Review

Toxicogenomics: challenges and opportunities

G. Orphanides *

Syngenta Central Toxicology Laboratory, Alderley Park, Macclesfield, Cheshire SK10 4TJ, UK

Received 15 September 2002; accepted 12 December 2002

Abstract

Toxicogenomics describes the measurement of global gene expression changes in biological samples exposed to

toxicants. This new technology promises to greatly facilitate research into toxicant mechanisms, with the possibility of

assisting in the detection of compounds with the potential to cause adverse health effects earlier in the development of

pharmaceutical and chemical products. In this short review, I discuss the opportunities presented by toxicogenomics,

the challenges we face in the application of these tools, and the progress we have made in realising the potential of these

new genomic approaches.

# 2003 Elsevier Science Ireland Ltd. All rights reserved.

Keywords: Toxicogenomics; Microarrays; Mechanistic toxicology; Predictive toxicology

1. Introduction

The publication of the draft sequence of the

human genome almost 2 years ago signalled the

arrival of the genomic era of the biological sciences

(International Human Genome Sequencing Con-

sortium, 2001). This newfound knowledge accel-

erated the development of tools that allow

biological processes to be examined on a global

scale. Among these tools are those that facilitate

the simultaneous measurement of the expression

levels of thousands of different genes, technologies

known collectively as gene expression profiling

(Duggan et al., 1999; Brown and Botstein, 1999).

Toxicologists quickly realised the potential of

these new tools to advance their discipline and a

new field was born. The application of gene

expression profiling to toxicology, termed toxico-

genomics, presents us with opportunities to define,

at unprecedented levels of detail, the molecular

events that precede and accompany toxicity,

promising to shed light on toxic mechanisms that

are presently poorly understood (Afshari et al.,

1999; Farr and Dunn, 1999; Nuwasyr et al., 1999;

Pennie, 2000; Pennie et al., 2000; Orphanides et al.,

2001; Gant, 2002; Ulrich and Friend, 2002).

Moreover, it is hoped that gene expression changes

induced upon chemical exposure will provide a

means of predicting mechanisms of toxicity more

rapidly.

Used in conjunction with existing tools available

to the toxicologist, toxicogenomics promises sig-

nificant advances in research and investigative

toxicology. These advances include:

* Tel.: �/44-1625-510803; fax: �/44-1625-590249.

E-mail address: george.orphanides@syngenta.com (G.

Orphanides).

Toxicology Letters 140�/141 (2003) 145�/148

www.elsevier.com/locate/toxlet

0378-4274/03/$ - see front matter # 2003 Elsevier Science Ireland Ltd. All rights reserved.

doi:10.1016/S0378-4274(02)00500-3

. a more detailed appreciation of molecularmechanisms of toxicity.

. faster screens for substance toxicity.

. enhanced extrapolation between experimental

animals and humans in the context of risk

assessment.

In this article, I discuss the use of toxicoge-

nomics in mechanistic and predictive toxicology.

In particular, I examine how far we have come

towards realising the full potential of these tools.

2. Use of toxicogenomics to predict mechanisms of

toxicity

A goal of modern toxicology is to protect the

human population from exposure to harmfulsubstances by identifying compounds with the

potential to cause toxicity. Most current testing

strategies measure the effects of long-term chemi-

cal exposure in experimental animals. Through the

identification of gene expression changes asso-

ciated with chemical exposure, the hope is that

toxicogenomics will facilitate the development of

methods that predict the long-term effects ofcompounds using short-term assays. The under-

lying assumption is that compounds that induce

toxicity through similar mechanisms will elicit

comparable changes in gene expression. It is,

therefore, possible that toxicant-induced expres-

sion changes will act as sensitive and specific

indicators of toxic mechanism. In this way, gene

expression ‘fingerprints’ can be identified formultiple mechanisms of toxic insult and entered

into a database. The gene expression profile of a

suspected toxicant can then be analysed for

similarity with the expression fingerprints of

known toxicants.

The predictive capacity of gene expression

profiling has been demonstrated most compel-

lingly in a clinical setting. A number of studieshave reported the classification of tumour type

using transcript profiling (reviewed by Clarke et

al., 2001). For example, van’t Veer et al. (2002)

identified a gene expression ‘fingerprint’ capable of

distinguishing metastatic and non-metastatic

breast tumours. This approach has also been

used successfully to predict chemical activity. Themost comprehensive study of this kind involved a

combination of chemical treatments and mutant

strains of the yeast Saccharomyces cerevisiae to

generate a gene expression database capable of

predicting the biological effects of exogenous

compounds (Hughes et al., 2000).

Two recent studies indicate that toxicogenomics

can be used to predict chemical mode of action intoxicologically relevant species (Waring et al.,

2001; Hamadeh et al., 2002). These reports de-

monstrate that the liver gene expression profiles

associated with exposure of rats to different

hepatotoxins segregate according to mechanisms

of toxicity. Thus, it appears that the assertion that

toxicogenomics has the potential to provide en-

hanced methods for predicting toxicity is wellfounded. The rodent liver is ideally suited for

demonstrating proof of principle: the hepatocyte is

the predominant cell type, therefore hepatotoxic

chemicals will induce mechanistically linked gene

expression changes in the majority of cells that

make up the organ. However, many toxicants

target only a small proportion of cells in an organ.

A challenge for the future application of toxico-genomics in a predictive context is the identifica-

tion of diagnostic gene expression changes

originating from cells that represent a minority

population. Nevertheless, it appears that this

general approach holds much promise.

3. Toxicogenomics as a mechanistic tool

The global analysis of gene expression levels has

found many diverse applications in modern biol-

ogy. A particular strength of this approach as

applied to toxicology is that it is holistic and,

therefore, provides an unbiased view of alterations

in cellular processes associated with chemical

insult. In this regard, global gene expressionprofiling is an ideal tool for hypothesis generation

in the context of mechanistic toxicology. Indivi-

dual genes, or entire pathways, implicated in a

mechanism of toxicity using this technology can be

further evaluated using more conventional ap-

proaches.

G. Orphanides / Toxicology Letters 140�/141 (2003) 145�/148146

A major challenge in the application of geneexpression technologies to mechanistic toxicology

is the identification of gene regulation events

linked directly to the mode of toxicity under

investigation. Successful application of toxicoge-

nomics in this context requires an understanding

of the link between gene expression changes and

phenotype (Smith, 2001). The simultaneous mea-

surement of changes in the expression levels of tensof thousands of genes is now becoming routine.

However, the increase in the rate at which gene

expression data can be generated has not been

accompanied by corresponding advances in our

ability to interpret them as biologically meaningful

information.

Any given toxicant is likely to induce alterations

in the expression levels of many different genes,and only some of these genes will play a role in the

mechanism of toxicity. Appropriate experimental

design can facilitate the identification of relevant

gene changes. For example, the use of animal

models in which pathways relevant to the mode of

action have been inactivated or modified can aid

the identification of gene expression changes

directly linked to the molecular mechanism of atoxicant. Transgenic ‘knock-out’ mice resistant to

the toxic effects of the compound being studied

can be used to identify genes whose regulation is

not directly related to the development of toxicity.

Changes in gene expression seen in these knock-

out mice exposed to toxicant are unlikely to be

linked to the adverse effects of the compound.

Therefore, any changes in gene expression thatoccur in a sensitive wild-type animal, but not in a

resistant knock-out animal, are more likely to be

directly associated with the mechanism of toxicity.

While, not all gene expression changes that match

this description will be directly involved in the

mode of action of a toxicant, this strategy focuses

attention on the most likely candidates. This

approach as been used to implicate the lactoferrinprotein in the mechanism of rodent non-genotoxic

hepatocarcinogenesis induced by peroxisome pro-

liferators (Hasmall et al., 2002).

Toxicant-induced gene expression changes are

often difficult to interpret in isolation. Careful

selection of compound dose and time of exposure

and the concurrent collection of conventional

toxicology data (e.g. biochemical, clinical andhistopathological data) can greatly facilitate the

interpretation of toxicogenomic data. A successful

toxicogenomic study will, therefore, be multi-

disciplinary, requiring the expert skills of the

toxicologist, pathologist and molecular biologist

(Orphanides et al., 2001).

4. Conclusions

Toxicogenomics is an evolving science. We have

witnessed many successes of the genomic sciencesin other fields of biology, and these tools are now

beginning to enhance our ability to understand

and predict mechanisms of toxicity. It is likely that

toxicogenomics, along with other global profiling

tools such as proteomics (Pandey and Mann, 2000)

and metabonomics (Nicholson et al., 2002), will

revolutionise research and investigative toxicol-

ogy, leading to a holistic appreciation of molecularresponses to toxicants. However, there is still a

long way to go before the full potential of

toxicogenomics is realised. The sheer weight of

data generated by gene expression profiling can be

overwhelming. Extraction of value from this data

will be facilitated by the development of toxicoge-

nomic databases capable of being interrogated by

expert and non-expert user alike. Moreover, theidentification of gene expression changes of pre-

dictive value or mechanistic significance often

requires the use of sophisticated computational

tools, which will evolve alongside gene expression

methodologies (Bassett et al., 1999). One thing we

can be confident about is that the tools of the

genomic era are here to stay. The toxicologist of

the future may feel equally at home with atoxicogenomic data set as with a histopathology

slide.

Acknowledgements

I thank Drs Ian Kimber and Jonathan Moggs

for critical comments on this article and apologise

to those authors whose work I have not cited due

to limitations on article length.

G. Orphanides / Toxicology Letters 140�/141 (2003) 145�/148 147

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