Biochemical modulation of tumor cell energy in vivo: II. A lower dose of adriamycin is required and a greater antitumor activity is induced when cellular energy is depressed.

A quadruple drug combination--consisting of a triple-drug combination of N-(phosphonacetyl)-L-aspartate (PALA) + 6-methylmercaptopurine riboside (MMPR) + 6-amino-nicotinamide (6-AN), designed to primarily deplete cellular energy in tumor cells, + Adriamycin (Adria)--yielded significantly enhanced anticancer activity (i.e., tumor regressions) over that produced by either Adria alone at maximum tolerated dose (MTD) or by the triple-drug combination, against large, spontaneous, autochthonous murine breast tumors. The adenosine triphosphate (ATP)-depleting triple-drug combination administered prior to Adria resulted in a 100% tumor regression rate (12% complete regression; 88% partial regression) of spontaneous tumors. Histological examination of treated tumors demonstrated that the treatment-induced mechanism of cancer cell death was by apoptosis. The augmented therapeutic results (100% tumor regressions) were obtained with approximately one-half the MTD of Adria as a single agent and suggest the potential clinical benefit of longer, more effective, and safer treatment by low doses of Adria when combined with the triple-drug combination. Two likely mechanisms of action are discussed: (1) prevention of DNA repair; (2) complementary disruption of biochemical pathways by both the triple-drug combination and the biochemical cascade of apoptosis that is induced by a DNA-damaging anticancer agents such as Adria.

Leucovorin enhances cytotoxicity of trimetrexate/fluorouracil, but not methotrexate/fluorouracil, in CCRF-CEM cells.

BACKGROUND: Increased response rates in studies of patients with colon cancer have indicated that the cytotoxic effects of fluorouracil (5-FU) are potentiated by leucovorin (LV) and by methotrexate (MTX). However, preliminary studies using a sequential combination of MTX, LV, and 5-FU showed no additional potentiation. PURPOSE: We hypothesized that the lack of additional cell kill with this combination could be due to competition of LV with MTX for cellular uptake and reduced folate polyglutamylation. We have tested this possibility by comparing the cytotoxicity of drug combinations containing MTX with that of drug combinations containing trimetrexate (TMTX), an antifolate that does not compete with LV for uptake or polyglutamylation. METHODS: Human lymphocytic leukemia CCRF-CEM cells were exposed to MTX or TMTX for 24 hours and to 5-FU during the last 4 hours of antifolate exposure. LV was administered 30 minutes before 5-FU. RESULTS: After 20 hours of exposure to TMTX or MTX, intracellular levels of phosphoribosyl pyrophosphate were elevated to a similar degree, and these levels did not decrease after a 30-minute exposure to LV. No additional cell kill was observed when LV was added to the MTX/5-FU combination, but cytotoxicity was enhanced when LV was added to the TMTX/5-FU combination. CONCLUSIONS: This study supports the hypothesis that the lack of additional cell kill when high-dose LV is added to the MTX/5-FU combination may be due to competition of MTX with LV for cellular uptake and/or competition of MTX or its polyglutamates with polyglutamylation of reduced folates. Inasmuch as TMTX does not compete with LV and reduced folates for uptake and polyglutamylation, the synergy obtained with the combination of TMTX plus 5-FU and high-dose LV further supports this hypothesis.

Failure of high-dose leucovorin to improve therapy with the maximally tolerated dose of 5-fluorouracil: a murine study with clinical relevance?

Almost all of the completed and ongoing phase III trials of the leucovorin/5-fluorouracil (LV/5-FU) combination have used either a single-agent 5-FU control arm in which the 5-FU was administered in a different schedule from the LV/5-FU arm or one in which the 5-FU was not at the maximally tolerated dose (MTD). Because both dose intensity and scheduling are known to affect drug activity, the LV/5-FU combination was evaluated in the preclinical CD8F1 murine model of advanced first-passage spontaneous breast tumors using the same dose (at MTD) and schedule for 5-FU alone and in the LV/5-FU combination arm. Overall, therapy with 5-FU at MTD was not improved by LV. Further, although the activity of 5-FU doses lower than the MTD could be increased by LV, the therapeutic result was comparable to that of single-agent 5-FU at MTD. In an evaluation with other modulators of 5-FU (e.g., uridine, PALA, methotrexate), therapy with various modulated 5-FU combinations at their MTD was not improved with LV. In conclusion, although LV can enhance the cytotoxicity of 5-FU in these in vivo preclinical studies, it does not confer enhanced selectivity to 5-FU, a conclusion at odds with many present clinical reports. Whether or not these murine findings have clinical relevance can be determined only by clinical trials designed with the MTD of 5-FU alone in the control arm, the MTD of 5-FU (or as close as tolerated) in the LV/5-FU arm, and identical schedules in both arms.