Skip to main content
NCBI home page
British Journal of Cancer logoLink to British Journal of Cancer
. 1999 Dec;81(7):1134–1141. doi: 10.1038/sj.bjc.6690820

8-Cl-cAMP antagonizes mitogen-activated protein kinase activation and cell growth stimulation induced by epidermal growth factor

A Budillon 1, E Di Gennaro 1, M Caraglia 2, D Barbarulo 1, A Abbruzzese 2, P Tagliaferri 3,1
PMCID: PMC2374321  PMID: 10584873

Abstract

The growth factor-activated mitogenic pathways are often disregulated in tumour cells and, therefore, they can provide specific molecular targets for novel anti-tumour approaches. 8-Chloro-cAMP (8-Cl-cAMP), a synthetic cAMP analogue, is a novel anti-tumour agent that has recently undergone clinical evaluation. We investigated the effects of 8-CI-cAMP on the epidermal growth factor (EGF)/EGF receptor (EGF-R) signalling in human epidermoid cancer KB cells, which are responsive to the mitogenic stimulus of EGF. We found that the growth-promoting activity of EGF was completely abolished when EGF treatment was performed in combination with 8-CI-cAMP. The inhibition of the EGF-induced proliferation by 8-CI-cAMP was paralleled by the blockade of the EGF-stimulated activation of mitogen-activated protein kinases (MAPK), ERK-1 and ERK-2. Conversely, we found an increase of EGF-R expression and EGF-R tyrosine phosphorylation when KB cells were growth inhibited by 8-Cl-cAMP. Moreover, the activity of Raf-1 and MEK-1 protein kinases, the activators upstream MAPK in the phosphorylation cascade induced by EGF, was not modified in 8-Cl-cAMP-treated cells. We concluded that the impairment of KB cell response to EGF, induced by 8-Cl-cAMP, resides in the specific inhibition of MAPK/ERKs activity while the function of the upstream elements in the EGF-R signalling is preserved. © 1999 Cancer Research Campaign

Keywords: cAMP analogue, protein kinase A, EGF, MAPK

Full Text

The Full Text of this article is available as a PDF (185.4 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aboud-Pirak E., Hurwitz E., Pirak M. E., Bellot F., Schlessinger J., Sela M. Efficacy of antibodies to epidermal growth factor receptor against KB carcinoma in vitro and in nude mice. J Natl Cancer Inst. 1988 Dec 21;80(20):1605–1611. doi: 10.1093/jnci/80.20.1605. [DOI] [PubMed] [Google Scholar]
  2. Bianco C., Tortora G., Baldassarre G., Caputo R., Fontanini G., Chinè S., Bianco A. R., Ciardiello F. 8-Chloro-cyclic AMP inhibits autocrine and angiogenic growth factor production in human colorectal and breast cancer. Clin Cancer Res. 1997 Mar;3(3):439–448. [PubMed] [Google Scholar]
  3. Bosanquet A. G., Burlton A. R., Bell P. B., Harris A. L. Ex vivo cytotoxic drug evaluation by DiSC assay to expedite identification of clinical targets: results with 8-chloro-cAMP. Br J Cancer. 1997;76(4):511–518. doi: 10.1038/bjc.1997.417. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Budillon A., Cereseto A., Kondrashin A., Nesterova M., Merlo G., Clair T., Cho-Chung Y. S. Point mutation of the autophosphorylation site or in the nuclear location signal causes protein kinase A RII beta regulatory subunit to lose its ability to revert transformed fibroblasts. Proc Natl Acad Sci U S A. 1995 Nov 7;92(23):10634–10638. doi: 10.1073/pnas.92.23.10634. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Budillon A., Tagliaferri P., Caraglia M., Torrisi M. R., Normanno N., Iacobelli S., Palmieri G., Stoppelli M. P., Frati L., Bianco A. R. Upregulation of epidermal growth factor receptor induced by alpha-interferon in human epidermoid cancer cells. Cancer Res. 1991 Feb 15;51(4):1294–1299. [PubMed] [Google Scholar]
  6. Caraglia M., Di Gennaro E., Barbarulo D., Marra M., Tagliaferri P., Abbruzzese A., Budillon A. Up-regulated EGF receptors undergo to rapid internalization and ubiquitin-dependent degradation in human cancer cells exposed to 8-Cl-cAMP. FEBS Lett. 1999 Mar 26;447(2-3):203–208. doi: 10.1016/s0014-5793(99)00292-6. [DOI] [PubMed] [Google Scholar]
  7. Caraglia M., Leardi A., Corradino S., Ciardiello F., Budillon A., Guarrasi R., Bianco A. R., Tagliaferri P. alpha-Interferon potentiates epidermal growth factor receptor-mediated effects on human epidermoid carcinoma KB cells. Int J Cancer. 1995 May 4;61(3):342–347. doi: 10.1002/ijc.2910610312. [DOI] [PubMed] [Google Scholar]
  8. Caraglia M., Tagliaferri P., Correale P., Genua G., Pinto A., Del Vecchio S., Esposito G., Bianco A. R. Cytosine arabinoside increases the binding of 125I-labelled epidermal growth factor and 125I-transferrin and enhances the in vitro targeting of human tumour cells with anti-(growth factor receptor) mAb. Cancer Immunol Immunother. 1993 Aug;37(3):150–156. doi: 10.1007/BF01525428. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Cho-Chung Y. S., Pepe S., Clair T., Budillon A., Nesterova M. cAMP-dependent protein kinase: role in normal and malignant growth. Crit Rev Oncol Hematol. 1995 Nov;21(1-3):33–61. doi: 10.1016/1040-8428(94)00166-9. [DOI] [PubMed] [Google Scholar]
  10. Cho-Chung Y. S. Site-selective 8-chloro-cyclic adenosine 3',5'-monophosphate as a biologic modulator of cancer: restoration of normal control mechanisms. J Natl Cancer Inst. 1989 Jul 5;81(13):982–987. doi: 10.1093/jnci/81.13.982. [DOI] [PubMed] [Google Scholar]
  11. Ciardiello F., Damiano V., Bianco R., Bianco C., Fontanini G., De Laurentiis M., De Placido S., Mendelsohn J., Bianco A. R., Tortora G. Antitumor activity of combined blockade of epidermal growth factor receptor and protein kinase A. J Natl Cancer Inst. 1996 Dec 4;88(23):1770–1776. doi: 10.1093/jnci/88.23.1770. [DOI] [PubMed] [Google Scholar]
  12. Cook S. J., McCormick F. Inhibition by cAMP of Ras-dependent activation of Raf. Science. 1993 Nov 12;262(5136):1069–1072. doi: 10.1126/science.7694367. [DOI] [PubMed] [Google Scholar]
  13. Crews C. M., Erikson R. L. Extracellular signals and reversible protein phosphorylation: what to Mek of it all. Cell. 1993 Jul 30;74(2):215–217. doi: 10.1016/0092-8674(93)90411-i. [DOI] [PubMed] [Google Scholar]
  14. Csar X. F., Ward A. C., Hoffmann B. W., Guy G. G., Hamilton J. A. cAMP suppresses p21ras and Raf-1 responses but not the Erk-1 response to granulocyte-colony-stimulating factor: possible Raf-1-independent activation of Erk-1. Biochem J. 1997 Feb 15;322(Pt 1):79–87. doi: 10.1042/bj3220079. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Denhardt D. T. Signal-transducing protein phosphorylation cascades mediated by Ras/Rho proteins in the mammalian cell: the potential for multiplex signalling. Biochem J. 1996 Sep 15;318(Pt 3):729–747. doi: 10.1042/bj3180729. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Fassina G., Aluigi M. G., Gentleman S., Wong P., Cai T., Albini A., Noonan D. M. The cAMP analog 8-Cl-cAMP inhibits growth and induces differentiation and apoptosis in retinoblastoma cells. Int J Cancer. 1997 Sep 17;72(6):1088–1094. doi: 10.1002/(sici)1097-0215(19970917)72:6<1088::aid-ijc25>3.0.co;2-#. [DOI] [PubMed] [Google Scholar]
  17. Faure M., Bourne H. R. Differential effects on cAMP on the MAP kinase cascade: evidence for a cAMP-insensitive step that can bypass Raf-1. Mol Biol Cell. 1995 Aug;6(8):1025–1035. doi: 10.1091/mbc.6.8.1025. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Frödin M., Peraldi P., Van Obberghen E. Cyclic AMP activates the mitogen-activated protein kinase cascade in PC12 cells. J Biol Chem. 1994 Feb 25;269(8):6207–6214. [PubMed] [Google Scholar]
  19. Hordijk P. L., Verlaan I., Jalink K., van Corven E. J., Moolenaar W. H. cAMP abrogates the p21ras-mitogen-activated protein kinase pathway in fibroblasts. J Biol Chem. 1994 Feb 4;269(5):3534–3538. [PubMed] [Google Scholar]
  20. Huang P. S., Heimbrook D. C. Oncogene products as therapeutic targets for cancer. Curr Opin Oncol. 1997 Jan;9(1):94–100. doi: 10.1097/00001622-199701000-00015. [DOI] [PubMed] [Google Scholar]
  21. Jetten A. M. Retinoids specifically enhance the number of epidermal growth factor receptors. Nature. 1980 Apr 17;284(5757):626–629. doi: 10.1038/284626a0. [DOI] [PubMed] [Google Scholar]
  22. Kharbanda S., Saleem A., Emoto Y., Stone R., Rapp U., Kufe D. Activation of Raf-1 and mitogen-activated protein kinases during monocytic differentiation of human myeloid leukemia cells. J Biol Chem. 1994 Jan 14;269(2):872–878. [PubMed] [Google Scholar]
  23. Langdon S. P., Ritchie A. A., Muir M., Dodds M., Howie A. F., Leonard R. C., Stockman P. K., Miller W. R. Antitumour activity and schedule dependency of 8-chloroadenosine-3',5'-monophosphate (8-ClcAMP) against human tumour xenografts. Eur J Cancer. 1998 Feb;34(3):384–388. doi: 10.1016/s0959-8049(97)00372-9. [DOI] [PubMed] [Google Scholar]
  24. Lange-Carter C. A., Vuillequez J. J., Malkinson A. M. 8-Chloroadenosine mediates 8-chloro-cyclic AMP-induced down-regulation of cyclic AMP-dependent protein kinase in normal and neoplastic mouse lung epithelial cells by a cyclic AMP-independent mechanism. Cancer Res. 1993 Jan 15;53(2):393–400. [PubMed] [Google Scholar]
  25. Langeveld C. H., Jongenelen C. A., Theeuwes J. W., Baak J. P., Heimans J. J., Stoof J. C., Peters G. J. The antiproliferative effect of 8-chloro-adenosine, an active metabolite of 8-chloro-cyclic adenosine monophosphate, and disturbances in nucleic acid synthesis and cell cycle kinetics. Biochem Pharmacol. 1997 Jan 24;53(2):141–148. doi: 10.1016/s0006-2952(96)00593-x. [DOI] [PubMed] [Google Scholar]
  26. Marshall C. J. Cell signalling. Raf gets it together. Nature. 1996 Sep 12;383(6596):127–128. doi: 10.1038/383127a0. [DOI] [PubMed] [Google Scholar]
  27. Marshall C. J. MAP kinase kinase kinase, MAP kinase kinase and MAP kinase. Curr Opin Genet Dev. 1994 Feb;4(1):82–89. doi: 10.1016/0959-437x(94)90095-7. [DOI] [PubMed] [Google Scholar]
  28. McKenzie F. R., Pouysségur J. cAMP-mediated growth inhibition in fibroblasts is not mediated via mitogen-activated protein (MAP) kinase (ERK) inhibition. cAMP-dependent protein kinase induces a temporal shift in growth factor-stimulated MAP kinases. J Biol Chem. 1996 Jun 7;271(23):13476–13483. doi: 10.1074/jbc.271.23.13476. [DOI] [PubMed] [Google Scholar]
  29. Miller W. R., Hulme M. J., Cho-Chung Y. S., Elton R. A. Types of cyclic AMP binding proteins in human breast cancers. Eur J Cancer. 1993;29A(7):989–991. doi: 10.1016/s0959-8049(05)80207-2. [DOI] [PubMed] [Google Scholar]
  30. Noguchi K., Murata T., Cho-Chung Y. S. 8-chloroadenosine 3',5'-monophosphate (8-Cl-cAMP) selectively eliminates protein kinase A type I to induce growth inhibition in c-ras-transformed fibroblasts. Eur J Cancer. 1998 Jul;34(8):1260–1267. doi: 10.1016/s0959-8049(98)00051-3. [DOI] [PubMed] [Google Scholar]
  31. Rohlff C., Clair T., Cho-Chung Y. S. 8-Cl-cAMP induces truncation and down-regulation of the RI alpha subunit and up-regulation of the RII beta subunit of cAMP-dependent protein kinase leading to type II holoenzyme-dependent growth inhibition and differentiation of HL-60 leukemia cells. J Biol Chem. 1993 Mar 15;268(8):5774–5782. [PubMed] [Google Scholar]
  32. Saunders M. P., Salisbury A. J., O'Byrne K. J., Long L., Whitehouse R. M., Talbot D. C., Mawer E. B., Harris A. L. A novel cyclic adenosine monophosphate analog induces hypercalcemia via production of 1,25-dihydroxyvitamin D in patients with solid tumors. J Clin Endocrinol Metab. 1997 Dec;82(12):4044–4048. doi: 10.1210/jcem.82.12.4410. [DOI] [PubMed] [Google Scholar]
  33. Scala S., Budillon A., Zhan Z., Cho-Chung Y. S., Jefferson J., Tsokos M., Bates S. E. Downregulation of mdr-1 expression by 8-Cl-cAMP in multidrug resistant MCF-7 human breast cancer cells. J Clin Invest. 1995 Aug;96(2):1026–1034. doi: 10.1172/JCI118088. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Schwartz D. A., Rubin C. S. Identification and differential expression of two forms of regulatory subunits (RII) of cAMP-dependent protein kinase II in Friend erythroleukemic cells. Differentiation and 8-bromo-cAMP elicit a large and selective increase in the rate of biosynthesis of only one type of RII. J Biol Chem. 1985 May 25;260(10):6296–6303. [PubMed] [Google Scholar]
  35. Tagliaferri P., Caraglia M., Muraro R., Pinto A., Budillon A., Zagonel V., Bianco A. R. Pharmacological modulation of peptide growth factor receptor expression on tumor cells as a basis for cancer therapy. Anticancer Drugs. 1994 Aug;5(4):379–393. doi: 10.1097/00001813-199408000-00001. [DOI] [PubMed] [Google Scholar]
  36. Tagliaferri P., Katsaros D., Clair T., Ally S., Tortora G., Neckers L., Rubalcava B., Parandoosh Z., Chang Y. A., Revankar G. R. Synergistic inhibition of growth of breast and colon human cancer cell lines by site-selective cyclic AMP analogues. Cancer Res. 1988 Mar 15;48(6):1642–1650. [PubMed] [Google Scholar]
  37. Tortora G., Damiano V., Bianco C., Baldassarre G., Bianco A. R., Lanfrancone L., Pelicci P. G., Ciardiello F. The RIalpha subunit of protein kinase A (PKA) binds to Grb2 and allows PKA interaction with the activated EGF-receptor. Oncogene. 1997 Feb 27;14(8):923–928. doi: 10.1038/sj.onc.1200906. [DOI] [PubMed] [Google Scholar]
  38. Tortora G., Pepe S., Cirafici A. M., Ciardiello F., Porcellini A., Clair T., Colletta G., Cho-Chung Y. S., Bianco A. R. Thyroid-stimulating hormone-regulated growth and cell cycle distribution of thyroid cells involve type I isozyme of cyclic AMP-dependent protein kinase. Cell Growth Differ. 1993 May;4(5):359–365. [PubMed] [Google Scholar]
  39. Tortora G., di Isernia G., Sandomenico C., Bianco R., Pomatico G., Pepe S., Bianco A. R., Ciardiello F. Synergistic inhibition of growth and induction of apoptosis by 8-chloro-cAMP and paclitaxel or cisplatin in human cancer cells. Cancer Res. 1997 Nov 15;57(22):5107–5111. [PubMed] [Google Scholar]
  40. Tortora G, Ciardiello F, Pepe S, Tagliaferri P, Ruggiero A, Bianco C, Guarrasi R, Miki K, Bianco AR. Phase I clinical study with 8-chloro-cAMP and evaluation of immunological effects in cancer patients. Clin Cancer Res. 1995 Apr;1(4):377–384. [PubMed] [Google Scholar]
  41. Ullrich A., Schlessinger J. Signal transduction by receptors with tyrosine kinase activity. Cell. 1990 Apr 20;61(2):203–212. doi: 10.1016/0092-8674(90)90801-k. [DOI] [PubMed] [Google Scholar]
  42. Vossler M. R., Yao H., York R. D., Pan M. G., Rim C. S., Stork P. J. cAMP activates MAP kinase and Elk-1 through a B-Raf- and Rap1-dependent pathway. Cell. 1997 Apr 4;89(1):73–82. doi: 10.1016/s0092-8674(00)80184-1. [DOI] [PubMed] [Google Scholar]
  43. Withers D. J., Bloom S. R., Rozengurt E. Dissociation of cAMP-stimulated mitogenesis from activation of the mitogen-activated protein kinase cascade in Swiss 3T3 cells. J Biol Chem. 1995 Sep 8;270(36):21411–21419. doi: 10.1074/jbc.270.36.21411. [DOI] [PubMed] [Google Scholar]
  44. Wu J., Dent P., Jelinek T., Wolfman A., Weber M. J., Sturgill T. W. Inhibition of the EGF-activated MAP kinase signaling pathway by adenosine 3',5'-monophosphate. Science. 1993 Nov 12;262(5136):1065–1069. doi: 10.1126/science.7694366. [DOI] [PubMed] [Google Scholar]
  45. Zuckier G., Tritton T. R. Adriamycin causes up regulation of epidermal growth factor receptors in actively growing cells. Exp Cell Res. 1983 Oct;148(1):155–161. doi: 10.1016/0014-4827(83)90195-7. [DOI] [PubMed] [Google Scholar]

Articles from British Journal of Cancer are provided here courtesy of Cancer Research UK

RESOURCES