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ME1111 is a novel antifungal agent currently under clinical development as a topical onychomycosis treatment. A major challenge in the application of topical onychomycotics is penetration and dissemination of antifungal agent into the infected nail plate and bed. In this study, pharmacokinetic/pharmacodynamic parameters of ME1111 that potentially correlate with clinical efficacy were compared with those of marketed topical onychomycosis antifungal agents: efinaconazole, tavaborole, ciclopirox, and amorolfine. An ME1111 solution and other launched topical formulations were applied to an in vitro dose model for 14 days based on their clinical dose and administration. Drug concentrations in the deep layer of the nail and within the cotton pads beneath the nails were measured using liquid chromatography-tandem mass spectrometry. Concentrations of ME1111 in the nail and cotton pads were much higher than those of efinaconazole, ciclopirox, and amorolfine. Free drug concentrations of ME1111 in deep nail layers and cotton pads were orders of magnitude higher than the MIC 90 value against Trichophyton rubrum ( n = 30). Unlike other drugs, the in vitro antifungal activity of ME1111 was not affected by 5% human keratin and under a mild acidic condition (pH 5.0). The in vitro antidermatophytic efficacy coefficients (ratio of free drug concentration to MIC 90 s against T. rubrum ) of ME1111, as measured in deep nail layers, were significantly higher than those of efinaconazole, tavaborole, ciclopirox, and amorolfine ( P < 0.05). This suggests that ME1111 has excellent permeation of human nails and, consequently, the potential to be an effective topical onychomycosis treatment., (Copyright © 2017 American Society for Microbiology.)

The effects of ME1111, a novel antifungal agent, on the hyphal morphology and ultrastructure of Trichophyton mentagrophytes were investigated by using scanning and transmission electron microscopy. Structural changes, such as pit formation and/or depression of the cell surface, and degeneration of intracellular organelles and plasmolysis were observed after treatment with ME1111. Our results suggest that the inhibition of energy production by ME1111 affects the integrity and function of cellular membranes, leading to fungal cell death., (Copyright © 2016 American Society for Microbiology.)

Fungal nail infection (onychomycosis) is a prevalent disease in many areas of the world, with a high incidence approaching 23%. Available antifungals to treat the disease suffer from a number of disadvantages, necessitating the discovery of new efficacious and safe antifungals. Here, we evaluate the in vitro antifungal activity and nail penetration ability of ME1111, a novel antifungal agent, along with comparator drugs, including ciclopirox, amorolfine, terbinafine, and itraconazole. ME1111 showed potent antifungal activity against Trichophyton rubrum and Trichophyton mentagrophytes (the major etiologic agents of onychomycosis) strains isolated in Japan and reference fungal strains with an MIC range of 0.12 to 0.5 mg/liter and an MIC50 and MIC90 of 0.5 mg/liter for both. Importantly, none of the tested isolates showed an elevated ME1111 MIC. Moreover, the antifungal activity of ME1111 was minimally affected by 5% wool keratin powder in comparison to the other antifungals tested. The ME1111 solution was able to penetrate human nails and inhibit fungal growth in a dose-dependent manner according to the TurChub assay. In contrast, 8% ciclopirox and 5% amorolfine nail lacquers showed no activity under the same conditions. ME1111 demonstrated approximately 60-fold-greater selectivity in inhibition of Trichophyton spp. than of human cell lines. Our findings demonstrate that ME1111 possesses potent antidermatophyte activity, maintains this activity in the presence of keratin, and possesses excellent human nail permeability. These results suggest that ME1111 is a promising topical medication for the treatment of onychomycosis and therefore warrants further clinical evaluation., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

Despite the existing treatment options for onychomycosis, there remains a strong demand for potent topical medications. ME1111 is a novel antifungal agent that is active against dermatophytes, has an excellent ability to penetrate human nails, and is being developed as a topical agent for onychomycosis. In the present study, we investigated its mechanism of action. Trichophyton mentagrophytes mutants with reduced susceptibility to ME1111 were selected in our laboratory, and genome sequences were determined for 3 resistant mutants. The inhibitory effect on a candidate target was evaluated by a spectrophotometric enzyme assay using mitochondrial fractions. Point mutations were introduced into candidate genes by a reverse genetics approach. Whole-genome analysis of the 3 selected mutants revealed point mutations in the structural regions of genes encoding subunits of succinate dehydrogenase (complex II). All of the laboratory-generated resistant mutants tested harbored a mutation in one of the subunits of succinate dehydrogenase (SdhB, SdhC, or SdhD). Most of the mutants showed cross-resistance to carboxin and boscalid, which are succinate dehydrogenase inhibitors. ME1111 strongly inhibited the succinate-2,6-dichlorophenolindophenol reductase reaction in Trichophyton rubrum and T. mentagrophytes (50% inhibitory concentrations [IC50s] of 0.029 and 0.025 μg/ml, respectively) but demonstrated only moderate inhibition of the same reaction in human cell lines. Furthermore, the target protein of ME1111 was confirmed by the introduction of point mutations causing the amino acid substitutions in SdhB, SdhC, and SdhD found in the laboratory-generated resistant mutants, which resulted in reduced susceptibility to ME1111. Thus, ME1111 is a novel inhibitor of the succinate dehydrogenase of Trichophyton species, and its mechanism of action indicates its selective profile., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

Horizontal transfer of the mutated ftsI gene from beta-lactamase-nonproducing ampicillin-resistant (BLNAR) Haemophilus influenzae to a susceptible strain was examined in vitro under selection with nine oral beta-lactams (ampicillin, amoxicillin, cefprozil, cefuroxime, cefpodoxime, cefdinir, cefcapene, cefditoren, and tebipenem). Compared to the penicillins and the carbapenem, the cephalosporins showed a wide selection window for the genetic transfer.

Horizontal gene transfer has been identified in only a small number of genes in Haemophilus influenzae, an organism which is naturally competent for transformation. This report provides evidence for the genetic transfer of the ftsI gene, which encodes penicillin-binding protein 3, in H. influenzae. Mosaic structures of the ftsI gene were found in several clinical isolates of H. influenzae. To identify the origin of the mosaic sequence, complete sequences of the corresponding gene from seven type strains of Haemophilus species were determined. Comparison of these sequences with mosaic regions identified a homologous recombination of the ftsI gene between H. influenzae and Haemophilus haemolyticus. Subsequently, ampicillin-resistant H. influenzae strains harboring identical ftsI sequences were genotyped by pulsed-field gel electrophoresis (PFGE). Divergent PFGE patterns among beta-lactamase-nonproducing ampicillin-resistant (BLNAR) strains from different hospitals indicated the potential for the genetic transfer of the mutated ftsI gene between these isolates. Moreover, transfer of the ftsI gene from BLNAR strains to beta-lactamase-nonproducing ampicillin-susceptible (BLNAS) H. influenzae strains was evaluated in vitro. Coincubation of a BLNAS strain (a rifampin-resistant mutant of strain Rd) and BLNAR strains resulted in the emergence of rifampin- and cefdinir-resistant clones at frequencies of 5.1 x 10(-7) to 1.5 x 10(-6). Characterization of these doubly resistant mutants by DNA sequencing of the ftsI gene, susceptibility testing, and genotyping by PFGE revealed that the ftsI genes of BLNAR strains had transferred to BLNAS strains during coincubation. In conclusion, horizontal transfer of the ftsI gene in H. influenzae can occur in an intraspecies and an interspecies manner.

The molecular mechanisms of reduced susceptibility to cefixime in clinical isolates of Neisseria gonorrhoeae, particularly amino acid substitutions in mosaic penicillin-binding protein 2 (PBP2), were examined. The complete sequence of ponA, penA, and por genes, encoding, respectively, PBP1, PBP2, and porin, were determined for 58 strains isolated in 2002 from Japan. Replacement of leucine 421 by proline in PBP1 and the mosaic-like structure of PBP2 were detected in 48 strains (82.8%) and 28 strains (48.3%), respectively. The presence of mosaic PBP2 was the main cause of the elevated cefixime MIC (4- to 64-fold). In order to identify the mutations responsible for the reduced susceptibility to cefixime in isolates with mosaic PBP2, penA genes with various mutations were transferred to a susceptible strain by genetic transformation. The susceptibility of partial recombinants and site-directed mutants revealed that the replacement of glycine 545 by serine (G545S) was the primary mutation, which led to a two- to fourfold increase in resistance to cephems. Replacement of isoleucine 312 by methionine (I312M) and valine 316 by threonine (V316T), in the presence of the G545S mutation, reduced susceptibility to cefixime, ceftibuten, and cefpodoxime by an additional fourfold. Therefore, three mutations (G545S, I312M, and V316T) in mosaic PBP2 were identified as the amino acid substitutions responsible for reduced susceptibility to cefixime in N. gonorrhoeae.

AG205 was identified from high-throughput screening as a potent inhibitor of FabK, the enoyl-ACP reductase in Streptococcus pneumoniae. Specific inhibition of lipid biosynthesis in a macromolecular biosynthesis assay and identification of an Ala141Ser substitution in FabK from spontaneous AG205-resistant mutants indicated that AG205 exerts antibacterial activity against S. pneumoniae through the specific inhibition of FabK.

The efficacy of ME1036, a novel parenteral carbapenem, was compared with that of vancomycin by using a rabbit model of methicillin-resistant Staphylococcus aureus (MRSA) endocarditis. Compared with vancomycin, ME1036 reduced the bacterial counts in the vegetations at a lower dosage or over a shorter period of administration when it was used for the treatment of MRSA endocarditis.