Alanine racemase a promising Helicobacter pylori drug target inhibited by propanoic acid.

Eradication of Helicobacter pylori, the class 1 carcinogen, faces several obstacles, which demand alternative options to conventional drug development methods. Alanine racemase (Alr) was proposed as H. pylori drug target, inhibited by propanoic acid (PA), in a previous in silico study. We investigated the possible treatment of H. pylori infection through Alr inhibition. A new model of H. pylori Alr was built, validated, and the binding of PA to the active site was modelled via molecular docking with a good docking score. PA minimum inhibitory concentration (MIC) against H. pylori ATCC 43504 and six H. pylori clinical isolates ranged from 312.5 to 416.7 ± 180 µg/ml and remained unchanged after 14 serial passages in increasing PA concentrations. The minimum bactericidal concentration of PA was 625 µg/ml. Selective Alr inhibition was confirmed by a significant PA MIC increase with increasing d-alanine concentrations. Similar PA MIC in other tested pathogens was recorded (312.5-625 µg/ml). PA lacked cytotoxicity in tested cell lines and efficiently eradicated H. pylori in a rat infection model. In conclusion, Alr is a promising broad-spectrum drug target, inhibited by PA without resistance development by repeated exposure for 14 serial passages.

Aspartate α-decarboxylase a new therapeutic target in the fight against Helicobacter pylori infection.

Effective eradication therapy for Helicobacter pylori is a worldwide demand. Aspartate α-decarboxylase (ADC) was reported as a drug target in H. pylori, in an in silico study, with malonic acid (MA) as its inhibitor. We evaluated eradicating H. pylori infection through ADC inhibition and the possibility of resistance development. MA binding to ADC was modeled via molecular docking. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of MA were determined against H. pylori ATCC 43504, and a clinical H. pylori isolate. To confirm selective ADC inhibition, we redetermined the MIC in the presence of products of the inhibited enzymatic pathway: ß-alanine and pantothenate. HPLC was used to assay the enzymatic activity of H. pylori 6x-his tagged ADC in the presence of different MA concentrations. H. pylori strains were serially exposed to MA for 14 passages, and the MICs were determined. Cytotoxicity in different cell lines was tested. The efficiency of ADC inhibition in treating H. pylori infections was evaluated using a Sprague-Dawley (SD) rat infection model. MA spectrum of activity was determined in different pathogens. MA binds to H. pylori ADC active site with a good docking score. The MIC of MA against H. pylori ranged from 0.5 to 0.75 mg/mL with MBC of 1.5 mg/mL. Increasing ß-alanine and pantothenate concentrations proportionally increased MA MIC. The 6x-his tagged ADC activity decreased by increasing MA concentration. No resistance to ADC inhibition was recorded after 14 passages; MA lacked cytotoxicity in all tested cell lines. ADC inhibition effectively eradicated H. pylori infection in SD rats. MA had MIC between 0.625 to 1.25 mg/mL against the tested bacterial pathogens. In conclusion, ADC is a promising target for effectively eradicating H. pylori infection that is not affected by resistance development, besides being of broad-spectrum presence in different pathogens. MA provides a lead molecule for the development of an anti-helicobacter ADC inhibitor. This provides hope for saving the lives of those at high risk of infection with the carcinogenic H. pylori.

Identification of Potential Drug Targets in Helicobacter pylori Using In Silico Subtractive Proteomics Approaches and Their Possible Inhibition through Drug Repurposing.

The class 1 carcinogen, Helicobacter pylori, is one of the World Health Organization's high priority pathogens for antimicrobial development. We used three subtractive proteomics approaches using protein pools retrieved from: chokepoint reactions in the BIOCYC database, the Kyoto Encyclopedia of Genes and Genomes, and the database of essential genes (DEG), to find putative drug targets and their inhibition by drug repurposing. The subtractive channels included non-homology to human proteome, essentiality analysis, sub-cellular localization prediction, conservation, lack of similarity to gut flora, druggability, and broad-spectrum activity. The minimum inhibitory concentration (MIC) of three selected ligands was determined to confirm anti-helicobacter activity. Seventeen protein targets were retrieved. They are involved in motility, cell wall biosynthesis, processing of environmental and genetic information, and synthesis and metabolism of secondary metabolites, amino acids, vitamins, and cofactors. The DEG protein pool approach was superior, as it retrieved all drug targets identified by the other two approaches. Binding ligands (n = 42) were mostly small non-antibiotic compounds. Citric, dipicolinic, and pyrophosphoric acid inhibited H. pylori at an MIC of 1.5-2.5 mg/mL. In conclusion, we identified potential drug targets in H. pylori, and repurposed their binding ligands as possible anti-helicobacter agents, saving time and effort required for the development of new antimicrobial compounds.