M. Wachsman et al., ANTICYTOMEGALOVIRAL ACTIVITY OF METHOTREXATE ASSOCIATED WITH PREFERENTIAL ACCUMULATION OF DRUG BY CYTOMEGALOVIRUS-INFECTED CELLS, Antimicrobial agents and chemotherapy, 40(2), 1996, pp. 433-436
We extend the observation that inhibitors of pyrimidine biosynthesis a
re active against human cytomegalovirus by demonstrating that methotre
xate (MTX) has preferential activity against cytomegalovirus replicati
on, The 50% and 90% inhibitory concentrations of MTX for inhibition of
cytomegaloviral DNA replication at 3 days postinfection in MRC-5 cell
s were 0.05 and 0.2 mu M, respectively. No cell toxicity was observed
in uninfected confluent cells at the highest concentration tested (1 m
u M). Under similar conditions (3 days of treatment with 0.2 mu M MTX)
, intracellular dTTP pools were diminished in cytomegalovirus-infected
cells (87% decrease relative to untreated infected cells, P < 0.001)
but were not reduced in uninfected cells. A potential explanation for
the preferential antiviral effect of MTX was that human cytomegaloviru
s-infected cells preferentially accumulated MTX. Increased intracellul
ar accumulation and increased polyglutamation of MTX were observed in
cytomegalovirus-infected cells compared with uninfected cells, Increas
ed uptake of [H-3]MTX by cytomegalovirus-infected cells was first obse
rved at 48 h postinfection, with threefold-higher accumulation within
infected cells. By 96 h, accumulation had increased to approximately f
ourfold in comparison with uninfected cells, The uptake of [H-3]MTX wa
s saturable and was blocked by addition of unlabelled MTX. Intracellul
ar MTX in infected cells was almost entirely in the polyglutamated for
m, as demonstrated by thin-layer chromatography, whereas intracellular
MTX was almost exclusively in the parent form in uninfected cells.