Abstract
4404
Among four forms of Ras proteins (H-Ras, N-Ras, K-Ras4A and K-Ras4B), K-Ras has been found to be resistant to farnesyl transferase (FT) inhibitors. The basis for the resistance may be related to high affinity of K-Ras to FT and its capacity to be prenylated by the related enzyme GGTase, in presence of FT. To reveal that the level of FT is not the only index to predict cell sensitivity to inhibitors of the farnesylation pathway and to demonstrate that there are alternative pathways to consider in explaining the relative resistance to the FT inhibitor, we selected 3 breast cancer cell lines with similar levels of p21ras and FT-α/GGT- α, detected by immunohistochemical method, to evaluate the in vitro effects of Pamidronate, an inhibitor of the farnesylation pathway at the level of farnesyl diphosphoate synthase. All three cell lines, SKBR-3, MDA-175 and MDA-231, were all negative for hormonal receptors but with different levels of HER-2/enu protein expression (high, mild and minimal levels, respectively). After incubating with Pamidronate for 4 days, proliferative rates of the three types of cells were determined using colorimetric method and the inhibitory rate was calculated (control-treated/control) as tabulated (*p<0.05 vs SKBR-3 and # p<0.05 vs MDA-175 by ANOVA, repeated 3 times). At both 30 and 90 μM of Pamidronate, MDA-175 cells showed higher resistance than SKBR-3 and MDA-231 cells. To demonstrate the additive effects of Pamidronate with Gleevec, an inhibitor of farnesylation independent platelet-derived growth factor receptor and c-kit signaling, Pamidronate at 30 μM and Gleevec at 20 μM were both added to MDA-175 cells to result in 75.2 ± 1.9 % inhibition; Pamidronate alone caused a 39.5 ± 2.2 %a inhibition, whereas Gleevec alone caused 50.2 ± 3.7 % a inhibition in MDA-175 cells (a, p<0.05 vs combined effects). To test whether the Akt pathway (associated with the mammalian target of rapamycin [m-TOR] and nuclear factor [NF]-kappaB) may also be an alternative farnesylation independent growth pathway associated with cell proliferation in MDA-175 cells, we combined rapamycin at 1.0 μM (an inhibitor of m-TOR) and Velcade at 30 nM (an inhibitor of proteasome and NF-kappaB), which additively resulted in 76.9 ± 0.9 % inhibition; rapamycin alone caused 28.1 ± 4.3 %a inhibition, whereas Velcade along caused 64.3 ± 1.3 % a inhibition. In summary, resistance to Pamidronate may not be simply determined by the level of FT and inhibition of other signal pathways not necessary related to Ras can provide additive effects overcoming the drug resistance.
[Proc Amer Assoc Cancer Res, Volume 45, 2004]
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