Raloxifene Attenuation of Methotrexate Cytotoxicity in Human Bone Marrow by Sequence-dependent Administration of Raloxifene, 5-FU/Methotrexate

Abstract

Background: Increasing evidence suggests that adjuvant systemic chemotherapy is necessary for the survival of breast cancer patients. Antitumor agents are more effective when used in combination with drugs exhibiting different mechanisms of action than when used alone. Previous studies from this laboratory have shown that raloxifene (RAL) attenuation of 5-fluorouracil/methotrexate (5-FU/MTX) cytotoxicity to breast cancer cells was sequence-dependent. The aim was to evaluate the same combination of RAL, 5-FU and MTX to determine the most effective regimes and cellular mechanisms of action to mitigate MTX cytotoxicity in human bone marrow cells. Materials and Methods: The sequence-dependent interaction among MTX, 5-FU and RAL on the proliferation and viability of human bone marrow HS-5 cells was determined by the MTT assay and the Trypan blue dye exclusion assay by exposing the cells to MTX, 5-FU and RAL alone, RAL 24 h prior to 5-FU followed 2 h later by MTX, and 5-FU 2 h prior to MTX followed 24 h later by RAL. The control cells were untreated. Results: The growth rate in MCF-7 in early RAL was 68±3.07% and late RAL 37±2.05% of the control rate, whereas in bone marrow the same drug combinations exhibit a significant protection against MTX cytotoxity, with the early RAL combination yielding 81±3.77% and late 54±2.74% of the control. The finding was further supported by cell flow cytometry and Western blot analysis. Conclusion: Sequence-dependent administration of RAL in combination with 5-FU/MTX may have maximum antineoplastic activity in breast cancer while at the same time provide protection to human bone marrow.