Semisynthetic pleuromutilin antimicrobials with therapeutic potential against methicillin-resistant Staphylococcus aureus by targeting 50S ribosomal subunit.

A series of pleuromutilin analogs with a substituted 1,2,4-triazole were designed, synthesized and assessed for their in vitro and in vivo antibacterial activity. Initially, the MIC of the synthesized derivatives against five strains of Staphylococcus aureus (MRSA ATCC 43300, S. aureus ATCC 29213, clinical isolation of S. aureus AD3, S. aureus 144 and S. aureus SA17) were tested by the broth dilution method. Compounds 30a, 31b and 32a were the most active antibacterial agents in vitro against MRSA (MIC = 0.0625 µg/mL). The results of the time-kill curves showed that compounds 30a and 32a could reduce the amount of MRSA in vitro quickly (-7.70 log10 CFU/mL and -7.10 log10 CFU/mL reduction). In the experiment to further evaluate the in vivo antibacterial activity of compound 30a against MRSA, compound 30a (-1.71 log10 CFU/g) was effective in reducing MRSA load in thigh infected mice. Compound 30a (survival rate was 50%) displayed superior in vivo efficacy to that of tiamulin (survival rate was 30%) in the mouse systemic model. The results of further pharmacokinetic studies on compound 30a showed that the half-life (t1/2), clearance rate (Cl) and the area under the plasma concentration time curve (AUC0→∞) of compound 30a were 0.37 h, 5.43 L/h/kg and 1.84 µg h/mL, respectively. After affinity measurement by surface plasmon resonance (SPR), compound 30a exhibited high affinity with the 50S ribosome, with KD value of 1.95 × 10-6 M. Furthermore, the results of molecular docking studies revealed that compound 30a was successfully localized inside the binding pocket of 50S ribosomal subunit (ΔGb = -9.40 kcal/mol). Meanwhile, most of these compounds had no significant inhibitory effect on RAW 264.7 cells and 16HBE cells at the concentration of 8 µg/mL. The obtained outcomes showed that compound 30a could be utilized as an encouraging perspective in the development of a new therapeutic candidate for bacterial infection.

Design, synthesis and biological evaluation of pleuromutilin-Schiff base hybrids as potent anti-MRSA agents in vitro and in vivo.

A series of pleuromutilin derivatives with 1,2,4-triazole-3-substituted Schiff base structure were designed and synthesized under mild conditions. The in vitro antibacterial activities of the synthesized derivatives against 4 strains of Staphylococcus aureus (MRSA ATCC 43300, S.aureus ATCC 29213, S.aureus 144 and S.aureus AD3) and 1 strain of E. coli (ATCC 25922) were evaluated by the broth dilution method. Among these derivatives, compound 60 exhibited superior in vitro antibacterial effect against MRSA (MIC = 0.25 µg/mL) than tiamulin (MIC = 0.5 µg/mL), and compound 60 (-2.28 log10 CFU/mL) also displayed superior in vivo antibacterial efficacy than tiamulin (-1.40 log10 CFU/mL) in reducing MRSA load in the mouse thigh infection model. The time-kill study and the post-antibiotic effect study indicated that compound 60 showed a faster bactericidal kinetic and longer PAE time (exposure to 2 × MIC and 4 × MIC for 2 h, the PAE was 4.06 and 4.27 h) against MRSA compared with tiamulin (exposure to 2 × MIC and 4 × MIC for 2 h, the PAE was 1.72 and 2.14 h). Meanwhile, most of these compounds had no significant inhibitory effect on RAW 264.7 cells and HepG2 cells at the concentration of 4 µg/mL. Additionally, the development of resistance study showed that MRSA did not easily develop resistance against compound 60 compared with tiamulin after induction for 8 passages.

In vivo efficacy of the boron-pleuromutilin AN11251 against Wolbachia of the rodent filarial nematode Litomosoides sigmodontis.

The elimination of filarial diseases such as onchocerciasis and lymphatic filariasis is hampered by the lack of a macrofilaricidal-adult worm killing-drug. In the present study, we tested the in vivo efficacy of AN11251, a boron-pleuromutilin that targets endosymbiotic Wolbachia bacteria from filarial nematodes and compared its efficacy to doxycycline and rifampicin. Doxycycline and rifampicin were previously shown to deplete Wolbachia endosymbionts leading to a permanent sterilization of the female adult filariae and adult worm death in human clinical studies. Twice-daily oral treatment of Litomosoides sigmodontis-infected mice with 200 mg/kg AN11251 for 10 days achieved a Wolbachia depletion > 99.9% in the adult worms, exceeding the Wolbachia reduction by 10-day treatments with bioequivalent human doses of doxycycline and a similar reduction as high-dose rifampicin (35 mg/kg). Wolbachia reductions of > 99% were also accomplished by 14 days of oral AN11251 at a lower twice-daily dose (50 mg/kg) or once-per-day 200 mg/kg AN11251 treatments. The combinations tested of AN11251 with doxycycline had no clear beneficial impact on Wolbachia depletion, achieving a > 97% Wolbachia reduction with 7 days of treatment. These results indicate that AN11251 is superior to doxycycline and comparable to high-dose rifampicin in the L. sigmodontis mouse model, allowing treatment regimens as short as 10-14 days. Therefore, AN11251 represents a promising pre-clinical candidate that was identified in the L. sigmodontis model, and could be further evaluated and developed as potential clinical candidate for human lymphatic filariasis and onchocerciasis.

Efficacy and Safety of Intravenous-to-oral Lefamulin, a Pleuromutilin Antibiotic, for the Treatment of Community-acquired Bacterial Pneumonia: The Phase III Lefamulin Evaluation Against Pneumonia (LEAP 1) Trial.

BACKGROUND: Lefamulin, a pleuromutilin antibiotic, is active against pathogens commonly causing community-acquired bacterial pneumonia (CABP). The Lefamulin Evaluation Against Pneumonia (LEAP 1) study was a global noninferiority trial to evaluate the efficacy and safety of lefamulin for the treatment of CABP. METHODS: In this double-blind study, adults with CABP of Pneumonia Outcomes Research Team risk class ≥III were randomized 1:1 to receive lefamulin at 150 mg intravenously (IV) every 12 hours or moxifloxacin at 400 mg IV every 24 hours. After 6 doses, patients could be switched to an oral study drug if prespecified improvement criteria were met. If methicillin-resistant Staphylococcus aureus was suspected, either linezolid or placebo was added to moxifloxacin or lefamulin, respectively. The US Food and Drug Administration primary endpoint was an early clinical response (ECR) 96 ± 24 hours after the first dose of the study drug in the intent-to-treat (ITT) population (noninferiority margin, 12.5%). The European Medicines Agency co-primary endpoints were an investigator assessment of clinical response (IACR) 5-10 days after the last dose of the study drug in the modified ITT (mITT) and clinically evaluable (CE) populations (noninferiority margin, 10%). RESULTS: There were 551 patients randomized (n = 276 lefamulin; n = 275 moxifloxacin). Lefamulin was noninferior to moxifloxacin for ECR (87.3% vs 90.2%, respectively; difference -2.9%, 95% confidence interval [CI] g -8.5 to 2.8) and IACR (mITT, 81.7% vs 84.2%, respectively; difference -2.6%, 95% CI -8.9 to 3.9; CE, 86.9% vs 89.4%, respectively; difference -2.5%, 95% CI -8.4 to 3.4). Rates of study drug discontinuation due to treatment-emergent adverse events were 2.9% for lefamulin and 4.4% for moxifloxacin. CONCLUSIONS: Lefamulin was noninferior to moxifloxacin for the primary efficacy endpoints and was generally safe and well tolerated. CLINICAL TRIALS REGISTRATION: NCT02559310.

Antibiotic susceptibility profiles of Mycoplasma synoviae strains originating from Central and Eastern Europe.

BACKGROUND: Mycoplasma synoviae causes infectious synovitis and respiratory diseases in chickens and turkeys and may lead to egg shell apex abnormalities in chickens; hence possesses high economic impact on the poultry industry. Control of the disease consists of eradication, vaccination or medication. The aim of the present study was to determine the in vitro susceptibility to 14 different antibiotics and an antibiotic combination of M. synoviae strains originating from Hungary and other countries of Central and Eastern Europe. RESULTS: Minimal inhibitory concentration (MIC) values of a total of 41 M. synoviae strains were determined by the microbroth dilution method. The strains were collected between 2002 and 2016 and originated from Hungary (n = 26), Austria (n = 3), the Czech Republic (n = 3), Slovenia (n = 3), Ukraine (n = 3), Russia (n = 2) and Serbia (n = 1). Tetracyclines (with MIC50 values of 0.078 µg/ml, ≤0.25 µg/ml and 0.5 µg/ml for doxycycline, oxytetracycline and chlortetracycline, respectively), macrolides (with MIC50 values of ≤0.25 µg/ml for tylvalosin, tylosin and tilmicosin), pleuromutilins (with MIC50 values of 0.078 µg/ml and ≤0.039 µg/ml for tiamulin and valnemulin) and the combination of lincomycin and spectinomycin (MIC50 1 µg/ml (0.333/0.667 µg/ml)) were found to be the most effective antibiotic agents against M. synoviae in vitro. High MIC values were detected in numerous strains for fluoroquinolones (with MIC50 values of 1.25 µg/ml and 2.5 µg/ml for enrofloxacin and difloxacin), neomycin (MIC50 32 µg/ml), spectinomycin (MIC50 2 µg/ml), lincomycin (MIC50 0.5 µg/ml) and florfenicol (MIC50 4 µg/ml). Nevertheless, strains with elevated MIC values were detected for most of the applied antibiotics. CONCLUSIONS: In the medical control of M. synoviae infections the preliminary in vitro antibiotic susceptibility testing and the careful evaluation of the data are crucial. Based on the in vitro examinations doxycycline, oxytetracycline, tylvalosin, tylosin and pleuromutilins could be recommended for the therapy of M. synoviae infections in the region.

A click chemistry approach to pleuromutilin derivatives. Part 3: extended footprinting analysis and excellent MRSA inhibition for a derivative with an adenine phenyl side chain.

Five promising pleuromutilin derivatives from our former studies, all containing adenine on various linkers, were supplemented with two new compounds. The binding to Escherichia coli ribosomes was verified by extensive chemical footprinting analysis. The ability to inhibit bacterial growth was investigated on two Staphylococcus aureus strains and compared to the pleuromutilin drugs tiamulin and valnemulin. Three of the compounds show an effect similar to tiamulin and one compound shows an excellent effect similar to valnemulin.

[The efficacy of pleuromutilins on Aegyptianella pullorum infections in chickens (author's transl)]. / Zur Wirksamkeit von Pleuromutilinen bei Aegyptianella pullorum-Infektionen der Küken.

Pleuromutilins have an aegyptianellicidal efficacy. A significant chemotherapeutic influence on the course of the infection in chickens could only be achieved after repeated oral and intramuscular applications of at least 150 mg and 75 mg/kg body weight respectively. However, even at these concentrations the infections were not cleared.