Preclinical Pharmacokinetics And Tissue Distribution Study Of BZG

Background:Hepatocellular carcinoma(HCC)is the most common primary malignancy of the liver with late discovery,high degree of malignancy and high mortality.Sorafenib is the only first-line molecular targeted agent for HCC treatment nowadays.Previous studies showed that sorafenib was able to significantly increase overall survival in patients with advanced hepatocellular carcinoma.However,Sorafeni is expensive and it costs each patient tens of thousands of yuan every month which is a heavy financial burden to the patient's family.In addition,sorafenib has some serious side effects.Therefore,it has become a hotspot in the field of drug chemistry to modify structure of sorafenib reasonably or carry out new drugs with similar active sites.BZG was one of multi-target enzyme inhibitors which was similar to sorafenib in chemical structure.Our previous researches proved that BZG could inhibit the proliferation of a panel of human cancer cells in vitro.Additionally,BZG significantly suppressed tumor growth in hepatocellular carcinoma xenograft model.Objectives:On the basis of our previous researches,this study intends to apply the preclinical model to elucidate absorption and distribution of BZG,and to clarify the preclinical metabolic kinetics of BZG.At the same time this study is also a preresearch for BZG clinical research.The ultimate goal of this study is to make BZG become a new drug with independent property rights,to provide a new choice for the patients with HCC in China.Furthermore,it will lessen the financial burden of patients and obtain good economic and social benefits.Methods:The UPLC-MS/MS was used for the determination of BZG in biological samples of rats under suitable chromatographic and mass spectrometric conditions.Imatinib was used as internal standard.The specificity,linearity,accuracy,precision,recovery,matrix effect and stability of the method were verified according to the FDA guidance for bioanalytical method.After oral administration of 20mg/kg BZG,the blood samples of rats were collected at different time points within 48 h.Then the concentrations of BZG in the blood samples were detected according to the UPLC-MS/MS method for biological samples.Pharmacokinetic parameters were calculated by DAS software.After oral administration of 20mg/kg BZG,the tissues of rats including intestine,liver,lung,kidney,pancreas,muscle,fat and brain were collected at lh,6h,12h,respectively.The concentrations of BZG in the tissue samples were detected according to the UPLC-MS/MS method for biological samples in order to clarify the tissue distribution of BZG.Results:The UPLC-MS/MS method was developed for the determination of BZG in biological samples of rats in this study.The results of method validation were as follows.In the concentration range(0.5ng/ml-2500ng/ml),the calibration curves of BZG in rat biological samples showed good linear(r2?0.994).The sample peaks of BZG possessed good specificity without any impurity peaks.RE of intra-day and inter-day accuracy of quality control samples at low,medium and high concentrations were from-11.65%to 11.76%,while RSD of intra-day and inter-day precision were from 0.34%to 14.64%.The recovery in concentration of low,medium and high quality control samples ranged from 80.42%to 89.46%,with the RSD ?10.87%.The matrix effect in concentration of low,medium and high quality control samples ranged from 84.62%to 92.41%,with the RSD?10.31%.The stabilities were between-15%and 15%under the following conditions:short-term stability of BZG in rat plasma room temperature for 8 h;long-term stability of BZG in rat plasma stored at-20? for 40 days;post-preparative stability of BZG in rat plasma for 24 h;freeze-thaw stability of BZG in rat plasma through three freeze-thaw cycles.After oral administration of 20mg/kg BZG,the main results of pharmacokinetics study were as follows.The pharmacokinetic behavior in rats of BZG was consistent with the one-compartment model.The maximum concentration was 587.08±84.08ng/ml at 6.00±0.35 hours after oral administration of BZG,and the elimination half-life(t1/2)was 2.86±2.49 hours.After oral administration of 20mg/kg BZG,the main results of tissue distribution were as follows.BZG was distributed to various tissues quickly and widely within the time course examined after oral administration.The highest concentration level was achieved after 6 h.The BZG concentration in small intestine was the highest,then followed by kidney,liver,lung,pancreas,muscle,fat,and brain.Conclusions:1.In this study we developed and validated a UPLC-MS/MS method for the determination of BZG in biological samples.2.After oral administration of 20mg/kg BZG,the peak serum concentration of rats was 587.08± 84.08ng/ml at 6.00 ±0.35 hours,and the elimination half-life(t1/2)was 2.86± 2.49 hours.The pharmacokinetic behavior in rats of BZG was consistent with the one-compartment model.3.After oral administration of 20mg/kg BZG,BZG was distributed to various tissues quickly and widely.The tissue concentration from high to low in order was intestine,kidney,liver,lung,pancreas,muscle,fat and brain.