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S5(R3) (MHLW Step 5)S5(R3) (ICH Step 4)


Adopted on 18 February 2020
This Guideline has been developed by the appropriate ICH Expert Working Group and has been subject to consultation by the regulatory parties, in accordance with the ICH Process. At Step 4 of the Process the final draft is recommended for adoption to the regulatory bodies of ICH regions.



1. 緒言及び一般原則

本ガイドラインは、1993年に発出されたICHガイドライン「S5 Detection of Toxicity to Reproduction for Medicinal Products」(医薬品の生殖発生毒性試験)の改定版である。本改定版では、他のICHガイドラインとの整合をとるとともに、用量設定における曝露マージンの利用について詳述し、リスク評価に関する項を設け、さらに適用範囲を拡大してワクチン及びバイオテクノロジー応用医薬品(以下、「バイオ医薬品」)も対象とする。また、代替試験法(以下、「代替法」)に関する適格性確認や使用可能なシナリオについて記載し、生殖発生毒性試験の延期に関する選択肢も提供する。医薬品の生殖発生への影響を評価するためには、一般的に、医薬品及び適切な場合はその代謝物(ICH M3 (1)、ICH S6 (2))の曝露による生殖発生の全ステージへの潜在的影響に関する情報を利用することができる。どのようなガイドラインにおいても、起こりうるすべての事例をカバーするために十分な情報は提供できないため、試験戦略には柔軟性が必要である。


The purpose of this document is to recommend international standards for, and promote harmonization of, the assessment of nonclinical developmental and reproductive toxicity (DART) testing required to support human clinical trials and marketing authorization for pharmaceuticals. The guideline describes potential strategies and study designs to supplement available data to identify, assess, and convey risk. General concepts and recommendations are also provided that should be considered when interpreting study data.
This is a revision of the ICH guideline “S5 Detection of Toxicity to Reproduction for Medicinal Products” that was originally published in 1993. This revision brings the guideline into alignment with other ICH guidelines, elaborates on the use of exposure margins in dose level selection, incorporates a section on risk assessment, and expands the scope to include vaccines and biopharmaceuticals. It also describes qualification of alternative assays, potential scenarios of use, and provides options for deferral of developmental toxicity studies.
To assess a human pharmaceutical’s effect on reproduction and development, there should generally be information available that addresses the potential impact of exposure to a pharmaceutical and, when appropriate, its metabolites (ICH M3 (1), ICH S6 (2)) on all stages of reproduction and development. No guideline can provide sufficient information to cover all possible cases, and flexibility in testing strategy is warranted.

1.1 試験の目的

A) 交尾前~受精(成熟雌雄動物の生殖機能、配偶子の発生及び成熟、交尾行動、受精)
B) 受精~着床(成熟雌動物の生殖機能、着床前発生、着床)
C) 着床~硬口蓋閉鎖(成熟雌動物の生殖機能、胚発生、主要な器官の形成)
D) 硬口蓋閉鎖~妊娠終了(成熟雌動物の生殖機能、胎児の発生と成長、器官の発生と発達)
E) 出生~離乳(分娩と授乳、新生児の子宮外生存への適応、離乳前の発生と成長)
F) 離乳~性成熟(離乳後の発生と成長、自立生存への適応、性成熟の開始と完全な性機能の確立、次世代への影響)

対象集団に関連しない生殖発生ステージを除き、全てのステージにおけるリスクを評価すべきである。各試験でカバーする生殖発生ステージは申請者の判断に委ねられるが、医薬品開発における試験の実施時期については、対象集団や薬剤の開発段階に依存する(ICH M3、ICH S6及びICH S9(3)参照)。

1.1. Aim of Studies
The aim of DART studies is to reveal any effect of the pharmaceutical on mammalian reproduction relevant for human risk assessment. As appropriate, the set of studies conducted should encompass observations through one complete life cycle (i.e., from conception in one generation through conception in the following generation), and permit detection of immediate and latent adverse effects. The following stages of reproduction are generally assessed:
A) Premating to conception (adult male and female reproductive functions, development and maturation of gametes, mating behavior, fertilization).
B) Conception to implantation (adult female reproductive functions, preimplantation development, implantation).
C) Implantation to closure of the hard palate (adult female reproductive functions, embryonic development, major organ formation).
D) Closure of the hard palate to the end of pregnancy (adult female reproductive functions, fetal development and growth, organ development and growth).
E) Birth to weaning (parturition and lactation, neonate adaptation to extrauterine life, pre-weaning development and growth).
F) Weaning to sexual maturity (post-weaning development and growth, adaptation to independent life, onset of puberty and attainment of full sexual function, and effects on second generation).

The risks to all stages should be assessed, unless the stage is not relevant to the intended population. The stages covered in individual studies are left to the discretion of the Sponsor, although the timing of studies within the pharmaceutical development process is dependent on study populations and phase of pharmaceutical development (see ICH M3, ICH S6 and ICH S9 (3)).

2. ガイドラインの適用範囲

本ガイドラインは、バイオ医薬品、感染症ワクチン(及び、ワクチンに含まれる新規構成成分)を含むすべての医薬品、及び新添加剤に適用される。なお、本ガイドラインでは、「医薬品」という用語を、これらすべての治療モダリティを含むものとして使用する。本ガイドラインは、細胞加工製品及び遺伝子治療用製品には適用されない。本ガイドラインで概説する方法論の原則(試験計画、用量設定及び動物種選択など)は、生殖発生毒性試験の実施が適切なすべての化合物に適用される。生殖発生毒性試験の要否及び実施時期を検討するにあたっては、本ガイドラインとICH M3、ICH S6及びICH S9を参照すべきである。


This guideline applies to all pharmaceuticals, including biopharmaceuticals, vaccines (and their novel constitutive ingredients) for infectious diseases, and novel excipients that are part of the final pharmaceutical product. For the purposes of this guideline, the term “pharmaceutical” is used to encompass all of these treatment modalities. This guideline does not apply to cellular therapies, gene therapies and tissue-engineered products. The methodological principles (e.g., study design, dose selection and species selection, etc.) outlined in this guideline apply to all compounds for which the conduct of reproductive and/or developmental toxicity studies is appropriate. This guideline should be read in conjunction with ICH M3, ICH S6, and ICH S9 regarding whether and when nonclinical DART studies are warranted.

3. 生殖毒性評価に関する一般的考慮事項

開発中のほとんどの医薬品については、いくつかの例外がありうるものの、上述した全ての生殖発生ステージを評価すべきである。臨床開発を進めるためには、一般的に以下の3種類のin vivo試験を用いて生殖発生ステージの評価が行われている:1)受胎能及び着床までの初期胚発生に関する試験(FEED試験)(ステージA~B)、2)2種の動物種を用いた胚・胎児発生に関する試験(EFD試験)(ステージC~D)、及び3)出生前及び出生後の発生並びに母体の機能に関する試験(PPND試験)(ステージC~F)。化合物ごとに評価する生殖発生ステージを決定し、実施すべき最も適切な試験を特定すべきである。生殖発生への影響を評価するにあたり、総合的に試験戦略を構築する上で考慮すべき重要な事項を以下に示す。

• 対象患者集団及び使用条件(特に生殖能力及び疾患の重篤度との関連性)
• 医薬品の剤型と臨床適用経路
• 毒性(in vitro、ex vivo及び非哺乳類を用いた試験、並びに構造活性相関も含む)、薬力学、薬物動態及び他の医薬品との薬理学的類似性に関連するデータ
• 医薬品の標的に関する生物学的特性や生殖発生における既知の役割

総合的なリスク評価を損なわない範囲で、動物の使用を最小限に抑える試験戦略をとるべきである。そのアプローチとしては、一般的なデザインの試験を組み合わせた試験の実施(7項参照)や、適切に適格性を確認された代替法(附属書2参照)を用いたリスク評価がある。第Ⅲ相臨床試験前に開発が断念される医薬品も多いことから、ICH M3に示されるように、検討中の臨床試験をサポートする試験(妊娠可能な女性を組み入れるための胚・胎児発生毒性データなど)を適切な時期に実施することで、動物の使用を減らすことが可能である。



The majority of pharmaceuticals being developed should be assessed for all stages of the reproductive cycle identified above, although there can be some exceptions which should be justified, as indicated below. To support clinical development, these stages have typically been evaluated using three in vivo study types: 1) a fertility and early embryonic development study (FEED – stages A and B), 2) embryo-fetal development studies in two species (EFD – stages C and D), and 3) a pre- and a postnatal development study (PPND – stages C through F). For each compound, the stages that are to be evaluated should be determined and the most appropriate studies to conduct should be identified. Key factors to consider when developing an overall integrated testing strategy to evaluate effects on reproduction and development include:
• The targeted patient population and conditions of use (especially in relation to reproductive potential and severity of disease);
• The formulation of the pharmaceutical and route(s) of administration intended for humans;
• Relevant data on toxicity (which can also include data from in vitro, ex vivo and non-mammalian studies, and structure-activity relationships), pharmacodynamics, pharmacokinetics, and pharmacological similarity to other pharmaceuticals;
• Aspects of the general biology of the pharmaceutical target, or known roles of the target in reproduction or development.
These concepts are discussed in more detail throughout the guideline.
To the extent that it does not diminish the overall risk assessment, the experimental strategy should minimize the use of animals. Approaches towards this goal can include the conduct of studies that combine typical study types (see Section 7), as well as appropriately qualified alternative assays for risk assessment (see Annex 2). Since many clinical development programs are terminated prior to Phase 3, animal use can also be reduced by appropriately timing studies to support ongoing clinical development (e.g., embryo-fetal developmental toxicity data to support enrollment of women of childbearing potential) as per ICH M3.
DART studies should, in general, be conducted according to Good Laboratory Practice (GLP) regulations, as they will contribute to the risk assessment. However, if a relevant DART risk is identified in a non-GLP study, repetition of the study to confirm the finding(s) under GLP conditions is not necessarily warranted. A relevant risk is one that occurs at or near intended clinical exposures and is of a nature that is reasonably likely to translate to humans (see Section 9). It is recognized that GLP compliance is not expected for some study types, or aspects of some studies, employing specialized test systems or methods. However, high quality scientific standards should be applied with data collection records readily available. Areas of non-compliance should be identified within the study report and their impact on study results/data interpretation should be considered relative to the overall safety assessment.

3.1 対象患者集団/適応症に関する考慮事項

3.1. Target Patient Population/ Therapeutic Indication Considerations
The intended patient population or therapeutic indication can influence the extent of DART testing. Studies evaluating all stages of reproduction and development are not warranted if the disease indicates that DART will have minimal impact on the risk of the pharmaceutical in the target population. For example, studies covering all stages are not necessarily appropriate for an exclusively post-menopausal female patient population, for use in the pediatric or juvenile pre-pubescent population, or for patient populations in hospitalized settings where pregnancy can be excluded.

3.2 薬理学的考慮事項

3.2. Pharmacology Considerations
Before designing a testing strategy, it should be determined if the intended pharmacologic effects of a pharmaceutical are known to be incompatible with fertility, normal EFD, or assessment of particular endpoints (e.g., a general anesthetic and assessment of mating behavior). This assessment can be based on data with other pharmaceuticals with similar pharmacology, known effects of target engagement, or on knowledge of effects in humans with related genetic diseases. For example, it would be appropriate to modify the design of a PPND study for a pharmaceutical developed to prevent pre-term labor. If the intended pharmacologic effects are incompatible with the study endpoints, testing for a particular reproductive endpoint is not warranted, with justification.

3.3 毒性に関する考慮事項

3.3. Toxicity Considerations
Repeated–dose toxicity studies with sexually mature animals can provide important information on toxicity to reproductive organs that can affect the design of a DART study. The existing toxicology data for the compound should always be considered, taking into account the dose levels, toxicokinetic profile, and dosing duration. For example, the standard fertility study design can be modified to alter the duration of dosing, or the start of cohabitation, for a compound that affects testicular tissue.

3.4 実施時期に関する考慮事項
生殖発生毒性試験の実施時期については、ICH M3、ICH S6及びICH S9に一般的なガイダンスが記載されている。特定の生殖発生毒性を評価する時期は、臨床試験又は対象患者集団において、当該医薬品を安全に使用するために、関連するデータが必要か否かを踏まえて検討すべきである。その結果、特定の生殖発生ステージへの影響を評価する時期を変更することが適切となる場合がある。その他の選択肢については4.2.2項及び4.2.3項で述べる。

3.4. Timing Considerations
General guidance on the timing for conduct of studies assessing reproductive and developmental endpoints is described in ICH M3, ICH S6, and ICH S9. The timing for when to conduct specific DART assessments should take into consideration the need for these data to support the safe use of the pharmaceutical in clinical trials or the intended patient population. Consequently, it can be appropriate to consider altering the timing of the assessment of specific reproductive stages. Additional options are discussed in Section 4.2.2 and 4.2.3.

3.5 トキシコキネティクス(TK)
TKデータの収集に関する一般的な考え方については、ICH S3A (4)に記載されている。

3.5. Toxicokinetics (TK)
Exposure data can be generated in either reproductive (dose range finding (DRF) or pivotal) or repeated-dose toxicity studies. However, given the potential for meaningful changes in TK parameters induced by pregnancy, it is recommended to determine if pregnancy alters exposure. If dose selection is based on exposure ratio (see section 6.1.3), GLP-compliant TK data in pregnant animals is expected. Sampling day(s) should be justified.
When warranted, determination of the pharmaceutical’s concentration in the embryo or fetus can facilitate interpretation of discordant or equivocal evidence of developmental hazard. This information can be collected in a separate study to determine the actual exposure. However, a direct comparison to the potential levels in the human conceptus is not appropriate.
Evidence of lactational excretion can be obtained, when warranted, by sampling milk or by demonstrating exposure in offspring during the pre-weaning period.
General concepts regarding TK data collection are discussed in ICH S3A (4).

4. 哺乳類を用いたin vivo試験のデザインと評価

医薬品の潜在的な生殖発生毒性リスクを評価するための戦略には、一般に、1種以上のin vivo試験が含まれる。一部の動物種(ヒト以外の霊長類(以下、「NHP」)など)では実施不可能であるが、全体としては、生殖発生の全ステージを網羅して評価することが重要である。ほとんどの医薬品では、通常、三試験計画法が適切となろうが、特定の製品ニーズに対応し、また使用動物数を削減するためには、これらの試験デザインを様々に組み合わせることも可能である。FEED試験、EFD試験及びPPND試験の詳細、又はそれらの組合せによる試験については附属書1を参照のこと。各試験でカバーする生殖発生ステージは申請者の判断に委ねられる。医薬品に関して入手し得るすべての薬理学的データ、トキシコキネティクス及び毒性学的データを考慮して、どの試験デザインを選択すべきか判断しなければならない。


The strategy to evaluate the potential reproductive and developmental risk of a pharmaceutical generally includes one or more in vivo studies. The key factor is that, in total, they leave no gaps between stages and allow for evaluation of all stages of the reproductive process, although in some species (e.g., the non-human primate (NHP)) it is not possible to evaluate all stages. For most pharmaceuticals, the 3-study design will usually be appropriate, although various combinations of these study designs can be conducted to address specific product needs and to reduce animal use. Study details for the FEED, EFD, and PPND studies, and combinations thereof, can be found in Annex 1. The stages covered in individual studies are left to the discretion of the sponsor. All available pharmacological, toxicokinetic, and toxicological data for the pharmaceutical should be considered in determining which study design(s) should be used.

4.1 受胎能及び初期胚発生(FEED)に関する戦略

4.1.1 バイオ医薬品に関する考慮事項
げっ歯類又はウサギにおいて薬理学的活性を有するバイオ医薬品の場合は、これらの動物種のいずれかを用いたFEED試験が推奨される。通常イヌやNHPなどの非げっ歯類を用いて交配による評価を行うことは現実的ではない。例えば、NHPが薬理学的に適切な唯一の動物種である場合(多くのモノクローナル抗体の場合など、ICH S6参照)には、少なくとも3カ月間の反復投与毒性試験で得られた生殖器の病理組織学的検査を受胎能の評価の代わりとして使用することができる。この際には、雌雄両方の生殖器の包括的な病理組織学的検査を含めるべきである

4.1. Strategy to Address Fertility and Early Embryonic Development (FEED)
The aim of the FEED study is to test for adverse effects resulting from treatment initiated prior to mating of males and/or females and continued through mating and implantation. This comprises evaluation of Stages A and B of the reproductive process. Results from repeated-dose toxicity studies of at least two weeks duration can often be used to design the fertility study without conducting further dose ranging studies, although studies of such short duration can be insufficient to reveal all adverse effects.
A mating phase is expected in most cases when a FEED study is warranted to support exposure of the target population. Such studies are typically performed in rodents. If no adverse effects on fertility are anticipated, both sexes can be treated and cohabited together in the same study. If effects on fertility are identified in the study, the affected sex should then be determined. In contrast, if adverse effects are anticipated based on mode of action or on the results of repeated-dose studies, each treated sex can be cohabited with untreated animals of the opposite sex. This can be achieved using separate treatment arms within a single study or by the conduct of two separate FEED studies. Reversibility of adverse effects on fertility and early embryonic development can have an important impact on risk assessment.
The FEED study design in female rodents (see Annex 1) allows for the detection of effects on the estrous cycle, tubal transport, implantation, and development of preimplantation stages of the embryo. When estrous/menstrual cycles are evaluated, it is important to obtain baseline cycle data (over 2 or 3 cycles minimum) to distinguish between treatment-related effects and inter/intra animal variability. The monitoring of estrous cyclicity should continue through the time of confirmation of mating.
The FEED study design for male rodents that includes 2 to 4 weeks of treatment prior to cohabitation allows for the detection of effects on spermatogenesis and epididymal transport. When data from repeated-dose studies suggest toxicity to the testis, it can be appropriate to extend the duration of pre-cohabitation treatment to 10 weeks; this permits assessment of effects on the full spermatogenic cycle as well as epididymal transport. The FEED study additionally permits detection of functional effects (e.g., on libido, epididymal sperm maturation, ejaculation) that cannot be detected by histological examinations of the male reproductive organs.
When there is cause for concern based on mode of action or data from previous studies, additional examinations can be included in repeated-dose toxicity and/or fertility studies (e.g., sperm collection for counts and morphology/motility assessments, measuring hormone levels, or monitoring of the estrous/menstrual cycle) to further characterize potential effects on fertility.

4.1.1. Considerations for Biopharmaceuticals
If the biopharmaceutical is pharmacologically active in rodents or rabbits, a FEED study in one of these species is recommended. Mating evaluations are not generally feasible in non-rodents such as dogs and NHPs. For example, if NHPs are the only pharmacologically relevant species (as for many monoclonal antibodies, see ICH S6), histopathological examinations of the reproductive tissues from the repeated-dose toxicity studies of at least three months duration can serve as a substitute for the fertility assessments. Such an approach should include a comprehensive histopathological examination of the reproductive organs from both male and female animals (Note 1). Unless the biopharmaceutical is intended to treat advanced cancer, in which case FEED studies are not warranted, animals should be sexually mature at study initiation in order for an adequate evaluation of the reproductive tissues to be made. These data would only provide information on the structure of the reproductive tissues, as no functional assessment of fertility can be made and predicting effects on fertility and early embryonic development is not always possible based solely on the results of histopathology assessments.

4.2 胚・胎児発生(EFD)に関する戦略
最大推奨臨床用量(Maximum Recommended Human Dose:MRHD)における推定臨床曝露量と同程度の曝露量で、形態異常や胚・胎児致死性(Malformations or Embryo-Fetal Lethality:MEFL)の誘発に関する明らかに陽性の結果が得られれば、開発している医薬品のリスク評価は当該動物種1種で十分と考えられる。

4.2.1 バイオ医薬品に関する考慮事項
バイオ医薬品のEFDへの影響は、2種の動物種(げっ歯類1種及び非げっ歯類1種)がいずれも薬理学的に適切であれば、通常、2種の動物種を用いて評価すべきである。しかしながら、げっ歯類は薬理学的に適切でない場合が多く、その場合には、薬理学的に適切な1種類の非げっ歯類のみを用いてEFDを評価することが可能である。適切な動物種が唯一NHPの場合には、EFD試験の代わりに、ePPND試験を実施することもできる。進行がんの治療を目的としたバイオ医薬品は、通常、薬理学的に適切な動物種1種を用いて評価するだけでよい(ICH S9参照)。生殖発生毒性試験に適したいずれの動物種を用いても、ヒトの標的分子と相同な配列を有する分子(オーソログ)にバイオ医薬品が相互作用せず、適切な動物種が特定できない場合には、ICH S6に記載されているように、サロゲート分子や遺伝子改変動物の使用を検討することが可能である。サロゲート分子を用いて臨床曝露量に対する安全域を算出することは適切ではない。適切な動物種、遺伝子改変動物又はサロゲート分子が利用できない場合には、in vivo生殖発生毒性試験の実施意義はない。その場合は、リスク評価に使用したアプローチ、又は試験を実施しないことの適切性を説明すべきである。

4.2.2 EFDリスクに対処するための代替アプローチ 代替法の利用
胚・胎児発生に対する潜在的ハザードを検出するために、in vitro、ex vivoや非哺乳類を用いたin vivoなどのいくつかの代替法が開発されている。これらの代替法はEFDに対する有害作用に関する創薬スクリーニングに使用され、毒性メカニズムの理解を深める一助となっており、(特にヒト特異的な標的について)非臨床データをヒトでのリスクに外挿する上で役立つ場合もある。これらの目的で代替法を継続的に利用することが推奨される。適格性が確認された場合、その代替法は、従来のin vivo試験の実施を延期又は(特定の状況において)代替する可能性がある。これには、使用動物数を削減できる可能性があるという更なる利点もある。代替法の適格性を確認する際に考慮すべき事項や、代替法の利用が適切なシナリオの例を附属書2に示す。代替法を取り入れたアプローチは、ヒトでの安全性を担保するにあたり、上述した現行試験の枠組みと比較して、少なくとも同等の信頼度を有するべきである。本文書作成時点での科学の進捗を考えると、規制当局の受入れを目的とする場合、段階的なアプローチや組合せによるアプローチの中で、複数の代替法が使用されることが想定される。各代替法の化学的な適用領域、及び代替法の対象となる生物学的メカニズムの特性評価によって使用の範囲が定められ、試験戦略の適格性(当局の受入れの可能性)は、各々の適用範囲内で判断される。

4.2.3 総合的試験戦略の一環としてin vivo本試験を延期することが可能なアプローチ

ICH M3では、2種の動物において予備的な胚・胎児発生(Preliminary EFD:pEFD)毒性データが得られている場合、EFD本試験を実施する前であっても、妊娠可能な女性(Women of Childbearing Potential:WOCBP)を限定的(最大150人のWOCBPを最長3カ月)に臨床試験に組み入れることが可能とされている。これらの考慮事項を踏まえ、本ガイドラインではICH M3を拡大し、第Ⅲ相臨床試験の前にWOCBPを臨床試験に組み入れることが許容されうる2つの追加的オプションを以下に記載する。
1) 1種の動物種における結果を予測する適格性が確認された代替法(附属書2参照)を、第二の動物種のpEFD試験データと組み合わせることで、WOCBPを限定的(最大150人のWOCBPを最長3カ月)に臨床試験に組み入れることを可能とする。その場合、通常、代替法と第二の動物種のpEFD試験データによって、げっ歯類と非げっ歯類の両方の動物種を評価されることになる。
2) 薬理学的に適切な動物種を用いてエンドポイントを追加し(特に、1群あたりの評価可能な同腹児数を増やし、胎児の骨格検査を含める)、GLP下で実施した少なくとも1つのpEFD試験が利用可能な場合、第二の動物種を用いたpEFD試験と組み合わせることにより、すべての地域において、第Ⅱ相までの臨床試験に組み入れるWOCBPの人数に制限を設けないことが可能となる。

4.2. Strategies to Address Embryo-Fetal Development (EFD)
The aim of the EFD studies is to detect adverse effects on the pregnant female and development of the embryo and fetus following treatment (Stage C) of the pregnant female during organogenesis. EFD studies include evaluation of fetal development and survival (Stages C through D).
For most small molecules, effects on EFD are typically evaluated in two species (i.e., rodent and non-rodent (typically rabbit)). At least one of the test species should exhibit the desired pharmacodynamic response. If the pharmaceutical is not pharmacodynamically active in any routinely used species (Section 5.1) then non-routine species (Section 5.2), genetically modified animals, or use of a species-specific surrogate molecule (Section 5.3) (e.g., in the case of oligonucleotides) can be considered, provided there is sufficient characterization of the model to ensure pharmacologic relevance. Genetically modified animals and surrogate molecules are generally most useful for hazard identification, but have limitations when used for risk assessment. Even when there are no relevant models (e.g., the pharmacological target only exists in humans, either normally or in the diseased state), EFD studies should be conducted in two species to detect the adversity of off-target effects or secondary pharmacology.
Clearly positive results for the induction of malformations or embryo-fetal lethality (MEFL), in a single species, at exposures similar to that at the projected clinical exposure at the maximum recommended human dose (MRHD) can be sufficient for risk assessment.
Under limited circumstances, other approaches can be used in place of definitive EFD studies (see Annex 2). Alternatively, there can be adequate information to communicate risk without conducting EFD studies. Evidence suggesting an adverse effect of the intended pharmacological mechanism on EFD (e.g., mechanism of action, phenotypic data from genetically modified animals) can be sufficient to communicate risk.

4.2.1. Considerations for Biopharmaceuticals
The effect of biopharmaceuticals on EFD should typically be assessed in two species (one rodent and one non-rodent) if both are pharmacologically relevant. However, the rodent is often not pharmacologically relevant, in which case EFD assessment in a single pharmacologically relevant non-rodent species can be conducted. In cases where the NHP is the only relevant species, an enhanced pre-and postnatal development (ePPND) study can be conducted instead of an EFD study. Biopharmaceuticals intended for the treatment of advanced cancer typically need only be assessed in a single pharmacologically relevant species (ICH S9).
When no relevant species can be identified because the biopharmaceutical does not interact with the orthologous target in any species relevant to reproductive toxicity testing, use of surrogate molecules or transgenic models can be considered, as described in ICH S6. Calculating safety margins relative to human exposures with surrogate molecules is not appropriate. If there are no relevant species, genetically modified animals or surrogates available, in vivo reproductive toxicity testing is not meaningful. In this case, the approach used for risk assessment, or rationale for not conducting studies, should be justified.

4.2.2. Alternative Approaches for Addressing EFD Risk Use of Alternative Assays
A number of alternative in vitro, ex vivo, and non-mammalian in vivo assays (alternative assays) have been developed to detect potential hazards to embryo-fetal development. They have been used as drug discovery screens for adverse effects on EFD and have assisted in the understanding of the mechanism of toxicity, which can be useful for translating nonclinical data to human risk (especially for human-specific targets).
The continued use of alternative assays for these purposes is encouraged.
If properly qualified, alternative assays have the potential to defer or replace (in certain circumstances) conventional in vivo studies. This has the added benefit of potentially reducing animal use. Concepts to consider when qualifying these assays, and examples when the use of such assays could be appropriate, appear in Annex 2. Approaches that incorporate alternative assays should provide a level of confidence for human safety assurance at least equivalent to that provided by the current testing paradigms described above. Based on the direction of scientific development as of the writing of this document, it is expected that for regulatory purposes multiple alternative assays will be used within a tiered or battery approach. These testing strategies will be qualified within a certain context of use, which is defined by the chemical applicability domain of the assay, and by characterization of the biological mechanisms covered by the assay.

4.2.3. Potential Approaches to Defer Definitive In Vivo Testing as Part of an Integrated Testing Strategy
The design of an appropriate testing strategy relies on a cumulative weight-of-evidence approach. ICH M3 allows preliminary embryo-fetal developmental (pEFD) toxicity data from two species to support the limited inclusion of women of childbearing potential (WOCBP) (up to 150 WOCBP for up to 3 months) before conducting definitive EFD studies. Based on these considerations, this guideline expands on ICH M3 by allowing two additional options to support inclusion of WOCBP prior to Phase 3 clinical trials:
1) Qualified alternative assays which predict the outcome in one species (see Annex 2), can be combined with a pEFD from a second species to enable the limited inclusion of WOCBP (up to 150 WOCBP for up to 3 months). The alternative assay and the second species should generally cover both a rodent and a non-rodent species.
2) Additional endpoints incorporated into at least one GLP pEFD study (specifically increasing the group size of evaluable litters with inclusion of skeletal examinations) performed in a pharmacologically relevant species, if available, combined with a pEFD in a 2nd species allows all regions to include an unlimited number of WOCBP in clinical trials through Phase 2.

4.3 出生前及び出生後の発生並びに母体の機能(PPND)に関する戦略
小児用医薬品の開発にあたり、改変されたPPNDやePPND試験デザインを検討している場合は、ICH S11(5)を参照すること。

4.3.1 バイオ医薬品に関する考慮事項
NHPのみで評価可能な医薬品に関しては、ePPND試験により限定的な出生後評価が可能であるが、出生児を成熟までの期間を通して評価することは現実的ではない(附属書1及びICH S6参照)。

4.3. Strategy to Address Effects on Pre- and Postnatal Development (PPND)
The aim of the PPND study is to detect adverse effects following exposure of the maternal animal from implantation through weaning to evaluate effects on the pregnant or lactating female and development of the offspring. Since manifestations of effects induced during this period can be delayed, development of the offspring is monitored through sexual maturity (i.e., Stages C to F). The rodent is usually used to assess PPND; however, other species can be used as appropriate (See Annex 1).
In most cases, a preliminary (dose range finding) PPND study is not warranted, because the appropriate information is generally available from prior studies. However, a preliminary PPND study with termination of the pups before or at weaning can be used to select dose levels or inform study design and/or to provide pup exposure data.
If a modified PPND/ePPND study design is being considered to support pediatric development, see ICH S11 (5).

4.3.1. Considerations for Biopharmaceuticals
For pharmaceuticals that can only be tested in the NHP, the ePPND study can provide a limited assessment of postnatal effects, but it is not generally feasible to follow the offspring through maturity (See Annex 1 and ICH S6).

5. 試験系の選択

5.1 通常の試験動物種

5.1.1 生殖発生毒性試験の動物種の選択

5.1.2 予防用及び治療用ワクチンのための動物種選択

5.2 通常用いられない動物種
NHPは通常用いられない試験動物種と考えるべきである。ICH S6に記載されているとおり、霊長類でのみ薬理学的活性を有するバイオ医薬品では、胚・胎児発生及び出生後早期の発達に対する影響の評価に通常、NHPが用いられる。ただし、NHPを用いて生殖発生毒性リスクを評価する際には、制約のあるエンドポイントがあることにも考慮する。(附属書1及びICH S6参照)。

5.3 病態モデル動物、遺伝子改変動物及びサロゲート分子の使用


5.1. Routine Test Species
Mammalian species should be used to detect DART. The use of the same species and strain as in already completed toxicity studies can eliminate the need to use additional animals or conduct additional studies to characterize pharmacokinetics and metabolism, and/or for dose range finding. The species used should be well-characterized and relevant for detecting effects on the endpoints in a particular study (e.g., with respect to health, fertility, fecundity, background rates of malformation and embryo-fetal death, etc.).

5.1.1. Selection of Species for DART Testing
The rat is generally appropriate for DART testing and is the most often used rodent species for reasons of practicality, general knowledge of pharmacology in this species, the extensive toxicology data usually available for interpretation of nonclinical observations and the large amount of historical background data. The mouse is also often used as the rodent species for many of the same reasons.
For assessment of EFD only, a second mammalian non-rodent species is typically evaluated, although there are exceptions (e.g., vaccines and biopharmaceuticals, see Sections 5.1.2 and 5.2, respectively). The rabbit has proven to be useful in identifying human teratogens that have not been detected in rodents and is routinely used as the non-rodent species based on the extensive historical background data, availability of animals, and practicality.

5.1.2. Species Selection for Preventative and Therapeutic Vaccines
The animal species selected for testing of vaccines (with or without adjuvants) should demonstrate an immune response to the vaccine. The type of developmental toxicity study conducted, and the choice of the animal model, should be justified based on the immune response observed and the ability to administer an appropriate dose. Typically, rabbits, rats, or mice are used in developmental toxicity studies for vaccines. Even though quantitative and qualitative differences can exist in the responses (e.g., in humoral and cellular endpoints) between species, it is usually sufficient to conduct developmental toxicity studies in a single species. Although the degree and time course of transfer of maternal antibodies across the placenta varies between species, a developmental toxicity study in rabbits, rats, or mice can still provide important information regarding potential embryo-fetal toxicity of the vaccine components/formulation and safety of the product during pregnancy. NHP should be used only if no other relevant animal species demonstrates an immune response.
When there is a lack of an appropriate animal model (including NHP), an EFD toxicity study in rabbits, rats, or mice can still provide important information regarding potential embryo-fetal toxicity of the vaccine components/formulation and safety of the product during pregnancy.

5.2. Non-routine Test Species
Species other than the rat, mouse or rabbit can be used to evaluate the effects of pharmaceuticals on various reproductive stages. When considering the use of other species, their advantages and disadvantages (summarized in Table 1 of Annex 1) should be considered in relation to the pharmaceutical being tested, the study design and selected endpoints, and the ability to extrapolate results to the human situation.
NHPs should be considered a non-routine test species. They are most typically used for evaluating effects on embryo-fetal development and early postnatal development for biopharmaceuticals that are only pharmacologically active in primates, as described in ICH S6. However, there are additional considerations that limit the utility of studies in NHPs for assessing some endpoints for DART risk assessment (see Annex 1 and ICH S6).

5.3. Use of Disease Models, Genetically Modified Models, and Surrogate Molecules
Animal models of disease, genetically modified models, and surrogate molecules can be valuable for investigating the effect of the intended pharmacology on development and reproduction. Studies in disease models can be of value in cases where the data obtained from healthy animals could be misleading or otherwise not apply to the disease conditions in the clinical setting. The model should be pharmacologically relevant and appropriate for the development and reproductive endpoints being assessed. The pathophysiology of the disease course in the model should be characterized. Some differences from the human pathophysiology would not preclude its use if these are unlikely to confound data interpretation. Animal-to-animal variability should be characterized and appropriate within the context of the study. If historical control information is limited, reference data for the study endpoints should be available or should be generated during the study to aid data interpretation.
Genetically modified models can be used to provide information about on-target effects of a pharmaceutical on DART parameters through permanent or conditional alterations in target activity. Such models can inform on whether the biology of the target is closely linked to adverse effects on reproduction and development in routine test species.
When the pharmaceutical does not have adequate activity against the target in the routine test species, surrogate molecules can be used to assess potential adverse effects on reproduction and development.

6. 用量設定、投与経路及び投与スケジュール

生殖発生毒性試験での用量、投与スケジュール及び投与経路は、試験デザインを検討する上で重要な事項であり、入手可能なあらゆる情報(薬理作用、反復投与毒性、薬物動態、用量設定試験など)に基づいて設定すべきである。低分子及びバイオ医薬品の用量設定の原則に関するガイダンスは、それぞれICH M3及びICH S6に示されている。試験系における忍容性に関する十分な情報が入手できない場合には、用量設定試験を実施することが望ましい。

6.1 用量設定

6.1.1 毒性に基づく用量設定指標

• 体重変化(変化量又は絶対値;増加・減少)。一過性のわずかな体重増加量又は体重の変化は、用量設定の根拠として適切ではない。体重変化の影響を評価する際には、試験における投与期間全体を考慮すべきである。
• 過剰な薬理作用(過度な鎮静や低血糖など)
• 毒性学的反応(けいれん、著しく高い胚・胎児死亡率、臨床病理学的な変動など)。計画した生殖発生毒性試験の評価に影響を及ぼす可能性がある特定の標的臓器毒性。

6.1.2 全身曝露の飽和に関する用量設定指標

6.1.3 曝露マージンに基づく用量設定指標
MRHDにおける曝露量に対して予測される曝露マージンを示すことで、用量設定の適切性を示すことは可能である。低分子の場合、MRHDにおけるAUC又はCmaxを十分に上回る全身曝露が得られるのであれば、高用量設定において曝露量を用量設定指標とすることは可能である。妊娠動物における曝露量が、MRHDにおける曝露量の25倍を超えるのであれば、通常、生殖発生毒性試験における最大用量として適切である(注2)。25倍の曝露マージンは、GLPに準拠した用量設定試験/pEFD試験又は本試験で確定すべきである。通常、当該曝露マージンは未変化体に基づいて計算するが、ヒトでの主要な代謝物についても十分な曝露マージンを確保することを検討すべきである(ICH M3及び ICH M3 Q&A参照)。プロドラッグにおいて、特に未変化体に対する活性代謝物の曝露量の比率がヒトと比べて低い試験動物種の場合、活性代謝物に基づいて曝露マージンを確定することがより適切である。当該曝露量の比較においては、未変化体又は代謝物を選択した根拠を示すべきである。
MRHDにおける曝露量の25倍を超える曝露でのみ、試験動物種において薬力学的活性がみられる医薬品では、過度な薬理作用の有害作用を評価する目的で、より高用量での評価が求められる場合があるが、リスク評価をする上で不適切なオフターゲット作用が発現しやすい。曝露量に基づくエンドポイントをEFD試験における用量設定の根拠とする場合には、GLP試験における妊娠動物のTKデータが求められる。当該曝露量として、結合型と非結合型を合わせた曝露量と非結合型の曝露量のいずれを選択するかについては適切性を示すべきであり、ICH S3Aに概説されているとおり、非臨床開発プログラム全体との整合性がなければならない。 バイオ医薬品における曝露量に基づくアプローチ
ICH S6に示されている通り、曝露量に基づくマージンを示すことにより、バイオ医薬品の用量設定の適切性を示すことが可能である。一般に、非臨床試験に用いる動物種において意図する薬理作用が最大となる用量、あるいはMRHDでの曝露量の10倍程度の曝露が得られる用量のいずれか高い方とすべきである。標的結合親和性の種差及びその他の関連要因による用量調節についてはICH S6を参照のこと。

6.1.4 投与可能な最大用量(MFD)に基づく用量設定指標
投与経路/投与頻度、及び試験動物種の解剖学的/生理学的特性に関連した原薬(又は製剤)の物理化学的特性によって、投与可能な医薬品の用量が制限される場合には、MFDを高用量の設定に用いることが可能である。ICH M3 Q&A (1)に示されている通り、MFDを用いて高用量を設定する際には、投与量を最大にするよりも、試験動物種での曝露が最大となるよう設定すべきである。なお、1日あたりの可能な総曝露量を増やすために、投与頻度の変更を検討することもできる(6.3項参照)。

6.1.5 限界量に基づく用量設定指標
1 g/kg/日未満の用量段階で、高用量設定のための要素が満たされない場合、一般に限界量として1 g/kg/日を適用することができる(その他の考慮事項についてはICH M3参照)。

6.1.6 高用量以外の用量設定

6.2 投与経路

6.3 投与スケジュール

6.4 ワクチンの用量設定及び試験デザイン


The choice of dose levels, schedule and route of administration are important study design considerations and should be based on all available information (e.g., pharmacology, repeated-dose toxicity, pharmacokinetics, and dose range finding studies). Guidance on the principles of dose selection for small molecules and biopharmaceuticals is given in ICH M3 and ICH S6, respectively. When sufficient information on tolerability in the test system is not available, dose range finding studies are advisable.

6.1. Dose Selection
There are a number of dose selection endpoints that can be used for DART studies. All endpoints discussed in this section are considered equally appropriate in terms of study design. The high dose in the definitive studies should be one that is predicted to comply with one or more of the concepts set forth in sections 6.1.1 to 6.1.5 below. The selected doses should take into account observations made in previous studies (e.g., repeated-dose, TK, DRF, etc.). There can be instances where fewer than three dose levels are sufficient to provide the necessary information for risk assessment.
Justification for high dose selection using endpoints other than those discussed below can be made on a case-by-case basis.

6.1.1. Toxicity–based Endpoint
This endpoint is based on inducing a minimal level of toxicity in the parental animals at the high dose. Factors limiting the high dose determined from previously conducted studies could include, but are not limited to:
• Alterations in body weight (gain or absolute; either reductions or increases). Minor, transient changes in body weight gain or body weight are not appropriate for dose selection. When assessing weight change effects, the entire dosing duration of the study should be considered.
• Exaggerated pharmacological responses (e.g., excessive sedation or hypoglycemia)
• Toxicological responses (e.g., convulsions, excessive embryo-fetal lethality, clinical pathology perturbations). Specific target organ toxicity that would interfere with the study endpoints within the duration of the planned DART study.

6.1.2. Saturation of Systemic Exposure Endpoint
High dose selection based on saturation of systemic exposure measured by systemic availability of pharmaceutical-related substances can be appropriate. There is little value in increasing the administered dose if it does not result in increased plasma concentration of parent or metabolites.

6.1.3. Exposure Margin Based Endpoint
It can be appropriate to select doses based on predicted exposure margins relative to the exposure at the MRHD. For small molecules, a systemic exposure representing a large multiple of the human AUC or Cmax at the MRHD can be an appropriate endpoint for high dose selection. Doses providing an exposure in pregnant animals > 25˗fold the exposure at the MRHD are generally considered appropriate as the maximum dose for DART studies (Note 2). The 25-fold exposure margin should be established in a GLP-compliant dose range finding/pEFD or definitive study. Usually this multiple should be determined based on parent drug levels; however, consideration should also be given to ensuring an adequate exposure margin to major human metabolites (see ICH M3 and ICH M3 Q&A). In the case of prodrugs, it can be more appropriate to establish the exposure multiple on the basis of the active metabolite, particularly if the test species has a lower ratio of active metabolite to prodrug, compared to humans. The basis for the moiety used for comparison (parent drug or metabolite) should be justified.
For pharmaceuticals that have demonstrated pharmacodynamic activity in the test species only at exposures > 25-fold that projected at the MRHD, higher doses can be warranted to assess adverse effects of exaggerated pharmacology. However, irrelevant off-target effects are more likely to be observed.
When exposure-based endpoints are used as the basis for selection of the dose levels for EFD studies, TK data from pregnant animals in a GLP-compliant study is expected. The choice for the use of total vs. fraction unbound pharmaceutical exposures should be justified and consistent with the entire nonclinical development program as outlined in ICH S3A. Exposure-based Approach for Biopharmaceuticals
Exposure-based margins can be appropriate to select doses for biopharmaceuticals as per ICH S6. Generally, the dose should provide the maximum intended pharmacological effect in the preclinical species or provide an approximately 10-fold exposure multiple over the maximum exposure to be achieved in the clinic, whichever is higher. ICH S6 should be consulted with regard to dose adjustment for differences in target binding affinity and other relevant factors.

6.1.4. Maximum Feasible Dose (MFD) Endpoint
The MFD can be used for high dose selection when the physico-chemical properties of the pharmaceutical (or formulation) associated with the route/frequency of administration and the anatomical/physiological attributes of the test species limit the amount of the pharmaceutical that can be administered. Use of the MFD should maximize exposure in the test species, rather than maximize the administered dose, as per ICH M3 Q&A (1). Note that changes to the frequency of dose administration can be considered to increase the total feasible daily exposure (see Section 6.3).

6.1.5. Limit Dose Endpoint
A limit dose of 1 g/kg/day can generally be applied when other dose selection factors have not been attained with lower dose levels (see also ICH M3 for other considerations).

6.1.6. Selection of Lower Dose Levels
It is generally desirable to establish a no observed adverse effect level (NOAEL) for DART. The selection of lower dose levels should take into account exposure, pharmacology, and toxicity, such that the dose-response of findings can be established when appropriate. The low dose should generally provide a low multiple (e.g., 1 to 5-fold) of the human exposure at the MRHD. Dose levels that yield exposures that are sub-therapeutic in humans should be justified.

6.2. Route
In general, the route of administration should be the clinical route. If, however, sufficient exposure cannot be achieved using the clinical route or the clinical route is not feasible, a different route should be considered. When multiple routes of administration are being evaluated in humans, a single route in the test species can be adequate provided that sufficient systemic exposure is achieved compared to that of all clinical routes and that there is adequate coverage for the metabolites.

6.3. Schedule
Dosing schedules used in the toxicity studies determine the exposure profile, which can be important in the risk assessment. Although mimicking the clinical schedule is often sufficient, a more or a less frequent schedule can be appropriate. For example, twice daily dosing can be warranted with compounds that are quickly metabolized in the test species, although pragmatic factors (e.g., study logistics, stress on animals) should be considered when a more frequent schedule is contemplated. It can also be important to alter the dosing schedule to ensure that adequate exposure is obtained at all critical stages of reproduction and/or development being evaluated in a given study.

6.4. Dose Selection and Study Designs for Vaccines
This guideline covers vaccines (adjuvanted or not) used in both preventative and therapeutic indications against infectious diseases. While not within the scope of this guideline, the principles outlined can be applicable to the nonclinical testing of vaccines for other indications as well (e.g., cancer).
The types of reproductive and/or developmental toxicity studies used for preventative and therapeutic vaccines depend on the target population for the vaccine and the relevant reproductive risk. Generally, DART studies are not warranted for vaccines being developed for neonates, pre-pubertal children, or geriatric populations.
For reproductive toxicity studies of vaccines, it is typically sufficient to assess a single dose level capable of eliciting an immune response in the animal model (Section 5.1.2), using the clinical route of administration. This single dose level should be the maximum human dose without correcting for body weight (i.e., 1 human dose = 1 animal dose). If it is not feasible to administer the maximum human dose to the animal because of a limitation in total volume that can be administered, or because of dose-limiting toxicity, whether local or systemic, a dose that exceeds the human dose on a mg/kg basis can be used. To use a reduced dose, justification as to why a full human dose cannot be used in an animal model should be provided.
The vaccination regimen should maximize maternal antibody titers and/or immune response throughout the embryonic, fetal, and early postnatal periods. Timing and number of doses will depend on the onset and duration of the immune response of the particular vaccine. When developing vaccines to be given during pregnancy, a justification should be provided for the specific study design, based upon its intended use (e.g., protecting the mother during pregnancy or protecting the child early postnatally).
Daily dosing regimens can lead to overexposure to the vaccine constituents. Episodic dosing of pregnant animals rather than daily dosing is recommended. Also, episodic dosing better approximates the proposed clinical immunization schedule for most preventive and therapeutic vaccines. Considering the short gestational period of routine animal species, it is generally recommended to administer a priming dose(s) to the animals several days or weeks prior to mating in order to elicit peak immune response during the critical phases of pregnancy (i.e., the period of organogenesis). The dosing regimen can be modified according to the intended vaccination schedule in humans.
At least one dose should be administered during early organogenesis to evaluate potential direct embryotoxic effects of the components of the vaccine formulation and to maintain a high antibody response throughout the remainder of gestation. If embryo-fetal toxicity is observed, this can be further assessed using subgroups of animals that are dosed at certain time points.
In cases where a vaccine includes a novel active constitutive ingredient (including novel adjuvants), consideration of additional testing strategies similar to those for non-vaccine products can be appropriate.

7. げっ歯類を用いた組合せによる試験計画法



Although three separate study designs, i.e., FEED (stages A and B), EFD (stages C through D) and PPND (stages C through F) have been employed to develop the majority of pharmaceuticals, various combinations of these study designs can be conducted to reduce animal use. The main advantage of combination designs is that all relevant stages of the reproductive process can be assessed using fewer animals. Combination studies can also better mimic the exposure duration in the clinic, especially for drugs with long half-lives. A common combination study design is a combined Fertility and EFD study (stages A through D) with a separate PPND study (stages C through F).
Designs and study details for FEED, EFD, and PPND studies, and combinations thereof, can be found in Annex 1.
In cases where no effects on male or female fertility are anticipated, or where extending the dosing period is appropriate due to observation of reproductive organ toxicity in a repeated-dose toxicity study, a combination design of repeated-dose and fertility studies can be considered. After a defined dosing period within the repeated-dose toxicity study, males can be paired with sexually mature females (whether untreated, or dosed for at least two weeks prior to mating). This combination study can reduce the number of animals used, but the number of mating pairs per group should be at least 16. Further, if treated, dosing of females can be extended until the end of organogenesis, thereby allowing evaluation of EFD endpoints (Annex 1).

8. データの報告及び統計

8.1 データの報告

8.2 統計


8.1. Data Reporting
Individual values should be tabulated in a clear concise manner to account for all animals in the study. The data tables should allow ready tracking of individual animals and their conceptuses, from study initiation through study conclusion.
Fetal morphologic abnormalities should be described using industry-harmonized terminology. All findings for each litter should be clearly listed by conceptus. Summary listings should be prepared by type of abnormality. The inclusion or exclusion of data from non-pregnant animals in summary tables should be clearly indicated.
Interpretation of study data relies primarily on comparison with the concurrent control group. Historical control/reference data can be used to assist data interpretation. Recent historical control data from the performing laboratory is preferable. Contemporary data typically from a five-year period is desirable and permits identification of genetic drift.

8.2. Statistics
Statistical testing to assess the significance of differences between the treated and control groups is expected in definitive studies. Many of the datasets from DART studies do not follow a normal distribution, necessitating the use of non-parametric statistical methods. Cesarean, fetal and postnatal data summary statistics should be calculated using the litter as the unit of analysis. Statistical significance need not convey a positive signal, nor lack of statistical significance impute absence of effect. Determination of biological plausibility, based on all available pharmacologic and toxicologic data, is often useful.

9. リスク評価の原則

これまでの項で述べたように、臨床試験及び製造販売承認後において、使用条件下でのヒトにおける潜在的な生殖発生リスクに対処するにあたっては、当該医薬品、関連化合物、ヒト遺伝学から得られた入手可能なデータ、及び当該医薬品の標的分子がもつヒトの生殖における役割に関する知識をすべて利用すべきである。制限事項(試験系の適切性、最大曝露量など)、不確実性、非臨床における生殖発生毒性データパッケージ内のデータの相違点については、いずれもその影響を評価すべきである。一般的に、十分な曝露量下で適切な動物種を用いて実施されたin vivo本試験の結果は、代替法や予備試験から得られる結果よりも重視される。随時新たな情報が得られるため、リスク評価は製品の開発期間を通じて継続的に行われる。生殖発生毒性試験で報告されるすべての所見が有害というわけではない。所見が有害だと思われる場合は、科学的根拠の重み付けにより、いくつかの要素を検討しながらリスク評価すべきである。これには、曝露マージン、生物学的妥当性、用量反応関係、回復性、用量を制限するような親動物毒性の可能性及び動物種間での一致が含まれる。稀な形態異常が認められた場合、用量相関性がないとしても、必ずしも懸念が低くなるとは限らない。
授乳に関して特に実施するリスク評価は、in vivo分娩試験(PPND又はePPND)により特定されたハザードに基づいて行う。これらのハザードには、乳汁中への薬物の分泌に起因する出生児の成長と発達に対する有害作用が含まれる。分娩試験で出生児の全身曝露データが得られた場合には、ヒト乳児で推定される授乳による曝露と比較することができる。乳汁の成分は動物種間で異なるため、動物の乳汁中薬物濃度をヒトの乳汁中薬物濃度と直接定量的に相関させることはできないが、動物の乳汁中に薬物が存在することは、一般に、ヒトの乳汁中にも薬物が存在することを示す。


As described in the preceding sections of this guideline, all available data garnered from the pharmaceutical, related compounds, human genetics, and knowledge of the role of target biology in human reproduction should be used to address potential reproductive risks in humans under the conditions of use, both during clinical trials and after marketing authorization. Any limitations (e.g., test system relevance, achieved exposure), uncertainties and data gaps in the available nonclinical DART data package should be addressed and their impact assessed. Generally, the results from definitive in vivo studies in an appropriate species with adequate exposures carry more weight than those from alternative assays or preliminary studies. Risk assessment is a continuous process through product development as more information becomes available.
Not all findings reported in DART studies are adverse. When a finding is deemed adverse, several factors should be considered in a weight-of-evidence evaluation for risk assessment. These can include exposure margins, biological plausibility, evidence of a dose-response relationship, potential for reversibility, the potential for confounding parental toxicity, and evidence for cross-species concordance. For rare malformations, the absence of increased frequency with dose does not always alleviate concern.
Comparison of pharmaceutical exposure at the NOAEL in the test species to the exposure at the MRHD is an important component of the risk assessment. This comparison should be based on the most relevant metric (e.g., AUC, Cmax, Cmin, body surface area-adjusted dose). In general, there is increased concern when the NOAEL occurs at exposures less than 10-fold the human exposure at the MRHD; above this threshold, concern is reduced. Effects that are limited to occurrence at more than 25-fold the human exposure at the MRHD are usually of minor concern for the clinical use of the pharmaceutical. The most relevant margin is generally the exposure metric in the most sensitive species, unless appropriately justified otherwise. Biological plausibility is assessed by comparison of pharmacologic mechanism of action with the known role of the target in reproduction or development. A finding that can be interpreted as a consequence of pharmacology suggests that it will be of concern for humans. This relationship is further strengthened by evidence that the finding is dose-related, whether characterized as increasing incidence or severity. Absence of biological plausibility does not preclude off-target toxicity, particularly if this is characterized by a dose-response relationship.
Understanding the potential for reversibility will alter the risk assessment. Effects on male and female fertility that are reversible after cessation of treatment are of less concern. Conversely, critical irreversible developmental endpoints, such as death or malformation, are of increased concern. Other forms of developmental toxicity (e.g., growth retardation, functional deficits), may or may not be reversible. Generally, transient findings (e.g., skeletal variations, such as wavy ribs in rodents) are of less concern when they occur in isolation. Similarly, variations that are indicative of growth retardation in the presence of reduced fetal weight are of less concern. However, an overall increase in the incidence of variations (qualitatively similar or not) can suggest increased concern for dysmorphogenesis in the presence of an equivocal increase in malformations.
The role of parental toxicity should be considered in determination of the relevance of findings. Embryo-fetal toxicity observed in the presence of maternal toxicity should be considered carefully to determine the likelihood that the finding is relevant for humans. Specifically, evaluation of the concordance between individual litter findings and the severity of maternal toxicity in the dam could be helpful in this assessment. It should not be assumed that developmental toxicity is secondary to maternal toxicity, unless such a relationship is demonstrated de novo, or relevant published literature can be cited.
Also, consistency of findings reported among studies, or between species can strengthen the concern for an adverse effect. Increased fetal lethality seen in a rodent EFD study that is consistent with decreased live litter sizes in the PPND study is an example of cross-study concordance. Observations of increased post implantation loss in rats and rabbits is an example of cross-species concordance. Further knowledge of the mechanism of reproductive or developmental effects identified in animal studies can help to explain differences in responses between species and provide information on the human relevance of the effect (e.g., corticosteroid-induced cleft palate in mice).
A specific risk assessment conducted for breastfeeding would be predicated on hazards identified by the in vivo littering study (PPND or ePPND). These hazards can include adverse effects on offspring growth and development that are attributed to excretion of the pharmaceutical in the milk. Systemic exposure data in the pups from the littering study, if available, can also be compared with projected lactational exposures in the human infant. While interspecies differences in milk composition preclude a direct quantitative correlation of animal milk levels to human milk levels of a pharmaceutical, the presence of pharmaceutical in animal milk generally indicates the presence of pharmaceutical in human milk.
Lastly, available human data can influence the overall assessment of human reproductive risk.

10. 注釈

注2:ヒトに対する催奇形性物質として既知あるいは推定される22の化合物を解析したところ、MEFLが認められたケースでは、少なくとも1種の動物種において、最小毒性量(LOAEL)での曝露量がMRHDでの曝露量の6倍未満であった(Andrews et al. (6))。このことは、EFD試験での高用量選択の際に、25倍を超える曝露量比を用いればこれらすべての医薬品に対する催奇形性のハザードを十分検出できることを示している。本解析では、動物でMEFLが検出されたヒト催奇形性物質に関して、少なくとも1種の動物種におけるNOAELでの曝露量がMRHDでの曝露量の4倍未満であったことも示された。
さらにIQ DruSafeリーダーシップグループによりEFD試験に関する調査が行われた(Andrews et al. (7))。この調査から、例えば、用量を制限するような母動物毒性が発現しない条件下において、ヒト(想定される治療用量での曝露量)に対して動物での未変化体の曝露量比が15倍以上に達していたEFD本試験は、ラットで153件、ウサギで128件であったことが明らかとなった。これらのデータによると、母動物毒性が認められない場合(認められれば高用量投与は制限される)、ヒト曝露量の25倍以上の曝露量を達成するよう動物へ投与しても、MEFLは稀にしか認められない。これらすべての場合において、MEFLは50倍を超える曝露量まで認められず、このような高曝露量での所見がヒトでのリスク評価に適しているとは考えられない。そのため、用量を制限するような母動物毒性が発現しない場合、EFD及びPPND試験の高用量として、MRHDでの総未変化体濃度におけるヒト血中曝露量に対する曝露量比が25倍を超える用量とすることは理にかなっており、ヒトのリスク評価に適した結果を検出するのに十分であると考えられる。


Note 1: In particular, the testes and epididymides should be sampled and processed using methods which preserve the tissue architecture of the seminiferous epithelium. A detailed qualitative microscopic evaluation with awareness of the spermatogenic cycle is a sensitive means to detect effects on spermatogenesis. While generally not warranted, additional experimental endpoints (e.g., immunohistochemistry, homogenization resistant spermatid counts, flow cytometry, quantitative analysis of staging) can be incorporated into the study design to further characterize any identified effects. In females, a detailed qualitative microscopic examination of the ovary (including follicles, corpora lutea, stroma, interstitium, and vasculature), uterus and vagina should be conducted with awareness of the reproductive cycle and the presence of primordial and primary follicles.
Note 2: An analysis of 22 known human or presumed human teratogens showed that if MEFL was observed, exposure at the lowest observed adverse effect level (LOAEL) in at least one species was < 6-fold the exposure at the MRHD (Andrews et al. (6)). This indicates that using a > 25-fold exposure ratio for high-dose selection in the EFD toxicity studies would have been sufficient to detect the teratogenic hazard for all these pharmaceuticals. The analysis also showed that for human teratogens that were detected in animal species, the exposure at the NOAEL in at least one species was < 4-fold the exposure at the MRHD.
In addition, a survey was conducted on EFD toxicity studies by the IQ DruSafe Leadership Group (Andrews et al. (7)). This survey identified 153 and 128 definitive rat and rabbit EFD studies, respectively, that achieved ≥ 15-fold animal to human parent drug exposure ratios (using human exposure at the intended therapeutic dose) in the absence of confounding (i.e., dose-limiting) maternal toxicity. These data show that dosing animals to achieve exposures ≥ 25-fold human exposures when there is no maternal toxicity (that would otherwise limit the high dose), only infrequently detects MEFL. In all these cases, MEFL findings were not observed until exposures exceeded 50-fold and findings at such high exposures are not believed to be relevant to human risk assessment. In the absence of confounding maternal toxicity, the selection of a high dose for EFD and PPND studies that represents a > 25-fold exposure ratio to human plasma exposure of total parent compound at the intended maximal therapeutic dose is therefore considered pragmatic and reasonably sufficient for detecting outcomes relevant for human risk assessment.

11. 用語

代替法:形態異常や胚・胎児致死性(MEFL参照)を予測することを目的としたin vitro、ex vivo又は非哺乳類in vivo試験法。


代替法の適格性確認(規制当局の受入れ目的):in vivoで認められるMEFLを特定する上での代替法の予測性の確認。



GD 0:妊娠0日。交尾成立が確認(げっ歯類では膣スメアによる精子確認/膣栓、ウサギでは交尾の確認など)された日。


予備的EFD(pEFD)毒性試験:器官形成期に曝露を行う胚・胎児発生毒性試験で、適切な用量段階を設定し、各群6匹以上の妊娠動物を用いて、胎児生存、胎児体重、外表・内臓の変化を評価する(ICH M3参照)。





Disclaimer: The definitions in this glossary are specific for their use within this guideline.
Alternative assay(s): In vitro, ex vivo or non-mammalian in vivo assay(s) intended to predict malformations or embryo-fetal lethality; see MEFL.
Applicability domain: refers to the definition of the physicochemical properties of the substances that can be reliably tested in the assay and the biological mechanisms of action covered by the assay.
Assay qualification (for regulatory use): Confirmation of the predictivity of an alternative assay(s) to identify MEFL, as observed in vivo.
Constitutive ingredients: Chemicals or biologic substances used as excipients, diluents, or adjuvants in a vaccine, including any diluent provided as an aid in the administration of the product and supplied separately.
Developmental toxicity: Any adverse effect induced prior to attainment of adult life. It includes effects induced or manifested from conception to postnatal life.
GD 0: The day on which positive evidence of mating is detected (e.g., sperm is found in the vaginal smear / vaginal plug in rodents, or observed mating in rabbits).
Malformation: Permanent structural deviation that generally is incompatible with or severely detrimental to normal development or survival.
Preliminary EFD (pEFD) toxicity study: An embryo-fetal developmental toxicity study that includes exposure over the period of organogenesis, has adequate dose levels, uses a minimum of 6 pregnant animals per group, and includes assessments of fetal survival, fetal weight, and external and soft tissue alterations (see ICH M3).
Surrogate molecule: A molecule showing similar pharmacologic activity in the test species as that shown by the human pharmaceutical in the human.
Vaccine: For the purpose of this guideline, this term refers to preventative or therapeutic vaccines for infectious diseases. Vaccine (inclusive of the term vaccine product) is defined as the complete formulation and includes antigen(s) (or immunogen(s)) and any additives such as adjuvants, excipients or preservatives. The vaccine is intended to stimulate the immune system and result in an immune response to the vaccine antigen(s). The primary pharmacological effect of the vaccine is the prevention and/or treatment of an infection or infectious disease.
Variation: Structural change that does not impact viability, development, or function (e.g., delays in ossification) which can be reversible, and are found in the normal population under investigation.