Clinical Implications of Dihydropyrimidine Dehydrogenase on 5-FU Pharmacology

Introduction

Dihydropyrimidine dehydrogenase (also known as DPD, dihydrouracil dehydrogenase, dihydrothymine dehydrogenase, uracil reductase, EC 1.3,1.2) is the initial rate-limiting enzymatic step in the catabolism of not only the naturally occurring pyrimidines, uracil and thymine, but also the widely used antimetabolite cancer chemotherapy drug, 5-fluorouracil (5-FU).[1,2] As shown in Figure 1, DPD occupies an important position in the overall metabolism of 5-FU, converting over 85% of a standard dose of administered 5-FU to dihydrofluorouracil (5-FUH₂), an inactive metabolite, in an enzymatic step that is essentially irreversible.[3]

It is true that anabolism is clearly critical for 5-FU cytotoxic action through conversion of 5-FU to the "active" nucleotides 5-fluorodeoxyuridine monophosphate (FdUMP), 5-fluorouridine triphosphate (FUTP), and 5-fluorodeoxyuridine triphosphate (FdUTP). These important active metabolites are, in turn, responsible for inhibition of cell replication through inhibition of thymidylate synthase, or through incorporation into RNA or DNA, respectively. Nevertheless, it is pyrimidine catabolism through the rate-limiting step, DPD, that controls the availability of 5-FU for anabolism and thus occupies a critical position in the overall metabolism of 5-FU.

Evidence of Importance of DPD Activity to 5-FU Pharmacology

DPD—as the rate-limiting step in pyrimidine catabolism—has recently been shown to play a critical role in determining the clinical pharmacology of 5-FU (Table 1). In particular, it has been demonstrated that DPD accounts for much of the variability that has been noted in clinical studies with 5-FU. This includes both intrapatient variability, as well as interpatient variability.

DPD activity has been observed to follow a circadian pattern in both animals and humans.[4-6] Studies in rats on a 12-hour light/12-hour dark schedule have demonstrated that hepatic levels of DPD follow a pattern that can be plotted on a cosine wave.[4] This pattern was completely reversed in another group of rats on an inverted 12-hour light/12-hour dark schedule. In patients receiving continuous-infusion 5-FU by automated pumps, sampling DPD activity in peripheral blood mononuclear cells over a 24-hour period has also been shown to exhibit circadian patterns when plotted on a cosine wave.[6] Serum samples obtained at the same time from the same patients have been shown to have serum 5-FU concentrations that were also characterized by a circadian pattern, which was essentially inverse to the DPD circadian pattern (Figure 2).

Data from this study suggested that perhaps DPD was responsible for the circadian variation in 5-FU, leading some chemotherapists to propose time-modified 5-FU infusions to optimize drug delivery during a 24-hour period. Such regimens have been suggested by some oncologists, particularly those in Western Europe, to have a potential benefit in the treatment of certain human cancers.[7]

What Causes the Variation?

For the past 4 decades, it has been unclear as to why the pharmacokinetics of 5-FU have been so variable, with half lives (t_1/2) ranging from around 4 minutes to 25 minutes after an intravenous bolus (Table 2).[8] Because of the critical position of DPD in the metabolic pathway, it was hypothesized that the variation in pharmacokinetic characteristics might be secondary to variability of DPD enzyme activity among different individuals. Population studies were undertaken assessing DPD enzyme activity initially in tissues (peripheral blood mononuclear cells and liver) from healthy individuals. DPD was shown to vary from individual to individual, with a normal distribution pattern (bell-shaped curve) showing a six-fold variation from the lowest to the highest values (Figure 3).[9,10]

Essentially the same pattern of DPD activity has also been observed in the peripheral blood mononuclear cells of both breast and colorectal cancer patients, although, interestingly, the normal distribution is shifted to the left, with lower median DPD activity in the breast cancer patient population.[11] The mechanism for this latter observation is unknown. The wide variation in DPD activity observed in the populations described above is thought to account for the wide variation in the half-life (and pharmacokinetics) observed in patients treated with 5-FU.[8]

Variation in DPD Activity

While most patients tolerate 5-FU reasonably well, over the past 4 decades, a number of patients have developed severe, at times life-threatening, toxicity after standard doses of 5-FU.[12-15] Because these patients demonstrated exaggerated normal toxic side effects (as if they had received an overdose of drug), it was hypothesized that these patients were deficient in a catabolic enzyme that resulted in more 5-FU being present over time.

Initial studies in these affected patients demonstrated elevated uracil and thymine levels, suggesting a deficiency in the first step in pyrimidine catabolism, DPD.[12-14] Subsequent studies demonstrated that many of these patients were indeed DPD deficient. It is now clear that an additional small percentage (< 3%) of such patients have DPD activity that is significantly below the normal distribution seen in the average person.[15] These individuals are at significant risk if they develop cancer and are given 5-FU. This is a true pharmacogenetic syndrome, with symptoms being unrecognizable until exposure to the drug.[15]

Variation in DPD activity has also recently been shown to be responsible for the apparent variable bioavailability of 5-FU, which over the years has led to the recommendation that 5-FU not be administered by the oral route. An explanation for the erratic bioavailability of 5-FU has previously been unclear, particularly since 5-FU is a small molecule with a pK_a that should favor excellent absorption and bioavailability. Recent studies using DPD inhibitors in animals have demonstrated that, following inhibition of DPD, the pharmacokinetic pattern resulting from oral administration of 5-FU is essentially the same as that produced by intravenous administration, resulting in essentially 100% bioavailability.[16]

While it has long been well known that variability in the activity of pyrimidine anabolic enzymes is important in determining the antitumor effectiveness of 5-FU, much less attention has focused on the variability in activity of the pyrimidine catabolic enzymes. Tumors that are resistant to 5-FU have been shown to express increased levels of DPD activity.[17] With the development of the quantitative polymerase chain reaction (PCR) to measure DPD mRNA, increased expression of DPD mRNA has been demonstrated in tumors of patients who were resistant to 5-FU.[18]

Developing a Pharmacologic Strategy Based on DPD

The variability in DPD levels in both normal and tumor tissues provides a basis for the chemotherapist to consider either altering the dose of the fluoropyrimidine drug or inhibiting DPD in order to eradicate the variability in 5-FU pharmacology. The presence of increased tumor DPD could lead to a decision to increase the dose of 5-FU (or a 5-FU prodrug) to overcome the increased catabolism of 5-FU within the tumor, thus increasing the amount of 5-FU that could be converted to active anabolites (Figure 1).

An alternative approach is to use known inhibitors of DPD, usually with a lower dose of 5-FU (or the use of a 5-FU prodrug; eg, capecitabine(Drug information on capecitabine) [Xeloda]), to directly inhibit 5-FU degradation within the tumor, thereby permitting the existing 5-FU (even if present in low concentrations) to be converted to active anabolites. Inhibiting DPD in 5-FU-susceptible host tissue, such as gastrointestinal mucosa and bone marrow, should also make dosing from patient to patient less variable, the latter being accomplished through lessening the variability in pharmacokinetics, bioavailability, and the resultant host toxicity.