James W. Watters

Cancer pharmacogenomics: current and future applications.

Heterogeneity in patient response to chemotherapy is consistently observed across patient populations. Pharmacogenomics is the study of inherited differences in interindividual drug disposition and effects, with the goal of selecting the optimal drug therapy and dosage for each patient. Pharmacogenomics is especially important for oncology, as severe systemic toxicity and unpredictable efficacy are hallmarks of cancer therapies. In addition, genetic polymorphisms in drug metabolizing enzymes and other molecules are responsible for much of the interindividual differences in the efficacy and toxicity of many chemotherapy agents. This review will discuss clinically relevant examples of gene polymorphisms that influence the outcome of cancer therapy, and whole-genome expression studies using microarray technology that have shown tremendous potential for benefiting cancer pharmacogenomics. The power and utility of the mouse as an experimental system for pharmacogenomic discovery will also be discussed in the context of cancer therapy.

Irinotecan Pharmacogenetics: Is It Time to Intervene?

The development of unpredictable systemic toxicity while attempting to achieve tumor response has been a common observation for as long as there has been anticancer therapy. The narrow therapeutic index of most chemotherapeutic agents and the severe consequences of both undertreatment and overdosing have led to a pressing need for molecular predictors of the toxicity and efficacy of cancer treatments. In the days when there were few options for the treatment of most human malignancies, it was acceptable to tolerate grade 4 toxicities in as many as 10% of patients in the search for optimal dose-intensity. Better that a patient experience side effects than the tumor be undertreated. However, much progress has been made in treating human malignancies, and there are now multiple treatment options with similar efficacy for nearly every type of cancer. Ideally, patients should receive a regimen that offers optimal efficacy with minimal chance of severe side effects. This can only be accomplished if there are means to determine the relative risks of both patient benefit and toxicity. In the context of regimens demonstrating similar efficacy in large populations of patients, but showing no individual predictive markers of benefit, toxicity prediction would be a meaningful way to achieve the goal of better therapy for individual patients. Irinotecan is an excellent candidate for individualized therapy. The drug has demonstrated potent activity against many types of human cancer, in particular, gastrointestinal and pulmonary malignancies. Indeed, the addition of irinotecan to first-line therapy with fluorouracil and leucovorin has led to improved survival in patients with advanced colorectal cancer. However, irinotecan does have significant side effects, including both acute and delayed diarrhea, neutropenia, and a vascular syndrome. The gastrointestinal and vascular syndromes have been associated with a high mortality rate in patients receiving the combination of irinotecan with bolus fluorouracil and leucovorin during the first 60 days of therapy. There are now combination therapy regimens with equal or superior efficacy, such as infusional fluorouracil and leucovorin with irinotecan or oxaliplatin. Hence the conundrum: can we identify patients who will receive antitumor benefit from irinotecan without experiencing severe, life-threatening, or fatal toxicity? Irinotecan is among the camptothecin class of topoisomerase 1 inhibitors. It is a prodrug and must be converted to SN-38 by carboxylesterase 2, resulting in a greater than 1,000-fold enhancement of cytotoxic activity. Before activation, irinotecan must run a disposition gauntlet of oxidation by cytochrome p450 enzymes, and transport by adenosine triphosphate– binding cassette efflux pumps. SN-38 must also undergo one of three fates: binding to the cellular target topoisomerase 1, exclusion from the cell via efflux pumps, or inactivation by the addition of a glucuronide moiety. While each of these steps has the potential to substantially regulate irinotecan activity, it is glucuronidation by the protein UGT1A1 that has the clearest potential impact on patient care. The glucuronidation of lipophilic compounds is catalyzed in vertebrates by the uradine diphosphate– glucuronosyltransferases (UGTs). This catalytic reaction utilizes UDP-glucuronic acid as a cosubstrate for the formation of glucuronides from various substrates, such as bilirubin, hormones, drugs, and other xenobiotics. This reaction leads to the formation of hydrophilic glucuronides from lipophilic substrates, facilitating the transport of these molecules to aqueous compartments of the body, and leading to elimination through the bile and urine. UGT glucuronidation is consequently regarded as a “detoxification” reaction, and UGTs therefore represent major phase II drug metabolizing enzymes. More than 16 human UGTs have been characterized and divided into two families—UGT1 and UGT2. All known family-1 members are encoded by the UGT1A locus JOURNAL OF CLINICAL ONCOLOGY E D I T O R I A L VOLUME 22 NUMBER 8 APRIL 15 2004

Pharmacogenomics: The Influence of Genomic Variation on Drug Response

Unpredictable efficacy and toxicity are major hurdles in the administration of many medications. By identifying inherited DNA polymorphisms that influence drug disposition and effects, pharmacogenomics is an exciting tool for the individualization of drug therapies. Single nucleotide polymorphisms (SNP) in genes encoding drug metabolizing enzymes, drug transporters, and DNA repair genes have recently been shown to influence drug toxicity and efficacy. This review will discuss clinically relevant examples of genetic polymorphisms that influence the outcome of drug therapy, and possibilities for future applications of pharmacogenomics.

Using genome-wide mapping in the mouse to identify genes that influence drug response

Differential drug response is most often likely to be a complex trait, controlled by the combined influences of multiple genes and environmental influences. As a result of theoretical and technical limitations, to date, most clinically useful pharmacogenomic studies in humans have been limited to a small number of candidate genes that have a relatively major impact on drug response. Here, the problems involved in identifying genes that underlie drug response in humans are discussed and the power of mouse genetics as a tool for pharmacogenomic discovery is highlighted.

Murine pharmacogenomics: using the mouse to understand the genetics of drug therapy

Pharmacogenomics seeks to understand the genetic basis of interindividual differences in drug disposition and effects. Differential drug response is likely to most often be a complex trait, in which multiple genes contribute with varying strengths to the therapeutic phenotype. Due to technical and economic limitations, pharmacogenomic studies in humans are mainly limited to a small number of candidate genes with relatively major influences on drug response. This review discusses the problems involved in mapping genes underlying drug response in humans and highlights the theoretical and applied uses of mouse genetics to address these important issues.

Analysis of variation in mouse TPMT genotype, expression and activity

Although the mouse has great potential for pharmacogenomic discovery, little is known about variation in drug response or genetic variation in pharmacologically relevant genes between inbred mouse strains. We therefore assessed variation in gene sequence, mRNA expression and protein activity of thiopurine methyltransferase (TPMT) in multiple inbred mouse strains. TPMT activity was measured by high-performance liquid chromatography detection of 6-MMP produced by incubation of liver homogenates with 6-MP. Genetic variation was assessed by resequencing and single nucleotide polymorphism (SNP) genotyping using pyrosequencing technology. mRNA expression was measured by real-time polymerase chain reaction. We observed an almost five-fold variation in TPMT activity, with strains falling into distinct low and high activity groups. This pattern of TPMT activity was highly correlated with expression of TPMT mRNA among strains, and high TPMT expression is dominant in F1 hybrids. To correlate genotype with phenotype, 29 SNPs and one insertion/deletion were genotyped throughout the TPMT gene and upstream 10 kb. Only two haplotypes were observed across all 30 polymorphisms, corresponding to the low and high activity groups. These results suggest that differential mouse TPMT activity is due to variation in mRNA expression. In addition, the identified pattern of low haplotype diversity suggests that the mouse is likely to be useful for pharmacogenomic discovery by associating haplotype blocks with drug response phenotypes among inbred strains.

Candidate gene selection in pharmacogenomics: biology leads the way

James W Watters & Howard L McLeod† †Author for correspondence Departments of Medicine, Genetics, and Molecular Biology and Pharmacology, Washington University School of Medicine and the Siteman Cancer Center, St Louis, MO, USA †Washington University School of Medicine, Department of Medicine, 660 South Euclid Ave, Campus Box 8069, St. Louis, MO 63110-1093, USA Tel: +1 (0)314 747 2060; Fax: +1 (0)314 362 3764; E-mail: hmcleod@ im.wustl.edu ‘There is a need for a more informed way to select candidate genes, so that only those candidates with the highest chance of showing association are analyzed in precious patient samples’