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Using a gene expression biomarker to identify DNA damage-inducing agents in microarray profiles.

ABSTRACT: High-throughput transcriptomic technologies are increasingly being used to screen environmental chemicals in vitro to provide mechanistic context for regulatory testing. The TGx-DDI biomarker is a 64-gene expression profile generated from testing 28 model chemicals or treatments (13 that cause DNA damage and 15 that do not) in human TK6 cells. While the biomarker is very accurate at predicting DNA damage inducing (DDI) potential using the nearest shrunken centroid method, the broad utility of the biomarker using other computational methods is not fully known. Here, we determined the accuracy of the biomarker used with the Running Fisher test, a nonparametric correlation test. In TK6 cells, the methods could readily differentiate DDI and non-DDI compounds with balanced accuracies of 87-97%, depending on the threshold for determining DDI positives. The methods identified DDI agents in the metabolically competent hepatocyte cell line HepaRG (accuracy = 90%) but not in HepG2 cells or hepatocytes derived from embryonic stem cells (60 and 80%, respectively). DDI was also accurately classified when the gene expression changes were derived using the nCounter technology (accuracy = 89%). In addition, we found: (1) not all genes contributed equally to the correlations; (2) the minimal overlap in genes between the biomarker and the individual comparisons required for significant positive correlation was 10 genes, but usually was much higher; and (3) different sets of genes in the biomarker can by themselves contribute to the significant correlations. Overall, these results demonstrate the utility of the biomarker to accurately classify DDI agents. Environ. Mol. Mutagen. 59:772-784, 2018. Published 2018. This article is a U.S. Government work and is in the public domain in the USA.

SUBMITTER: Corton JC

PROVIDER: S-EPMC7875442 | biostudies-literature | 2018 Dec

REPOSITORIES: biostudies-literature

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Using a gene expression biomarker to identify DNA damage-inducing agents in microarray profiles.

Corton J Christopher JC Williams Andrew A Yauk Carole L CL

Environmental and molecular mutagenesis 20181017 9

High-throughput transcriptomic technologies are increasingly being used to screen environmental chemicals in vitro to provide mechanistic context for regulatory testing. The TGx-DDI biomarker is a 64-gene expression profile generated from testing 28 model chemicals or treatments (13 that cause DNA damage and 15 that do not) in human TK6 cells. While the biomarker is very accurate at predicting DNA damage inducing (DDI) potential using the nearest shrunken centroid method, the broad utility of th ...[more]

PMID: 30329178

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Project description:BackgroundCurrent biosensors are designed to target and react to specific nucleic acid sequences or structural epitopes. These 'target-specific' platforms require creation of new physical capture reagents when new organisms are targeted. An 'open-target' approach to DNA microarray biosensing is proposed and substantiated using laboratory generated data. The microarray consisted of 12,900 25 bp oligonucleotide capture probes derived from a statistical model trained on randomly selected genomic segments of pathogenic prokaryotic organisms. Open-target detection of organisms was accomplished using a reference library of hybridization patterns for three test organisms whose DNA sequences were not included in the design of the microarray probes.ResultsA multivariate mathematical model based on the partial least squares regression (PLSR) was developed to detect the presence of three test organisms in mixed samples. When all 12,900 probes were used, the model correctly detected the signature of three test organisms in all mixed samples (mean(R²)) = 0.76, CI = 0.95), with a 6% false positive rate. A sampling algorithm was then developed to sparsely sample the probe space for a minimal number of probes required to capture the hybridization imprints of the test organisms. The PLSR detection model was capable of correctly identifying the presence of the three test organisms in all mixed samples using only 47 probes (mean(R²)) = 0.77, CI = 0.95) with nearly 100% specificity.ConclusionsWe conceived an 'open-target' approach to biosensing, and hypothesized that a relatively small, non-specifically designed, DNA microarray is capable of identifying the presence of multiple organisms in mixed samples. Coupled with a mathematical model applied to laboratory generated data, and sparse sampling of capture probes, the prototype microarray platform was able to capture the signature of each organism in all mixed samples with high sensitivity and specificity. It was demonstrated that this new approach to biosensing closely follows the principles of sparse sensing.

Project description:DNA microarray is a powerful tool in biomedical research. However, transcriptomic profiling using DNA microarray is subject to many variations including biological variability. To evaluate the different sources of variation in mRNA gene expression profiles, gene expression profiles were monitored using the Affymetrix RatTox U34 arrays in cultured primary hepatocytes derived from six rats over a 26 hour period at 6 time points (0 h, 2h, 5h, 8h, 14 h and 26 h) with two replicate arrays at each time point for each animal. In addition, the impact of sample size on the variability of differentially expressed gene lists and the consistency of biological responses were also investigated. Excellent intra-animal reproducibility was obtained at all time points with 0 out of 370 present probe sets across all time points showing significant difference between the 2 replicate arrays (3-way ANOVA, p <or= 0.0001). However, large inter-animal biological variation in mRNA expression profiles was observed with 337 out of 370 present probe sets showing significant differences among 6 animals (3-way ANOVA, p <or= 0.05). Principal Component Analysis (PCA) revealed that time effect (PC1) in this data set accounted for 47.4% of total variance indicating the dynamics of transcriptomics. The second and third largest effects came from animal difference, which accounted for 16.9% (PC2 and PC3) of the total variance. The reproducibility of gene lists and their functional classification was declined considerably when the sample size was decreased. Overall, our results strongly support that there is significant inter-animal variability in the time-course gene expression profiles, which is a confounding factor that must be carefully evaluated to correctly interpret microarray gene expression studies. The consistency of the gene lists and their biological functional classification are also sensitive to sample size with the reproducibility decreasing considerably under small sample size.

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Using a gene expression biomarker to identify DNA damage-inducing agents in microarray profiles.

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Using a gene expression biomarker to identify DNA damage-inducing agents in microarray profiles.

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