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Cancer Chemotherapy and Pharmacology

, Volume 63, Issue 5, pp 807–818 | Cite as

Characterization of kinase suppressor of Ras-1 expression and anticancer drug sensitivity in human cancer cell lines

  • Scott M. Stoeger
  • Kenneth H. CowanEmail author
Original Article

Abstract

Previous studies have indicated that the ERK1/2 MAP kinase signaling pathway plays an important role not only in cell growth, cell cycle regulation, and differentiation, but also in determining the sensitivity of cells to anticancer agents as well. Furthermore, expression of kinase suppressor of Ras-1 (KSR1), a molecular scaffold that modulates signaling through the ERK1/2 MAP kinase pathway, has been shown to influence the cellular sensitivity to the anticancer agent cisplatin. To further define the role of KSR1 expression on drug sensitivity, the expression of KSR1 was examined in the NCI60 anticancer drug screen, a panel of cancer cell lines representing nine tissue types, established by the Developmental Therapeutics Program (DTP) at the National Cancer Institute (NCI). The expression of thousands of molecular targets has been examined in the NCI60 panel as well as the cellular toxicity for greater than 400,000 compounds. KSR1 expression varied almost 30-fold difference between the highest and lowest expressing cell lines in the NCI60. Using the COMPARE analysis algorithm, KSR1 expression was correlated with sensitivity of the compounds screened by DTP and several novel agents were identified whose sensitivity correlated with KSR1 expression in the NCI60 panel. Cytotoxicity of two agents, cytochalasin H and tunicamycin, identified through the COMPARE analysis of KSR1 expression and drug sensitivity, was also examined in wild type (KSR+/+) mouse embryo fibroblasts (MEFs) and MEFs deficient in KSR1 expression (KSR1−/−). These studies demonstrated enhanced sensitivity, as well as increased ERK activation, in KSR−/− MEFs following exposure to tunicamycin or cytochalasin H compared to KSR+/+ MEFs. Furthermore, restoration of KSR1 expression in KSR−/− MEFs following stable transduction of cells with a KSR1 expression vector, enhanced sensitivity of cells to tunicamycin and cytochalasin H and decreased ERK1/2 activation following exposure to these drugs. In addition, the sensitivity to cytochalasin H and tunicamycin of breast cancer cell lines with low KSR1 expression, (HS578T and MDA-MB-231/ATCC), was increased relative to the sensitivity of breast cancer cells with higher levels of KSR1 (MCF7). These studies indicate that KSR1 may play an important role in the determination of cellular sensitivity to anticancer agents.

Keywords

KSR1 NCI anticancer drug screen Drug resistance DNA damage Signal transduction 

Notes

Acknowledgments

We would like to thank Dr. Nick Scudiero for the frozen samples of each cell line and the select cell lines described in "Materials and methods", Dr. Robert Schultz for the compounds, and Dr. Susan Holbeck for help with the COMPARE analysis. We would also like to thank the laboratory of Dr. Robert Lewis for supplying the KSR1−/−, KSR1+/+, and the KSR1/GFP MEFs, as well as assistance in the use of the Li-Cor Odyssey system. SMS was supported by a Program of Excellence fellowship from the University of Nebraska Medical Center.

References

  1. 1.
    Allred DC, Clark GM, Tandon AK, Molina R, Tormey DC, Osborne CK, Gilchrist KW, Mansour EG, Abeloff M, Eudey L et al (1992) HER-2/neu in node-negative breast cancer: prognostic significance of overexpression influenced by the presence of in situ carcinoma. J Clin Oncol 10:599–605PubMedGoogle Scholar
  2. 2.
    Alvarez M, Paull K, Monks A, Hose C, Lee JS, Weinstein J, Grever M, Bates S, Fojo T (1995) Generation of a drug resistance profile by quantitation of mdr-1/P-glycoprotein in the cell lines of the National Cancer Institute Anticancer Drug Screen. J Clin Invest 95:2205–2214PubMedCrossRefGoogle Scholar
  3. 3.
    Berton G, Fumagalli L, Laudanna C, Sorio C (1994) Beta 2 integrin-dependent protein tyrosine phosphorylation and activation of the FGR protein tyrosine kinase in human neutrophils. J Cell Biol 126:1111–1121PubMedCrossRefGoogle Scholar
  4. 4.
    Boyd MR (2004) The NCI human tumor cell line (60-cell) screen: concept, implementation, and applications. In: Teicher B, Monks AP (eds) Anticancer drug development guide. Humana Press, Totowa, pp 41–61Google Scholar
  5. 5.
    Burt RK, Garfield S, Johnson K, Thorgeirsson SS (1988) Transformation of rat liver epithelial cells with v-H-ras or v-raf causes expression of MDR-1, glutathione-S-transferase-P and increased resistance to cytotoxic chemicals. Carcinogenesis 9:2329–2332PubMedCrossRefGoogle Scholar
  6. 6.
    Cacace AM, Michaud NR, Therrien M, Mathes K, Copeland T, Rubin GM, Morrison DK (1999) Identification of constitutive and ras-inducible phosphorylation sites of KSR: implications for 14-3-3 binding, mitogen-activated protein kinase binding, and KSR overexpression. Mol Cell Biol 19:229–240PubMedGoogle Scholar
  7. 7.
    Carlson BA, Dubay MM, Sausville EA, Brizuela L, Worland PJ (1996) Flavopiridol induces G1 arrest with inhibition of cyclin-dependent kinase (CDK) 2 and CDK4 in human breast carcinoma cells. Cancer Res 56:2973–2978PubMedGoogle Scholar
  8. 8.
    Continolo S, Baruzzi A, Majeed M, Caveggion E, Fumagalli L, Lowell CA, Berton G (2005) The proto-oncogene Fgr regulates cell migration and this requires its plasma membrane localization. Exp Cell Res 302:253–269PubMedCrossRefGoogle Scholar
  9. 9.
    Dickstein B, Valverius EM, Wosikowski K, Saceda M, Pearson JW, Martin MB, Bates SE (1993) Increased epidermal growth factor receptor in an estrogen-responsive, adriamycin-resistant MCF-7 cell line. J Cell Physiol 157:110–118PubMedCrossRefGoogle Scholar
  10. 10.
    Dickstein BM, Wosikowski K, Bates SE (1995) Increased resistance to cytotoxic agents in ZR75B human breast cancer cells transfected with epidermal growth factor receptor. Mol Cell Endocrinol 110:205–211PubMedCrossRefGoogle Scholar
  11. 11.
    Gee JM, Robertson JF, Ellis IO, Nicholson RI (2001) Phosphorylation of ERK1/2 mitogen-activated protein kinase is associated with poor response to anti-hormonal therapy and decreased patient survival in clinical breast cancer. Int J Cancer 95:247–254PubMedCrossRefGoogle Scholar
  12. 12.
    Gerlier D, Thomasset N (1986) Use of MTT colorimetric assay to measure cell activation. J Immunol Methods 94:57–63PubMedCrossRefGoogle Scholar
  13. 13.
    Gusterson BA, Gelber RD, Goldhirsch A, Price KN, Save-Soderborgh J, Anbazhagan R, Styles J, Rudenstam CM, Golouh R, Reed R et al (1992) Prognostic importance of c-erbB-2 expression in breast cancer. International (Ludwig) breast cancer study group. J Clin Oncol 10:1049–1056PubMedGoogle Scholar
  14. 14.
    Hagan MP, Yacoub A, Dent P (2007) Radiation-induced PARP activation is enhanced through EGFR-ERK signaling. J Cell Biochem 12:12Google Scholar
  15. 15.
    Hu P, Han Z, Couvillon AD, Exton JH (2004) Critical role of endogenous Akt/IAPs and MEK1/ERK pathways in counteracting endoplasmic reticulum stress-induced cell death. J Biol Chem 279:49420–49429 (Epub 2004 Aug 31)PubMedCrossRefGoogle Scholar
  16. 16.
    Huang C, Jacobson K, Schaller MD (2004) MAP kinases and cell migration. J Cell Sci 117:4619–4628PubMedCrossRefGoogle Scholar
  17. 17.
    Hung CC, Ichimura T, Stevens JL, Bonventre JV (2003) Protection of renal epithelial cells against oxidative injury by endoplasmic reticulum stress preconditioning is mediated by ERK1/2 activation. J Biol Chem 278:29317–29326 (Epub May 8 2003)PubMedCrossRefGoogle Scholar
  18. 18.
    Jiang CC, Chen LH, Gillespie S, Wang YF, Kiejda KA, Zhang XD, Hersey P (2007) Inhibition of MEK sensitizes human melanoma cells to endoplasmic reticulum stress-induced apoptosis. Cancer Res 67:9750–9761PubMedCrossRefGoogle Scholar
  19. 19.
    Kaminski WE, Piehler A, Wenzel JJ (2006) ABC A-subfamily transporters: structure, function and disease. Biochim Biophys Acta 1762:510–524 (Epub 2006 Feb 28)PubMedGoogle Scholar
  20. 20.
    Katagiri K, Matsuura S (1971) Antitumor activity of cytochalasin D. J Antibiot (Tokyo) 24:722–723Google Scholar
  21. 21.
    Kelsell DP, Norgett EE, Unsworth H, Teh MT, Cullup T, Mein CA, Dopping-Hepenstal PJ, Dale BA, Tadini G, Fleckman P, Stephens KG, Sybert VP, Mallory SB, North BV, Witt DR, Sprecher E, Taylor AE, Ilchyshyn A, Kennedy CT, Goodyear H, Moss C, Paige D, Harper JI, Young BD, Leigh IM, Eady RA, O’Toole EA (2005) Mutations in ABCA12 underlie the severe congenital skin disease harlequin ichthyosis. Am J Hum Genet 76:794–803 (Epub 2005 Mar 8)PubMedCrossRefGoogle Scholar
  22. 22.
    Kim M, Yan Y, Kortum RL, Stoeger SM, Sgagias MK, Lee K, Lewis RE, Cowan KH (2005) Expression of kinase suppressor of Ras1 enhances cisplatin-induced extracellular signal-regulated kinase activation and cisplatin sensitivity. Cancer Res 65:3986–3992PubMedCrossRefGoogle Scholar
  23. 23.
    Kim YK, Kim HJ, Kwon CH, Kim JH, Woo JS, Jung JS, Kim JM (2005) Role of ERK activation in cisplatin-induced apoptosis in OK renal epithelial cells. J Appl Toxicol 25:374–382PubMedCrossRefGoogle Scholar
  24. 24.
    Koo HM, Monks A, Mikheev A, Rubinstein LV, Gray-Goodrich M, McWilliams MJ, Alvord WG, Oie HK, Gazdar AF, Paull KD, Zarbl H, Vande Woude GF (1996) Enhanced sensitivity to 1-beta-D-arabinofuranosylcytosine and topoisomerase II inhibitors in tumor cell lines harboring activated ras oncogenes. Cancer Res 56:5211–5216PubMedGoogle Scholar
  25. 25.
    Kornfeld K, Hom DB, Horvitz HR (1995) The ksr-1 gene encodes a novel protein kinase involved in Ras-mediated signaling in C. elegans. Cell 83:903–913PubMedCrossRefGoogle Scholar
  26. 26.
    Kortum RL, Costanzo DL, Haferbier J, Schreiner SJ, Razidlo GL, Wu MH, Volle DJ, Mori T, Sakaue H, Chaika NV, Chaika OV, Lewis RE (2005) The molecular scaffold kinase suppressor of Ras 1 (KSR1) regulates adipogenesis. Mol Cell Biol 25:7592–7604PubMedCrossRefGoogle Scholar
  27. 27.
    Kortum RL, Johnson HJ, Costanzo DL, Volle DJ, Razidlo GL, Fusello AM, Shaw AS, Lewis RE (2006) The molecular scaffold kinase suppressor of Ras 1 is a modifier of RasV12-induced and replicative senescence. Mol Cell Biol 26:2202–2214PubMedCrossRefGoogle Scholar
  28. 28.
    Kortum RL, Lewis RE (2004) The molecular scaffold KSR1 regulates the proliferative and oncogenic potential of cells. Mol Cell Biol 24:4407–4416PubMedCrossRefGoogle Scholar
  29. 29.
    Kurokawa H, Lenferink AE, Simpson JF, Pisacane PI, Sliwkowski MX, Forbes JT, Arteaga CL (2000) Inhibition of HER2/neu (erbB-2) and mitogen-activated protein kinases enhances tamoxifen action against HER2-overexpressing, tamoxifen-resistant breast cancer cells. Cancer Res 60:5887–5894PubMedGoogle Scholar
  30. 30.
    Lefevre C, Audebert S, Jobard F, Bouadjar B, Lakhdar H, Boughdene-Stambouli O, Blanchet-Bardon C, Heilig R, Foglio M, Weissenbach J, Lathrop M, Prud’homme JF, Fischer J (2003) Mutations in the transporter ABCA12 are associated with lamellar ichthyosis type 2. Hum Mol Genet 12:2369–2378 (Epub 2003 Jul 15)PubMedCrossRefGoogle Scholar
  31. 31.
    Lozano J, Xing R, Cai Z, Jensen HL, Trempus C, Mark W, Cannon R, Kolesnick R (2003) Deficiency of kinase suppressor of Ras1 prevents oncogenic ras signaling in mice. Cancer Res 63:4232–4238PubMedGoogle Scholar
  32. 32.
    Matheny SA, Chen C, Kortum RL, Razidlo GL, Lewis RE, White MA (2004) Ras regulates assembly of mitogenic signalling complexes through the effector protein IMP. Nature 427:256–260PubMedCrossRefGoogle Scholar
  33. 33.
    McCubrey JA, Steelman LS, Chappell WH, Abrams SL, Wong EW, Chang F, Lehmann B, Terrian DM, Milella M, Tafuri A, Stivala F, Libra M, Basecke J, Evangelisti C, Martelli AM, Franklin RA (2006) Roles of the Raf/MEK/ERK pathway in cell growth, malignant transformation and drug resistance. Biochim Biophys Acta 7:7Google Scholar
  34. 34.
    Miyata Y (2005) Hsp90 inhibitor geldanamycin and its derivatives as novel cancer chemotherapeutic agents. Curr Pharm Des 11:1131–1138PubMedCrossRefGoogle Scholar
  35. 35.
    Monks A, Scudiero D, Skehan P, Shoemaker R, Paull K, Vistica D, Hose C, Langley J, Cronise P, Vaigro-Wolff A et al (1991) Feasibility of a high-flux anticancer drug screen using a diverse panel of cultured human tumor cell lines. J Natl Cancer Inst 83:757–766PubMedCrossRefGoogle Scholar
  36. 36.
    Morrison DK (2001) KSR: a MAPK scaffold of the Ras pathway? J Cell Sci 114:1609–1612PubMedGoogle Scholar
  37. 37.
    Moscow J, Schneider E, Sikic B, Morrow C, Cowan K (2006) Drug resistance and its clinical circumvention. In: Kufe D, Bast R, Hait W, Hong W, Pollock R, Weichselbaum R, Holland J, Frei E (eds) Cancer medicine, vol 7. BC Decker Inc, Hamilton, pp 630–647Google Scholar
  38. 38.
    Moscow JA, Connolly T, Myers TG, Cheng CC, Paull K, Cowan KH (1997) Reduced folate carrier gene (RFC1) expression and anti-folate resistance in transfected and non-selected cell lines. Int J Cancer 72:184–190PubMedCrossRefGoogle Scholar
  39. 39.
    Muller J, Ory S, Copeland T, Piwnica-Worms H, Morrison DK (2001) C-TAK1 regulates Ras signaling by phosphorylating the MAPK scaffold, KSR1. Mol Cell 8:983–993PubMedCrossRefGoogle Scholar
  40. 40.
    Natarajan P, May JA, Sanderson HM, Zabe M, Spangenberg P, Heptinstall S (2000) Effects of cytochalasin H, a potent inhibitor of cytoskeletal reorganisation, on platelet function. Platelets 11:467–476PubMedCrossRefGoogle Scholar
  41. 41.
    Nguyen A, Burack WR, Stock JL, Kortum R, Chaika OV, Afkarian M, Muller WJ, Murphy KM, Morrison DK, Lewis RE, McNeish J, Shaw AS (2002) Kinase suppressor of Ras (KSR) is a scaffold which facilitates mitogen-activated protein kinase activation in vivo. Mol Cell Biol 22:3035–3045PubMedCrossRefGoogle Scholar
  42. 42.
    Nguyen DT, Kebache S, Fazel A, Wong HN, Jenna S, Emadali A, Lee EH, Bergeron JJ, Kaufman RJ, Larose L, Chevet E (2004) Nck-dependent activation of extracellular signal-regulated kinase-1 and regulation of cell survival during endoplasmic reticulum stress. Mol Biol Cell 15:4248–4260 (Epub 2004 Jun 16)PubMedCrossRefGoogle Scholar
  43. 43.
    Paull KD, Shoemaker RH, Hodes L, Monks A, Scudiero DA, Rubinstein L, Plowman J, Boyd MR (1989) Display and analysis of patterns of differential activity of drugs against human tumor cell lines: development of mean graph and COMPARE algorithm. J Natl Cancer Inst 81:1088–1092PubMedCrossRefGoogle Scholar
  44. 44.
    Persons DL, Yazlovitskaya EM, Cui W, Pelling JC (1999) Cisplatin-induced activation of mitogen-activated protein kinases in ovarian carcinoma cells: inhibition of extracellular signal-regulated kinase activity increases sensitivity to cisplatin. Clin Cancer Res 5:1007–1014PubMedGoogle Scholar
  45. 45.
    Persons DL, Yazlovitskaya EM, Pelling JC (2000) Effect of extracellular signal-regulated kinase on p53 accumulation in response to cisplatin. J Biol Chem 275:35778–35785PubMedCrossRefGoogle Scholar
  46. 46.
    Razidlo GL, Kortum RL, Haferbier JL, Lewis RE (2004) Phosphorylation regulates KSR1 stability, ERK activation, and cell proliferation. J Biol Chem 279:47808–47814 (Epub 2004 Sep 13)PubMedCrossRefGoogle Scholar
  47. 47.
    Rubinstein LV, Shoemaker RH, Paull KD, Simon RM, Tosini S, Skehan P, Scudiero DA, Monks A, Boyd MR (1990) Comparison of in vitro anticancer-drug-screening data generated with a tetrazolium assay versus a protein assay against a diverse panel of human tumor cell lines. J Natl Cancer Inst 82:1113–1118PubMedCrossRefGoogle Scholar
  48. 48.
    Sausville EA, Arbuck SG, Messmann R, Headlee D, Bauer KS, Lush RM, Murgo A, Figg WD, Lahusen T, Jaken S, Jing X, Roberge M, Fuse E, Kuwabara T, Senderowicz AM (2001) Phase I trial of 72-hour continuous infusion UCN-01 in patients with refractory neoplasms. J Clin Oncol 19:2319–2333PubMedGoogle Scholar
  49. 49.
    Shoemaker RH (2006) The NCI60 human tumour cell line anticancer drug screen. Nat Rev Cancer 6:813–823PubMedCrossRefGoogle Scholar
  50. 50.
    Steelman LS, Pohnert SC, Shelton JG, Franklin RA, Bertrand FE, McCubrey JA (2004) JAK/STAT, Raf/MEK/ERK, PI3 K/Akt and BCR-ABL in cell cycle progression and leukemogenesis. Leukemia 18:189–218PubMedCrossRefGoogle Scholar
  51. 51.
    Sundaram M, Han M (1995) The C. elegans ksr-1 gene encodes a novel Raf-related kinase involved in Ras-mediated signal transduction. Cell 83:889–901PubMedCrossRefGoogle Scholar
  52. 52.
    Szakacs G, Annereau JP, Lababidi S, Shankavaram U, Arciello A, Bussey KJ, Reinhold W, Guo Y, Kruh GD, Reimers M, Weinstein JN, Gottesman MM (2004) Predicting drug sensitivity and resistance: profiling ABC transporter genes in cancer cells. Cancer Cell 6:129–137PubMedCrossRefGoogle Scholar
  53. 53.
    Szegezdi E, Fitzgerald U, Samali A (2003) Caspase-12 and ER-stress-mediated apoptosis: the story so far. Ann N Y Acad Sci 1010:186–194PubMedCrossRefGoogle Scholar
  54. 54.
    Therrien M, Chang HC, Solomon NM, Karim FD, Wassarman DA, Rubin GM (1995) KSR, a novel protein kinase required for RAS signal transduction. Cell 83:879–888PubMedCrossRefGoogle Scholar
  55. 55.
    Vicentini L, Mazzi P, Caveggion E, Continolo S, Fumagalli L, Lapinet-Vera JA, Lowell CA, Berton G (2002) Fgr deficiency results in defective eosinophil recruitment to the lung during allergic airway inflammation. J Immunol 168:6446–6454PubMedGoogle Scholar
  56. 56.
    Wang X, Martindale JL, Holbrook NJ (2000) Requirement for ERK activation in cisplatin-induced apoptosis. J Biol Chem 275:39435–39443PubMedCrossRefGoogle Scholar
  57. 57.
    Wang X, Studzinski GP (2001) Phosphorylation of raf-1 by kinase suppressor of ras is inhibited by “MEK-specific” inhibitors PD 098059 and U0126 in differentiating HL60 cells. Exp Cell Res 268:294–300PubMedCrossRefGoogle Scholar
  58. 58.
    Wang X, Studzinski GP (2004) Kinase suppressor of RAS (KSR) amplifies the differentiation signal provided by low concentrations 1, 25-dihydroxyvitamin D3. J Cell Physiol 198:333–342PubMedCrossRefGoogle Scholar
  59. 59.
    Wang X, Wang TT, White JH, Studzinski GP (2006) Induction of kinase suppressor of RAS-1(KSR-1) gene by 1, alpha25-dihydroxyvitamin D3 in human leukemia HL60 cells through a vitamin D response element in the 5′-flanking region. Oncogene. 25:7078–7085 (Epub 2006 May 29)PubMedCrossRefGoogle Scholar
  60. 60.
    Wei SQ, Sui LH, Zheng JH, Zhang GM, Kao YL (2004) Role of ERK1/2 kinase in cisplatin-induced apoptosis in human ovarian carcinoma cells. Chin Med Sci J 19:125–129PubMedGoogle Scholar
  61. 61.
    Wells JM, Cutler HG, Cole RJ (1976) Toxicity and plant growth regulator effects of cytochalasin H isolated from Phomopsis sp. Can J Microbiol 22:1137–1143PubMedCrossRefGoogle Scholar
  62. 62.
    Xing H, Kornfeld K, Muslin AJ (1997) The protein kinase KSR interacts with 14–3-3 protein and Raf. Curr Biol 7:294–300PubMedCrossRefGoogle Scholar
  63. 63.
    Xing HR, Cordon-Cardo C, Deng X, Tong W, Campodonico L, Fuks Z, Kolesnick R (2003) Pharmacologic inactivation of kinase suppressor of ras-1 abrogates Ras-mediated pancreatic cancer. Nat Med 9:1266–1268 (Epub 2003 Sep 7)PubMedCrossRefGoogle Scholar
  64. 64.
    Yahara I, Harada F, Sekita S, Yoshihira K, Natori S (1982) Correlation between effects of 24 different cytochalasins on cellular structures and cellular events and those on actin in vitro. J Cell Biol 92:69–78PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2008

Authors and Affiliations

  1. 1.Eppley Institute for Research in Cancer and Allied Diseases, 986805 Nebraska Medical CenterUniversity of Nebraska Medical CenterOmahaUSA

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