20060628 EMEA/CHMP/QWP/251344/2006
基因毒性杂质限度指南（中英文对照） London, 28 June 2006
CPMP/SWP/5199/02
EMEA/CHMP/QWP/251344/2006
1. INTRODUCTION 介绍............................................................................................................... 3
2. SCOPE 范围 ............................................................................................................................... 3
3. LEGAL BASIS法 律依据............................................................................................................ 3
4. TOXICOLOGICAL BACKGROUND 毒理学背景.................................................................... 4
5. RECOMMENDATIONS 建议..................................................................................................... 4
5.1 Genotoxic Compounds With Sufficient Evidence for a Threshold-Related Mechanism
具有充分证据证明其阈值相关机理的基因毒性化合物......................................................... 4
5.2 Genotoxic Compounds Without Sufficient Evidence for a Threshold-Related Mechanism
不具备充分证据支持其阈值相关机理的基因毒性化合物...................................................... 5
5.2.1 Pharmaceutical Assessment药学评价.................................................................................. 5
5.2.2 Toxicological Assessment毒理学评价................................................................................... 5
5.2.3 Application of a Threshold of Toxicological Concern 毒理学担忧阈值应用........................ 5
5.3 Decision Tree for Assessment of Acceptability of Genotoxic Impurities
基因毒性杂质可接受性评价决策树.......................................................................................... 7
REFERENCES. 参考文献................................................................................................................ 8
EXECUTIVE SUMMARY 内容摘要
The toxicological assessment of genotoxic impurities and the determination of acceptable
limits for such impurities in active substances is a difficult issue and not addressed in sufficient
detail in the existing ICH Q3X guidances. The data set usually available for genotoxic impurities is quite variable and is the main factor that dictates the process used for the assessment of acceptable
limits. In the absence of data usually needed for the application of one of the established risk
assessment methods, i.e. data from carcinogenicity long-term studies or data providing evidence
for a threshold mechanism of genotoxicity, implementation of a generally applicable approach as
defined by the Threshold of Toxicological Concern (TTC) is proposed. A TTC value of 1.5 μg/day
intake of a genotoxic impurity is considered to be associated with an acceptable risk (excess cancer
risk of
permitted level in the active substance can be calculated based on the expected daily dose. Higher
limits may be justified under certain conditions such as short-term exposure periods.
基因毒性杂质的毒理学评估和这些杂质在活性药物中的可接受标准的测定是一件困难的
事情，并且在现有的ICH Q3X指南中也没有详细的规定。现有的关于基因毒性杂质的相关
数据是容易变化的，也是对杂质可接受标准如何进行评价的主要影响因素。如果缺少风险评估方法所需要的数据，比如，致癌作用的长期研究数据，或为基因毒性的阀值提供证据的数据，一般建议使用一般通用的被定义为毒理学关注的阈值（TTC ）的方法。一个“1.5μg/day”的TTC 值，即相当于每天摄入1.5μg 的基因毒性杂质，被认为对于大多数药品来说是可以
接受的风险（一生中致癌的风险小于十万分之1）。按照这个阀值，可以根据这个预期的每日摄入量计算出活性药物中可接受的杂质水平。较高的临界值可以在特定的条件下，如短期暴露周期等，进行推算。
1. INTRODUCTION 介绍
A general concept of qualification of impurities is described in the guidelines for active
substances (Q3A, Impurities in New Active Substances) or medicinal products (Q3B, Impurities in New Medicinal Products), whereby qualification is defined as the process of acquiring and
evaluating data that establishes the biological safety of an individual impurity or a given impurity
profile at the level(s) specified. In the case of impurities with a genotoxic potential, determination
of acceptable dose levels is generally considered as a particularly critical issue, which is not
specifically covered by the existing guidelines.
在原料药（Q3A ）和药物制剂（Q3B ）的杂质指导原则中，杂质限度确定的依据包括各
个杂质的生物安全性数据或杂质在某特定含量水平的研究概况。而对于遗传毒性杂质限度的
确定，通常都认为是特别关键的问题，但目前尚无相关的指导原则。
2. SCOPE 范围
This Guideline describes a general framework and practical approaches on how to deal with
genotoxic impurities in new active substances. It also relates to new applications for existing active substances, where assessment of the route of synthesis, process control and impurity profile does
not provide reasonable assurance that no new or higher levels of genotoxic impurities are
introduced as compared to products currently authorised in the EU containing the same active
substance. The same also applies to variations to existing Marketing Authorisations pertaining to
the synthesis. The guideline does, however, not need to be applied retrospectively to authorised
products unless there is a specific cause for concern.
本指导原则阐述了如何处理新原料药中遗传毒性杂质的一般框架和实际方法。该指导原则也适用于已有原料药的新申请，如果其合成路线、过程控制和杂质研究尚无法确保不会产生新的或更高含量的遗传毒性杂质（与EU 目前批准的相同原料药相比）。该指导原则同样适用于已上市原料药有关合成方面的补充申请。除非有特殊原因，本指导原则不适用于已上市的产品。
In the current context the classification of a compound (impurity) as genotoxic in general means that there are positive findings in established in vitro or in vivo genotoxicity tests with the main focus on DNA reactive substances that have a potential for direct DNA damage. Isolated in vitro findings may be assessed for in vivo relevance in adequate follow-up testing. In the absence of such information in vitro genotoxicants are usually considered as presumptive in vivo mutagens and carcinogens.
目前对于基因毒性杂质的分类主要是指：在以DNA 反应物质为主要研究对象的体内体外试验中，如果发现它们对DNA 有潜在的破坏性，那可称之为基因毒性。如果有足够的后续试验，可由单独的体外试验结果，对它的体内关联性进行评估。在缺乏这样的信息时，体外基因毒性物质经常被考虑为假定的体内诱变剂和致癌剂。
3. LEGAL BASIS 法规依据
This guideline has to be read in conjunction with Directive 2001/83/EC (as amended) and all relevant CHMP Guidance documents with special emphasis on:
在阅读该指南时有必要参考“Directive 2001/83/EC”以及相关的CHMP 指南文件，特别是以下几个指南：
Impurities Testing Guideline: Impurities in New Drug Substances (CPMP/ICH/2737/99, ICHQ3A(R))
Note for Guidance on Impurities in New Drug Products (CPMP/ICH/2738/99, ICHQ3B (R)) Note for Guidance on Impurities: Residual Solvents (CPMP/ICH/283/95)
Note for Guidance on Genotoxicity: Guidance on Specific Aspects of Regulatory Genotoxicity Tests for Pharmaceuticals (CPMP/ICH/141/95, ICHS2A)
Note for Guidance on Genotoxicity: A Standard Battery for Genotoxicity Testing of
Pharmaceuticals (CPMP/ICH/174/95, ICHS2B)
4. TOXICOLOGICAL BACKGROUND 毒理学背景
According to current regulatory practice it is assumed that (in vivo) genotoxic compounds have the potential to damage DNA at any level of exposure and that such damage may
lead/contribute to tumour development. Thus for genotoxic carcinogens it is prudent to assume that there is no discernible threshold and that any level of exposure carries a risk.
根据目前的研究实践，具有（体内）遗传毒性的化合物在任何暴露量下都有可能对DNA 产生损伤，而这种损伤可能会引发肿瘤。因此，对于遗传毒性致癌物质，应谨慎认为不存在明确的阈值，任何暴露量下都存在风险。
However, the existence of mechanisms leading to biologically meaningful threshold effects is increasingly acknowledged also for genotoxic events. This holds true in particular for compounds interacting with non-DNA targets and also for potential mutagens, which are rapidly detoxified before coming into contact with critical targets. The regulatory approach to such chemicals can be based on the identification of a critical no-observed-effect level (NOEL) and use of uncertainty factors.
然而，对于一些遗传毒性事件，其产生生物学意义的阈值效应的机理正越来越为人所了解。对于非DNA 靶点的化合物和潜在致突变剂更是如此，因为它们在与关键靶点接触前就已经去毒化了。对于这些化合物，研究的基础可以是确定关键的未观察到影响的剂量（NOEL ）和采用不确定因子。
Even for compounds which are able to react with the DNA molecule, extrapolation in a linear manner from effects in high-dose studies to very low level (human) exposure may not be justified due to several protective mechanisms operating effectively at low doses. However, at present it is extremely difficult to experimentally prove the existence of threshold for the genotoxicity of a given mutagen. Thus, in the absence of appropriate evidence supporting the existence of a
threshold for a genotoxic compound making it difficult to define a safe dose it is necessary to adopt a concept of a level of exposure that carries an acceptable risk.
即使对能与DNA 分子发生反应的化合物，由于低剂量时有多种有效的保护机制存在，而不能将高剂量下的影响以线性方式外推到很低的（人）暴露水平。不过，目前要用实验方法证明某诱变剂的遗传毒性阈值仍然非常困难。所以，在缺乏恰当的证据支持遗传毒性阈值存在的情况下，确定安全剂量很困难，因此非常有必要采用一个可接受风险的暴露水平概念。
5. RECOMMENDATIONS 建议
As stated in the Q3A guideline, actual and potential impurities most likely to arise during synthesis, purification and storage of the new drug substance should be identified, based on a
sound scientific appraisal of the chemical reactions involved in the synthesis, impurities associated with raw materials that could contribute to the impurity profile of the new drug substance, and possible degradation products. This discussion can be limited to those impurities that might reasonably be expected based on knowledge of the chemical reactions and conditions involved. Guided by existing genotoxicity data or the presence of structural alerts, potential genotoxic impurities should be identified. When a potential impurity contains structural alerts, additional genotoxicity testing of the impurity, typically in a bacterial reverse mutation assay, should be considered (Dobo et al. 2006, Müller et al. 2006). While according to the Q3A guideline such studies can usually be conducted on the drug substance containing the impurity to be controlled, studies using isolated impurities are much more appropriate for this purpose and highly
recommended.
正如Q3A 指导原则所述，根据合理的化学反应机理分析，在新的原料药合成、纯化和贮存过程中很有可能产生实际的和潜在的杂质。依据现有的“可能引起遗传毒性的结构”数据库，潜在的遗传毒性杂质应能被确认。如果潜在的杂质含有可引起遗传毒性的结构单元，该杂质应考虑进行遗传毒性试验（一般是细菌回复突变试验）（Dobo 等，2006）。虽然Q3A 指导原则认为这些研究采用含有那些需控制杂质的原料药进行是可行的，但用分离出来的杂质进行这些研究更恰当，也是高度推荐的方法。
For determination of acceptable levels of exposure to genotoxic carcinogens considerations of possible mechanisms of action and of the dose-response relationship are important components. Based on the above considerations genotoxic impurities may be distinguished into the following two classes:
根据以上论述，遗传毒性杂质可以归纳成以下两类：
- Genotoxic compounds with sufficient (experimental) evidence for a threshold-related
mechanism
有充分阈值相关机理证据（实验）的遗传毒性化合物
- Genotoxic compounds without sufficient (experimental) evidence for a threshold-related mechanism
无充分阈值相关机理证据（实验）的遗传毒性化合物
5.1 Genotoxic Compounds With Sufficient Evidence for a Threshold-Related Mechanism 具有充分证据证明其阈值相关机理的基因毒性化合物
Examples of mechanisms of genotoxicity that may be demonstrated to lead to non-linear or thresholded dose-response relationships include interaction with the spindle apparatus of cell
division leading to aneuploidy, topoisomerase inhibition, inhibition of DNA synthesis, overloading of defence mechanisms, metabolic overload and physiological perturbations (e.g. induction of erythropoeisis, hyper- or hypothermia).
非线性或阈值明确的剂量效应关系的遗传毒性机理包括：与细胞分化过程中纺锤体相互作用；拓扑异构酶抑制；DNA 合成抑制；过度的防御机制；代谢过度和生理性干扰（如诱导红血球生成，高体温和低体温）。
For (classes of) compounds with clear evidence for a thresholded genotoxicity, exposure levels which are without appreciable risk of genotoxicity can be established according to the procedure as outlined for class 2 solvents in the Q3C Note for Guidance on Impurities: Residual Solvents. This approach calculates a “Permitted Daily Exposure” (PDE), which is derived from the NOEL, or the lowestobserved effect level (LOEL) in the most relevant (animal) study using “uncertainty factors” (UF).
有明确遗传毒性阈值的化合物，不产生遗传毒性风险的暴露水平可以被确定，方法可参照Q3C“杂质指导原则”中二类溶剂的限度确定方法。该方法可计算 “每日最大允许暴露量”（PDE ），数据来源于 “不确定因数”动物研究中的NOEL （未观察到效果的最低水平）或观察到效果的最低水平（LOEL ）。
5.2 Genotoxic Compounds Without Sufficient Evidence for a Threshold-Related
Mechanism 不具备充分证据支持其阈值相关机理的基因毒性化合物
The assessment of acceptability of genotoxic impurities for which no threshold mechanisms are identified should include both pharmaceutical and toxicological evaluations. In general, pharmaceutical measurements should be guided by a policy of controlling levels to “as low as reasonably practicable” (ALARP principle), where avoiding is not possible. Levels considered being consistent with the ALARP principle following pharmaceutical assessment should be assessed for acceptability from a toxicological point of view (see decision tree & following sections).
对于此类遗传毒性杂质，研究应包括药学和毒理学评估。总之，如果杂质无法避免，药
学方面的控制应遵循“合理可行的最低限量”原则（ALARP 原则）。符合ALARP 原则的杂质水平再经毒理学方面的进一步评估，以验证其合理性（见决策树和以下章节）。
5.2.1 Pharmaceutical Assessment 药学评价
A specific discussion – as part of the overall discussion on impurities (see Q3A(R)) – should be provided in the application with regard to impurities with potential genotoxicity.
申请材料应提供关于潜在遗传毒性杂质的特别讨论资料（见Q3A （R ））。
A rationale of the proposed formulation/manufacturing strategy should be provided based on available formulation options and technologies. The applicant should highlight, within the chemical process and impurity profile of active substance, all chemical substances, used as
reagents or present as intermediates, or side-products, known as genotoxic and/or carcinogenic (e.g. alkylating agents).
需要根据现在的配方选择和技术，提供证明所选的配方/生产策略合理性的证据。申请人应在合成工艺和杂质研究部分重点指出所有的化学物质，包括用到的试剂、中间体、副产物，哪些是已知遗传毒性和/或致癌性物质（如烷化剂）。
More generally, reacting substances and substances which show “alerting structure” in terms of genotoxicity which are not shared with the active substance should be considered (see e.g. Dobo et al. 2006). Potential alternatives which do not lead to genotoxic residues in the final product, should be used if available.
值得关注的是，虽然有些含有“可能引起遗传毒性的结构” （alerting structure）的反应试剂与最终活性物质并没有共同结构，但也要考虑它们的遗传毒性(see e.g. Dobo et al. 2006).。如果有可能，应该对它们进行一些替代研究，以使最终产品中不会引入基因毒性残留。
A justification needs to be provided that no viable alternative exists, including alternative routes of synthesis or formulations, different starting materials. This might for instance include cases where the structure, which is responsible for the genotoxic and/or carcinogenic potential is equivalent to that needed in chemical synthesis (e.g. alkylation reactions).
需要提供充分的论证来说明没有可行的替代方法存在，包括可替代的合成路线或配方，不同的起始物料等。比如，应证明具有遗传毒性和/或致癌性的结构在化学合成中（如烷化反应）是必需的。
If a genotoxic impurity is considered to be unavoidable in a drug substance, technical efforts
(e.g. purification steps) should be undertaken to reduce the content of the genotoxic residues in the final product in compliance with safety needs or to a level as low as reasonably practicable (see safety assessment). Data on chemical stability of reactive intermediates, reactants, and other components should be included in this assessment.
如果遗传毒性杂质在原料中不可避免，则应该采取适当的技术（如纯化步骤）降低该杂质的含量，以满足安全性要求，或符合“合理可行的最低限量”原则（见安全评估）。药学评估还应包括反应中间体、反应物和其它组件等的化学稳定性研究。
Detection and/or quantification of these residues should be done by state-of-the-art analytical techniques.
应该使用比较先进的分析检测技术来检测和量化这些残留的杂质。
5.2.2 Toxicological Assessment 毒理学评价
The impossibility of defining a safe exposure level (zero risk concept) for genotoxic
carcinogens without a threshold and the realization that complete elimination of genotoxic
impurities from drug substances is often unachievable, requires implementation of a concept of an acceptable risk level, i.e. an estimate of daily human exposure at and below which there is a negligible risk to human health.
鉴于在没有明确阈值的前提下定义安全暴露水平（零风险）是不可能的，且从原料药中完全除去遗传毒性杂质经常是很难做到的，所以有必要提出一个“可接受风险水平”
（acceptable risk level）的概念，比如估算一个“每日最大暴露量”值，低于该暴露量时就可以忽略其对人体健康的风险。
Procedures for the derivation of acceptable risk levels are considered in the Appendix 3 of the Q3C Note for Guidance on Impurities: Residual Solvents for Class 1 solvents. However, these approaches require availability of adequate data from long-term carcinogenicity studies.
对于可接受风险水平的推导过程请参见Q3C （杂质指南注释: 一类溶液残留）中的附件
三。然而，应用这些方法必须有足够多的长期致癌性研究数据。
In most cases of toxicological assessment of genotoxic impurities only limited data from in vitro studies with the impurity (e.g. Ames test, chromosomal aberration test) are available and thus established approaches to determine acceptable intake levels cannot be applied. Calculation of “safety multiples” from in vitro data (e.g. Ames test) are considered inappropriate for justification of acceptable limits. Moreover, negative carcinogenicity and genotoxicity data with the drug
substance containing the impurity at low ppm levels do not provide sufficient assurance for setting acceptable limits for the impurity due to the lack of sensitivity of this testing approach. Even potent mutagens and carcinogens are most likely to remain undetected when tested as part of the drug substance, i.e. at very low exposure levels. A pragmatic approach is therefore needed which recognises that the presence of very low levels of genotoxic impurities is not associated with an unacceptable risk.
大多数情况下，遗传毒性杂质的毒理学评估只是局限于杂质的体外研究（如Ames 试验，染色体畸变试验），但这些方法并不适用于确定杂质可接受的摄入水平。也就是说，根据体外数据（如Ames 试验）计算杂质的“安全倍数（safety multiples）”、进而确定可接受的限度，是不合适的。此外，用含有较低（ppm 级）杂质水平的原料药研究其致癌性和遗传毒性，即使得出阴性结果也不足以确保该杂质限度的合理性，因为这种试验方法缺少必要的灵敏度。有些具有很强致突变性和致癌性物质与原料药一起进行试验时，因为在非常低的暴露水平情况下，很有可能因为低于检测限而无法检出。所以，如果认识到含量非常低的遗传毒性杂质不存在“不可接受的风险”（unacceptable risk），那么可以采取实用的方法来控制该杂质。
5.2.3 Application of a Threshold of Toxicological Concern 毒理学相关的阈值应用
A threshold of toxicological concern (TTC) has been developed to define a common exposure level for any unstudied chemical that will not pose a risk of significant carcinogenicity or other toxic effects (Munro et al. 1999, Kroes and Kozianowski 2002). This TTC value was estimated to be 1.5 μg/person/day. The TTC, originally developed as a “threshold of regulation” at the FDA for food contact materials (Rulis 1989, FDA 1995) was established based on the analysis of 343 carcinogens from a carcinogenic potency database (Gold et al. 1984) and was repeatedly confirmed by evaluations expanding the database to more than 700 carcinogens (Munro 1990, Cheeseman et al. 1999, Kroes et al. 2004). The probability distribution of carcinogenic potencies has been used to derive an estimate of a daily exposure level (μg/person) of most carcinogens which would give rise to less than a one in a million (1 x 10-6) upper bound lifetime risk of cancer (“virtually safe dose”). Further analysis of subsets of high potency carcinogens led to the suggestion of a 10-fold lower TTC (0.15 μg/day) for chemicals with structural alerts that raise concern for potential genotoxicity (Kroes et al. 2004).
“毒理学关注的阈值”用于定义那些不会产生显著致癌性或其他毒性作用、但又未明确研究的化合物的“常见暴露量”（common exposure level）(Munro et al. 1999, Kroes and
Kozianowski 2002)。该TTC 估计值是1.5μg/人/日。TTC 概念最早来源于FDA 关于食品接触
材料的“规定阈值”（a threshold of regulation）（Rulis 1989, FDA 1995），该阈值根据对致癌能力数据库(Gold et al. 1984)中343种致癌物质的分析结果得出。随后该数据库扩大到700多个致癌性物质(Munro 1990, Cheeseman et al. 1999, Kroes et al. 2004)，这种分析结果不断得到重复验证。通过对致癌能力的概率分布进行评价，可以得到一个对大多数致癌物质适用的“日常摄入水平(μg/person)”，此水平造成的一生中患癌症的风险小于正常风险水平的上限1 x 10-6（真实的安全剂量）。对于含有“可能引起遗传毒性结构” 的化合物，其TTC 应严格10倍（0.15μg/日）（Kroes et al. 2004）。
However, for application of a TTC in the assessment of acceptable limits of genotoxic
impurities in drug substances a value of 1.5 μg/day, corresponding to a 10-5 lifetime risk of cancer can be justified as for pharmaceuticals a benefit exists. It should be recognized in this context that the methods on which the TTC value is based, are generally considered very conservative since they involved a simple linear extrapolation from the dose giving a 50% tumour incidence (TD50) to a 1 in 106 incidence, using TD50 data for the most sensitive species and most sensitive site (several “worst case” assumptions) (Munro et al. 1999).
然而，用TTC 评估原料药中的遗传毒性杂质限度，1.5μg/日（相当于10万分之一的患癌风险）是可以接受的。应该承认，基于TTC 值控制遗传毒性杂质是非常保守的，因为这只是根据从产生50%肿瘤发生率（TD50）到百万分之一致癌率的剂量线性推导得到的，而且TD50数据是用最敏感的动物和最敏感的部位研究得到的（几个“最坏条件”假设）（Munro et al. 1999）。
Some structural groups were identified to be of such high potency that intakes even below the TTC would be associated with a high probability of a significant carcinogenic risk (Cheeseman et al. 1999, Kroes et al. 2004). This group of high potency genotoxic carcinogens comprises
aflatoxin-like-, nitroso-, and azoxy-compounds that have to be excluded from the TTC approach. Risk assessment of members of such groups requires compound-specific toxicity data.
有几个结构基团被认定为具有非常高的基因毒性，它们即使被摄入低于TTC 值的量也会面临非常高的基因毒性风险(Cheeseman et al. 1999, Kroes et al. 2004)。这些高致癌性物质包括黄曲霉素类、N-亚硝基物和偶氮类化合物，不适用TTC 方法。这类化合物的风险评估需采用专门的毒性数据。
There may be reasons to deviate from the TTC value based on the profile of genotoxicity results.
根据基因杂质概况，有些情况下会偏离TTC 值。
Positive result from in vitro studies only may allow to exempt an impurity from limitation at TTC level if lack of in vivo relevance of the findings is convincingly demonstrated based on a weight-ofevidence approach (see ICH S2 guidelines). This approach will usually need negative results with the impurity from some additional in vitro and/or appropriate in vivo testing.
假如按照证据权衡法能充分证明“结果缺乏体内相关性”，体外试验的阳性结果也仅能在TTC 水平上排除一个杂质(参见 ICH 指南S2) 。这种方法经常需要在额外的体外试验和/或合理的体内试验，并且得到杂质的阴性结果。
A TTC value higher than 1.5 μg/day may be acceptable under certain conditions, e.g.
short-term exposure, for treatment of a life-threatening condition, when life expectancy is less than 5 years, or where the impurity is a known substance and human exposure will be much greater from other sources (e.g. food). Genotoxic impurities that are also significant metabolites may be assessed based on the acceptability of the metabolites.
某些情况下TTC 值高于1.5μg/日也是可以接受的，如短期用药；用于治疗威胁生命疾病的药物；或人的存活期少于5年；或该杂质是已知物质，人体从其他途经（如食物）获得的暴露量远远高于药物途经。如果遗传毒性杂质本身就是重要的代谢物，那么该杂质可以根据代谢物的可接受限度进行控制。
The concentration limits in ppm of genotoxic impurity in drug substance derived from the TTC can be calculated based on the expected daily dose to the patient using equation (1).
采用下列公式，从TTC 值和日服用剂量，可以计算出原料药中的基因毒性杂质的浓度限度。
(1) Concentration limit (ppm) = TTC [μg/day]/dose (g/day]
浓度限度（ppm ） = TTC [μg/day]/剂量(g/day]
The TTC concept should not be applied to carcinogens where adequate toxicity data
(long-term studies) are available and allow for a compound-specific risk assessment.
对于有确切毒性数据（长期毒性研究）的致癌性物质不宜使用TTC 概念，应进行特定化合物风险评估。
It has to be emphasized that the TTC is a pragmatic risk management tool using a probabilistic methodology, i.e. there is a high probability that a 10-5 lifetime cancer risk will not be exceeded if the daily intake of a genotoxic impurity with unknown carcinogenic potential/potency is below the
TTC value. The TTC concept should not be interpreted as providing absolute certainty of no risk.
应强调，TTC 是一个实用性的风险管理方法，是按概率方法学估算的。比如按这一概念，如果某未知致癌性遗传毒性杂质的摄入量低于TTC 值，那么就可以保证患癌风险控制在十万分之一之内。但TTC 概念不能被理解为确保绝对无风险。
5.3 Decision Tree for Assessment of Acceptability of Genotoxic Impurities 基因毒性可接受性评价决策树
(shaded boxes = pharmaceutical assessment, white boxes = toxicological assessment)
（阴影框 = 药学评价，白框 = 毒理学评价）
1) Impurities with structural relationship to high potency carcinogens (see text) are to be excluded from the TTC approach
1) 结构上与高致癌性物质有关的杂质（见正文）不能采用TTC 法。
2) If carcinogenicity data available: Does intake exceed calculated 10-5 cancer lifetime risk?
2) 如果有致癌性数据：摄入量超过10-5患癌风险吗？
3) Case-by-case assessment should include duration of treatment, indication, patient
population etc (see text)
3) 具体情况具体分析，包括用药时间长短、适应症、患者人群等（见正文）。
*) Abbreviations: 缩写
NOEL/UF – No Observed Effect Level/Uncertainty Factor, 未观察到效果水平/不确定因素 PDE – Permitted Daily Exposure, 允许日接触量
TTC – Threshold of Toxicological Concern 毒理学关注的域值
REFERENCES 参考文献
Cheeseman M.A., Machuga E.J., Bailey A.B., A tiered approach to threshold of regulation, Food Chem Toxicol 37, 387-412, 1999
食品化学毒性，1999，
Dobo K.L., Greene N., Cyr M.O., Caron S., Ku W.W., The application of structure-based assessment to support safety and chemistry diligence to manage genotoxic impurities in active pharmaceutical ingredients during drug development, Reg Tox Pharm 44, 282-293, 2006.
Gold L.S., Sawyer C.B., Magaw R., Backman G.M., de Veciana M., Levinson R., Hooper N.K., Havender W.R., Bernstein L., Peto R., Pike M.C., Ames B.N., A carcinogenic potency database of the standardized results of animal bioassays, Environ Health Perspect 58, 9-319, 1984.
Kroes R., Renwick A.G., Cheeseman M., Kleiner J., Mangelsdorf I., Piersma A., Schilter B., Schlatter J., van Schothorst F., Vos J.G., Würtzen G., Structure-based threshold of toxicological concern (TTC): guidance for application to substances present at low levels in the diet, Food Chem Toxicol 42, 65-83, 2004.
Kroes R., Kozianowski G., Threshold of toxicological concern (TTC) in food safety
assessment, Toxicol Letters 127, 43-46, 2002.
Müller L., Mauthe R.J., Riley C.M., Andino M.M., De Antonis D., Beels C., DeGeorge J., De Knaep A.G.M., Ellison D., Fagerland J.A., Frank R., Fritschel B., Galloway S., Harpur E.,
Humfrey C.D.N., Jacks A.S.J., Jagota N., Mackinnon J., Mohan G., Ness D.K., O’Donovan M.R., Smith M.D., Vudathala G., Yotti L., A rationale for determining, testing, and controlling specific impurities in pharmaceuticals that possess potential for genotoxicity, Reg Tox Pharm 44, 198-211, 2006
一种确定、检测和控制药品中可能具有基因毒性的特定杂质的理论
Munro I.C., Safety assessment procedures for indirect food additives: an overview. Report of a workshop. Reg Tox Pharm 12, 2-12, 1990.
Munro I.C., Kennepohl E., Kroes R., A procedure for the safety evaluation of flavouring substances, Food Chem Toxicol 37, 207-232, 1999. Rulis A.M., Establishing a threshold of regulation. In Risk Assessment in Setting National Priorities (J.J. Bonin and D.E. Stevenson, Eds.) Plenum, New York, 271-278, 1989.
U.S. Food and Drug Administration (FDA), Food additives: Threshold of regulation for substances used in food-contact articles (final rule), Fed. Regist. 60, 36582-36596, 1995.
20060628 EMEA/CHMP/QWP/251344/2006
基因毒性杂质限度指南（中英文对照） London, 28 June 2006
CPMP/SWP/5199/02
EMEA/CHMP/QWP/251344/2006
1. INTRODUCTION 介绍............................................................................................................... 3
2. SCOPE 范围 ............................................................................................................................... 3
3. LEGAL BASIS法 律依据............................................................................................................ 3
4. TOXICOLOGICAL BACKGROUND 毒理学背景.................................................................... 4
5. RECOMMENDATIONS 建议..................................................................................................... 4
5.1 Genotoxic Compounds With Sufficient Evidence for a Threshold-Related Mechanism
具有充分证据证明其阈值相关机理的基因毒性化合物......................................................... 4
5.2 Genotoxic Compounds Without Sufficient Evidence for a Threshold-Related Mechanism
不具备充分证据支持其阈值相关机理的基因毒性化合物...................................................... 5
5.2.1 Pharmaceutical Assessment药学评价.................................................................................. 5
5.2.2 Toxicological Assessment毒理学评价................................................................................... 5
5.2.3 Application of a Threshold of Toxicological Concern 毒理学担忧阈值应用........................ 5
5.3 Decision Tree for Assessment of Acceptability of Genotoxic Impurities
基因毒性杂质可接受性评价决策树.......................................................................................... 7
REFERENCES. 参考文献................................................................................................................ 8
EXECUTIVE SUMMARY 内容摘要
The toxicological assessment of genotoxic impurities and the determination of acceptable
limits for such impurities in active substances is a difficult issue and not addressed in sufficient
detail in the existing ICH Q3X guidances. The data set usually available for genotoxic impurities is quite variable and is the main factor that dictates the process used for the assessment of acceptable
limits. In the absence of data usually needed for the application of one of the established risk
assessment methods, i.e. data from carcinogenicity long-term studies or data providing evidence
for a threshold mechanism of genotoxicity, implementation of a generally applicable approach as
defined by the Threshold of Toxicological Concern (TTC) is proposed. A TTC value of 1.5 μg/day
intake of a genotoxic impurity is considered to be associated with an acceptable risk (excess cancer
risk of
permitted level in the active substance can be calculated based on the expected daily dose. Higher
limits may be justified under certain conditions such as short-term exposure periods.
基因毒性杂质的毒理学评估和这些杂质在活性药物中的可接受标准的测定是一件困难的
事情，并且在现有的ICH Q3X指南中也没有详细的规定。现有的关于基因毒性杂质的相关
数据是容易变化的，也是对杂质可接受标准如何进行评价的主要影响因素。如果缺少风险评估方法所需要的数据，比如，致癌作用的长期研究数据，或为基因毒性的阀值提供证据的数据，一般建议使用一般通用的被定义为毒理学关注的阈值（TTC ）的方法。一个“1.5μg/day”的TTC 值，即相当于每天摄入1.5μg 的基因毒性杂质，被认为对于大多数药品来说是可以
接受的风险（一生中致癌的风险小于十万分之1）。按照这个阀值，可以根据这个预期的每日摄入量计算出活性药物中可接受的杂质水平。较高的临界值可以在特定的条件下，如短期暴露周期等，进行推算。
1. INTRODUCTION 介绍
A general concept of qualification of impurities is described in the guidelines for active
substances (Q3A, Impurities in New Active Substances) or medicinal products (Q3B, Impurities in New Medicinal Products), whereby qualification is defined as the process of acquiring and
evaluating data that establishes the biological safety of an individual impurity or a given impurity
profile at the level(s) specified. In the case of impurities with a genotoxic potential, determination
of acceptable dose levels is generally considered as a particularly critical issue, which is not
specifically covered by the existing guidelines.
在原料药（Q3A ）和药物制剂（Q3B ）的杂质指导原则中，杂质限度确定的依据包括各
个杂质的生物安全性数据或杂质在某特定含量水平的研究概况。而对于遗传毒性杂质限度的
确定，通常都认为是特别关键的问题，但目前尚无相关的指导原则。
2. SCOPE 范围
This Guideline describes a general framework and practical approaches on how to deal with
genotoxic impurities in new active substances. It also relates to new applications for existing active substances, where assessment of the route of synthesis, process control and impurity profile does
not provide reasonable assurance that no new or higher levels of genotoxic impurities are
introduced as compared to products currently authorised in the EU containing the same active
substance. The same also applies to variations to existing Marketing Authorisations pertaining to
the synthesis. The guideline does, however, not need to be applied retrospectively to authorised
products unless there is a specific cause for concern.
本指导原则阐述了如何处理新原料药中遗传毒性杂质的一般框架和实际方法。该指导原则也适用于已有原料药的新申请，如果其合成路线、过程控制和杂质研究尚无法确保不会产生新的或更高含量的遗传毒性杂质（与EU 目前批准的相同原料药相比）。该指导原则同样适用于已上市原料药有关合成方面的补充申请。除非有特殊原因，本指导原则不适用于已上市的产品。
In the current context the classification of a compound (impurity) as genotoxic in general means that there are positive findings in established in vitro or in vivo genotoxicity tests with the main focus on DNA reactive substances that have a potential for direct DNA damage. Isolated in vitro findings may be assessed for in vivo relevance in adequate follow-up testing. In the absence of such information in vitro genotoxicants are usually considered as presumptive in vivo mutagens and carcinogens.
目前对于基因毒性杂质的分类主要是指：在以DNA 反应物质为主要研究对象的体内体外试验中，如果发现它们对DNA 有潜在的破坏性，那可称之为基因毒性。如果有足够的后续试验，可由单独的体外试验结果，对它的体内关联性进行评估。在缺乏这样的信息时，体外基因毒性物质经常被考虑为假定的体内诱变剂和致癌剂。
3. LEGAL BASIS 法规依据
This guideline has to be read in conjunction with Directive 2001/83/EC (as amended) and all relevant CHMP Guidance documents with special emphasis on:
在阅读该指南时有必要参考“Directive 2001/83/EC”以及相关的CHMP 指南文件，特别是以下几个指南：
Impurities Testing Guideline: Impurities in New Drug Substances (CPMP/ICH/2737/99, ICHQ3A(R))
Note for Guidance on Impurities in New Drug Products (CPMP/ICH/2738/99, ICHQ3B (R)) Note for Guidance on Impurities: Residual Solvents (CPMP/ICH/283/95)
Note for Guidance on Genotoxicity: Guidance on Specific Aspects of Regulatory Genotoxicity Tests for Pharmaceuticals (CPMP/ICH/141/95, ICHS2A)
Note for Guidance on Genotoxicity: A Standard Battery for Genotoxicity Testing of
Pharmaceuticals (CPMP/ICH/174/95, ICHS2B)
4. TOXICOLOGICAL BACKGROUND 毒理学背景
According to current regulatory practice it is assumed that (in vivo) genotoxic compounds have the potential to damage DNA at any level of exposure and that such damage may
lead/contribute to tumour development. Thus for genotoxic carcinogens it is prudent to assume that there is no discernible threshold and that any level of exposure carries a risk.
根据目前的研究实践，具有（体内）遗传毒性的化合物在任何暴露量下都有可能对DNA 产生损伤，而这种损伤可能会引发肿瘤。因此，对于遗传毒性致癌物质，应谨慎认为不存在明确的阈值，任何暴露量下都存在风险。
However, the existence of mechanisms leading to biologically meaningful threshold effects is increasingly acknowledged also for genotoxic events. This holds true in particular for compounds interacting with non-DNA targets and also for potential mutagens, which are rapidly detoxified before coming into contact with critical targets. The regulatory approach to such chemicals can be based on the identification of a critical no-observed-effect level (NOEL) and use of uncertainty factors.
然而，对于一些遗传毒性事件，其产生生物学意义的阈值效应的机理正越来越为人所了解。对于非DNA 靶点的化合物和潜在致突变剂更是如此，因为它们在与关键靶点接触前就已经去毒化了。对于这些化合物，研究的基础可以是确定关键的未观察到影响的剂量（NOEL ）和采用不确定因子。
Even for compounds which are able to react with the DNA molecule, extrapolation in a linear manner from effects in high-dose studies to very low level (human) exposure may not be justified due to several protective mechanisms operating effectively at low doses. However, at present it is extremely difficult to experimentally prove the existence of threshold for the genotoxicity of a given mutagen. Thus, in the absence of appropriate evidence supporting the existence of a
threshold for a genotoxic compound making it difficult to define a safe dose it is necessary to adopt a concept of a level of exposure that carries an acceptable risk.
即使对能与DNA 分子发生反应的化合物，由于低剂量时有多种有效的保护机制存在，而不能将高剂量下的影响以线性方式外推到很低的（人）暴露水平。不过，目前要用实验方法证明某诱变剂的遗传毒性阈值仍然非常困难。所以，在缺乏恰当的证据支持遗传毒性阈值存在的情况下，确定安全剂量很困难，因此非常有必要采用一个可接受风险的暴露水平概念。
5. RECOMMENDATIONS 建议
As stated in the Q3A guideline, actual and potential impurities most likely to arise during synthesis, purification and storage of the new drug substance should be identified, based on a
sound scientific appraisal of the chemical reactions involved in the synthesis, impurities associated with raw materials that could contribute to the impurity profile of the new drug substance, and possible degradation products. This discussion can be limited to those impurities that might reasonably be expected based on knowledge of the chemical reactions and conditions involved. Guided by existing genotoxicity data or the presence of structural alerts, potential genotoxic impurities should be identified. When a potential impurity contains structural alerts, additional genotoxicity testing of the impurity, typically in a bacterial reverse mutation assay, should be considered (Dobo et al. 2006, Müller et al. 2006). While according to the Q3A guideline such studies can usually be conducted on the drug substance containing the impurity to be controlled, studies using isolated impurities are much more appropriate for this purpose and highly
recommended.
正如Q3A 指导原则所述，根据合理的化学反应机理分析，在新的原料药合成、纯化和贮存过程中很有可能产生实际的和潜在的杂质。依据现有的“可能引起遗传毒性的结构”数据库，潜在的遗传毒性杂质应能被确认。如果潜在的杂质含有可引起遗传毒性的结构单元，该杂质应考虑进行遗传毒性试验（一般是细菌回复突变试验）（Dobo 等，2006）。虽然Q3A 指导原则认为这些研究采用含有那些需控制杂质的原料药进行是可行的，但用分离出来的杂质进行这些研究更恰当，也是高度推荐的方法。
For determination of acceptable levels of exposure to genotoxic carcinogens considerations of possible mechanisms of action and of the dose-response relationship are important components. Based on the above considerations genotoxic impurities may be distinguished into the following two classes:
根据以上论述，遗传毒性杂质可以归纳成以下两类：
- Genotoxic compounds with sufficient (experimental) evidence for a threshold-related
mechanism
有充分阈值相关机理证据（实验）的遗传毒性化合物
- Genotoxic compounds without sufficient (experimental) evidence for a threshold-related mechanism
无充分阈值相关机理证据（实验）的遗传毒性化合物
5.1 Genotoxic Compounds With Sufficient Evidence for a Threshold-Related Mechanism 具有充分证据证明其阈值相关机理的基因毒性化合物
Examples of mechanisms of genotoxicity that may be demonstrated to lead to non-linear or thresholded dose-response relationships include interaction with the spindle apparatus of cell
division leading to aneuploidy, topoisomerase inhibition, inhibition of DNA synthesis, overloading of defence mechanisms, metabolic overload and physiological perturbations (e.g. induction of erythropoeisis, hyper- or hypothermia).
非线性或阈值明确的剂量效应关系的遗传毒性机理包括：与细胞分化过程中纺锤体相互作用；拓扑异构酶抑制；DNA 合成抑制；过度的防御机制；代谢过度和生理性干扰（如诱导红血球生成，高体温和低体温）。
For (classes of) compounds with clear evidence for a thresholded genotoxicity, exposure levels which are without appreciable risk of genotoxicity can be established according to the procedure as outlined for class 2 solvents in the Q3C Note for Guidance on Impurities: Residual Solvents. This approach calculates a “Permitted Daily Exposure” (PDE), which is derived from the NOEL, or the lowestobserved effect level (LOEL) in the most relevant (animal) study using “uncertainty factors” (UF).
有明确遗传毒性阈值的化合物，不产生遗传毒性风险的暴露水平可以被确定，方法可参照Q3C“杂质指导原则”中二类溶剂的限度确定方法。该方法可计算 “每日最大允许暴露量”（PDE ），数据来源于 “不确定因数”动物研究中的NOEL （未观察到效果的最低水平）或观察到效果的最低水平（LOEL ）。
5.2 Genotoxic Compounds Without Sufficient Evidence for a Threshold-Related
Mechanism 不具备充分证据支持其阈值相关机理的基因毒性化合物
The assessment of acceptability of genotoxic impurities for which no threshold mechanisms are identified should include both pharmaceutical and toxicological evaluations. In general, pharmaceutical measurements should be guided by a policy of controlling levels to “as low as reasonably practicable” (ALARP principle), where avoiding is not possible. Levels considered being consistent with the ALARP principle following pharmaceutical assessment should be assessed for acceptability from a toxicological point of view (see decision tree & following sections).
对于此类遗传毒性杂质，研究应包括药学和毒理学评估。总之，如果杂质无法避免，药
学方面的控制应遵循“合理可行的最低限量”原则（ALARP 原则）。符合ALARP 原则的杂质水平再经毒理学方面的进一步评估，以验证其合理性（见决策树和以下章节）。
5.2.1 Pharmaceutical Assessment 药学评价
A specific discussion – as part of the overall discussion on impurities (see Q3A(R)) – should be provided in the application with regard to impurities with potential genotoxicity.
申请材料应提供关于潜在遗传毒性杂质的特别讨论资料（见Q3A （R ））。
A rationale of the proposed formulation/manufacturing strategy should be provided based on available formulation options and technologies. The applicant should highlight, within the chemical process and impurity profile of active substance, all chemical substances, used as
reagents or present as intermediates, or side-products, known as genotoxic and/or carcinogenic (e.g. alkylating agents).
需要根据现在的配方选择和技术，提供证明所选的配方/生产策略合理性的证据。申请人应在合成工艺和杂质研究部分重点指出所有的化学物质，包括用到的试剂、中间体、副产物，哪些是已知遗传毒性和/或致癌性物质（如烷化剂）。
More generally, reacting substances and substances which show “alerting structure” in terms of genotoxicity which are not shared with the active substance should be considered (see e.g. Dobo et al. 2006). Potential alternatives which do not lead to genotoxic residues in the final product, should be used if available.
值得关注的是，虽然有些含有“可能引起遗传毒性的结构” （alerting structure）的反应试剂与最终活性物质并没有共同结构，但也要考虑它们的遗传毒性(see e.g. Dobo et al. 2006).。如果有可能，应该对它们进行一些替代研究，以使最终产品中不会引入基因毒性残留。
A justification needs to be provided that no viable alternative exists, including alternative routes of synthesis or formulations, different starting materials. This might for instance include cases where the structure, which is responsible for the genotoxic and/or carcinogenic potential is equivalent to that needed in chemical synthesis (e.g. alkylation reactions).
需要提供充分的论证来说明没有可行的替代方法存在，包括可替代的合成路线或配方，不同的起始物料等。比如，应证明具有遗传毒性和/或致癌性的结构在化学合成中（如烷化反应）是必需的。
If a genotoxic impurity is considered to be unavoidable in a drug substance, technical efforts
(e.g. purification steps) should be undertaken to reduce the content of the genotoxic residues in the final product in compliance with safety needs or to a level as low as reasonably practicable (see safety assessment). Data on chemical stability of reactive intermediates, reactants, and other components should be included in this assessment.
如果遗传毒性杂质在原料中不可避免，则应该采取适当的技术（如纯化步骤）降低该杂质的含量，以满足安全性要求，或符合“合理可行的最低限量”原则（见安全评估）。药学评估还应包括反应中间体、反应物和其它组件等的化学稳定性研究。
Detection and/or quantification of these residues should be done by state-of-the-art analytical techniques.
应该使用比较先进的分析检测技术来检测和量化这些残留的杂质。
5.2.2 Toxicological Assessment 毒理学评价
The impossibility of defining a safe exposure level (zero risk concept) for genotoxic
carcinogens without a threshold and the realization that complete elimination of genotoxic
impurities from drug substances is often unachievable, requires implementation of a concept of an acceptable risk level, i.e. an estimate of daily human exposure at and below which there is a negligible risk to human health.
鉴于在没有明确阈值的前提下定义安全暴露水平（零风险）是不可能的，且从原料药中完全除去遗传毒性杂质经常是很难做到的，所以有必要提出一个“可接受风险水平”
（acceptable risk level）的概念，比如估算一个“每日最大暴露量”值，低于该暴露量时就可以忽略其对人体健康的风险。
Procedures for the derivation of acceptable risk levels are considered in the Appendix 3 of the Q3C Note for Guidance on Impurities: Residual Solvents for Class 1 solvents. However, these approaches require availability of adequate data from long-term carcinogenicity studies.
对于可接受风险水平的推导过程请参见Q3C （杂质指南注释: 一类溶液残留）中的附件
三。然而，应用这些方法必须有足够多的长期致癌性研究数据。
In most cases of toxicological assessment of genotoxic impurities only limited data from in vitro studies with the impurity (e.g. Ames test, chromosomal aberration test) are available and thus established approaches to determine acceptable intake levels cannot be applied. Calculation of “safety multiples” from in vitro data (e.g. Ames test) are considered inappropriate for justification of acceptable limits. Moreover, negative carcinogenicity and genotoxicity data with the drug
substance containing the impurity at low ppm levels do not provide sufficient assurance for setting acceptable limits for the impurity due to the lack of sensitivity of this testing approach. Even potent mutagens and carcinogens are most likely to remain undetected when tested as part of the drug substance, i.e. at very low exposure levels. A pragmatic approach is therefore needed which recognises that the presence of very low levels of genotoxic impurities is not associated with an unacceptable risk.
大多数情况下，遗传毒性杂质的毒理学评估只是局限于杂质的体外研究（如Ames 试验，染色体畸变试验），但这些方法并不适用于确定杂质可接受的摄入水平。也就是说，根据体外数据（如Ames 试验）计算杂质的“安全倍数（safety multiples）”、进而确定可接受的限度，是不合适的。此外，用含有较低（ppm 级）杂质水平的原料药研究其致癌性和遗传毒性，即使得出阴性结果也不足以确保该杂质限度的合理性，因为这种试验方法缺少必要的灵敏度。有些具有很强致突变性和致癌性物质与原料药一起进行试验时，因为在非常低的暴露水平情况下，很有可能因为低于检测限而无法检出。所以，如果认识到含量非常低的遗传毒性杂质不存在“不可接受的风险”（unacceptable risk），那么可以采取实用的方法来控制该杂质。
5.2.3 Application of a Threshold of Toxicological Concern 毒理学相关的阈值应用
A threshold of toxicological concern (TTC) has been developed to define a common exposure level for any unstudied chemical that will not pose a risk of significant carcinogenicity or other toxic effects (Munro et al. 1999, Kroes and Kozianowski 2002). This TTC value was estimated to be 1.5 μg/person/day. The TTC, originally developed as a “threshold of regulation” at the FDA for food contact materials (Rulis 1989, FDA 1995) was established based on the analysis of 343 carcinogens from a carcinogenic potency database (Gold et al. 1984) and was repeatedly confirmed by evaluations expanding the database to more than 700 carcinogens (Munro 1990, Cheeseman et al. 1999, Kroes et al. 2004). The probability distribution of carcinogenic potencies has been used to derive an estimate of a daily exposure level (μg/person) of most carcinogens which would give rise to less than a one in a million (1 x 10-6) upper bound lifetime risk of cancer (“virtually safe dose”). Further analysis of subsets of high potency carcinogens led to the suggestion of a 10-fold lower TTC (0.15 μg/day) for chemicals with structural alerts that raise concern for potential genotoxicity (Kroes et al. 2004).
“毒理学关注的阈值”用于定义那些不会产生显著致癌性或其他毒性作用、但又未明确研究的化合物的“常见暴露量”（common exposure level）(Munro et al. 1999, Kroes and
Kozianowski 2002)。该TTC 估计值是1.5μg/人/日。TTC 概念最早来源于FDA 关于食品接触
材料的“规定阈值”（a threshold of regulation）（Rulis 1989, FDA 1995），该阈值根据对致癌能力数据库(Gold et al. 1984)中343种致癌物质的分析结果得出。随后该数据库扩大到700多个致癌性物质(Munro 1990, Cheeseman et al. 1999, Kroes et al. 2004)，这种分析结果不断得到重复验证。通过对致癌能力的概率分布进行评价，可以得到一个对大多数致癌物质适用的“日常摄入水平(μg/person)”，此水平造成的一生中患癌症的风险小于正常风险水平的上限1 x 10-6（真实的安全剂量）。对于含有“可能引起遗传毒性结构” 的化合物，其TTC 应严格10倍（0.15μg/日）（Kroes et al. 2004）。
However, for application of a TTC in the assessment of acceptable limits of genotoxic
impurities in drug substances a value of 1.5 μg/day, corresponding to a 10-5 lifetime risk of cancer can be justified as for pharmaceuticals a benefit exists. It should be recognized in this context that the methods on which the TTC value is based, are generally considered very conservative since they involved a simple linear extrapolation from the dose giving a 50% tumour incidence (TD50) to a 1 in 106 incidence, using TD50 data for the most sensitive species and most sensitive site (several “worst case” assumptions) (Munro et al. 1999).
然而，用TTC 评估原料药中的遗传毒性杂质限度，1.5μg/日（相当于10万分之一的患癌风险）是可以接受的。应该承认，基于TTC 值控制遗传毒性杂质是非常保守的，因为这只是根据从产生50%肿瘤发生率（TD50）到百万分之一致癌率的剂量线性推导得到的，而且TD50数据是用最敏感的动物和最敏感的部位研究得到的（几个“最坏条件”假设）（Munro et al. 1999）。
Some structural groups were identified to be of such high potency that intakes even below the TTC would be associated with a high probability of a significant carcinogenic risk (Cheeseman et al. 1999, Kroes et al. 2004). This group of high potency genotoxic carcinogens comprises
aflatoxin-like-, nitroso-, and azoxy-compounds that have to be excluded from the TTC approach. Risk assessment of members of such groups requires compound-specific toxicity data.
有几个结构基团被认定为具有非常高的基因毒性，它们即使被摄入低于TTC 值的量也会面临非常高的基因毒性风险(Cheeseman et al. 1999, Kroes et al. 2004)。这些高致癌性物质包括黄曲霉素类、N-亚硝基物和偶氮类化合物，不适用TTC 方法。这类化合物的风险评估需采用专门的毒性数据。
There may be reasons to deviate from the TTC value based on the profile of genotoxicity results.
根据基因杂质概况，有些情况下会偏离TTC 值。
Positive result from in vitro studies only may allow to exempt an impurity from limitation at TTC level if lack of in vivo relevance of the findings is convincingly demonstrated based on a weight-ofevidence approach (see ICH S2 guidelines). This approach will usually need negative results with the impurity from some additional in vitro and/or appropriate in vivo testing.
假如按照证据权衡法能充分证明“结果缺乏体内相关性”，体外试验的阳性结果也仅能在TTC 水平上排除一个杂质(参见 ICH 指南S2) 。这种方法经常需要在额外的体外试验和/或合理的体内试验，并且得到杂质的阴性结果。
A TTC value higher than 1.5 μg/day may be acceptable under certain conditions, e.g.
short-term exposure, for treatment of a life-threatening condition, when life expectancy is less than 5 years, or where the impurity is a known substance and human exposure will be much greater from other sources (e.g. food). Genotoxic impurities that are also significant metabolites may be assessed based on the acceptability of the metabolites.
某些情况下TTC 值高于1.5μg/日也是可以接受的，如短期用药；用于治疗威胁生命疾病的药物；或人的存活期少于5年；或该杂质是已知物质，人体从其他途经（如食物）获得的暴露量远远高于药物途经。如果遗传毒性杂质本身就是重要的代谢物，那么该杂质可以根据代谢物的可接受限度进行控制。
The concentration limits in ppm of genotoxic impurity in drug substance derived from the TTC can be calculated based on the expected daily dose to the patient using equation (1).
采用下列公式，从TTC 值和日服用剂量，可以计算出原料药中的基因毒性杂质的浓度限度。
(1) Concentration limit (ppm) = TTC [μg/day]/dose (g/day]
浓度限度（ppm ） = TTC [μg/day]/剂量(g/day]
The TTC concept should not be applied to carcinogens where adequate toxicity data
(long-term studies) are available and allow for a compound-specific risk assessment.
对于有确切毒性数据（长期毒性研究）的致癌性物质不宜使用TTC 概念，应进行特定化合物风险评估。
It has to be emphasized that the TTC is a pragmatic risk management tool using a probabilistic methodology, i.e. there is a high probability that a 10-5 lifetime cancer risk will not be exceeded if the daily intake of a genotoxic impurity with unknown carcinogenic potential/potency is below the
TTC value. The TTC concept should not be interpreted as providing absolute certainty of no risk.
应强调，TTC 是一个实用性的风险管理方法，是按概率方法学估算的。比如按这一概念，如果某未知致癌性遗传毒性杂质的摄入量低于TTC 值，那么就可以保证患癌风险控制在十万分之一之内。但TTC 概念不能被理解为确保绝对无风险。
5.3 Decision Tree for Assessment of Acceptability of Genotoxic Impurities 基因毒性可接受性评价决策树
(shaded boxes = pharmaceutical assessment, white boxes = toxicological assessment)
（阴影框 = 药学评价，白框 = 毒理学评价）
1) Impurities with structural relationship to high potency carcinogens (see text) are to be excluded from the TTC approach
1) 结构上与高致癌性物质有关的杂质（见正文）不能采用TTC 法。
2) If carcinogenicity data available: Does intake exceed calculated 10-5 cancer lifetime risk?
2) 如果有致癌性数据：摄入量超过10-5患癌风险吗？
3) Case-by-case assessment should include duration of treatment, indication, patient
population etc (see text)
3) 具体情况具体分析，包括用药时间长短、适应症、患者人群等（见正文）。
*) Abbreviations: 缩写
NOEL/UF – No Observed Effect Level/Uncertainty Factor, 未观察到效果水平/不确定因素 PDE – Permitted Daily Exposure, 允许日接触量
TTC – Threshold of Toxicological Concern 毒理学关注的域值
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