an antihelmintic drug that inhibits both of these signaling pathways. SSc was induced in BALB/c mice by daily s.c. injections of hypochlorous acid (HOCl). Mice were treated or not every other day, 5 d a week, for 6 wk, by niclosamide. Skin and lung fibrosis as well as immunological features were studied. Mice exposed to HOCl developed a diffuse cutaneous SSc with pulmonary fibrosis and anti-DNA topoisomerase 1 autoantibodies. STAT3, AKT, and Wnt/ß-catenin pathways were hyperactivated in the skin and the lungs of diseased mice. Niclosamide reversed fibrosis of the skin and the lungs. Beneficial immunological effects were also observed because niclosamide decreased the activation of CD4+ and CD8+ T cells, autoimmune B cell activation, as well as IL-4 and IL-13 production in the skin. The improvement permitted by niclosamide in the mouse model of HOCl-induced SSc as well as the well-documented safety profile of this drug provide a rationale for the evaluation of niclosamide in the management of patients affected by this disease.