Decrease in susceptibility toward induction of apoptosis and alteration inG(1) checkpoint function as determinants of resistance of human lung cancer cells against the antisignaling drug UCN-01 (7-hydroxystaurosporine)
K. Sugiyama et al., Decrease in susceptibility toward induction of apoptosis and alteration inG(1) checkpoint function as determinants of resistance of human lung cancer cells against the antisignaling drug UCN-01 (7-hydroxystaurosporine), CANCER RES, 59(17), 1999, pp. 4406-4412
7-Hydroxystaurosporine (UCN-01) is a protein kinase inhibitor that is under
development as an anticancer agent in the United States and Japan. Long-te
rm exposure of human A549 non-small cell lung cancer cells to UCN-01 furnis
hed cells (A549/UCN) with acquired resistance against UCN-01. In this study
, the sensitivity of these cells toward the growth-arresting properties of
certain conventional cytotoxic agents was explored. Cells were not cross-re
sistant against adriamycin, Taxol, staurosporine, and UCN-02, but they disp
layed 14- and 4.4-fold resistance against cisplatin and mitomycin C, respec
tively. Previous studies on the mechanism(s) of action of UCN-01 suggest th
at induction of apoptosis and G(1) phase accumulation are important for its
anticancer activity; therefore, we compared induction of apoptosis and cel
l cycle distribution caused by UCN-01 in wild-type A549 and A549/UCN cells
using flow cytometry. UCN-01 (0.4 mu M) induced apoptosis (62% terminal deo
xynucleotidyl transferase mediated nick end labeling-positive cells) in A54
9 cells, but not in A549/UCN cells. The percentages of cells that accumulat
ed in G(1) when exposed to UCN-01 (0.4 mu M) were 22% in A549 cells and 67%
in A549/UCN cells. These results suggest that acquired resistance of cance
r cells against UCN-01 is characterized by attenuation of apoptosis inducti
on associated with reinforcement of the G(1) checkpoint and that apoptosis
regulation is drastically altered in A549/UCN cells as compared with A549 c
ells. Cyclin-dependent kinase (CDK) inhibitor proteins p21 and p27 in A549/
UCN cells were up-regulated, which was accompanied by overexpression of G(1
) cyclins D1 and E, but UCN-01 hardly affected levels of these proteins. In
contrast, cyclin A, cyclin B1, retinoblastoma, and CDK2 proteins were appa
rently down-regulated, without changes in CDK4/6, UCN-01 hardly affected th
e expression level of cyclin B1 and induced dephosphorylation of retinoblas
toma in both cell types. UCN-01 induced down-regulation of cyclin A level a
nd CDK2 activity accompanied with its dephosphorylation in A549/UCN cells,
but not in A549 cells. The antiapoptotic protein bcl-2 was apparently up-re
gulated in A549/UCN cells, however, bcl-xL, another antiapoptotic protein,
was down-regulated, without changes in bak and bar. Taken together, these r
esults are consistent with the notion that induction of apoptosis and block
of cell cycle in G(1) are important determinants of the sensitivity of can
cer cells to UCN-01 and suggest that inhibition of CDK2 activity accompanie
d by its dephosphorylation and decrease of expression level of cyclin A mig
ht play an important role in the G(1) phase accumulation induced by UCN-01.