本計畫為為期三年期的連續性研究，主要是在探討、研究並評估一新prokinetic agent –Morsapride為大鼠細胞色素3A 體內外活性探針試藥之可能性。此三年期計劃各分年的目標與執行進度如下：第一年： 開發體外微粒體與大鼠血漿藥品濃度的確效方法及靜脈劑量範圍的確定，並完成體外(肝及腸)微粒體酵素動力學的解析，期望藉由體外表現特定細胞色素各基因型之supersome 代謝活性與大鼠肝腸微粒體酵素動力學解析，進一步了解CYP3A 在本藥代謝所扮演的貢獻。第二年: 預計以靜脈注射方法給大鼠5 種不同劑量。又因CYP3A 表現量有性別差異，故公鼠與母鼠皆須研究。再藉由血中濃度之原型藥與代謝物藥動學解析，欲達成單點法的開發。為增大體內細胞色素3A 含量變化的範圍也需加入誘發劑與不可逆抑制劑組別(預定各兩組)，並利用西方點墨技術來測量肝臟中細胞色素CYP3A 的表現量，進而評估二者之關係。第三年: 與第二年方法類似，但欲改以口服方式給藥，評估是否亦能預測肝臟、腸道 CYP3A 的表現量。目前第一年度計畫已順利進行。並已將成果發表於Biomedical Chromatography 期刊。此方法也成功應用在體外microsome酵素動力學參數的決定。並能確實應用在大鼠藥物動力學之研究。第二年度部分實際執行(1, 5, 10, 15 mg/kg) 四種劑量已可完整描述所需臨床濃度範圍。母鼠部分則仍持續進行中。誘發劑與不可逆抑制劑組也已完成。結果顯示，以靜脈注射方式投予大鼠mosapride後，只要利用1至4個時間點的血中濃度就可以對mosapride的AUC有準確的預測，亦即能對本藥之清除率（主要反應肝清除率）能有好的預測。本法利用mosapride少量時間採點的方式，提供大鼠活體內肝臟CYP3A2活性評估之一方便、便宜、易取得與節省時間的方式。目前，並已開始執行口服組的藥動實驗預試驗，也同時開發CYP3A2西方點墨法在腸樣本定量方法的開發。期能在三年期限內盡量完成所有實驗。 In this project, it was proposed that “Investigating mosapride as a potential CYP3A probe in rats: from in vitro to in vivo”. In brief, in the first year, an analytical HPLC method will be validated. Enzyme kinetics of mosapride and other common CYP3A probes (e.g., midazolam, and testosterone) in rat liver and intestinal microsome will be assessed. Production of metabolites via specific recombinant rat and human microsomes were employed to evaluate the contribution of CYP3A on the active metabolite (M1). In the second year, pharmacokinetics of mosapride and M1 after intravenous injection (5 different doses) will be determined in male and female rats. It is aimed to find a proper dose to establish a single time point to assess hepatic CYP3A activity. In the third year, pharmacokinetics of mosapride and M1 after oral administration (5 different doses) will be investigated in male and female rats. It is aimed to find a single time point to assess intestinal CYP3A activity. Last year, we finished the first year parts. The developed and validated HPLC method for simultaneous determination of mosapride and its major metabolite in small volume rat plasma was published in Biomedical Chromatography. This method was also successfully applied to in vitro supersome/microsome enzyme kinetic analysis. This year (the second year), the linear pharmacokinetics characteristics were established between 1-15 mg/kg dose of mosapride via i.v. bolus route. It was found that the hepatic CYP3A2 protein contents showed strongly correlation with mosapride clearance. Mosapride AUC can be well predicted by using one to four points of plasma concentrations. To establish the better correlation, female rats will remain for further studies. Preliminary studies for oral route administration were in progress currently to bridge the third year goals.
