Imexon-based combination chemotherapy in A375 human melanoma and RPMI 8226 human myeloma cell lines

Abstract

Purpose: This study evaluated the cytotoxic effects of imexon (NSC-714597) in tumor cells when combined with a broad panel of chemotherapeutic drugs.

Methods: The sulforhodamine B (SRB) and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cytotoxicity assays were used to analyze the degree of growth inhibition for the combination studies in the A375 human malignant melanoma and RPMI 8226 human multiple myeloma cell lines, respectively. Cells were continuously exposed to both drugs at a constant molar ratio for 4-5 days. Combination effects were analyzed using the Median Effect method. Statistical significance was inferred if the 95% confidence interval for the combination interaction (C.I.) values for a particular two-drug combination did not include 1.0 (additivity). Synergy was inferred for C.I. values<1.0 and antagonism for CI values>1.0.

Results: Imexon was synergistic when combined with DNA-binding agents (cisplatin, dacarbazine, melphalan) and pyrimidine-based antimetabolites (cytarabine, fluorouracil, gemcitabine) in both cell lines. Antagonistic combinations with imexon included methotrexate and the topoisomerase I (TOPO I) and II (TOPO II) inhibitors irinotecan, doxorubicin, mitoxantrone and etoposide. Docetaxel was synergistic with imexon in both cell lines whereas paclitaxel and fludarabine showed a mixed result. Dexamethasone and the proteasome inhibitor bortezomib showed synergy in myeloma cells and additivity in the melanoma cells. The vinca alkaloid, vinorelbine, and the multi-targeted antifol, pemetrexed, were additive with imexon in both cell lines.

Discussion: The consistent synergy seen for imexon and alkylating agents may relate to the sulfhydryl-lowering effect of imexon, which would render cells more sensitive to electrophilic species from the alkylators. The marked synergy noted with pyrimidine-based antimetabolites was unexpected and may relate to the induction of cell cycle arrest in S-phase. The strong antagonism noted for imexon with topoisomerase I and II inhibitors may be due to the effect of imexon at increasing oxidant levels which are known to antagonize the cytotoxic effects of topoisomerase poisons. In contrast, the synergy seen with bortezomib in myeloma cells may be related to an increase in reactive oxygen species (ROS) from both drugs. These results suggest that combinations of imexon with alkylating agents and pyrimidine-based antimetabolites are rational to pursue in therapeutic studies in vivo.

Fig. 1

Combination indices of representative DNA-binding agents with imexon. The mean combination indexes (logarithmic y-axis) are plotted versus the fraction of cells effected (% of control growth, x-axis), for DNA-binding agents combined with imexon in RPMI 8226 multiple myeloma cells (closed symbols) and in A375 malignant melanoma cells (open symbols). The vertical bars indicate one standard deviation for the combinations of imexon with: a cisplatin, b dacarbazine, c melphalan and d mitomycin. The horizontal bar at 1.0 indicates the line of simple additivity

Fig. 2

Combination indices of antimetabolites with imexon. The mean combination indexes (logarithmic y-axis) are plotted versus the fraction of cells effected (% of control growth, x-axis), for pyrimidine-based antimetabolites combined with imexon in RPMI 8226 multiple myeloma cells (closed symbols) and in A375 malignant melanoma cells (open symbols). The vertical bars indicate one standard deviation for the combinations of imexon with: a cytarabine, b fludarabine, c gemcitabine and d fluorouracil

Fig. 3

Combination indices of topoisomerase inhibitors with imexon. The mean combination indexes (logarithmic y-axis) are plotted versus the fraction of cells effected (% of control growth, x-axis), for agents that were antagonist when combined with imexon in RPMI 8226 multiple myeloma cells (closed symbols) and in A375 malignant melanoma cells (open symbols). The vertical bars indicate one standard deviation for the combinations of imexon with: a methotrexate, b irinotecan, c doxorubicin and d etoposide