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

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 COLO320-DM human colon adenocarcinoma which harbours a wild-type K- ras gene. Immunoblot analysis of isolated cell membranes and total cellular lysates of COLO320-DM cells demonstrated that manumycin dose-dependently reduced p21 ras farnesylation with a 50% inhibitory concentration (IC50) of 2.51 ± 0.11 μM and 2.68 ± 0.20 μM, respectively, while the geranylgeranylation of p21 rhoA and p21 rap1 was not affected. Manumycin dose-dependently inhibited (IC50= 2.40 ± 0.67 μM) the phosphorylation of the mitogen-activated protein kinase/extracellular-regulated kinase 2 (p42MAPK/ERK2), the main cytoplasmic effector of p21 ras, as well as COLO320-DM cell growth (IC50= 3.58 ± 0.27 μM) without affecting the biosynthesis of cholesterol. Mevalonic acid (MVA, 100 μM), a substrate of the isoprenoid synthesis, was unable to protect COLO320-DM cells from manumycin cytotoxicity. Finally, manumycin 1–25 μM for 24–72 h induced oligonucleosomal fragmentation in a dose- and time-dependent manner and MVA did not protect COLO320-DM cells from undergoing 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 cells and the effect on cell growth does not require the presence of a mutated ras gene for maximal expression of chemotherapeutic activity. © 2000 Cancer Research Campaign

[1]  C. Der,et al.  Farnesyltransferase inhibitors and cancer treatment: targeting simply Ras? , 1997, Biochimica et biophysica acta.

[2]  A. Levitzki.  Targeting signal transduction for disease therapy , 1997, Medical oncology.

[3]  F. Larcher,et al.  Up-regulation of vascular endothelial growth factor/vascular permeability factor in mouse skin carcinogenesis correlates with malignant progression state and activated H-ras expression levels. , 1996, Cancer research.

[4]  A. Marchetti,et al.  BRONCHIOLOALVEOLAR LUNG CARCINOMAS: K‐ras MUTATIONS ARE CONSTANT EVENTS IN THE MUCINOUS SUBTYPE , 1996, The Journal of pathology.

[5]  D. Samid,et al.  Phenylacetate inhibits protein isoprenylation and growth of the androgen-independent LNCaP prostate cancer cells transfected with the T24 Ha-ras oncogene. , 1996, Molecular pharmacology.

[6]  J. Veerkamp,et al.  Effects of HMG-CoA reductase inhibitors on growth and differentiation of cultured rat skeletal muscle cells. , 1996, Biochimica et biophysica acta.

[7]  A. Levitzki.  Targeting signal transduction for disease therapy. , 1996, Current opinion in cell biology.

[8]  Y. Matsuzawa,et al.  Inhibition of cell growth of human hepatoma cell line (HepG2) by a farnesyl protein transferase inhibitor: A preferential suppression of ras farnesylation , 1996, International journal of cancer.

[9]  T. Nagasu,et al.  Inhibition of human tumor xenograft growth by treatment with the farnesyl transferase inhibitor B956. , 1995, Cancer research.

[10]  A. Kikuchi,et al.  rap1 p21 regulates the interaction of ras p21 with RGL, a new effector protein of ras p21 , 1995, FEBS letters.

[11]  M. Marshall,et al.  Ras target proteins in eukaryotic cells , 1995, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[12]  A. Wittinghofer,et al.  Ras‐effector interactions, the problem of specificity , 1995, FEBS letters.

[13]  A. Wittinghofer,et al.  The 2.2 Å crystal structure of the Ras-binding domain of the serine/threonine kinase c-Raf1 in complex with RaplA and a GTP analogue , 1995, Nature.

[14]  R. Danesi,et al.  Paclitaxel (taxol) inhibits protein isoprenylation and induces apoptosis in PC-3 human prostate cancer cells. , 1995, Molecular pharmacology.

[15]  J. Pollard,et al.  Peripheral neuropathy associated with simvastatin. , 1995, Journal of neurology, neurosurgery, and psychiatry.

[16]  Min Han,et al.  Ras farnesyltransferase inhibitors suppress the phenotype resulting from an activated ras mutation in Caenorhabditis elegans. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[17]  M. Okuma,et al.  A new method for quantitative estimation of the degree of DNA fragmentation utilizing agarose gel electrophoresis. , 1994, International journal of hematology.

[18]  F. Giancotti,et al.  Integrin-mediated adhesion and signaling in tumorigenesis. , 1994, Biochimica et biophysica acta.

[19]  N. Kohl,et al.  Farnesyltransferase inhibitors: Ras research yields a potential cancer therapeutic , 1994, Cell.

[20]  F. Mollinedo,et al.  Inhibition of isoprenoid biosynthesis induces apoptosis in human promyelocytic HL-60 cells. , 1994, Biochemical and biophysical research communications.

[21]  F. Tamanoi,et al.  Inhibitors of Ras farnesyltransferases. , 1993, Trends in biochemical sciences.

[22]  C J Marshall,et al.  Protein prenylation: a mediator of protein-protein interactions. , 1993, Science.

[23]  F. Tamanoi,et al.  Identification of Ras farnesyltransferase inhibitors by microbial screening. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[24]  W. Maltese.  Posttranslational modification of proteins by isoprenoids in mammalian cells , 1990, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[25]  J. L. Bos,et al.  ras oncogenes in human cancer: a review. , 1989, Cancer research.

[26]  Oliver H. Lowry,et al.  Protein measurement with the Folin phenol reagent. , 1951, The Journal of biological chemistry.

[27]  L. V. van't Veer,et al.  Mutational activation of the K-ras oncogene and the effect of chemotherapy in advanced adenocarcinoma of the lung: a prospective study. , 1997, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[28]  J. Gibbs,et al.  The potential of farnesyltransferase inhibitors as cancer chemotherapeutics. , 1997, Annual review of pharmacology and toxicology.

[29]  C. Marshall.  Ras effectors. , 1996, Current opinion in cell biology.

[30]  A. Catapano,et al.  Pharmacology of competitive inhibitors of HMG-CoA reductase. , 1995, Pharmacological research.

[31]  B R Goldspiel,et al.  Paclitaxel (Taxol) , 1994, Pharmacotherapy.

[32]  D. Lowy,et al.  Function and regulation of ras. , 1993, Annual review of biochemistry.