Effectiveness of swapping to ustekinumab after vedolizumab failure in patients with multi-refractory Crohn's disease.

BACKGROUND: Ustekinumab (UST) and vedolizumab (VDZ) are biologic therapies for moderate-to-severe Crohn's disease (CD) in patients who failed or had contraindication to anti-TNF treatment. AIMS: To evaluate ustekinumab efficacy as third-line treatment after swapping from VDZ for failure. METHODS: We conducted a monocentric, retrospective, observational study where CD patients were followed for 12 months from the beginning of UST therapy. We assessed clinical activity (HBI) and laboratory markers (CRP) at the initiation of UST therapy (T0) and after 2(T2), 6(T6) and 12(T12) months. Endoscopic activity was recorded at T0 and T12. We registered data regarding their clinical history and previous biologic treatments. Steroid-free clinical remission was defined as HBI ≤ 4 without need for steroids. Clinical response was defined as HBI reduction of at least three points or the suspension of steroids. RESULTS: 27 CD patients treated with UST after VDZ failure had a minimum follow up of 12 months and were included. All patients had previously been treated with anti-TNF agents. After 12 months, steroid-free clinical remission was evident in 15 (55.5%) patients, 5 (18.5%) had clinical response, while 7 (26%) had suspended for failure or persisted on treatment after optimization. CONCLUSIONS: Ustekinumab should be considered as third-line biologic treatment in multi-refractory CD patients.

The identification of farnesoid X receptor modulators as treatment options for nonalcoholic fatty liver disease.

INTRODUCTION: The farnesoid-x-receptor (FXR) is a ubiquitously expressed nuclear receptor selectively activated by primary bile acids. AREA COVERED: FXR is a validated pharmacological target. Herein, the authors review preclinical and clinical data supporting the development of FXR agonists in the treatment of nonalcoholic fatty liver disease. EXPERT OPINION: Development of systemic FXR agonists to treat the metabolic liver disease has been proven challenging because the side effects associated with these agents including increased levels of cholesterol and LDL-c and reduced HDL-c raising concerns over their long-term cardiovascular safety. Additionally, pruritus has emerged as a common, although poorly explained, dose-related side effect with all FXR ligands, but is especially common with OCA. FXR agonists that are currently undergoing phase 2/3 trials are cilofexor, tropifexor, nidufexor and MET409. Some of these agents are currently being developed as combination therapies with other agents including cenicriviroc, a CCR2/CCR5 inhibitor, or firsocostat an acetyl CoA carboxylase inhibitor. Additional investigations are needed to evaluate the beneficial effects of combination of these agents with statins. It is expected that in the coming years, FXR agonists will be developed as a combination therapy to minimize side effects and increase likelihood of success by targeting different metabolic pathways.

Bile Acid Signaling in Inflammatory Bowel Diseases.

Bile acids are a group of chemically different steroids generated at the host/microbial interface. Indeed, while primary bile acids are the end-product of cholesterol breakdown in the host liver, secondary bile acids are the products of microbial metabolism. Primary and secondary bile acids along with their oxo derivatives have been identified as signaling molecules acting on a family of cell membrane and nuclear receptors collectively known as "bile acid-activated receptors." Members of this group of receptors are highly expressed throughout the gastrointestinal tract and mediate the bilateral communications of the intestinal microbiota with the host immune system. The expression and function of bile acid-activated receptors FXR, GPBAR1, PXR, VDR, and RORγt are highly dependent on the structure of the intestinal microbiota and negatively regulated by intestinal inflammation. Studies from gene ablated mice have demonstrated that FXR and GPBAR1 are essential to maintain a tolerogenic phenotype in the intestine, and their ablation promotes the polarization of intestinal T cells and macrophages toward a pro-inflammatory phenotype. RORγt inhibition by oxo-bile acids is essential to constrain Th17 polarization of intestinal lymphocytes. Gene-wide association studies and functional characterizations suggest a potential role for impaired bile acid signaling in development inflammatory bowel diseases (IBD). In this review, we will focus on how bile acids and their receptors mediate communications of intestinal microbiota with the intestinal immune system, describing dynamic changes of bile acid metabolism in IBD and the potential therapeutic application of targeting bile acid signaling in these disorders.

Bile acid-activated receptors and the regulation of macrophages function in metabolic disorders.

Bile acids are produced in the liver by the cholesterol breakdown and further metabolized by the intestinal microbiota to generate a group of chemically heterogeneous steroids that bind and activate a family of cells surface and nuclear receptors, collectively known as the bile acid-activated receptors (BARs). The two best characterized members of this family are the farnesoid-x-receptor (FXR) and G protein Bile Acid Receptor (GPBAR1). Both receptors are expressed by cells of innate immunity including liver-resident and intestinal-resident macrophages and monocytes-derived macrophages. Because FXR and GPBAR1 knockout mice are biased toward a pro-inflammatory phenotype, it appears the both receptors might have a role in the development and maintenance of a tolerogenic phenotype. FXR and GPBAR1 ligands have been proven effective in the treatment in inflammatory and metabolic disorders and ligands for these receptors are currently under development for the treatment of non-alcoholic steato-hepatitis and diabetes.

Opposite effects of the FXR agonist obeticholic acid on Mafg and Nrf2 mediate the development of acute liver injury in rodent models of cholestasis.

The farnesoid-X-receptor (FXR) is validated target in the cholestatic disorders treatment. Obeticholic acid (OCA), the first in class of FXR agonist approved for clinical use, causes side effects including acute liver decompensation when administered to cirrhotic patients with primary biliary cholangitis at higher than recommended doses. The V-Maf avian-musculoaponeurotic-fibrosarcoma-oncogene-homolog-G (Mafg) and nuclear factor-erythroid-2-related-factor-2 (Nrf2) mediates some of the downstream effects of FXR. In the present study we have investigated the role of FXR/MafG/NRF2 pathway in the development of liver toxicity caused by OCA in rodent models of cholestasis. Cholestasis was induced by bile duct ligation (BDL) or administration of α-naphtyl-isothiocyanate (ANIT) to male Wistar rats and FXR-/- and FXR+/+ mice. Treating BDL and ANIT rats with OCA exacerbated the severity of cholestasis, hepatocytes injury and severely downregulated the expression of basolateral transporters. In mice, genetic ablation FXR or its pharmacological inhibition by 3-(naphthalen-2-yl)-5-(piperidin-4-yl)-1,2,4-oxadiazole rescued from negative regulation of MRP4 and protected against liver injury caused by ANIT. By RNAseq analysis we found that FXR antagonism effectively reversed the transcription of over 2100 genes modulated by OCA/ANIT treatment, including Mafg and Nrf2 and their target genes Cyp7a1, Cyp8b1, Mat1a, Mat2a, Gss. Genetic and pharmacological Mafg inhibition by liver delivery of siRNA antisense or S-adenosylmethionine effectively rescued from damage caused by ANIT/OCA. In contrast, Nrf2 induction by sulforaphane was protective. CONCLUSIONS: Liver injury caused by FXR agonism in cholestasis is FXR-dependent and is reversed by FXR and Mafg antagonism or Nrf2 induction.

Identification of cysteinyl-leukotriene-receptor 1 antagonists as ligands for the bile acid receptor GPBAR1.

The cysteinyl leukotrienes (CysLTs), i.e. LTC4, LTD4 and LTE4, are a family of proinflammatory agents synthesized from the arachidonic acid. In target cells, these lipid mediators bind to the cysteinyl leukotriene receptors (CysLTR), a family of seven transmembrane G-protein coupled receptors. The CysLT1R is a validated target for treatment of pulmonary diseases and several selective antagonists for this receptor, including montelukast, zafirlukast and pranlukast, have shown effective in the management of asthma. Nevertheless, others CysLT1R antagonists, such as the alpha-pentyl-3-[2-quinolinylmethoxy] benzyl alcohol (REV5901), have been extensively characterized without reaching sufficient priority for clinical development. Since drug reposition is an efficient approach for maximizing investment in drug discovery, we have investigated whether CysLT1R antagonists might exert off-target effects. In the report we demonstrate that REV5901 interacts with GPBAR1, a well characterized cell membrane receptor for secondary bile acids. REV5901 transactivates GPBAR1 in GPBAR1-transfected cells with an EC50 of 2.5 µM and accommodates the GPBAR1 binding site as shown by in silico analysis. Exposure of macrophages to REV5901 abrogates the inflammatory response elicited by bacterial endotoxin in a GPBAR1-dependent manner. In vivo, in contrast to montelukast, REV5901 attenuates inflammation and immune dysfunction in rodent models of colitis. The beneficial effects exerted by REV5901 in these models were abrogated by GPBAR1 gene ablation, confirming that REV5901, a shelved CysLT1R antagonist, is a GPBAR1 ligand. These data ground the basis for the development of novel hybrid ligands designed for simultaneous modulation of CysTL1R and GPBAR1.

The Bile Acid Receptor GPBAR1 Modulates CCL2/CCR2 Signaling at the Liver Sinusoidal/Macrophage Interface and Reverses Acetaminophen-Induced Liver Toxicity.

Drug-induced liver injury caused by acetaminophen (acetyl-para-aminophenol [APAP]) is the main cause of acute liver failure and liver transplantation in several Western countries. Whereas direct toxicity exerted by APAP metabolites is a key determinant for early hepatocytes injury, the recruitment of cells of innate immunity exerts a mechanistic role in disease progression, determining the clinical outcomes. GPBAR1 is a G protein-coupled receptor for secondary bile acids placed at the interface between liver sinusoidal cells and innate immunity. In this report, using genetic and pharmacological approaches, we demonstrate that whereas Gpbar1 gene deletion worsens the severity of liver injury, its pharmacological activation by 6ß-ethyl-3a,7b-dihydroxy-5b-cholan-24-ol rescues mice from liver injury caused by APAP. This protective effect was supported by a robust attenuation of liver recruitment of monocyte-derived macrophages and their repolarization toward an anti-inflammatory phenotype. Macrophage depletion by gadolinium chloride pretreatment abrogated disease development, whereas their reconstitution by spleen-derived macrophage transplantation restored the sensitivity to APAP in a GPBAR1-dependent manner. RNA sequencing analyses demonstrated that GPBAR1 agonism modulated the expression of multiple pathways, including the chemokine CCL2 and its receptor, CCR2. Treating wild-type mice with an anti-CCL2 mAb attenuated the severity of liver injury. We demonstrated that negative regulation of CCL2 production by GPBAR1 agonism was promoter dependent and involved FOXO1. In conclusion, we have shown that GPBAR1 is an upstream modulator of CCL2/CCR2 axis at the sinusoidal cell/macrophage interface, providing a novel target in the treatment of liver damage caused by APAP.

The Aryl Hydrocarbon Receptor (AhR) Mediates the Counter-Regulatory Effects of Pelargonidins in Models of Inflammation and Metabolic Dysfunctions.

Pelargonidins are anthocyanidins thought to be beneficial for the human health, although controversies exist over the doses needed and the unclear mechanism of action, along with poor systemic bioavailability. One putative target of pelargonidins is the aryl hydrocarbon receptor (AhR). A synthetic pelargonidin (Mt-P) was synthesized by the methylation of the pelargonidin (the natural compound indicated as P). Mt-P transactivated the AhR with an EC50 of 1.97 µM and was ~2-fold more potent than the natural compound. In vitro Mt-P attenuated pro-inflammatory activities of Raw264.7 macrophage cells in an AhR-dependent manner. In vivo, administration of the Mt-P in Balb/c mice resulted in a dose-dependent attenuation of signs and symptoms of colitis induced by TNBS. A dose of 5 mg/kg Mt-P, but not the natural compound P, reversed intestinal inflammation and increased expression of Tnf-α, Ifn-Æ´, and Il-6, while promoted the expansion of regulatory T cells and M2 macrophages. In C57BL/6J mice fed a high fat diet (HFD), Mt-P attenuated body weight gain, intestinal and liver inflammation, and ameliorated insulin sensitivity, while worsened liver steatosis by up-regulating the liver expression of Cd36 and Apo100b. These effects were abrogated by AhR gene ablation. Mt-P is a synthetic pelargonidin endowed with robust AhR agonist activity that exerts beneficial effects in murine models of inflammation and metabolic dysfunction.

Epidemiological features and disease-related concerns of a large cohort of Italian patients with active Crohn's disease.

BACKGROUND-AIMS: The SOLE study was conducted on a large cohort of Italian patients with moderate-severe Crohn's disease (CD) to assess epidemiological and disease characteristics and their correlation with disease-related worries, treatment satisfaction and adherence, workability. METHODS: The following tools were used over 12 months to assess: Results were correlated with demographic and clinical variables with linear regression models. RESULTS: 552 patients with active CD (51% men) were recruited. Higher worries were having an ostomy bag and undergoing surgery. Variables associated with a higher RFIPC score included female sex, higher disease activity, lower treatment adherence (p < 0.001), previous surgical treatments (p = 0.003). 60% of patients claimed difficulties with activities of daily living. Lower VAS scores were reported by patients with disease duration >6years; treatment satisfaction/adherence was higher with anti-TNF-α treatment. Decreased hospitalizations during follow-up and improved workability/daily activities occurred with adalimumab, infliximab, azathioprine (p < 0.001). CONCLUSION: Worries included having an ostomy bag, undergoing surgery, developing cancer: conditions significantly associated with worsened disease activity and low treatment adherence. Higher treatment adherence scores/greater workability improvements were observed in patients treated with anti-TNF-α agents.

IL-9 and Mast Cells Are Key Players of Candida albicans Commensalism and Pathogenesis in the Gut.

Candida albicans is implicated in intestinal diseases. Identifying host signatures that discriminate between the pathogenic versus commensal nature of this human commensal is clinically relevant. In the present study, we identify IL-9 and mast cells (MCs) as key players of Candida commensalism and pathogenicity. By inducing TGF-ß in stromal MCs, IL-9 pivotally contributes to mucosal immune tolerance via the indoleamine 2,3-dioxygenase enzyme. However, Candida-driven IL-9 and mucosal MCs also contribute to barrier function loss, dissemination, and inflammation in experimental leaky gut models and are upregulated in patients with celiac disease. Inflammatory dysbiosis occurs with IL-9 and MC deficiency, indicating that the activity of IL-9 and MCs may go beyond host immunity to include regulation of the microbiota. Thus, the output of the IL-9/MC axis is highly contextual during Candida colonization and reveals how host immunity and the microbiota finely tune Candida behavior in the gut.

Colonic motility in ulcerative colitis.

BACKGROUND: Inflammatory conditions affecting the gut may cause motility disturbances, and ulcerative colitis - one of the main disorders among the inflammatory bowel diseases - may display abnormal colonic motility. AIM: To review the abnormalities of the large bowel in ulcerative colitis, by considering the motility, laboratory (in vitro) and pathological studies dealing with this topic. METHODS: A comprehensive online search of Medline and the Science Citation Index was carried out. RESULTS: Patients with ulcerative colitis frequently display colonic motor abnormalities, including lack of contractility, an increase of propulsive contractile waves, an excessive production of nitric oxide, vasoactive intestinal polypeptide nerves, interleukin 1 beta, neurotensin, tachykinins levels and the weaker action of substance P, likely related to a neuromuscular dysfunction due to the inflammatory process. CONCLUSIONS: A better understanding of the pathophysiological grounds of altered colonic motility in ulcerative colitis may lead to a more in-depth knowledge of the accompanying symptoms and to better and more targeted therapeutic approaches.

Intestinal superinfections in patients with inflammatory bowel diseases.

BACKGROUND: Intestinal superinfections may occur in the setting of inflammatory bowel diseases (IBD), complicating the clinical picture and triggering flares of disease. AIMS: To report our experience with intestinal superinfections in IBD patients over a three-year period. METHODS: Charts of patients hospitalized for moderate-to-severe active disease during the observation period were reviewed, and data of patients with flares due to infections collected and analyzed. RESULTS: Overall, 15 out of 113 IBD patients (13.3%) had flare-ups related to intestinal infections; 143 acute flare-ups were thus documented, with 17 episodes (12%) related to infective agents, represented by Campylobacter jejuni (3 infections), Clostridium difficile (7 infections), and Cytomegalovirus (7 infections). All but two infections occurred in ulcerative colitis patients, and all responded to appropriate treatment. CONCLUSIONS: Intestinal superinfections may complicate the clinical picture of IBD patients, increasing the diagnostic and therapeutic burden. Appropriate early diagnostic and therapeutic measures are thus needed in these patients.

Ultrasonographic assessment of colonic wall in moderate-severe ulcerative colitis: comparison with endoscopic findings.

BACKGROUND: Bowel ultrasound has been shown to be a useful tool to evaluate patients with inflammatory bowel disease, especially Crohn's disease. However, such data are still scarce in ulcerative colitis patients. AIMS: To establish the value of bowel ultrasound in moderate to severe ulcerative colitis patients, and compare these data with endoscopic findings. PATIENTS AND METHODS: Endoscopic, ultrasound and C-reactive protein data from 51 patients with moderate to severe ulcerative colitis observed during a 3-year period were retrospectively obtained and analysed. RESULTS: All patients displayed pathological thickness (>4 mm) of the colon wall. This value strongly correlated with C-reactive protein values (p=0.0001) and the endoscopic score (p<0.0001). Also, a strong correlation (p<0.0001) was found between CRP values and endoscopic score. CONCLUSIONS: Bowel ultrasound, in expert hands, may represent a useful adjunctive (or first line) tool for the evaluation of patients with moderate to severe ulcerative colitis.

Small intestinal bacterial overgrowth and warfarin dose requirement variability.

The dose of warfarin needed to obtain a therapeutic anticoagulation level varies widely among patients and can undergo abrupt changes for unknown reasons. Drug interactions and genetic factors may partially explain these differences. Intestinal flora produces vitamin K2 (VK2) and patients with small intestinal bacterial overgrowth (SIBO) rarely present reduced INR values due to insufficient dietary vitamin K. The present study was undertaken to investigate whether SIBO occurrence may affect warfarin dose requirements in anticoagulated patients. Based on their mean weekly dose of warfarin while on stable anticoagulation, 3 groups of 10 patients each were defined: low dose (LD, or=70 mg/wk). Each patient underwent a lactulose breath test to diagnose SIBO. Plasma levels of warfarin and vitamin K-analogues were also assessed. Patients with an altered breath test were 50% in the VHD group, 10% in the HD group, and none in the LD group (P=0.01). Predisposing factors to SIBO were more frequent in the VHD group, while warfarin interfering variables were not. VHD patients were younger and had a higher plasma vitamin K1 (VK1) concentration (P>0.05). On the contrary, the plasma VK2 levels tended to be lower. This pilot study suggests that SIBO may increase a patient's warfarin dose requirement by increasing dietary VK1 absorption through the potentially damaged intestinal mucosa rather than increasing intestinal VK2 biosynthesis. Larger studies are needed to confirm these preliminary data and to evaluate the effects of SIBO decontamination on warfarin dosage.

A simplified method for anal ultrasonography: preliminary report.

BACKGROUND: Anal endosonography (AES) is able to reliably visualize and identify anal sphincter abnormalities. However, dedicated probes are quite expensive. AIM: We describe a simple and less costly method to perform AES in a unit that already has echoendoscopes available by inserting the endoscope through a disposable anoscope filled with standard ultrasound gel. PATIENTS: Subjects without anal abnormalities and patients with anal disease (abscesses, fistulas) were evaluated. RESULTS: Good-quality images were obtained in both controls and patients, with optimal visualization of the anatomic structures and pathologic features. The latter (abscesses, fistulas) were always confirmed by magnetic resonance imaging. CONCLUSIONS: This simple and less costly method allows to perform good-quality AES in units having echoendoscopes availability, without the need of a more expensive dedicated probe.

Pasta naturally enriched with isoflavone aglycons from soy germ reduces serum lipids and improves markers of cardiovascular risk.

Most studies of soy and cholesterol have tested foods made from purified soy proteins containing mainly isoflavone glycosides. Fermented soy foods have mainly isoflavone aglycons and account for a high proportion of the soy protein source in Asia, where there is an inverse relationship between soy intake and serum cholesterol. The aim of this study was to compare a novel soy germ pasta, naturally enriched in isoflavone aglycons as a result of the manufacturing process, with conventional pasta for effects on serum lipids and other cardiovascular risk markers. In this randomized, controlled, parallel study design of 62 adults with hypercholesterolemia who consumed a Step II diet that included one 80-g serving/d of pasta, we measured serum lipids, high sensitivity C-reactive protein (hsCRP), urinary isoprostanes, and brachial artery flow-mediated vasodilatation at baseline and after 4 and 8 wk. The pasta delivered 33 mg of isoflavones and negligible soy protein and led to a serum isoflavone concentration of 222 +/- 21 nmol/L; 69% of subjects were equol producers. Soy germ pasta reduced serum total and LDL cholesterol by 0.47 +/- 0.13 mmol/L (P = 0.001) and 0.36 +/- 0.10 mmol/L (P = 0.002) more than conventional pasta, representing reductions from baseline of 7.3% (P = 0.001) and 8.6% (P = 0.002), respectively. Arterial stiffness (P = 0.003) and hsCRP (P = 0.03) decreased and improvements in all the above risk markers were greatest in equol producers. All measures returned to baseline when patients were switched to conventional pasta. In conclusion, pasta naturally enriched with isoflavone aglycons and lacking soy protein had a significant hypocholesterolemic effect beyond a Step II diet and improved other cardiovascular risk markers.

3alpha-6alpha-Dihydroxy-7alpha-fluoro-5beta-cholanoate (UPF-680), physicochemical and physiological properties of a new fluorinated bile acid that prevents 17alpha-ethynyl-estradiol-induced cholestasis in rats.

3alpha-6alpha-Dihydroxy-7alpha-fluoro-5beta-cholanoate (UPF-680), the 7alpha-fluorine analog of hyodeoxycholic acid (HDCA), was synthesized to improve bioavailability and stability of ursodeoxycholic acid (UDCA). Acute rat biliary fistula and chronic cholestasis induced by 17alpha-ethynyl-estradiol (17EE) models were used to study and compare the effects of UPF-680 (dose range 0.6-6.0 micromol/kg min) with UDCA on bile flow, biliary bicarbonate (HCO(3)(-)), lipid output, biliary bile acid composition, hepatic enzymes and organic anion pumps. In acute infusion, UPF-680 increased bile flow in a dose-related manner, by up to 40.9%. Biliary HCO(3)(-) output was similarly increased. Changes were observed in phospholipid secretion only at the highest doses. Treatment with UDCA and UPF-680 reversed chronic cholestasis induced by 17EE; in this model, UDCA had no effect on bile flow in contrast to UPF-680, which significantly increased bile flow. With acute administration of UPF-680, the biliary bile acid pool became enriched with unconjugated and conjugated UPF-680 (71.7%) at the expense of endogenous cholic acid and muricholic isomers. With chronic administration of UPF-680 or UDCA, the main biliary bile acids were tauro conjugates, but modification of biliary bile acid pool was greater with UPF-680. UPF-680 increased the mRNA for cytochrome P450 7A1 (CYP7A1) and cytochrome P450 8B (CYP8B). Both UDCA and UPF-680 increased the mRNA for Na(+) taurocholate co-transporting polypeptide (NCTP). In conclusion, UPF-680 prevented 17EE-induced cholestasis and enriched the biliary bile acid pool with less detergent and cytotoxic bile acids. This novel fluorinated bile acid may have potential in the treatment of cholestatic liver disease.

Genomic and non-genomic effects of different glucocorticoids on mouse thymocyte apoptosis.

Glucocorticoids, widely used therapeutic agents for several pathologies, act upon diverse cells and tissues, including the lympho-haemopoietic system. Glucocorticoid-mediated apoptosis has been described as one of the mechanisms underlying their pharmacological and physiological effects. Glucocorticoids induce apoptosis in thymocytes through genomic and non-genomic signals. We tested thymocyte apoptosis rates as induced by a panel of glucocorticoids. Using four glucocorticoids that are widely adopted in clinical practice we compared their induction of thymocyte apoptosis and activation of non-genomic and genomic signals, including phosphatidylinositol-specific phospholipase C (PI-PLC), caspase-8, -9 and -3, and Glucocorticoid-Induced Leucine Zipper (GILZ). GILZ is a protein that is rapidly induced by glucocorticoids treatment and involved in apoptosis modulation. Results indicate different glucocorticoids have different apoptotic activity which is related to their ability to induce both genomic, evaluated as caspases activation and GILZ expression, and non-genomic effects, evaluated as PI-PLC phosphorylation.