Skip to main content

Advertisement

SpringerLink
Log in

Temozolomide may induce cell cycle arrest by interacting with URG4/URGCP in SH-SY5Y neuroblastoma cells

  • Research Article
  • Published:
Tumor Biology

Abstract

Temozolomide (TMZ) is an alkylating drug used usually in glioma treatment by inducing the apoptosis in glioma cell. The aim of the study is to investigate the anticancer mechanism of TMZ in SH-SY5Y human neuroblastoma cell line. Cytotoxic effects of TMZ were determined by using XTT assay. IC50 doses in the SH-SY5Y were detected as 5 mM. Expression profiles of novel genes URG4/URGCP, CCND1, CCND2, CDK4, and BCL2 were determined by real-time PCR. The apoptotic effects of TMZ were evaluated with TUNEL method. Furthermore, effects of TMZ on colony formation and invasion were investigated in this study. It was observed that TMZ in SH-SY5Y cell line caused a significant decrease in the gene expressions of URG4/URGCP, CCND1, CCND2, CDK4, and BCL2. According to TUNEL assay results, TMZ markedly induced apoptosis in SH-SY5Y cell line. It was found that TMZ in SH-SY5Y cell line suppressed invasion and colony formation using matrigel invasion chamber and colony formation assay, respectively. To conclude, it is thought that TMZ demonstrates anticarcinogenesis activity by affecting cell cycle arrest, apoptosis, invasion, and colony formation on SH-SY5Y cells. TMZ may be an effective agent for treatment of neuroblastoma as a single or in combination with other drugs.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

Abbreviations

T-ALL:

T-acute lymphocytic leukemia

CDK:

Cyclin-dependent kinase

CNS:

Central nervous system tumors

DMSO:

Dimethylsulfoxide

NB:

Neuroblastoma

TMZ:

Temozolomide

URG4:

Upregulated gene 4

URG4CP:

Upregulator of cell proliferation

XTT:

(2-Methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide

TMZ:

Temozolomide

TUNEL:

Mediated deoxyuridine triphosphate (dUTP)-digoxigenin nick-end labeling

References

  1. Brodeur GM. Neuroblastoma: biological insights into a clinical enigma. Nat Rev Cancer. 2003;3(3):203–16.

    Article  CAS  PubMed  Google Scholar 

  2. Maris JM, Hogarty MD, Bagatell R, Cohn SL. Neuroblastoma. Lancet. 2007;369(9579):2106–20.

    Article  CAS  PubMed  Google Scholar 

  3. Jiang M, Stanke J, Lahti JM. The connections between neural crest development and neuroblastoma. Curr Top Dev Biol. 2011;94:77–127.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Prasad G, Sottero T, Yang X, Mueller S, James CD, Weiss WA, et al. Inhibition of PI3K/mTOR pathways in glioblastoma and implications for combination therapy with temozolomide. Neuro Oncol. 2011;13(4):384–92.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Kaina B, Christmann M, Naumann S, Roos WP. MGMT: key node in the battle against genotoxicity, carcinogenicity and apoptosis induced by alkylating agents. DNA Repair (Amst). 2007;6(8):1079–99.

    Article  CAS  Google Scholar 

  6. Tisdale MJ. Antitumor imidazotetrazines – XV. Role of guanine O6 alkylation in the mechanism of cytotoxicity of imidazotetrazinones. Biochem Pharmacol. 1987;36(4):457–62.

    Article  CAS  PubMed  Google Scholar 

  7. Stupp R, Dietrich PY, Ostermann Kraljevic S, Pica A, Maillard I, Maeder P, et al. Promising survival for patients with newly diagnosed glioblastoma multiforme treated with concomitant radiation plus temozolomide followed by adjuvant temozolomide. J Clin Oncol. 2002;20(5):1375–82.

    Article  CAS  PubMed  Google Scholar 

  8. Mutter N, Stupp R. Temozolomide: a milestone in neurooncology and beyond? Expert Rev Anticancer Ther. 2006;6(8):1187–204.

    Article  CAS  PubMed  Google Scholar 

  9. Estlin EJ, Lashford L, Ablett S, et al. Phase I study of temozolomide in paediatric patients with advanced cancer. United Kingdom Children’s Cancer Study Group. Br J Cancer. 1998;78(5):652–61.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Middlemas DS, Stewart CF, Kirstein MN, et al. Biochemical correlates of temozolomide sensitivity in pediatric solid tumor xenograft models. Clin Cancer Res. 2000;6(3):998–1007.

    CAS  PubMed  Google Scholar 

  11. De Sio L, Milano GM, Castellano A, et al. Temozolomide in resistant or relapsed pediatric solid tumors. Pediatr Blood Cancer. 2006;47(1):30–6.

    Article  PubMed  Google Scholar 

  12. Rubie H, Chisholm J, Defachelles AS, et al. Phase II study of temozolomide in relapsed or refractory high-risk neuroblastoma: a joint Socie´te´ Franc_aise des Cancers de l’Enfant and United Kingdom Children Cancer Study Group-New Agents Group Study. J Clin Oncol. 2006;24(33):5259–64.

    Article  CAS  PubMed  Google Scholar 

  13. Dodurga Y, Gundogdu G, Tekin V, Koc T, Satiroglu-Tufan NL, Bagci G, et al. Valproic acid inhibits the proliferation of SHSY5Y neuroblastoma cancer cells by downregulating URG4/URGCP and CCND1 gene expression. Mol Biol Rep. 2014;41(7):4595–9.

    Article  CAS  PubMed  Google Scholar 

  14. Tufan NL, Lian Z, Liu J, et al. Hepatitis Bx antigen stimulates expression of a novel cellular gene, URG4, that promotes hepatocellular growth and survival. Neoplasia. 2002;4(4):355–68.

    Article  CAS  PubMed  Google Scholar 

  15. Song J, Xie H, Liany Z, Yang G, Du R, Du Y, et al. Enhanced cell survival of gastric cells by a novel gene URG4. Neoplasia. 2006;8(12):995–1002.

    Article  PubMed  PubMed Central  Google Scholar 

  16. Huang J, Zhu B, Lu L, Lian Z, et al. The expression of novel gene URG4 in osteosarcoma: correlation with patients’ prognosis. Pathology. 2009;41(2):149–54.

    Article  CAS  PubMed  Google Scholar 

  17. Dodurga Y, Oymak Y, Gu¨ndu¨z C, Tufan NL, Vergin C, Cetingul N, et al. Leukemogenesis as a new approach to investigate the correlation between up regulated gene 4/upregulator of cell proliferation (URG4/URGCP) and signal transduction genes in leukemia. Mol Biol Rep. 2013;40(4):3043–8.

    Article  CAS  PubMed  Google Scholar 

  18. Satiroglu-Tufan NL, Dodurga Y, Gok D, Cetinkaya A, Feitelson MA. RNA interference-mediated URG4 gene silencing diminishes cyclin D1 mRNA expression in HepG2 cells. Genet Mol Res. 2010;9(3):1557–67.

    Article  CAS  PubMed  Google Scholar 

  19. Osoba D, Braba M, Yung YKA, Prados MP. Health-related quality of life in patients treated with temozolomide versus procarbazine for recurrent glioblastoma multiforme. J Clin Oncol. 2000;18(7):1481–91.

    Article  CAS  PubMed  Google Scholar 

  20. Johnson DR, O'Neill BP. Glioblastoma survival in the United States before and during the temozolomide era. J Neurooncol. 2012;107(2):359–64.

    Article  CAS  PubMed  Google Scholar 

  21. Shirai K, Chakravarti A. Towards personalized therapy for patients with glioblastoma. Expert Rev Anticancer Ther. 2011;11(12):1935–44.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Pyrko P, Schonthal AH, Hofman FM, Chen TC, Lee AS. The unfolded protein response regulator GRP78/BiP as a novel target for increasing chemosensitivity in malignant gliomas. Cancer Res. 2007;67(20):9809–16.

    Article  CAS  PubMed  Google Scholar 

  23. Kil WJ, Cerna D, Burgan WE, et al. In vitro and in vivo radiosensitization induced by the DNA methylating agent temozolomide. Clin Cancer Res. 2008;14(3):931–8.

    Article  CAS  PubMed  Google Scholar 

  24. Gao S, Yang XJ, Zhang WG, et al. Mechanism of thalidomide to enhance cytotoxicity of temozolomide in U251-MG glioma cells in vitro. Chin Med J (Engl). 2009;122(11):1260–6.

    CAS  Google Scholar 

  25. Carmo A, Carvalheiro H, Crespo I, et al. Effect of temozolomide on the U-118 glioma cell line. Oncol Lett. 2011;2(6):1165–70.

    CAS  PubMed  PubMed Central  Google Scholar 

  26. Osoba D, Brada M, Yung WKA, et al. Health-related quality of life in patients with anaplastic astrocytoma during treatment with temozolomide. Eur J Cancer. 2000;36(14):1788–95.

    Article  CAS  PubMed  Google Scholar 

  27. Osoba D, Brada M, Prados MD, et al. Effect of disease burden on health-related quality of life in patients with malignant gliomas. Neuro Oncol. 2000;2(4):221–8.

    CAS  PubMed  PubMed Central  Google Scholar 

  28. Quinn JA, Reardon DA, Friedman AH, et al. Phase II trial of temozolomide in patients with progressive low-grade glioma. J Clin Oncol. 2003;21(4):646–51.

    Article  CAS  PubMed  Google Scholar 

  29. Estlin EJ, Lashford L, Ablett S, et al. Phase I study of temozolomide in paediatric patients with advanced cancer. United Kingdom Children’s Cancer Study Group. Br J Cancer. 1998;78(5):652–61.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Daniel RA, Rozanska AL, Thomas HD, Mulligan EA, Drew Y, Castelbuono DJ, et al. Inhibition of poly(ADP-ribose) polymerase-1 enhances temozolomide and topotecan activity against childhood neuroblastoma. Clin Cancer Res. 2009;15(4):1241–9.

    Article  CAS  PubMed  Google Scholar 

  31. Di Giannatale A, Dias-Gastellier N, Devos A, Mc Hugh K, Boubaker A, Courbon F, et al. Phase II study of temozolomide in combination with topotecan (TOTEM) in relapsed or refractory neuroblastoma: a European Innovative Therapies for Children with Cancer-SIOP-European Neuroblastoma study. Eur J Cancer. 2014;50(1):170–7.

    Article  PubMed  Google Scholar 

  32. Shen W, Hu JA, Zheng JS. Mechanism of temozolomide induced antitumour effects on glioma cells. J Int Med Res. 2014;42(1):164–72.

    Article  CAS  PubMed  Google Scholar 

  33. Woo SR, Ham Y, Kang W, Yang H, Kim S, Jin J, et al. KML001, a telomere targeting drug, sensitizes glioblastoma cells to temozolomide chemotherapy and radiotherapy trough DNA damage and apoptosis. Biomed Res Int. 2014;2014:747415.

    Article  PubMed  PubMed Central  Google Scholar 

  34. Peurala E, Koivunen P, Haapasaari KM, Bloigu R, Jukkola-Vuorinen A. The prognostic significance and value of cyclin D1, CDK4 and p16 in human breast cancer. Breast Cancer Res. 2013;15(1):R5.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Musgrove EA, Caldon CE, Barraclough J, Stone A, Sutherland RL. Cyclin D as a therapeutic target in cancer. Nat Rev Cancer. 2011;11(8):558–72.

    Article  CAS  PubMed  Google Scholar 

  36. Paternot S, Bockstaele L, Bisteau X, Kooken H, Coulonval K, Roger PP. Rb inactivation in cell cycle and cancer: the puzzle of highly regulated activating phosphorylation of CDK4 versus constitutively active CDK activating kinase. Cell Cycle. 2010;9(4):689–99.

    Article  CAS  PubMed  Google Scholar 

  37. Molenaar JJ, Ebus ME, Koster J, van Sluis P, van Noesel CJ, Versteeg R, et al. Cyclin D1 and CDK4 activity contribute to the undifferentiated phenotype in neuroblastoma. Cancer Res. 2008;68(8):2599–609.

    Article  CAS  PubMed  Google Scholar 

  38. Rader J, Russell MR, Hart LS, Nakazawa MS, Belcastro LT, Martinez D, et al. Dual CDK4/CDK6 inhibition induces cell-cycle arrest and senescence in neuroblastoma. Clin Cancer Res. 2013;19(22):6173–82.

    Article  CAS  PubMed  Google Scholar 

  39. Kanzawa T, Germano IM, Komata T, et al. Role of autophagy in temozolomide-induced cytotoxicity for malignant glioma cells. Cell Death Differ. 2004;11(4):448–57.

    Article  CAS  PubMed  Google Scholar 

  40. Günther W, Pawlak E, Damasceno R, Arnold H, Terzis AJ. Temozolomide induces apoptosis and senescence in glioma cells cultured as multicellular speroids. Br J Cancer. 2003;88(3):463–9.

    Article  PubMed  PubMed Central  Google Scholar 

  41. Dai C, Zhang B, Liu X, Guo K, Ma S, Cai F, et al. Pyrimethamine sensitizes pituitary adenomas cells to temozolomide through cathepsin B-dependent and caspase-dependent apoptotic pathways. Int J Cancer. 2013;133(8):1982–93.

    Article  CAS  PubMed  Google Scholar 

  42. Fukai J, Koizumi F, Nakao N. Enhanced antitumor effect of zoledronic acid combined with temozolomide against human malignant glioma cell expressing O6-methylguanine DNA methyltransferase. PLoS One. 2014;9(8):e104538. doi:10.1371/journal.pone.0104538.

    Article  PubMed  PubMed Central  Google Scholar 

  43. Yang J, Liu X, Bhalla K, et al. Prevention of apoptosis by Bcl-2: release of cytochrome c from mitochondria blocked. Science. 1997;275(5303):1129–32.

    Article  CAS  PubMed  Google Scholar 

  44. Ma S, Liu X, Yao Y, et al. Effect of temozolomide on cell viability in gonadotroph adenoma cell lines. Oncol Rep. 2011;26(3):543–50.

    PubMed  Google Scholar 

  45. Lavictoire SJ, Parolin DA, Klimowicz AC, Kelly JF, Lorimer IA. Interaction of Hsp90 with the nascent form of the mutant epidermal growth factor receptor EGFRvIII. J Biol Chem. 2003;278(7):5292–9.

    Article  CAS  PubMed  Google Scholar 

  46. Choi EJ, Cho BJ, Lee DJ, Hwang YH, Chun SH, Kim HH, et al. Enhanced cytotoxic effect of radiation and temozolomide in malignant glioma cells: targeting PI3K-AKT-mTORsignaling, HSP90 and histone deacetylases. BMC Cancer. 2014;14:17. doi:10.1186/1471-2407-14-17.

    Article  PubMed  PubMed Central  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Neurosurgery, Pamukkale University School of Medicine, Denizli, Turkey

    Veli Çıtışlı

  2. Department of Medical Biology, Pamukkale University School of Medicine, Denizli, Turkey

    Yavuz Dodurga, Canan Eroğlu & Mücahit Seçme

  3. Department of Medical Biology, Ege University School of Medicine, Izmir, Turkey

    Çığır Biray Avcı

  4. Department of Forensic Medicine, Ankara University School of Medicine, Ankara, Turkey

    N. Lale Şatıroğlu-Tufan

Authors
  1. Veli Çıtışlı
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yavuz Dodurga
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Canan Eroğlu
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Mücahit Seçme
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Çığır Biray Avcı
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. N. Lale Şatıroğlu-Tufan
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Yavuz Dodurga.

Additional information

Highlights

1. TMZ is inhibiting cell proliferation in SH-SY5Y neuroblastoma cells by inducing cell cycle arrest via CCND1, CCND2, CDK4, and URG4/URGCP gene expression changes.

2. TMZ is stimulating apoptotic mechanisms in SH-SY5Y.

3. First mechanisms of TMZ and URG4/URGCP relationship in neuroblastoma cells were presented in this study.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Çıtışlı, V., Dodurga, Y., Eroğlu, C. et al. Temozolomide may induce cell cycle arrest by interacting with URG4/URGCP in SH-SY5Y neuroblastoma cells. Tumor Biol. 36, 6765–6772 (2015). https://doi.org/10.1007/s13277-015-3373-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s13277-015-3373-7

Keywords

Search

Navigation