Manumycin inhibits ras signal transduction pathway and induces apoptosis in COLO320-DM human colon tumour cells

The aim of the present study was to assess the cytotoxicity of manumycin, a specific inhibitor of farnesyl:protein transferase, as well as its effects on protein isoprenylation and kinase-dependent signal transduction in COLO 320-DM human colon adenocarcinoma which harbours a wild-type K-ras gene. Im munoblot analysis of isolated cell membranes and total cellular lysates of COLO320-DM cells demonstrated that manumycin dose-dependently reduced p21ra s farnesylation with a 50% inhibitory concentration (IC50) of 2.51 +/- 0.11 mu M and 2.68 +/- 0.20 mu M, respectively, while the geranylgeranylation o f p21rhoA and p21rap1 was not affected. Manumycin dose-dependently inhibite d (IC50 = 2.40 +/- 0.67 mu M) the phosphorylation of the mitogen-activated protein kinase/extracellular-regulated kinase 2 (p42MAPK/ERK2), the main cy toplasmic effector of p21 ras, as well as COLO320-DM cell growth (IC50 = 3. 58 +/- 0.27 mu M) without affecting the biosynthesis of cholesterol. Mevalo nic acid (MVA, 100 mu M), a substrate of the isoprenoid synthesis, was unab le to protect COLO320-DM cells from manumycin cytotoxicity. Finally, manumy cin 1-25 mu M for 24-72 h induced oligonucleosomal fragmentation in a dose- and time-dependent manner and MVA did not protect COLO320-DM cells from un dergoing DNA cleavage. The present findings indicate that the inhibition of p21 ras processing and signal transduction by manumycin is associated with marked inhibition of cell proliferation and apoptosis in colon cancer cell s and the effect on cell growth does not require the presence of a mutated ras gene for maximal expression of chemotherapeutic activity. (C) 2000 Canc er Research Campaign.