Computational protocol: Identification of chemical modulators of the constitutive activatedreceptor (CAR) in a gene expression compendium

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Protocol publication

[…] Analyses were performed using BRB-ArrayTools version 4.2.1 Stable Release developed by Dr. Richard Simon and BRB-ArrayTools Development Team (http://linus.nci.nih.gov/BRB-ArrayTools.html) (). These studies were carried out independently of the use of the CAR biomarker signature described below. The procedures used were similar to those described previously (). “All samples used in the training and testing sets were first log2 normalized using RMA in the RMAExpress software environment (http://rmaexpress.bmbolstad.com/). The cel files from the three Affymetrix array types (mouse 430A, mouse 430_2 and mouse 430PM arrays) were normalized separately. Normalized expression values of common probesets (22,626) were then combined into one master file. Prior to classification, probesets were excluded under any of the following conditions: 1) minimum fold change - less than 20% of the expression data values have at least a 1.5-fold change in either direction from the median value of the genes, 2) variance is in the bottom 75th percentile, or 3) percent missing exceeds 50%. Filtering using these criteria resulted in 5644 probesets used in the classification study. The 7 models used for class prediction included Compound Covariate Predictor, Bayesian Compound Covariate, Diagonal Linear Discriminant Analysis, 1- and 3-Nearest Neighbor Classifications, Nearest Centroid, and Support Vector Machines with Linear Kernel. The models incorporated genes that were differentially expressed at p ≤ 0.001 significance level, as assessed by the random variance t-test. The prediction error of each model was estimated using 10-fold cross-validation.” Two training sets were used for predicting CAR activation: the samples from wild-type and CAR-null mice from and the same dataset lacking the control and treated CAR-null samples. The derived classifiers of 110 or 247 probesets, respectively, were then used to predict CAR activation of the remaining samples. A test set of 80 and 239 samples known to be positive or negative, respectively, for CAR activation came from a number of studies in which mice or mouse primary hepatocytes were exposed to CAR activators or control substances (; ; Study 1 and Study 3 (above)). […]

Pipeline specifications

Software tools BRB-ArrayTools, RMAExpress
Application Gene expression microarray analysis
Organisms Mus musculus, Martes pennanti
Diseases Fatty Liver, Liver Neoplasms
Chemicals Acetaminophen, Phenobarbital, Silicon Dioxide