ArticleLiterature Review

Mutagenic impurities in pharmaceuticals: A critical assessment of the cohort of concern with a focus on N-nitrosamines

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Abstract

The TTC (Threshold of Toxicological Concern; set at 1.5 μg/day for pharmaceuticals) defines an acceptable patient intake for any unstudied chemical posing a negligible risk of carcinogenicity or other toxic effects. A group of high potency mutagenic carcinogens, defined solely by the presence of particular structural alerts, are referred to as the "cohort of concern" (CoC); aflatoxin-like-, N-nitroso-, and alkyl-azoxy compounds are considered to pose a significant carcinogenic risk at intakes below the TTC. Kroes et al.2004, derived values for the TTC and CoC in the context of food components, employing a non-transparent dataset never placed in the public domain. Using a reconstructed all-carcinogen dataset from relevant publications, it is now clear that there are exceptions for all three CoC structural classes. N-Nitrosamines represent 62% of the N-nitroso class in the reconstructed dataset. Employing a contemporary dataset, 20% are negative in rodent carcinogenicity bioassays with less than 50% of N-nitrosamines estimated to fall into the highest risk category. It is recommended that CoC nitrosamines are identified by compound-specific data rather than structural alerts. Thus, it should be possible to distinguish CoC from non-CoC N-nitrosamines in the context of mutagenic impurities described in ICH M7 (R1).

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... If a mutagenic impurity exceeds the permitted limit, corrective actions may be required, such as process optimization or additional purification steps. To identify and measure mutagenic contaminants, sophisticated analytical methods like nuclear magnetic resonance, mass spectrometry and high-performance liquid chromatography are utilized [40]. By adhering to regulatory guidelines and implementing effective control strategies, pharmaceutical companies aim to minimize the potential risks associated with mutagenic impurities. ...
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Information in the 1999 Physician's Desk Reference as well as from the peer-reviewed published literature was used to evaluate the genotoxicity of marketed pharmaceuticals. This survey is a compendium of genotoxicity information and a means to gain perspective on the inherent genotoxicity of structurally diverse pharmaceuticals. Data from 467 marketed drugs were collected. Excluded from analysis were anti-cancer drugs and nucleosides, which are expected to be genotoxic, steroids, biologicals and peptide-based drugs. Of the 467 drugs, 115 had no published gene-tox data. This group was comprised largely of acutely administered drugs such as antibiotics, antifungals, antihistamines decongestants and anesthetics. The remaining 352 had at least one standard gene-tox assay result. Of these, 101 compounds (28.7%) had at least one positive assay result in the pre-ICH/OECD standard four-test battery (bacterial mutagenesis, in vitro cytogenetics, mouse lymphoma assay (MLA), in vivo cytogenetics). Per assay type, the percentage of positive compounds was: bacterial mutagenesis test, 27/323 (8.3%); in vitro cytogenetics 55/222 (24.8%); MLA 24/96 (25%); in vivo cytogenetics 29/252 (11.5%). Of the supplemental genetic toxicology test findings reported, the sister chromatid exchange (SCE) assay had the largest percentage of positives 17/39 (43.5%) and mammalian mutagenesis assays (excluding MLA) had the lowest percentage of positives 2/91 (2.2%). The predictive value of genetic toxicology findings for 2-year bioassay outcomes is difficult to assess since carcinogenicity can occur via non-genotoxic mechanisms. Nevertheless, the following survey findings were made: 201 drugs had both gene-tox data and rodent carcinogenicity data. Of these, 124 were negative and 77 were equivocal or positive for carcinogenicity in at least 1 gender/1 species. Of the 124 non-carcinogens, 100 had no positive gene-tox findings. Of the remaining 24, 19 were positive in in vitro cytogenetics assays. Among the 77 compounds that exhibited equivocal or positive effects in carcinogenesis studies, 26 were positive in gene-tox assays and 51 were negative. Of the 51 negatives, 47 had multiple negative gene-tox assay results suggesting that these are probably non-genotoxic carcinogens. Statistical analyses suggested that no combination of gene-tox assays provided a higher predictivity of rodent carcinogenesis than the bacterial mutagenicity test itself.
Article
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The threshold of toxicological concern (TTC) is a pragmatic risk assessment tool that is based on the principle of establishing a human exposure threshold value for all chemicals, below which there is a very low probability of an appreciable risk to human health. The concept that there are levels of exposure that do not cause adverse effects is inherent in setting acceptable daily intakes (ADIs) for chemicals with known toxicological profiles. The TTC principle extends this concept by proposing that a de minimis value can be identified for many chemicals, in the absence of a full toxicity database, based on their chemical structures and the known toxicity of chemicals which share similar structural characteristics. The establishment and application of widely accepted TTC values would benefit consumers, industry and regulators. By avoiding unnecessary toxicity testing and safety evaluations when human intakes are below such a threshold, application of the TTC approach would focus limited resources of time, cost, animal use and expertise on the testing and evaluation of substances with the greatest potential to pose risks to human health and thereby contribute to a reduction in the use of animals. An Expert Group of the European branch of the International Life Sciences Institute-ILSI Europe-has examined the TTC principle for its wider applicability in food safety evaluation. The Expert Group examined metabolism and accumulation, structural alerts, endocrine disrupting chemicals and specific endpoints, such as neurotoxicity, teratogenicity, developmental toxicity, allergenicity and immunotoxicity, and determined whether such properties or endpoints had to be taken into consideration specifically in a step-wise approach. The Expert Group concluded that the TTC principle can be applied for low concentrations in food of chemicals that lack toxicity data, provided that there is a sound intake estimate. The use of a decision tree to apply the TTC principle is proposed, and this paper describes the step-wise process in detail. Proteins, heavy metals and polyhalogenated-dibenzodioxins and related compounds were excluded from this approach. When assessing a chemical, a review of prior knowledge and context of use should always precede the use of the TTC decision tree. The initial step is the identification and evaluation of possible genotoxic and/or high potency carcinogens. Following this step, non-genotoxic substances are evaluated in a sequence of steps related to the concerns that would be associated with increasing intakes. For organophosphates a TTC of 18microg per person per day (0.3 microg/kg bw/day) is proposed, and when the compound is not an OP, the TTC values for the Cramer structural classes III, II and I, with their respective TTC levels (e.g. 1800, 540 and 90 microg per person per day; or 30, 9 and 1.5 microg/kg bw /day), would be applied sequentially. All other endpoints or properties were shown to have a distribution of no observed effect levels (NOELs) similar to the distribution of NOELs for general toxicity endpoints in Cramer classes I, II and III. The document was discussed with a wider audience during a workshop held in March 2003 (see list of workshop participants).
Article
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The Carcinogenic Potency Database (CPDB) is a systematic and unifying resource that standardizes the results of chronic, long-term animal cancer tests which have been conducted since the 1950s. The analyses include sufficient information on each experiment to permit research into many areas of carcinogenesis. Both qualitative and quantitative information is reported on positive and negative experiments that meet a set of inclusion criteria. A measure of carcinogenic potency, TD(50) (daily dose rate in mg/kg body weight/day to induce tumors in half of test animals that would have remained tumor-free at zero dose), is estimated for each tissue-tumor combination reported. This article is the ninth publication of a chronological plot of the CPDB; it presents results on 560 experiments of 188 chemicals in mice, rats, and hamsters from 185 publications in the general literature updated through 1997, and from 15 Reports of the National Toxicology Program in 1997-1998. The test agents cover a wide variety of uses and chemical classes. The CPDB Web Site (http://potency.berkeley.edu/) presents the combined database of all published plots in a variety of formats as well as summary tables by chemical and by target organ, supplemental materials on dosing and survival, a detailed guide to using the plot formats, and documentation of methods and publications. The overall CPDB, including the results in this article, presents easily accessible results of 6153 experiments on 1485 chemicals from 1426 papers and 429 NCI/NTP (National Cancer Institute/National Toxicology program) Technical Reports. A tab-separated format of the full CPDB for reading the data into spreadsheets or database applications is available on the Web Site.
Article
The detection of N-nitrosamines, derived from solvents and reagents and, on occasion, the active pharmaceutical ingredient (API) at higher than acceptable levels in drug products, has led regulators to request a detailed review for their presence in all medicinal products. In the absence of rodent carcinogenicity data for novel N-nitrosamines derived from amine-containing APIs, a conservative class limit of 18 ng/day (based on the most carcinogenic N-nitrosamines) or the derivation of acceptable intakes (AIs) using structurally related surrogates with robust rodent carcinogenicity data is recommended. The guidance has implications for the pharmaceutical industry given the vast number of marketed amine-containing drugs. In this perspective, the rate-limiting step in N-nitrosamine carcinogenicity, involving cytochrome P450-mediated α-carbon hydroxylation to yield DNA-reactive diazonium or carbonium ion intermediates, is discussed with reference to the selection of read-across analogs to derive AIs. Risk-mitigation strategies for managing putative N-nitrosamines in the preclinical discovery setting are also presented.
Article
This article reports the outcome of an in silico analysis of more than 12,000 small molecule drugs and drug impurities, identifying the nitrosatable structures, assessing their potential to form nitrosamines under relevant conditions and the challenges to determine compound-specific AIs based on data available or read-across approaches for these nitrosamines and their acceptance by health authorities. Our data indicate that the presence of nitrosamines in pharmaceuticals is likely more prevalent than originally expected. In total, 40.4 % of the analyzed APIs and 29.6 % of the API impurities are potential nitrosamine precursors. Most structures identified through our workflow could form complex API-related nitrosamines, so-called nitrosamine drug substance related impurities (NDSRIs), although we also found structures that could release the well-known small and potent nitrosamines NDMA, NDEA, and others. Due to common structural motifs including secondary or tertiary amine moieties, whole essential drug classes such as beta blockers and ACE inhibitors are at risk. To avoid the risk of drug shortages or even the complete loss of therapeutic options, it will be essential that the well-established ICH M7 principles remain applicable for nitrosamines and that that the industry and regulatory authorities keep an open communication not only about the science but also to make sure there is a good balance between risk and benefit to patients.
Article
Recently, the formation of genotoxic and carcinogenic N-nitrosamines impurities during drug manufacturing of tetrazole-containing angiotensin-II blockers has been described. However, drug-related (complex) nitrosamines may also be generated under certain conditions, i.e., through nitrosation of vulnerable amines in drug substances in the presence of nitrite. An investigation of valsartan drug substance showed that a complex API-related N-nitrosamine chemically designated as (S)-2-(((2'-(1H-tetrazol-5-yl)-[1,1′-biphenyl]-4-yl)methyl)(nitroso)amino)-3-methylbutanoic acid (named 181–14) may be generated. 181–14 was shown to be devoid of a mutagenic potential in the Non-GLP Ames test. According to ICH M7 (R1) (2018), impurities that are not mutagenic in the Ames test would be considered Class 5 impurities and limited according to ICH Q3A (R2) and B (R2) (2006) guidelines. However, certain regulatory authorities raised the concern that the Ames test may not be sufficiently sensitive to detect a mutagenic potential of nitrosamines and requested a confirmatory in vivo study using a transgenic animal genotoxicity model. Our data show that 181–14 was not mutagenic in the transgenic gene mutation assay in MutaTMMice. The data support the conclusion that the Ames test is an adequate and sensitive test system to assess a mutagenic potential of nitrosamines.
Article
The detection of N-nitrosodimethylamine (NDMA) in several marketed drugs led regulatory agencies to require that N-nitrosamine risk assessments be performed on all marketed medical products [EMA/351053/2019 rev 1 (2019)]. Regulation of N-nitrosamine impurity levels in pharmaceutical drug substances and products is described in the ICH M7(R1) guideline where they are referred to as “cohort-of-concern” compounds as several are potent rodent carcinogens [Kroes et. al. 2004]. EMA, U.S. FDA and other regulatory agencies have set provisional acceptable daily intake limits for N-nitrosamines calculated from rodent carcinogenicity TD50 values for experimentally measured N-nitrosamines or the measured TD50 values of close analogs. The class-specific limit can be adjusted based upon a structure activity relationship analysis (SAR) and comparison with analogs having established carcinogenicity data [EMA/369136/2020, (2020)]. To investigate whether improvements in SARs can more accurately predict N-nitrosamine carcinogenic potency, an ad hoc workgroup of 23 companies and universities was established with the goals of addressing several scientific and regulatory issues including: reporting and review of N-nitrosamine mutagenicity and carcinogenicity reaction mechanisms, collection and review of available, public relevant experimental data, development of structure-activity relationships consistent with mechanisms for prediction of N-nitrosamine carcinogenic potency categories, and improved methods for calculating acceptable intake limits for N-nitrosamines based upon mechanistic analogs. Here we describe this collaboration and review our progress to date towards development of mechanistically based structure-activity relationships. We propose improving risk assessment of N-nitrosamines by first establishing the dominant reaction mechanism prior to retrieving an appropriate set of close analogs for use in read-across exercises.
Article
The control of potentially mutagenic impurities in pharmaceutical products is of key importance in assessing carcinogenic risk to humans. The recent discovery of nitrosamine impurities in several marketed pharmaceuticals has increased interest in their mutagenic and carcinogenic potential. This chemical class is considered part of a ‘cohort of concern’, indicating that standard control protocols, such as the use of a threshold of toxicological concern (TTC), cannot be applied. Whilst some nitrosamines are known to be exceptionally potent carcinogens, it's not clear whether this is a property of all members of the class. To investigate the mutagenic and carcinogenic potential of nitrosamines, data was extracted from published literature to augment that already present in the Vitic and Lhasa Carcinogenicity Databases. This data was analysed to assess the application of the ICH M7 guideline to nitrosamine impurities, with respect to the predictivity of the Ames test for carcinogenic potential and the distribution of carcinogenic potency. It was found that 18% of nitrosamines were considered non-carcinogenic. Nitrosamines showed a greater correlation between mutagenicity and carcinogenicity compared to non-nitrosamine compounds. Whilst nitrosamines, in general, are more potent carcinogens than non-nitrosamines, there is a significant overlap between the two distributions of TD50s for each class.
Article
The Microbial mutagenicity Ames test is a bacterial bioassay accomplished in vitro to evaluate the mutagenicity of various environmental carcinogens and toxins. While Ames test is used to identify the revert mutations which are present in strains, it can also be used to detect the mutagenicity of environmental samples such as drugs, dyes, reagents, cosmetics, waste water, pesticides and other substances which are easily solubilized in a liquid suspension. We present the protocol for conducting Ames test in the laboratory.
Article
p>Synthetic and naturally occurring substances present in food and feed, together with their possible breakdown or reaction products, represent a large number of substances, many of which require risk assessment. EFSA’s Scientific Committee was requested to evaluate the threshold of toxicological concern (TTC) approach as a tool for providing scientific advice about possible human health risks from low level exposures, its applicability to EFSA’s work, and to advise on any additional data that might be needed to strengthen the underlying basis of the TTC approach. The Scientific Committee examined the published literature on the TTC approach, undertook its own analyses and commissioned an in silico investigation of the databases underpinning the TTC approach. The Scientific Committee concluded that the TTC approach can be recommended as a useful screening tool either for priority setting or for deciding whether exposure to a substance is so low that the probability of adverse health effects is low and that no further data are necessary. The following human exposure threshold values are sufficiently conservative to be used in EFSA’s work; 0.15 μg/person per day for substances with a structural alert for genotoxicity, 18 μg/person per day for organophosphate and carbamate substances with anti-cholinesterase activity, 90 μg/person per day for Cramer Class III and Cramer Class II substances, and 1800 μg/person per day for Cramer Class I substances, but for application to all groups in the population, these values should be expressed in terms of body weight, i.e. 0.0025, 0.3, 1.5 and 30 μg/kg body weight per day, respectively. Use of the TTC approach for infants under the age of 6 months, with immature metabolic and excretory systems, should be considered on a case-by-case basis. The Committee defined a number of exclusion categories of substances for which the TTC approach would not be used.</p
Article
Information in the 1999 Physician's Desk Reference as well as from the peer-reviewed published literature was used to evaluate the genotoxicity of marketed pharmaceuticals. This survey is a compendium of genotoxicity information and a means to gain perspective on the inherent genotoxicity of structurally diverse pharmaceuticals. Data from 467 marketed drugs were collected. Excluded from analysis were anti-cancer drugs and nucleosides, which are expected to be genotoxic, steroids, biologicals and peptide-based drugs. Of the 467 drugs, 115 had no published gene-tox data. This group was comprised largely of acutely administered drugs such as antibiotics, antifungals, antihistamines decongestants and anesthetics. The remaining 352 had at least one standard gene-tox assay result. Of these, 101 compounds (28.7%) had at least one positive assay result in the pre-ICH/OECD standard four-test battery (bacterial mutagenesis, in vitro cytogenetics, mouse lymphoma assay (MLA), in vivo cytogenetics). Per assay type, the percentage of positive compounds was: bacterial mutagenesis test, 27/323 (8.3%); in vitro cytogenetics 55/222 (24.8%); MLA 24/96 (25%); in vivo cytogenetics 29/252 (11.5%). Of the supplemental genetic toxicology test findings reported, the sister chromatid exchange (SCE) assay had the largest percentage of positives 17/39 (43.5%) and mammalian mutagenesis assays (excluding MLA) had the lowest percentage of positives 2/91 (2.2%). The predictive value of genetic toxicology findings for 2-year bioassay outcomes is difficult to assess since carcinogenicity can occur via non-genotoxic mechanisms. Nevertheless, the following survey findings were made: 201 drugs had both gene-tox data and rodent carcinogenicity data. Of these, 124 were negative and 77 were equivocal or positive for carcinogenicity in at least 1 gender/1 species. Of the 124 non-carcinogens, 100 had no positive gene-tox findings. Of the remaining 24, 19 were positive in in vitro cytogenetics assays. Among the 77 compounds that exhibited equivocal or positive effects in carcinogenesis studies, 26 were positive in gene-tox assays and 51 were negative. Of the 51 negatives, 47 had multiple negative gene-tox assay results suggesting that these are probably non-genotoxic carcinogens. Statistical analyses suggested that no combination of gene-tox assays provided a higher predictivity of rodent carcinogenesis than the bacterial mutagenicity test itself.
Article
The results of studies on the structure and reactivity of spiro[5.2]oct-5,7-diene-4yl carbocation [phenonium ion] have had a significant impact on the course of discussion about the distinction between classical and nonclassical carbocations. This minireview will present a brief overview of the structure, bonding, and reactivity of ring-substituted phenonium ions (X-4+), with an emphasis on work completed since 2004. The discussion will focus on the development of new experimental protocol for determination of the selectivity for addition of nucleophilic anions to X-4+ in aqueous solution. The existing relationships between carbocation lifetime and nucleophilic selectivity, and the known lifetime of ca 140s for spiro[2.5]oct-4,7-diene-6-one provide rough estimates of the lifetimes for 4-Me-4+ and 4-MeO-4+ in aqueous solution. Evidence is presented that nucleophile addition to X-4+ proceeds through an "exploded" transition state, with relatively weak bonding of the nucleophile and leaving group, and the development of significant positive charge at the reacting primary cyclopropyl carbon.
Article
Methyl-t-butylnitrosamine (MtBN) was tested for carcinogenicity by s.c. injection in the Syrian golden hamster. No tumors were observed at 160, 80 or 40 mg/kg/week, in accord with Druckrey's report that ethyl-t-butylnitrosamine (EtBN), the homolog of MtBN, was not carcinogenic in the rat. Acetoxymethyl-t-butylnitrosamine, an activated form of MtBN, was also chronically tested by s.c. injection and found to be non-carcinogenic. It was demonstrated that 2-methyl-propane diazotic acid, the proposed “ultimate” reactive metabolite of MtBN, afforded the carbenium ion product t-butyl alcohol in 63% yield. These results are consistent with the hypothesis that MtBN and EtBN are not carcinogenic because the proposed “ultimate” metabolite in their metabolic activation cannot interact with the critical cellular site due to steric reasons.
Article
Potentially mutagenic impurities in new pharmaceuticals are controlled to levels with negligible risk, the TTC (threshold of toxicological concern, 1.5 μg/day for a lifetime). The TTC was based on the more potent rodent carcinogens, excluding the highly potent "cohort of concern" (COC; for mutagenic carcinogens these are N-nitroso, Aflatoxin-like, and azoxy structures). We compared molecules with DEREK "structural alerts" for mutagenicity used in drug syntheses with the mutagenic carcinogens in the Gold Carcinogenicity Potency Database. Data from 108 diverse synthetic routes from 13 companies confirm that many "alerting" or mutagenic chemicals are in structural classes with lower carcinogenic potency than those used to derive the TTC. Acceptable daily intakes can be established that are higher than the default TTC for many structural classes (e.g., mono-functional alkyl halides and certain aromatic amines). Examples of ADIs for lifetime and shorter-term exposure are given for chemicals of various potencies. The percentage of chemicals with DEREK alerts that proved mutagenic in the Ames test ranged from 36 to 83%, depending on structural class, demonstrating that such SAR analysis to "flag" potential mutagens is conservative. We also note that aromatic azoxy compounds need not be classed as COC, which was based on alkyl azoxy chemicals.
Article
Buffered trifluoroacetolysis of 2-phenylethyl tosylate at 72° produces pure 2-phenylethyl trifluoroacetate in virtually quantitative yield. When the reaction is carried out with 2-phenylethyl-1,1-d2 tosylate for approximately one half-life, complete methylene-group equilibration is found in the trifluoroacetate product, whereas the recovered tosylate is only slightly rearranged. At 75.0° (unbuffered) trifluoroacetolysis of 2-phenylethyl tosylate is 3040 times as fast as that of ethyl tosylate. These results indicate the reaction to proceed entirely via ethylenephenonium ion. Buffered trifluoroacetolysis of 1-phenyl-2-propyl tosylate at 25.0° gives a high yield of unrearranged trifluoroacetate plus a trace of 1-phenyl-1-propyl trifluoroacetate, and proceeds 20.1 times as fast as that of 2-propyl tosylate.
Article
An analysis is presented in which are evaluated correlations among chemical structure, mutagenicity to Salmonella, and carcinogenicity to rats and mice among 301 chemicals tested by the U.S. NTP. Overall, there was a high correlation between structural alerts to DNA reactivity and mutagenicity, but the correlation of either property with carcinogenicity was low. If rodent carcinogenicity is regarded as a singular property of chemicals, then neither structural alerts nor mutagenicity to Salmonella are effective in its prediction. Given this, the database was fragmented and new correlations sought between the derived sub-groups. First, the 301 chemicals were segregated into six broad chemical groupings. Second, the rodent cancer data were partially segregated by target tissue.
Article
The mutagenic and carcinogenic potency of α-CH(n)-nitrosamines such as N-nitrosodimethylamine (NDMA) is caused by their P450-catalyzed α-hydroxylation and subsequent dealkylation, yielding alkyl diazonium ions (R-N≡N(+)) as potent electrophiles. Alternatively, P450s may also catalyze their denitrosation as metabolic detoxification. DFT calculations at the UB3LYP/LANL2DZ(Fe)/6-31G+**(H,C,N,O,S)//LANL2DZ(Fe)/6-31G(H,C,N,O,S) level of theory show that H-abstraction from the α-C of NDMA as initial metabolic step yields an α-nitrosamino radical (•CH(2)N(CH(3))NO) as common first intermediate for both the oxidative dealkylation (toxification) and denitrosation (detoxification) pathways. In particular, the calculated kinetic isotope effect for the P450-mediated dealkylation of NDMA is in good agreement with experimental information. The results show further that the initial α-hydroxylation of NDMA may proceed in two spin states. Besides a stepwise high-spin (HS, quartet) route with a separate rebound barrier, there is a concerted low-spin (LS, doublet) pathway. Interestingly, the resultant two-state reactivity appears to discriminate between metabolic toxification and detoxification: Evaluation of calculated free energy barriers of the H-abstraction (ΔG(‡)) through the Eyring equation suggests that the dealkylation:denitrosation product ratio is governed by the LS:HS ratio of the overall metabolic process. Moreover, inclusion of three further α-CH(n)-nitrosamines in the computational analysis demonstrates that the initial H-abstraction barrier is proportional to the C-H bond dissociation enthalpy (BDE) of the substrates, which enables the estimation of spin-averaged reaction barriers through ground-state BDE calculations. The discussion includes also reductive denitrosation pathways that according to current computational evidence appear to be unlikely for aliphatic nitrosamines.
Article
An index of carcinogenic potency for chemicals tested in chronic animal experiments is described. By analogy with the well-known 'lethal dose 50' (LD50) of quantal bioassay, a 'tumorigenic dose 50' (TD50) may be defined (in the absence both of tumors in the control group and of intercurrent deaths) as that (daily) dose of chemical which gives 50% of the test animals tumors by some fixed age. Tumors in the control (zero-dose) group are handled exactly as for the LD50, and intercurrent deaths are handled by life-table methods. Nonparametric procedures are developed for estimating the TD50 and for constructing confidence intervals. These are based on likelihoods which assume that the tumor hazard is linear in dose.
Article
The mutagenic activities of a diverse group of N-nitrosamines were measured in Salmonella typhimurium TA 100 under conditions designed to maximize metabolism of N-nitrosamines and enhance their mutagenic effects. These conditions were also chosen since some of the carcinogens were previously reported to be non-mutagenk or of questionable mutagenic activity and some only became mutagenic after the bacteria were exposed to a “threshold dose” of metabolites. The mutagenic potencies spanned a range of 105-fold and correlated well with semiquantitative carcinogenic potencies taken from the literature. This correlation appears to be the strongest yet reported for any particular class of compounds. In addition, the mutagenic activities of a number of carcinogens, previously reported to be non-mutagenk, were determined. Among the structural features necessary for high mutagenic activity in this group of compounds was a potential, unsubstituted methylating or ethylating group. Substitution of alkyI, hydroxyl, methoxyl, and cyano moieties at the α or β carbon of these groups reduced mutagenic activity.
Article
This paper presents methods for extending the principle of a single "threshold of regulation" to a range of dietary concentrations between 0.5 and 15 parts per billion by using structure-activity relationships, genotoxicity, and short-term toxicity data. The database used to develop the FDA's threshold of regulation was examined to determine whether structural parameters or the result of certain short-term toxicity tests could be used to define a subset of less potent substances that supports higher threshold levels. In addition, results of reproductive toxicity tests for 3306 compounds and other multidose toxicity tests for 2542 compounds were compared with the database of carcinogenic potencies to establish that carcinogenic endpoints are the most conservative toxicity endpoint for establishing thresholds of regulation.
Article
The concept that "safe levels of exposure" for humans can be identified for individual chemicals is central to the risk assessment of compounds with known toxicological profiles. The Threshold of Toxicological Concern (TTC) is a concept that refers to the establishment of a level of exposure for all chemicals, whether or not there are chemical-specific toxicity data, below which there would be no appreciable risk to human health. The concept proposes that a low level of exposure with a negligible risk can be identified for many chemicals, including those of unknown toxicity, based on knowledge of their chemical structures. The present paper aims to describe the history of the TTC principle, its use to date, its potential future applications and the incorporation of the TTC principle in the Risk Assessment paradigm.
Article
The synthesis of pharmaceutical products frequently involves the use of reactive reagents and the formation of intermediates and by-products. Low levels of some of these may be present in the final drug substance and drug product as impurities. Such chemically reactive impurities may have at the same time the potential for unwanted toxicities including genotoxicity and carcinogenicity and hence can have an impact on product risk assessment. This paper outlines a procedure for testing, classification, qualification, toxicological risk assessment, and control of impurities possessing genotoxic potential in pharmaceutical products. Referencing accepted principles of cancer risk assessment, this document proposes a staged threshold of toxicological concern (TTC) approach for the intake of genotoxic impurities over various periods of exposure. This staged TTC is based on knowledge about tumorigenic potency of a wide range of genotoxic carcinogens and can be used for genotoxic compounds, for which cancer data are limited or not available. The delineated acceptable daily intake values of between approximately 1.5 microg/day for approximately lifetime intake and approximately 120 microg/day for < or = 1 month are virtually safe doses. Based on sound scientific reasoning, these virtually safe intake values do not pose an unacceptable risk to either human volunteers or patients at any stage of clinical development and marketing of a pharmaceutical product. The intake levels are estimated to give an excess cancer risk of 1 in 100,000 to 1 in a million over a lifetime, and are extremely conservative given the current lifetime cancer risk in the population of over 1 in 4 (http://seer.cancer.gov/statfacts/html.all.html). The proposals in this document apply to all clinical routes of administration and to compounds at all stages of clinical development. It is important to note that certain types of products, such as those for life-threatening indications for which there are no safer alternatives, allow for special considerations using adaptations of the principles outlined in this paper.
Article
Prevention of environmentally induced cancers is a major health problem of which solutions depend on the rapid and accurate screening of potential chemical hazards. Lately, theoretical approaches such as the one proposed here - Quantitative Structure-Activity Relationship (QSAR) - are increasingly used for assessing the risks of environmental chemicals, since they can markedly reduce costs, avoid animal testing, and speed up policy decisions. This paper reports a QSAR study based on the Topological Substructural Molecular Design (TOPS-MODE) approach, aiming at predicting the rodent carcinogenicity of a set of nitroso-compounds selected from the Carcinogenic Potency Data Base (CPDB). The set comprises nitrosoureas (14 chemicals), N-nitrosamines (18 chemicals) C-nitroso-compounds (1 chemical), nitrosourethane (1 chemical) and nitrosoguanidine (1 chemical), which have been bioassayed in male rat using gavage as the route of administration. Here we are especially concerned in gathering the role of both parameters on the carcinogenic activity of this family of compounds. First, the regression model was derived, upon removal of one identified nitrosamine outlier, and was able to account for more than 84% of the variance in the experimental activity. Second, the TOPS-MODE approach afforded the bond contributions -- expressed as fragment contributions to the carcinogenic activity -- that can be interpreted and provide tools for better understanding the mechanisms of carcinogenesis. Finally, and most importantly, we demonstrate the potentialities of this approach towards the recognition of structural alerts for carcinogenicity predictions.
The Carcinogenic Potency Database: analyses of 4000 chronic animal cancer experiments published in the general literature and by the U.S. National Cancer Institute/National Toxicology Program
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