Simultaneous determination of novel ß-lactamase inhibitor WCK 4234 and Meropenem in dog plasma by LC-MS/MS and its application to preclinical pharmacokinetic study.

A precise and accurate liquid chromatography-tandem mass spectrometric (LC-MS/MS) bioanalytical method has been developed and validated for the simultaneous quantification of WCK 4234 and meropenem (MEM) in dog plasma. Protein precipitation using acetonitrile was employed as a sample preparation approach. Cefepime was used as an internal standard. The developed method was selective, sensitive (limit of quantification, 0.075 µg/ml for both drugs), accurate (recovery > 90%), precise (CV < 10%) and linear (r2  ≥ 0.99, concentration range 0.075-120 µg/ml for both analytes). The developed method was successfully applied for the determination of both drugs in plasma to assess the pharmacokinetics in beagle dogs. WCK 4234 + MEM in a 1:1 ratio at 15 + 15 and 30 + 30 mg/kg doses were administered by the intravenous route. The mean plasma concentration and area under the concentration-time curve of WCK 4234 ranged from 38.3 to 77.4 µg/ml and from 47.8 to 77.1 µg h/ml, respectively, and the values for MEM ranged from 52.2 to 115.3 µg/ml and 70.5 to 133.6 µg h/ml respectively. The elimination half-life of WCK 4234 and MEM was around 0.8 h.

Assessment of the in vitro cytochrome P450 (CYP) inhibition potential of nafithromycin, a next generation lactone ketolide antibiotic.

Nafithromycin is a next generation lactone ketolide antibiotic slated to enter phase III clinical development in India for the treatment of CABP as a shorter 800 mg-OD X3 day therapeutic regimen. Nafithromycin exhibits potent activity against MDR Streptococcus pneumoniae including erythromycin and telithromycin-resistant resistant strains. Older macrolides/ketolides are reported to be potent inhibitors of CYP3A4/5. To facilitate comparative assessment of drug-drug interaction potential, CYP inhibitory activities of nafithromycin was evaluated in comparison with clarithromycin, telithromycin, cethromycin and solithromycin. CYP inhibitory activities were assessed against key CYP isoforms (CYP1A2, 2B6, 2C8, 2C9, 2C19, 2D6 and CYP3A4/5) using human liver microsomes. Additionally, time-dependent inhibition (TDI), metabolism-based inhibition (MBI) and k inact /K I activities were also investigated for CYP3A4/5. Nafithromycin did not inhibit key CYP enzymes and was found to be a weak inhibitor of CYP3A4/5. Comparator antibiotics were found to be potent inhibitors with 2- to 50-fold leftward shifts in CYP3A4/5 IC50 values, while such shift was not noted for nafithromycin. k inact /K I ratio of nafithromycin was 3- to 153-fold lower than comparator drugs, further substantiating its lower affinity for CYP3A4/5. In sum, weaker inhibition and lower k inact /K I ratio for CYP3A4/5, points towards nafithromycin's lower propensities towards clinical drug-drug interactions as compared to other macrolides/ketolides antibiotics.

Assessment of in vitro inhibitory effects of novel anti MRSA benzoquinolizine fluoroquinolone WCK 771 (levonadifloxacin) and its metabolite on human liver cytochrome P450 enzymes.

WCK 771 (INN: levonadifloxacin) is a novel antibacterial agent belonging to benzoquinolizine subclass of fluoroquinolones which is under clinical development as a parenteral formulation and its prodrug WCK 2349 (INN: alalevonadifloxacin) as an oral option. Both the drugs have been approved recently in India based on phase III trial completed for ABSSSI.In vitro CYP inhibition potential of levonadifloxacin and its sulfate metabolite (WCK 2146) were assessed in this study. The inhibitory effects of levonadifloxacin and its sulfate metabolite were assessed for seven key human liver CYP isoforms 1A2, 2B6, 2C8, 2C9, 2C19, 2D6 and 3A4 using human liver microsome (HLM) employing validated LC-MS/MS method.The results showed that levonadifloxacin and its metabolite did not inhibit enzyme activity of any of the key CYP isoforms (1A2, 2B6, 2C8, 2C9, 2C19, 2D6 and 3A4) even at supra therapeutic concentrations (12-24X, Clinical Cmax: 25-35µg/mL).These in vitro CYP inhibition studies of levonadifloxacin and its sulfate metabolite indicate lack of potential for pharmacokinetic drug interactions of levonadifloxacin when co-administered with drugs which are substrate of these isoforms. Therefore, further clinical studies evaluating CYP mediated drug-drug interactions are not warranted for levonadifloxacin and alalevonadifloxacin.

In Vivo Pharmacokinetic/Pharmacodynamic Targets of Levonadifloxacin against Staphylococcus aureus in a Neutropenic Murine Lung Infection Model.

Levonadifloxacin is a novel benzoquinolizine subclass of fluoroquinolone, active against quinolone-resistant Staphylococcus aureus A phase 3 trial for levonadifloxacin and its oral prodrug was recently completed. The present study identified area under the concentration-time curve for the free, unbound fraction of a drug divided by the MIC (fAUC/MIC) as an efficacy determinant for levonadifloxacin in a neutropenic murine lung infection model. Mean plasma fAUC/MIC requirement for static and 1 log10 kill effects against 9 S. aureus were 8.1 ± 6.0 and 25.8 ± 12.3, respectively. These targets were employed in the selection of phase 3 doses.

Simultaneous determination of zidebactam and cefepime in dog plasma by LC-MS/MS and its application to pre-clinical pharmacokinetic study.

A precise and accurate liquid chromatography-tandem mass spectrometric (LC-MS/MS) bioanalytical method has been developed and validated for the simultaneous quantification of zidebactam (ZID) and cefepime (FEP) in dog plasma. Ceftazidime was used as an internal standard. Protein precipitation method was used as sample preparation approach. The calibration curve obtained was linear (r ≥ 0.99) over the concentration range 0.156-80 µg/mL for ZID and 0.312-160 µg/mL for FEP. The method was validated as per US Food and Drug Administration guidelines and the results met the acceptance criteria. A run time of 3.5 min for each sample made it possible to analyze the maximum number of samples per day. The proposed method was successfully applied for pharmacokinetic study in beagle dogs.