Pharmacokinetics Study Of Vortioxetine Crystal Forms

Depression is a kind of mental illness that seriously affects patients’work and life.It is very important in clinical practice because of its high prevalence,high recurrence rate and high mortality.Vortioxetine is a new antidepressant drug approved by the US FDA on September 30,2013 to treat adult Major Depressive Disorder（MDD）in the form of vortioxetine hydrobromide.Compared with traditional antidepressants,vortioxetine exerts antidepressant effects through two mechanisms of action:regulating receptor activity and inhibiting serotonin transporter reuptake,with higher efficacy,safety and tolerability.Due to the poor water solubility（0.1 mg/mL）of vortioxetine itself,our laboratory has prepared a new formulation,vortixetine hemihydrochloride,which can be developed as a new drug candidate for further study.Drug metabolism research is an indispensable part of the development of new drugs,including in vitro metabolism studies and in vivo metabolism studies.In vitro metabolism studies include determining the enzymatic kinetic parameters and determining the P450enzyme phenotype which is used to predict the metabolic clearance process and drug interaction in vivo,providing experimental basis for the metabolic process of drugs in the body.In vivo metabolic studies include the pharmacokinetics study and metabolites identification in the body,providing a basis for drug safety and effectiveness.In this paper,vortioxetine hemihydrochloride was selected as a potential antidepressant drug candidate for in vitro and in vivo metabolism studies,and it was systematically compared with the original vortexine,vortioxetine hydrobromide and vortioxetine hydrochloride.The differences in in vitro solubility and in vivo absorption of different salt crystal forms of vortioxetine were compared,and the relationship between in vitro solubility and oral absorption in vivo was discussed to provide a basis for the preparation of vortioxetine.The main contents of the research are:（1）Solubility study Determinated the solubility of vortioxetine and its three salts（hydrobromide,hydrochloride and semi-hydrochloride）in hydrochloric acid solution（0.1mol/L,PH1.0）,acetate buffer（pH 4.5）,phosphate buffer（pH 6.5）,water,SDS solutions（0.2%and 0.5%）by HPLC-UV method and compared the differences among them.The results showed that the solubility of vortioxetine and its three salts was basically the same in hydrochloric acid solution（pH 1.0）and phosphate buffer（pH 6.5）;In acetate buffer（pH 4.5）,the solubility of vortioxetine is the highest;The solubility from high to low is as follow:VOT>VOT-HCl>VOT-HBr>VOT-0.5HCl;In water,solubility from high to low is as follow:VOT-HCl>VOT-HBr>VOT-0.5HCl>VOT;In 0.5%SDS,solubility from high to low is arranged as follow:VOT-HBr>VOT=VOT-HCl=VOT-0.5HCl.（2）In vivo pharmacokinetic study of vortioxine and its salt crystals was performed by ultra-high performance liquid chromatography coupled with triple quadrupole mass spectrometry（UHPLC-MS/MS）.A new method for the determination of vortioxetine in rat plasma was established and validated systematically.The results show that the analytical method has good specificity and linearity,the matrix effect,precision,accuracy and stability were all meet the requirements and can be used for quantitative analysis of vortioxetine in rat plasma.Its pharmacokinetics in rats were studied by oral administration of 10.07μmol/kg and33.57μmol/kg of vortioxetine and its 3 salt crystal forms by equal molar.The study found no statistical difference in the pharmacokinetic parameters of vortioxetine and its three salt crystal forms at the two doses.However,the test results showed that the blood concentration of vortioxetine was the highest and the blood drug concentration was maintained for the longest time,suggesting that different crystalline substances may have different clinical effects.Although there were some differences in the in vitro solubility of vortioxetine and its three salt crystal forms,there was no significant difference in pharmacokinetic parameters.From the data above,the solubility of vortioxetine and the three crystal forms is similar in hydrochloric acid solution（pH 1.0）and the phosphate buffer（pH 6.5）,this may be the main reason why there was no statistical difference in the pharmacokinetic parameters of vortixetine crystal polymorphism in rats.（3）Analysis and identification of metabolites of vortioxetine Rats’plasma,urine and feces were collected by oral administration of vortixetine semi-hydrochloride in the dose of10 mg/kg.The metabolites of vortioxetine in rat plasma,urine and feces were screened and identified by ultra performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry（UHPLC-Q-TOF-MS）.As a result,29 metabolites were found,of which17 were in plasma,9 in urine,and 12 in feces,and 11 metabolites were found for the first time.The main metabolic pathways are methylation,hydroxylation,oxidation and glucuronidation.（4）Metabolic kinetics study and CYP450 subtypes involved in vortioxetine metabolism identification A series concentration of vortioxetine were co-incubated with rat liver microsomes,the vortioxetine remaining in the incubation solution was determined by UHPLC-MS/MS,and the enzyme kinetic parameter was fitted and calculated using GraphPad Prism 6.0 software.Metabolic enzyme subtypes of vortioxetine were determined by selective chemical inhibitors,and the major metabolic enzyme subtypes involved in the metabolism of vortioxetine in rat liver microsomes were investigated.The Vmax,Km and CL int of vortioxetine metablized in rat liver microsome were 23.76±2.99 ng·min^-1·mg^-1;88.04±31.84 ng·mL^-11 and 0.27 mL·min^-1·mg^-1,respectively.In addition to ciprofloxacin,the other six inhibitors of metabolic enzymes have a significant inhibitory effect on the metabolism of vortioxetine.This indicates that vortioxetine is widely metabolized in rat liver microsomes,including phase I CYP450 enzymes（CYP2C9,CYP2D6,CYP3A4,CYP2C19）,and phase Ⅱ enzymes such as SULT and UGT.