Genetic Toxicity Testing Services for the NTP

Integrated Laboratory Systems, Llc, Research Triangle Park NC

Investigators

Abstract

This genetic toxicology testing contract has supported the overall mission of NTP to provide detailed toxicological profiles, including potential for carcinogenicity, of compounds of interest for over 30 years. Assessment of genetic damage, an important factor in the etiology of cancer as well as neurodegenerative diseases such as ALS, neurological conditions such as autism, aging, birth defects such as Down syndrome, mitochondrial diseases, and other adverse human health effects, is a critical component of any in-depth toxicological profile of a chemical or product. This Genetic Toxicity Testing contract provides information on exposure-related genetic damage to the NTP using standard tests as well as innovative approaches when necessary. The capabilities of the contract are continuously updated to remain current and consistent with international guidelines for conducting these kinds of tests. Testing systems employed include both in vitro (animal cell-based and bacterial) and in vivo (rats and mice) assays. Three main tests are conducted routinely: in vitro bacterial mutation assays, in vivo rodent erythrocyte blood micronucleus (MN) assays, and in vivo rodent DNA damage (comet) assays in multiple tissues including, for example, liver, brain, stomach, kidney, and lung. Using peripheral blood as the sample source, MN and comet studies can also be conducted in humans to translate findings from animal studies to human exposure scenarios to better characterize potential adverse human health effects from a particular exposure. We have conducted such studies in the past, looking into potential genetic effects of particular drugs and botanical products that had demonstrated adverse effects in NTP animal studies. The NTP is increasingly employing in vitro MN and comet assays as a substitute for in vivo rodent MN assays, where possible, to reduce our use of animals in toxicology testing. A new addition to the NTP battery of genotoxicity assays, an animal mutation endpoint that holds promise for application in human clinical and biomonitoring studies in the future, is now routinely assessed in the testing laboratory: the Pig-a gene mutation assay (phosphatidylinositol glycan anchor biosynthesis, class A gene). Mutations in this gene are easily detected in red blood cell samples from laboratory rodents, and updated methods have been developed to allow easy integration of this endpoint into ongoing animal toxicity tests. Where possible, we multiplex this assay with the in vivo MN and comet assays, increasing the genetic damage information we obtain from test animals to provide a comprehensive profile of the genetic toxicity potential of a chemical. During the past year, we investigated whether frozen tissue samples obtained for the comet assay and stored long term were suitable for generating gene expression patterns, another indication of biological effect of chemical exposure. We found that we could obtain high quality data from these frozen tissue samples, providing yet another approach to assessment of genetic effects induced by chemical exposure. In addition to our cell-based and animal model studies, human blood samples provided by collaborators such as the NIEHS Clinical Research Unit may also be examined for specific genetic endpoints (MN, comet, Pig-a) following an environmental exposure that is suspected of inducing genetic damage, or has been shown to do so in NTP animal studies. The number of genetic toxicity studies of each type described above varies annually, depending on the needs of the NTP. During the past year, historical control data bases have been established for MN and Pig-a data from a new strain of mouse ? the CD-1 mouse ? that is used in NTP reproductive (transgenerational) studies. In addition, a Pig-a historical control database has been established for the Sprague Dawley rat and the CD-1 mouse models used in in these reproductive toxicity studies. We now are able to place our rat Pig-a and our CD-1 mouse MN and Pig-a data into a broader context, which is helpful in the interpretation of the observed responses. This contract has the ability to be highly flexible in response to NTP needs, and in situations where rapid generation of genetic damage data are required, as, for example, in response to an environmental disaster, resources can be concentrated on generating the required data quickly, such as occurred with the West Virginia Elk River water contamination event. Key words: genetics, mutation, chromosome damage, cancer, non-cancer effects, neurodevelopmental disorders, heritable disease, in vitro testing, chemical exposure, toxicology, biotechnology, DNA damage, birth defects, genetic disease, genomics, DNA sequencing, gene expression, botanical products, dietary supplements, dietary supplements