Gemcitabine (Gemzar) is a pyrimidine antimetabolite that is anabolized sequentially to the nucleoside mono-, di-, and triphosphate intracellularly. Difluorodeoxycytidine triphosphate (dFdCTP) is incorporated into DNA, resulting in chain termination. In addition, gemcitabine(Drug information on gemcitabine) inhibits ribonucleotide reductase, an enzyme that catalyzes the formation of deoxynucleosides required for DNA synthesis. Gemcitabine has broad activity in a variety of solid tumors, including breast cancer, and is approved for the treatment of pancreatic and non-small-cell lung cancers. The clinical pharmacology of gemcitabine has been reviewed recently.
Pemetrexed disodium (Alimta, LY231514) is a novel antimetabolite that targets at least three enzymes involved in the synthesis of purines and pyrimidines. A primary enzyme target is thymidylate synthase (TS),[4-6] a folate-dependent enzyme, whose inhibition results in decreased thymidine necessary for DNA synthesis.
The other known enzyme targets of pemetrexed(Drug information on pemetrexed) disodium are dihydrofolate reductase (DHFR) and glycinamide ribonucleotide formyl transferase (GARFT). The dependence of the cytotoxicity of pemetrexed disodium on both DHFR and GARFT is supported by the fact that thymidine and hypoxanthine are both required to rescue cells from the cytotoxic effects of this agent in vitro.
Key Enzyme Targets
The three enzyme targets for pemetrexed disodium are shown in Figure 1. As illustrated, pemetrexed disodium is not only similar to methotrexate(Drug information on methotrexate) (a DHFR inhibitor), fluorouracil(Drug information on fluorouracil) (5-FU), and raltitrexed (Tomudex, a TS inhibitor), but also inhibits GARFT, which currently has no clinically relevant inhibitor. Because of its ability to inhibit multiple enzymes, pemetrexed disodium may prove to have superior clinical activity in comparison to other antifolates and TS inhibitors.
Pemetrexed is transported into cells via the reduced folate carrier, and is polyglutamated in a reaction that is catalyzed by folylpolyglutamate synthase (FPGS). The predominant intracellular glutamated form of pemetrexed disodium is the pentaglutamate, which is greater than 60-fold more potent in its inhibition of TS than the monoglutamate. The pharmacology and clinical activity of pemetrexed disodium has been comprehensively reviewed.
Folate Status and Toxicity
Folic acid and its derivatives have been traditionally utilized to reverse the toxicity of antifolate agents. However, there has not been a consistent correlation between cellular or serum folate levels of patients and the incidence of toxicity from antifolate chemotherapeutic agents. Recent data have provided some insight into this apparent contradiction by indicating that plasma homocysteine is a much more sensitive measure of the functional folate status of patients than the traditionally used measures of red blood cell counts or serum folate levels. In view of these data, serum homocysteine and vitamin metabolite levels have been evaluated as predictors of pemetrexed disodium toxicity in phase I and phase II studies.
The s-adenosylmethionine (SAM) cycle responsible for critical single-carbon transfer reactions (through methyl groups) in mammalian systems is illustrated in Figure 2. The transfer of a methyl group from N5-methyltetrahydrofolate (CH3FH4) to homocysteine generates methionine, which in turn generates SAM. When humans are folate-deficient, the lack of CH3FH4 leads to an increase in the levels of plasma homocysteine, which is an early, sensitive, and reliable indicator of folate deprivation.
Vitamins B12 and B6
Cobalamin (vitamin B12) and pyridoxine(Drug information on pyridoxine) (vitamin B6) deficiencies can also lead to an increase in plasma homocysteine levels. Cobalamin is a cofactor for two synthetic processes in mammals. The first process is the synthesis of methionine from homocysteine and CH3FH4, catalyzed by methionine synthase. The second process is the formation of succinyl co-enzyme A from L-methylmalonic acid co-enzyme A. Serum methylmalonic acid levels are therefore elevated in cobalamin deficiency, but not in folate deficiency, and are useful in the differential diagnosis of cobalamin and folate deficiency in the setting of elevated plasma homocysteine levels.
Pyridoxine is involved in the conversion of homocysteine to cystathionine, and the subsequent conversion of cystathionine to cysteine(Drug information on cysteine) and alphaketoglutaric acid. Cystathionine levels are elevated to a much greater extent in vitamin B6 deficiency, compared to folate and B12 deficiency.[15,16] Isolated vitamin B6 deficiency, however, occurs very rarely.
Phase I and Phase II Studies
With the emerging information described above, plasma homocysteine, cystathionine, and methylmalonic acid levels have been measured and included in a multivariate analysis of potential prognostic factors that predict serious toxicity in patients treated with pemetrexed disodium. In phase II trials, 139 patients with a variety of solid tumors were treated with pemetrexed disodium at doses of 600 mg/m2 every 3 weeks.
Plasma homocysteine, other vitamin deficiency markers, serum albumin, and hepatic enzymes were measured at baseline and once each cycle thereafter. Preliminary data find baseline plasma homocysteine concentrations to be the only statistically significant prognostic factors for serious toxicities, predominantly myelosuppression. Mucositis and diarrhea were also correlated with plasma concentrations of methylmalonic acid. A threshold baseline homocysteine value of 10 µmol/L was used to differentiate between high- and low-risk populations.
Plasma concentrations > 10 µmol/L predicted a greatly increased rate of toxicities, even though a continuum existed for various serum levels. These studies support the notion that individuals who are folate-deficient are at subclinically or clinically increased risk of severe toxicity when treated with standard doses of pemetrexed disodium.
In an ongoing phase I trial, pemetrexed disodium is administered with high-dose intermittent folic acid(Drug information on folic acid) supplementation (5 mg orally, days -2 to +2). Available results suggest that this folic acid supplementation schedule permits marked dose escalation of pemetrexed disodium with minimal toxicity. Minimally pretreated patients have tolerated pemetrexed disodium doses of up to 925 mg/m2.
A strict interpretation of these findings would suggest that patients with reduced folate pools should receive folic acid supplementation before receiving pemetrexed disodium. Since plasma homocysteine is a continuous predictor for severe toxicities, it may be assumed that folic acid supplementation significantly improves the tolerability of pemetrexed disodium in all patients. Early evidence that supplementation with low-dose daily oral folic acid and quarterly IM vitamin B12 significantly reduced toxicities lends support to this idea.
Pemetrexed disodium has been investigated as a single agent in advanced breast cancer without folic acid and vitamin B12 support. Preliminary results from two completed phase II studies in Europe have been reported.
In one study, most patients (33 out of 38) had been previously treated with chemotherapy. Of these, 26 patients had previously received anthracyclines, while 4 had been treated with taxanes; 16 patients received therapy in the adjuvant setting. Out of 36 evaluable patients, 10 achieved a partial response, and 1 patient achieved a complete response, for an overall response rate of 31% (95% confidence interval [CI]: 16%-46%). Responses were seen in three out of the four patients previously treated with taxanes and in three out of the 26 patients previously treated with anthracyclines. The median survival was 13 months, and median time to disease progression was 5 months. The median response duration was > 9 months.
In the second study, all enrolled patients had been previously treated with anthracyclines. In all, 26 patients had documented disease progression ≤ 30 days after anthracycline treatment (anthracycline refractory) and 43 patients developed progressive disease > 30 days after stopping treatment (anthracycline failures); 29 patients had also received a taxane. The overall response rate in 69 evaluable patients was 23% in anthracycline failures and 19% in anthracycline refractory patients.
In the cohort of patients who received both anthracyclines and taxanes, objective responses were documented in 8 out of a total of 29 patients for an overall response rate of 28%. The median response duration was 6 months, the median time to progression was 4 months, and the 1-year survival was 46%.
These early results indicate that pemetrexed disodium may not demonstrate complete cross resistance with taxanes or anthracyclines. To test this hypothesis, a phase II study of pemetrexed disodium in patients with stage IV breast cancer who have previously received anthracyclines and taxanes is underway in the United States and Europe. This study enrolled 40 patients prior to a protocol amendment that added folic acid and vitamin B12 supplementation to the remaining patients enrolled. Preliminary data report a 19% objective response rate with grade 3/4 neutropenia as the most frequent toxicity. Final reports of the analysis from this study are awaited.