RT Journal Article
SR Electronic
T1 Toxicity of Methotrexate and Metoprine in a Dihydrofolate Reductase Gene-Amplified Mouse Cell Line
JF Molecular Pharmacology
JO Mol Pharmacol
FD American Society for Pharmacology and Experimental Therapeutics
SP 637
OP 643
VO 20
IS 3
A1 MICHAEL HAMRELL
A1 JOHN LASZLO
A1 OLIVER E. BROWN
A1 W. DAVID SEDWICK
YR 1981
UL http://molpharm.aspetjournals.org/content/20/3/637.abstract
AB The comparative effects of methotrexate (MTX) and the lipid-soluble antifolate metoprine [DDMP, 2,4-diamino-5-(3',4'-dichlorophenyl)-6-methylpyrimidine] on DNA synthesis and cell viability were studied in an MTX-resistant mouse fibroblast 3T6R400 cell line. MTX levels as high as 1000 µM inhibited deoxyuridine incorporation into cellular DNA by only 28%, whereas a relatively low concentration (3 µM) of DDMP produced 50% inhibition. Inhibition of DNA synthesis and of cytotoxicity (measured by cloning efficiency) were approximately proportional with both drugs. Partially purified dihydrofolate reductase (DHFR) from the mutant cell was about 100 times more resistant to both MTX and DDMP than was enzyme from the wild-type cell. Furthermore, the resistant cell line took up MTX as efficiently as did the sensitive cell line. However, since the mutant cell contained very high levels of drug-resistant DHFR, a normal rate of drug entry apparently did not permit sufficient MTX to saturate the dihydrofolate reductase binding sites; consequently, cells escaped toxicity. Low concentrations of DDMP (relative to MTX) inhibit deoxyuridine incorporation by resistant cells. This inhibition occurs presumably because DDMP is not dependent on active transport. DDMP also was slightly more inhibitory (relative to MTX) for the DHFR of the mutant cell line than for this enzyme from the wild-type cells. Because toxicity to antifolates is mediated by a variety of factors in this mutant cell line, its observed sensitivity to lipid-soluble antifolates lends added importance for further developmental work on these compounds. We suggest that such drugs may be useful against tumors where resistance is a result of the complex interplay of transport limitations, altered target enzyme affinity, and changes in the amount of target enzyme.