The Toxicologist - Society of Toxicology

oligomers synthesized with both CPD and oxoG lesions were then used in in vitro
assays to evaluate both polymerase activity and fidelity during replication of undamaged
and damaged DNA. We anticipate that some or all of these β-strand mutations
will alter the activity and fidelity of Pol η compared to wild-type and that
the results from these studies will suggest that these amino acids provide an important
structural component to the enzyme necessary for the successful bypass of
DNA lesions during replication as well as organism survival. Furthermore, we believe
that the findings will contribute to an explanation for the low bypass fidelity
of Pol η past oxoG lesions.
123 COMMONALITY AND STOCHASTICITY IN GENE
EXPRESSION PROFILES DURING AGING PROCESS.
T. Inoue 1 and Y. Hirabayashi 2 . 1 Center for Biological Safety & Research, National
Institute of Health Sciences, Tokyo, Japan and 2 Division of Cellular & Molecular
Toxicology, Center for Biological Safety & Research, National Institute of Health
Sciences, Tokyo, Japan.
Senescence is conceptually considered to be xenobiotic responses to “time”, where
essential biological responses during aging may be based on a lifetime-dependent
relationship between living creature and xenobiotic materials in relationship with
oxidative stresses and photo-stimulation, among others. When one examines a hundred
senescent mice, senescent phenotypes may be generally observed nearly in all
mice. Among them, there may be common changes in the most on one hand, there
may be also probabilistic phenotypes observed individually in case by case manner.
When one compares those phenotypes in senescent mice comparing with those of
less senescent young mice by gene chip microarray, two different categories of gene
expression profiles are recognized; i.e., one corresponds to common aging profiles
(CAPs) selected by two-way analysis of variance (ANOVA) and the latter corresponds
to stochastic aging profiles (SAPs) selected by principal component analysis
(PCA). Reason we focused such stochastic gene expression was because we recognized
that gene expression profiles in microarray were mostly uniform (common)
when those in RT-PCRs were uniform, whereas those profiles in microarrays individually
vary (stochastic) when those data in RT-PCRs were divergent individually.
CAPs are definitive expression profiles to senescent mice whereas SAPs are probabilistic
expression profiles observed in one senescent mouse to the other. Gene expression
profiles compared bone marrow cells from 2- and 21-month-old male
mice of the C3H/He strain provided 122 probe sets of CAPs and 1005 of SAPs,
resp.. When one examines the relationships between CAP-related and SAP-related
signalings using network database, stochastic gene expressions scattered in the networks
occasionally merged downstream toward the CAP genes, stochastically. CAPs
seem to be conciliatorily regulated by positive and negative signals from various stochastic
genes differing from one case to another.
124 COMPARISON OF TOXICOGENOMICS PROFILES TO
DISCOVER CO-REGULATED GENES.
Y. Igarashi 1 , N. Nakatsu 1 , H. Yamada 1 , A. Ono 2 , Y. Ohno 2 and T. Urushidani 1,
3 . 1 National Institute of Biomedical Innovation, Ibaraki-City, Osaka, Japan,
2 National Institute of Health Sciences, Setagaya-ku, Tokyo, Japan and 3 Doshisha
Women’s College of Liberal Arts, Kyotanabe-City, Kyoto, Japan.
We have developed in silico methodology for toxicogenomics data to compare a
pair of probe sets considering with their time courses and dose effects. This method
can be applied to any kinds of gene expression data which is obtained in a systematic
manner. The toxicogenomics project in Japan, started in 2002, produced
genome-wide gene expression data of the liver of rat in vivo and vitro using 150
pharmaceutical drugs. The gene expression data has been obtained at 8 time points
and 4 dose levels in vivo, and 3 time points and 4 dose levels in vitro. One of the
reasons the identification of safety biomarkers is difficult is that transcriptional responses
following treatment are embedded in the feed back loops of signaling pathways.
Those responses would be varied at different time points and dose levels even
if same drug is treated. Therefore, it is high opportunity to identify co-regulated
genes when the expression levels are similar at continuous time points and dose levels.
We have developed simple and flexible algorithm to compare genome-wide
transcriptional profiling consisting of multiple time and dose points. Our method
compares gene expression profiles between a pair of probe sets. The similarity is calculated
using a pair of vectors based on the expression levels at continuous 2 time
points and 2 dose levels. For example, a probe which consists of 8 time points and
4 dose levels has 21 vectors for the comparison. Using this method, we have estimated
clusters of co-expressed gene sets including several known co-regulated genes
by treatment of lipopolysaccharide. This method has a great potential to identify
co-regulated genes which lead to identification of safety biomarkers.
26 SOT 2011 ANNUAL MEETING
125 USE OF PATHWAY ANALYSES TO IDENTIFY
POTENTIAL TARGET SAFETY RISKS FOR ANTIBODY
DRUG CONJUGATES (ADC).
B. Lu1 , L. Obert1 , A. Hooper2 and M. Gosink1 . 1Drug Safety Research &
Development, Pfizer, Groton, CT and 2Center for Integrative Biology &
Biotherapeutics, Pfizer, Pearl River, NY.
Antibody drug conjugates (ADC), which use an antibody to deliver a cytotoxic
drug selectively into a target cell, represent a new and powerful therapeutic approach
against diseases such as cancer. Different from a traditional drug’s mode of
action (MOA), an ADC achieves efficacy largely by selectively eliminating targetexpressing
cells. Assessing the target safety for an ADC program at a early development
stage (e.g. pre-lead development) would be difficult because there is a lack of
in vivo as well as in vitro assays to experimentally approach such issues. Here we
present an in silico approach to use pathway analyses to identify potential target
safety risks for Antibody drug conjugates (ADC). We propose that the primary target
safety risks for ADC should first be analyzed at the cellular level; essentially the
target-expressing cell should be approached as an integral unit. Secondary risks
should be evaluated using knowledge of pathway databases to study the ADC target
and its downstream pathways. Because ADC therapy can remove all cells expressing
a particular target, any extracellular signaling performed by target expressing cells
would also be ablated. The absence of those signaling molecules could have a significant
impact on neighboring cells as well as other tissues. We introduced such a
working concept by using Ingenuity pathway analysis software on two ADC targets
CD19 and CD30. We further ran the analysis on a hypothetical ADC target:
EGFR and found target risk for EGFR ADC therapy may be greater than those
risks of conventional anti-EGF therapy because in conventional anti-EGF therapy
feedback mechanism could partially compensate for the loss of EGF. Given the
huge interest and investment in ADCs, the knowledge driven approach presented
here might help to highlight some potential target safety risks in their early development
stage.
126 COMPARISON OF NEXT GENERATION SEQUENCING
AND MICROARRAY ANALYSES OF GENE EXPRESSION
IN KIDNEY OF THE RATS TREATED WITH
CARCINOGEN ARISTOLOCHIC ACID.
T. Chen 1 , Z. Li 1 , Z. Su 2 and L. Shi 3 . 1 Division of Genetic and Molecular
Toxicology, U.S. FDA-NCTR, Jefferson, AR, 2 Z-Tech, U.S. FDA-NCTR, Jefferson,
AR and 3 Division of System Biology, U.S. FDA-NCTR, Jefferson, AR.
Next-generation sequencing (NGS) has been used to evaluate gene expression level.
In this study, we compared gene expression profiles of aristolochic acid (AA) in rat
kidney generated from NGS and microarray technologies. AA is the active component
of herbal drugs derived from Aristolochia species that have been used for medicinal
purposes since antiquity. AA, however, induced nephropathy and urothelial
cancer in humans and malignant tumors in the kidney, urinary tract and other tissues
in rodents. Rats were treated with 10 mg/kg or vehicle control for 12 weeks
and the kidney tissues were quickly isolated and frozen immediately after the treatment.
Four samples for the treatment and 4 for the control were used for the comparison.
The transcriptome analysis of the samples was conducted using the
Affymetrix Rat Genome 230 2.0 Microarray for the microarray method and
Illumina Genome Analyzer II for the NGS approach. The gene expression data
generated from both platforms were processed using Significance Analysis of
Microarray (SAM) for determining the deferentially expressed genes (DEG) and
Ingenuity Pathway Analysis for defining the altered biological functions. The gene
expression profiles resulted from both platforms showed that AA treatment produced
a large number of DEGs, 1297 for microarray and 4247 for NGS. There
were 925 DEGs that were commonly identified by both platforms. Functional
analysis demonstrated that the major biological processes dysregulated by the treatment
by both platforms were very similar and related to chemical carcinogenesis.
Tumorigenesis and apoptosis were identified as the top functional pathways for
both platforms. These results indicate that while the alteration of gene expression
determined by NGS is comparable to that by microarray in terms of the biological
functional analysis, NGS is more sensitive than microarray for detecting DEGs.
127 THE ARYL HYDROCARBON RECEPTOR PROTEIN
INTERACTION NETWORK (AHR-PIN).
J. J. LaPres 1, 2 , D. M. Tappenden 1, 2 , L. Yang 3 and R. S. Thomas 3 . 1 Biochemistry,
Michigan State University, East Lansing, MI, 2 Center for Integrative Toxicology,
Michigan State University, East Lansing, MI and 3 The Hamner Institutes for Health
Sciences, Research Triangle Park, NC.
The aryl hydrocarbon receptor (AHR), a ligand-activated PAS super-family transcription
factor member, is responsible for controlling most, if not all, of the toxic
effects of environmental pollutants, such as polyaromatic hydrocarbons (PAHs).