Introduction

Until irinotecan (Camptosar) became
available, 5-fluorouracil (5-FU) and 5-fluoro-2'-deoxy-uridine (FUDR) comprised
the only commercially available treatment options that could be considered
standard of care for advanced colorectal cancer. An understanding of the
intracellular metabolism of 5-FU and its mechanism of action has suggested
potential therapeutic strategies to improve its clinical activity. A variety of
approaches to modulate 5-FU in patients with colorectal cancer have been tested
in randomized studies over the past 15 years.

Leucovorin Modulation of 5-FU

Inhibition of thymidylate synthase by the 5-FU metabolite
5-fluoro-2'-deoxyuridine monophosphate (FdUMP) is one of the principal
mechanisms of 5-FU action.[1] Thymidylate synthase is an important therapeutic
target because it catalyzes the de novo formation of thymidylate (dTMP) from
deoxyuridylate (dUMP). During this process, a methyl group is transferred from
the reduced folate cofactor 5,10-methylenetetrahydrofolate to the carbon-5
position of the uracil moiety. Thymidylate is subsequently converted to
thymidine triphosphate, which is required for DNA synthesis and repair. In
contrast to the interaction of thymidylate synthase with its physiological
substrate, the TS-FdUMP-folate complex is only slowly reversible.

The stability of the ternary complex is influenced by the
intracellular level of 5,10-methylenetetrahydrofolate and the extent of its
polyglutamation. Pharmacologic concentrations of 5-formyltetrahydrofolate
(leucovorin) expand the intracellular pools of 5,10-methylenetetrahydrofolate,
thereby increasing the extent and duration of 5-FU-mediated thymidylate synthase
inhibition. This strategy for biochemical modulation of 5-FU by leucovorin has
proven successful in the clinical setting. The two most commonly employed
5-FU/leucovorin regimens involve either weekly administration of IV bolus 5-FU
delivered at the midpoint of a 2-hour IV infusion of high-dose leucovorin, or a
monthly (daily for 5 days every 4-5 weeks) schedule of IV bolus leucovorin and
5-FU.

Intravenous Bolus Administration

Published results are currently available from 14 randomized
trials that compared 5-FU administered by IV bolus either as a single agent or
modulated by leucovorin.[2-16] The response rates are based on the number of
eligible and assessable patients reported by each investigator, and therefore
are somewhat higher than they would be if all patients randomized to that arm
were included in the denominator. Two trials included patients with
nonmeasurable disease.[8,12] The combined data for all 14 trials indicate a
response rate of 25.3% ± 2.4% (95% confidence interval) among 1,262 patients
treated with 5-FU/leucovorin compared to 13.8% ± 2.2% among 995 patients
treated with 5-FU alone. For time to progression, the median of the reported
values was 6.0 months with 5-FU/leucovorin and 4.2 months for 5-FU alone (Figure
1). The median of the reported survival times was 12.4 months for
5-FU/leucovorin and 11.0 months for 5-FU alone.

The original meta-analysis that assessed the value of bolus
5-FU/leucovorin in patients with advanced colorectal cancer employed individual
data provided by the principal investigators from nine randomized trials.[6] All
1,381 patients entered in these trials were included on an intent-to-treat
basis. There was no significant difference in median survival (11.5 vs 11.0
months, odds ratio 0.97).

Finding the Optimal Schedule

A second generation of trials sought to establish the optimal
5-FU/LV schedule. A randomized trial conducted by the North Central Cancer
Treatment Group (NCCTG) compared a monthly schedule of low-dose
leucovorin-modulated 5-FU with a weekly schedule of high-dose
leucovorin-modulated 5-FU.[17] In this trial, the overall response rate among
102 assessable patients on the monthly schedule was 34% and among 98 patients on
the weekly schedule, it was 31%. Both the median time to tumor progression (~
4.4 and ~ 6.3 months) and median survival (9.3 and 10.7 months) were somewhat
longer with the weekly schedule, but these differences were not statistically
significant. If the response rates for the monthly and weekly schedules for 5-FU
alone or with leucovorin from the 14 randomized studies cited above are compared
side by side, the results are virtually identical (Figure
2). Thus, these two
schedules of 5-FU/leucovorin can be considered therapeutically equivalent in
advanced colorectal cancer, although the spectrum of toxicity differs.

Dose-limiting diarrhea occurred more frequently with the weekly
schedule, while severe mucositis was uncommon. In contrast, mucositis tended to
be dose-limiting with the monthly schedule; however, severe diarrhea also
occurred. The monthly schedule was considered the "gold-standard" for
pivotal trials in colorectal cancer in the United States since an NCCTG study
showed a survival advantage with leucovorin-modulated 5-FU.[12] Furthermore,
because of the cost of leucovorin, the low-dose monthly regimen was also less
expensive. However, the weekly schedule remains popular as it is more convenient
for some patients and is perceived as easier to manage because treatment can be
interrupted if early signs of toxicity occur. In contrast, with the monthly
schedule, clinical toxicity generally does not develop until after all 5 days of
treatment have been completed.

High- or Low-Dose Leucovorin

A number of randomized trials focused on ascertaining whether
high-dose leucovorin was necessary. Three randomized trials compared 20 mg/m² of
racemic (d,l-) leucovorin or 10 mg/m² of pure l-leucovorin with a ten-fold
higher dose administered with 5-FU (370-425 mg/m²) on the monthly
schedule.[18-20] None of the trials showed an advantage for high-dose leucovorin
in terms of either response rate, time to progression, or survival. Two other
trials compared 20 or 25 mg/m² leucovorin to 500
mg/m² leucovorin given as a
2-hour infusion with 5-FU at 500 or 600 mg/m² IV bolus at the midpoint.[3,21]
Although the response rate was somewhat higher with high-dose leucovorin, it was
not significant. Further, there was no difference in survival.

The impact of bolus 5-FU/leucovorin on survival has been greater
in the adjuvant setting. Randomized trials demonstrating a disease-free and
overall survival benefit with weekly bolus 5-FU/leucovorin as adjuvant therapy
in patients with colorectal cancer used high-dose leucovorin (500 mg/m²) given
as a 2-hour infusion.[22,23] Because the benefit of low-dose leucovorin with
weekly bolus 5-FU has not been tested in the adjuvant setting, caution should be
exercised in extrapolating the data from advanced disease to the adjuvant
setting. In contrast, in adjuvant colon cancer trials evaluating the monthly
schedule, a benefit was demonstrated with both high-dose (200 mg/m²) and
low-dose (20 mg/m²) leucovorin.

Methotrexate/5-FU

Other strategies have been investigated to modulate the activity
of 5-FU. Sequence-dependent synergy was noted in preclinical studies with
methotrexate preceded by 5-FU, while the opposite sequence was
antagonistic.[1,24] Inhibition of thymidylate synthase reduces consumption of
reduced folate cofactors that would otherwise be used in the conversion of
deoxyuridine monophosphate to thymidine monophosphate. The availability of the
reduced folates permits continued purine biosynthesis despite
methotrexate-mediated inhibition of dihydrofolate reductase. Administration of
methotrexate prior to 5-FU establishes inhibition of purine biosynthesis. As a
consequence, intracellular levels of phosphoribosylpyrophosphate increase, and
this promotes the anabolism of 5-FU to FUMP by orotate
phosphoribosyltransferase.

A number of phase III trials in advanced colorectal cancer have
compared methotrexate given at various intervals prior to 5-FU, with mixed
results. One pharmacodynamic study suggested that a 24-hour interval between
methotrexate and 5-FU was optimal for expansion of phosphorbosyl pyrophosphate
(PRPP) pools.[25] A clinical study in advanced colorectal cancer that directly
compared a 24-hour interval between administration of methotrexate and 5-FU with
a 1-hour interval demonstrated that the longer interval was superior.[26] A
meta-analysis of eight randomized trials involving 1,178 patients that compared
5-FU alone with methotrexate-modulated 5-FU (± leucovorin rescue) revealed a
significantly higher response rate (19% vs 10%, odds ratio 0.51, P < .0001)
and survival advantage (median, 10.7 vs 9.1 months, odds ratio 0.87, P = .024)
in favor of methotrexate/5-FU.[27] However, because many of the
methotrexate-containing arms included leucovorin rescue, at issue is whether the
apparent benefit might have been due to leucovorin.