| Sitafloxacin is a fluoroquinolone antibacterial with in-vitro activity against a broad range of Gram-positive and-negative bacteria, including anaerobic bacteria, as well as against atypical pathogens, and the activity of sitafloxacin appeares greater than that of levofloxacin, moxifloxacin or ciprofloxacin against the main pathogenic bacteria in respiratory tract infection, such as streptococcus pneumonia, haemophilus influenza, moraxella catarrhalis, klebsiellla pneumoniae, et al. Besides, sitafloxacin was also more active against legionella pneumophila, mycoserum and chlamydia pneumonia than comparator fluoroquinolones. Sitafloxacin inhibits DNA gyrase and topoisomerase IV and the inhibitory activity of sitafloxacin against these enzymes was greater than that of comparator fluoroquinolones, including levofloxacin, ciprofloxacin and moxifloxacin, et al. Sitafloxacin was well tolerated in healthy Japanese volunteers after single (25,50,100, and 200 mg) oral dose, and the serum concentrations increased when the dose goes up. Since June 2008, this drug has been used clinically in Japan for a number of conditions including pneumonia, cystitis, and pyelonephritis. The dosage and administration in adult patients is 50 or 100 mg, administrated orally, twice daily. But whether the indication and the dosage regimen are suitable for our Chinese infection patient population? It is necessary to clarify the Pharmacokinetics (PK, the drug absorption, distribution, metabolism and excretion) in the Chinese population, and to know the Pharmacodynamics (PD) of sitafloxacin on domestic target pathogens, and then a better regimen can be selected based on the PK/PD study. The selected regimen could be applied to the further clinical study in the target patients. All of these studies can provide scientific data of sitafloxacin for the later clinical use. Thus, we carried on the pharmacokinetic study following single oral dose of sitaflocaxin (50 mg, 100 mg and 200 mg) under fasting state. Also, we conducted oral 100 mg multiple-dose pharmacokinetic and tolerance study of satifloxacin for ten days. Furthermore, a PK/PD study was carried out based on our previous PD data. This study included four parts:Part I. Establishment and validation of the method for the determination of satifloxacin in human serum and urineAn ultra high performance liquid chromatographic-tandem mass spectrometric (UPLC-MS/MS) method was developed and validated to determine satifloxacin concentrations both in serum and urine samples, in order to provide a simple and convenient method to quantitative analysis in the single dose and multiple-dose oral pharmacokinetic study of satifloxacin. Waters UPLC instrument and AB QTRAP-4000 tandem mass spectrometer were applied in the process of quantitative analysis. Satifloxacin was separated by using a Waters ACQUITY HSS T3 column (100 mm×2.1 mm I.D.,1.8 μm). Mobile phase was consisted of A (acetonitrile)/B(10 mmol ammonium formate solution, pH 3.0), and was deliverd at a flow rate of 0.4 mL/min under a gradient elution procedure. The gradient used was 0-1.6min 30%A; 1.6-2.6min 90%A; 2.6-2.7min 30%A; 3.5min, stop. Column temperature:30℃. The biological samples were pretreated by Solid Phase Extraction (SPE). Both sitafloxacin and internal standard were determined using electrospray ionization and the MS data acquisition via the multiple reaction monitoring in positive ion mode. Results showed that:① The lower limit of quantification for serum was 0.00500 μg/mL and the calibration curves were linear in the concentration range of 0.00500-1.00 μg/mL. The lower limit of quantification for urine was 0.100 μg/mL and the calibration curves were linear in the concentration range of 0.100-10.0 μg/mL. ②The calculated matrix effect factors of blank serum and blank urine for sitafloxacin were (97.1～98.4)% and (98.8-107.0)%, RSD were(1.8～5.6)% and (2.3～5.9)%, respectively.③ For serum and urine samples, the recoveries of sitafloxacin were (88.9～103.3)% and (90.0～92.8)%, RSD were (3.5～7.8)% and (3.6～8.9)%, respectively. ④The intra-day precision of serum and urine were (1.2～3.9)% and(2.2～4.1)%, and the inter-day precision of serum and urine were (1.1～4.1)% and(1.8～4.7)%. ⑤The accuracy of 1:10 diluted serum samples was 101.3%, RSD was 2.7%, and the accuracy of 1:10,1:20 and 1:100 diluted urine samples was (96.7～111.5)%, RSD was (1.5-2.6)%. ⑥ Satifloxacin quality control samples of serum and urine kept stable over 6h and 8h at room temperature before pretreatment, while they both kept stable over 48 h post-treatment at 10℃. ⑦ Satifloxacin quality control samples of serum and urine kept stable over three and four cycles of freeze (-40℃) and thawing (room temperature) respectively. The long term stability test showed that sitafloxacin in serum and urine were stable for 102 days and 82 days respectively.Part Ⅱ. Pharmacokinetic and safety study of satifloxacin following single oral dose at 50mg, 100mg,200mg in healthy volunteers under fasting conditionsThis research was single center, open, consists of three dose groups (50mg, 100mg,200mg). 36 healthy volunteers (female/male= 1:1) were enrolled in this study, and were administered orally 50 mg,100 mg and 200 mg satifloxacin tablets under fasting conditions. The serum and urine samples were collected on schedule according to the clinical trial protocol, and were determined using the method described in the Part I. Pharmacokinetic software Phoenix WinNonlin6.0 was applied to analysis data of biological samples and calculate the parameters of pharmacokinetics. Gender was also considered during the analysis of the PK character of satifloxacin. What’s more, adverse events were given enough attention during the whole study.The results showed that: ①non-compartmental analysis showed that, after a single dose of 50mg, 100mg and 200mg, Cmax were (0.718±0.189)μg/mL, (1.39±0.36)μg/mL and (2.32±0.78) μg/mL; Tmax were (0.92±0.47)h, (1.42±1.14)h and (1.21±0.94)h. AUC0-∞ were (3.67±0.66)μg-h/mL, (8.19±1.15)μg-h/mL and (14.22±3.21)μg-h/mL; Vdz/F N were (2.24±0.34) L/kg、(2.14±0.33) L/kg and (2.99±1.28) L/kg; CLt/F were (14.03±2.51) L/h、(12.42±1.67) L/h and (14.87±4.05) L/h;T1/2 were (6.67±0.72)h, (6.84±1.17)h and (8.39±2.00)h. ②The results of compartmental analysis demonstrated that the pharmacokinetic feature of satifloxacin fits a two compartment model. The volume of central compartment (V1/F) increased with dose going up from 50 mg to 200 mg, which were 52.66L,68.10L and 84.99L respectively. T1/2,α also became longer when the dose increased,and the values were 0.98h,1.26h and 1.43h respectively. The tendency of other PK parameters was irregular. ③Accumulation excretion (Ae) of sitafloxacin in urine was similar in three groups(50mg, 100mg and 200mg), which were(69.95±11.31)%, (69.03±9.2)% and (61.36±15.71)%. ④The PK character of satifloxacin was linear and was not affected by gender. ⑤There were 12 people (20 cases) adverse events after taking sitafloxacin tablets of 50mg, 100mg and 200mg, among which 3 people(4 cases) were drug relevent. All the adverse events were mild and transient. The results showed that the tolerance of sitafloxacin is good.Part Ⅲ. Pharmacokinetics and safety of sitafloxacin following postpranial oral multiple-dose of 100mg in healthy volunteersThis research was single center, open, one single dose group trial.12 healthy volunteers (female/male=1:1) were enrolled in this study, and were administered orally 100 mg satifloxacin tablets, twice daily. The serum and urine samples were collected on schedule according to the protocol. The full course PK samples were collected on day 1 and day 10 after dosing, while trough serum samples were also collected prior to dosing on day 4-day 9, respectively. The UPLC-MS/MS method was applied in the determination of satifloxacin concentration in biological samples. Pharmacokinetic analysis was carried out by WinNonlin 6.0 software, and to evaluate the pharmacokinetic feature of multiple-dose oral pharmacokinetic study of satifloxacin, to explore whether gender has an influence on the pharmacokinetic, and to evaluate the accumulation effect. What’s more, safety of sitafloxacin was also been evaluated during the research.The results showed that: ①he non-compartmental analysis showed that, after 10-days dose of 100mg, Cmax on dayl and day10 were (1.01±0.19) μg/mL and (1.26±0.20) μg/mL, respectively; Tmax were (1.75±0.62) and (2.08±0.76) h, respectively; AUC0-12were (5.13±0.63) μg·h/mL and (6.88±0.81) μg·h/mL, respectively; Vdz/F N were (2.63±0.70) L/kg and (2.38±1.00) L/kg, respectively; CLt/F were (14.72±1.48) L/h and (10.65±1.69) L/h, respectively;T1/2 were (7.53±1.63)h and (9.37±2.83) h, respectively. Rcmax and RAUC were 1.26±021 and 1.35±016, respectively, which indicated that sitafloxacin accumulated slightly in body after 10-days dosing. ②The results of compartmental analysis demonstrated that the volume of peripheral compartment (V2/F) increased significantly (P<0.01) compared with that of the single dose of sitafloxacin; k21 (P<0.01) and ka (P<0.05) decreased remarkably; while T1/2,α (P<0.01) and T1/2,β (P<0.01) prolonged. These results demonstrated that the process of absorption, distribution and excretion go slower after multiple-dose of satifloxacin. ③ccumulation excretion (Ae) of sitafloxacin in urine on day1 and day10 were (67.64±9.6)% and (99.11±17.5)%, respectively, the higher Ae on day 10 was due to the twice-daily dose from day3 to day9.④Tmax in female subjects was longer than that in male subjects (2.58h±0.66h vs 1.58h±0.49h, P<0.05); CLSS/F N and CLr N in female subjects were also higher (P<0.05); while RAUC in female subjects was lower(P<0.05). The results indicated female subjects have higher excretion rate and lower accumulation of sitafloxacin. ⑤here were 6 people (11 cases) adverse events, among which 4 people(7 cases) were drug relevent among, only one was moderate, others were all mild, and all the adverse events were transient. The results showed that the drug tolerance of sitafloxacin is good.Part Ⅳ PK/PD study of satifloxacin and evaluation of the dose regimensFluoroquinolone drugs are concentration dependent according to the concept of PK/PD. The main PK/PD parameters are Cmax/MIC (ratio by the maximum plasma concentration and minimal inhibitory concentration) and AUC/MIC (ratio by the area under the concentration-time curve and minimal inhibitory concentration).Sitafloxacin is a new kind of fluoroquinolone antibacterial drugs, so it is concentration dependent like other fluoroquinolone drugs, and the PK/PD parameters are Cmax/MIC and AUC/MIC.Based on the results from our pharmacokinetics studies in Part Ⅲ and the previous pharmacodynamic study, we estimated the cumulative fraction of response (CFR) of satifloxacin against streptococcus pneumoniae and the probability of target attainment (PTA) under various MIC levels using single point estimation and Monte Carlo simulation. We assumed that when Cmax/MIC≥2 and AUC0-24/MIC≥30, good clinical and microbiological efficacy could be obtained. Results showed that, apart from MRSA, for all the other bacteria(Streptococcus pneumonia, Haemophilus influenza, Moraxella catarrhalis, Escherichia coli and Acinetobacter baumannii), Cmax/MIC≥8.1 and AUC0-24/MIC≥89.0. CFR was almost 100% for PSSP, PISP and PRSP, PTA>99% when MIC<0.125 μg/mL.Conclusions1. Sitafloxacin was absorbed rapidly, distributed extendedly and excreted slowly, most of sitafloxacin was excreted though urine without metabolism after oral administration in healthy subjects. Sitafloxacin accumulated slightly after multiple doses of 100mg, twice daily. The PK character of sitafloxacin in Chinese healthy subjects was slightly affected by gender, but the excretion from urine was not affected.2. Sitafloxacin tablets 100mg, administered twice a day for 10 days, tolerance is good.3. The PK/PD parameters of common pathogens in lung infections (Streptococcus pneumonia, Haemophilus influenza and Moraxella catarrhalis) and common pathogens in urinary tract infections (Escherichia coli and Acinetobacter baumannii) were very high when the dosing regimen (100mg, twice a day) is used, and the regimen was expected to achieve good clinical and microbiological efficacy. |
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