High-performance liquid chromatography/mass spectrometry characterization of Ki4B-Ras in PSN-1 cells treated with the prenyltransferase inhibitor L-778,123
Ca. Buser et al., High-performance liquid chromatography/mass spectrometry characterization of Ki4B-Ras in PSN-1 cells treated with the prenyltransferase inhibitor L-778,123, ANALYT BIOC, 290(1), 2001, pp. 126-137
Cellular transformation by Ras oncoproteins requires the posttranslation mo
dification of farnesylation in a reaction catalyzed by farnesyl protein tra
nsferase (FPTase). Thus, inhibitors of FPTase have been developed as potent
ial anticancer agents. However, recent studies with selective inhibitors of
FPTase have shown that Ki4B-Ras retains its ability to transform cells by
undergoing alternative prenylation by the related geranylgeranyl protein tr
ansferase I (GGPTase-I) in human tumor cells. Ne have developed a high-perf
ormance liquid chromatography/mass spectrometry assay for the detection and
quantitation of the different processing states of Ki4B-Ras isolated from
PSN-1 cells (a human pancreatic cell line with an activating Gly12 to Arg m
utation) treated with the prenyltransferase inhibitor, L-778,123, Recently
tested in the clinic, L-778,123 is a potent inhibitor of FPTase (in vitro I
C50 = 2 nM) with some activity against GGPTase-I tin vitro IC50 = 98 nM). W
e find primarily farnesylated-Ki4B-Ras in vehicle-treated PSN-1 cells, a mi
xture of farnesylated- and geranylgeranylated-Ki4B-Ras in cells treated wit
h nanomolar concentrations of L-778,123, and a mixture of unprocessed, farn
esylated, and geranylgeranylated-Ki4B-Ras in cells treated with micromolar
concentrations of compound. Of importance, this technique does not require
metabolic labeling and may be used as a pharmacodynamic assay for Ki4B-Ras
processing in mouse models. (C) 2001 Academic Press.