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Pharmacogenetics
EDITED BY

Ian P. Hall
University Hospital of Nottingham
Nottingham, U.K.

Munir Pirmohamed
University of Liverpool
Liverpool, U.K.

New York London

Taylor & Francis is an imprint of the

Taylor & Francis Group, an informa business
Foreword

Pharmacogenetics seeks to determine how a person’s genetic makeup affects their

response to medicines. Rapid advances in genetic technologies have opened up the possi-
bility of genetic testing on a large scale at an affordable cost, thus facilitating the exploita-
tion of pharmacogenetics not only in assessing risks, benefits, and indications of
medicines, but also as a valuable tool in drug discovery.
Since the beginning of modern drug therapy, it has been realized that dose is a
poor predictor of therapeutic response and that there is substantial variability in both
therapeutic efficacy and the occurrence of adverse effects of most medicines. One of
the main aims of clinical pharmacology is to understand the basis of this variability.
Whether it is due to differences in drug disposition caused by variations in absorption,
distribution, transport, metabolism, or elimination, or by differences in end-organ
responsiveness, many studies have shown that genetic variability is a most important
determinant of each of these.
In the field of drug discovery, the application of genetics was originally focused on
the discovery of novel disease genes that could become the targets for new drug discovery
programs. But the rapid expansion of understanding of the variability of human DNA
sequences has given rise to an appreciation that this approach may be more challenging
than originally thought. Currently, more use is made of population-based genetic
association studies to identify disease intervention targets.
As the recent report of the Royal Society “Personalised Medicines: Hopes and
Realities” makes abundantly clear, it is easy to exaggerate the importance of pharma-
cogenetics on clinical practice. While there are a few medicines on the market where
there is evidence for the benefits of genetic testing for patient selection, mainly in the
field of cancer, there are as yet no marketed products that are the result of genetic-
based discovery. It seems likely that this situation will radically change, but the time
scale is not yet clear. It seems more likely that, for the foreseeable future, the impact
of pharmacogenetics will be most profound in the development of diagnostic tests that
will allow for the better use of medicines, both in terms of efficacy and safety.
This book is a timely account of the state of the science and clinical application
of pharmacogenetics. Its format should make it attractive and easy to use for the clinician

iii
iv Foreword

in search of detailed information about specific disease areas, and also for those wishing
to be updated in the field of pharmacogenetics in general.

Alasdair Breckenridge
Medicines and Healthcare Products Regulatory Agency
London, U.K.
Preface

This book aims to provide a comprehensive account of pharmacogenetics with particular

reference to its potential role in specific disease areas. While there have been previous
books on pharmacogenetics, these have concentrated on the broader principles (often
almost exclusively dealing with pharmacokinetic issues). We felt that a book that provides
an account of the generic issues and then provides a disease-area-by-disease-area account
would be of value.
This approach has an obvious advantage in that a reader specifically interested in
the possible role of pharmacogenetics in dealing with patients with cardiovascular
disease, for example, will be able to find all the relevant information in a single
chapter, as opposed to having to read about specific enzymes or receptor systems and
pick out those bits of information relevant to the disease area. It has, however, caused
some difficulty in editing the text because obviously some issues (e.g., the role of
cytochrome P450 polymorphism) are generic to many disease areas. In general, we left
essential information in each chapter to ensure that the reader can gain all the relevant
details without constantly having to cross-refer to other chapters. This means that there
is a small amount of repetition in some of the chapters, but wherever possible we have
reduced this to the minimum required to maintain the flow of the text.
The other obvious difficulty in this fast-moving subject area is ensuring that the
account is up to date. While many of the broad issues have not changed over recent
years, many new studies are underway. We tried to ensure that the book is as comprehen-
sive as possible.
Looking through the contents readers will note that there is no specific chapter
dealing with neurological disease; however, the relevant issues are dealt with either in
the chapter on adverse drug reactions (anticonvulsants) or psychiatric disease. Similarly,
the relatively small amount of information on renal disease pharmacogenetics is covered
in the sections on metabolic and cardiovascular disease.
We hope that this book provides a useful starting point for both clinicians and
non-clinicians with an interest in pharmacogenetics. This is an exciting time to be
involved in the study of pharmacogenetics. Although the concept of personalized prescribing
has been around for about 50 years, the next 10 years will prove whether or not this approach
to improving the effectiveness of prescribing by achieving high efficacy while reducing the
risk of severe adverse drug reactions is a viable generic approach to patient management.

Ian P. Hall
Munir Pirmohamed
v
Contents

Foreword Alasdair Breckenridge . . . . iii

Preface . . . . v
Contributors . . . . xi

Part I: Principles of Pharmacogenetics

1. Identification of Treatment Response Genes . . . . . . . . . . . . . . . . . . . . . . . . 1
Eva Halapi and Hakon Hakonarson
Introduction . . . . 1
Pharmacogenetics of Metabolizing Enzymes: CYP450 and
Drug Transporter Genes . . . . 2
Pharmacogenetic Targets in Cancer . . . . 3
Pharmacogenetic Targets in Central Nervous System Disorders . . . . 6
Pharmacogenetic Targets in Respiratory Disorders . . . . 8
Pharmacogenetic Targets in Cardiovascular Disease . . . . 9
Microarray and Disease Response Genes . . . . 10
Conclusion . . . . 12
References . . . . 13

2. Methods of Genotyping . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21
Matthias Wjst and Monika Werner
Introduction . . . . 21
High-Throughput Genotyping Methods . . . . 22
Test Principles . . . . 22
50 Nuclease Assay . . . . 27
Special Genotyping Applications . . . . 30
Future Developments . . . . 32
References . . . . 32

3. Pharmacoproteomics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35
Alaisdair C. Stamps and Jonathan A. Terrett
Introduction . . . . 35

vii
viii Contents

Historical Perspective . . . . 35
Proteomic Techniques . . . . 38
Current Activities in Pharmacoproteomics . . . . 43
Conclusions . . . . 47
References . . . . 47

4. Potential Social, Ethical, and Legal Issues Raised by the Development

of Pharmacogenetics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 51
Paul Martin, Andrew Smart, and Robert Dingwall
Introduction—Assessing the Potential Impact of
Pharmacogenetics . . . . 51
Bioethics and Its Limitations . . . . 52
Possible Application of Pharmacogenetics . . . . 53
Social and Ethical Issues . . . . 56
Conclusion: The Social and Ethical Issues Raised by
Pharmacogenetics—Implications for Policy . . . . 62
References . . . . 63

Part II: Clinical Practice and Pharmacogenetics

5. Pharmacogenetics of Adverse Drug Reactions . . . . . . . . . . . . . . . . . . . . . . 65
Ana Alfirevic, B. Kevin Park, and Munir Pirmohamed
Introduction . . . . 65
Definitions and Classification of Adverse Drug Reactions . . . . 66
Historical Overview . . . . 66
Variability in Drug Handling and Adverse Drug Reactions . . . . 67
Genetic Variability in Drug Metabolizing Enzymes . . . . 67
Genetic Variability in Drug Transporters and Adverse
Drug Reactions . . . . 73
Immunogenetic Polymorphisms . . . . 74
Receptors . . . . 76
Miscellaneous Drug Targets . . . . 77
Summary and Future Directions . . . . 78
References . . . . 80

6. Pharmacogenetics of Respiratory Disease . . . . . . . . . . . . . . . . . . . . . . . . . . 91

Ian P. Hall and Stephen B. Liggett
Introduction . . . . 91
b2 Adrenoceptor Polymorphism . . . . 91
Clinical Consequences of b2AR Polymorphisms . . . . 98
Pharmacogenetics of Theophylline . . . . 102
Cys Leukotriene Receptor Pathway Polymorphism . . . . 103
Glucocorticoids and Treatment Response . . . . 105
Pharmacogenetics of Muscarinic Receptor Antagonists . . . . 106
Summary . . . . 106
References . . . . 107

7. Hematological Pharmacogenetics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111

Howard L. McLeod, Charles Eby, and Brian Gage
Introduction . . . . 111
Contents ix

Warfarin Pharmacogenetics . . . . 112

Genetic Polymorphisms in Hemostatic Systems . . . . 116
Thiopurine Pharmacogenetics . . . . 121
The Future . . . . 123
References . . . . 123

8. Pharmacogenetics in Oncology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 129

Romano Danesi, Antonello Di Paolo, Mario Del Tacca, and
Filippo De Braud
Introduction . . . . 129
Pharmacogenetics of Drug Targets: Theoretical Background . . . . 131
Pharmacogenetics of Drug Targets and
Enzymes of Drug Metabolism . . . . 133
Pharmacogenetics of DNA Repair Systems
and Apoptosis . . . . 141
The ATP-Binding Cassette Superfamily of Drug
Transporters . . . . 144
Clinical Pharmacogenetics . . . . 144
Conclusions and Future Prospects . . . . 147
References . . . . 147

9. Pharmacogenetics in Infectious Diseases . . . . . . . . . . . . . . . . . . . . . . . . . . 155

Belle L. Lee, Laurent A. Decosterd, Reinhold Kerb, and
Amalio Telenti
Introduction . . . . 155
Pharmacogenetics of Specific Infectious Diseases . . . . 157
Other Pharmacogenetic Markers of Interest . . . . 169
Immunogenetics in Infectious Diseases . . . . 175
Validation of Pharmacogenetic Markers . . . . 176
Conclusions . . . . 177
References . . . . 178

10. Pharmacogenetics in Rheumatology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 189

Samantha L. Hider, Catharine Morgan, Wendy Thomson, and
Ian N. Bruce
Introduction . . . . 189
Therapeutic Options in the Treatment of Rheumatoid
Arthritis . . . . 189
Efficacy and Toxicity of Antirheumatic Therapies . . . . 190
Nonsteroidal Anti-Inflammatory Drugs . . . . 191
Methotrexate . . . . 193
Sulfasalazine . . . . 195
D- Penicillamine . . . . 195
Gold . . . . 196
Azathioprine . . . . 196
Leflunomide . . . . 198
Antimalarial Drugs . . . . 199
Biological Therapies . . . . 200
Summary and Future Directions . . . . 202
References . . . . 202
x Contents

11. Polymorphisms in Cardiovascular Medicine: The Role of Genetic

Variants in Disease Diagnosis and Drug Response . . . . . . . . . . . . . . . . . . 209
Yasmin, David N. Cooper, and John R. Cockcroft
Introduction . . . . 209
Polymorphism in the Human Genome . . . . 210
Polymorphism in Cardiovascular Disease . . . . 211
Genetic Polymorphisms and Response to Drugs . . . . 225
A Perspective on Pharmacogenetics in
Cardiovascular Medicine . . . . 232
References . . . . 232

12. Pharmacogenetics and Metabolic Disease . . . . . . . . . . . . . . . . . . . . . . . . . 243

Simon Constable and Munir Pirmohamed
Introduction . . . . 243
Diabetes Mellitus . . . . 243
Hyperlipidemia . . . . 251
Osteoporosis . . . . 257
Conclusions . . . . 261
References . . . . 262

13. Pharmacogenetics in Gastroenterology . . . . . . . . . . . . . . . . . . . . . . . . . . . 273

Guruprasad Padur Aithal
Introduction . . . . 273
Application of Pharmacogenetics in Clinical Gastroenterology
and Hepatology . . . . 273
Candidate Gene Case – Control Association Studies . . . . 284
Future Developments . . . . 286
References . . . . 286

14. The Genetics of Antipsychotic Response: Pharmacogenetic and

Pharmacogenomic Investigations and Clinical Applications . . . . . . . . . . . 295
Maria J. Arranz, Dalu Mancama, and Robert W. Kerwin
Introduction . . . . 295
Pharmacogenetic and Pharmacogenomic Research . . . . 295
Clinical Applications of Pharmacogenetics
and Pharmacogenomics . . . . 302
Future Research: Investigations on Genetic Expression . . . . 304
References . . . . 304

Index . . . . 311
Contributors

Guruprasad Padur Aithal Queen’s Medical Centre University Hospital,

Nottingham, U.K.
Ana Alfirevic Department of Pharmacology and Therapeutics, University of
Liverpool, Liverpool, U.K.
Maria J. Arranz Section of Clinical Neuropharmacology, Institute of
Psychiatry—King’s College, London, U.K.
Ian N. Bruce arc Epidemiology Unit, University of Manchester, Manchester, U.K.
John R. Cockcroft Wales Heart Research Institute, Cardiff University, Cardiff, U.K.
Simon Constable Department of Pharmacology and Therapeutics, University
of Liverpool, Liverpool, U.K.
David N. Cooper Institute of Medical Genetics, Cardiff University, Cardiff, U.K.
Romano Danesi Division of Pharmacology and Chemotherapy, Department of
Oncology, University of Pisa, Pisa, Italy
Filippo De Braud Clinical Pharmacology and New Drug Development Unit,
European Institute of Oncology, Milano, Italy
Laurent A. Decosterd Division of Clinical Pharmacology, University of Lausanne,
Lausanne, Switzerland
Mario Del Tacca Division of Pharmacology and Chemotherapy, Department
of Oncology, University of Pisa, Pisa, Italy
Antonello Di Paolo Division of Pharmacology and Chemotherapy, Department
of Oncology, University of Pisa, Pisa, Italy
Robert Dingwall Institute for the Study of Genetics, Biorisks and Society,
University of Nottingham, Nottingham, U.K.
Charles Eby Department of Medicine, Washington University School of Medicine,
Saint Louis, Missouri, U.S.A.
Brian Gage Department of Medicine, Washington University School of Medicine,
Saint Louis, Missouri, U.S.A.
xi
xii Contributors

Hakon Hakonarson Division of Inflammation and Pharmacogenomics, deCODE

Genetics, Inc., Reykjavik, Iceland
Eva Halapi Division of Inflammation and Pharmacogenomics, deCODE
Genetics, Inc., Reykjavik, Iceland
Ian P. Hall Division of Therapeutics and Molecular Medicine, University Hospital
of Nottingham, Nottingham, U.K.
Samantha L. Hider arc Epidemiology Unit, University of Manchester,
Manchester, U.K.
Reinhold Kerb Experimental Medicine, Astra Zeneca, Mölndal, Sweden
Robert W. Kerwin Section of Clinical Neuropharmacology, Institute of
Psychiatry—King’s College, London, U.K.
Belle L. Lee University of California, San Francisco, California, U.S.A., and Division
of Infectious Diseases, Department of Clinical Research, University Hospital, Berne,
Switzerland
Stephen B. Liggett Departments of Medicine and Physiology, University of
Maryland School of Medicine, Baltimore, Maryland, U.S.A.
Dalu Mancama Section of Clinical Neuropharmacology, Institute of
Psychiatry—King’s College, London, U.K.
Paul Martin Institute for the Study of Genetics, Biorisks and Society, University
of Nottingham, Nottingham, U.K.
Howard L. McLeod Department of Medicine, Washington University School
of Medicine, Saint Louis, Missouri, U.S.A.
Catharine Morgan arc Epidemiology Unit, University of Manchester,
Manchester, U.K.
B. Kevin Park Department of Pharmacology and Therapeutics, University of
Liverpool, Liverpool, U.K.
Munir Pirmohamed Department of Pharmacology and Therapeutics, University
of Liverpool, Liverpool, U.K.
Andrew Smart Institute for the Study of Genetics, Biorisks and Society, University
of Nottingham, Nottingham, U.K.
Alaisdair C. Stamps UCB-Celltech, Slough, U.K.
Amalio Telenti Institute of Microbiology, University Hospital, University
of Lausanne, Lausanne, Switzerland
Jonathan A. Terrett Medarex, Inc., Milpitas, California, U.S.A.
Wendy Thomson arc Epidemiology Unit, University of Manchester,
Manchester, U.K.
Contributors xiii

Monika Werner GSK Glaxo Smith Kline Beecham, Munich, Germany

Matthias Wjst Gruppe Molekulare Epidemiologie, GSF—Forschungszentrum für
Umwelt und Gesundheit, Neuherberg/Munich, Germany
Yasmin Clinical Pharmacology Unit, University of Cambridge, Cambridge, U.K.
PART I: PRINCIPLES OF PHARMACOGENETICS

1
Identification of Treatment
Response Genes
Eva Halapi and Hakon Hakonarson
Division of Inflammation and Pharmacogenomics, deCODE Genetics, Inc.,
Reykjavik, Iceland

INTRODUCTION

Genetic diversity contributes to both disease susceptibility and variability in response to

drug therapy. Pharmacogenomics is a discipline focused on examining the genetic basis
for individual variations in response to therapeutics (1 – 4). Although the task of develop-
ing individualized medicines tailored to patient’s genotypes poses a major scientific chal-
lenge, pharmacogenomics is already starting to influence how physicians/scientists design
clinical trials and its impact on the practice of medicine is forthcoming (5,6). Recent evi-
dence suggests that most prescribed medications are effective in no more than 60% of the
individuals in whom they are used, and a significant number of patients also develop major
adverse effects. Better understanding of the genetic factors that regulate patient’s respon-
siveness to drugs is therefore needed to elucidate the molecular mechanisms involved and
allow for development of new therapeutic strategies that match each patient and the most
suitable drug (7 – 9).
While drug treatment constitutes the mainstay of medicine, for most drugs, there is
considerable variability in patient’s therapeutic response (2,3). In other cases, unforeseen
serious side effects may occur (10,11). For the patient this represents a dangerous and
potentially life-threatening situation, and, at the societal level, adverse drug reactions
are the most common cause of hospital admissions in the elderly and represent a
leading cause of disease and death (12). In some cases, genetic variations have been
shown to influence both efficacy and safety profiles, as in the case of dicumarol, warfarin,
or isoniazid, wherein patient variation in response to these drugs can largely be attributed
to polymorphisms in the CYP450 gene family that confer rapid versus slow acetylation of
these drugs (10). Since genetic variations can lead to differences in the regulatory func-
tions of genes, variability in their mRNA and/or protein expressions may follow. Pharma-
cogenomics is charged with measuring these differences in mRNA and protein messages
in response to drugs, and although relatively few examples of success exist, this approach
holds the promise that we may be able to profile these variations in individuals’ genetic
makeup and accurately predict response to drugs addressing both efficacy and safety
issues (3,4,6,9,10).

1
2 Halapi and Hakonarson

In recent years, microarray technology has revolutionized almost all fields of

biomedical research by enabling high-throughput gene expression profiling in a single
experiment, thereby allowing thousands of genes in different species to be examined
in the context of organ differentiation and development in search for disease susceptibility
genes and new targets in drug discovery. With the use of expression microarray, the
future holds the promise that we may soon gain more global understanding of gene
expression changes with respect to disease susceptibility and progression and that
biomarkers will become increasingly used as diagnostic and prognostic indicators of
treatment response and also provide new insights into the process of new target discovery.
A brief overview of some of the key examples of pharmacogenetic effects follows.
This is designed to give the reader an initial idea of the potential for pharmacogenetic
effects to contribute to disease management. Fuller accounts of the range of pharmacoge-
netic effects relevant to each major disease group can be found in the chapters that follow.

PHARMACOGENETICS OF METABOLIZING ENZYMES: CYP450 AND

DRUG TRANSPORTER GENES

The cytochrome P450 (CYP450) enzyme system is involved in various metabolic and bio-
synthetic processes and constitutes a superfamily of heme enzymes found in most organ-
isms (i.e., bacteria to humans). These enzymes are estimated to account for the
biotransformation of approximately 60% of the most commonly prescribed drugs in the
United States. A few representative examples are discussed in the following, and
additional details on some of these examples (and others) are contained in later chapters.

CYP2D6
Three main phenotypes have been identified that relate to the oxidative metabolisms of
drug substrates by CYP2D6. The slow metabolizers (with defective CYP2D6 alleles),
the normal metabolizers (wild type), and the ultra-rapid metabolizers all have variable
number of genes for the functional CYP2D6 enzyme (13 –15). Within the Caucasian popu-
lation, approximately 7% of the CYP2D6 alleles are defective, resulting in potentially
increased concentration of various drug metabolites at conventional therapeutic doses.
Some of the most commonly used drugs, such as the beta-adrenoceptor blockers and tri-
cyclic antidepressants, are substrates for CYP2D6 (13 – 15). The latter are known to cause
adverse reactions that are attributed to increased levels of tricyclic antidepressants. These
include, but are not limited to, such events as life-threatening arrhythmias and other
cardiotoxic effects that result from decreased activity of CYP2D6 metabolism. Diagnostic
tests are now available to identify beforehand those who are at risk.

CYP2C9
Three defective alleles have been reported in the CYP2C9 enzyme, two of which
confer decreased activity. Their frequency ranges from 1% to13% in different populations
(13 –15). Substrates of CYP2C9 include nonsteroidal anti-inflammatory drugs and hypo-
glycemic agents (13 – 15). The clinical relevance of CYP2C9 is particularly noticed in the
metabolism of drugs that are used to treat Type II diabetes, wherein decreased clearance of
these drugs may result in severe hypoglycemia. Another relevant example is S-warfarin,
where major bleeding may occur (10).
Identification of Treatment Response Genes 3

CYP2C19
CYP2C19 is a highly polymorphic enzyme system, with approximately 3% of the Cauca-
sians having allele variation that renders them as slow metabolizers. In contrast, almost
20% of the Asian population carries the slow allele. Allele 9 confers total abolishment
of enzymatic activity (13 – 15). For most pharmacological compounds that are substrates
for CYP2C19, this poses a limited problem because the majority of the drugs are metab-
olized by several CYP450 enzymes. An important exemption is the drug omeprazole,
which is only partially metabolized by CYP3A4 and shows up to 12-fold larger area
under the concentration – time curve (AUC) in slow versus fast metabolizers (13 –15).

Drug Transporters
Blood and tissue concentrations of most drugs are influenced by interindividual variation
in the structure and function of the metabolizing enzyme and transporter genes. Transpor-
ters are genes that control drug uptake, distribution, and elimination. The multidrug resist-
ance gene (MDR1) encodes for a P-glycoprotein (PgP), which belongs to the large
adenosine triphosphate (ATP)-binding cassette (ABC) protein. The MDR1 gene was orig-
inally discovered as the protein causing cross-resistance of tumors to many different cyto-
toxic agents (16). Multiple substrates are transported by PgP, including the
chemotherapeutics tamoxifen and mitoxantrone, the antibiotics cefotetan and cefazolin,
the immunosuppressant cyclosporin A, the antiarrytmic drug quindine, the cardiac stimu-
lants digoxin, and such opioid drugs as morphine, to name a few (17). Many cancers are
known to overexpress the PgP protein, and this has been correlated with poor prognosis,
particularly in patients with leukemia (18). Several SNPs have been reported in the MDR1
gene, some of which have been correlated with PgP protein expression, notably including
the C/T polymorphism in exon 26 (19). In a recent study (20), antiviral response to nelfi-
navir and efavirenez was shown to correlate with the allelic variant, 3435C/T, of the
MDR1 gene. Patients who were homozygotes, carrying two copies of the 3435 T allele,
demonstrated lower serum concentration, faster recovery in CD4 T cell count, and more
rapid decrease in viral load, suggesting that the MDR1 3435C/T variant may be predictive
of immune recovery after antiviral treatment in HIV patients (20). Approximately 50% of
Caucasians are heterozygote (C/T) at the 3435 MDR1 polymorphic site, while a homozy-
gote state (C/C) or (T/T), is seen in 25% of individuals, respectively. In contrast, the
frequency of the CC genotype in African Americans is 67% to 83%, whereas the fre-
quency of the TT allele is very low. Increased expression of PgP has also been correlated
with variation in clinical response to glucocorticoids in patients with inflammatory bowel
disease (21) and systemic lupus erythematosus (22). Collectively, these studies suggest
that there are important variations in the MDR1 gene that regulate tumor resistance,
immune function, and metabolism of multiple drugs.

PHARMACOGENETIC TARGETS IN CANCER

For most cancers, conventional histopathologic evaluation, encompassing tumor

grade and stage, is inadequate to accurately predict the biological behavior of the tumor
(23 –25). Considerable effort is underway to identify and characterize the biological
potential of various cancers at the molecular level. The need to predict response to
therapy and determine which tumors are most likely to progress or recur or which invasive
4 Halapi and Hakonarson

tumors will metastasize has prompted intensive efforts in search for prognostic biomarkers
and markers that correlate with patients’ responses to anticancer therapy.

Estrogen and Progesterone Receptors

In the United States and other western societies, approximately one in 10 women develops
breast cancer. Given the unacceptably high mortality rate of approximately 40%, it is cri-
tically important that the most effective therapy at any given time is administered to each
patient. Although both adjuvant chemo- and hormonal therapies reduce the risk of metas-
tasis by approximately one-third, the best indicators for clinical progression available
today, including lymph node status, tumor size and histological grade, are unable to accu-
rately predict the outcome.

SERM
The presence of estrogen and/or progesterone receptors on tumors is considered favorable
because these patients are eligible for hormonal treatment. Tamoxifen, is a selective estro-
gen-receptor modulator (SERM), which acts as an estrogen antagonist in normal breast
tissue and breast cancer cells but as an antagonist in liver and bone cells. Apart from low-
ering serum cholesterol and preventing postmenopausal osteoporosis (26), tamoxifen is
the most effective and extensively used hormonal treatment for all stages of breast
cancer. More recently, the drug was approved for prevention of breast cancer in high-
risk individuals. In a recent meta-analysis, which included information on 37,000
women in 55 clinical trials of adjuvant tamoxifen therapy for five years (27), reduction
in recurrence and mortality rates were 47% and 26%, respectively, over a 10-year
period. Fifty percent decrease was also observed in the incidence of contralateral breast
cancer in patients receiving tamoxifen, regardless of the ER status of the primary tumor
(27). Several SERMs are currently in clinical trials. Toremifene is a relatively new
SERM drug with properties similar to that of tamoxifen. However, unlike tamoxifen, tor-
emifene does not seem to increase the risk of endometrial cancer. Based on information
available to date, the Food and Drug Administration (FDA) has restricted the use of tor-
emifene to postmenopausal women with metastatic breast cancer. Raloxifene is another
antiestrogen SERM that has received approval for the treatment of osteoporosis in
women beyond menopause.

Aromatase Inhibitors
Aromatase, a cytochrome P-450 enzyme that catalyzes the conversion of androgens to
estrogens, is the major source of estrogen synthesis in postmenopausal women. Inhibition
of aromatase, the terminal step in estrogen biosynthesis, provides a mechanism to inter-
vene in hormone-dependent breast cancer in postmenopausal women. Compared to
tamoxifen, both nonsteroidal (such as anastrozole and letrozole) and steroidal (such as
exemestane) aromatase inhibitors (AIs) provide superior efficacy and better toxicity
profile as first- and second-line therapy of metastatic disease. Early results from the
ATAC study (anastrozole, tamoxifen, alone or in combination trial), encompassing
9300 women with early-stage disease at 381 research and medical centers in 21 countries,
suggest that anastrozole is superior to tamoxifen as measured by disease-free survival in
receptor-positive patients and in reducing the incidence of contralateral breast cancer (28).
Identification of Treatment Response Genes 5

ER Antagonists
Fulvestrant (faslodex) is a potent antiestrogen drug that mediates its effects by estrogen
receptor (ER) downregulation. It acts as a pure antiestrogen and exhibits none of the nega-
tive side effects associated with the partial agonist activity of tamoxifen and related drugs.
It has been shown to have comparable efficacy to that of the oral AI, anastrozole, in post-
menopausal women with advanced breast cancer who have failed to respond to tamoxifen
or related drugs (29). It therefore provides the clinician with an alternative therapeutic
strategy following the development of tamoxifen resistance. Fulvestrant might also be
beneficial as a follow-on therapy after tamoxifen in an adjuvant setting to palliate some
of the concerns surrounding a long-term therapy with tamoxifen (five years).
Antiestrogens are among the most potent therapies in preventing cancer and reducing
the risk of recurrence in high-risk patients and in treating metastatic disease. Thus, ER
expression has become a valuable marker in predicting treatment response in breast cancer.

Epidermal Growth Factor Receptor Family

Epidermal Growth Factor Receptor
The epidermal growth factor (EGF) pathway has been identified as a key regulator of cell
growth and replication (30,31). Cumulative evidence shows that the epidermal growth
factor receptor (EGFR) pathway is actively involved in a wide variety of solid tumurs,
including non– small cell lung cancer (NSCLC), prostate cancer, breast cancer, stomach
cancer, colon cancer, ovarian cancer, and tumors of the head and neck (30 – 32). Overex-
pression of the EGFR in cancer cells has been associated with more advanced disease,
development of metastatic phenotypes, and poor prognosisents (32). IressaTM , a new tyro-
sine kinase inhibitor, directly blocks the signals for cell growth and division and is cur-
rently licensed for the treatment of inoperable or recurrent NSCLC in Asia and is being
tested in clinical trials for other solid tumors (33). Other drugs targeting the EGFR are
TarcevaTM and ErbituxTM , which are currently in clinical trials.

HER2/neu
HER2/neu has been shown to be overexpressed in 20% to 30% of breast cancer patients
(34,35). Recent evidence supports a clear association between HER2 overexpression and
reduced overall and disease-free survival, especially in patients with node-positive
disease (34 –38). Tumors displaying HER2 amplification show a correlation with poor
prognosis (39,40). Thus, the greatest value of HER2 as a predictive marker lies in the pre-
diction of response to therapies that target HER2, notably herceptin. Indeed, patients with
strongly HER2-positive breast cancer get significant clinical benefits from herceptin
therapy, and HER2 testing has become an integral part of the optimal management of
breast cancer patients. It is important to determine HER2 status of all primary breast
cancers at the time of diagnosis and recurrence because HER2 overexpression and ampli-
fication can be used to identify patients for herceptin therapy (41). Thus, HER2 has
approached a clinically validated status as a prognostic factor and also as a predictive
factor for response to therapy, and it is already part of the routine assessment for breast
cancer patients. A prior knowledge of HER2 status is therefore an absolute requirement
for herceptin therapy.

Thiopurine S-Methyltransferase
The thiopurine S-methyltransferase (TPMT ) gene metabolizes thiopurine medications,
such as mercaptopurine, azathioprine, and thioguanine. TPMT activity is polymorphic
6 Halapi and Hakonarson

with 10% of the subjects being heterozygous and about 1/300 with low or deficient
activity (42,43). Patients with low or deficient TPMT activity are at high risk for severe
hematological toxicity from standard doses of thiopurine medications. Thus, it is import-
ant to be able to identify those patients who are at risk for such complications. The mol-
ecular basis for altered TPMT activity is well characterized with three distinct alleles
accounting for up to 95% of both heterozygous and homozygous mutant patients (43).
However, significant ethnic differences have been identified in both the frequency of
low TPMT activity and in the mutations that account for them (43). Although extended
clinical studies are needed to better understand and quantitate the dosing of these drugs,
TPMT genotype-specific dosing guidelines have been proposed for the use of mercapto-
purine in leukemia patients (44). Given the severity of thiopurine medication-related tox-
icity in TPMT-deficient patients, screening of the entire patient populations for TPMT
polymorphism prior to prescribing these drugs has been shown to be cost-effective. By
determining thiopurine transferase activity in patients before they receive thiopurines,
enzyme efficacy can be determined beforehand and life-threatening complications,
which are strongly related to the genetically determined activity of this enzyme, can be
avoided. Accordingly, screening for genetic variations in the TPMT gene presents an
ideal model for the translation of genomic information to guide patient therapeutics.

N-Acetyltransferase (NAT)
NAT was first identified as the enzyme responsible for inactivation of the antitubercular
drug isoniazid (45). NAT also plays an important role in carcinogen metabolism. The
N-acetylation metabolizing pathway is a major route for the conjugative metabolism
of many drugs and chemicals (46). Functional polymorphisms in the NAT gene were
initially associated with differences in the susceptibility to occupational and smoking-
related bladder cancer (47). Based on the substrate, individuals can be phenotyped as
either “fast” or “slow” acetylators. Individuals with the slow phenotype are homozygotes
for the slow allele, whereas subjects with the fast phenotype are either heterozygotes or
homozygotes for the fast allele. The frequency of slow acetylator varies worldwide,
ranging from 5% to 10% in Asia and reaching 90% frequency in certain European popu-
lations (13). Two functionally relevant human NAT genes, NAT1 and NAT2, have been
identified that are highly polymorphic and are encoded at multi-allelic loci. The relation-
ship between these polymorphisms and the resulting phenotypes is well established (48).

PHARMACOGENETIC TARGETS IN CENTRAL NERVOUS

SYSTEM DISORDERS

Although pharmacotherapy in patients with affective disorders has improved the outcome
of millions of patients worldwide, medical treatment of mental depression is efficacious in
no more than two-thirds of the cases, and there are no biological markers of treatment
response. Apart from identification of genomic markers of treatment response, which
would constitute an enormous clinical advantage of public health value, the application
of pharmacogenmoics may also uncover new targets for the development of novel and
hopefully more efficacious drugs with favorable side-effect profile.

Serotonin Transporter
Selective serotonin reuptake inhibitors (SSRIs) are widely used for the treatment of
depression because of their efficacy and relatively favorable side-effect profile compared
Identification of Treatment Response Genes 7

with those of the tricyclic antidepressants. SSRIs act by interfering with the activity of the
serotonin transporter (SERT) (49). A number of polymorphisms have been reported in the
SERT gene (50 – 52), and genetic variations in the SERT promoter have been linked to
altered functions, such as the association between the short (S) allele (44 bp deletion)
of the SERT-PR site and poor response to fluvoxamine and paroxetine in patients
treated for major depression (53,54).

Dopamine Transporter
Attention deficit/hyperactivity disorder (ADHD) affects between 3% and 5% of school-
age children. Treatment of major symptoms, such as inattention, hyperactivity, and
impulsive behavior, has been effectively achieved using psychostimulants, of which
methylphenidate (Ritalinw) is the most commonly prescribed in the United States.
These drugs are beneficial in many cases; however, inter-individual variation in clinical
response and adverse events is well documented (55 –57). Methylphenidate binds to
and directly inhibits the dopamine transporter (DAT1). Accordingly, variations in genes
involved in dopamine action and metabolism (such as DAT1, D2, and the D4 receptor
genes) have been examined in search for explanation of the variability in clinical response
to methylphenidate and other psychostimulants in ADHD patients. A recent study (58)
demonstrated significant association between the 10/10 genotype in DAT1 and lack of
response to methylphenidate but failed to demonstrate any association with polymorphism
in the dopamine receptors (58). These results have recently been confirmed in a larger
patient cohort (59).

Dopamine and Serotonin Receptors

Clozapine is a potent drug in the treatment of schizophrenia; however, not all patients
benefit from treatment, and some patients react adversely to therapy while others fail to
respond adequately. Several studies have reported an association between clozapine
response and polymorphisms in the dopamine receptor 3 (D3) gene (60 –62), dopamine
receptor 4 (D4) gene (63,64), and in the serotonin receptor 2A (65,66) and 5A genes
(67). In a recent study, Arranz et al. (68) screened for the combination of upto 19 poly-
morphisms that predicted clinical response to clozapine in schizophrenic patients with
high accuracy. These 19 single-nucleotide polymorphisms (SNPs) were located in eight
receptor genes and one transporter gene, including the a2A-adrenoceptor, dopamine D3
receptor, 5-HT2A, 5-HT2C, 5-HT3A, 5-HT5A, histamine H1 receptor, histamine H2
receptor, and the serotonin transporter gene. A combination of six polymorphisms, includ-
ing the 5-HT2A 102T/C, His452Tyr, 5-HT2C-330GT/-244CT, Cys23Ser, 5-HTTLPR,
H2-1018G/A, predicted clinical response with 76.86% accuracy (x2 ¼ 35.8; P ¼0.0001)
with sensitivity of 95.89 (+0.04), suggesting that these polymorphisms could be used to
identify patients who are most likely to show a satisfactory improvement with treatment.

Apolipoprotein E
Apart from its role in cardiovascular disease, the apoliprotein E (ApoE) protein has also been
associated with late onset and sporadic Alzheimer’s disease (AD) (69,70). In the central
nervous system (CNS), ApoE plays a key role in mobilization and redistribution of choles-
terol and phospholipids during membrane remodeling (70). In a study assessing the influence
of the ApoE genotype on therapeutic response to tacrine (acetylcholinesterase inhibitor) in a
responder/nonresponder cohort, more than 80% of ApoE4 negative carriers demonstrated
8 Halapi and Hakonarson

marked clinical response to tacrine, as measured by the AD assessment scale (ADAS),

whereas ApoE4 positive carriers presented ADAS scores that were worse compared with
the baseline levels (69). The ApoE genotype has also been implicated in modulating response
to drugs targeting vasopressinergic activity (71). Thus, ApoE is emerging as a potentially
useful marker in predicting clinical response to tacrine.

PHARMACOGENETIC TARGETS IN RESPIRATORY DISORDERS

There are four major classes of asthma pharmacotherapy currently in widespread use (72):
(i) b2-agonists (b-agonist) used by inhalation for the relief of airway obstruction
(e.g., albuterol, salmeterol, fenoterol); (ii) glucocorticoids for both inhaled and systemic
use (e.g., fluticazone, beclomethasone, triamcinolone, prednisone); (iii) theophylline
and its derivatives; and (iv) inhibitors and receptor antagonists of the cysteinyl-
leukotriene pathway (e.g., montelukast, pranlukast, zafirlukast, zileuton). The following
section summarizes pharmacogenetic work reported on b-agonists and inhibitors of the
cysteinyl-leukotriene pathway in asthma; comprehensive accounts of these effects are
contained in the chapter on respiratory disease.

b2 Adrenergic Receptor
Four polymorphisms of the coding block of b2 adrenergic receptor (b2AR) have been
found, three of which result in receptors that have different properties compared with
the wild-type (73). These polymorphisms include Arg16 ! Gly, Gln27 ! Glu,
Val34 ! Met, and Thr164 ! Ile, of which the first two are the most common. Most
studies have found no differences in the frequencies of these polymorphisms between
asthma patients and healthy nonasthmatic controls (74,75). Thus, the genetic variability
of the b2AR does not appear to play a major causative role in asthma. However, these poly-
morphisms, although not causative, could modify the disease. Other studies have assessed
the relationship between b2AR polymorphisms at positions 16 and 27 and atopy, including
IgE levels (76). A significant association between the Glu27 form of the b2 receptor and log
serum IgE was reported, suggesting that b2AR polymorphisms may act to modify the asth-
matic phenotype (77).
Several studies have assessed the modulatory role of b2AR polymorphisms on the
treatment response to b-agonists (2,78) in relation to bronchial hyperreactivity (79) and
control of asthma (80). These studies demonstrate that certain b2AR polymorphisms
affect the clinical response to b-agonist therapy, which may impact the asthmatic pheno-
type, rendering these variants candidates that may ultimately provide for individualized
therapy in asthma.

Leukotriene Pathway Pharmacogenetics

The leukotrienes are eicosatetraenoic acid compounds that are derived from arachidonic
acid and exhibit a wide range of pharmacological and physiological actions (81). Three
enzymes are exclusively involved in the formation of the leukotrienes, including the
5-lipoxygenase (ALOX5), the leukotriene C4 synthase (LTC4), and LTA4 epoxide hydro-
lase. ALOX5 is the central enzyme required for the production of both the cysteinyl-
leukotrienes (LTC4, LTD4, and LTE4) and the potent neutrophil chemo-attractant, LTB4.
Drugs that inhibit ALOX5 activity or antagonize the action of the cysteinyl-leukotrienes
at their receptor site have been shown to attenuate broncho-constriction in asthma
patients (82).
Identification of Treatment Response Genes 9

Mutations in the ALOX5 gene were found to have significant functional conse-
quences in the context of promoter – reporter constructs and patients with variable
number of tandem repeats (VNTRs) other than the wild-type (i.e., five repeats of the
sequence 2GGGCGG2 in the core promoter) have been shown to have diminished tran-
scription of the ALOX5 gene and produce lower levels of leukotrienes (83,84).
The leukotriene LTC4 synthase is another enzyme with a known SNP in its promoter
region (A 2444C) with a C allele frequency that is reportedly higher in patients with
severe asthma (71,72), wherein the 2444C variant is associated with enhanced cysteinyl-
leukotriene production, suggesting that patients with the A/A genotype may have leuko-
triene-driven asthma. These findings provide possible evidence that, apart from the
ALOX5, there may be another pharmacogenetic locus that can modulate the leukotriene
pathway. Whether DNA sequence variants that are associated with decrease in cysteinyl-
leukotriene synthesis will also be associated with a decreased response to therapy
remains to be determined.

PHARMACOGENETIC TARGETS IN CARDIOVASCULAR DISEASE

Atherosclerosis is, at least in part, attributed to an underlying immune-mediated process

with onset early in life, ultimately leading to severe clinical manifestations, such as myo-
cardial infarction, unstable angina, and cerebral stroke. The increased incidence of cardio-
vascular events in the western societies is attributed to the underlying immune process,
which is amplified by additional cardiovascular risk factors, such as hypercholesterolemia,
hypertension, smoking, diabetes, and obesity, which by themselves have their own genetic
background. This section provides a summary of some of the key issues; for a full discus-
sion, see the chapter on cardiovascular disease.

Apolipoprotein E
Statins act primarily by inhibiting hydroxy-methylglutaryl coenzyme A (HGM-CoA)
reductase activity (85 –87). Although statins are among the most effective cholesterol-
lowering agents available, hyperlipidemic individuals display marked variability in
lipid-lowering response which may, at least in part, be due to genetic differences (88).
Among the multiple candidate genes that have been shown to be involved in lipid metab-
olism, most attention has been focused on the ApoE locus. ApoE is essential for the normal
catabolism of triglyceride-rich lipoprotein constituents. Three major ApoE isoforms are
encoded by three common alleles at the ApoE locus (89). Genetic variations in the
ApoE gene at the ApoE locus have been associated with plasma lipoprotein concentrations
in both fasting and in the postprandial states (89,90). In this regard, the E2 allele is associ-
ated with lower and the E4 allele with higher total plasma cholesterol and low-density
lipoprotein (LDL) cholesterol levels compared with the E3 allele, whereas ApoE E2 car-
riers have been reported to be more responsive to lipid-lowering statin therapy (91). The
ApoE E4 allele has been shown in some studies to be associated with increased response to
dietary intervention.

Angiotensin-Converting Enzyme
Angiotensin-converting enzyme (ACE) plays a critical role in blood pressure regulation.
Restenosis after coronary artery stent is a major health problem that is primarily attributed
to intimal hyperplasia, which can be attenuated by ACE inhibitors. Identification of patients
who are at higher risk of developing restenosis is therefore desirable. In this regard, an
10 Halapi and Hakonarson

insertion/deletion (I/D) polymorphism in intron 16 of the ACE gene has attracted signifi-
cant attention due to its correlation with serum ACE activity (92). An association was
recently reported between carriers of the D allele and reduced risk of restenosis following
coronary stenting (93,94).

MICROARRAY AND DISEASE RESPONSE GENES

Studies using commercially available whole genome or targeted arrays as well as

“in-house” spotted arrays have generated a wealth of information that importantly
contributes to our understanding of human biology in health and disease. These studies
are particularly important in addressing molecular pathways involved in organ differen-
tiation and development. More recently, their value in screening for disease susceptibility
genes and new drug targets has become evident (95 – 99). Indeed, the DNA microarray
technique has proven to play a fundamental role in high-throughput screening of the
genome. Microarrays are commonly used for global assessment of mRNA expression
levels in various tissues and cell culture systems (100 – 107) and also for polymorphism
scoring (108 – 111). The two major technology platforms for microarray analysis consists
of: (i) spotted microarrays in which pre-synthesized single- or double-stranded DNA are
bound onto glass slides and; (ii) high density oligonucleotide arrays where sets of
oligomers are synthesized directly on wafers using photolabile nucleotide chemistry
(95 –98,112). More novel application for this technology, includes the array-based com-
parative genomic hybridization (CGH) (113,114) and high-density protein microarrays
(115,116), which allow for assessment of protein– protein, protein– DNA, protein –
RNA, and protein– ligand interactions.
As the last decade has witnessed an explosion in various applications of microarray,
an attempt to cover this area thoroughly is not feasible, so the following section highlights
only a few studies aiming at identifying genes that may serve as biomarkers of treatment
response and also as possible new drug targets.

Predicting Treatment Response/Clinical Outcome

Apart from their important contribution to cancer classification (103,107), DNA micro-
arrays have been utilized to define distinct gene expression profiles that are associated
with treatment response to various anticancer drugs in diseases, such as diffuse large B-
cell lymphomas (107,117) and primary breast cancer (104 – 106).
Approximately 35% to 40% of diffuse large B-cell lymphomas respond to anthracy-
clins. By analyzing molecular signatures using cDNA arrays, Alizadeh et al. (107) demon-
strated that diffuse large B-cell lymphoma consisted of two major groups: a germinal
center B-cell-like type and an activated B-cell-like type. Patients with germinal centre
B-cell-like diffuse large B-cell lymphoma demonstrated significantly better 5-year
overall survival of 76%, compared with 16% of those with the activated form.
More recently, Rosenwald et al. (117), distinguished three different subtypes of gene
expression profiles—germinal center B-cell-like, activated B-cell-like, and a type-3
profile—in biopsies from 240 diffuse large B-cell lymphomas. Clinical and gene
expression data were then used to identify genes that predicted the outcome. A molecular
predictor consisting of 17 genes was constructed using gene expression data from 160
patients, and the predictor was validated on an independent set of 80 patients. Accord-
ingly, the gene expression profiling approach generates valuable biomarkers that allow
physicians to target patients who are most likely to benefit from conventional therapy
and focus on alternative therapy in those who do not.
Identification of Treatment Response Genes 11

In a study by van’t Veer et al. (105), the expression patterns of 98 primary tumors
from young patients with lymph node-negative breast cancer were examined, and a set of
70 genes, “classifier,” whose expression profile correlated most accurately with the short-
est interval to appearance of distant metastases (i.e., poor prognosis signature), was ident-
ified and then analyzed in a larger cohort of patients. The latter study included 295 primary
breast cancer patients (106), who were either lymph node-negative or -positive. Of those,
180 tumors were assigned the poor prognosis profile, and 115 were classified as having a
good prognosis profile. The mean overall 10-year survival rate was 54.6% and 94.5% in
the poor and favorable tumor expression profile cohorts, respectively. Interestingly,
lymph node involvement did not correlate with the expression profile, whereas patient’s
age, ER status, and histological grade were associated with specific expression profile
patterns, with younger age and higher histological grade being more prevalent in
the poor prognosis group, whereas positive ER status was indicative of a favorable prog-
nosis profile.
In a study by Sotiriou et al. (118), fine-needle aspirates from breast tumors were used
to investigate whether sufficient RNA for microarray analysis could be obtained and
whether specific gene expression profiles could be used to distinguish between patients
with differential response to chemotherapy. Although a small set of patients were ana-
lyzed, the authors identified a set of 37 genes that distinguished response status from
lack of response in pretreatment samples. As drug resistance to chemotherapy poses a
major problem, establishment of predictive markers for drug response are of great
value. The use of neoadjuvant treatment of advanced breast cancer prior to operation is
growing, and biomarkers of response would facilitate the selection of patients who are
eligible for such treatment. Many breast cancer patients receive unnecessary treatment
for possible tumor spread after the removal of the primary tumor. Molecular profiling
would offer more accurate predictions of who may need such treatment.

Discovery of Novel Drug Targetable Genes

Several studies have made an attempt to associate glucocorticoid (GC) resistance to
known polymorphic variations in genes that constitute the GC response pathway (119 –
123). Although both structural and functional alterations in the glucocorticoid receptor
units or their response elements are important determinants of glucocorticoid responsive-
ness, no relevant clinical prediction has emerged from these studies. In a recent study
using microarray to examine gene expression profiles in peripheral blood cells (PBM)
cells obtained from asthmatic patients who were either glucocorticoid therapy responders
or nonresponders, glucocorticoid responders could be separated from nonresponders with
more than 85% accuracy using only a few genes (124). The glucocorticoid-resistant
patients were also clustered into families and examined for linkage (125).
A set of genes unique for psoriatic skin disease was recently reported in a study using
DNA microarray. By comparing lesional and uninvolved skin from patients with psoriasis,
a set of 159 genes that showed twofold or greater difference in their expression level was
defined. Of these, several were mapped to disease-associated loci (126). The gene set pre-
dicted expression patterns unique to normal versus lesional skin with 100% accuracy, and a
subset of these genes was also useful for monitoring treatment response (126).
Microarray has also been applied in combination with genetic linkage studies to
dissect out disease susceptibility genes in experimental models. A recent study using quan-
titative trait loci (QTL) analysis in a mouse model of experimental allergic asthma
revealed two distinct loci on chromosome 2 that controlled allergen-induced airway hyper-
responsiveness (AHR), abhr1 and abhr2 (127). In a study comparing expression profiles in
12 Halapi and Hakonarson

RNA from lung obtained from susceptible (A/J), resistant (C3H/HeJ), and backcross
mice with various phenotypes, Karp et al. (128) identified 21 differently expressed
genes, including complement factor 5 (C5) that mapped to either locus. Expression
levels of C5 were significantly higher in AHR-resistant mice than in AHR-
susceptible mice. Sequence analysis of the C5 gene in the parental strain revealed a
2 bp deletion in the 50 untranlated region of A/J mice, resulting in reduced C5 mRNA
level and lack of functional C5 protein (128). In a backcross progeny, more extreme
AHR was seen in mice that were homozygotes for the susceptible A/J-derived C5
allele, compared with the low responders that were heterozygotes. The authors concluded
that C5 deficiency might interfere with monocyte/macrophage production of interleukin-
12, thereby altering immunoregulatory mechanisms that determine susceptibility to
asthma (128).
Linkage analysis in combination with microarray expression analysis has also been
applied to identify candidate genes in cardiovascular disease. In this regard, a QTL regu-
lating high-density lipoprotein cholesterol (HDL-C) phenotype, was localized in baboons
at chromosome 18 (129). An array consisting of genes from the chromosomal region was
created by assembling a contig of bacterial artificial chromosome (BAC) clones for the
region of interest. This chromosomal region is highly conserved between species allowing
human BACs to be used to assemble the contig (129). Liver cDNA from sibling baboons
of contrasting HDL1-C phenotypes that were exposed to different diets was used to screen
for differently expressed genes. The authors identified 53 differentially expressed genes of
keen interest (129).
In a recent study, Lock et al. (130) selected genes that were expressed differently in
acute and chronic lesions obtained from autopsy samples of patients with multiple
sclerosis (MS). The genes were used as targets for therapy in experimental autoimmune
encephalitis (EAE), an experimental model for MS. Of note, the granulocyte colony-
stimulating factor (GM-CSF) was overexpressed in acute plaques but not in chronic
plaques, whereas the Fcg receptor was upregulated in chronic plaques and not increased
in acute plaques. Administration of GM-CSF prior to induction of EAE halted the onset
and lowered the disease score compared with the control. EAE was also ameliorated in
FcgR knockout mice compared with the wild-type mice (130).

CONCLUSION

The powerful combination of genetic linkage and microarray expression profiling presents
an integrated approach that is likely to facilitate discovery of genes involved in common
complex disease processes and the regulation of therapeutic response to drugs, thereby
facilitating the development of new drugs, DNA-based disease diagnostic products, and
pharmacogenomic tests to the market. Such tests will play an important role in delivering
more personalized medicine, contributing to the development of more effective means of
diagnosing and treating disease by matching each patient and the most suitable drug.
Accordingly, the new scope of genetics is charged with the promises to transform the prac-
tice of medicine by enabling physicians to assess the risks of disease, permit early detec-
tion of disease, determine likely responses to medication, choose the best courses of
therapy, and have at their disposal new therapies that target the disease process itself.
This chapter has provided a general introduction to the subject and given some key
examples of what is currently known about the potential value of pharmacogenetic infor-
mation and the potential use of such information in the near future. The following chapters
in this book consider both general issues and disease-specific pharmacogenetics in detail.
Identification of Treatment Response Genes 13

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