In Vitro Permeation Test Studies for Topical Drug Products Submitted in ANDAs Guidance for Industry

工業(yè)指南中ANDAs申請(qǐng)?zhí)峤坏耐庥弥苿┑捏w外滲透試驗(yàn)研究

U.S. Department of Health and Human Services Food and Drug Administration Center for Drug Evaluation and Research (CDER) October 2022 Generic Drugs

美國(guó)衛(wèi)生與公眾服務(wù)部食品和藥物管理局藥物評(píng)估與研究中心（CDER）2022年10月仿制藥

IINTRODUCTION

This guidance is intended to assist applicants who are submitting abbreviated new drug applications (ANDAs) for liquid-based and/or other semisolid products applied to the skin, including integumentary and mucosal (e.g., vaginal) membranes, which are hereinafter called “topical products.” Because of the complex route of delivery associated with these products, which are typically locally acting, and the potential complexity of certain formulations, topical products (other than topical solutions) are classified as complex products. This guidance provides recommendations for in vitro permeation test (IVPT) studies comparing a proposed generic (test) topical product and its reference standard (RS) for the purpose of supporting a demonstration of bioequivalence (BE) to the reference listed drug (RLD). The reference standard ordinarily is the RLD

本指南旨在幫助申請(qǐng)人提交用于皮膚的液體和/或其他半固體產(chǎn)品申請(qǐng)ANDA，包括皮膚和粘膜（如陰道），以下稱為“局部產(chǎn)品”。由于這些產(chǎn)品復(fù)雜的遞送途徑，通常是局部起效的，以及某些制劑的潛在復(fù)雜性，外用制劑（外用溶液除外）被歸類為復(fù)雜制劑。本指南為體外滲透試驗(yàn)（IVPT）研究提供了建議，該研究比較了擬申報(bào)仿制制劑及其參考制劑（RS），以支持證明與參比制劑的（RLD）生物等效性（BE）。參比制劑通常為RLD
 

This guidance does not address drug products that are administered via ophthalmic, otic, nasal, inhalation, oral, or injection-based routes, or that are transdermal or topical delivery systems (including products known as patches, topical patches, or extended release films).

本指南不適用于通過(guò)眼、耳、鼻、吸入、口服或注射途徑給藥的藥物，也不適用于透皮或局部給藥系統(tǒng)（包括貼片、局部貼片或緩釋膜）。

It is beyond the scope of this guidance to discuss specific topical products to which this guidance applies. FDA recommends that applicants consult this guidance and any relevant product-specific guidances (PSGs) and any other relevant guidances for industry, when considering the design and conduct of IVPT studies that, in conjunction with other studies, as deemed necessary, may be appropriate to support a demonstration that a proposed generic topical product and its RLD are bioequivalent. FDA also recommends that applicants routinely refer to FDA’s guidance web pages, because additional guidances may become available that could assist in the development of a generic topical product.

本指南是不討論適用該指南的特定外用制劑的指南。FDA建議申請(qǐng)人在考慮IVPT研究的設(shè)計(jì)和實(shí)施時(shí)，參考本指南和任何相關(guān)的特定產(chǎn)品指南（PSG）以及任何其他相關(guān)的行業(yè)指南，如有必要，結(jié)合其他研究，IVPT研究可能適合支持建議的通用外用制劑及其RLD具有生物等效性的證明。FDA還建議申請(qǐng)人定期查閱FDA的指導(dǎo)網(wǎng)頁(yè)，因?yàn)榭赡軙?huì)有更多的指導(dǎo)，有助于仿制藥的開發(fā)。

In general, FDA’s guidance documents do not establish legally enforceable responsibilities. Instead, guidances describe the Agency’s current thinking on a topic and should be viewed only as recommendations, unless specific regulatory or statutory requirements are cited. The use of the word should in Agency guidance means that something is suggested or recommended, but not required.

一般來(lái)說(shuō)，F(xiàn)DA的指導(dǎo)文件并沒有確立法律上可強(qiáng)制執(zhí)行的責(zé)任。相反，指南描述了機(jī)構(gòu)目前對(duì)某一主題的想法，并且應(yīng)僅視為建議，除非引用了具體的監(jiān)管或法定要求。在機(jī)構(gòu)指南中使用“應(yīng)該”一詞意味著有人建議或建議，但不是必須的。

II. BACKGROUND

This guidance has been developed as part of FDA’s “Drug Competition Action Plan,” which, in coordination with the Generic Drug User Fee Amendments (GDUFA) program and other FDA activities, is intended to increase competition in the market place for prescription drugs, facilitate the entry of high-quality and affordable generic drugs, and improve public health.

本指南是作為FDA“藥品競(jìng)爭(zhēng)行動(dòng)計(jì)劃”的一部分制定的，該計(jì)劃配合GDUFA計(jì)劃和其他FDA舉措，旨在促進(jìn)處方藥市場(chǎng)的競(jìng)爭(zhēng)，促進(jìn)高質(zhì)量和實(shí)惠的藥品的進(jìn)入市場(chǎng)，并改善公眾醫(yī)療。

The Federal Food, Drug, and Cosmetic Act (FD&C Act) generally requires an ANDA to contain, among other things, information to show that the proposed generic drug product 1) is the same as the RLD with respect to the active ingredient(s), conditions of use, route of administration, dosage form, strength, and labeling (with certain permissible differences) and 2) is bioequivalent to the RLD. Thus, an ANDA will not be approved if the information submitted in the ANDA is insufficient to show that the test product is bioequivalent to the RLD.

《聯(lián)邦食品、藥品和化妝品法案》（FD&C法案）通常要求ANDA包含合適的信息，說(shuō)明：仿制藥與RLD相比1）具有相同活性成分、使用條件、給藥途徑、劑型、規(guī)格和標(biāo)簽（允許存在某些差異），和2）與RLD具有生物等效性。因此，如果ANDA中提交的信息不足以證明自制制劑與RLD具有生物等效性，則ANDA將不予批準(zhǔn)。

An IVPT study may be used to assess the rate and extent to which a drug (i.e., an active ingredient) from a topical product becomes available at or near a site of action in the skin, and may be used to characterize and compare the rate and extent of bioavailability for a drug from a test topical product and RS. The IVPT flux profiles resemble pharmacokinetic profiles and can be analyzed using unique IVPT endpoints that are somewhat analogous to the pharmacokinetic endpoints of maximum concentration (Cmax) and the area under the concentration-time curve(AUC). Yet, IVPT studies characterize the rate and extent of absorption, not the distribution, metabolism and excretion that occurs in vivo. Therefore, while it is relevant to characterize the kinetics of topical drug bioavailability monitored by IVPT studies, the use in this guidance of the term “cutaneous pharmacokinetics” should not be construed to embody all aspects of pharmacokinetics—only those related to the absorption component that directly controls the rate and extent to which a topically applied drug becomes available locally at the site of action. This guidance focuses on general considerations and recommendations for the method development, method validation, and conduct of IVPT studies that are submitted in ANDAs and intended to support a demonstration of BE.

IVPT研究可用于評(píng)估局部產(chǎn)品中的藥物（即活性成分）在皮膚作用部位或附近可用的速率和程度，并可用于表征和比較測(cè)試局部產(chǎn)品和RS中藥物的生物利用率和程度。IVPT通量曲線類似于藥代動(dòng)力學(xué)曲線，可以使用*的IVPT終點(diǎn)進(jìn)行分析，這些終點(diǎn)與最大濃度（Cmax）的藥代動(dòng)力學(xué)終點(diǎn)和濃度-時(shí)間曲線下的面積（AUC）有些相似。然而，IVPT研究表征的是吸收的速率和程度，而不是體內(nèi)的分布、代謝和排泄。因此，雖然表征IVPT研究監(jiān)測(cè)的局部藥物生物利用度的動(dòng)力學(xué)是相關(guān)的，本指南中使用的術(shù)語(yǔ)“皮膚藥代動(dòng)力學(xué)”不應(yīng)被解釋為僅體現(xiàn)與吸收成分相關(guān)的藥代動(dòng)力學(xué)的所有方面，吸收成分直接控制局部應(yīng)用藥物在作用部位局部可用的速率和程度。本指南側(cè)重于在ANDA中提交的用于支持BE演示的方法開發(fā)、方法驗(yàn)證和IVPT研究的一般考慮和建議。

III. IVPT METHOD DEVELOPMENT

The development of an IVPT method that is suitable to support a demonstration of BE for a specific topical product routinely involves a systematic series of exploratory studies. Inappropriate or insufficient efforts to develop an IVPT method that is suitable for its intended purpose increases the likelihood that the subsequent IVPT validation, pilot, and pivotal studies will ultimately be inadequate to support a demonstration of BE. By contrast, appropriate and systematic IVPT method development studies help to identify IVPT study designs and protocol (method) parameters which reliably produce flux profiles that can facilitate a comparison of the cutaneous pharmacokinetics of a drug delivered topically to the skin from test topical products and RSs.

IVPT方法的開發(fā)適用于支持特定局部產(chǎn)品的BE演示，通常涉及一系列系統(tǒng)的探索性研究。開發(fā)適合其預(yù)期目的的IVPT方法的努力不當(dāng)或不足，增加了后續(xù)IVPT驗(yàn)證、試點(diǎn)和關(guān)鍵研究最終不足以支持BE演示的可能性。相比之下，適當(dāng)和系統(tǒng)的IVPT方法開發(fā)研究有助于確定IVPT研究設(shè)計(jì)和方案（方法）參數(shù)，這些設(shè)計(jì)和方案能夠可靠地產(chǎn)生通量曲線，從而有助于比較從測(cè)試局部產(chǎn)品和RS局部遞送至皮膚的藥物的皮膚藥代動(dòng)力學(xué)。

A detailed and well-organized IVPT method development report should be submitted in an ANDA to show how the IVPT method was optimized, and to support a demonstration that the method parameters selected for the IVPT are appropriate or necessary, particularly in situations where the method parameters are different from the methods recommended in this guidance). The Agency’s interest in reviewing the method development report is to understand why specific IVPT method parameters were selected and whether the resulting IVPT method is suitably sensitive and reproducible. This method development report should clearly indicate/distinguish the method parameters used for each set of data, illustrate the efforts made to optimize the IVPT method, and demonstrate that the method parameters selected for the IVPT are appropriate.

應(yīng)在ANDA中提交一份詳細(xì)且組織良好的IVPT方法開發(fā)報(bào)告，以說(shuō)明IVPT方法是如何優(yōu)化的，并支持為IVPT選擇的方法參數(shù)是適當(dāng)或必要的，特別是在方法參數(shù)與本指南中推薦的方法不同的情況下）。機(jī)構(gòu)審查方法開發(fā)報(bào)告的目的是了解為什么選擇了特定的IVPT方法參數(shù)，以及所得IVPT方法是否具有適當(dāng)?shù)拿舾行院椭噩F(xiàn)性。該方法開發(fā)報(bào)告應(yīng)明確指出/區(qū)分每組數(shù)據(jù)使用的方法參數(shù)，說(shuō)明為優(yōu)化IVPT方法所做的努力，并證明為IVPT選擇的方法參數(shù)是合適的。

Applicants are encouraged to use the recommendations in this guidance, and if an applicant elects to use methods that are different from those recommended in this guidance, the IVPT method development report should demonstrate why it is scientifically justified to use an alternative approach than what is recommended in this guidance to optimize the IVPT method. Some examples of recommended procedures are described in subsequent sections, to help applicants identify circumstances when information should be submitted in the ANDA to explain why a different procedure was utilized.

鼓勵(lì)申請(qǐng)人使用本指南中的建議，如果申請(qǐng)人選擇使用與本指南中建議的方法不同的方法，IVPT方法開發(fā)報(bào)告應(yīng)說(shuō)明為什么使用本指南推薦的替代方法來(lái)優(yōu)化IVPT方法在科學(xué)上是合理的。建議程序的一些示例在隨后的章節(jié)中描述，以幫助申請(qǐng)人確定應(yīng)在ANDA中提交信息的情況，以解釋為什么使用不同的程序。

A. IVPT Method Parameters

All relevant parameters of the final IVPT method should be summarized (e.g., in a table) and submitted in the ANDA. Also, information should be provided to briefly explain the choice of the final IVPT method parameters like the equipment (e.g., a vertical diffusion cell (VDC)), skin source (e.g., cadaver), skin type (e.g., posterior torso), skin preparation (e.g., dermatomed), skin barrier integrity test (e.g., trans-epidermal water loss (TEWL) measurement), skin barrier integrity test acceptance criteria (e.g., < 15 grams/meter2 /hour (g/m2 /hr)), topical product dose amount (e.g., 15 milligrams/centimeter2 (mg/cm2 )), dose duration (e.g., 6 hours), study duration (e.g., 24 hours, 48 hours, etc.), receptor solution sampling times (e.g., 1, 2, 4, 6, 8, 12, 16, 20, and 24 hours), etc.

應(yīng)總結(jié)最終IVPT方法的所有相關(guān)參數(shù)（例如，在表格中），并在ANDA中提交。此外，應(yīng)提供信息，以簡(jiǎn)要解釋最終IVPT方法參數(shù)的選擇，如設(shè)備（例如，垂直擴(kuò)散池（VDC））、皮膚源（例如，尸體）、皮膚類型（例如，后軀干）、皮膚準(zhǔn)備（例如，皮膚科）、皮膚屏障完整性測(cè)試（例如，經(jīng)表皮水分損失（TEWL）測(cè)量）、，皮膚屏障完整性測(cè)試驗(yàn)收標(biāo)準(zhǔn)（例如，<15克/米2/小時(shí)（g/m2/小時(shí)））、局部產(chǎn)品劑量（例如，15毫克/厘米2（mg/cm2））、劑量持續(xù)時(shí)間（例如，6小時(shí)）、研究持續(xù)時(shí)間（如，24小時(shí)、48小時(shí)等）、受體溶液取樣時(shí)間（例如1、2、4、6、8、12、16、20和24小時(shí)）等。《IVPT測(cè)試中的皮膚研究》

B. IVPT Method Considerations

The choice of some IVPT method parameters like the equipment, skin source, skin type, skin preparation, and skin barrier integrity test procedures may be based upon investigator experience or convenience, like the availability of specific equipment or instrumentation in a laboratory, established tissue supply agreements, or other logistical considerations. However, if the chosen IVPT method parameters do not appear to be well-suited for a specific IVPT method, it is the applicant’s responsibility to systematically evaluate alternative method parameters, and ultimately, to validate that the IVPT method parameters chosen are suitable for the intended purpose. The recommended procedures for IVPT method validation are detailed in section IV of this guidance.

一些IVPT方法參數(shù)的選擇，如設(shè)備、皮膚來(lái)源、皮膚類型、皮膚準(zhǔn)備和皮膚屏障完整性測(cè)試程序，可能基于研究人員的經(jīng)驗(yàn)或便利性，如實(shí)驗(yàn)室中特定設(shè)備或儀器的可用性、既定的組織供應(yīng)協(xié)議或其他后勤考慮。然而，如果選定的IVPT方法參數(shù)似乎不適合特定的IVPT法，則申請(qǐng)人有責(zé)任系統(tǒng)地評(píng)估備選方法參數(shù)，并最終驗(yàn)證選定的IVPT方法參數(shù)是否適合預(yù)期用途。IVPT方法驗(yàn)證的推薦程序詳見本指南第四節(jié)。

The choice of other IVPT method parameters like the topical product dose amount, dose duration, study duration (which may be longer than the dose duration), sampling schedule, sampling procedures, receptor solution composition, and sample analytical method may be different for each IVPT method, and such parameters of IVPT methods should be systematically developed, optimized, and/or validated for the relevant topical product, as appropriate. The IVPT method development studies should characterize how differences in these method parameters influence the resulting IVPT flux profile so that optimal study conditions can be objectively selected from among those evaluated.

其他IVPT方法參數(shù)的選擇，如局部產(chǎn)品劑量、劑量持續(xù)時(shí)間、研究持續(xù)時(shí)間（可能長(zhǎng)于劑量持續(xù)時(shí)間）、取樣計(jì)劃、取樣程序、受體溶液組成和樣品分析方法，對(duì)于每種IVPT方法可能有所不同，應(yīng)系統(tǒng)開發(fā)、優(yōu)化IVPT方法的此類參數(shù)，和/或?qū)ο嚓P(guān)局部產(chǎn)品進(jìn)行驗(yàn)證。IVPT方法開發(fā)研究應(yīng)描述這些方法參數(shù)的差異如何影響最終的IVPT通量分布，以便從評(píng)估的條件中客觀選擇最佳研究條件。

The selection of the dose amount used in the study should be assessed for each IVPT method based upon studies performed during IVPT method development. Different dose amounts may be compared in parallel on replicate skin sections from the same set of donors to optimize the dose amount for the IVPT study. Considerations for selecting an optimal dose amount may include (1) the consistency with which the dose can be applied (potentially using different dispensing and/or spreading techniques), (2) the reproducibility of the flux profiles, (3) the influence of dose amount and dose duration on the shape of the flux profile, and (4) the approximate range of drug concentrations in receptor solution samples at different time points (relative to the sample analytical method limits of quantification).

應(yīng)根據(jù)IVPT方法開發(fā)期間進(jìn)行的研究，評(píng)估每種IVPT方法在研究中使用的劑量選擇?？梢栽趤?lái)自同一組供體的復(fù)制皮膚切片上平行比較不同劑量，以優(yōu)化IVPT研究的劑量。選擇最佳劑量的考慮因素可以包括（1）可以施加劑量的一致性（可能使用不同的分配和/或散布技術(shù)），（2）通量分布的再現(xiàn)性，（3）劑量和劑量持續(xù)時(shí)間對(duì)通量分布形狀的影響，和（4）不同時(shí)間點(diǎn)受體溶液樣品中藥物濃度的近似范圍（相對(duì)于樣品分析方法的定量限值）。

The selected sampling schedule and study duration should be sufficient to characterize the cutaneous pharmacokinetics of the drug, which ideally includes a sufficiently complete flux profile to identify the maximum (peak) flux and a decline in the flux thereafter across multiple subsequent time points. A dose that remains on the skin for the duration of the study may continue to deliver the drug for a sustained period and may not necessarily exhibit a suitable decline in the flux at later time points. In such instances, it may be appropriate to develop an IVPT method that involves wiping off the applied dose after a suitable duration on the skin and continuing to monitor the receptor solution for an extended period thereafter, during which the decline in the flux profile can be characterized. The sampling frequency should be selected to provide a suitable resolution for the flux profile, and a minimum of eight non-zero sampling time points is recommended across the study duration (e.g., 48 hours).

所選擇的采樣時(shí)間表和研究持續(xù)時(shí)間應(yīng)足以表征藥物的皮膚藥代動(dòng)力學(xué)，理想情況下，包括足夠完整的通量曲線，以確定最大（峰值）通量以及隨后在多個(gè)后續(xù)時(shí)間點(diǎn)的通量下降。在研究持續(xù)時(shí)間內(nèi)留在皮膚上的劑量可能會(huì)持續(xù)給藥一段時(shí)間，并且不一定會(huì)在以后的時(shí)間點(diǎn)出現(xiàn)適當(dāng)?shù)牧髁肯陆怠Ｔ谶@種情況下，開發(fā)IVPT方法可能是合適的，該方法包括在皮膚上適當(dāng)?shù)某掷m(xù)時(shí)間后擦去施用的劑量，并在此后的較長(zhǎng)時(shí)間內(nèi)繼續(xù)監(jiān)測(cè)受體溶液，在此期間可以表征通量分布的下降。應(yīng)選擇采樣頻率，以便為通量分布提供合適的分辨率，并且建議在整個(gè)研究持續(xù)時(shí)間內(nèi)（例如，48小時(shí)）至少八個(gè)非零采樣時(shí)間點(diǎn)。

C. IVPT Method Procedures and Controls

Suitable technical procedures and control parameters should be established during method development. These may include procedures for preparing and mounting the skin on the diffusion cell in a consistent manner, determining the instrument settings that regulate the skin surface temperature within the specified range, performing the barrier integrity test appropriately, controlling the accuracy and precision of the dose amount dispensed on each skin section.

在方法開發(fā)過(guò)程中，應(yīng)建立適當(dāng)?shù)募夹g(shù)程序和控制參數(shù)。這些可以包括以一致的方式制備皮膚并將其安裝在擴(kuò)散池上的程序，確定將皮膚表面溫度調(diào)節(jié)在規(guī)定范圍內(nèi)的儀器設(shè)置，適當(dāng)?shù)剡M(jìn)行屏障完整性測(cè)試，控制分配到每個(gè)皮膚部分的劑量的準(zhǔn)確性和精密度。

For example, a dosing procedure may be developed that uses a positive displacement pipette to dispense a volumetrically controlled amount of a topical product, targeting the deposition on the skin of a certain mass (e.g., 15 mg/cm2 ) of topical product. If the inner diameter of the orifice in the dosing compartment of the diffusion cell is 15 millimeters (mm), and the effective dose area is ~1.77 cm2 , this would indicate a target dose of ~26.5 mg of topical product per diffusion cell. Experiments during method development may establish that, based upon the density of the topical product, a specific volumetric setting on a specific model of positive displacement pipette with a specific pipette tip repeatedly dispenses ~27.5 mg of topical product (e.g., characterized by multiple replicate pipette dispensations into a weigh boat on a fine balance). This pipette setting may be optimal for a dosing procedure where the dose spreading instrument, like the flat bottom of a high performance liquid chromatography (HPLC) glass vial, or the rounded end of a glass rod or capillary tube, is subsequently used to spread the dispensed dose evenly upon the skin section mounted in the diffusion cell, and where repeatedly weighing the dose-spreading instrument before and after the dose spreading indicates that the residual topical product remaining on the bottom of the glass vial after the dose spreading reproducibly amounts to ~1.0 mg of topical product (indicating that ~26.5 mg of the topical product would reproducibly be dosed to each skin section). Such characterizations of the technical procedures and control parameters for the IVPT method, like the reproducibility of the dosing procedure, should be established during method development and may not need to be demonstrated thereafter each time the same procedure is used.

例如，可以開發(fā)一種劑量程序，其使用正位移移液管來(lái)分配體積控制量的局部產(chǎn)品，靶向一定質(zhì)量（例如，15mg/cm2）的局部產(chǎn)品在皮膚上的沉積。如果擴(kuò)散池給藥室的孔口內(nèi)徑為15毫米（mm），有效劑量面積約為1.77 cm2，則表明每個(gè)擴(kuò)散池的目標(biāo)劑量為約26.5 mg局部產(chǎn)品。方法開發(fā)過(guò)程中的實(shí)驗(yàn)可以確定，基于外用產(chǎn)品的密度，具有特定移液管的特定型號(hào)正移液管的特定體積設(shè)置可重復(fù)分配約27.5 mg外用產(chǎn)品（例如，以多次重復(fù)移液管分配為特征，將其分配到精密天平上的稱量舟中）。這種移液管設(shè)置對(duì)于劑量分配過(guò)程可能是最佳的，其中劑量分配儀器，如高效液相色譜（HPLC）玻璃瓶的平底，或玻璃棒或毛細(xì)管的圓形端，隨后用于將分配的劑量均勻地分配到安裝在擴(kuò)散池中的皮膚部分上，并且其中在劑量散布之前和之后重復(fù)稱量劑量散布儀器表明，在劑量散布之后殘留在玻璃瓶底部的殘留局部產(chǎn)品可重復(fù)地達(dá)到約1.0mg局部產(chǎn)品（表明將可重復(fù)地將約26.5mg局部產(chǎn)品施用到每個(gè)皮膚部分）。IVPT方法的技術(shù)程序和控制參數(shù)的此類特征，如給藥程序的再現(xiàn)性，應(yīng)在方法開發(fā)過(guò)程中確定，此后可能不需要在每次使用相同程序時(shí)進(jìn)行證明。‍

‍D. ‍‍IVPT Skin Barrier Integrity Testing: Common Methods

The technical procedures for the skin barrier integrity test should be established during IVPT method development. Three types of barrier integrity tests are common, however, there are currently no applicable compendial standard protocols or acceptance criteria for any of these three types of human skin barrier integrity tests. Nonetheless, recommended parameters for the three common types of barrier integrity tests are discussed below.

應(yīng)在IVPT方法開發(fā)期間制定皮膚屏障完整性測(cè)試的技術(shù)程序。三種類型的屏障完整性測(cè)試是常見的，然而，目前沒有適用于這三種類型人體皮膚屏障完整性檢測(cè)的藥典標(biāo)準(zhǔn)協(xié)議或驗(yàn)收標(biāo)準(zhǔn)。盡管如此，下面討論了三種常見類型的屏障完整性測(cè)試的推薦參數(shù)。IVPT測(cè)試中的皮膚研究

1. Trans-Epidermal Water Loss Skin Barrier Integrity Test

A TEWL skin barrier integrity test involves a measurement near the outer surface of the skin of the rate at which water (vapor) is fluxing through the skin barrier from the underside of the skin section. For the test, the skin section is mounted in a diffusion cell (e.g., clamped in place between the donor and receptor compartments), with the underside of the skin in contact with the receptor solution in the receptor compartment (e.g., phosphate buffered saline, pH 7.4), and equilibrated to a skin surface temperature of 32°C ± 1°C. If skin sections are cut large enough to cover the flange of the diffusion cell in which they are mounted, then after they have equilibrated for several hours at a skin surface temperature of 32°C ± 1°C, it may be feasible to gently remove the donor compartment without disrupting a skin section’s adherence to the lower flange of the diffusion cell, thereby allowing the TEWL probe to be placed directly on the skin surface, instead of being placed atop the donor compartment. Typically, a minimum of three replicate measurements are made on each skin section, which are recorded after the measurements have stabilized.

TEWL皮膚屏障完整性測(cè)試包括在皮膚外表面附近測(cè)量水（蒸汽）從皮膚部分下側(cè)流過(guò)皮膚屏障的速率。對(duì)于試驗(yàn)，將皮膚部分安裝在擴(kuò)散池中（例如，夾在供體和受體室之間的適當(dāng)位置），皮膚下側(cè)與受體室中的受體溶液（例如，磷酸鹽緩沖鹽水，pH 7.4）接觸，并平衡至32°C±1°C的皮膚表面溫度。如果皮膚部分被切割得足夠大，足以覆蓋其所安裝的擴(kuò)散池的法蘭，那么在皮膚表面溫度為32°C±1°C的條件下平衡數(shù)小時(shí)后，可以在不破壞皮膚部分與擴(kuò)散池下法蘭的粘附性的情況下輕輕移除供體隔室，從而允許TEWL探針直接放置在皮膚表面上而不是放置在供體隔室的頂部。通常，在每個(gè)皮膚部分上至少進(jìn)行三次重復(fù)測(cè)量，并在測(cè)量穩(wěn)定后進(jìn)行記錄。（下圖，皮膚測(cè)試用經(jīng)皮水分流失測(cè)量?jī)x）

Commercially available devices to measure TEWL may differ in design and operational principles. The TEWL measured by devices with certain designs (e.g., an open chamber versus a closed chamber) may be relatively more susceptible to the influence of environmental conditions. Therefore, environmental temperature and humidity are typically controlled as precisely as possible (e.g., a temperature range of 21°C ± 2°C and a humidity range of 50% ± 20% relative humidity are ideal, if feasible). More precise control of the relative humidity (e.g., in the range of 40% – 50%) may reduce the variability of TEWL measurements for devices with certain designs. Certain designs of measurement probes and adapters for in vitro use are available by the manufacturers of TEWL devices, and may be appropriate to use. Inconsistency in the diameters for the measurement probe chamber, the measurement probe orifice, the in vitro adapters, and the skin area being measured, as well as variation in the distance of the probe sensor(s) from the skin surface, potentially because of the (variable) height of donor compartments (when applicable), could increase the variability of TEWL measurements. Inconsistent control of the alignment of the TEWL measurement device in relation to the donor compartment and/or the skin section may also increase the variability of TEWL measurements. Also, the TEWL measured by devices with certain designs may be relatively more susceptible to the influence of heat transfer from the hand that holds the probe. Applicants should follow relevant instructions in the manufacturer’s user manual for the specific TEWL measurement device used.

用于測(cè)量TEWL的市售設(shè)備在設(shè)計(jì)和操作原理上可能有所不同。由具有特定設(shè)計(jì)（例如，開放室與封閉室）的裝置測(cè)量的TEWL可能相對(duì)更容易受到環(huán)境條件的影響。因此，通常盡可能精確地控制環(huán)境溫度和濕度（例如，如果可行，理想的溫度范圍為21°C±2°C，濕度范圍為50%±20%相對(duì)濕度）。更精確地控制相對(duì)濕度（例如，在40%–50%的范圍內(nèi)）可以減少具有特定設(shè)計(jì)的設(shè)備的TEWL測(cè)量的可變性。TEWL裝置的制造商可提供用于體外使用的測(cè)量探針和適配器的某些設(shè)計(jì)，并且可能適合使用。測(cè)量探頭室、測(cè)量探頭孔口、體外適配器和被測(cè)皮膚面積的直徑不一致，以及探頭傳感器與皮膚表面的距離變化，可能是由于供體隔室的（可變）高度（如適用），可能會(huì)增加TEWL測(cè)量的變異性。TEWL測(cè)量裝置相對(duì)于供體隔室和/或皮膚部分的對(duì)準(zhǔn)的不一致控制也可能增加TEWL測(cè)定的可變性。此外，由具有特定設(shè)計(jì)的設(shè)備測(cè)量的TEWL可能相對(duì)更容易受到握住探頭的手的熱傳遞的影響。申請(qǐng)人應(yīng)遵循制造商用戶手冊(cè)中關(guān)于所用特定TEWL測(cè)量設(shè)備的相關(guān)說(shuō)明。

No more than approximately 15 grams of water per square meter per hour (i.e., ≤ 15 g/m2 /hr) could be a reasonable skin barrier integrity acceptance (cutoff) criterion for a TEWL barrier integrity test on human torso or thigh skin; if this was selected as the cutoff criterion, skin sections with a TEWL > 15 g/m2 /hr would fail the test. Skin sections that fail a barrier integrity test should not be dosed, but may serve as non-dosed control skin sections. A higher cutoff (e.g., ≤ 20 g/m2 /hr) may also be reasonable if justified by experimental data demonstrating that the selected acceptance criterion appropriately discriminates skin sections with a compromised barrier integrity from those with a competent barrier integrity.

每平方米每小時(shí)不超過(guò)約15克水（例如，≤ 15g/m2/hr）可能是人體軀干或大腿皮膚上TEWL屏障完整性測(cè)試的合理皮膚屏障完整性驗(yàn)收（截止）標(biāo)準(zhǔn)；如果將此作為截止標(biāo)準(zhǔn)，TEWL>15 g/m2/hr的皮膚切片將無(wú)法通過(guò)測(cè)試。未通過(guò)屏障完整性測(cè)試的皮膚部分不應(yīng)給藥，但可作為未給藥的對(duì)照皮膚部分。較高的截止（例如，≤ 20g/m2/hr）也可能是合理的，如果實(shí)驗(yàn)數(shù)據(jù)證明所選驗(yàn)收標(biāo)準(zhǔn)適當(dāng)?shù)貐^(qū)分屏障完整性受損的皮膚部分和屏障完整性合格的皮膚部分。

However, TEWL measurements for skin sections with a competent barrier integrity can vary depending upon the TEWL measurement device, the manner in which it is operated, and the environmental conditions (e.g., higher ambient humidity or greater distance from the skin surface may decrease the value of the TEWL measurement). Precise control of environmental and device/operational factors can minimize variability in TEWL measurements. Therefore, the technical procedures for measuring TEWL should be optimized during IVPT method development (or based upon prior optimization in the laboratory performing the test). Also, the TEWL measurement device should be appropriately calibrated (by the manufacturer, and for some devices, also before each set of tests). Applicants may provide information about the relevant calibration procedures specified by the manufacturer for the specific TEWL device used; this can be submitted in the ANDA along with the IVPT method development report, to support the appropriateness of the technical procedures established by the laboratory for TEWL measurements. When a TEWL barrier integrity test is used in any study phase (IVPT method development, pilot study, validation, and/or pivotal study) the ambient laboratory temperature and humidity during the TEWL barrier integrity test should be monitored and reported.

然而，對(duì)于具有合格屏障完整性的皮膚部分的TEWL測(cè)量可以根據(jù)TEWL測(cè)試設(shè)備、其操作方式和環(huán)境條件而變化（例如，較高的環(huán)境濕度或離皮膚表面更大的距離可能會(huì)降低TEWL的測(cè)量值）。精確控制環(huán)境和設(shè)備/操作因素可以最大限度地減少TEWL測(cè)量的變化。因此，應(yīng)在IVPT方法開發(fā)過(guò)程中優(yōu)化TEWL測(cè)量的技術(shù)程序（或基于實(shí)驗(yàn)室進(jìn)行試驗(yàn)的事先優(yōu)化）。此外，TEWL測(cè)量設(shè)備應(yīng)進(jìn)行適當(dāng)校準(zhǔn)（由制造商進(jìn)行校準(zhǔn)，對(duì)于某些設(shè)備，也應(yīng)在每組測(cè)試之前進(jìn)行校準(zhǔn)）。申請(qǐng)人可提供制造商為所用特定TEWL裝置規(guī)定的相關(guān)校準(zhǔn)程序的信息；這可以與IVPT方法開發(fā)報(bào)告一起提交在ANDA中，以支持實(shí)驗(yàn)室為TEWL測(cè)量建立的技術(shù)程序的適當(dāng)性。當(dāng)TEWL屏障完整性測(cè)試用于任何研究階段（IVPT方法開發(fā)、試點(diǎn)研究、驗(yàn)證和/或關(guān)鍵研究）時(shí)，應(yīng)監(jiān)測(cè)和報(bào)告TEWL阻隔完整性測(cè)試期間的實(shí)驗(yàn)室環(huán)境溫度和濕度。

2.Tritiated Water Skin Barrier Integrity Test

An example of a recommended approach to a tritiated water skin barrier integrity test would be to mount the skin in a diffusion cell (e.g., clamped in place between the donor and receptor compartments) and allow it to equilibrate to a skin surface temperature of 32°C ± 1°C with the stratum corneum exposed to the air in the donor compartment and the underside of the skin in contact with the receptor solution (e.g., phosphate buffered saline, pH 7.4).

氚化水-皮膚屏障完整性測(cè)試的推薦方法的一個(gè)例子是將皮膚安裝在擴(kuò)散池中（例如，夾在供體和受體室之間的適當(dāng)位置），并使其平衡至32°C±1°C的皮膚表面溫度，使角質(zhì)層暴露在供體室中的空氣中，皮膚下側(cè)與受體溶液（例如磷酸鹽緩沖鹽水，pH 7.4）。

A small amount of tritiated water (sufficient to cover the entire surface of the skin section) would be briefly dosed on the stratum corneum. This dose of tritiated water would be left on the surface for a precisely controlled and relatively brief period (e.g., 5 minutes) after which it would be removed from the skin surface (e.g., using a pipette to remove the bulk volume and then an absorbent low lint laboratory tissue to gently blot dry). The receptor solution would then be sampled at a precise duration after the removal of the tritiated water from the skin surface (e.g., 30 minutes after the removal of the 5-minute dose of tritiated water from the skin surface).

將少量的氚水（足以覆蓋皮膚部分的整個(gè)表面）短暫地施加于角質(zhì)層。該劑量的氚水將在表面上停留一段精確控制且相對(duì)較短的時(shí)間（例如，5分鐘），之后將其從皮膚表面去除（例如，使用移液管去除大量體積，然后使用實(shí)驗(yàn)室吸水紙輕輕吸干）。然后在從皮膚表面去除氚水之后的精確持續(xù)時(shí)間（例如，從皮膚表面除去5分鐘劑量的氚水后30分鐘）對(duì)受體溶液進(jìn)行取樣。

While the entire volume of the receptor compartment may be removed and replenished, typically only an aliquot of the receptor solution (e.g., phosphate buffered saline, pH 7.4) is transferred to a suitable volume of scintillation fluid for counting. The volume of the aliquot typically depends upon the type of scintillation fluid used and the maximum amount of aqueous fluid that is suitable to mix with the scintillation fluid. A scintillation counter is then used to quantify the amount of radioactivity in the aliquot sampled, which can be used to calculate the amount of tritiated water that permeated into the larger (entire) volume of receptor solution; the calculation is performed using the specific activity of the tritiated water to equate a given amount of radioactivity to the equivalent volume of tritiated water that permeated per square centimeter of skin surface area.

雖然可以移除并補(bǔ)充整個(gè)體積的受體室，但通常僅將一等分的受體溶液（例如，磷酸鹽緩沖鹽水，pH 7.4）轉(zhuǎn)移到合適體積的閃爍液中進(jìn)行計(jì)數(shù)。等分試樣的體積通常取決于所使用的閃爍流體的類型以及適合與閃爍流體混合的水性流體的最大量。然后，使用閃爍計(jì)數(shù)器來(lái)定量取樣的等分試樣中的放射性量，可用于計(jì)算滲入較大（整個(gè)）體積的受體溶液中的氚水的量；使用氚水的比活度進(jìn)行計(jì)算，以將給定的放射性量等同于每平方厘米皮膚表面積滲透的氚水當(dāng)量體積。

Approximately 1.5 equivalent(eq.) microliter (µL) of tritiated water per cm2 (i.e., ~1.5 eq. µL/cm2 or ~1.5 eq. mg/cm2 ) would be a reasonable skin barrier integrity acceptance (cutoff) criterion for a tritiated water barrier integrity test that involves a 5-minute dose followed by a 30- minute sampling duration (i.e., sampling 30 minutes after dose removal) on human torso or thigh skin. Skin sections with a tritiated water test result of > 1.5 eq. mg/cm2 would fail the test and be excluded from the population of skin sections dosed with the topical product; skin sections that fail a barrier integrity test should not be dosed, but may serve as non-dosed control skin sections. Other acceptance criteria may also be reasonable if justified by experimental data demonstrating that the selected acceptance criterion appropriately discriminates skin sections with a compromised barrier integrity from those with a competent barrier integrity.

人體軀干或大腿皮膚上氚水屏障完整性測(cè)試的合理皮膚屏障完整性驗(yàn)收（截止）標(biāo)準(zhǔn)是每cm2約1.5當(dāng)量（eq）微升（µL）的氚水（例如，約1.5 eq. µL/cm2或約1.5 eq. mg/cm2）。該測(cè)試涉及5分鐘劑量，隨后30分鐘采樣持續(xù)時(shí)間（即，去除劑量后30分鐘采樣）。氚水測(cè)試結(jié)果大于1.5 eq. mg/cm2的皮膚切片判定為未通過(guò)測(cè)試，并被排除在使用局部產(chǎn)品的皮膚切片人群之外；未通過(guò)屏障完整性測(cè)試的皮膚部分不應(yīng)給藥，但可作為未給藥的對(duì)照皮膚部分。如果實(shí)驗(yàn)數(shù)據(jù)證明所選驗(yàn)收標(biāo)準(zhǔn)能夠適當(dāng)區(qū)分屏障完整性受損的皮膚部分和屏障完整性合格的皮膚部分，則其他驗(yàn)收標(biāo)準(zhǔn)也可能合理。（下圖，放射性測(cè)試用閃點(diǎn)計(jì)數(shù)器）

When calculating the results for a tritiated water barrier integrity test, it may be important to account for the surface area dosed. For example, if using an acceptance criterion of 1.5 eq. mg/cm2 with a diffusion cell that has an orifice diameter of 15 mm and a skin surface area of 1.77 cm2 , the mass of tritiated water that would be calculated to have permeated into the receptor compartment would be ~2.7 eq. mg/cm2 of tritiated water.

當(dāng)計(jì)算氚水屏障完整性測(cè)試的結(jié)果時(shí)，給藥表面積是一個(gè)很重要的影響因素。例如，如果使用1.5 eq. mg/cm2的驗(yàn)收標(biāo)準(zhǔn)和孔直徑為15 mm、皮膚表面積為1.77 cm2的擴(kuò)散池，則計(jì)算出滲入受體室的氚水質(zhì)量為約2.7 eq. mg/cm2的氚。

3.Electrical Based Skin Barrier Integrity Tests

There are several variations of electrical based skin barrier integrity tests that report the test result as a measure of the resistance, conductance, or a related electrical concept that characterizes the bulk flow of electrical current across the skin. Transepithelial electrical resistance tests involving the skin may be referred to more specifically as Trans-Epidermal Electrical Resistance (TEER) skin barrier integrity tests. The test results may be described in units of conductance, which is the reciprocal of resistance. Electrical based skin barrier integrity tests often use instruments that are designed to measure the inductance (L), capacitance (C), and resistance (R) of electronic circuits or electrical components; these instruments are commonly known as LCR meters and have different settings (test parameters) that can be adjusted.

基于電的皮膚屏障完整性測(cè)試有幾種變體，將測(cè)試結(jié)果報(bào)告為電阻、電導(dǎo)或相關(guān)電概念的測(cè)量值，以表征穿過(guò)皮膚的整體電流。涉及皮膚的經(jīng)上皮電阻測(cè)試可以更具體地稱為經(jīng)表皮電阻（TEER）皮膚屏障完整性測(cè)試。測(cè)試結(jié)果可用電導(dǎo)單位表示，電導(dǎo)是電阻的倒數(shù)?；陔姎獾钠つw屏障完整性測(cè)試通常使用設(shè)計(jì)用于測(cè)量電子電路或電氣部件的電感（L）、電容（C）和電阻（R）的儀器；這些儀器通常被稱為L(zhǎng)CR儀表，并具有可調(diào)整的不同設(shè)置（測(cè)試參數(shù)）。

An example of a recommended approach to a TEER skin barrier integrity test would be to mount the skin in a diffusion cell (e.g., clamped in place between the donor and receptor compartments) and allow it to equilibrate to a skin surface temperature of 32°C ± 1°C with the stratum corneum exposed to the air in the donor compartment and the underside of the skin in contact with an ionic solution (e.g., phosphate buffered saline, pH 7.4).

TEER皮膚屏障完整性測(cè)試推薦方法的一個(gè)示例是將皮膚安裝在擴(kuò)散池中（例如，夾在供體和受體隔室之間的適當(dāng)位置），并使其恒定在32°C±1°C的皮膚表面溫度，角質(zhì)層暴露于供體隔室中的空氣中，皮膚下側(cè)與離子溶液接觸（例如，磷酸鹽緩沖鹽水，pH 7.4）。

A small amount of the ionic solution (sufficient to cover the entire surface of the skin section) would be briefly dosed on the stratum corneum. Then, one lead/electrode from an LCR meter would be placed in contact with the solution in the receptor compartment while the other lead/electrode would be placed in contact with the solution in the donor compartment. After measuring the resistance across the skin (e.g., in kΩ, normalized for area, noting that resistance is inversely proportional to area) the solution in the donor compartment would be removed and the skin surface would be gently blotted dry with an absorbent low lint laboratory tissue. The skin (still mounted in the diffusion cell) would then be allowed to equilibrate with the dry air above for a sufficient duration to normalize the hydration state of the stratum corneum before being dosed with the test topical product or RS.

將少量離子溶液（足以覆蓋皮膚部分的整個(gè)表面）短暫地施加到角質(zhì)層上。然后，LCR測(cè)量?jī)x的一根導(dǎo)線/電極與受體室中的溶液接觸，而另一根導(dǎo)線或電極與供體室中的液體接觸。測(cè)量皮膚上的電阻（例如，kΩ，按面積歸一化，注意電阻與面積成反比）后，除去供體室中的溶液，并用吸水性低皮棉實(shí)驗(yàn)室紙巾輕輕吸干皮膚表面。然后讓皮膚（仍安裝在擴(kuò)散池中）與上面的干燥空氣平衡足夠長(zhǎng)的時(shí)間，以使角質(zhì)層的水合狀態(tài)正常化。

The results for a TEER skin barrier integrity test can vary substantially depending on the LCR meter settings (e.g., frequency) and the technical procedures used for the test. The acceptance criterion for a specific electrical based skin barrier integrity test method may be justified by experimental data demonstrating that the selected acceptance criterion appropriately discriminates skin sections with a compromised barrier integrity from those with a competent barrier integrity.

TEER皮膚屏障完整性測(cè)試的結(jié)果可能會(huì)根據(jù)LCR儀表設(shè)置（例如頻率）和測(cè)試使用的技術(shù)程序而產(chǎn)生較大差異?？梢酝ㄟ^(guò)實(shí)驗(yàn)數(shù)據(jù)證明，特定基于電氣的皮膚屏障完整性測(cè)試方法的驗(yàn)收標(biāo)準(zhǔn)可以適當(dāng)區(qū)分屏障完整性受損的皮膚部分和屏障完整性合格的皮膚部分。

E.IVPT Skin Barrier Integrity Testing: General Considerations

There are three general considerations for the development or adoption of technical procedures for any skin barrier integrity test method during IVPT method development:

在IVPT方法開發(fā)過(guò)程中，開發(fā)或采用任何皮膚屏障完整性測(cè)試方法的技術(shù)程序有三個(gè)一般考慮因素：

i. The technical procedures should not irreversibly alter the skin barrier. It may be acceptable to temporarily alter the hydration state of the stratum corneum by briefly depositing an aqueous solution on the surface of the skin, as long as sufficient time is afforded for the hydration of the stratum corneum to normalize before dosing of the topical product. The procedure described above for a brief (e.g., 5-minute) exposure of the skin surface to tritiated water followed by a 30-minute duration during which the hydration state of the stratum corneum is re-equilibrating would likely be appropriate. By contrast, a 30-minute exposure of the skin surface to an aqueous solution for an electricalbased test method, followed within 5 minutes by dosing of the topical product, may not be appropriate without further characterization of the influence of the hydration state of the stratum corneum on the discrimination sensitivity of the skin to differences in topical bioavailability. Similarly, if a portable lamp were placed close to the skin to improve visibility while study procedures were being performed, the heat from the lamp may alter the local (micro)environment of the skin in a manner that is not representative of the ambient environmental conditions in the laboratory; this should be avoided.

i、 技術(shù)程序不應(yīng)給皮膚屏障造成不可逆轉(zhuǎn)地改變。只要有足夠的時(shí)間使角質(zhì)層的水合作用正?；?，就可以通過(guò)在皮膚表面短暫沉積水溶液來(lái)暫時(shí)改變角質(zhì)層的水合狀態(tài)。上述將皮膚表面短暫（例如5分鐘）暴露于氚水，然后持續(xù)30分鐘，在此期間角質(zhì)層的水合狀態(tài)重新平衡的過(guò)程可能是合適的。相比之下，如果不進(jìn)一步表征角質(zhì)層的水合狀態(tài)對(duì)皮膚對(duì)局部生物利用度差異的辨別敏感性的影響，將皮膚表面暴露于基于電的測(cè)試方法的水溶液中30分鐘，然后在5分鐘內(nèi)給藥局部產(chǎn)品可能是不合適的。類似地，如果在研究時(shí)將便攜式燈靠近皮膚放置以提高能見度，燈的熱量可能會(huì)改變皮膚的局部(微)環(huán)境，而這種方式不能代表實(shí)驗(yàn)室中的環(huán)境條件，應(yīng)該避免。

ii. The acceptance criterion should be a cutoff value for the test result, at which a skin section fails the test. Skin sections that fail a barrier integrity test should not be dosed but may serve as non-dosed control skin sections. Skin sections with a passing barrier integrity test result may be considered to have a competent barrier integrity and may be dosed. This acceptance criterion should be selected based upon an understanding of the distribution of test results (among multiple replicate skin sections from multiple donors) for the specific barrier integrity test procedure performed with the specific type and preparation of skin under conditions relevant to the IVPT pivotal studies submitted in the ANDA. The intention of the barrier integrity test is to identify (and exclude) skin sections whose barrier integrity (intactness) is compromised. The intent is not to reduce the inherent variability in barrier function (permeability) in human skin that is representative of real variation in the human population. Also, the relative permeability of the skin to a drug from a topical product may not necessarily correlate with the permeability of the skin to water, and therefore, constraining the variability of the skin permeability to water (using a stricter acceptance criterion that excludes a larger number of skin sections) may not necessarily reduce the variability in the IVPT study results.

ii．驗(yàn)收標(biāo)準(zhǔn)應(yīng)為皮膚部分未通過(guò)測(cè)試結(jié)果時(shí)的臨界值。未通過(guò)屏障完整性測(cè)試的皮膚部分不應(yīng)給藥，但可作為未給藥的對(duì)照皮膚部分。屏障完整性測(cè)試結(jié)果合格的皮膚部分可被視為具有合格的屏障完整性，并可給藥。應(yīng)基于對(duì)測(cè)試結(jié)果分布(來(lái)自多個(gè)供體的多個(gè)重復(fù)皮膚切片)的理解來(lái)選擇此接受標(biāo)準(zhǔn)，該測(cè)試程序是在與ANDA中提交的IVPT關(guān)鍵研究相關(guān)的條件下，使用特定類型和制備的皮膚進(jìn)行的。屏障完整性測(cè)試的目的是識(shí)別（并排除）屏障完整性（完整性）受損的皮膚部分。其目的并不是減少人體皮膚屏障功能（滲透性）的固有變異性，這這代表了人群的真實(shí)變化。此外，皮膚對(duì)局部產(chǎn)品的藥物的相對(duì)滲透性可能不一定與皮膚對(duì)水的滲透性相關(guān)，因此，限制皮膚對(duì)水滲透性的可變性（使用更嚴(yán)格的接受標(biāo)準(zhǔn)，排除更多的皮膚部分）不一定會(huì)降低IVPT研究結(jié)果的可變性。

iii. The acceptance criterion should be able to discriminate skin sections with a compromised barrier integrity. This may be demonstrated by measuring the barrier integrity of skin sections mounted and equilibrated in a diffusion cell before and after deliberately compromising the skin barrier (e.g., by repeatedly using adhesive tape to strip away increasing amounts of the stratum corneum, piercing the skin several times with a 30 gauge needle, or using other physical or chemical insults to damage the skin barrier). Based upon the acceptance criterion selected, the test result for skin sections that pass the test before being damaged should fail the test after the damage.

iii.驗(yàn)收標(biāo)準(zhǔn)應(yīng)能夠區(qū)分屏障完整性受損的皮膚部分。這可以通過(guò)在故意破壞皮膚屏障之前和之后測(cè)量在擴(kuò)散池中安裝和平衡的皮膚部分的屏障完整性來(lái)證明（例如，通過(guò)反復(fù)使用膠帶剝離越來(lái)越多的角質(zhì)層，用30號(hào)針刺穿皮膚幾次，或使用其他物理或化學(xué)侮辱來(lái)破壞皮膚屏障）。根據(jù)選定的接受標(biāo)準(zhǔn)，受損前測(cè)試結(jié)果合格的皮膚，破損后的測(cè)試結(jié)果應(yīng)不合格。

F.Differences Between IVPT Method Development and Validation

1.Optimization of an IVPT Method Prior to Advancing to IVPT Method Validation

Different study designs and method parameters may be evaluated during the IVPT method development phase. For example, if the selected study parameters initially involve a dose duration of 48 hours and a study duration of 48 hours, and the flux profile is measurable, but it is not feasible to identify the maximum (peak) flux and a decline in the flux thereafter across multiple subsequent time points, then it may be appropriate to evaluate other study parameters as part of the IVPT method development. For example, a different target dose of the topical product and/or a longer sampling duration may be evaluated. An alternate study design may involve a shorter dose duration (e.g., 4–6 hours) after which the applied dose is removed from the skin, and the receptor solution continues to be sampled across a study duration that is sufficient to identify the maximum (peak) flux and a decline in the flux thereafter across multiple subsequent time points. While shorter dose durations can help to improve the shape of IVPT flux profiles, the removal of the topical product dose from the skin surface can be challenging and often requires its own method development and optimization. Also, the design of sensitivity studies for such an IVPT study design may require a more sophisticated understanding of IVPT studies. While reasonable efforts should be made to develop an IVPT method that produces a well-defined maximum (peak) flux and a decline in the flux thereafter across multiple subsequent time points, this may not be feasible for certain topical products even with study durations of 96 hours, or, at least, may not be feasible to produce reliably in all donors. In such circumstances, the IVPT method development report should detail the systematic efforts made to optimize the IVPT method.

在IVPT方法開發(fā)階段，可以評(píng)估不同的研究設(shè)計(jì)和方法參數(shù)。例如，如果所選的研究參數(shù)最初涉及48小時(shí)的劑量持續(xù)時(shí)間和48小時(shí)的研究持續(xù)時(shí)間，并且通量分布是可測(cè)量的，但無(wú)法確定最大（峰值）通量和通量下降，則可以評(píng)估其他研究參數(shù)作為IVPT方法開發(fā)的一部分。例如，可以評(píng)估局部產(chǎn)品的不同目標(biāo)劑量和/或更長(zhǎng)的取樣持續(xù)時(shí)間。另一種研究設(shè)計(jì)可能涉及更短的劑量持續(xù)時(shí)間（例如，4-6小時(shí)），之后去除皮膚的上樣劑量，并且在研究持續(xù)時(shí)間內(nèi)繼續(xù)對(duì)受體溶液進(jìn)行采樣，該研究持續(xù)時(shí)間足以確定最大（峰值）通量以及隨后在多個(gè)后續(xù)時(shí)間點(diǎn)的通量下降。雖然更短的劑量持續(xù)時(shí)間有助于改善IVPT通量分布曲線的形狀，但從皮膚表面去除局部產(chǎn)品劑量可能具有挑戰(zhàn)性，通常需要自己開發(fā)方法和優(yōu)化。此外，此類IVPT研究設(shè)計(jì)的敏感性研究設(shè)計(jì)可能需要對(duì)IVPT研究有更復(fù)雜的理解。盡管應(yīng)盡合理努力開發(fā)一種IVPT方法，該方法可在隨后的多個(gè)時(shí)間點(diǎn)產(chǎn)生明確定義的最大（峰值）通量和通量下降，但這對(duì)于某些局部產(chǎn)品來(lái)說(shuō)可能不可行，即使研究持續(xù)時(shí)間為96小時(shí)，或者至少不可能在所有供體中穩(wěn)定地產(chǎn)生。在這種情況下，IVPT方法開發(fā)報(bào)告應(yīng)詳細(xì)說(shuō)明為優(yōu)化IVPT方法所做的系統(tǒng)努力。

The IVPT method development studies, being exploratory in nature, are often performed using a sample analytical method that is not validated (e.g., an HPLC or ultrahigh performance liquid chromatography (UPLC) method, often involving mass spectrometry (MS)); also, IVPT method development studies are often conducted in a manner that is not compatible with a quality management system which would otherwise make the evidence generated suitable to support valid conclusions. Such method development studies would not be suitable to demonstrate the validity of an IVPT method, or associated results. Therefore, although it may appear to be redundant, certain experiments performed during IVRT method development may need to be repeated during IVPT method validation, using appropriate controls, like a validated analytical method and procedures that are compatible with a suitable quality management system.

IVPT方法開發(fā)研究本質(zhì)上是探索性的，通常使用未經(jīng)驗(yàn)證的樣品分析方法（例如，HPLC或超高效液相色譜（UPLC）方法，通常涉及質(zhì)譜（MS））；此外，IVPT方法開發(fā)研究通常與質(zhì)量管理體系不兼容，否則會(huì)使生成的證據(jù)適合支持有效結(jié)論。此類方法開發(fā)研究不適合證明IVPT方法或相關(guān)結(jié)果的有效性。因此，盡管IVRT方法開發(fā)過(guò)程中進(jìn)行的某些實(shí)驗(yàn)看起來(lái)可能是多余的，但在IVPT方法驗(yàn)證過(guò)程中可能需要重復(fù)進(jìn)行，使用適當(dāng)?shù)目刂疲缃?jīng)過(guò)驗(yàn)證的與合適的質(zhì)量管理系統(tǒng)兼容的分析方法和程序。。

It is important to clearly segregate and consistently identify those experiments and results that were part of IVPT method development separately from those that were part of IVPT method validation. It is also important to consistently identify all relevant method parameters and experimental conditions/controls for each set of IVPT results. Information in the method development report should clearly identify/distinguish when the results for apparently similar sets of experiments may have been obtained using different method parameters. Method development reports should clarify which sets of diffusion cells were run in parallel or separately (e.g., on separate days). In addition, the sample analytical method parameters used to analyze the samples from each set of IVPT experiments should be specified, and the report should indicate whether or not the sample analytical method was validated (either at the time of sample analysis or subsequently).

重要的是要明確的將IVPT方法開發(fā)中的實(shí)驗(yàn)和結(jié)果與IVPT方法驗(yàn)證中的實(shí)驗(yàn)與結(jié)果分開，并加以一致識(shí)別。對(duì)于每組IVPT結(jié)果，一致識(shí)別所有相關(guān)的方法參數(shù)和實(shí)驗(yàn)條件/對(duì)照同樣重要。方法開發(fā)報(bào)告中的信息應(yīng)清楚地識(shí)別/區(qū)分什么時(shí)候明顯相似的實(shí)驗(yàn)結(jié)果可能是使用不同的方法參數(shù)獲得的。方法開發(fā)報(bào)告應(yīng)闡明哪些擴(kuò)散測(cè)試池組是平行或單獨(dú)運(yùn)行的（例如，在不同的日子）。此外，應(yīng)規(guī)定用于分析每組IVPT實(shí)驗(yàn)樣品的樣品分析方法參數(shù)，報(bào)告應(yīng)說(shuō)明樣品分析方法是否經(jīng)過(guò)驗(yàn)證（在樣品分析時(shí)或隨后）。

IV.IVPT METHOD VALIDATION

When all the relevant parameters of the IVPT method have been established, a pilot study should be performed using the final IVPT method and using a validated sample analytical method. The purpose of the pilot study is to validate the suitability of the selected IVPT method parameters by demonstrating that the performance characteristics of the IVPT method are appropriate to compare the cutaneous pharmacokinetics of a drug delivered topically from a test product and RS. The results from the pilot study, thereby, support the systematic validation of the IVPT method, which proceeds as a distinct study phase following IVPT method development.

當(dāng)IVPT方法的所有相關(guān)參數(shù)都已確定后，應(yīng)使用最終IVPT方法和經(jīng)驗(yàn)證的樣品分析方法進(jìn)行試點(diǎn)研究。試點(diǎn)研究的目的是通過(guò)證明IVPT方法的性能特征適用于比較在測(cè)試產(chǎn)品和RS中局部傳遞的藥物的皮膚藥代動(dòng)力學(xué)，來(lái)驗(yàn)證所選IVPT方法參數(shù)的適用性。因此，試點(diǎn)研究結(jié)果支持IVPT方法系統(tǒng)驗(yàn)證，這在IVPT方法開發(fā)之后作為一個(gè)特殊的研究階段進(jìn)行。

The results from this IVPT pilot study can help to estimate the number of donors that may be needed to adequately power the IVPT pivotal study. In addition to the test topical product and RS evaluated in the pilot study, a parallel assessment should be performed with a third topical product or formulation that is known or designed to be different from the RS, to validate the selectivity of the IVPT method to discriminate differences in bioavailability. The IVPT pilot study results should be plotted with error bars, comparing the permeation profiles for the three treatment groups in the pilot study. Separate plots should be prepared for average flux results and average cumulative permeation results. These data can be used to support specific IVPT method validation parameters (e.g., permeation profile and range).

IVPT試點(diǎn)研究的結(jié)果有助于估計(jì)可能需要的供體數(shù)量，以充分支持IVPT關(guān)鍵研究。除了試點(diǎn)研究中評(píng)估的試驗(yàn)外用產(chǎn)品和RS外，還應(yīng)使用已知或設(shè)計(jì)與RS不同的第三種外用產(chǎn)品或配方進(jìn)行平行評(píng)估，以驗(yàn)證IVPT方法對(duì)區(qū)分生物利用度差異的選擇性。IVPT試點(diǎn)研究結(jié)果應(yīng)繪制誤差條，比較中試研究中三個(gè)治療組的滲透曲線。應(yīng)為平均通量結(jié)果和平均累積滲透結(jié)果繪制單獨(dú)的圖。這些數(shù)據(jù)可用于支持特定的IVPT方法驗(yàn)證參數(shù)（例如滲透圖譜和范圍）。

A pilot IVPT study performed with multiple skin donors (e.g., 4–6 skin donors) and a minimum of four replicate skin sections per donor per treatment group is recommended. As skin from an increasing number of donors is evaluated in the pilot study, the accuracy of the estimated number of donors needed to adequately power the IVPT pivotal study may improve. While skin from the same donors evaluated in the pilot study may also be used in the IVPT pivotal study, the results from the pilot study should not be combined with the results from the IVPT pivotal study for the purpose of statistical analysis.

建議對(duì)多個(gè)皮膚供者（例如4-6個(gè)皮膚捐贈(zèng)者）進(jìn)行IVPT試驗(yàn)研究，每個(gè)治療組每個(gè)供者至少四個(gè)重復(fù)皮膚切片。隨著越來(lái)越多的供者的皮膚在試點(diǎn)研究中得到評(píng)估，為IVPT關(guān)鍵研究提供足夠動(dòng)力所需捐獻(xiàn)者估計(jì)數(shù)量的準(zhǔn)確性可能會(huì)提高。雖然在試點(diǎn)研究中評(píng)估的同一供者的皮膚也可用于IVPT關(guān)鍵研究，但試點(diǎn)研究的結(jié)果不應(yīng)與IVPT關(guān)鍵性研究的結(jié)果結(jié)合起來(lái)進(jìn)行統(tǒng)計(jì)分析。

The equipment, methodologies, and study conditions used in the IVPT pilot study (and the eventual IVPT pivotal study) should be appropriately validated or qualified. If an applicant elects to use equipment, methodologies, or study conditions that are different from those recommended in this guidance, the applicant should demonstrate why it was necessary and scientifically justified to do so. Detailed protocols and well-controlled study procedures are recommended to ensure the precise control of dosing, sampling, and other IVPT study parameters, as well as potential sources of experimental bias.

IVPT試點(diǎn)研究（以及最終的IVPT關(guān)鍵研究）中使用的設(shè)備、方法和研究條件應(yīng)經(jīng)過(guò)適當(dāng)驗(yàn)證或鑒定。如果申請(qǐng)人選擇使用與本指南中建議的不同的設(shè)備、方法或研究條件，申請(qǐng)人應(yīng)證明這樣做的必要性和科學(xué)合理性。建議采用詳細(xì)的方案和控制良好的研究程序，以確保精確控制給藥、取樣和其他IVPT研究參數(shù)，以及實(shí)驗(yàn)偏差的潛在來(lái)源。

The validation of the IVPT method should incorporate specific qualifications and controls (described below), performed using a validated sample analytical method, as applicable. The qualification of an IVPT method parameter refers to the process of defining what attributes make it suitable to perform its function in the IVPT method. For example, when repeated measurements of the temperature at the surface of skin mounted in a diffusion cell demonstrate that an IVPT equipment can maintain the skin surface temperature in the range of 32°C ± 1°C, the results can support a demonstration that the equipment is qualified to perform its function in an IVPT method for which a method parameter is the control of skin surface temperature in the range of 32°C ± 1°C across the relevant study duration.

IVPT方法的驗(yàn)證應(yīng)包括具體的限定條件和控制（如下所述），如適用，使用經(jīng)驗(yàn)證的樣品分析方法進(jìn)行。IVPT方法參數(shù)的限定指的是定義哪些屬性適合在IVPT方法中執(zhí)行其功能的過(guò)程。例如，當(dāng)對(duì)安裝在擴(kuò)散池中的皮膚表面溫度重復(fù)測(cè)量的結(jié)果表明IVPT設(shè)備可以將皮膚表面溫度保持在32°C±1°C的范圍內(nèi)時(shí)，結(jié)果可以證明該設(shè)備有資格在IVPT方法中執(zhí)行其功能，其中方法參數(shù)是在相關(guān)研究持續(xù)時(shí)間內(nèi)將皮膚表面溫度控制在32°C±1°C范圍內(nèi)。

A.Equipment Qualification

Suitable equipment for the IVPT method includes various models of VDCs and flow-through diffusion cells. The operating principles and specific test procedures differ among the various equipment; relevant procedures from the manufacturer may be used for installation, operational, and performance qualifications. The laboratory qualification of each diffusion cell should, at minimum, include 1) measurements of the diffusional area of the orifices of the donor and receptor compartments between which the skin is mounted, 2) the empirically measured volume of the receptor solution compartment in each VDC or the empirically measured outflow tube length for each flow-through diffusion cell, 3) the stability of the temperature measured at the skin surface (e.g., 32°C ± 1°C) across a relevant duration (e.g., 48 hours), and 4) the rate of stirring or agitation in VDCs, or the flow rate for flow-through diffusion cells, as applicable.

適用于IVPT方法的設(shè)備包括各種型號(hào)的VDC和流通擴(kuò)散池。不同設(shè)備的工作原理和具體測(cè)試程序不同；制造商的相關(guān)程序可用于安裝、操作和性能鑒定。每個(gè)擴(kuò)散池的實(shí)驗(yàn)室鑒定至少應(yīng)包括：1）測(cè)量皮膚安裝在供體和受體隔間的孔口擴(kuò)散面積，2）每個(gè)VDC中受體溶液隔間的經(jīng)驗(yàn)測(cè)量體積或每個(gè)流通擴(kuò)散池的經(jīng)驗(yàn)測(cè)量流出管長(zhǎng)度，3）在相關(guān)持續(xù)時(shí)間（例如48小時(shí)）內(nèi)在皮膚表面測(cè)量的溫度（例如32°C±1°C）的穩(wěn)定性，以及4）VDC中的攪拌或攪拌速率，或流通擴(kuò)散池的流速（如適用）。

If information related to the diffusional area of the orifices and the volume of the receptor solution compartment for each diffusion cell is available from the manufacturer, that information should be provided for each relevant diffusion cell, in addition to the empirical measurements made by the laboratory performing the IVPT studies. The equipment should control the diffusion cell temperature so that the skin surface temperature is verified to be stable (e.g., 32°C ± 1°C) for each diffusion cell before dosing (e.g., measured by a calibrated infrared thermometer), and monitored periodically throughout the duration of the experiment by repeatedly measuring the skin surface temperature of a non-dosed control diffusion cell that is run in parallel with the other replicate dosed diffusion cells and connected to the same water bath or thermoregulation system.

如果制造商提供了與每個(gè)擴(kuò)散池的孔口擴(kuò)散面積和受體溶液隔室體積相關(guān)的信息，則除了實(shí)驗(yàn)室進(jìn)行IVPT研究的經(jīng)驗(yàn)測(cè)量之外，還應(yīng)提供每個(gè)相關(guān)擴(kuò)散池的信息。設(shè)備應(yīng)控制擴(kuò)散池溫度，以便在給藥前驗(yàn)證每個(gè)擴(kuò)散池的皮膚表面溫度穩(wěn)定（例如，32°C±1°C）（例如，通過(guò)校準(zhǔn)的紅外溫度計(jì)測(cè)量），并在整個(gè)實(shí)驗(yàn)期間通過(guò)重復(fù)測(cè)量非劑量控制擴(kuò)散池的皮膚表面溫度進(jìn)行周期性監(jiān)測(cè)，該擴(kuò)散池與其他重復(fù)劑量給藥擴(kuò)散池并聯(lián)運(yùn)行并連接到相同的水浴或溫度調(diào)節(jié)系統(tǒng)。

B.Membrane (Skin) Qualification

Excised human skin is recommended as the membrane for the IVPT study. The validity of each skin section dosed in the study should be qualified using an appropriate test procedure to evaluate the stratum corneum barrier integrity. Acceptable barrier integrity tests may be based upon tritiated water permeation, TEWL, or electrical impedance/conductance measured across the skin. The test parameters and acceptance criteria used for the skin barrier integrity test should be justified for the specific method and instrumentation that is used during the study. The skin from all donors whose skin is included in the study should be prepared in a consistent manner and dermatomed to a relatively consistent thickness, within limits specified in the study protocol. The skin thickness should be measured and reported for each skin section included in the study. The assignment of replicate skin sections from a donor to each treatment group should be randomized, as feasible. It is acceptable to balance the distribution of skin thicknesses in each treatment group (test topical product or RS) by a procedure specified in the study protocol.

建議將切除的人體皮膚作為IVPT研究的膜。通過(guò)適當(dāng)?shù)臏y(cè)試程序來(lái)評(píng)估角質(zhì)層屏障的完整性，來(lái)鑒定研究中使用的每個(gè)皮膚切片的有效性?？山邮艿钠琳贤暾詼y(cè)試可基于氚水滲透、TEWL或皮膚上測(cè)量的電阻抗/電導(dǎo)。用于皮膚屏障完整性測(cè)試的測(cè)試參數(shù)和驗(yàn)收標(biāo)準(zhǔn)應(yīng)針對(duì)研究期間使用的特定方法和儀器進(jìn)行驗(yàn)證。研究中包括的所有捐獻(xiàn)者的皮膚應(yīng)以一致的方式制備，并在研究方案中規(guī)定的范圍內(nèi)保持相對(duì)一致的皮膚厚度。應(yīng)測(cè)量并報(bào)告研究中每個(gè)皮膚切片的皮膚厚度。在可行的情況下，應(yīng)將供體的復(fù)制皮膚切片隨機(jī)分配給每個(gè)治療組。也可以通過(guò)研究方案中規(guī)定的程序來(lái)平衡每個(gè)治療組（測(cè)試局部產(chǎn)品或RS）的皮膚厚度分布。

C.Receptor Solution Qualification

The composition and pH of the receptor solution used for the IVPT study should be qualified in relation to its compatibility with the skin as well as the stability and solubility of the drug in that receptor solution. The stability of the drug in the receptor solution samples should be validated as part of the receptor sample analytical method validation. The solubility of the drug in the IVPT receptor solution should be empirically determined in triplicate, to illustrate that the solubility of the drug in the receptor solution exceeds the highest sample concentration in the IVPT pivotal study, ideally by an order of magnitude. The solubility of the drug in the IVPT receptor solution should be sufficient to characterize the higher amounts of drug permeating from the increased drug delivery condition evaluated in the IVPT sensitivity assessment during IVPT method validation.

用于IVPT研究的受體溶液的組成和pH應(yīng)符合其與皮膚的相容性以及藥物在該受體溶液中的穩(wěn)定性和溶解度。受體溶液樣品中藥物的穩(wěn)定性應(yīng)作為受體樣品分析方法驗(yàn)證的一部分進(jìn)行驗(yàn)證。藥物在IVPT受體溶液中的溶解度應(yīng)根據(jù)經(jīng)驗(yàn)確定三份，以說(shuō)明藥物在受體溶液中溶解度超過(guò)IVPT關(guān)鍵研究中的最高樣品濃度，理想情況下為一個(gè)數(shù)量級(jí)。藥物在IVPT受體溶液中的溶解度應(yīng)足以表征IVPT方法驗(yàn)證期間IVPT敏感性評(píng)估中評(píng)估的藥物遞送條件增加導(dǎo)致的藥物滲透量增加。

The inclusion of 0.1% polyoxyethylene[20]oleyl ether (also known as Oleth-20, Volpo-20, or Brij-20; CAS number 9004-98-2) is recommended to enhance the solubility of physiological buffer based (aqueous) receptor solutions for hydrophobic drugs. If additional solubility is needed, small increases in the concentration of polyoxyethylene[20]oleyl ether (e.g., from 0.1% or 0.2%, which is typically adequate for most hydrophobic drugs, to higher concentrations) are recommended, but should not exceed 6% polyoxyethylene[20]oleyl ether. Other strategies to improve the solubility of the drug in the receptor solution that may have the potential to alter the permeability of the skin (e.g., inclusion of organic solvents and alcohols in the receptor solution) are not recommended and may invalidate the IVPT method.

建議加入0.1%聚氧乙烯[20]油酸酯（也稱為Oleth-20、Volpo-20或Brij-20；CAS號(hào)9004-98-2），以提高疏水性藥物生理緩沖液基（水）受體溶液的溶解度。如果需要額外的溶解度，建議將聚氧乙烯[20]油酸酯的濃度小幅度增加（例如，從0.1%或0.2%，這對(duì)于大多數(shù)疏水性藥物來(lái)說(shuō)通常是足夠的，到更高的濃度），但不應(yīng)超過(guò)6%的聚氧乙烯[20]油酸酯。不建議采用其他可能改變皮膚滲透性的提高藥物在受體溶液中溶解度的策略（例如，在受體溶液內(nèi)加入有機(jī)溶劑和醇），可能使IVPT方法無(wú)效。

The inclusion of an anti-microbial agent in the receptor solution (e.g., ~0.1% sodium azide or ~ 0.01% gentamicin sulfate) is recommended to mitigate potential bacterial decomposition of the dermis and/or epidermis in the diffusion cell, regardless of the study duration. Other antimicrobial agents may also be acceptable, and if used, information should be included in the ANDA to explain the reason for their selection (and for the concentration at which they were used).

無(wú)論研究持續(xù)時(shí)間如何，建議在受體溶液中加入抗菌劑（例如，約0.1%*氮化鈉或約0.01%硫酸慶大霉素），以減輕擴(kuò)散測(cè)試池中真皮和/或表皮的潛在細(xì)菌分解。其他抗菌劑也可以接受，如果使用，應(yīng)在ANDA中包含相關(guān)信息，以解釋選擇的原因（以及使用的濃度）。

D.Receptor Solution Sampling Qualification

The accuracy and precision of receptor solution sample collection at each time point should be appropriately qualified. Evidence to qualify a sampling procedure should illustrate that the sampling technique can reliably collect a consistent volume of the sample from the well-mixed volume of the receptor compartment at each sampling event, and that no artifacts are likely to be created by the sampling technique. Information should be included describing the equipment manufacturer’s specification for the accuracy and precision of receptor solution sampling, when available.

應(yīng)適當(dāng)限定每個(gè)時(shí)間點(diǎn)受體溶液樣品采集的準(zhǔn)確度和精密度。鑒定采樣程序的證據(jù)應(yīng)表明，采樣技術(shù)可以在每次采樣中從一定體積、均勻混合的受體室中收集一致體積的樣品，并且采樣技術(shù)不會(huì)產(chǎn)生污染。如果可用，應(yīng)包括描述設(shè)備制造商關(guān)于受體溶液取樣準(zhǔn)確性和精密度的規(guī)范的信息。

For IVPT studies using a flow-through diffusion cell, it may be appropriate to qualify the lengths of tubing, and their associated dead volumes, to accurately calculate the lag time before a sample elutes through the tubing and is collected. For IVPT studies using a VDC, removal of the entire receptor solution volume and full volume replacement of the receptor solution at each time point may provide optimal solubility sink conditions. The sampling of small aliquots of the receptor solution for an IVPT study may introduce anomalous measurements of apparently negative flux in certain regions of the IVPT study and produce flux profiles that are difficult to interpret.

對(duì)于使用流通擴(kuò)散池的IVPT研究，可能需要確定管道長(zhǎng)度及其相關(guān)死體積，以準(zhǔn)確計(jì)算樣品通過(guò)管道洗脫并收集之前的滯后時(shí)間。對(duì)于使用VDC的IVPT研究，在每個(gè)時(shí)間點(diǎn)去除整個(gè)受體溶液體積并全部置換受體溶液可以提供最佳溶解度漏槽條件。在IVPT研究中對(duì)受體溶液的小等份取樣可能會(huì)在IVPT研究某些區(qū)域引入明顯負(fù)通量的異常測(cè)量，并產(chǎn)生難以解釋的通量分布。

E.Environmental Control

Ambient laboratory temperature and humidity during the study should be monitored and reported. An environmentally controlled temperature range of 21°C ± 2°C is recommended, and a humidity range of 50% ± 20% relative humidity is recommended, if feasible.

研究期間應(yīng)監(jiān)測(cè)和報(bào)告實(shí)驗(yàn)室環(huán)境溫度和濕度。環(huán)境溫度宜控制在21°C±2°C，相對(duì)濕度宜控制在為50%±20%。

F.Permeation Profile and Range

The flux profile and cumulative permeation profile for the IVPT pilot study should be plotted across a range of sampling times, which corresponds to the IVPT pivotal study duration. The calculation of flux and cumulative total permeation is discussed in more detail below. The results of the IVPT pilot study should validate that the selected study parameters are suitable to adequately characterize the permeation profile (the cutaneous pharmacokinetics) of the drug within the selected study duration (the range of sampling time points).

IVPT試點(diǎn)研究的通量分布和累積滲透分布應(yīng)在一定取樣時(shí)間范圍內(nèi)繪制，這與IVPT關(guān)鍵研究持續(xù)時(shí)間相對(duì)應(yīng)。下面將更詳細(xì)地討論通量和累積總滲透的計(jì)算。IVPT先導(dǎo)研究的結(jié)果應(yīng)驗(yàn)證所選研究參數(shù)適合于在所選研究時(shí)間(采樣時(shí)間點(diǎn)范圍)內(nèi)充分表征藥物的滲透特性（皮膚藥代動(dòng)力學(xué)）。

A sufficiently complete flux profile should be adequate to identify the maximum (peak) flux and a decline in the flux thereafter across multiple subsequent time points in the IVPT pilot study. The results of the IVPT pilot study should also validate that the sampling frequency provides suitable resolution to adequately characterize the permeation profile (particularly the flux profile).

在IVPT試點(diǎn)研究中，一個(gè)足夠完整的通量分布應(yīng)足以確定多個(gè)后續(xù)時(shí)間點(diǎn)的最大（峰值）通量和此后通量的下降。IVPT試點(diǎn)研究的結(jié)果還應(yīng)驗(yàn)證采樣頻率提供了適當(dāng)?shù)姆直媛?，以充分表征滲透曲線（尤其是通量曲線）。

G.Precision and Reproducibility

The flux and cumulative permeation results from the IVPT pilot study (and the eventual IVPT pivotal study) should be calculated, tabulated, and reported for each diffusion cell at each time point, with summary statistics to describe the intra-donor average, standard deviation, and percent coefficient of variation (%CV) among replicates, as well as the inter-donor average, standard error, and %CV. Complete results for all data values used in the calculations should be reported in a clear and organized manner, to facilitate the reconstruction of the flux and cumulative permeation results. The design of the study should be detailed and clear, and data values should be clearly associated with specific donors, replicates, treatment groups, time points, etc

IVPT先導(dǎo)研究（以及最終的IVPT關(guān)鍵研究）的通量和累積滲透結(jié)果應(yīng)在每個(gè)時(shí)間點(diǎn)為每個(gè)擴(kuò)散池進(jìn)行計(jì)算、制表和報(bào)告，并提供匯總統(tǒng)計(jì)數(shù)據(jù)，以描述供體內(nèi)平均值、標(biāo)準(zhǔn)偏差和重復(fù)之間的變異系數(shù)百分比（%CV），以及供體間平均值和%CV。計(jì)算中使用的所有數(shù)據(jù)值的完整結(jié)果應(yīng)以清晰和有組織的方式報(bào)告，以便于重建通量和累積滲透結(jié)果。研究的設(shè)計(jì)應(yīng)詳細(xì)、清晰，數(shù)據(jù)值應(yīng)與特定捐獻(xiàn)者、重復(fù)、治療組、時(shí)間點(diǎn)等明確相關(guān)。

H.Dose Depletion

The recovery of permeated drug in the receptor solution should be characterized in each diffusion cell as the cumulative total permeation of the drug in the receptor solution over the IVPT duration. This may be expressed as a percentage of the nominal amount of drug in the applied dose (which may be estimated based upon the nominal strength of the drug in the topical product and the approximate mass of topical product dosed on the skin).

受體溶液中滲透藥物的回收應(yīng)為在每個(gè)擴(kuò)散池中表征為藥物在IVPT持續(xù)時(shí)間內(nèi)在受體溶液中的累積總滲透。這可以表示為藥物標(biāo)稱量在應(yīng)用劑量中的百分比（可以基于局部產(chǎn)品中藥物的標(biāo)稱強(qiáng)度和皮膚上施用的局部產(chǎn)品的近似質(zhì)量來(lái)估計(jì)）。

For example, if 10 mg of a topical product containing 5% drug was dosed on the membrane, the amount of drug in the applied dose may be estimated to be 0.5 mg (or 500 µg). If a cumulative total of 10 µg of drug diffused into the receptor solution across a 48-hour duration of the IVPT, it would be possible to estimate that the 500 µg dose would have been depleted by approximately 10 µg, amounting to an approximately 2% dose depletion. The average percentage dose depletion may thereby be estimated (not accounting for skin content) and should be reported.

例如，如果將含有5%藥物的10mg局部產(chǎn)品應(yīng)用在膜上，則應(yīng)用劑量中的藥物量估計(jì)為0.5mg（或500µg）。如果在IVPT的48小時(shí)持續(xù)時(shí)間內(nèi)，累計(jì)10µg藥物擴(kuò)散到受體溶液中，可以估計(jì)500µg劑量將消耗約10µg，相當(dāng)于約2%的劑量消耗。由此可以估計(jì)平均劑量消耗百分比（不考慮皮膚含量），并應(yīng)予以報(bào)告。

I.Discrimination Sensitivity and Selectivity

The discrimination ability of the IVPT method may be described using two concepts: sensitivity and selectivity. The IVPT sensitivity studies are necessarily performed during IVPT method development to establish IVPT method parameters like the dose amount, dose duration, study duration, etc. However, the analysis of the results from these studies is qualitative in nature, and they need not be repeated during the IVPT method validation phase.

IVPT方法的鑒別能力可以用兩個(gè)概念來(lái)描述：靈敏度和選擇性。IVPT方法開發(fā)期間必須進(jìn)行IVPT靈敏性研究，以確定IVPT方法參數(shù)，如劑量、劑量持續(xù)時(shí)間、研究持續(xù)時(shí)間等。然而，這些研究結(jié)果的分析本質(zhì)上是定性的，在IVPT方法驗(yàn)證階段無(wú)需重復(fù)驗(yàn)證。

The IVPT sensitivity studies are typically performed toward the end of the IVPT method development phase, and a key purpose of these studies is to incorporate the final IVPT method parameters for the target dose and dose duration to be used in the pivotal study so that the IVPT sensitivity studies can support a demonstration of the validity of the final IVPT method. Therefore, IVPT sensitivity studies are described within this section of the guidance in the context of IVPT validation (rather than method development) to avoid dissociating the discussions of IVPT sensitivity (which is performed to establish the suitability of the final IVPT method parameters) and IVPT selectivity (which is performed once the final IVPT method parameters are established, and which is based upon the IVPT pilot study that is performed as part of the IVPT method validation). With the exception of the alternative dose amounts or dose durations used in the IVPT sensitivity study, it is important that the IVPT method parameters are consistent across the IVPT sensitivity, pilot, and pivotal studies (including the anatomical region specified in the study protocol (e.g., posterior torso), the skin source, and skin preparation).

IVPT靈敏度研究通常在IVPT方法開發(fā)階段的末尾進(jìn)行，這些研究的一個(gè)關(guān)鍵目的是將最終IVPT方法參數(shù)的目標(biāo)劑量和劑量時(shí)間用于關(guān)鍵研究，IVPT靈敏性研究可以支持證明最終IVPT方法的有效性。因此本節(jié)指南在IVPT驗(yàn)證（而非方法開發(fā)）的背景下描述了IVPT靈敏度研究，以避免將IVPT靈敏度（用于確定最終IVPT方法參數(shù)的適用性）和IVPT選擇性（一旦確定了最終IVPT方法參數(shù)，即進(jìn)行該試驗(yàn)，并基于IVPT試驗(yàn)研究，該試驗(yàn)研究是IVPT方法驗(yàn)證的一部分）的討論分開。除了IVPT敏感性研究中使用的替代劑量或劑量持續(xù)時(shí)間外，重要的是IVPT方法參數(shù)在IVPT靈敏度、先導(dǎo)性和關(guān)鍵性研究（包括研究方案中規(guī)定的解剖區(qū)域（如軀干后部）、皮膚來(lái)源和皮膚制備）中保持一致。

1.IVPT Sensitivity

IVPT sensitivity is the ability of the IVPT method to detect changes in the cutaneous pharmacokinetics of the drug as a function of differences in drug delivery. If the IVPT method consistently demonstrates higher and lower flux profiles (i.e., higher and lower values for IVPT endpoints) in response to increased and decreased drug delivery, respectively (or in response to other conditions expected to increase and decrease drug delivery, respectively), the IVPT method may be considered sensitive.

IVPT靈敏度是IVPT方法檢測(cè)藥物的皮膚藥代動(dòng)力學(xué)變化作為藥物傳遞差異的功能的能力。如果IVPT方法始終顯示出較高和較低的通量分布（即IVPT終點(diǎn)的高和低），分別對(duì)藥物遞送的增加和減少作出反應(yīng)（或?qū)︻A(yù)期分別增加和減少藥物遞送量的其他條件作出反應(yīng)），則IVPT方法可能被認(rèn)為是靈敏的。

There are a few potential approaches by which to produce the differences in drug delivery that can be differentiated by a suitably discriminating IVPT method. Regardless of the approach used, the differences in the IVPT permeation profiles are not necessarily expected to be specifically proportional to differences in the dose amount, dose duration, or product strength. For example, three-fold differences in the dose amount (even if outside the recommended target dose range) may provide distinct flux curves but may not result in three-fold differences in the IVPT endpoints because the skin barrier may be rate-limiting both in vitro and in vivo.

有幾種潛在的方法可以產(chǎn)生藥物遞送的差異，這些差異可以通過(guò)適當(dāng)鑒別的IVPT方法進(jìn)行區(qū)分。無(wú)論采用何種方法，IVPT滲透曲線的差異不一定與劑量、劑量持續(xù)時(shí)間或產(chǎn)品強(qiáng)度的差異成正比。例如，劑量的三倍差異（即使在推薦的目標(biāo)劑量范圍之外）可以提供不同的通量曲線，但不會(huì)導(dǎo)致IVPT終點(diǎn)的三倍差別，因?yàn)槠つw屏障在體外和體內(nèi)都可能是限制速率的因素。

In other words, if the target dose for the pivotal IVPT study was 10 mg/cm2 , a 3-fold lower dose would be ~3 mg/cm2 and a 3-fold higher dose would be 30 mg/cm2 ; thus, an IVPT sensitivity study might compare the flux profiles from 3, 10, and 30 mg/cm2 doses of the topical product. Similarly, if the target dose for the pivotal IVPT study was 15 mg/cm2 , a 3-fold lower dose would be 5 mg/cm2 and a 3-fold higher dose would be 45 mg/cm2 ; thus, an IVPT sensitivity study might compare the flux profiles from 5, 15, and 45 mg/cm2 doses of the topical product. An IVPT sensitivity study performed with multiple skin donors (e.g., 4–6 skin donors) and a minimum of four replicate skin sections per donor per treatment group is recommended.

換言之，如果關(guān)鍵IVPT研究的目標(biāo)劑量為10 mg/cm2，則低3倍的劑量為~3 mg/cm2且高3倍的濃度為30 mg/cm2；因此，IVPT敏感性研究可以比較3、10和30 mg/cm2劑量的局部產(chǎn)品的通量分布。類似地，如果關(guān)鍵IVPT研究的目標(biāo)劑量為15 mg/cm2，低3倍的劑量為5 mg/cm2而高3倍的濃度為45 mg/cm2；因此，IVPT敏感性研究可能會(huì)比較5、15和45 mg/cm2劑量的局部產(chǎn)品的通量分布。建議對(duì)多個(gè)皮膚捐獻(xiàn)者（例如4-6個(gè)皮膚捐贈(zèng)者）進(jìn)行IVPT靈敏度研究，每個(gè)治療組每個(gè)捐獻(xiàn)者至少四個(gè)重復(fù)皮膚切片。

Modulation of Dose Amount: An IVPT method development study with different dose amounts may provide supportive evidence that the IVPT methodology is sensitive to differences in drug delivery.

劑量調(diào)節(jié)：不同劑量的IVPT方法開發(fā)研究可能提供支持性證據(jù)，證明IVPT方法對(duì)藥物遞送的差異敏感。

This approach is well suited to topical products that contain volatile components that evaporate from the formulation following dose application to the skin. Modulating the dose amount for such topical products effectively alters the thickness of the applied dose. The majority of volatile components from a thinner dose will tend to evaporate more rapidly (compared to a thicker dose), and a thinner dose will tend to deliver less drug into the skin (and/or for a shorter duration) compared to a thicker dose.

這種方法非常適合于含有揮發(fā)性成分的局部產(chǎn)品，這些揮發(fā)性成分在涂抹到皮膚后從配方中蒸發(fā)。調(diào)節(jié)這種局部產(chǎn)品的劑量有效地改變了應(yīng)用劑量的厚度。較薄劑量的大部分揮發(fā)性成分往往會(huì)蒸發(fā)得更快（與較厚劑量相比），較薄劑量進(jìn)入皮膚的藥量較少（和/或持續(xù)時(shí)間更短）。

Modulating the dose amount can be an effective technique to modulate differences in drug delivery for formulations with volatile components, like gels, lotions, and many creams. However, modulating the dose amount may not necessarily produce perceptible differences in drug delivery for topical products like petrolatum-based ointments, or other types of topical products that do not evaporate on the skin, or that may not experience dose-dependent differences in metamorphosis that can alter the rate and extent of drug delivery.

調(diào)節(jié)劑量可以是一種有效的技術(shù)，可以調(diào)節(jié)含有揮發(fā)性成分的制劑（如凝膠、乳液和許多乳膏）的藥物遞送差異。然而，調(diào)節(jié)劑量并不一定會(huì)對(duì)局部產(chǎn)品（如基于凡士林的軟膏）或其他類型的局部產(chǎn)品（不會(huì)在皮膚上蒸發(fā)）產(chǎn)生明顯的藥物遞送差異，或者可能不會(huì)經(jīng)歷可改變藥物遞送速率和程度的劑量依賴性**差異。

Modulation of Dose Duration: For many topical products, it may be more effective to modulate the dose duration, instead of the dose amount, to produce differences in drug delivery and associated changes in the cutaneous pharmacokinetics of the drug.

劑量持續(xù)時(shí)間的調(diào)節(jié)：對(duì)于許多外用產(chǎn)品，調(diào)節(jié)劑量持續(xù)時(shí)間而不是劑量可能更有效，以產(chǎn)生藥物遞送的差異和藥物的皮膚藥代動(dòng)力學(xué)的相關(guān)變化。

An IVPT method development study with a controlled dose amount (e.g., 15 mg/cm2 ) dosed for different durations (e.g., 2 hours, 6 hours, and 12 hours) may be well suited to provide supportive evidence that the IVPT methodology is sensitive to differences in drug delivery from many topical products. The scenario described in this example would support an IVPT study design where a topical product dose of 15 mg/cm2 is dosed for 6 hours (the target duration for the IVPT study) and then wiped off. The applied dose may be removed with a series of cotton-tipped swabs, one or more of which may be dry and one or more of which may be moistened (e.g., with a soap solution or water). The initial (dry) swab typically removes the bulk of the dose and subsequent swabs are used to remove the residual dose (i.e., the residue of the topical product which may otherwise continue to deliver drug into the skin) and/or to rinse the skin.

具有不同持續(xù)時(shí)間（例如，2小時(shí)、6小時(shí)和12小時(shí)）的受控劑量（例如，15 mg/cm2）的IVPT方法開發(fā)研究可能非常適合提供支持性證據(jù)，證明IVPT方法對(duì)許多局部產(chǎn)品的藥物遞送差異敏感。本例中描述的方案將支持IVPT研究設(shè)計(jì)，其中局部產(chǎn)品劑量為15 mg/cm2，持續(xù)6小時(shí)（IVPT研究的目標(biāo)持續(xù)時(shí)間），然后擦掉。可使用一系列棉簽去除應(yīng)用的劑量，其中一個(gè)或多個(gè)棉簽可能干燥，一個(gè)或更多棉簽可能濕潤(rùn)（例如，用肥皂溶液或水）。初始（干）拭子通常去除大部分劑量，隨后的拭子用于去除剩余劑量（即，局部產(chǎn)品的殘留物，否則可能繼續(xù)將藥物輸送到皮膚中）和/或沖洗皮膚。

To support a demonstration of the sensitivity of the IVPT study, the permeation profile produced by the target dose duration for the IVPT study (e.g., 6 hours) should be compared with a shorter dose duration (e.g., 2 hours) that is expected to perceptibly decrease the drug delivery, and also be compared with a longer dose duration (e.g., 12 hours) that is expected to perceptibly increase the drug delivery. Thereby, the three dose durations compared in the IVPT sensitivity study are designed to produce perceptible changes in the cutaneous pharmacokinetics of the drug as a function of differences in drug delivery, and thereby support a demonstration of the sensitivity of the IVPT method.

為了證明IVPT研究的靈敏度，應(yīng)將IVPT研究目標(biāo)劑量持續(xù)時(shí)間（例如6小時(shí)）產(chǎn)生的滲透曲線與預(yù)期明顯降低藥物遞送的更短劑量持續(xù)時(shí)間進(jìn)行比較，并且還與預(yù)期顯著增加藥物遞送的更長(zhǎng)劑量持續(xù)時(shí)間（例如12小時(shí)）相比較。因此，IVPT靈敏度研究中比較的三個(gè)劑量持續(xù)時(shí)間旨在根據(jù)藥物遞送差異產(chǎn)生藥物的皮膚藥代動(dòng)力學(xué)的明顯變化，從而支持IVPT方法的靈敏度證明。

The specific dose durations may be selected based upon an initial exploratory IVPT study performed during IVPT method development that characterizes the permeation profile when the dose is left on the skin for a longer duration (e.g., 24 or 48 hours). An important feature of the results from such an IVPT study is the duration of the initial phase of the permeation profile, when the flux is increasing at a relatively rapid rate.

具體劑量持續(xù)時(shí)間可以基于IVPT方法開發(fā)期間進(jìn)行的初始探索性IVPT研究來(lái)選擇，該研究表征了當(dāng)劑量在皮膚上停留更長(zhǎng)時(shí)間（例如24或48小時(shí)）時(shí)的滲透曲線。這種IVPT研究結(jié)果的一個(gè)重要特征是當(dāng)通量以相對(duì)較快的速率增加時(shí)，滲透曲線初始階段的持續(xù)時(shí)間。

For example, if such an exploratory study indicates that the flux increases on a steep slope until approximately 12 hours, and then continues to deliver the drug at a gradually increasing rate thereafter, it may suggest that the permeation profile for a dose duration of longer than 12 hours (e.g., 24 hours) may not be perceptibly different from that of the 12- hour dose duration, especially when compared in a relatively small number of donors and replicates (e.g., four donors with four replicates each per dose duration). It may also suggest that a 12-hour dose duration may be a good choice for the longest of the three dose durations in the IVPT sensitivity study.

例如，如果這樣的探索性研究表明，通量以陡峭的斜率增加，直到大約12小時(shí)，然后繼續(xù)以逐漸增加的速率遞送藥物，這可能表明，超過(guò)12小時(shí)（例如24小時(shí)）的劑量持續(xù)時(shí)間的滲透曲線可能與12小時(shí)劑量持續(xù)時(shí)間沒有明顯差異，特別是在相對(duì)較少數(shù)量的供體和復(fù)制體（例如每個(gè)劑量持續(xù)時(shí)間具有四個(gè)復(fù)制體的四個(gè)供體）中進(jìn)行比較時(shí)。這也可能表明，在IVPT敏感性研究中，12小時(shí)的劑量持續(xù)時(shí)間可能是三個(gè)劑量持續(xù)時(shí)間中最長(zhǎng)的一個(gè)很好的選擇。

The target dose duration should be selected based upon considerations like the sensitivity of the sample analytical method, the ability to produce a permeation profile that can be perceptibly discriminated from that produced by the longer (12 hour) dose duration, and/or the labeled use of the topical product (which may indicate that the topical product should be reapplied every 4–6 hours).

目標(biāo)劑量持續(xù)時(shí)間的選擇應(yīng)基于以下考慮因素，如樣品分析方法的靈敏度、產(chǎn)生滲透曲線的能力，該滲透曲線可明顯區(qū)別于較長(zhǎng)（12小時(shí)）劑量持續(xù)時(shí)間產(chǎn)生的滲透曲線，和/或局部產(chǎn)品的標(biāo)記使用（這可能表明局部產(chǎn)品應(yīng)每4-6小時(shí)重新給藥一次）。

The shortest of the three dose durations in the IVPT sensitivity study should be selected based upon the sensitivity of the sample analytical method and its ability to produce a permeation profile that can be perceptibly discriminated from that produced by the target (6 hour) dose duration.

在IVPT靈敏度研究中，三個(gè)劑量持續(xù)時(shí)間中最短的時(shí)間選擇，應(yīng)根據(jù)樣品分析方法的靈敏度，以及可明顯區(qū)別于目標(biāo)維持時(shí)間（6小時(shí)）所產(chǎn)生的滲透曲線的能力來(lái)選擇。

· Modulation of Product Strength: To validate the sensitivity, specificity, and selectivity of an in vitro release test (IVRT) method, altered strength formulations are routinely prepared. While it may seem convenient to use these altered strength formulations in an attempt to demonstrate the sensitivity and selectivity of an IVPT method, doing so may not produce the desired outcomes. There may be circumstances when this strategy may produce perceptibly different permeation profiles, however, in many instances, the resulting permeation profiles may not be perceptibly different when compared in a relatively small number of donors and replicates (e.g., four donors with four replicates each per topical product strength). In general, the modulation of topical product strength to support a demonstration of IVPT sensitivity is not recommended because it may not consistently produce the expected increase or decrease in drug delivery; however, in certain situations, higher and lower strength formulations (relative to the nominal strength of the RS) may suitably increase and decrease the drug delivery and cutaneous pharmacokinetics relative to that from the nominal strength topical product.

•調(diào)節(jié)產(chǎn)品規(guī)格：為驗(yàn)證IVRT方法的靈敏度、專屬性和選擇性，通常采用改變配方制劑規(guī)格的方法。雖然采用改變配方制劑的規(guī)格的方法來(lái)證明IVPT方法的靈敏度和選擇性似乎很方便，但這種方式可能不會(huì)產(chǎn)生預(yù)期的結(jié)果。在某些情況下，這種策略可能會(huì)產(chǎn)生明顯不同的滲透曲線，然而，在很多情況下，由于在對(duì)比研究中采用的供體組和重復(fù)數(shù)較少（如，每種外用制劑規(guī)格采用4個(gè)供體組，每組供體4個(gè)重復(fù)），并不能產(chǎn)生明顯不同的滲透曲線。通常，不建議通過(guò)調(diào)整外用制劑規(guī)格的方法以證明IVPT的靈敏度，因?yàn)樗赡懿粫?huì)持續(xù)產(chǎn)生預(yù)期的增加或減少的藥物遞送；然而，在某些情況下，相較于外用制劑的目標(biāo)規(guī)格，較高和較低規(guī)格的配方制劑（相對(duì)于對(duì)照制劑（RS）的規(guī)格）可以適當(dāng)?shù)卦黾雍徒档退幬镞f送和皮膚藥代動(dòng)力學(xué)。

2.IVPT Selectivity/IVPT選擇性

IVPT selectivity is the ability of the IVPT method to discriminate the cutaneous pharmacokinetics of the drug between the RS and a topical product or formulation that exhibits differences in drug delivery relative to the RS. The IVPT pilot study with the parallel assessment of the RS, the test topical product, and a third topical product or formulation that is known or designed to be different from the RS may provide supportive evidence that the IVPT methodology is selective for differences in drug delivery. Topical product batch information for all topical product lots used in IVPT method development, validation and pilot studies, as applicable, should be submitted in the study reports. The topical product information should include, but not be limited to, information about the batch formula, manufacturing date, batch size, altered manufacturing processes (if applicable) and, if available, potency and content uniformity. The evaluation of inequivalence may be based upon a qualitative or quantitative comparison of the permeation profiles and/or the IVPT endpoints.

IVPT選擇性是IVPT方法區(qū)分“對(duì)照制劑（RS）“和“與RS在藥物遞送方面存在差異的外用制劑或配方”在藥物皮膚藥代動(dòng)力學(xué)差異的能力。在IVPT初步研究中，通過(guò)將RS、自研外用制劑和“已知或設(shè)計(jì)不同于RS”的第三種外用制劑或配方進(jìn)行平行評(píng)估，提供支持性證據(jù)，說(shuō)明IVPT方法對(duì)藥物遞送的差異具有選擇性。如適用，應(yīng)在研究報(bào)告中遞交，IVPT方法開發(fā)、驗(yàn)證和初步研究中用到的所有外用制劑的相關(guān)批信息，包括但不限于：批配方、生產(chǎn)日期、批量、變更的生產(chǎn)工藝（如適用），以及效價(jià)和含量均勻度（如有）。可基于滲透曲線和/或IVPT終點(diǎn)的定性或定量比較進(jìn)行不等效性評(píng)估。

J.Robustness/耐用性

A primary assumption related to robustness testing is that the test system performs consistently when all system variables (e.g., temperature, stirring rate) are at nominal settings. A value of robustness testing is that it can verify whether the system continues to provide a consistent output when specific variables are slightly altered, thereby qualifying operational ranges for those variables. However, the variability inherent in the permeability of human skin, whether in vitro or in vivo, may not be compatible with the primary assumption related to the consistency of the test system.

與耐用性測(cè)試相關(guān)的一個(gè)主要假設(shè)是，當(dāng)所有系統(tǒng)變量（如：溫度、攪拌速率）處于標(biāo)準(zhǔn)設(shè)置時(shí)，測(cè)試系統(tǒng)的性能一致。耐用性測(cè)試意義在于，它可以驗(yàn)證當(dāng)特定變量略有改變時(shí)，系統(tǒng)是否能提供一致的輸出結(jié)果，從而確定這些變量的操作范圍。然而，無(wú)論是體外還是體內(nèi)，人體皮膚滲透性固有的變異性可能與與測(cè)試系統(tǒng)一致性相關(guān)的主要初始的假設(shè)不兼容。

Nonetheless, results from studies during IVPT method development that appear to support the robustness of the IVPT method or system should be reported, if relevant. For example, an IVPT method may be robust to substantial variations in the stirring rate of the receptor compartment. Similarly, the permeation profile of a drug into and through human skin may appear to be robust to certain differences in the topical product strength. Ultimately, because it may not always be feasible to validate the robustness of IVPT method parameters, IVPT study procedures should be controlled as precisely as possible.

盡管如此，如果相關(guān)，應(yīng)報(bào)告IVPT方法開發(fā)期間得到的關(guān)于支持IVPT方法或系統(tǒng)耐用性的研究結(jié)果。例如，IVPT方法對(duì)于接收室攪拌速率的顯著變化具有耐用性。與其類似地，藥物進(jìn)入并通過(guò)人體皮膚的滲透曲線，可能對(duì)外用制劑規(guī)格的改變表現(xiàn)出耐用性。所以，對(duì)于驗(yàn)證IVPT方法參數(shù)的耐用性并不總是可行的，在IVPT研究中應(yīng)盡可能精準(zhǔn)地控制IVPT研究程序。

V. SAMPLE ANALYTICAL METHOD VALIDATION/樣品分析方法驗(yàn)證

While exploratory studies performed during IVPT method development may use an unvalidated sample analytical method, it is essential that all studies conducted as part of the IVPT method validation use a validated sample analytical method. A validated IVPT method should use a validated sample analytical method (e.g., HPLC/MS or UPLC/MS). Therefore, a discussion of the sample analytical method for the IVPT method is included in this guidance under this section on IVPT method validation

雖然在IVPT方法開發(fā)的探索性研究中，可采用未經(jīng)驗(yàn)證的樣品分析方法，但作為IVPT方法驗(yàn)證的一部分，所進(jìn)行的IVPT研究都必須采用經(jīng)過(guò)驗(yàn)證的樣本分析方法。經(jīng)驗(yàn)證的IVPT方法應(yīng)使用已驗(yàn)證的樣品分析方法（如：HPLC/MS或UPLC/MS）。因此，本指南中關(guān)于IVPT方法驗(yàn)證的章節(jié)中包含了對(duì)IVPT方法樣品分析方法的討論。

However, the study protocols and reports related to the IVPT method are distinct from those for the sample analytical method that is used to quantify drug concentrations in IVPT receptor solution samples. The validation of a sample analytical method, in and of itself, does not demonstrate the validity of an IVPT method. Separate and specific reports should be submitted for the validation of the sample analytical method (e.g., HPLC/MS or UPLC/MS) and for the validation of the IVPT method.

然而，與IVPT方法相關(guān)的研究方案和報(bào)告與用于量化IVPT接收液樣品定量的分析方法不同。樣品分析方法的驗(yàn)證本身并不能證明IVPT方法的有效性。因此，應(yīng)分別提交IVPT方法驗(yàn)證報(bào)告和樣品分析方法（如：HPLC/MS或UPLC/MS）。

Any results from studies of the IVPT method that are performed during method development using a different sample analytical method than that which is ultimately validated, cannot support a demonstration of the validity of the IVPT method. Information should be provided in the IVPT method validation report referencing the (separate) sample analytical method validation, and clearly indicate that all relevant results in the IVPT method validation report were obtained using a validated sample analytical method (as opposed to a sample analytical method with different parameters than those which were validated).

如果在方法開發(fā)過(guò)程中，使用的樣品分析方法與最終驗(yàn)證的樣品分析方法不同，那么與其相關(guān)的IVPT方法研究結(jié)果則不能用于支持論證IVPT方法的有效性。應(yīng)在IVPT方法驗(yàn)證報(bào)告中，提供參考的樣品分析方法驗(yàn)證的信息，并明確表明IVPT方法驗(yàn)證報(bào)告中的所有相關(guān)結(jié)果均采用經(jīng)驗(yàn)證的樣品分析方法（不是與經(jīng)驗(yàn)證的樣品分析法有不同參數(shù)的樣品分析放法）獲得的。

The receptor sample analysis procedures (e.g., typically involving an HPLC/MS or UPLC/MS system) should be performed using chromatography software (e.g., a chromatography data system) with audit trails, and should include a multi-point (6–8 concentration) calibration curve with suitable quality control samples, and should be validated in a manner compatible with the FDA guidance for industry Bioanalytical Method Validation (May 2018).

接收液樣品的分析方法（如：通常為HPLC/MS或UPLC/MS系統(tǒng)），應(yīng)使用具有審計(jì)追蹤的色譜軟件（如，色譜數(shù)據(jù)系統(tǒng)），并應(yīng)包括具有適當(dāng)質(zhì)量控制樣品的多點(diǎn)（6～8個(gè)）校準(zhǔn)曲線，并應(yīng)符合FDA生物分析方法驗(yàn)證行業(yè)指南要求。

The validation of the receptor sample analytical method should include relevant qualifications of dilution integrity, if applicable, as well as stability assessments with the highest relevant temperature in the receptor solution for the longest relevant duration; the highest relevant temperature may be warmer than 32°C because the temperature of the receptor solution is often higher than the temperature at the surface of the skin, and the longest relevant duration may be the longest interval between sampling time points for methods in which the entire receptor solution is replaced at each sampling time point, or it could be longer in scenarios with only partial sampling of the receptor solution (e.g., 34°C for 48 hours).

接收液中樣品分析方法的驗(yàn)證應(yīng)包括稀釋完整性確認(rèn)（如適用），以及樣品在最高相關(guān)溫度的接收液中放置最長(zhǎng)時(shí)間的穩(wěn)定性評(píng)估；最高相關(guān)溫度可略高于32°C，因?yàn)榻邮找旱臏囟韧ǔ８哂谄つw表面的溫度。對(duì)于在每個(gè)取樣時(shí)間點(diǎn)替換整個(gè)接收液的情況，最長(zhǎng)相關(guān)時(shí)間為各相鄰取樣時(shí)間點(diǎn)中，間隔最長(zhǎng)時(shí)間；但對(duì)于接收液部分取樣的情況，時(shí)間可能更長(zhǎng)（例如34°C放置 48h）。

If the samples are processed in specific ways for analysis (e.g., by drying and reconstituting the receptor samples in a smaller volume to concentrate the sample and increase the effective analytical sensitivity, or by dilution of receptor solution samples into the validated curve range of the sample analytical method) those procedures should be validated (e.g., by qualifying the dilution integrity during the sample analytical method validation). The stability of the drug in the receptor solution sample should be validated in a receptor solution matrix that has been exposed to the underside of the skin in a diffusion cell under conditions relevant to the IVPT pivotal study.

如果樣品以特定的分析方法進(jìn)行處理（如：通過(guò)將樣品干燥后重新以較小的體積溶劑配制以提高有效分析靈敏度，或通過(guò)稀釋接收液樣品，使其達(dá)到已驗(yàn)證分析方法的曲線范圍內(nèi)），則應(yīng)確認(rèn)其處理方法（如：在樣品分析方法驗(yàn)證期間確認(rèn)稀釋完整性）。接收液樣品中藥物的穩(wěn)定性，應(yīng)在與IVPT正式研究相關(guān)的條件下，在擴(kuò)散池中皮膚下側(cè)接觸的接收液基質(zhì)中進(jìn)行。

VI. IVPT PIVOTAL STUDY/IVPT正式研究

The IVPT pivotal study protocol should incorporate considerations relevant to BE studies, in general

IVPT正式研究方案通常應(yīng)包括與BE研究相關(guān)的考慮因素

A. Handling and Retention of Samples/樣品的處理與保存

Refer to 21 CFR 320.38, 320.63 and the FDA guidances for industry Handling and Retention of BA and BE Testing Samples (May 2004) and Compliance Policy for the Quantity of Bioavailability and Bioequivalence Samples Retained Under 21 CFR 320.38(c) (August 2020), as applicable, regarding considerations for retention of study drug samples and to 21 CFR 320.36 for requirements for maintenance of records of BE testing. Retention samples should be randomly selected from the drug supplies received before allocating topical product units for use in an IVPT study in which the test topical product and RS are compared.

參考21 CFR 320.38，320.63和FDA行業(yè)指南《生物利用度（BA）和生物等效性（BE）研究中試驗(yàn)樣品的處理和保存》（2004年5月）和《21CFR 320.38(c) BE樣品留存的數(shù)量及生物利用度》（2020年8月），關(guān)于保留研究藥物樣品的考慮以及21 CFR 320.36關(guān)于保存BE檢測(cè)記錄的要求（如適用）。在采用自研外用制劑和RS進(jìn)行IVPT對(duì)比研究前，應(yīng)從收到的藥品中隨機(jī)選擇樣品進(jìn)行留樣。

B. Control of Study Procedures/研究程序的控制

Study procedures that have the potential to influence the results of the study should be appropriately controlled. Also, experimental observations that may have the potential to influence the interpretation of the study results, as well as any protocol or standard operating procedure (SOP) deviations, should be reported.

應(yīng)對(duì)可能影響研究結(jié)果的研究程序進(jìn)行適當(dāng)控制。此外，應(yīng)報(bào)告可能影響研究結(jié)果解釋的實(shí)驗(yàn)觀察結(jié)果，以及任何方案或標(biāo)準(zhǔn)操作程序（SOP）發(fā)生偏差。

Control of procedures related to the skin include the consistent control across the study of the skin preparation (e.g., dermatoming of skin sections) and the thickness of skin sections mounted on diffusion cells, as well as the skin storage conditions, including the duration for which the skin was frozen and the number of freeze-thaw cycles to which the skin was exposed. Skin from the same anatomical location should be used from all donors, and the demographics (age, race, sex) should be reported for all donors. Also, the IVPT sensitivity, pilot, and pivotal studies should use skin from the same anatomical site; otherwise, if skin from different anatomical sites is used across the different study phases, it may not be possible for the results of the IVPT sensitivity and pilot studies to support a demonstration of the discrimination ability of the IVPT method used for the pivotal study because the method parameters would not be aligned across the respective studies. Similarly, if a non-rate-limiting support membrane is used beneath the skin section (e.g., a filter membrane used in a validated IVRT method for the same topical product) then it should be used in a consistent manner for the IVPT sensitivity, pilot, and pivotal studies.

與皮膚相關(guān)的程序的控制，包括相同的皮膚處理方法（如：皮膚的剝離）和安裝在擴(kuò)散測(cè)試池上的皮膚切片的厚度的研究中的一致性，及皮膚儲(chǔ)存條件，例如皮膚冷凍的儲(chǔ)存時(shí)間和取出的凍融循環(huán)次數(shù)。所有供給皮膚均應(yīng)采用相同解剖位置，并報(bào)告所有供體的人口統(tǒng)計(jì)信息（年齡、種族、性別）。此外，在IVPT靈敏度、初步實(shí)驗(yàn)和正式研究，應(yīng)采用同一解剖部位的皮膚；否則，如果在不同的研究階段使用不同位置解剖部位，IVPT靈敏度和初步試驗(yàn)的結(jié)果可能不足以支持用于正式研究的IVPT方法的區(qū)分力的論證，因?yàn)榉椒▍?shù)在各個(gè)研究中不一致。同樣，如果在皮膚部分下方使用非速率限制性支撐膜（如：同類IVRT方法驗(yàn)證研究中使用的濾膜），則與IVPT靈敏度、初步試驗(yàn)和正式研究中一致。

Control of procedures related to the dose include the control of the area of dose application, the dose amount, the dosing technique, the dose duration, and the blinding and randomization procedures for dosing. The test topical product and RS should be dosed in an identical and consistent manner for all diffusion cells in the study. Differences in dosing technique may alter the metamorphosis of the dosage form on the skin, and inconsistencies in the diameter of the area dosed on each diffusion cell may significantly influence the dosed area and contribute to errors in the calculation of flux.

與劑量或上樣相關(guān)的控制程序，包括：對(duì)上樣面積、上樣量、上樣技術(shù)、劑量維持時(shí)間的控制，以及在研究中每個(gè)擴(kuò)散池采用的盲法和隨機(jī)化方法的一致性。不同的上樣技術(shù)可能改變上樣到皮膚上劑型的形態(tài)，擴(kuò)散池孔口擴(kuò)散面積的不一致可能會(huì)顯著影響上樣面積，從而導(dǎo)致通量計(jì)算的偏差。

Control of procedures related to sampling include the control of sampling time precision, the sampling technique, the duration of sampling and replacement of receptor solution, the sample volume or flow rate, and sample handling and storage.

與取樣相關(guān)的程序控制包括取樣時(shí)間精度、取樣技術(shù)、取樣和替換接收液的時(shí)間間隔、取樣體積或流速，以及樣品的處理和存儲(chǔ)。

Control of procedures related to the pivotal study should include a non-dosed control skin section from each skin donor, which should be mounted in a diffusion cell and otherwise treated identically to the dosed skin sections, including sampling of the receptor solution at all time points to ensure that drug concentrations monitored in the receptor solution are associated with the dose applied in the IVPT pivotal study, and not drug contamination in the skin from that donor that might permeate into the receptor solution across the duration of the study. A pre-dose “zero” sample collected from each diffusion cell is also recommended, which may identify potential contamination associated with each skin section and/or each diffusion cell.

與正式研究相關(guān)的控制程序，應(yīng)包括來(lái)自每個(gè)皮膚供體的非劑量對(duì)照皮膚部分，包括在所有時(shí)間點(diǎn)對(duì)接收液進(jìn)行取樣，以確保接收液中監(jiān)測(cè)的藥物濃度與IVPT正式研究中應(yīng)用的劑量相關(guān)，而不是研究期間由于皮膚的藥物污染滲透進(jìn)入。建議從每個(gè)擴(kuò)散池中采集“零”濃度樣品，用于識(shí)別每個(gè)皮膚切片和/或每個(gè)擴(kuò)散池的潛在污染。

In addition, investigators should perform the IVPT validation and pivotal studies within a quality management system that includes, but is not limited to, documented procedures for:

此外，研究人員應(yīng)在一定的質(zhì)量管理體系條件下，進(jìn)行IVPT驗(yàn)證和正式研究，該體系包括但不限于：

·Study personnel identification, training, qualification, and responsibilities/ 研究人員的識(shí)別、培訓(xùn)、資格和職責(zé)

·Study management and study management personnel responsibilities/研究管理和研究管理人員職責(zé)

· Quality control (QC) and QC personnel responsibilities/質(zhì)量控制（QC）和QC人員職責(zé)

· Quality assurance (QA) and QA personnel responsibilities/質(zhì)量保證（QA）和QA人員職責(zé)

·Use of SOPs /標(biāo)準(zhǔn)操作規(guī)程（SOP）的制定

· Use of study protocols/研究方案的制定

· Use of study reports/研究報(bào)告的制定

· Maintenance and control of the study facility environment and systems/研究設(shè)施環(huán)境和系統(tǒng)的維護(hù)和控制。

·Qualification and calibration of instruments and computerized systems/儀器和計(jì)算機(jī)系統(tǒng)的鑒定和校準(zhǔn)。

·Good documentation practices including, but not limited to, contemporaneous documentation of study procedures and recording of experimental observations or deviations from procedures specified in the study protocol or in relevant SOPs/良好的文件規(guī)范，包括但不限于：研究程序的及時(shí)記錄、實(shí)驗(yàn)觀察及相關(guān)SOPs中規(guī)定的程序的偏差。

· Maintenance of suitable records that facilitate the reconstruction of study events and procedures, including study sample handling and storage records (e.g., sample tracking logs), audit trails for sample analysis procedures, control of study materials and reagents, and electronic data control/維護(hù)有助于重建研究事件和程序的適當(dāng)記錄，包括研究樣本處理和存儲(chǔ)記錄（如：樣本跟蹤日志）、樣本分析過(guò)程的審計(jì)跟蹤、研究物料和試劑的控制以及電子數(shù)據(jù)控制。

·Archival of study records研究記錄存檔

C. Blinding Procedure/盲法程序

A detailed description of the blinding procedure should be provided in the study protocol and final report. The packaging of the test topical product and RS should be similar in appearance to maintain adequate blinding of the investigator and any experimental operators.

研究方案和最終報(bào)告中應(yīng)提供盲法程序的詳細(xì)說(shuō)明。試驗(yàn)外用制劑和RS的包裝應(yīng)保持相似的外，以確保研究者和任何實(shí)驗(yàn)操作人員的充分致盲。

D.Randomization/隨機(jī)化

The method of randomization should be described in the protocol of the IVPT study and the randomization schedule provided, preferably in a SAS data set in .xpt format (created using the SAS XPORT procedure). It is recommended that an independent third party generate and hold the randomization code throughout the conduct of the study to minimize bias. The applicant may generate the randomization code if not involved in the packaging and labeling of the test topical product and RS dosed in the study. A sealed copy of the randomization scheme should be retained at the study site and should be available to FDA investigators at the time of site inspection to allow for verification of the treatment identity of each skin section.

應(yīng)在IVPT研究方案中描述隨機(jī)化方法并提供隨機(jī)化計(jì)劃表，推薦以.xpt格式的SAS數(shù)據(jù)集形式（使用SAS XPORT程序創(chuàng)建）。建議由獨(dú)立的第三方在整個(gè)研究過(guò)程中生成并保存隨機(jī)化代碼，以減少偏差。如果研究中未涉及自研外用制劑和RS的包裝和標(biāo)簽，申請(qǐng)人可以自行生成隨機(jī)代碼。隨機(jī)方案的密封副本應(yīng)保存在研究現(xiàn)場(chǎng)，并應(yīng)在現(xiàn)場(chǎng)檢查時(shí)提供給FDA研究人員，以便確認(rèn)每個(gè)皮膚切片的具體信息。

E.Dosing/上樣

In the IVPT pivotal study, the test topical product and RS should be dosed in an alternating pattern on successive diffusion cells (skin sections) from each donor. One of two dosing sequences (illustrated below) may be randomly assigned for each donor:

在IVPT正式研究中，對(duì)于每個(gè)供體組，可以在擴(kuò)散池（皮膚切片）上以交替給藥方式依次放置自研外用制劑和RS。對(duì)于每個(gè)供體組，可采用下述兩個(gè)方式中的一個(gè)進(jìn)行隨機(jī)放樣（如下所示）：

a. ABABAB…

b. BABABA…

F. Study Design/  研究設(shè)計(jì)

The IVPT pivotal study should compare the cutaneous pharmacokinetics of the drug from the test topical product versus that from the RS using excised human skin with a competent skin barrier mounted on a qualified diffusion cell system. The IVPT pivotal study should use a design that directly compares the test topical product and RS on skin from the same set of donors, each with the same number of replicate skin sections per donor per treatment group (dosed with either test topical product or RS topical), using the same IVPT method parameters.

在IVPT正式研究中，應(yīng)采用皮膚屏障完整性良好的離體人類皮膚和經(jīng)驗(yàn)證的擴(kuò)散池系統(tǒng)，對(duì)自研外用制劑和RS的皮膚藥代動(dòng)力學(xué)進(jìn)行對(duì)比研究。在自研外用制劑和RS的對(duì)比研究中，每個(gè)試驗(yàn)組應(yīng)采用相同的皮膚供體、相同的皮膚切片重復(fù)數(shù)，并采用相同的IVPT方法參數(shù)。

The IVPT pivotal study design, methodology, and diffusion cell equipment considerations relating to sampling precision should be controlled as precisely as possible. For example, it may be appropriate to stagger the dose application on successive diffusion cells and to synchronize the sampling time points with the dosing time for that diffusion cell, to ensure consistent durations between dosing and sampling of all diffusion cells.

應(yīng)盡可能精確地控制IVPT正式研究設(shè)計(jì)、方法和與擴(kuò)散池設(shè)備取樣精度。例如，在連續(xù)的擴(kuò)散池上，錯(cuò)開上樣時(shí)間點(diǎn)，根據(jù)上樣時(shí)間的差異，從而同步取樣時(shí)間，以確保所有擴(kuò)散池給藥和取樣之間的時(shí)間間隔一致。

G. Inclusion Criteria/納入標(biāo)準(zhǔn)

In general, the following inclusion criteria should apply: healthy, normal, barrier-competent skin from male and/or female donors of at least 18 years of age. Inclusion criteria related to donor demographics (e.g., age, race, sex) should be specified in the study protocol and demographic information should be reported for each donor. Additional criteria may be added by the applicant

通常，以下入選標(biāo)準(zhǔn)應(yīng)適用：來(lái)自至少18歲的健康、正常、屏障功能皮膚，雄性和/或雌性供體。應(yīng)在研究方案中規(guī)定符合納入標(biāo)準(zhǔn)的人口統(tǒng)計(jì)信息（如，年齡、種族、性別），并報(bào)告每個(gè)供體的個(gè)人信息。申請(qǐng)人可以增加其它納入標(biāo)準(zhǔn)。

The skin may be harvested following excision from patients undergoing a surgical procedure or excised from cadavers. A consistent source is recommended for all the skin used. The anatomical region specified in the study protocol (e.g., posterior torso) should be consistent for all donors whose skin is included in the study.

皮膚可以在外科手術(shù)的患者或尸體上獲取。所有使用的皮膚應(yīng)有一致的來(lái)源。研究方案中規(guī)定的解剖區(qū)域（如軀干后部）應(yīng)與研究中包括皮膚的所有供體一致。

The study protocol should specify the inclusion (acceptance) criteria for skin sections based upon the barrier integrity test result, which should be reported for each skin section.

研究方案應(yīng)根據(jù)屏障完整性測(cè)試結(jié)果規(guī)定皮膚部分的納入（接受）標(biāo)準(zhǔn)。

The study protocol should specify inclusion criteria related to the temperature and duration of skin storage as well as the number of freeze-thaw cycles, all of which should be reported for each donor’s skin.

研究方案應(yīng)規(guī)定與皮膚儲(chǔ)存的溫度和儲(chǔ)存時(shí)間以及凍融循環(huán)次數(shù)納入相關(guān)的標(biāo)準(zhǔn)。

The study protocol should specify the inclusion criteria related to the skin harvesting/processing procedures and skin thickness (e.g., dermatomed skin of 500 μm ± 250 μm thickness) used in the IVPT study.

研究方案應(yīng)規(guī)定與IVPT研究中使用的皮膚采集/處理程序和皮膚厚度（例如，500μm±250μm厚度的皮膚）。

H. Exclusion Criteria/排除標(biāo)準(zhǔn)

In general, the following exclusion criteria should apply. Skin from subjects with a known (history of) dermatological disease should be excluded from the study. Skin with tattoos, stretch marks, or any visible sign of abnormality should be excluded from the study. Skin exhibiting a significant density of terminal hair is not recommended and should be excluded from the study. Additional criteria may be added by the applicant.

通常，采用以下排除標(biāo)準(zhǔn)?；加幸阎つw?。ú∈罚┑墓w皮膚；有紋身、妊娠紋或任何明顯異常跡象的皮膚應(yīng)；皮膚表現(xiàn)出明顯的末梢毛發(fā)密度等。申請(qǐng)人可增加其他標(biāo)準(zhǔn)。

While gentle washing or rinsing of the skin surface is appropriate, submerging the skin in an aqueous solution for more than a few minutes may damage the skin barrier and should be avoided; such skin sections should be excluded from the study. Also, skin that has been subjected to shaving with a blade; abrasive polishing; tape-stripping; or cleansing with alcohols, solvents, or other strong solutions that could damage the skin barrier should be excluded from the study.

雖然可以采用溫和地清洗或漂洗皮膚表面，但將皮膚浸泡在水溶液中幾分鐘以上可能會(huì)破壞皮膚屏障，應(yīng)避免該方式。另外，不建議采用用刀片刮過(guò)的皮膚；磨料拋光；膠帶剝離；或使用酒精、溶劑或其他可能破壞皮膚屏障的強(qiáng)極性溶液進(jìn)行清潔。

Skin from donors with significant background levels of the drug or other compounds that may interfere with the quantification of the drug in receptor solution samples should be excluded from the study.

來(lái)自供體的皮膚具有顯著的藥物背景水平或其他可能干擾受體溶液樣品中藥物定量的化合物應(yīng)排除在研究之外。

Skin from donors exhibiting a high barrier integrity test failure rate among replicate skin sections may be excluded from the study, and skin from an alternative donor may be used instead.

重復(fù)皮膚切片中屏障完整性測(cè)試失敗率高的供體皮膚可能被排除在研究之外，應(yīng)采用替代供體皮膚。

I. IVPT Endpoints/  IVPT終點(diǎn)

The endpoints for the IVPT pivotal study are based upon parameters that characterize the rate and extent to which the drug permeates into and through the skin and becomes available in the receptor solution. Specifically, the rate of drug permeation is characterized by the flux (J) and the extent of drug permeation is characterized by the total cumulative amount (AMT) of drug permeated into the receptor solution across the study duration.

IVPT正式研究的終點(diǎn)，是基于表征藥物滲透和通過(guò)皮膚中，并進(jìn)入接收液的速率和程度的參數(shù)。具體來(lái)說(shuō)，藥物滲透速率以通量（J）進(jìn)行表征，而藥物滲透程度采用整個(gè)研究期間滲透到接收液中的藥物的總累積量（AMT）進(jìn)行表征。

The flux (rate of drug permeation) should be plotted as J on the Y-axis in units of mass/area/time (e.g., nanograms (ng)/cm2 /hr) versus time on the X-axis. Flux profiles commonly resemble plasma pharmacokinetic profiles, however, it is important to distinguish that the flux is a rate, rather than a concentration. The extent of drug permeation should also be plotted, as the total cumulative amount (AMT) of drug permeated on the Y-axis in units of mass/area (e.g., ng/cm2 ) versus time on the X-axis.

以通量（藥物滲透速率）為縱坐標(biāo)Y-軸，時(shí)間為橫坐標(biāo)X-軸作圖；其中Y-軸用“J”標(biāo)識(shí)，以質(zhì)量/面積/時(shí)間（如，納克/平方厘米/小時(shí)）為單位。通量分布模型通常類似于血漿藥代動(dòng)力學(xué)分布模型，盡管通量是一個(gè)速率單位，并不代表濃度。也應(yīng)對(duì)藥物的滲透程度作圖，以藥物累積滲透量（單位為質(zhì)量/面積，如納克/平方厘米）為縱坐標(biāo)Y-軸，時(shí)間為橫坐標(biāo)X-軸。

The flux should be calculated based upon: the receptor sample concentration (e.g., 2.0 ng/mL) at each time point; the precise, empirically measured volume of that specific diffusion cell (e.g., 6.0 mL) which may vary between individual cells; the area of dose application (e.g., 1 cm2 ); and the duration for which the receptor volume was accepting the drug. For example, if the sample exemplified here represented a 2-hour period following dosing, then J would be calculated based upon the values above as:

流量的計(jì)算應(yīng)基于：每個(gè)時(shí)間點(diǎn)的接收液中的樣品濃度（如：2.0 ng/mL）；精確、經(jīng)驗(yàn)性的測(cè)量擴(kuò)散池的體積（如：6.0mL），每個(gè)擴(kuò)散池體積或有不同；上樣區(qū)域面積（如：1cm2）；研究的持續(xù)時(shí)間。例如，如果這里例舉的樣品表示給藥后2小時(shí)內(nèi)的情況，則J將基于上述值計(jì)算為：J = [(2.0 ng/mL) x (6.0 mL)]/(1 cm2 )/(2 hrs) = 6 ng/cm2 /hr

This flux should be calculated and reported for each diffusion cell for each sampling interval and plotted across the entire study duration to generate the flux profile for each diffusion cell. The rate calculated above may be plotted at the 2-hour time point, or at the midpoint between 0 and 2 hours (i.e., 1 hour).

應(yīng)計(jì)算并報(bào)告各擴(kuò)散池的所有相鄰采樣點(diǎn)之間的通量，并繪制整個(gè)研究期間所有擴(kuò)散池的通量曲線。上面計(jì)算的速率可以在2小時(shí)的實(shí)際時(shí)間點(diǎn)進(jìn)行繪制，或者在0到2小時(shí)（即1小時(shí)）之間的中點(diǎn)繪制。

In addition, the AMT should be calculated and reported for each diffusion cell. This cumulative amount of drug that has permeated (in total across the entire study) should be reported as the AMT endpoint, rather than using a trapezoid rule to calculate the area under the flux curve.

此外，應(yīng)計(jì)算并報(bào)告每個(gè)擴(kuò)散池的總累計(jì)滲透量（AMT）。AMT終點(diǎn)為藥物滲透的累計(jì)量（整個(gè)研究期間的滲透總量），而不是采用梯形法則計(jì)算的通量曲線下的面積。

The maximum flux (Jmax) at the peak of the drug flux profile and the AMT should both be compared for locally-acting test topical products and RSs. This is somewhat analogous to the comparison of the Cmax and AUC for systemically-acting test products and RSs, inasmuch as the pair of endpoints in each case facilitates a comparison of the rate and extent to which the drug from each type of product (locally-acting or systemically-acting) becomes available at the site of action.

應(yīng)將局部起效的自研制劑和RSs的最大通量（Jmax，藥物通量曲線的最高峰）和AMT進(jìn)行對(duì)比。這類似于全身起效的自研制劑和RSs的Cmax和AUC的比較，因?yàn)檫@些終點(diǎn)可分別用于表征各劑型（局部起效或全身起效）藥物到達(dá)作用部位的速率和程度。

A confidence interval (CI) should be calculated for each IVPT endpoint: 應(yīng)計(jì)算每個(gè)IVPT終點(diǎn)的置信區(qū)間（CI）：

a. the natural log-transformed maximum flux (Jmax) 自然對(duì)數(shù)換算后的最大通量（Jmax）

b. the natural log-transformed total cumulative amount (AMT) permeated自然對(duì)數(shù)換算后的總累積滲透量（AMT）

It is the responsibility of the applicant to determine the number of donors required to adequately power the IVPT pivotal study, however, a minimum of four dosed replicates per donor per treatment group (test product or RS) is recommended.

申請(qǐng)人有責(zé)任確定足以支持IVPT關(guān)鍵研究所需的供體數(shù)量，然而，建議每個(gè)試驗(yàn)組（自研制劑或RS）每個(gè)供體至少4個(gè)重復(fù)劑量。

At the completion of the study, if the number of skin replicates is the same for all donors in the test topical product and RS treatment groups in the IVPT study, a statistical analysis for a balanced design is recommended. If skin sections or diffusion cells are excluded from the final statistical analysis because of experimental loss/issues, and the resulting data set is unbalanced, a statistical analysis for an unbalanced design is recommended.

在IVPT研究結(jié)束時(shí)，如果自研外用制劑和RS試驗(yàn)組中所有供體的皮膚切片重復(fù)數(shù)均一致，建議采用平衡設(shè)計(jì)法進(jìn)行統(tǒng)計(jì)分析。如果皮膚切片或擴(kuò)散測(cè)試池因?qū)嶒?yàn)損失/問(wèn)題而被排除在最終統(tǒng)計(jì)分析之外，且所得數(shù)據(jù)集不平衡，則建議對(duì)不平衡設(shè)計(jì)進(jìn)行統(tǒng)計(jì)分析。

Approaches to statistical analysis of the pivotal study are described in section VIII of this guidance. Appendix I provides example SAS code for determining BE with both a balanced dataset and an unbalanced dataset. Appendix II provides numerical examples with simulated data sets. Appendix III provides example R code for determining BE.

在本指南的第VIII部分，對(duì)正式研究的統(tǒng)計(jì)分析方法進(jìn)行了描述。附錄I中提供了用于測(cè)定平衡數(shù)據(jù)集和非平衡數(shù)據(jù)集BE的SAS代碼示例。附錄II中提供了模擬數(shù)據(jù)集的具體示例。附錄III中提供了測(cè)定BE的R代碼示例。

VII. SUBMITTING INFORMATION ON IVPT STUDIES IN AN ANDA/ANDA中遞交的IVPT研究信息

For IVPT studies with topical products submitted in ANDAs that are intended to support a demonstration of BE, detailed study protocols, relevant SOPs, and detailed reports should be submitted for the IVPT method validation (including the IVPT pilot study) and the IVPT pivotal study. In addition, a detailed report describing the IVPT method development should be submitted. These protocols, SOPs, and reports should be submitted in module 5.3.1.2 of the electronic Common Technical Document (eCTD) and should describe experimental procedures, study controls, quality management procedures, and data analyses.

對(duì)于在ANDA中提交的用于支持BE演示的外用制劑的IVPT研究，應(yīng)提交詳細(xì)的研究方案、相關(guān)SOP和詳細(xì)的報(bào)告，用于IVPT方法驗(yàn)證（包括IVPT初步研究）和IVPT關(guān)鍵研究。此外，還應(yīng)提交一份詳細(xì)的IVPT方法開發(fā)報(bào)告。這些方案、SOPs和報(bào)告應(yīng)在電子通用技術(shù)文件（eCTD）的模塊5.3.1.2中遞交，并對(duì)相關(guān)的試驗(yàn)方法、研究控制、質(zhì)量管理程序和數(shù)據(jù)分析進(jìn)行描述。

Note that the study protocols, SOPs, and reports related to the IVPT method are distinct from those for the sample analytical method that is used to quantify drug concentrations in IVPT receptor solution samples (e.g., an HPLC/MS or UPLC/MS method). Separate protocols and SOPs should be submitted for the sample analytical method validation. Sample analytical method development and validation reports, pilot and pivotal IVPT study sample analysis reports, as well as associated SOPs and protocols relevant to the sample analysis of an IVPT study with human skin should be submitted in Module 5.3.1.4 of the eCTD.

注意，用于IVPT接收液中樣品濃度的定量分析方法（如，HPLC/MS或UPLC/MS方法）與IVPT方法相關(guān)的研究方案、SOPs和報(bào)告不同。樣品分析方法驗(yàn)證應(yīng)提交單獨(dú)的方案和標(biāo)準(zhǔn)操作規(guī)程。樣本分析方法開發(fā)和驗(yàn)證報(bào)告、試點(diǎn)和關(guān)鍵IVPT研究樣本分析報(bào)告以及與人體皮膚IVPT研究的樣本分析相關(guān)的相關(guān)SOP和協(xié)議應(yīng)在eCTD的模塊5.3.1.4中提交。

VIII.IVPT PIVOTAL STUDY STATISTICAL ANALYSIS/IVPT正式研究統(tǒng)計(jì)分析

The two treatment groups would correspond to the test topical product (T) and the RS (R). The statistical analysis should consider a sample of n donors, for which r^T replicate skin sections from the j ^thdonor (j = 1, ? , n) are available for the T group and r^R replicate skin sections from the j ^th donor (j= 1, ? , n) are available for the R group. Each replicate (i) from each donor (j) should have been randomly assigned to each product.

兩個(gè)治療組將對(duì)應(yīng)于自研外用制劑（T）和RS（R）。進(jìn)行統(tǒng)計(jì)分析應(yīng)考慮以下樣本n 組供體，其中r^T復(fù)制第j個(gè)供者的皮膚切片(j = 1.? , n) 適用于T組和r^R復(fù)制第j個(gè)供體的皮膚切片(j = 1.? , n) 可用于R組。每個(gè)供體（j）的每個(gè)復(fù)制品（i）應(yīng)隨機(jī)分配給每個(gè)產(chǎn)品。

Define the following quantities: 定義如下：

T_ij= the natural log-transformed IVPT endpoint (Jmax or AMT) dosed with the test topical product for the i^th skin replicate from the j^th donor (i = 1, 2, ? ,r^T，j =1, 2, ? , n)

T_ij=第j個(gè)供體的第i個(gè)皮膚復(fù)制品的試驗(yàn)外用產(chǎn)品給藥的自然對(duì)數(shù)轉(zhuǎn)換IVPT終點(diǎn)（Jmax或AMT）(i = 1, 2, ? , r^T，j =1, 2, ? , n)

R_ij = the natural log-transformed IVPT endpoint( Jmax or AMT) dosed with the RS for the i^th skin replicate from the j^th donor (i= 1, 2, ? , r^R，

j = 1, 2, ? , n)

R_ij=第j個(gè)供體第i次皮膚復(fù)制的RS給藥的自然對(duì)數(shù)轉(zhuǎn)換IVPT終點(diǎn)（Jmax或AMT）(i= 1, 2, ? , r^R，j= 1, 2, ? , n)

r^T_j= the number of skin replicates from the j th donor dosed with the test topical product ( j = 1, 2, ? , n)

r^T_j=第j個(gè)供體用外用制劑（j = 1, 2, ? , n)

r^Rj = the number of skin replicates from the j th donor dosed with the RS ( j=1, 2, ? , n)

r^Rj=第j個(gè)供體用RS( j=1, 2, ? , n)給藥的皮膚切片的重復(fù)數(shù)

r^∗=r^R₁+r^R₂+……+r^R_n = the total number of skin replicates in the R group

r^∗ =r^R₁+r^R₂+……+r^R_n=R組皮膚切片總數(shù)

n = the number of donors

n = 供體組的數(shù)量

If the numbers of skin replicates available for the final statistical analysis are the same for the n donors for the T group and the R group, the resulting data set is balanced. For simplicity of notation, the common number of skin replicates for one donor for one treatment group in a balanced data set is denoted as r=r^T₁+r^T₂+……=r^T_n=r^R1+r^R2=……r^R_n

如果可用于最終統(tǒng)計(jì)分析的皮膚切片重復(fù)次數(shù)相同的n 組和T組和R組的供體中，所得數(shù)據(jù)集是平衡的。為方便標(biāo)記，在平衡數(shù)據(jù)集中，一個(gè)供體對(duì)一個(gè)治療組的皮膚重復(fù)次數(shù)表示為r=r^T₁+r^T₂+……=r^T_n=r^R1+r^R2=……r^R_n

A diffusion cell may be excluded from among the replicates in a data set when there is a documented observation of a failure (e.g., visual observation that a skin section tears and leaks during the study) or a protocol deviation (e.g., the receptor compartment in a diffusion cell is discovered to be empty at the first sampling time point). In such instances, if sufficient skin remains from the same donor, and no samples from that diffusion cell have been analyzed, a replacement diffusion cell can be set up and studied. Otherwise (if the diffusion cell cannot be replaced) the resulting data set becomes unbalanced.

當(dāng)有失敗的觀察記錄（如：在研究期間觀察到皮膚可見撕裂和泄漏）或方案的偏差（如：擴(kuò)散測(cè)試池中的接收室在第一個(gè)取樣時(shí)間點(diǎn)被發(fā)現(xiàn)為空）時(shí)，擴(kuò)散測(cè)試池可從數(shù)據(jù)集中的重復(fù)數(shù)中排除。在這種情況下，如果來(lái)自同一供體的足夠皮膚仍充足，并且沒有對(duì)該擴(kuò)散池的樣品進(jìn)行分析，則可以采用替代擴(kuò)散池。若沒有可替代擴(kuò)散池，則該結(jié)果數(shù)據(jù)集是不平衡的。

The statistical analysis methods for assessing BE in the cases of a balanced data set and an unbalanced data set are described below. For a donor to be included in the statistical analysis, there should be at least 3 replicate skin sections from the donor for each (T and R) treatment group.

下文描述了在平衡數(shù)據(jù)集和非平衡數(shù)據(jù)集情況下評(píng)估BE的統(tǒng)計(jì)分析方法。對(duì)于納入統(tǒng)計(jì)分析的供體，每個(gè)（T和R）治療組的供體應(yīng)至少有3個(gè)重復(fù)皮膚切片。

Step 1. Determine S_wr,the estimated within-donor standard deviation of the RS, for each of the natural log-transformed IVPT endpoints Jmax and AMT:

步驟1.確定S_wr,，每個(gè)自然對(duì)數(shù)變換IVPT終點(diǎn)Jmax和AMT的RS的估計(jì)供體內(nèi)標(biāo)準(zhǔn)偏差：

(a) If S_wr≥0.294, use the scaled average BE (SABE) approach to determine BE for the individual IVPT endpoint(s) in Steps 2, 3.1, and 4.1

（a） 如果S_wr ≥0.294，使用縮放平均BE（SABE）方法確定步驟2、3.1和4.1中單個(gè)IVPT終點(diǎn)的BE

(b) If S_wr＜0.294, use the regular average BE (ABE) approach through the two one-sided tests (TOST) procedure (Schuirmann, 1987) to determine BE fo the individual IVPT endpoint(s) in Steps 2, 3.2, and 4.2

（b） 如果S_wr＜0.294，使用常規(guī)平均BE（ABE）方法，通過(guò)兩個(gè)單側(cè)測(cè)試（TOST）程序（Schuirmann，1987），在步驟2、3.2和4.2中確定單個(gè)IVPT終點(diǎn)的BE

Step 2. Determine the point estimate for the mean difference of T and R products (I^{^}), its standard error (se(I^{^})), and the corresponding degrees of freedom (df∗).

確定T和R乘積平均差的點(diǎn)估計(jì)(I^{^}), 其標(biāo)準(zhǔn)誤差(se(I^{^}）），以及相應(yīng)的自由度(df∗)

For a balanced data set, determine I^{^}, se(I^{^}) and df∗ by the following: 對(duì)于平衡數(shù)據(jù)集，確定I^{^}, se(I^{^})和df∗通過(guò)以下方式：

For an unbalanced data set, approximate I^{^}, se(I^{^}) and df∗ by using PROC MIXED  (or PROC GLM) in SAS. The example code is provided in Appendix I.

對(duì)于不平衡數(shù)據(jù)集，近似I^{^}, se(I^{^})和df∗ 在SAS中使用PROC MIXED（或PROC GLM）。附錄I中提供了示例代碼。

Step 3.1. Scaled Average BE (SABE) Approach標(biāo)度平均BE（SABE）方法

In the SABE approach, the hypotheses to be tested are: 在SABE方法中，要測(cè)試的假設(shè)是：

Rejection of the null hypothesis supports the conclusion of equivalence of the two products. 否定零假設(shè)支持兩種產(chǎn)品等價(jià)的結(jié)論。

Determine (1-α)*100% upper confidence bound for (μT-μR)2-θσ²_WRbased on Howe’s Approximation (Howe, 1974) (α = 0.05):

Note that t（1-α）,df∗ is （1-α） ∗ 100^th percentile of the Student’s t-distribution with df∗ degrees of freedom and x²（1-α），（r*-n） is （1-α） ∗ 100th percentile of the Chi- square distribution with （r*-n）degrees of freedom

Step 3.2. Regular Average BE (ABE) Approach常規(guī)平均BE（ABE）方法

In the ABE approach, the hypotheses to be tested are: 在ABE方法中，要測(cè)試的假設(shè)是：

Rejection of the null hypothesis supports the conclusion of equivalence of the two products. 否定零假設(shè)支持兩種產(chǎn)品等價(jià)的結(jié)論。

Determine the (1 − 2α)*100% confidence interval for μT-μR (α = 0.05):

where t（1-α）,df∗ is （1-α） ∗ 100th  percentile of the Student’s t-distribution with df∗ degrees of freedom.

Step 4.1. BE Determination with SABE Approach用SABE方法確定BE

For the test product to be bioequivalent to the reference standard, both of the following conditions must be satisfied for each IVPT endpoint tested: 為使自制制劑與參比制劑具有生物等效性，每個(gè)IVPT試驗(yàn)終點(diǎn)必須滿足以下兩個(gè)條件：

a.the 95% upper confidence bound for (μ_T-μ_R)²-θσ²_WR must be less than or equal to zero (numbers should be kept to a minimum of four significant figures for comparison).

b.the point estimate of the T/R geometric mean ratio must fall within the pre- specified limits [1/m，m], where m = 1.2500.

Step 4.2. BE Determination with ABE Approach 采用ABE法確定BE測(cè)定

For the test product to be bioequivalent to the reference standard, the 90% confidence interval for μ_T-μ_R must be contained within the limits [1/m，m] in the original scale for each IVPT endpoint tested, where m = 1.2500.

在IVPT重點(diǎn)測(cè)試中，如果μ_T-μ_R的90%置信區(qū)間必須包含在每個(gè)測(cè)試IVPT終點(diǎn)原始量表中的限值[1/m，m]內(nèi)，其中m=1.2500，則證明自研制劑與對(duì)照制劑具有生物等效