Increased expression of heat shock protein 70 in adherent ovarian cancer and mesothelioma following treatment with manumycin, a farnesyl transferase inhibitor.
Since farnesylation by farnesyltransferase (FTase) is the first obligatory step in the signaling transduction pathway of ras p21, FTase has been the target of new anticancer treatment modalities. Famesyl transferase inhibitors (FTIs), a novel class of antitumor drugs that blocks the oncogenic activity of ras, were first developed as therapy for ras-mutated human tumors. FTIs are also capable of inhibiting tumors without ras mutations, while different mechanisms of cell growth inhibition by FTI have been proposed. In cell culture and in animal models, FTIs have been shown to be potent inhibitors of cancer cell growth. We showed that manumycin, a farnesyl pyrophosphate competitive inhibitor, inhibits ovarian and mesothelioma cancer cell growth. To examine the molecular changes after exposure to manumycin, total proteins from treated and untreated 2774 cell line were extracted and separated by two dimensional gel electrophoresis (2-DE) and detected by colloidal coomassie blue staining. The 2-DE was found reliable for comparison of protein profiles before and after manumycin treatment. Two protein spots (spot 1 and 2) appeared after manumycin treatment. Generated peptide peaks were matched by database query (prowl.rockefeller.edu/cgi-bin/ProFound) to a heat shock protein (HSP) and a modified HSP, both with MW 70 KD. The expression status of HSP 70 was confirmed by Western blot and HSP 70 ELISA using an antibody against HSP 70. The expression of HSP70 increased with the length of exposure to, and the dose of manumycin as demonstrated by ELISA. Increase in HSP70 expression was also observed by Western blot after a 48-hour treatment with manumycin in ovarian cancer cell line OVCAR3 and three other cell cultures derived from samples of patients diagnosed with ovarian cancer and mesothelioma. Our study is the first report demonstrating an up-regulation of HSP 70 in ovarian cancer cell lines and mesothelioma cell cultures after treatment with the FTI, manumycin. This up-regulation of HSP 70 may be a cellular mechanism of cell self-protection against the apoptotic process and cell death, and it may enhance resistance and account for the altered sensitivity of certain cancers to FTIs agents. HSPs may therefore be an important molecular target for drug intervention strategies.
Hu
Wei
W
Department of Gynecologic and Medical Oncology, The University of Texas M.D. Anderson Cancer Center, Houston 77030, USA. weihu@mdanderson.org
Wu
Weiguo
W
Yeung
Sai-Ching Jim
SC
Freedman
Ralph S
RS
Kavanagh
John J
JJ
Verschraegen
Claire F
CF
D019883:Alkyl and Aryl Transferases / Q000037:antagonists & inhibitors*; D015153:Blotting, Western; D015180:Electrophoresis, Gel, Two-Dimensional; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004797:Enzyme-Linked Immunosorbent Assay; D051231:Farnesyltranstransferase; D005260:Female; D018840:HSP70 Heat-Shock Proteins / Q000096:biosynthesis*; D006801:Humans; D013058:Mass Spectrometry; D008654:Mesothelioma / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D010051:Ovarian Neoplasms / Q000188:drug therapy / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology; D011090:Polyenes / Q000494:pharmacology*; D053284:Polyunsaturated Alkamides; D014407:Tumor Cells, Cultured; D015854:Up-Regulation / Q000187:drug effects
D004791:Enzyme Inhibitors; D018840:HSP70 Heat-Shock Proteins; D011090:Polyenes; D053284:Polyunsaturated Alkamides; D019883:Alkyl and Aryl Transferases; D051231:Farnesyltranstransferase; C054474:manumycin