It is readily accepted that genetic factors play an important role in influencing a child’s potential physical characteristics such as height, weight, or hair color. Genetic factors are also important (although not sole) determinants of inter- and intraindividual variability in susceptibility to pediatric diseases as well as in the disposition of and response to medications used to treat those diseases. Pharmacotherapy in adults has benefited from knowledge of pharmacogenetic principles acquired over the past 50 to 60 years, but application to pediatric therapeutics is still in its infancy. On April 14, 2003, the International Human Genome Sequencing Consortium announced successful completion of the Human Genome Project initiated in 1990, 2 years after being presented in draft form (1). There is considerable hope that morbidity and mortality will be decreased through the development of more effective strategies to diagnose, treat, and prevent human disease, and society has every right to expect that children and adults will benefit equally from this investment of public funds. However, children present unique challenges in this context since developmental changes in drug disposition and response are superimposed upon a basal level of pharmacogenetic variability. The purpose of this chapter is to introduce the concepts of pharmacogenetics, pharmacogenomics, and pharmacoproteomics (and other “-omic” fields of endeavor spawned in the genome era) in the context of the changes in growth and development characteristic of the pediatric population.

In 1841, Alexander Ure reported that hippuric acid was formed from benzoic acid in the body leading physiological chemists to discover that many foreign substances excreted by humans were chemically altered relative to the forms that had been administered—the process we now refer to as drug biotransformation (or less properly, drug metabolism). At the beginning of the 20th century, Archibald Garrod proposed that enzymes were implicated in the detoxification of foreign substances. A key element of his later work was the concept that disproportionate responses to foreign substances could result from deficiency of the required detoxifying enzyme. A variation in the theme of altered responses to foreign substances (xenobiotics) became apparent with the synthesis of phenylthiocarbamide in 1931 when, in the process searching out artificial sweeteners, A. L. Fox discovered that some people found the chemical intensely bitter whereas others found it tasteless (2). It was not until the 1950s, however, that certain adverse drug reactions, such as unusually prolonged respiratory muscle paralysis due to succinylcholine, hemolysis associated with antimalarial therapy, and isoniazid-induced neurotoxicity, were recognized to be a consequence of inherited variation in enzyme activities as reviewed by Arno Motulsky in 1957 (3).

In 1959, Fridriech Vogel coined the term pharmacogenetics to describe the study of genetically determined variations in drug response and the first book on the subject was published in 1962 by Werner Kalow (4). Through a series of twin studies conducted during the late 1960s and early 1970s, Elliott Vesell illustrated the importance of genetic variation in drug disposition by observing that the half-lives of several drugs were more similar in monozygotic twins than in dizygotic twins (5). With the discovery of the debrisoquine/sparteine hydroxylase polymorphism (6,7), due to inherited defects in the cytochrome P450 2D6 gene (CYP2D6), and mephenytoin hydroxylase deficiency (CYP2C19) (8) in the late 1970s and early 1980s, the importance of genetic polymorphisms in drug-metabolizing enzymes has become increasingly apparent. This is particularly true in recent years due to an enhanced awareness of the number of clinically useful drugs that are
metabolized by polymorphically expressed enzymes and the proportion of treated patients who are affected.

In the 1970s, an increasing appreciation of genetic influences on variability in drug disposition and response was accompanied by heightened awareness that environmental factors (e.g., diet, smoking status, and concomitant drug or toxicant exposure), physiologic variables (e.g., age, gender, disease, and pregnancy), and patient compliance also played important roles. Advances in analytic tools to accurately measure drugs and drug metabolites in biological fluids and the development of mathematical models to characterize and predict changes in drug concentration over time (pharmacokinetics) led to the application of pharmacokinetic principles to optimize drug therapy in individual patients. Introduction of therapeutic drug monitoring programs was the first application of personalized medicine—recognition that all patients were unique and that utilizing serum concentration–time data for an individual patient theoretically could be used to optimize pharmacotherapy was a significant advance over the concept of “one dose fits all.” However, routine therapeutic drug monitoring does not necessarily translate to improved patient outcome in all situations (9).

At a molecular level, the pharmacokinetic properties of a drug are determined by the genes that control its disposition in the body (e.g., absorption, distribution, metabolism, and excretion) with drug-metabolizing enzymes and drug transporters assuming particularly important roles. Over the past 25 years, the functional consequences of genetic variation in several drug-metabolizing enzymes have been described in subjects representative of different ethnic groups (10). Whereas the most common clinical
manifestation of pharmacogenetic variability in drug biotransformation is an increased risk of concentration-dependent toxicity due to reduced clearance and drug accumulation, it has become more apparent in recent years that the concentration–effect relationship (pharmacodynamics) is more relevant for optimizing drug efficacy. Therefore, the pharmacogenetics of drug receptors and other target proteins involved in signal transduction or disease pathogenesis can also be expected to contribute significantly to interindividual variability in drug response (11,12). However, the most important concept is that the pharmacogenetic determinants of drug response involve multiple genes and therefore, for a particular individual, polymorphisms in a single gene are unlikely to be predictive of response.

In 1987, the term “genomics” was introduced to describe the study of the structure and function of the entire complement of genetic material—the genome—including chromosomes, genes, and DNA (13). In 1990, the Human Genome Project was initiated as a $3 billion public investment with the goal of sequencing the entire complement of human genes by the year 2005 but more importantly, with the expectation that decreased morbidity and mortality through the development of more effective strategies to diagnose, treat, and prevent human disease would be the return on that investment (Fig. 5.1). The publicly funded initiative was forced to accelerate its efforts when J. Craig Venter announced that his company, Celera Genomics, would sequence the human genome first. The two efforts resulted in simultaneous publication of initial draft sequences in 2001 (1,14), and the International Human Genome Sequencing Consortium announced completion of the task on April 15, 2003 with an estimated accuracy of one error in 100,000 bases (15). The genome consists of 3 gigabases (three billion bases) of DNA sequence that code for approximately 30,000 genes, far fewer than was originally expected. However, it appears that this number of genes encodes 100,000 proteins through the process of alternative splicing whereby a gene’s exons or coding regions are spliced together in different ways to produce variant mRNA molecules that are translated into different proteins or isoforms of the same protein. Thus, the vast amounts of genomic data generated by the Human Genome Project have laid the foundation for an “-omic” revolution that includes, but is not limited to, the transcriptome, the set of expressed genes from a genome (16,17), the proteome, the set of proteins encoded by the genome (18), and the physiome, in which biochemical, biophysical, and anatomic information from cells, tissues, and organs will be integrated using computational methods to provide a model of the human body (19). Metabolomics and metabonomics are related terms that are sometimes used interchangeably in the literature. Metabolomics refers to the complete set of low–molecular-weight molecules (metabolites) present in a living system (cell, tissue, organ, or organism) at a particular developmental or pathological state. Metabonomics has been defined as the study of how the metabolic profile of biological systems changes in response to perturbations due to pathophysiologic stimuli, toxic exposures, dietary changes, among others (20,21). Pharmacometabonomics has been defined as “the prediction of the outcome, efficacy or toxicity, of a drug or xenobiotic intervention in an individual based on a mathematical model of preintervention metabolite signatures” (22), Chemogenomics is the application of combinatorial chemistry to generate libraries of small molecular weight compounds that can serve both as probes to investigate biological mechanisms and as lead compounds for drug development (23). Several resources that define these fields and their potential application to human health and disease are available on the Internet (Table 5.1).

Genetic variability results from gene mutation and the exchange of genetic information between chromosomes that occurs during meiosis. With the exception of sex-linked genes (genes occurring on the X or Y chromosomes), every individual carries two copies of each gene he or she
possesses. All copies of a specific gene present within a population may not have identical nucleotide sequences and these genetic polymorphisms contribute to the variability observed in that population. The presence of different nucleotides at a given position within a gene is called a single nucleotide polymorphism or “SNP” and SNPs are rapidly becoming an important component of the genomics lexicon. More recently, focus has shifted to characterizing haplotypes, collections of SNPs, and other allelic variations that are located close to each other and inherited together; creating a catalogue of haplotypes or “HapMap” is also a goal of the Human Genome Project (24). In genes where polymorphisms have been detected, alternative forms of the gene are called alleles. When the alleles at a particular gene locus are identical, a homozygous state exists whereas the term “heterozygous” refers to the situation in which different alleles are present at the same gene locus. The term genotype refers to an individual’s genetic constitution while the observable characteristics or physical manifestations constitute the phenotype, which is the net consequence of genetic and environmental effects.

Human genetic variation can take many forms but is broadly divided into two general classes of variation: single nucleotide variations and structural variations. SNPs are the most prevalent class of genetic variation, and it has been estimated that there are approximately 11 million SNPs in the human genome. Structural variations encompass all differences in DNA sequence that involve more than one nucleotide. Insertions–deletions variants (indels) occur when a contiguous set of one or more nucleotides is absent in some individuals and present in others. Block substitutions involve variation of a string of contiguous nucleotides (or “block”) that differs between two genomes. Sequence inversions occur when the order of an entire block of nucleotides is reversed in a specific region of the genome. Finally, copy number variations (CNVs) refer to the deletion or duplication of identical or near-identical DNA sequences that may be thousands to millions of bases in size. Although they occur less frequently than SNPs, structural variations may constitute 0.5% to 1% of an individual’s genome and thus are the subject of intensive investigation for their contribution to phenotypic variation (25,26).

Pharmacogenetics, the study of the role of genetic factors in drug disposition, response, and toxicity, essentially relates allelic variation in human genes to variability in drug responses at the level of the individual patient. In other words, the promise of pharmacogenetics is to identify the right drug for the right patient. The field of pharmacogenetics classically has focused on the phenotypic consequences of allelic variation in single genes but often in the past, there was confusion between genotypic and phenotypic definitions of “polymorphism,” and thus a need to clarify the relationship between genetic concepts and the clinical relevance of a given phenotype. In 1991, Meyer proposed that pharmacogenetic polymorphism be defined as a monogenic trait caused by the presence in the same population of more than one allele at the same locus and more than one phenotype in regard to drug interaction with the organism. The frequency of the least common allele should be at least 1% (27). According to this definition, the key elements of pharmacogenetic polymorphisms are heritability, the involvement of a single gene locus, and the fact that distinct phenotypes are observed within the population only after drug challenge.

The vast majority of our current understanding of pharmacogenetic polymorphisms involves enzymes responsible for drug biotransformation. Clinically, individuals are classified as being “fast,” “rapid” or “extensive” metabolizers at one end of the spectrum, and “slow” or “poor” metabolizers at the other end of a continuum that may, depending on the particular enzyme, also include an intermediate metabolizer group. Pediatric pharmacogenetics involves an added measure of complexity since fetuses and newborns may be phenotypically “slow” or “poor” metabolizers for certain drug-metabolizing pathways, acquiring a phenotype consistent with their genotype at some point later in the developmental process as those pathways mature [e.g., glucuronidation, some cytochrome P450 activities (28,29,30)].

Although some authors use the terms “pharmacogenetics” and “pharmacogenomics” interchangeably, the latter term represents the marriage of pharmacology with genomics and is therefore considerably broader in scope. Pharmacogenomics can be defined as the study of the genome-wide response to small molecular weight compounds administered with therapeutic intent—finding the right drug for the right disease. Proteomics represents the systematic investigation of qualitative and quantitative changes in protein expression in a cell or tissue in response to disease or disease treatment. In this context, pharmacoproteomics involves characterizing the response of the proteome to therapeutic agents. Similarly, toxicogenomics and toxicoproteomics investigate the analogous response to environmental contaminants and other toxicants (31,32). In contrast to the focus of “pharmacogenetics” on single gene events, “pharmacogenomics” involves understanding how interacting systems or networks of genes influence drug responses (33). This definition is particularly appealing to pediatric health and disease since the concept of many genes acting in concert captures the essence of the developmental processes that characterize maturation from the time of birth through adulthood while retaining a focus on the individual.

It is safe to say that application of pharmacogenomic principles to pediatric medicine has received far less attention than its application to diseases affecting adults, and the scope of the field remains to be completely defined. However, developmental and pediatric pharmacogenomics necessarily must take into consideration the dynamic changes in gene expression that accompany maturation from embryonic life through fetal development, the neonatal period, infancy, childhood, and adolescence, for example, during organogenesis, as receptor systems and neural networks become established, and functional drug biotransformation capacity is acquired, among others. In other words, patterns of gene expression and the nature of the gene interactions that contribute to the pathogenesis of pediatric diseases (and thereby serve as potential targets for pharmacologic intervention) may only be discernable or relevant at specific critical points in the developmental continuum. Furthermore, variability in drug disposition (i.e., pharmacokinetics) and action (i.e., pharmacodynamics) that
ultimately impact drug response in pediatric patients can also be expected to change as children grow and develop. Finally, developmental and pediatric pharmacogenomic investigations can be distinguished from similar studies conducted in adults by the fact that drug or toxicant exposure at critical points in development may disrupt or alter the normal patterns of development—a genome-wide response to drug/toxicant exposure. This may have immediate, observable consequences, for example, fetal demise or major structural abnormalities such as those associated with retinoids (34) or other human teratogens. Of equal concern, however, is the possibility that drug exposure, or lack of effective drug treatment (35), may have unintended consequences on cognitive or behavioral development that do not manifest until much later in maturation. The remainder of this chapter will highlight examples of how pharmacogenomic approaches are currently improving pediatric pharmacotherapy and present several opportunities for future application.

Completion of the Human Genome Project was facilitated by several technological advances; the demands of genomic, proteomic, pharmacogenetic, and pharmacogenomic analyses have driven the development of an industry dedicated to the discovery and refinement of technologies capable of generating large datasets of information derived from DNA, RNA, proteins, and small endogenous molecules. These tools are used widely to investigate disease pathogenesis but are equally applicable to investigations of variability in drug disposition and response.