viernes, 15 de febrero de 2019

NCTR Research Highlights—Genetic and Epigenetic Research

  • Effect of Carcinogens on Transcriptomic and Epigenetic Alterations in Liver Cells
  • Epigenome-Wide Association Study of Lupus Based on Ethnicity 
  • Minimally Invasive Gene-Mutation Assay May Identify Mutagens and Carcinogens 
  • Detection of Rare Genomic Mutations Using Next-Generation Sequencing

Genetic and Epigenetic Research at NCTR

Effect of Carcinogens on Transcriptomic and Epigenetic Alterations in Liver Cells
NCTR scientists, in collaboration with the University of New Mexico Comprehensive Cancer Center, investigated the utility of high-throughput microarray gene expression and next-generation sequencing for the in vitro identification of genotoxic and non-genotoxic carcinogens. This approach may substantially enhance the identification and assessment of potential liver carcinogens. The increasing number of man-made chemicals in the environment that may pose a carcinogenic risk highlights the need for developing reliable time- and cost-effective approaches for carcinogen detection and identification.  
Transcriptomic analysis of human-liver HepaRG cells treated at minimally toxic concentrations with three different carcinogens generated distinct gene-expression profiles. In contrast to transcriptomic alterations, treatment of liver cells with the carcinogenic and non-carcinogenic chemicals resulted in profound changes in the DNA methylation footprint; however, the correlation between gene-specific DNA methylation and gene expression changes was minimal. Among the carcinogen-altered genes, transferrin (TF) emerged as a sensitive marker for an initial screening of chemicals for their potential liver carcinogenicity. Potential liver carcinogens (i.e., chemicals causing altered TF gene expression) could then be subjected to gene-expression analyses to differentiate genotoxic from non-genotoxic liver carcinogens. Information about this study can be found in Food and Chemical Toxicologydisclaimer icon.
For more information, contact Volodymyr Tryndyak, Ph.D. or Igor Pogribny, M.D., Ph.D., Division of Biochemical Toxicology, FDA/NCTR.      
Epigenome-Wide Association Study of Systemic Lupus Erythematosus (SLE) Based on Ethnicity
NCTR scientists published findings from an epigenome-wide association study of lupus and non-lupus patients, and within those populations examined methylation profiles between African-American and European-American women. In addition, they studied the SLE disease activity status of those individuals — those with less disease (SLE score<6 disease="" increased="" score="" those="" to="" with="">6). Autoimmune diseases affect women at a higher ratio (9:1) than men and at a higher rate among African-American women. Lupus is an autoimmune disease where the body attacks its own cells and organs. The SLE form of lupus is known to cause damage to several organs such as the kidney, heart, lung, and to joints in the body. Lupus was thought to be a genetic disease because it often occurred in families, but now research has established the importance of epigenetics (no alterations in gene structure) in lupus pathogenesis.
Until 2011, FDA had not approved a drug for lupus in fifty years, so research to spur development of lupus therapeutics is greatly needed. Different DNA profiles were observed in genes involved in the Type-1 interferon pathway in lupus versus age-matched control subjects. Furthermore, African-American women with lupus had a more robust DNA profile than European women with lupus. The identification of these epigenetic biomarkers can greatly improve diagnosis of this disease in lupus patients. Several epigenetic drugs are currently in various stages of development in clinical trials for other diseases, but data from these trials and other data suggest that epigenetic drugs may be potentially useful in lupus patients. This study was recently published in the Journal of Autoimmunitydisclaimer icon.
For additional information, contact Beverly Lyn-Cook, Ph.D., Division of Biochemical Toxicology, FDA/NCTR.
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Minimally Invasive Gene-Mutation Assay May Identify Mutagens and Carcinogens 
The Pig-a assay is an emerging test for rapidly monitoring gene mutation(s) in rats, mice, and humans. The assay is minimally invasive, conducted on erythrocytes from peripheral blood. One drop of blood contains enough erythrocytes to conduct the most sensitive assays.
NCTR scientists are currently validating this assay as a regulatory test suitable for identifying mutagens and potential carcinogens. A recent study has developed a novel version of the assay for rat bone-marrow erythroid cells. These cells are the direct precursors of red blood cells found in circulating blood. Mammalian erythrocytes lack genomic DNA; therefore, confirmation of mutation induction — a necessary step for assay validation — is not possible in erythrocytes. The data from an NCTR study conclusively demonstrate that the Pig-a mutant erythrocytes (red blood cells) measured in the circulating blood of mutagenized rats were descendent from cells containing Pig-a mutations. In contrast, bone marrow erythroid precursor cells have nuclei and DNA, and thus are suitable for sequencing Pig-a mutations. The two publications below describing these findings are now available in Environmental Molecular Mutagenesis.
For more information, please contact Javier Revollo, Ph.D. or Azra Nfn, Ph.D., Division of Genetic and Molecular Toxicology, FDA/NCTR.
Detection of Rare Genomic Mutations Using Next-Generation Sequencing (NGS) 
FDA scientists from NCTR and the Center for Drug Evaluation and Research are developing a sensitive method to detect mutations induced by chemicals. Mutations are changes in the DNA sequence of an organism, ranging from small point mutations to large chromosome alterations that can cause adverse health effects, such as cancer and genetic disease. The goal of the ongoing study is to establish a new next generation sequencing (NGS) assay that may become a powerful, rapid, and practical tool to routinely evaluate the mutagenicity of FDA-regulated products. The scientists treated bacteria and mammalian cells with different types of mutagenic chemicals, then the DNA from cloned cells was isolated and sequenced using NGS. The sequencing data were analyzed by a bioinformatics pipeline established for the study.
For the bacterial part of the study, the NGS results showed that the mutagens caused significant mutation induction over the controls. The frequencies and types of mutations were like those previously reported with traditional mutation assays. For the mammalian cell part of the study, mouse lymphoma cells were treated with mutagenic chemicals, then the DNA samples from large and small colony clones were isolated for whole genome sequencing. The mutagens induced mutation frequencies greater than their concurrent untreated controls and the types of mutations suggested a mechanism for the mutagenesis. Data from this study have been interpreted and reported in Archives of Toxicologydisclaimer icon.
For more information, please contact Tao Chen, Ph.D., Division of Genetic and Molecular Toxicology, FDA/NCTR.

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