Dr. Stephanie Smith-Roe of the Division of the National Toxicology Program (NTP), National Institute of Environmental Health Science will present results of an effort involving ILS, TwinStrand Biosciences, and the NTP at this year’s virtual Environmental Mutagenesis and Genomics Society’s annual meeting. ILS scientists Cheryl A. Hobbs, Ph.D., Leslie Recio, Ph.D., DABT, Michael A. Streicker, LATG, and Miriam V. Rivas, Ph.D. are co-authors of the presented work. The study was supported by ILS Genetic Toxicology and Investigational Toxicology technical staff, and was funded through the NIEHS/NTP’s Genetic Toxicity Testing Contract with ILS. TwinStrand BioSciences provided data analytics and interpretation.
Dr. Smith-Roe’s presentation is entitled, “Adopting Duplex Sequencing™ Technology for Genetic Toxicity Testing: A Proof-of-Concept Mutagenesis Experiment with N-Ethyl-N-Nitrosourea (ENU)-Exposed Rats.”
The work will be featured as a platform talk in Platform Session 7: Detecting Rare Mutations, Gene and Germ Cell Toxicity on Wednesday, September 16, 2020 (1:30 PM – 3:30 PM CDT) and a poster in Block 1: Applied Genetic Toxicology during which Dr. Smith-Roe will be available for a live Q&A session on Saturday, September 12th from 8:00 – 8:30 CDT.
Here is a full abstract detailing the study:
Duplex sequencing (DuplexSeq) is an error-corrected sequencing method in which molecular barcodes are used to preserve the directionality of DNA strands, enabling precise identification and removal of errors arising during amplification and sequencing by comparing complementary sequences. The resulting background of one artifactual mutation per 107 nucleotides allows detection of true, somatic mutations. TwinStrand Biosciences is developing a Duplex Sequencing Rat Mutagenesis Assay to sample the entire rat genome in an unbiased, representative manner, for use in genotoxicity testing. To evaluate the sensitivity of the assay for early detection of mutagenesis, male Hsd:Sprague Dawley SD rats (3/group) received a single oral gavage administration of 40 mg/kg N-ethyl-N-nitrosourea (ENU), and mutation frequency and spectrum were analyzed in blood, bone marrow, liver, and stomach tissues at 3 h, 24 h, 7 days, and 28 days post-exposure. In vehicle control rats (sampled at 28 days), mutation frequency and spectrum were similar for all four tissues. Significant increases in mutation frequency were observed in ENU-exposed rats as early as 3 and 24 h for stomach and bone marrow, respectively, and at 7 days for liver and blood. The mutational signature of ENU – increases in T>A and T>C mutations – was established by 7 days post-exposure in all four tissues. However, an increased fraction of non-canonical C>T mutations was observed in all four tissues at the two earlier timepoints. These results demonstrate that DuplexSeq can detect significant increases in chemical-induced mutations within 24 h of exposure and can monitor the evolution of mutational spectra over time.