物质平衡的实质及要点

药物、制剂吸收药物（给药部位）（血浆）血浆蛋白结合（靶器官等全身组织）药物代谢产物（肝细胞等生物转化部位）药物代谢产物代谢分布代谢产物代谢产物药物（肾、胆道等）排泄（尿粪等）排出体外药物、制剂吸收药物（给药部位）（血浆）血浆蛋白结合（靶器官等全身组织）药物代谢产物（肝细胞等生物转化部位）药物代谢产物代谢分布代谢产物代谢产物药物（肾、胆道等）排泄（尿粪等）排出体外物质平衡=总排除量（药物+代谢产物）总给药量X100%物质平衡=总排除量（药物+代谢产物）总给药量X100%HPLC-MSHPLC-紫外HPLC-荧光三种常用方法均需标准品才能进行定量分析，-因而无法获得代谢产物或所有代谢产物的排泄量物质平衡=总排除量（药物+代谢产物）总给药量X100%?(代谢产物)110100100004812162024283236404448TimePostDose(hr)Concentration(ng/mL,ngEq/mLforTRA)TotalRadioactivityCC-10004原药总放射性0.0010.0020.0030.0040.0050.0060.0070.00min0500100015002000250030003500(CPM)0.0010.0020.0030.0040.0050.0060.0070.00min010002000300040005000600070008000900010000(CPM)原药0-24小时尿液0-24小时粪便提取物物质不平衡的真正意义应用LC-MS等方法检测结果显示“物质不平衡”，实质是该候选药在动物或人体内的生物转化没搞清楚，需深入研究药物在体内的代谢，弄清主要代谢产物及排除途径，以支持药效和毒性研究结果。WHATISTHEOBJECTIVEOFTHEMASSBALANCESTUDY?ARETROSPECTIVEANALYSISOFDATAINANIMALANDHUMANEXCRETIONSTUDIESEMPLOYINGRADIOLABELEDDRUGSDrugMetabolismReviews,39:17–43,2007SarahJ.Roffey,R.ScottObach,JennyI.Gedge,andDennisA.SmithPharmacokinetics,Dynamics,andMetabolism,Pfizer,Inc.,Sandwich,Kent,UK,andGroton,CT,USAMassbalanceexcretionstudiesinlaboratoryanimalsandhumansusingradiolabeledcompoundsrepresentastandardpartofthedevelopmentprocessfornewdrugs.Fromthesestudies,thetotalfateofdrug-relatedmaterialisobtained:massbalance,routesofexcretion,and,withadditionalanalyses,metabolicpathways.使用放射性标记化合物进行实验室动物和人体物质平衡排泄研究代表新药发展阶段一个标准部分。从这些实验可获得药物相关物质的总去向：物质平衡、排泄途径，加上样品分析获得代谢途径.However,rarelydoesthemassbalanceinradiolabeledexcretionstudiestrulyachieve100%recovery.Manydefinitionsofcutoffcriteriaformassbalancethatidentifyacceptableversusunacceptablerecoveryhavebeenpresentedasadhocstatementswithoutastrongrationale.…Overall,meanrecoverywasgreaterinratsanddogsthaninhumans.Whenthecirculatinghalf-lifeoftotalradioactivityisgreaterthan50h,therecoverytendstobelower....Overall,fromthesedata,recoveryofradiolabelwouldnormallybeequaltoorgreaterthan90%,85%,and80%inrat,dog,andhuman,respectively.然而，几乎没有放射性排泄实验得到100%回收率。提出过许多接受或不接受标准但均缺乏有根据的理由。……总体来说，大鼠的平均回收率高于高于狗，狗高于人。当循环系统总放射性的半衰期长于50小时，回收率较低。……总体上，大鼠、狗和人的放射性回收率通常分别大于或等于90%、85%和80%。Sinceseveraltechnicallimitationscanunderliealackofmassbalanceandsincemassbalancedataarenotsensitiveindicatorsofthepotentialfortoxicityarisingviatissuesequestration,absoluterecoveryinhumansshouldnotbeusedasamajordecisioncriteriaastowhetheraradiolabeledstudyhasmetitsobjectives.Instead,thestudyshouldbeseenasanintegralpartofdrugdevelopmentansweringfourprincipalquestions:由于一些技术限制，更因为物质平衡不能明确指示组织残留导致的毒性，绝对回收率不应作为判断人体放射性实验达标的主要决定性标准。作为药物开发整体的一部分，该实验需要回答以下四个问题:1)Istheproposedclearancemechanismsufficientlysupportedbytheidentitiesofthedrugrelatedmaterialsinexcreta,soastoprovideacompleteunderstandingofclearanceandpotentialcontributorstointer-patientvariabilityanddrug-druginteractions?在排泄物中鉴定的代谢产物是否支持提出的清除机制，以提供一个对清除和可能产生的患者间差异和药物相互作用的全面解释。2)Whatarethedrug-relatedentitiespresentincirculationthataretheactiveprincipalscontributingtoprimaryandsecondarypharmacology?什么主要循环代谢产物与药物主要和次要的药理学相关3)Aretherefindings(lowextractionrecoveryofradiolabelfromplasma,metabolitestructuresindicativeofchemicallyreactiveintermediates)thatsuggestpotentialsafetyissuesrequiringfurtherriskassessment这些发现（低放射性血浆提取回收率、代谢产物结构显示存在化学活性中间体）指示潜在的安全性问题而要求进一步评价风险4)Doquestions2and3haveappropriatepreclinicalsupportintermsofpharmacology,safetypharmacologyandtoxicology第2、3问题是否有临产前药理、安全药理和毒理恰当的数据支持0.0010.0020.0030.0040.0050.0060.0070.00min0500100015002000250030003500(CPM)0.0010.0020.0030.0040.0050.0060.0070.00min010002000300040005000600070008000900010000(CPM)原药0-24小时尿液0-24小时粪便提取物1.鉴定胆汁、尿液和粪便中的主要代谢产物2.合成主要代谢产物标准品3.建立LC-MS定量方法4.测定排泄物中原药及代谢产物的含量5.计算回收率药物非临床药代动力学研究技术指导原则（CFDACDE2014年5月）8、物质平衡在临床前和临床早期阶段，特别是毒性剂量和有效治疗剂量范围确定的情况下运用放射性标记化合物，可通过收集动物和人体血浆、粪、尿以及胆汁以研究药物的物质平衡。这些研究能够获得化合物的排泄速率和途径等资料，而且有助于代谢产物的性质鉴定，并通过有限的数据比较它们的体内吸收和分布特点通过体外和动物样品中分离出的代谢产物可用作参比品用于临床和非临床的定量研究。同时，大鼠组织分布研究和动物胆管插管收集的胆汁能够提供药物的组织分布资料和明确胆汁清除特点。一般应采用放射性同位素标记技术研究物质平衡。有关试验方法的介绍及相关考虑见附录2放射性标记技术对于代谢广泛或代谢产物标准品不易获得的药物进行药物DMPK研究十分有用大鼠的物质平衡回收率很低5%、血浆暴露量也很低16RNO2CCH**ExcretaTime（h）Recovery（%）G5-01-MG5-02-MG5-03-MG5-04-FG5-05-FMeanSDUrine0-870.5274.5468.4057.4067.7167.716.358-246.446.627.5413.6510.528.953.0924-480.821.371.101.781.621.340.3948-720.351.320.590.720.610.720.36（0-72）78.1383.8577.6373.5580.4678.723.80Feces0-2413.722.708.959.738.818.783.9524-481.270.353.422.162.681.981.2048-720.260.960.740.270.420.530.31（0-72）15.254.0113.1112.1611.9111.294.28Cagewash0-244.491.782.754.222.353.121.1824-480.642.141.150.600.521.010.6848-720.250.480.460.311.570.610.54144-1680.280.401.710.810.550.750.57（0-168）5.664.806.075.944.995.490.57Total99.0492.6696.8191.6597.3695.503.19Mainlyexcretedwithin48hours0.005.0010.0015.0020.0025.0030.0035.0040.0045.0050.0055.0001000200030004000500060007000ARC(DPM)MinBkg0.005.0010.0015.0020.0025.0030.0035.0040.0045.0050.0055.0001002003004005006007008009001000ARC(DPM)Min0.005.0010.0015.0020.0025.0030.0035.0040.0045.0050.0055.00050100150200250300350ARC(CPM)Min尿液血浆粪便0.005.0010.0015.0020.0025.0030