Pınar OBAKAN, Gizem ALKURT, Betsi KÖSE, Ajda ÇOKER GÜRKAN, Elif Damla ARISAN, Deniz COŞKUN, Zeynep Narçin ÜNSAL

Downregulation of c-Myc mediated ODC expression after purvalanol treatment is under control of upstream MAPK signaling axis in MCF-7 breast cancer cells

Roscovitine and purvalanol are specific cyclin-dependent kinase (CDK) inhibitors, which induce apoptosis by triggering cell cycle arrest in various cancer cells such as colon, prostate, and breast cancer cells. Although the apoptotic action of roscovitine was clarified at the molecular level, the exact mechanism of purvalanol-induced apoptosis is still under investigation. The mitogen-activated protein kinase (MAPK) signaling cascade is activated by different inducers related to growth, proliferation, differentiation processes, or environmental stress factors. Recent reports showed that modulation of MAPKs might lead to regulation of c-Myc, which is a transcription factor for the polyamine (PA) biosynthesis enzyme, ornithine decarboxylase (ODC). PAs are amine-derived cationic molecules that play crucial roles in cell proliferation, growth, and differentiation. In this study, we investigated the potential role of the MAPK signaling cascade in the purvalanol-induced apoptosis mechanism by comparing the results of roscovitine in MCF-7 and MDA-MB-231 breast cancer cells. We found that CDK inhibitors decreased the cell viability in a dose- and time-dependent manner in MCF-7 and MDA-MB-231 cancer cells. Although both CDK inhibitors induced cell cycle arrest, which led to apoptosis by activating caspases and PARP cleavage in MCF-7 breast cancer cells, the apoptotic effect of purvalanol was less than that of roscovitine in MDA-MB-231 cells. Inhibition of MAPKs prevented CDK inhibitor-induced cell viability loss in both cell lines. We determined that purvalanol downregulated c-Myc and ODC expression levels, which led to sharp decrease in the PA pool in MCF-7 cells. On the contrary, purvalanol did not significantly alter c-Myc expression levels, which led to de novo biosynthesis of ODC in a time-dependent manner in MDA-MB-231 cells. Therefore, we suggest that a purvalanol-mediated resistance phenotype might be a possible outcome of c-Myc-mediated ODC expression level in MDA-MB-231 cells.

Anahtar Kelimeler:

Breast cancer, cyclin-dependent kinase inhibitors, purvalanol, mitogen-activated protein kinases, polyamines, apoptosis, c-Myc

Downregulation of c-Myc mediated ODC expression after purvalanol treatment is under control of upstream MAPK signaling axis in MCF-7 breast cancer cells

Keywords:

Breast cancer, cyclin-dependent kinase inhibitors, purvalanol, mitogen-activated protein kinases, polyamines, apoptosis, c-Myc,

PDF

___

Arellano M, Moreno S (1997). Regulation of CDK/cyclin complexes during the cell cycle. Int J Biochem Cell Biol 29: 559–573.
Arisan ED, Obakan P, Coker A, Palavan-Unsal N (2012). Inhibition of ornithine decarboxylase alters the roscovitine-induced mi- tochondrial-mediated apoptosis in MCF-7 breast cancer cells. Mol Med Rep 5: 1323–1329.
Arisan ED, Obakan P, Coker-Gurkan A, Calcabrini A, Agostinelli E, Unsal NP (2014). CDK inhibitors induce mitochondria-medi- ated apoptosis through the activation of polyamine catabolic pathway in LNCaP, DU145 and PC3 prostate cancer cells. Curr Pharm Des 20: 180–188.
Bachrach U (2010). The early history of polyamine research. Plant Physiol Biochem 48: 490–495.
Besson A, Dowdy SF, Roberts JM (2008). CDK inhibitors: cell cycle regulators and beyond. Dev Cell 14: 159–169.
Bodur C, Kutuk O, Karsli-Uzunbas G, Isimjan TT, Harrison P, Ba- saga H (2013). Pramanicin analog induces apoptosis in human colon cancer cells: critical roles for Bcl-2, Bim, and p38 MAPK signaling. PLoS One 8: e56369.
Brown L, Benchimol S (2006). The involvement of MAPK signaling pathways in determining the cellular response to p53 activa- tion: cell cycle arrest or apoptosis. J Biol Chem 281: 3832–3840.
Chen YR, Wang X, Templeton D, Davis RJ, Tan TH (1996). The role of c-Jun N-terminal kinase (JNK) in apoptosis induced by ul- traviolet C and gamma radiation. Duration of JNK activation may determine cell death and proliferation. J Biol Chem 271: 31929–31936.
Coller HA, Grandori C, Tamayo P, Colbert T, Lander ES, Eisenman RN, Golub TR (2000). Expression analysis with oligonucle- otide microarrays reveals that MYC regulates genes involved in growth, cell cycle, signaling, and adhesion. P Natl Acad Sci USA 97: 3260–3265.
Dang CV (1999). c-Myc target genes involved in cell growth, apopto- sis, and metabolism. Mol Cell Biol 19: 1–11.
Deacon K, Mistry P, Chernoff J, Blank JL, Patel R (2003). p38 Mito- gen-activated protein kinase mediates cell death and p21-acti- vated kinase mediates cell survival during chemotherapeutic drug-induced mitotic arrest. Mol Biol Cell 14: 2071–2087.
Deng YT, Huang HC, Lin JK (2010). Rotenone induces apoptosis in MCF-7 human breast cancer cell-mediated ROS through JNK and p38 signaling. Mol Carcinog 49: 141–151.
Evan G, Littlewood T (1998). A matter of life and cell death. Science 281: 1317–1322.
Frey RS, Singletary KW (2003). Genistein activates p38 mitogen-ac- tivated protein kinase, inactivates ERK1/ERK2 and decreases Cdc25C expression in immortalized human mammary epithe- lial cells. J Nutr 133: 226–231.
Gan FY, Gesell MS, Alousi M, Luk GD (1993). Analysis of ODC and c-myc gene expression in hepatocellular carcinoma by in situ hybridization and immunohistochemistry. J Histochem Cyto- chem 41: 1185–1196.
Goodyear S, Sharma MC (2007). Roscovitine regulates invasive breast cancer cell (MDA-MB231) proliferation and survival through cell cycle regulatory protein cdk5. Exp Mol Pathol 82: 25–32.
Gurkan AC, Arisan ED, Obakan P, Palavan-Unsal N (2013). Inhibi- tion of polyamine oxidase prevented cyclin-dependent kinase inhibitor-induced apoptosis in HCT 116 colon carcinoma cells. Apoptosis 18: 1536–1547.
Haan C, Behrmann I (2007). A cost effective non-commercial ECL- solution for Western blot detections yielding strong signals and low background. J Immunol Methods 318: 11–19.
Hahntow IN, Schneller F, Oelsner M, Weick K, Ringshausen I, Fend F, Peschel C, Decker T (2004). Cyclin-dependent kinase inhibi- tor roscovitine induces apoptosis in chronic lymphocytic leu- kemia cells. Leukemia 18: 747–755.
Hanson KD, Shichiri M, Follansbee MR, Sedivy JM (1994). Effects of c-myc expression on cell cycle progression. Mol Cell Biol 14: 5748–5755.
Heby O, Persson L (1990). Molecular genetics of polyamine synthesis in eukaryotic cells. Trends Biochem Sci 15: 153–158.
Henfling ME, Ramaekers FC, Schutte B (2004). Proteasomes act in the pre-mitochondrial signal transduction route towards roscovitine-induced apoptosis. Int J Oncol 25: 1437–1446.
Henriksson M, Luscher B (1996). Proteins of the Myc network: es- sential regulators of cell growth and differentiation. Adv Can- cer Res 68: 109–182.
Hsieh WC, Hsu PC, Liao YF, Young ST, Wang ZW, Lin CL, Tsay GJ, Lee H, Hung HC, Liu GY (2010). Overexpression of ornithine decarboxylase suppresses thapsigargin-induced apoptosis. Mol Cells 30: 311–318.
Hsu PC, Hung HC, Liao YF, Liu CC, Tsay GJ, Liu GY (2008). Orni- thine decarboxylase attenuates leukemic chemotherapy drugs- induced cell apoptosis and arrest in human promyelocytic HL- 60 cells. Leuk Res 32: 1530–1540.
Hsu YL, Kuo PL, Lin LT, Lin CC (2005). Asiatic acid, a triterpene, induces apoptosis and cell cycle arrest through activation of ex- tracellular signal-regulated kinase and p38 mitogen-activated protein kinase pathways in human breast cancer cells. J Phar- macol Exp Ther 313: 333–344.
Johnson GL, Lapadat R (2002). Mitogen-activated protein kinase pathways mediated by ERK, JNK, and p38 protein kinases. Sci- ence 298: 1911–1912.
Kahana C, Reiss Y (2005). Cell-free assay for ubiquitin-independent proteasomal protein degradation. Methods Mol Biol 301: 83– 96.
Komina O, Nosske E, Maurer M, Wesierska-Gadek J (2011). Rosco- vitine, a small molecule CDK inhibitor induces apoptosis in multidrug-resistant human multiple myeloma cells. J Exp Ther Oncol 9: 27–35.
Meijer L (1996). Chemical inhibitors of cyclin-dependent kinases. Trends Cell Biol 6: 393–397.
Mgbonyebi OP, Russo J, Russo IH (1999). Roscovitine induces cell death and morphological changes indicative of apoptosis in MDA-MB-231 breast cancer cells. Cancer Res 59: 1903–1910.
Morgan DO, Fisher RP, Espinoza FH, Farrell A, Nourse J, Chamber- lin H, Jin P (1998). Control of eukaryotic cell cycle progression by phosphorylation of cyclin-dependent kinases. Cancer J Sci Am 4 (Suppl. 1): S77-83.
Noguchi K, Yamana H, Kitanaka C, Mochizuki T, Kokubu A, Kuchi- no Y (2000). Differential role of the JNK and p38 MAPK path- way in c-Myc- and s-Myc-mediated apoptosis. Biochem Bio- phys Res Commun 267: 221–227.
Obakan P, Arisan ED, Ozfiliz P, Coker-Gurkan A, Palavan-Unsal N (2014). Purvalanol A is a strong apoptotic inducer via activat- ing polyamine catabolic pathway in MCF-7 estrogen receptor positive breast cancer cells. Mol Biol Rep 41: 145–154.
Paulovich AG, Toczyski DP, Hartwell LH (1997). When checkpoints fail. Cell 88: 315–321.
Pearson G, Robinson F, Beers Gibson T, Xu BE, Karandikar M, Ber- man K, Cobb MH (2001). Mitogen-activated protein (MAP) kinase pathways: regulation and physiological functions. En- docr Rev 22: 153–183.
Ravanko K, Jarvinen K, Paasinen-Sohns A, Holtta E (2000). Loss of p27Kip1 from cyclin E/cyclin-dependent kinase (CDK) 2 but not from cyclin D1/CDK4 complexes in cells transformed by polyamine biosynthetic enzymes. Cancer Res 60: 5244–5253.
Ringer L, Sirajuddin P, Yenugonda VM, Ghosh A, Divito K, Trabosh V, Patel Y, Brophy A, Grindrod S, Lisanti MP et al. (2010). VMY-1-103, a dansylated analog of purvalanol B, induces cas- pase-3-dependent apoptosis in LNCaP prostate cancer cells. Cancer Biol Ther 10: 320–325.
Sarfaraz S, Siddiqui IA, Syed DN, Afaq F, Mukhtar H (2008). Gug- gulsterone modulates MAPK and NF-kappaB pathways and inhibits skin tumorigenesis in SENCAR mice. Carcinogenesis 29: 2011–2018.
Siepi F, Gatti V, Camerini S, Crescenzi M, Soddu S (2013). HIPK2 catalytic activity and subcellular localization are regulated by activation-loop Y354 autophosphorylation. Biochim Biophys Acta 1833: 1443–1453.
Troppmair J, Bruder JT, Munoz H, Lloyd PA, Kyriakis J, Banerjee P, Avruch J, Rapp UR (1994). Mitogen-activated protein kinase/ extracellular signal-regulated protein kinase activation by on- cogenes, serum, and 12-O-tetradecanoylphorbol-13-acetate requires Raf and is necessary for transformation. J Biol Chem 269: 7030–7035.
Villerbu N, Gaben AM, Redeuilh G, Mester J (2002). Cellular effects of purvalanol A: a specific inhibitor of cyclin-dependent kinase activities. Int J Cancer 97: 761–769.
Walhout AJ, Gubbels JM, Bernards R, van der Vliet PC, Timmers HT (1997). c-Myc/Max heterodimers bind cooperatively to the E-box sequences located in the first intron of the rat ornithine decarboxylase (ODC) gene. Nucleic Acids Res 25: 1493–1501.
Wallace HM, Duthie J, Evans DM, Lamond S, Nicoll KM, Heys SD (2000). Alterations in polyamine catabolic enzymes in human breast cancer tissue. Clin Cancer Res 6: 3657–3661.
Wallace HM, Fraser AV (2004). Inhibitors of polyamine metabolism: review article. Amino Acids 26: 353–365.
Wang X, Feith DJ, Welsh P, Coleman CS, Lopez C, Woster PM, O’Brien TG, Pegg AE (2007). Studies of the mechanism by which increased spermidine/spermine N1-acetyltransferase activity increases susceptibility to skin carcinogenesis. Carci- nogenesis 28: 2404–2411.
Wang X, Martindale JL, Holbrook NJ (2000). Requirement for ERK activation in cisplatin-induced apoptosis. J Biol Chem 275: 39435–39443.
Wesierska-Gadek J, Gueorguieva M, Wojciechowski J, Horky M (2004). Cell cycle arrest induced in human breast cancer cells by cyclin-dependent kinase inhibitors: a comparison of the ef- fects exerted by roscovitine and olomoucine. Pol J Pharmacol 56: 635–641.
Wesierska-Gadek J, Schmitz ML, Ranftler C (2007). Roscovitine- activated HIP2 kinase induces phosphorylation of wt p53 at Ser-46 in human MCF-7 breast cancer cells. J Cell Biochem 100: 865–874.
Wu CL, Liao YF, Hung YC, Lu KH, Hung HC, Liu GY (2011). Or- nithine decarboxylase prevents dibenzoylmethane-induced apoptosis through repressing reactive oxygen species genera- tion. J Biochem Mol Toxicol 25: 312–319.
Xia Z, Dickens M, Raingeaud J, Davis RJ, Greenberg ME (1995). Op- posing effects of ERK and JNK-p38 MAP kinases on apoptosis. Science 270: 1326–1331.
Zhang T, Jiang T, Zhang F, Li C, Zhou YA, Zhu YF, Li XF (2010). Involvement of p21Waf1/Cip1 cleavage during roscovitine- induced apoptosis in non-small cell lung cancer cells. Oncol Rep 23: 239–245.
Zhang Z, Leonard SS, Huang C, Vallyathan V, Castranova V, Shi X (2003). Role of reactive oxygen species and MAPKs in vana- date-induced G(2)/M phase arrest. Free Radic Biol Med 34: 1333–1342.