Uptake, cytotoxicity and metabolism of m-azidopyrimethamine and related lipophilic antifolates in SV-K14 human keratinocytes in vivo.
The growth-inhibitory properties of a series of lipophilic diaminopyrimidine antifolates were evaluated in comparison with methotrexate (MTX) against SV-K14 human keratinocytes in vitro under folate-dependent and folate-independent conditions. Under folate-dependent conditions metoprine (DDMP) proved more cytotoxic than MTX, despite the greater inhibitory activity of the latter compound against mammalian dihydrofolate reductase (DHFR), possibly reflecting differences in cellular accumulation. The significantly lower activity of both compounds under folate-independent conditions indicated DHFR as the primary target. Pyrimethamine (PYM), m-azidopyrimethamine (MZP) and m-aminopyrimethamine (MAP), a metabolite of MZP, were approximately equiactive but less cytotoxic than MTX or DDMP. The unexpected activity of MAP, an inferior DHFR inhibitor, suggests differences in the mechanism of action or cellular transport of the drug, although the reduction of cytotoxicity observed under folate-independent conditions indicate folate metabolism as the cytotoxic locus. In contrast, the cytotoxicity of PYM or MZP was not reduced under folate-independent conditions implying an alternative mechanism of action. The uptake of 2-[14C]pyrimethamine by SV-K14 keratinocytes was rapid with steady-state intracellular concentrations being observed after approximately 100 min, partition of drug into the plasma membrane preceding redistribution and extensive accumulation within the particulate cell components. The previously reported NADPH-dependent metabolism of MZP to MAP by murine liver microsome preparations was not observed with SV-K14 keratinocytes nor with murine skin homogenates in the present study.
Baker
N D
ND
Pharmaceutical Sciences Institute, Aston University, Birmingham U.K.
Griffin
R J
RJ
Irwin
W J
WJ
Reichert
U
U
Schmidt
R
R
D000818:Animals; D000875:Anthralin / Q000494:pharmacology; D001386:Azides / Q000378:metabolism / Q000494:pharmacology* / Q000627:therapeutic use; D002455:Cell Division / Q000187:drug effects; D002461:Cell Line, Transformed / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D003470:Culture Media; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical; D005493:Folic Acid Antagonists; D006801:Humans; D066298:In Vitro Techniques; D015603:Keratinocytes / Q000187:drug effects* / Q000201:enzymology / Q000648:ultrastructure; D008099:Liver / Q000187:drug effects; D008727:Methotrexate / Q000494:pharmacology; D051379:Mice; D008812:Mice, Hairless; D011739:Pyrimethamine / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology / Q000627:therapeutic use; D012044:Regression Analysis; D012867:Skin / Q000187:drug effects* / Q000201:enzymology
D001386:Azides; D003470:Culture Media; D005493:Folic Acid Antagonists; C006308:metoprine; C055910:3-azidopyrimethamine; D000875:Anthralin; D008727:Methotrexate; D011739:Pyrimethamine