title: Reverse transformation of Harvey murine sarcoma virus-transformed NIH/3T3 cells by site-selective cyclic AMP analogs.
abstract: Eighteen site-selective cAMP analogs modified at either the C-8 position or the C-6 position were tested for their growth regulatory effects on the Harvey murine sarcoma virus-transformed NIH/3T3 clone 13-3B-4 cells grown in a serum-free defined medium. All 18 analogs, when tested individually, exhibited an appreciable growth inhibitory effect at micromolar concentrations. The most potent growth inhibitory analogs contained a thio moiety at the C-8 position. In general, C-6 analogs required 5-10-fold greater concentrations than C-8 analogs to produce the same degree of growth inhibition. The growth inhibition induced by these analogs was accompanied by a change in cell morphology; cells treated with the analogs exhibited the morphology characteristic of untransformed fibroblasts, while untreated cells retained a transformed phenotype. The regulatory subunit of cAMP-dependent protein kinase, the cAMP receptor protein, has two different intrachain cAMP binding sites, and cAMP analogs modified at the C-8 position (C-8 analogs) are generally selective for Site 1, while analogs modified at the C-6 position (C-6 analogs) are generally selective for Site 2. Thus, C-8 and C-6 analogs were tested in combination to enhance the growth regulatory effect. Both growth inhibition and morphological change were enhanced synergistically by a combination of the C-6 and C-8 analogs. Two C-6 analogs or two C-8 analogs added together did not cause synergism. For both growth inhibition and phenotypic change, C-8 thio analogs acted far more synergistically than C-8 amino analogs when cells were treated in combination with C-6 analogs, suggesting a response of the RII rather than the RI cAMP receptor protein. DEAE-cellulose chromatography revealed that the growth inhibition, in fact, correlates with an increase of the RII cAMP receptor protein and a decrease of the RI receptor protein. The growth inhibitory effect of the site-selective analogs was not due to the cytotoxic effect of adenosine metabolites as shown by the different behavior of 8-Cl-cAMP compared with 8-Cl-adenosine in 1) cell cycle effects and 2) release from growth inhibition. It is concluded that the observed growth inhibition and phenotypic reversion of 13-3B-4 cells is most likely mediated through the cellular effector, the RII cAMP receptor protein.
authors.affiliation: Cellular Biochemistry Section, National Cancer Institute, Bethesda, Maryland 20892.
authors.forename: P
authors.initials: P
authors.lastname: Tagliaferri
authors.forename: D
authors.initials: D
authors.lastname: Katsaros
authors.forename: T
authors.initials: T
authors.lastname: Clair
authors.forename: L
authors.initials: L
authors.lastname: Neckers
authors.forename: R K
authors.initials: RK
authors.lastname: Robins
authors.forename: Y S
authors.initials: YS
authors.lastname: Cho-Chung
mesh_terms: D000818:Animals; D002471:Cell Transformation, Neoplastic / Q000187:drug effects*; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000031:analogs & derivatives* / Q000494:pharmacology*; D004357:Drug Synergism; D006251:Harvey murine sarcoma virus / Q000235:genetics*; D007700:Kinetics; D051379:Mice; D008815:Mice, Inbred Strains; D009053:Sarcoma Viruses, Murine / Q000235:genetics*; D013329:Structure-Activity Relationship
chemical_list: D000242:Cyclic AMP