Effects of Tegoprazan, a Novel Potassium-Competitive Acid Blocker, on Rat Models of Gastric Acid-Related Disease.

Tegoprazan, a novel potassium-competitive acid blocker (P-CAB), is a next-generation therapeutics developed for the treatment of acid-related gastrointestinal diseases such as gastroesophageal reflux disease (GERD) and peptic ulcers. In the present study, the in vitro and in vivo pharmacological properties of tegoprazan were compared with those of esomeprazole, a representative proton pump inhibitor. In vitro enzyme assays were performed using ion-leaky vesicles containing gastric H+/K+-ATPases isolated from pigs. The in vivo efficacies of tegoprazan were evaluated in rat models of GERD and peptic ulcer. Tegoprazan inhibited the activity of porcine H+/K+-ATPase with an IC50 value of 0.53 µM in a reversible manner, whereas esomeprazole showed weak and irreversible inhibition with an IC50 value of 42.52 µM. In a GERD model, tegoprazan showed dose-dependent efficacy in inhibiting esophageal injury and gastric acid secretion with an ED50 of 2.0 mg/kg, which was 15-fold more potent than that of esomeprazole. In peptic ulcer models, tegoprazan exhibited superior antiulcer activity compared with esomeprazole. The ED50 of tegoprazan in the naproxen-, ethanol-, and water-immersion restraint stress-induced peptic ulcer models were 0.1, 1.4, and 0.1 mg/kg, respectively. In the acetic acid-induced peptic ulcer model, the curative ratio of tegoprazan at 10 mg/kg was higher than that of esomeprazole at 30 mg/kg (44.2% vs. 32.7%, respectively), after 5 days of repeated oral administration. Thus, tegoprazan is a novel P-CAB that shows potent and reversible inhibition of gastric H+/K+-ATPase and may provide stronger efficacy compared with previous proton pump inhibitors.

Randomised phase 3 trial: tegoprazan, a novel potassium-competitive acid blocker, vs. esomeprazole in patients with erosive oesophagitis.

BACKGROUND: Tegoprazan is a novel potassium-competitive acid blocker that has a fast onset of action and can control gastric pH for a prolonged period, which could offer clinical benefit in acid-related disorders. AIM: To confirm the non-inferiority of tegoprazan to esomeprazole in patients with erosive oesophagitis (EE). METHODS: In this multicentre, randomised, double-blind, parallel-group comparison study, 302 Korean patients with endoscopically confirmed EE (Los Angeles Classification Grades A-D) were randomly allocated to either tegoprazan (50 or 100 mg) or esomeprazole (40 mg) treatment groups for 4 or 8 weeks. The primary endpoint was the cumulative proportion of patients with healed EE confirmed by endoscopy up to 8 weeks from treatment initiation. Symptoms, safety and tolerability were also assessed. RESULTS: The cumulative healing rates at week 8 were 98.9% (91/92), 98.9% (90/91) and 98.9% (87/88) for tegoprazan 50 mg, tegoprazan 100 mg and esomeprazole 40 mg, respectively. Both doses of tegoprazan were non-inferior to esomeprazole 40 mg. The incidence of adverse events was comparable among the groups, and tegoprazan was well-tolerated. CONCLUSION: Once daily administration of tegoprazan 50 or 100 mg showed non-inferior efficacy in healing EE and tolerability to that of esomeprazole 40 mg.

An Optimized Combination of Ginger and Peony Root Effectively Inhibits Amyloid-ß Accumulation and Amyloid-ß-Mediated Pathology in AßPP/PS1 Double-Transgenic Mice.

The progressive aggregation of amyloid-ß protein (Aß) into senile plaques is a major pathological factor of Alzheimer's disease (AD) and is believed to result in memory impairment. We aimed to investigate the effect of an optimized combination of ginger and peony root (OCGP), a standardized herbal mixture of ginger and peony root, on Aß accumulation and memory impairment in amyloid-ß protein precursor (AßPP)/presenilin 1 (PS1) double-transgenic mice. In an in vitro thioflavin T fluorescence assay, 100 µg/ml OCGP inhibited Aß accumulation to the same extent as did 10 µM curcumin. Furthermore, AßPP/PS1 double-transgenic mice treated with OCGP (50 or 100 mg/kg/day given orally for 14 weeks) exhibited reduced Aß plaque accumulation in the hippocampus and lower levels of glial fibrillary acid protein and cyclooxygease-2 expression compared with vehicle-treated controls. These results suggest that OCGP may prevent memory impairment in AD by inhibiting Aß accumulation and inflammation in the brain.

Metformin enhances the anti-adipogenic effects of atorvastatin via modulation of STAT3 and TGF-ß/Smad3 signaling.

Adipocyte accumulation is associated with the development of obesity and obesity-related diseases. Interactions of master transcription factors and signaling cascades are required for adipogenesis. Regulation of excessive adipogenic processes may be an attractive therapeutic for treatment of obesity and obesity-related diseases. In this study, we found that atorvastatin exerts an anti-adipogenic activity in 3T3-L1 pre-adipocytes, and that this activity is elevated in combination with metformin. Expression of the adipogenic master regulators PPARγ and C/EBPα, and their target gene aP2, was suppressed by atorvastatin. Furthermore, atorvastatin treatment resulted in increased activation of the key master regulator of cellular energy homeostasis, AMPK. These biological activities of atorvastatin were elevated in combination with metformin. These anti-adipogenic activities were associated with regulation of the STAT3 and TGF-ß signaling cascades, resulting in the regulation of the expression of STAT3 target genes, such as KLF5, p53, and cyclin D1, and TGF-ß signaling inhibitory genes, such as SMAD7. Our results suggest that combination therapy with atorvastatin and metformin may have therapeutic potential for the treatment of obesity and obesity-related diseases caused by excessive adipogenesis.

Cold shock induces qnrA expression in Shewanella algae.

Plasmid-carried quinolone resistance genes, like qnrA, are widespread in Enterobacteriaceae. To gain insight into its little-understood native functions, we studied the effect of environmental conditions on chromosomal qnrA expression in Shewanella algae. Among conditions of DNA damage, oxidative and osmotic stress, starvation, heat, and cold, only cold shock increased gene expression, as measured by quantitative reverse transcription-PCR (qRT-PCR). Induction was graded and occurred during growth arrest, suggesting that qnrA may contribute to the adaptation of Shewanella to low temperatures.

Transferable quinolone resistance in Vibrio cholerae.

Ciprofloxacin was introduced for treatment of patients with cholera in Bangladesh because of resistance to other agents, but its utility has been compromised by the decreasing ciprofloxacin susceptibility of Vibrio cholerae over time. We correlated levels of susceptibility and temporal patterns with the occurrence of mutation in gyrA, which encodes a subunit of DNA gyrase, followed by mutation in parC, which encodes a subunit of DNA topoisomerase IV. We found that ciprofloxacin activity was more recently further compromised in strains containing qnrVC3, which encodes a pentapeptide repeat protein of the Qnr subfamily, members of which protect topoisomerases from quinolone action. We show that qnrVC3 confers transferable low-level quinolone resistance and is present within a member of the SXT integrating conjugative element family found commonly on the chromosomes of multidrug-resistant strains of V. cholerae and on the chromosomes of Escherichia coli transconjugants constructed in the laboratory. Thus, progressive increases in quinolone resistance in V. cholerae are linked to cumulative mutations in quinolone targets and most recently to a qnr gene on a mobile multidrug resistance element, resulting in further challenges for the antimicrobial therapy of cholera.

oqxAB encoding a multidrug efflux pump in human clinical isolates of Enterobacteriaceae.

The genes for multidrug efflux pump OqxAB, which is active on fluoroquinolones, were found in human clinical isolates on a plasmid in Escherichia coli and on the chromosome of Klebsiella pneumoniae. IS26-like sequences flanked the plasmid-mediated oqxAB genes, suggesting that they had been mobilized as part of a composite transposon.

Prevalence of plasmid-mediated quinolone resistance determinants over a 9-year period.

Recently, several plasmid-mediated quinolone resistance (PMQR) genes conferring low levels of quinolone resistance have been discovered. To evaluate the temporal change in the prevalence of PMQR genes over a decade in a tertiary hospital in the Republic of Korea, we selected every fifth isolate of Escherichia coli and Klebsiella pneumoniae and every third isolate of Enterobacter cloacae between 1998 and 2001 and between 2005 and 2006 from a collection of blood isolates. Six PMQR genes [qnrA, qnrB, qnrC, qnrS, aac(6')-Ib-cr, and qepA] were screened by multiplex PCR and then confirmed by direct sequencing, and the aac(6')-Ib-positive PCR products were digested with BtsCI to identify the aac(6')-Ib-cr variant. Of 461 isolates, 37 (8%) had one of the six PMQR genes; 13 (5%) of 261 E. coli strains, 13 (10%) of 135 K. pneumoniae strains, and 11 (17%) of 65 E. cloacae strains. qnrB was the most common PMQR gene and was found as early as 1998, whereas qnrS, aac(6')-Ib-cr, and qepA emerged after 2000. None of the isolates carried qnrA or qnrC. Ciprofloxacin resistance increased over time (P < 0.001), and the overall prevalence of PMQR genes tended to increase (P = 0.20). PMQR-positive isolates had significantly higher ciprofloxacin resistance and multidrug resistance rates (P = 0.005 and P < 0.001, respectively). The increasing frequency of ciprofloxacin resistance in Enterobacteriaceae was associated with an increasing prevalence of PMQR genes, and this change involved an increase in the diversity of the PMQR genes and also an increase in the prevalence of the mutations in gyrA, parC, or both in PMQR-positive strains but not PMQR-negative strains.

Mechanistic and structural analysis of aminoglycoside N-acetyltransferase AAC(6')-Ib and its bifunctional, fluoroquinolone-active AAC(6')-Ib-cr variant.

Enzymatic modification of aminoglycoside antibiotics mediated by regioselective aminoglycoside N-acetyltransferases is the predominant cause of bacterial resistance to aminoglycosides. A recently discovered bifunctional aminoglycoside acetyltransferase (AAC(6')-Ib variant, AAC(6')-Ib-cr) has been shown to catalyze the acetylation of fluoroquinolones as well as aminoglycosides. We have expressed and purified AAC(6')-Ib-wt and its bifunctional variant AAC(6')-Ib-cr in Escherichia coli and characterized their kinetic and chemical mechanism. Initial velocity and dead-end inhibition studies support an ordered sequential mechanism for the enzyme(s). The three-dimensional structure of AAC(6')-Ib-wt was determined in various complexes with donor and acceptor ligands to resolutions greater than 2.2 A. Observation of the direct, and optimally positioned, interaction between the 6'-NH 2 and Asp115 suggests that Asp115 acts as a general base to accept a proton in the reaction. The structure of AAC(6')-Ib-wt permits the construction of a molecular model of the interactions of fluoroquinolones with the AAC(6')-Ib-cr variant. The model suggests that a major contribution to the fluoroquinolone acetylation activity comes from the Asp179Tyr mutation, where Tyr179 makes pi-stacking interactions with the quinolone ring facilitating quinolone binding. The model also suggests that fluoroquinolones and aminoglycosides have different binding modes. On the basis of kinetic properties, the pH dependence of the kinetic parameters, and structural information, we propose an acid/base-assisted reaction catalyzed by AAC(6')-Ib-wt and the AAC(6')-Ib-cr variant involving a ternary complex.

Prevalence in the United States of aac(6')-Ib-cr encoding a ciprofloxacin-modifying enzyme.

Among 313 Enterobacteriaceae from the United States with a ciprofloxacin MIC of >/=0.25 microg/ml and reduced susceptibility to ceftazidime, aac(6')-Ib was present in 50.5% of isolates, and of these, 28% carried the cr variant responsible for low-level ciprofloxacin resistance. aac(6')-Ib-cr was geographically widespread, stable over time, most common in Escherichia coli, equally prevalent in ciprofloxacin-susceptible and -resistant strains, and not associated with qnr genes.

Plasmid-mediated quinolone resistance in non-Typhi serotypes of Salmonella enterica.

BACKGROUND: Serious infections with Salmonella species are often treated with fluoroquinolones or extended-spectrum beta-lactams. Increasingly recognized in Enterobacteriaceae, plasmid-mediated quinolone resistance is encoded by qnr genes. Here, we report the presence of qnr variants in human isolates of non-Typhi serotypes of Salmonella enterica (hereafter referred to as non-Typhi Salmonella) from the United States National Antimicrobial Resistance Monitoring System for Enteric Bacteria. METHODS: All non-Typhi Salmonella specimens from the United States National Antimicrobial Resistance Monitoring System for Enteric Bacteria collected from 1996 to 2003 with ciprofloxacin minimum inhibitory concentrations > or = 0.06 microg/mL (233 specimens) and a subset with minimum inhibitory concentrations < or = 0.03 microg/mL (102 specimens) were screened for all known qnr genes (A, B, and S) by polymerase chain reaction. For isolates with positive results, qnr and quinolone resistance-determining region sequences were determined. Plasmids containing qnr genes were characterized by conjugation or transformation. RESULTS: Conjugative plasmids harboring qnrB variants were detected in 7 Salmonella enterica serotype Berta isolates and 1 Salmonella enterica serotype Mbandaka isolate. The S. Mbandaka plasmid also had an extended-spectrum beta -lactamase. Variants of qnrS on nonconjugative plasmids were detected in isolates of Salmonella enterica serotype Anatum and Salmonella enterica serotype Bovismorbificans. CONCLUSIONS: Plasmid-mediated quinolone resistance appears to be widely distributed, though it is still uncommon in non-Typhi Salmonella isolates from the United States, including strains that are quinolone susceptible by the criteria of the Clinical and Laboratory Standards Institute (formerly the National Committee for Clinical Laboratory Standards). The presence of this gene in non-Typhi Salmonella that causes infection in humans suggests potential for spread through the food supply, which is a public health concern.

Fluoroquinolone-modifying enzyme: a new adaptation of a common aminoglycoside acetyltransferase.

Antimicrobial-modifying resistance enzymes have traditionally been class specific, having coevolved with the antibiotics they inactivate. Fluoroquinolones, antimicrobial agents used extensively in medicine and agriculture, are synthetic and have been considered safe from naturally occurring antimicrobial-modifying enzymes. We describe reduced susceptibility to ciprofloxacin in clinical bacterial isolates conferred by a variant of the gene encoding aminoglycoside acetyltransferase AAC(6')-Ib. This enzyme reduces the activity of ciprofloxacin by N-acetylation at the amino nitrogen on its piperazinyl substituent. Although approximately 30 variants of this gene have been reported since 1986, the two base-pair changes responsible for the ciprofloxacin modification phenotype are unique to this variant, first reported in 2003 and now widely disseminated. An intense increase in the medical use of ciprofloxacin seems to have been accompanied by a notable development: a single-function resistance enzyme has crossed class boundaries, and is now capable of enzymatically undermining two unrelated antimicrobial agents, one of them fully synthetic.

Hetero- and autoprocessing of the extracellular metalloprotease (Mpr) in Bacillus subtilis.

Most proteases are synthesized as inactive precursors which are processed by proteolytic cleavage into a mature active form, allowing regulation of their proteolytic activity. The activation of the glutamic-acid-specific extracellular metalloprotease (Mpr) of Bacillus subtilis has been examined. Analysis of Mpr processing in defined protease-deficient mutants by activity assay and Western blotting revealed that the extracellular protease Bpr is required for Mpr processing. pro-Mpr remained a precursor form in bpr-deficient strains, and glutamic-acid-specific proteolytic activity conferred by Mpr was not activated in bpr-deficient strains. Further, purified pro-Mpr was processed to an active form by purified Bpr protease in vitro. We conclude that Mpr is activated by Bpr in vivo, and that heteroprocessing, rather than autoprocessing, is the major mechanism of Mpr processing in vivo. Exchange of glutamic acid for serine in the cleavage site of Mpr (S93E) allowed processing of Mpr into its mature form, regardless of the presence of other extracellular proteases, including Bpr. Thus, a single amino acid change is sufficient to convert the Mpr processing mechanism from heteroprocessing to autoprocessing.

Streptomyces griseus trypsin is stabilized against autolysis by the cooperation of a salt bridge and cation-pi interaction.

Streptomyces griseus trypsin (SGT) is a bacterial trypsin that lacks the conserved disulphide bond surrounding the autolysis loop. We investigated the molecular mechanism by which SGT is stabilized against autolysis. The autolysis loop connects to another surface loop via a salt bridge (Glu146-Arg222), and the Arg222 residue also forms a cation-pi interaction with Tyr217. Elimination of these bonds by site-directed mutagenesis showed that the surface salt bridge at Glu146-Arg222 is the main force stabilizing the enzyme against autolysis. The effect of the cation-pi interaction at Tyr217-Arg222 is small, however, its presence increases the half-life by about five hours and enhances the protein stability more than three-fold considering the catalytic activity in the presence of the salt bridge. The melting temperature also showed cooperation between the salt bridge and cation-pi interaction. These findings show that S. griseus trypsin is stabilized against autolysis through a cooperative network of a salt bridge and cation-pi interaction, which compensate for the absence of the conserved C136-C201 disulphide bond.