In vitro anti-helicobacter pylori activity of a new artemisinin derivative

Introduction: Helicobacter pylori (H. pylori) is a microaerophilic spiral bacterium that is associated with the pathogenesis of active and chronic gastritis, peptic ulcer, and gastric carcinoma. H. pylori eradication therapy is compromised by the increasing drug resistance. This is particularly alarming since in vivo studies demonstrate that antibiotic therapy targeted at eliminating the microbe, may also contribute to the prevention of stomach cancer in humans. The aim of this study was to evaluate the in vitro activity of a new artemisinin derivative, GC012 alone and in combination with two antibiotics commonly used in anti-H. pylori therapy. Moreover, since the antibiotics degradation can occur in an acidic environment, the stability of the tested compound was investigated at different pH values. Materials and Methods: Twenty-four clinical strains of H. pylori with different antibiotic susceptibility pattern, were used in this study. A reference strain of H. pylori ATCC 43504 was used as control. The Minimal Inhibitory Concentrations (MICs) and Minimal Bactericidal Concentrations (MBCs) were performed by modified broth dilution method; the same method was used for MICs and MBCs determination at different pH values (2.5; 5.5 and 7.5). The bactericidal activity of GC012 was evaluated by time-to-kill assay. H. pylori ATCC 43504 and a clinical isolate resistant to clarithromycin (CAM), were used to test the efficacy of GC012 at 0.5 (0.5X), 1 (1X) and 2 times (2X) the MIC value. The combination effect of standard drugs and GC012 was examined by using the checkerboard method against H. pylori ATCC 45304 and five clinical isolates having a different antimicrobial susceptibility pattern against CAM and metronidazole (MNZ). Of these, H. pylori ATCC 45304 was MNZ resistant, one was MNZ and CAM resistant, one was CLR resistant, and the other two were CLR, AMX and MNZ susceptible. Results: H. pylori activity was assessed in term of bacteriostatic and bactericidal effect. GC012 showed good in vitro antimicrobial activity, with MIC50 of 0.06 mg/L, MIC90 of 0.125 mg/L, and MBC50 of 0.125 mg/L. Interestingly, the pH variation did not alter the activity of GC012 in all the strains tested. Killing kinetic data demonstrated that GC012 at 1X MIC (0,125 µg/ml for both strains) had bactericidal effect on both the reference and clinical strains, leading to a total killing after 48 h. Using the fractional inhibitory concentration index, GC012 showed synergism with amoxicillin and metronidazole in 80% of the strains, and with clarithromycin in 60%. Conclusion: GC012 shows excellent antibacterial activity against H. pylori, no alteration of MIC value was detected under acid conditions. So, GC012 may be useful for future development in the treatment of H. pylori infection.