Inhibition of cell proliferation by putative metabolites and non-degradable analogs of methotrexate-gamma-dimyristoylphosphatidylethanolamine.
Previous investigations have shown that untargeted liposomes, in which methotrexate is anchored to the lipid bilayers as methotrexate-gamma-dimyristoylphosphatidylethanolamine (methotrexate-gamma-DMPE), can inhibit in vitro cell proliferation. To test the possibility that this inhibition may involve extracellular metabolism of methotrexate-gamma-DMPE, we have degraded it chemically (dilute alkali) or enzymatically (phospholipase A2, phospholipase C, phospholipase C plus phosphatase), and assayed the products using human lymphoblastoid T cells or a subline that has a defective methotrexate transport system. Neither methotrexate-gamma-(1-myristoyl)-glycerophosphorylethanolamine, methotrexate-gamma-glycerophosphorylethanolamine, methotrexate-gamma-phosphorylethanolamine, nor methotrexate-gamma-ethanolamine resemble methotrexate-gamma-DMPE sensitized liposomes or the free derivative in their ability to block tritiated deoxyuridine incorporation into DNA. When added extracellularly, these putative metabolites manifest a higher ID50 concentration and/or, unlike the liposomes or unincorporated methotrexate-gamma-DMPE, utilize the methotrexate transport system to enter cells. Additionally, we have synthesized methotrexate-gamma-dihexadecylphosphatidylethanolamine and methotrexate-gamma-hexadecylphosphorylethanolamine, analogs of methotrexate-gamma-DMPE that cannot be hydrolyzed by phospholipases A2, C and D; liposomes prepared with these derivatives are markedly less potent cytotoxic agents than methotrexate-gamma-DMPE sensitized liposomes. All together, these results are consistent with the conclusion that methotrexate-gamma-DMPE must undergo intracellular metabolism to exert optimal inhibition; they also bear on possible mechanisms by which methotrexate-gamma-DMPE may enter cells.
D002455:Cell Division / Q000187:drug effects*; D002460:Cell Line; D003857:Deoxyuridine / Q000378:metabolism; D006801:Humans; D008081:Liposomes / Q000378:metabolism; D008727:Methotrexate / Q000031:analogs & derivatives* / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology; D010714:Phosphatidylethanolamines / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology*; D013601:T-Lymphocytes / Q000378:metabolism
D008081:Liposomes; D010714:Phosphatidylethanolamines; C047978:methotrexate-gamma-1,2-dimyristoylphosphatidylethanolamine; D003857:Deoxyuridine; D008727:Methotrexate