Pharmacokinetics Study Of Everolimus Tablet In Healthy Volunteers

Objective:1、To offer a specific, sensitive, efficient and robust method using the online solid phase extraction-high performance liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) system for the quantitative determination of everolimus in the whole blood.2、In order to offer the basis for application and delaration, making the pharmacokinetics study of Everolimus tablet in healthy volunteers which provided by Shandong new era Pharmaceutical Co., Ltd.Methods:1、After mixed with IS,200 uL of the whole blood samples were precipitated by 0.4 M Zinc sulphate solution (containing 80% methanol), and the supernatant was analyzed by the online SPE LC-MS/MS system. The analyte was separated on Waters SunFireTM C]8 (4.6×150 mm,3.5μm) with 10% water which 0.1% formic acid was included and 5mM ammonium formate (A)-methanol containing 0.1%fomic acid(B) (10:90,v/v) as the mobile phase.50 uL of analyte was injected onto the column. Positive electrospray ionization and multiple reaction monitoring (MRM) mode was employed. The transition of m/z 975.8→908.6 for everolimus and m/z 809.6→756.7 for the IS were monitored. Each run was 5 min.2、Two parts are included:(1) Pharmacokinetic study on single dose(11),according to 3×3 crossover trial design,11 healthy adult volunteers were given three oral dosage levels (0.75 mg,1.5 mg,3.0 mg) of everolimus tablets respectively.(2)Pharmacokinetic study on Multiple doses (11), according to 2×2 crossover trial design (0.75mg,1.5mg). The samples were detected by online-SPE-LC-MS/MS, and the pharmacokinetic parameters were calculated by WinNonlin 6.3 of Pharsight.Results:1、 The linear range of the calibration curve (0.201-100.7 ng·mL-1) with a good correlation coefficient (r=0.9998±0.0002) was obtained. The lower limit of quantification was 0.201ng·mL-1, average recovery was 99.16-105.4% and the intra-and inter-day precision represented by RSD were<8%. Results for process efficiency test and all stability tests met the acceptance criteria.2、(1) The main pharmacokinetic parameter of oral everolimus 0.75mg,1.5mg and 3.0 mg as below:the values of Tmax were (0.82±0.16) h、(0.64±0.17) h and (0.73±0.21) h, Cmax were (4.42±1.18) ng·mL-1、(9.47±1.72) ng·mL-1 and (17.30±5.19) ng·mL-1, AUC0-120 were (22.86±5.80) ng·mL-1·h、(59.10±12.57)ng·mL-1·h and(118.14±25.96) ng·mL-1·h,AUC0-∞ were29.99±7.87) ng·mL-1·h、(70.95±13.15) ng·mL-1·h and (129.90±25.67) ng·mL-1·h,t1/2z were (20.90±9.24) h、(32.06±6.94) h and (31.82±5.51) h,CLz/F were (26.88±8.02) L·h-1、(21.83±4.10) L·h-1 and (23.72±3.54) L·h-1,Vz/F were (728.93±184.25) L、(981.33±142.98) L and (1087.52±259.48) L,MRT0-120 were (10.44±3.45) h、(18.16±5.58) h and (22.04±3.71) h,MRT0-∞ were (24.09±9.54) h、(34.23±8.01) h and (32.96±6.10) h.(2) The main pharmacokinetic parameter of oral everolimus (0.75mg,1.5mg) 11 times as below:the values of Tmax were (0.68±0.20) h and (0.61±0.13) h,Css_max were (8.07±2.94) ng·mL-1 and (17.14±6.88) ng·mL-1,Css_min were (2.19±0.77) ng·mL-1 and (3.93±1.47) ng·mL-1,Cac were (3.44±1.08) ng·mL-1 and (6.69±2.29) ng·mL-1 AUCss were (41.26±12.97) ng·mL-1·h and (80.25±27.49) ng·mL-1·h,AUC0-120 were (126.27±48.96) ng·mL-1·hand (246.42±105.77) ng·mL-1 h,AUC0-∞ were (140.42±52.21) ng·mL-1 ·h and (264.46±111.76) ng·mL-1·h,ti/2z were (33.06±4.19) h and (29.16±5.22) h, CLss/F were (19.95Q6.62) L·h-1 and (21.43±9.58) L·h-1, Vz/F were (938.65±281.99) L and (898.42±455.30) L, MRT0-∞ were (33.08±4.14) h and (31.46±5.65) h, DF were (169.62±34.16)% and (194.55±44.31)%, Accumulation Index were 4.50±0.50 and 4.03±0.62.Conclusion:1、The method is simple and quick, with high repeatability, specificity and sensitivity, which can be used to analyze clinical samples and determine the pharmacokinetics of everolimus.2、The Cmax and AUCS of single dose (0.75 mg,1.5 mg and 3.0 mg)showed a dose dependent, the main pharmacokinetic parameters are similar to foreign literature reports.There was no difference between sexes in the main pharmacokinetic parameters.After taking everolimus for 11 times(0.75 mg,1.5 mg),the Cmax、AUC0-t、AUC0-∞ had statistical significance with a single dose (P<0.05), which showed that everolimus accumulated in the body.During the trial, there were 2 adverse events, but no serious, showing the everolimus tablets will betolerated well.