内源性物质药物的定量测定及生物等效性评价研究进展

从定量分析和方法验证的角度来看,生物样品中内源性化合物的定量测定是非常复杂和困难的,而内源性药物的生物等效性评价同样也有其特殊性。文中通过对国内外文献资料进行调研、分析和总结,详细综述了内源性药物的生物分析方法及生物等效性试验设计和评价方法,并对面临的不同问题提出针对性解决方法。本文提供的一些思路和方法,对内源性药物的生物样品分析及生物等效性评价有重要意义。     

生物等效性评价及相关问题

生物等效性在新药研究、非专利药开发和药品上市后评价中具有重要作用。本文从实验方法学、药剂因素、内源性物质、活性代谢物、立体异构药物和高变异性药物等方面对生物等效性研究在技术指南及实际应用中需要特别注意的问题进行了具体论述,同时对生物药剂学分类系统等生物等效性研究的相关进展做了初步介绍。     

内源性药物生物等效性试验规程管理

作为机体内原有的物质,因自身调节机制,外源性成分、机体内源性物质水平等多种干扰因素,内源性药物生物等效性试验方案设计复杂,增加试验实施难度。本文从受试者的选择和管理、饮食饮水的控制、样本采集、安全监护方面,对试验实施的难点及特殊性进行分析,总结和探讨质量控制措施,对内源性药物生物等效性临床试验执行管理提供参考。     

特殊生物等效性评价方法介绍

本文我们将着重讨论一些特殊的仿制药品的生物等效性试验方法.此类药品包括代谢物具有生物活性类的药物、对映异构体、内源性物质、高变异性药物和局部作用药物等仿制药品.     

FDA药物体内生物等效性指导原则介绍

１９９５年出版的美国药典（ＵＳＰ）第２３版收载了美国食品药品管理局（ＦＤＡ）制定的药物体内生物等效性试验指导原则。该指导原则分为两部分，一部分是关于生物等效性试验的指导原则，另一部分是几个具体药物的生物利用度的实验规程。该指导原则由ＦＤＡ的基本药物办公室（ＯＧＤ）下属的生物等效性评价处（ＤｉｖｉｓｉｏｎｏｆＢｉｏｅｑｕｉｖａ－ｌｅｎｃｅ）制订。本文介绍第一部分内容。１引言美国ＯＧＤ下属的生物等效性评价处通常选用健康受试者比较试验制剂和参比制剂的体内吸收速率和吸收度来评价其生物等效性。一个标准的体…     

美国FDA《特定药物的生物等效性指导原则》 高变异性药物生物等效性指导原则调研

目的：研究美国食品药品管理局（FDA）《特定药物的生物等效性指导原则》对高变异性药物生物等效性研究相关规定,为我国仿制药质量和疗效一致性评价工作提供借鉴和帮助。方法：从剂型、给药方式、试验设计、受试者选择、给药条件、检测物质选择、豁免条件、体外溶出试验等多个方面对美国FDA公布的高变异性药物《特定药物的生物等效性指导原则》进行详细分析,并特别指出涉及我国仿制药质量与疗效一致性评价首批品种的高变异性药物。结果：美国FDA公布的涉及高变异性药物《特定药物的生物等效性指导原则》对具体化学仿制药的生物等效性评价从多个方面进行较为详细的规范,是对美国FDA相关生物等效性总则的补充和解读,对仿制药的发展有重要的推动作用。结论：在我国国家食品药品监督管理总局（CFDA）尚未颁布针对具体高变异性药物相关生物等效性指导原则的背景下,美国FDA《特定药物的生物等效性指导原则》中对高变异性药物相关规范对我国正在进行的仿制药质量和疗效一致性评价具有一定指导和借鉴意义。     

一致性评价中部分品种BCS分类不明确问题的解决方案

基于生物药剂学分类系统（BCS）的生物等效性豁免旨在减少对体内生物等效性研究的需求,即可以提供一种体内生物等效性的替代方法。基于BCS分类的生物等效性豁免需提供药物的溶解性、渗透性和体外溶出数据。主要论述药物溶解性、渗透性和体外溶出度测定的方法,希望建立药物相关特性的标准化、规范化测定流程,以保证企业在申报生物等效性豁免时,所提供的数据是准确、可靠的。     

体外群体生物等效性的统计分析方法

美国食品药品监督管理局指导原则表明原研药以及仿制药的生物等效性可用体内外数据或体内数据评价,而体外数据的评价方法依赖于体外群体生物等效性(PBE)方法。关于体外群体生物等效性的执行在美国食品药品监督管理部门出台的相关法案中已有明确要求,其运用越来越得到申办者的重视,但国内尚属起步阶段且未付诸实用。因此,本文通过介绍体外群体生物等效性的评价准则,判断界限,相关参数的讨论以及计算方法并列举实例说明,以期为国内群体生物等效性评价提供科学性参考。     

在线生物样品处理与LC-MS／MS联用新技术

1在线生物样品处理新技术概况 生物样品预处理是体内药物分析中的一个重要环节,也是分析中最困难、最繁琐的工作。在体内药物分析中,生物样品成分复杂,除被测组分外,往往含有大量内源性物质、代谢产物及共存药物等干扰物质,且被测组分的含量很低,要准确地测定生物样品中的药物浓度,必须除去妨碍测定的杂质,对样品进行预处理。     

内源性药物尿药生物等效性的试验设计和分析方法

目的以枸橼酸钾为例,介绍内源性药物的尿药生物等效性试验(BE)的设计和分析方法。方法根据FDA关于内源性药物BE研究相关指南及相关文献,以枸橼酸钾尿药BE研究为实例,探讨其研究流程、质量监控、剂量选择、样本采集以及数据分析等关键问题。结果在枸橼酸钾BE研究中,用单剂量、双交叉设计,对试验期间的饮食和活动进行标准化控制,用同一周期的给药前的内源性物质作为基线,校正尿药排泄量,最后用24 h尿药累积排泄量(Ae0-24h)、最大尿药排泄速率(Rmax)进行生物等效性分析。结论内源性药物BE研究,在设计上更为复杂,饮食和生理因素对结果影响大,需要基线校正,数据波动明显。尿药参数以Ae0-t和Rmax为主,而Tmax和T1/2没有实际意义。     

Assessing the bioequivalence of analogues of endogenous substances (��endogenous drugs��): considerations to optimize study design

BACKGROUND

Assessment of the bioequivalence of generic versions of certain reference drugs is complicated by the presence of endogenous levels of said compounds which cannot be distinguished from externally derived compound levels following drug administration. If unaccounted for, the presence of endogenous compound biases towards equivalence in bioequivalence studies of these drugs. Bioequivalence assessments may be complicated further as disposition of the exogenous analogue can be subject to various endogenous processes resulting in nonlinear pharmacokinetics. To overcome these inherent biases a number of different strategies have been employed.

AIMS

To critically review methods used to overcome confounding biases in bioequivalence studies of ‘endogenous’ drugs.

METHODS

A literature search of the EMBASE and PubMed databases was performed.

RESULTS

The following strategies were identified: ablation/modulation of baseline endogenous substance levels; recruitment of ‘substance-deficient’ populations; restriction of dietary intake of the relevant substance; standardization of conditions with the potential to affect relevant homeostatic mechanisms; correction for baseline substance levels; and administration of supra-therapeutic drug doses.

CONCLUSIONS

On the basis of this review key study design concepts, intended to optimize the design of future bioequivalence studies of these so-called ‘endogenous drugs’, are described. The dual stable isotope method, which could be used in a specific context, is also discussed.     

Bioequivalence Behind the Scenes

This paper focuses on some unresolved issues in bioequivalence. In these cases, intrinsic difficulties not clearly considered in EU and US FDA operating guidelines are involved. Careful attention to these issues and wide experience is required for tailoring specific bioequivalence study protocols. The aim is to reach a bioequivalence or bioinequivalence conclusion on the basis of test versus reference performance, avoiding less definite, borderline conclusions attributable to intrinsic difficulties in the specific study. In discussing these unresolved issues, the author suggests potential solutions, where possible, on the basis of personal experience and specific bibliographic references. The difference in titer between test and reference would normally be in the range of 95–105%, but in some cases it would fall within the expanded range of 90–120%. This would call for a titer normalization in establishing bioequivalence. A long elimination half-life (t1/2) [e.g. 50 days with amiodarone] calls for decision making about study design, namely crossover or parallel group design, the latter design being more adequate for drugs cleared with a t1/2 >10 days. Ethical problems concerning drug administration in healthy individuals (e.g. cytostatic agents in single dose, and other drugs in a repeated-dose regimen) is another issue. Drugs that produce unacceptable side effects in a repeated-dose regimen should be studied in healthy volunteers only in single dose. Drugs characterized by high variability require a high number of participants in the trial. Endogenous substances, most of which are eliminated faster than they are absorbed, have plasma concentration-time behavior not predictive of bioavailability. When the endogenous substance is cleared via urine, cumulative urinary excretion is a better evaluation parameter than area under the plasma concentration curve. In other cases a repeated dose regimen to steady state can allow post-dose concentration without baseline subtraction to be considered in assessing bioequivalence. Drugs producing one or more active metabolite(s) raise different opinions on whether bioequivalence should be assessed for the drug, metabolite(s) or both. All active moiety, namely parent drug and metabolite(s) should be assayed. Bioequivalence, however, should be assessed on only one substance, preferably the parent drug. In the presence of a prevalent active metabolite, this metabolite should be preferred in assessing bioequivalence. Insufficient sensitivity of the bioassay often compels an alternative solution, such as working in steady state or assaying the compound in urine. Polymorphic metabolism calls for a preliminary classification of volunteers in order to exclude poor metabolizers from bioequivalence studies of repeated-dose regimens. Topical application of drugs for topical effect calls for a therapeutic bioequivalence; this is because circulating concentrations are not an expression of activity, but only of safety. When there are several strengths of a given drug on the market, the replicator can profit from a waiver allowing only one strength to be checked when some conditions hold. Reversible metabolites should be considered active metabolites. This is because they can revert into the parent compound. Reversibility of metabolites is very common with endogenous substances, but it is also present with several exogenous substances. The approaches selected to solve bioequivalence problems must not conflict with US FDA and EU operating guidelines, which, however, in some instances do not overlap. This would mean that it is mandatory to operate on a case-by-case basis.     

Open questions on bioequivalence: some problems and some solutions.

This paper focuses on some specific situations where bioequivalence requires careful attention and tailored protocols in order to overcome intrinsic difficulties either marginally covered or fully neglected by operating guidelines. Some problems congregate with serious difficulties, namely high variability, very poorly absorbed drugs and endogenous substances with their own baseline. With endogenous substances, the dilemma faced is whether to subtract baseline from post-dose values in assessing bioequivalence. Either approach has intrinsic problems and is somewhat puzzling. In an attempt to resolve other existing problems, the most appropriate approach should be selected on a case-by-case basis, ensuring that the adopted procedure does not conflict with operating guidelines and scientific literature on the matter. Problematic cases include the management of trials with a predominant active metabolite, the absence of a reliable analytical bioassay, the availability of various strengths of the same drug on the market, a wide acceptability titre range, the management of studies on topical drugs that are devoid of systemic activity, the management of drugs that cannot be given for ethical reasons to healthy subjects or that may cause adverse events, especially when a steady state design is required. The parallel group study design appears to be more appropriate than the cross-over or the individual bioequivalence design in assessing drugs with a long half-life. Some pharmacokinetic and statistical analysis-related issues are also discussed such as the sequence/period interaction sometimes encountered in these trials, which, in the absence of the carry-over effect, does not bias the bioequivalence results and the need to process data with non-compartmental pharmacokinetic analysis.     

Bioequivalence of endogenous substances facing homeostatic equilibria: an example with potassium

Oral administration of endogenous substances in most cases results in negligible net increases in baseline plasma concentrations, associated with high variability. This poses the problem of their bioequivalence. Using the data obtained from a bioequivalence investigation of potassium aspartate (test vs reference formulation), the authors demonstrate the inconsistency of bioequivalence based on plasma concentrations and standard methods. Potassium aspartate was given orally at a dose of 15.8 mmoles to 12 healthy volunteers as test and reference values according to a two-period, two-formulation, two-sequence design. The individual net values of the area under the curve of plasma concentration (AUC) and cumulative urinary excretion (CUE), both obtained with the test formulation as post-dose minus baseline, were multiplied by 2, 3, 4, 5 and 6 and added to the baseline in order to simulate the administration of increasing single doses of the test, assuming dose-linear kinetics. Data generated with the test formulation were compared with original data of the reference according to 90% confidence intervals. With AUC, bioequivalence of test and reference formulations was demonstrated with 1 : 1, 2 : 1 and 3 : 1 test to reference dose ratios. With CUE only the 1 : 1 dose ratio comparison produced bioequivalence. The authors conclude that bioequivalence of endogenous substances conducted with standard procedures in most cases is a useless exercise. With potassium and more generally with drugs cleared via urine, urinary excretion would reflect the extent of absorption more faithfully than AUC.     

A perspective for biowaivers of human bioequivalence studies on the basis of the combination of the ratio of AUC to the dose and the biopharmaceutics classification system

The ratio of AUC to the dose (AUC/dose) was previously found as a parameter that predicts a risk of bioinequivalence of oral drug products. On the basis of the combination of this parameter and the biopharmaceutics classification system (BCS), a perspective for biowaivers of human bioequivalence studies is discussed. Databases of bioequivalence studies using immediate-release solid oral dosage forms were disclosed by 6 Japanese generic pharmaceutical companies, and the number of subjects required for demonstrating bioequivalence between generic and reference products was plotted as a function of AUC/dose for each BCS category. A small variation in the number of subjects was constantly observed in bioequivalence studies using dosage forms containing an identical BCS class 1 or class 3 drug, even though formulations of the generic product differ between companies. The variation was extremely enlarged when the drugs were substituted with BCS class 2 drugs. Rate-determining steps in oral absorption of highly water-soluble BCS class 1 and class 3 drugs are independent of formulations when there is no significant difference in the in vitro dissolution profiles between formulations. The small variation observed for both BCS categories indicates that the number of subjects converges into one value for each drug. Our analysis indicates the appropriateness of biowaiver of bioequivalence studies for immediate-release solid oral dosage forms containing not only BCS class 1 drugs but also class 3 drugs.     

A modified large sample approach in the assessment of population bioequivalence

The U.S. Food and Drug Administration (FDA) requires pharmaceutical companies to show bioequivalence between different formulations or generic companies to show bioequivalence between generic drugs and brand drugs before approval. In a recent FDA guidance on bioequivalence, new criteria were proposed for assessment of population and individual bioequivalence. In this article, computer simulation is used to compare a modified large sample (MLS) upper bound for the population bioequivalence ratio with the bootstrap upper bound recommended by the FDA. The comparison criteria are the ability to maintain the stated confidence level and the estimated power of tests based on these bounds.     

Impact of truncated area under the curve on failed bioequivalence studies: a computer simulation analysis

The common measures used in a bioequivalence study are area under the curve (AUC) and the maximum plasma concentration. Estimation of AUC requires frequent blood samples. For long half-life drugs, sampling for long periods of time may become cumbersome. To resolve this issue some investigators have suggested the use of truncated AUC in bioequivalence studies for long half-life drugs. The suggested length of time for the truncated AUC is 72 hours. Many studies have been conducted to show that truncated AUC till 72 hours is a suitable approach. However, the suitability of truncated AUC for failed bioequivalence study has not been demonstrated. This report of simulated plasma concentration versus time data evaluates the suitability of truncated AUC for failed bioequivalence study of two hypothetical drugs. The results of the study indicate that the truncated approach for the estimation of the AUC for long half-life drugs in bioequivalence studies may be useful but it also increases the probability of accepting drugs as being bioequivalent when they are not.