Pharmacokinetics And Safety Of Adefovir Dipivoxil Tablet After A Single And Multi-oral Dose In Chinese Healthy Volunteers

Adefovir dipivoxil(ADV)is a novel nucleoside analog which is different fromothers. It is not dependent on the cell peculiarity nucleoside kinase when changinginto activated PMEApp because it contains one phosphoric acid. Therefore, ADV hasstronger activity for various cells. The data of abroad clinical trials was indicated thatADV is effective not only in the treatment of wild chronic hepatitis patients with wildtype but also lamivudine-resistant hepatitis B virus(HBV) and that the occurrence ofviral resistance to 72～136 weeks ADV therapy is rare. So, ADV becomes thereplacement and complementary drugs to cure lamivudine-resistant HBV infection,can be used for combination therapy with HBV drugs, and can effectivly resolve theresistance to nucleoside analogue. It has broad clinical future. But, renal dysfunctionwhich is one of ADV adverse drug reaction is crucial issues for ADV extensiveapplication. Nephrotoxicity of ADV happens having lagging nature, which is relatedto dosages and the course of treatment and mainly happens in high dosages, forexample clinical trial curing human immunodeficiency virus (HIV) infection(60mg.d^-1 or 120mg.d^-1) or curing HBV infection (30 mg.d^-1). Because of this,whether ADV can be used in Chronic Hepatitis B (CHB) or not is determined by itsdosages, course of treatment and safety. ADV was empoldered by Gilead Sciences Incin USA. In September 2002, ADV appears in the market approved by FDA in USA to cure CHB. In our country, ADV still begins to develop and the pharmacokinetics ofADV in Chinese isn't yet published. In this paper, Chinese healthy volunteers weregiven a single oral dose of 5,10,30 mg ADV or 10mg ADV daily for 6 days, theplasma concentrations and urine concentrations were determined by HPLC/MS/MS,pharmacokinetic parameters of ADV were calculated, and the adverse events wereobserved. Thereby, to study the pharmacokinetics of ADV in Chinese healthyvolunteers, assess the safety of ADV, and know the dosages, course of treatment andsafety of ADV therapy to HBV. In the text, ADV tablet, as a new drug whichdeveloped by a manufacturer in the homeland was chosen as the study object. Thepoint being innovative of study drug is that has designed a new combiningpreparation route, at the same time adopted new crystal, method in preparation, so thecrystal shape is somewhat also different from Gilead Sciences Inc. These datademonstrate that ADV absorbed rapidly in volunteers, distribute in the whole bodyfluid, C_maxand AUC_0-t are directly proportion to doses, but CL/F isn't proportion toV/F. ADV eliminates mainly by kidney. There is not accumulation of ADV in plasmaafter multiple-dose of 10mg. The plasma concentrations reach stabilization at days 4.These pharmacokinetic parameters are approximately consistent with Gilead SciencesInc publication. These observations indicate that ADV single or multiple-dose is safein Chinese healthy subjects, but the long range put into use need paying attention tothe liver, kidney function and the change of sour phosphorus enzyme muscle kinase(CK). The research is first to study the pharmacokinetic parameters of domestic ADVin the China, measure method for adefovir (HPLC/MS/MS) was created first in thehomeland, and will announce the pharmacokinetics of ADV in Chinese healthyvolunteer. The results will provided the science basis of using ADV safely, rationally,effectively in clinic and have important theory significance and gigantic applicationvalue.The following parts comprise the study:1. To develop method for analyzing plasma and urine sample Plasma and urineadefovir was quantitated by using a proprietary high-pressure liquidchromatography-MS/MS methodology, in which sample was pretreated by solidphase extraction, the mass analyzer was operated under positive mode usingatmospheric pressure chemical ionization and[9-[2-(phospho-nomethoxy) propyl] adenine was selected as internal standard. The method was assessed for its speciality,precision, accuracy, sensitivity, linearity and stability of biological samples. ADV is aprodrugs and is quickly and completely hydrolyzed to monoesters metabolites andadefovir into the body. ADV and monoesters metabolites can't be detected in plasmaafter oral ADV, so it measures the concentrations of the adefovir. The result showsunder these conditions, the retention times were approximately 1.9 and 2.6 min foradefovir and internal standard respectively and there is no interference in the plasmaand urine. This assay has a lower limit of quantitation of 1ng.mL^-1,50ng.mL^-1foradefovir at the plasma and urine respectively with calibration curve ranging from 1 to200 ng.mL^-1in plasma. Blood, urine samples and extracted samples can be stable atroom temperature (25℃) for 24 hours. This method is highly specific and has goodrepeatability, high accuracy and sensitivity and the pretreatment is greatly simplified.2. To establish standard operate procedure(SOP) for ADV pharmacokinetics andsafety trial These SOP includes SOP for Phase I clinical trials, SOP for collection ofADV blood and urine sample, SOP for pretreatment of ADV biological samples et al.A total of eight SOP are conducive to the quality control of the trial.3. To study the pharmacokinetics and safety of ADV tablet in Chinese healthyvolunteers This was a single center, randomized, Latin square design, three-waycrossover study. Twelve Chinese healthy male volunteers were randomly divided intothree groups (n=4), who were given a single oral dose of 5(A),10(B),30(C)mg ADVon three separate occasions. Sequence administration was ABC, BCA, CABrespectively with dose interval of seven days. After the final cycle, subjects werefollow-up of seven days. Blood was collected at 0, 0.5, 0.75, 1, 1.5, 2, 2.5, 3, 4, 6, 8,12, 24, 36 and 48h postadministration. Plasma was separated by centrifugation andfrozen at-30℃until analysis. Pharmacokinetic analysis was performed usingnoncompartmental methods with the WinNonlin software (version 4.1; PharsightCorp., Mountain View, Calif.). The correlation among AUC_0-t, C_max and dose wasanalyzed by SPSS (10.0) after a single dose administration and Pharmacokineticparameters were analyzed by the analysis of variance. After a complete physicalexamination during screening, the volunteers were identified as qualified. At theadministration eve of each test cycle, the subjects were assessed the following majorinclusion criteria: the update medical history, physical examination, vital signs, 12-lead electrocardiogram, adverse events. Before every administration, blood, urinewas collected for clinical laboratory testing. In the study within 48 hours afteradministration the subjects were observed for adverse events during their stay at thestudy centers. After every 48 hours for medical administration, the subjects wereassessed the following major inclusion criteria: physical examination, vital signs,12-lead electrocardiogram, and blood, urine were collected for clinical laboratorytesting. Adverse events were observed in the entire process until the end. The mainpharmacokinetic parameters of adefovir after oral 5,10,30 mg were as follow: AUC_0-twere (102.7±51.7), (235.0±82.3), (715.4±267.6) ng.h.mL^-1, C_max were (11.4±3.7), (25.4±8.2), (76.3±23.0) ng.mL^-1, t_max were (1.69±1.41), (0.90±0.56),(0.94±0.50) h, t_1/2were (10.50±5.38), (9.17±2.83), (8.45±1.38) h, V/Fwere (447.22±197.53); (518.74±153.87), (543.22±173.06) L, CL/F were(30.45±5.40), (41.04±12.44), (45.81±17.00)L.h^-1, respectively. Analysis ofvariance showed that normalization of Log C_max, AUC_0-8was from the impact ofdose and administration cycle (P＞0.05), but by the impact of individual volunteers(P＜0.05). After 12 healthy volunteers oral single dose of ADV, three cases occurredalanine aminotransferase(ALT), aspartate aminotransferase(AST), creatinephosphokinase(CK) increased which were clinically relevant changes. Butwithdrawal can get back to normal without handling. The result shows that ADV 5,10and 30 mg were safe in Chinese male healthy subjects.4. To study the pharmacokinetics and safety of multiple oral administration ofADV tablet in Chinese healthy volunteers This was a single center, investigator andsubject-blind, randomized, placebo controlled study. Fifteen healthy volunteers werestratified randomly divided into the experimental group (6 male and 6 female) and theplacebo group (2 male and 1 female) by sex. On days 1, 15 volunteers received 10mgoral ADV (or placebo) once, were collected blood, urine samples within 48 hours. Ondays 3 to 7, 15 volunteers was given 10mg ADV or placebo once a day for 5 days andwere followed for 7 days after administration. On first medication (days 1) and thelast medication (days 7), blood was collected at 0, 0.5, 0.75, 1, 1.5, 2, 2.5, 3, 4, 6, 8,12, 24, 36 and 48h postadministration for study pharmacokinetics after a single doseand repeat doses respectively. Plasma was separated by centrifugation and frozen at-30℃until analysis. On days 3 to 6, blood samples were collected before taking the medication and after 1h for determining peak and trough concentrations in plasma.On days 1, urine sample was collected at 0-4,4-8,8-12,12-24,24-36,36-48hpostadministration, measured total urination of each time, transferred 5mL urinemixed of each time, frozen at-30℃until analysis for calculating the renal clearanceof adefovir. Pharmacokinetic analysis was performed using noncompartmentalmethods with the WinNonlin software (version 4.1). After a complete physicalexamination during screening, the volunteers were identified as qualified. At the eveof the first and last administration, the subjects were assessed the following majorinclusion criteria: the update medical history, physical examination, vital signs,12-lead electrocardiogram, adverse events. Before the first medication (days 1), 48hours after the first medication, before the medication (days 4) and 48 hours after thelast medication (days 7) after 48 hours, blood, urine was collected for clinicallaboratory testing. On days 14, the subjects were assessed the following majorinclusion criteria: physical examination, vital signs, 12-lead electrocardiogram, andwere collected blood, urine for clinical laboratory testing. Adverse events wereobserved in the entire process until the end. The main pharmacokinetic parameters ofadefovir after oral a single dose (10mg) and repeat doses(10mg/day, for 5days) were asfollow: AUC_0-t were (205.8±53.1), (288.8±90.6) ng.h.mL^-1, C_maxwere (21.5±5.1), (26.7±7.2) ng.mL^-1, t(max) were (1.06±0.41), (0.94±0.44) h, t_1/2 were(8.12±2.02), (9.68±2.84) h, V/F were (545.9±186.5), (464.2±119.5) L, CL/Fwere (46.2±8.9), (34.8±9.4) L.h^-1, respectively. In 48h following a single dose of10mg ADV, the dose eliminated by kidney was (67.8±15.7)％, which demonstratesthat adefovir eliminates mainly by kidney. The plasma concentrations reachstabilization at days 4. C_max, and AUC compared with the observed accumulation (thedays 7 AUC_0-24 divided by the days 1 AUC_0-24) were 1.23 and 1.28 respectively andAUC of the steady accumulation (the days 7 AUC_0-24 divided by the days 1 AUC_0-8)was 1.13, suggesting that there is not accumulation of adefovir in plasma aftermultiple-dose of 10mg for 5 days. There weren't serious adverse events within thestudy. In the test group, one patient had ALT, AST increased slightly, six cases forcontinue medication felt drowsiness, malaise, two cases had a sense of throatdiscomfort, one had stomach pain and distension after medication, one casescomplained aching joints. But these adverse events were mild, short duration of treatment and may disappear without medication. The result shows that ADVmultiple-dose of 10mg was safe in Chinese healthy subjects.