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Background: Low-dose aspirin's mechanism of action for preventing colorectal cancer (CRC) is still debated, and the optimal dose remains uncertain. We aimed to optimize the aspirin dose for cancer prevention in CRC patients through deep phenotyping using innovative biomarkers for aspirin's action. Methods: We conducted a Phase II, open-label clinical trial in 34 CRC patients of both sexes randomized to receive enteric-coated aspirin 100 mg/d, 100 mg/BID, or 300 mg/d for 3 ± 1 weeks. Biomarkers were evaluated in blood, urine, and colorectal biopsies at baseline and after dosing with aspirin. Novel biomarkers of aspirin action were assessed in platelets and colorectal tissues using liquid chromatography-mass spectrometry to quantify the extent of cyclooxygenase (COX)-1 and COX-2 acetylation at Serine 529 and Serine 516, respectively. Results: All aspirin doses caused comparable % acetylation of platelet COX-1 at Serine 529 associated with similar profound inhibition of platelet-dependent thromboxane (TX)A2 generation ex vivo (serum TXB2) and in vivo (urinary TXM). TXB2 was significantly reduced in CRC tissue by aspirin 300 mg/d and 100 mg/BID, associated with comparable % acetylation of COX-1. Differently, 100 mg/day showed a lower % acetylation of COX-1 in CRC tissue and no significant reduction of TXB2. Prostaglandin (PG)E2 biosynthesis in colorectal tumors and in vivo (urinary PGEM) remained unaffected by any dose of aspirin associated with the variable and low extent of COX-2 acetylation at Serine 516 in tumor tissue. Increased expression of tumor-promoting genes like VIM (vimentin) and TWIST1 (Twist Family BHLH Transcription Factor 1) vs. baseline was detected with 100 mg/d of aspirin but not with the other two higher doses. Conclusion: In CRC patients, aspirin 300 mg/d or 100 mg/BID had comparable antiplatelet effects to aspirin 100 mg/d, indicating similar inhibition of the platelet's contribution to cancer. However, aspirin 300 mg/d and 100 mg/BID can have additional anticancer effects by inhibiting cancerous tissue's TXA2 biosynthesis associated with a restraining impact on tumor-promoting gene expression. EUDRACT number: 2018-002101-65. [ABSTRACT FROM AUTHOR]

Helicobacter pylori infection constitutes a silent pandemic of global concern. In the last decades, the alarming increase in multidrug resistance evolved by this pathogen has led to a marked drop in the eradication rates of traditional therapies worldwide. By using a high-throughput screening strategy, in combination with in vitro DNA binding assays and antibacterial activity testing, we identified a battery of novel drug-like HsrA inhibitors with MIC values ranging from 0.031 to 4 mg/L against several antibiotic-resistant strains of H. pylori, and minor effects against both Gram-negative and Gram-positive species of human microbiota. The most potent anti-H. pylori candidate demonstrated a high therapeutic index, an additive effect in combination with metronidazole and clarithromycin as well as a strong antimicrobial action against Campylobacter jejuni, another clinically relevant pathogen of phylum Campylobacterota. Transcriptomic analysis suggests that the in vivo inhibition of HsrA triggers lethal global disturbances in H. pylori physiology including the arrest of protein biosynthesis, malfunction of respiratory chain, detriment in ATP generation, and oxidative stress. The novel drug-like HsrA inhibitors described here constitute valuable candidates to a new family of narrow-spectrum antibiotics that allow overcoming the current resistome, protecting from dysbiosis, and increasing therapeutic options for novel personalized treatments against H. pylori. [ABSTRACT FROM AUTHOR]

Simple Summary: Aspirin has been studied for a long time in familial adenomatous polyposis (FAP), but the data on the prevention of colorectal adenoma formation and recurrence remain controversial. We conducted a biomarker-based clinical study in eight FAP patients treated with low-dose Aspirin to clarify the drug's mechanism of action. Aspirin appropriately acetylated platelet cyclooxygenase (COX)-1 but left persistently high systemic thromboxane(TX)A2 and prostaglandin (PG)E2 biosynthesis (assessed by measuring their primary urinary metabolites, i.e., 11-dehydro-TXB2 and PGEM) associated with the incomplete acetylation of COX-1 in colorectal mucosa and adenomas. Proteomics of colorectal adenomas of FAP patients showed that the vimentin upregulation and hemoglobin subunit beta downregulation distinguished the FAP patients in two groups with high vs. low residual 11-dehydro-TXB2 levels, possibly identifying Aspirin nonresponders and responders, respectively. Novel chemotherapeutic strategies in FAP may involve drugs that affect TXA2 and PGE2 signaling in the colorectum with receptor antagonists, and clinical studies should be performed to verify the drugs' impact. Background: The results of Aspirin prevention of colorectal adenomas in patients with familial adenomatous polyposis (FAP) are controversial. Methods: We conducted a biomarker-based clinical study in eight FAP patients treated with enteric-coated low-dose Aspirin (100 mg daily for three months) to explore whether the drug targets mainly platelet cyclooxygenase (COX)-1 or affects extraplatelet cellular sources expressing COX-isozymes and/or off-target effects in colorectal adenomas. Results: In FAP patients, low-dose Aspirin-acetylated platelet COX-1 at Serine529 (>70%) was associated with an almost complete inhibition of platelet thromboxane (TX) B2 generation ex vivo (serum TXB2). However, enhanced residual urinary 11-dehydro-TXB2 and urinary PGEM, primary metabolites of TXA2 and prostaglandin (PG)E2, respectively, were detected in association with incomplete acetylation of COX-1 in normal colorectal biopsies and adenomas. Proteomics of adenomas showed that Aspirin significantly modulated only eight proteins. The upregulation of vimentin and downregulation of HBB (hemoglobin subunit beta) distinguished two groups with high vs. low residual 11-dehydro-TXB2 levels, possibly identifying the nonresponders and responders to Aspirin. Conclusions: Although low-dose Aspirin appropriately inhibited the platelet, persistently high systemic TXA2 and PGE2 biosynthesis were found, plausibly for a marginal inhibitory effect on prostanoid biosynthesis in the colorectum. Novel chemotherapeutic strategies in FAP can involve blocking the effects of TXA2 and PGE2 signaling with receptor antagonists. [ABSTRACT FROM AUTHOR]

The increasing occurrence of multidrug-resistant strains of the gastric carcinogenic bacterium Helicobacter pylori threatens the efficacy of current eradication therapies. In a previous work, we found that several 1,4-dihydropyridine (DHP)-based antihypertensive drugs exhibited strong bactericidal activities against H. pylori by targeting the essential response regulator HsrA. To further evaluate the potential of 1,4-DHP as a scaffold for novel antimicrobials against H. pylori , we determined the antibacterial effects of 12 novel DHP derivatives that have previously failed to effectively block L- and T-type calcium channels. Six of these molecules exhibited potent antimicrobial activities (MIC ≤ 8 mg/L) against three different antibiotic-resistant strains of H. pylori , while at least one compound resulted as effective as metronidazole. Such antimicrobial actions appeared to be specific against Epsilonproteobacteria , since no deleterious effects were appreciated on Escherichia coli and Staphylococcus epidermidis. The new bactericidal DHP derivatives targeted the H. pylori regulator HsrA and inhibited its DNA binding activity according to both in vitro and in vivo analyses. Molecular docking predicted a potential druggable binding pocket in HsrA, which could open the door to structure-based design of novel anti- H. pylori drugs. [ABSTRACT FROM AUTHOR]

Chronic inflammation takes part in the pathogenesis of some malignancies of the gastrointestinal tract including colorectal (CRC), gastric, and esophageal cancers. The use of ω3 polyunsaturated fatty acid (ω3-PUFA) supplements for chemoprevention or adjuvant therapy of gastrointestinal cancers is being investigated in recent years. Most evidence has been reported in CRC, although their protective role has also been reported for Helicobacter pylori -induced gastric cancer or Barrett's esophagus-derived adenocarcinoma. Studies based on ω3-PUFA supplementation in animal models of familial adenomatous polyposis (FAP) and CRC revealed positive effects on cancer prevention, reducing the number and size of tumors, down-regulating arachidonic acid-derived eicosanoids, upregulating anti-oxidant enzymes, and reducing lipid peroxidation, whereas contradictory results have been found in induced colitis and colitis-associated cancer. Beneficial effects have also been found in FAP and ulcerative colitis patients. Of special interest is their positive effect as adjuvants on radio- and chemo-sensitivity, specificity, and prevention of treatment complications. Some controversial results obtained in CRC might be justified by different dietary sources, extraction and preparation procedures of ω3-PUFAs, difficulties on filling out food questionnaires, daily dose and type of PUFAs, adenoma subtype, location of CRC, sex differences, and genetic factors. Studies using animal models of inflammatory bowel disease have confirmed that exogenous administration of active metabolites derived from PUFAs called pro-resolving mediators like lipoxin A4, arachidonic acid-derived, resolvins derived from eicosapentaenoic (EPA), docosahexaenoic (DHA), and docosapentaenoic (DPA) acids as well as maresin 1 and protectins DHA- and DPA-derived improve disease and inflammatory outcomes without causing immunosuppression or other side effects. [ABSTRACT FROM AUTHOR]

Recent evidence has reported that proton pump inhibitors (PPIs) can exert antineoplastic effects through the disruption of pH homeostasis by inhibiting vacuolar ATPase (H+-VATPase), a proton pump overexpressed in several tumor cells, but this aspect has not been deeply investigated in EAC yet. In the present study, the expression of H+-VATPase was assessed through the metaplasia-dysplasia-adenocarcinoma sequence in Barrett's esophagus (BE) and the antineoplastic effects of PPIs and cellular mechanisms involved were evaluated in vitro. H+-VATPase expression was assessed by immunohistochemistry in paraffined-embedded samples or by immunofluorescence in cultured BE and EAC cell lines. Cells were treated with different concentrations of PPIs and parameters of citotoxicity, oxidative stress, and autophagy were evaluated. H+-VATPase expression was found in all biopsies and cell lines evaluated, showing differences in the location of the pump between the cell lines. Esomeprazole inhibited proliferation and cell invasion and induced apoptosis of EAC cells. Production of reactive oxygen species (ROS) seemed to be involved in the cytotoxic effects observed since the addition of N-acetylcysteine significantly reduced esomeprazole-induced apoptosis in EAC cells. Esomeprazole also reduced intracellular pH of tumor cells, whereas only disturbed the mitochondrial membrane potential in OE33 cells. Esomeprazole induced autophagy in both EAC cells, but also triggered a blockade in autophagic flux in the metastatic cell line. These data provide in vitro evidence supporting the potential use of PPIs as novel antineoplastic drugs for EAC and also shed some light on the mechanisms that trigger PPIs cytotoxic effects, which differ upon the cell line evaluated. [ABSTRACT FROM AUTHOR]

Background: Genetic factors influencing the prognosis of gastric adenocarcinoma (GAC) are not well known. Given the relevance of cytokines and other pro-inflammatory mediators in cancer progression and invasiveness, we aimed to assess the prognostic role of several functional cytokine and cyclooxygenase gene polymorphisms in patients with GAC. Methodology: Genomic DNA from 380 Spanish Caucasian patients with primary GAC was genotyped for 23 polymorphisms in pro-inflammatory (IL1B, TNFA, LTA, IL6, IL12p40), anti-inflammatory (IL4, IL1RN, IL10, TGFB1) cytokine, and cyclooxygenase (PTGS1 and PTGS2) genes by PCR, RFLP and TaqMan assays. Clinical and histological information was collected prospectively. Survival curves were estimated by the Kaplan-Meier method and compared using the log rank test. Outcome was determined by analysis of Cox proportional hazards, adjusting for confounding factors. Results: The median follow-up period and median overall survival (OS) time were 9.9 months (range 0.4-120.3) and 10.9 months (95% CI: 8.9-14.1), respectively. Multivariate analysis identified tumor stages III (HR, 3.23; 95% CI:2-5.22) and IV (HR, 5.5; 95% CI: 3.51-8.63) as independent factors associated with a significantly reduced OS, whereas surgical treatment (HR: 0.44; 95%CI: 0.3-0.6) was related to a better prognosis of the disease. Concerning genetic factors, none of the 23 polymorphisms evaluated in the current study did influence survival. Moreover, no gene-environment interactions on GAC prognosis were observed. Conclusions: Our results show that, in our population, the panel of selected pro- and anti-inflammatory cytokine, and cyclooxygenase gene polymorphisms are not relevant in determining the prognosis of gastric adenocarcinoma. [ABSTRACT FROM AUTHOR]

Abstract: Accumulating evidence suggests that COX-2-derived prostaglandin E2 (PGE2) plays an important role in esophageal adenocarcinogenesis. Recently, PGE2 receptors (EP) have been shown to be involved in colon cancer development. Since it is not known which receptors regulate PGE2 signals in esophageal adenocarcinoma, we investigated the role of EP receptors using a human Barrett''s-derived esophageal adenocarcinoma cell line (OE33). OE33 cells expressed COX-1, COX-2, EP1, EP2 and EP4 but not EP3 receptors as determined by real time RT-PCR and Western-blot. Treatment with 5-aza-dC restored expression, suggesting that hypermethylation is involved in EP3 downregulation. Endogenous PGE2 production was mainly due to COX-2, since this was significantly suppressed with COX-2 inhibitors (NS-398 and SC-58125), but not COX-1 inhibitors (SC-560). Cell proliferation (3H-thymidine uptake) was significantly inhibited by NS-398 and SC-58125, the EP1 antagonist SC-51322, AH6809 (EP1/EP2 antagonist), and the EP4 antagonist AH23848B, but was not affected by exogenous PGE2. However, treatment with the selective EP2 agonist Butaprost or 16,16-dimethylPGE2 significantly inhibited butyrate-induced apoptosis and stimulated OE33 cell migration. The effect of exogenous PGE2 on migration was attenuated when cells were first treated with EP1 and EP4 antagonists. These findings suggest a potential role for EP selective antagonists in the treatment of esophageal adenocarcinoma. [Copyright &y& Elsevier]

Background and Aims: Ulceration is a common feature of inflammatory bowel diseases, where subepithelial cell growth is frequently necessary for resolution. In order to further understand the role of colonic fibroblasts in this process, we have used an in vitro model of wound repair to study the response of human colonic fibroblasts to several growth factors expressed in colonic tissues. Methods: Proliferation was determined by [[sup 3] H]thymidine incorporation into DNA in subconfluent fibroblast cultures. In vitro wound repair was determined in confluent fibroblast monolayers after mechanical denudation. The presence of growth factors secreted by fibroblasts was studied in conditioned medium by heparin affinity chromatography and immunodetection with specific antibodies. Results: Serum and platelet-derived growth factor (PDGF-BB) induced a dramatic increase in both colonic fibroblast proliferation and closure of wounded cell monolayers. Epidermal growth factor (EGF) stimulated both fibroblast activities, but the effect was less potent. However, colonic fibroblasts did not respond to transforming growth factor-β[sub 1] . Conditioned medium stimulated fibroblast proliferation and wound repair activity, which was reverted by the addition of suramin. Furthermore, a PDGF-like factor was isolated from colonic fibroblast-conditioned medium. Conclusions: EGF and PDGF-BB promote human colonic fibroblast-dependent wound repair activities. Human colonic fibroblasts may exert an autocrine regulation via the production of growth factors.Copyright © 2000 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Unlike gastric mucosa, it has been considered that lipoxygenase metabolites protect the esophageal mucosa and that prostaglandins are only secreted in the presence of esophageal inflammation. The aim of this study was to determine the profile of arachidonic acid metabolites and their response to regulatory compounds in rabbit esophageal mucosal cells in culture. Eicosanoids secreted into the medium were extracted and identified by HPLC and RIA. Esophageal mucosal cells in culture metabolized arachidonic acid mainly through the cycloxygenase pathway and PGE2 was the major arachidonic acid metabolite secreted. The addition of IL-1β and A23187 (calcium ionophore) stimulated PGE2 synthesis. In basal conditions neither leukotrienes nor HETEs were detected. However, the addition of the NDGA induced the secretion of lipoxygenase metabolites identified as 12–15 HETEs. In conclusion, rabbit esophageal epithelial cells in culture metabolize arachidonic acid via both cycloxygenase and lipoxygenase pathways. In our system, PGE2 was the main arachidonic acid metabolite. [ABSTRACT FROM AUTHOR]

Abstract: Colorectal cancer (CRC) is one of the most common neoplasms and a leading cause of death related to cancer worldwide. Hereditary nonpolyposis colorectal cancer (HNPCC) is the most frequent autosomal dominant predisposition to the development of CRC, accounting for approximately 2.5% of the total CRC burden in Spain. Genomic rearrangements in the MSH2 and MLH1 genes have been reported to account for an important proportion of the mutation spectrum in HNPCC, and DNA dosage techniques have been developed facilitating molecular screening of such deletions/duplications. We screened for MSH2 and MLH1 genomic rearrangements by multiplex ligation-dependent probe amplification (MLPA) in 142 Spanish patients at risk for HNPCC prior to the exon-by-exon mutation scanning and found a deletion encompassing exons 9–16 of MSH2 and a duplication encompassing exons 11–16 of MSH2, both only in one case. These results showed that MSH2/MLH1 rearrangements in Spanish patients at risk for HNPCC seem to be a less frequent mutational event than in other populations. [Copyright &y& Elsevier]