- Research Article
Correlation between the levels of survivin and survivin promoter-driven gene expression in cancer and non-cancer cells
Cellular & Molecular Biology Lettersvolume 14, Article number: 70 (2008)
Survivin, a member of the inhibitor of apoptosis (IAP) protein family, is associated with malignant transformation and is over-expressed in most human tumors. Using lipoplex-mediated transfection, we evaluated the activity of the reporter enzyme, luciferase, expressed from plasmids encoding the enzyme under the control of either the cytomegalovirus (CMV) or survivin promoters, in tumor- and non-tumor-derived human and murine cells. We also examined whether there is a correlation between the survivin promoter-driven expression of luciferase and the level of endogenous survivin. Human cancer cells (HeLa, KB, HSC-3, H357, H376, H413), oral keratinocytes, GMSM-K, and chemically immortalized human mammary cells, 184A-1, were transfected with Metafectene at 2 μl/1 μg DNA. Murine squamous cell carcinoma cells, SCCVII, mouse embryonic fibroblasts, NIH-3T3, and murine immortalized mammary cells, NMuMG, were transfected with Metafectene PRO at 2 μl/1 μg DNA. The expression of luciferase was driven by the CMV promoter (pCMV.Luc), the human survivin promoter (pSRVN.Luc-1430), or the murine survivin promoters (pSRVN.Luc-1342 and pSRVN.Luc-194). Luciferase activity was measured, using the Luciferase Assay System and expressed as relative light units (RLU) per ml of cell lysate or per mg of protein. The level of survivin in the lysates of human cells was determined by ELISA and expressed as ng survivin/mg protein. In all cell lines, significantly higher luciferase activity was driven by the CMV promoter than by survivin promoters. The expression of luciferase driven by the CMV and survivin promoters in murine cells was much higher than that in human cells. The cells displayed very different susceptibilities to transfection; nevertheless, high CMV-driven luciferase activity appeared to correlate with high survivin-promoter driven luciferase expression. The survivin concentration in lysates of cancer cells ranged from 5.8 ± 2.3 to 24.3 ± 2.9 ng/mg protein (mean, 13.7 ng/mg). Surprisingly, elevated survivin protein was determined in lysates of non-tumor-derived cells. Survivin levels for GMSM-K and 184A-1 cells, were 16.7 ± 8.7 and 13.5 ± 6.2 ng/mg protein, respectively. The expression of endogenous survivin did not correlate with the level of survivin promoter-driven transgene activity in the same cells. The expression of survivin by non-tumorigenic, transformed cell lines may be necessary for their proliferative activity. The level of survivin promoter-driven gene expression achieved via liposomal vectors in OSCC cells was too low to be useful in cancer-cell specific gene therapy.
Dulbecco’s modified Eagle’s MEM medium
fetal bovine serum
head and neck squamous cell carcinoma
inhibitor of apoptosis
oral squamous cell carcinoma
relative light units
Hanahan, D. and Weinberg, R.A. The hallmarks of cancer. Cell 100 (2000) 57–70.
Evan, G.I. and Vousden, K.H. Proliferation, cell cycle and apoptosis in cancer. Nature 411 (2001) 342–348.
Salvesen, G.S. and Duckett, C.S. IAP proteins: blocking the road to death’s door. Natl. Rev. Mol. Cell Biol. 3 (2002) 401–410.
Li, F. and Altieri, D.C. The caner anti-apoptosis mouse survivin gene: characterization of locus and transcriptional requirements of basal and cell cycle-dependent expression. Cancer Res. 59 (1999) 3142–3151.
Altieri, D.C. Validating survivin as a cancer therapeutic target. Nat. Rev. Cancer 3 (2003) 46–54.
Altieri, D.C. Molecular circuits of apoptosis regulation and cell division control: The survivin paradigm. J. Cell. Biochem. 92 (2004) 656–663.
Altieri, D.C. Targeted therapy by disabling crossroad signaling networks: the survivin paradigm. Mol. Cancer Ther. 5 (2006) 478–482.
Dohi, T., Beltrami, E., Wall, N.R., Plescia, J. and Altieri, D.C. Mitochondrial survivin inhibits apoptosis and promotes tumorigenesis. J. Clin. Invest. 114 (2004) 1117–1127.
Ambrosini, G., Adida, C. and Altieri, D.C. A novel anti-apoptotic gene, survivin, expressed in cancer and lymphoma. Nat. Med. 3 (1997) 917–921.
Adida, C., Crotty, P.L., McGrath, J., Berrebi, D., Diebold, J. and Altieri, D.C. Developmentally regulated expression of the novel cancer antiapoptosis gene survivin in human and mouse differentiation. Am. J. Pathol. 152 (1998) 43–49.
Fukuda, S. and Pelus, L.M. Regulation of the inhibitor-of-apoptosis family member survivin in normal cord blood and bone marrow CD34+ cells by hematopoietic growth factors: Implications of survivin expression in normal hematopoiesis. Blood 98 (2001) 2091–2100.
Altieri, D.C. Survivin and apoptosis control. Adv. Cancer Res. 88 (2003) 31–52.
Johnson, M.E. and Howerth, E.W. Survivin: a bifunctional inhibitor of apoptosis protein. Vet. Pathol. 41 (2004) 599–607.
Lo Muzio, L., Pannone, G., Leonardi, R., Staibano, S., Mignogna, M.D., De Rosa, G., Kudo, Y., Takata, T. and Altieri, D.C. Survivin, a potential early predictor of tumor progression in the oral mucosa. J. Dent. Res. 82 (2003) 923–928.
Lo Muzio, L., Pannone, G., Staibano, S., Mignogna, M.D., Rubini, C., Mariggiò, M.A., Procaccini, M., Ferrari, F., De Rosa, G. and Altieri, D.C. Survivin expression in oral squamous cell carcinoma. Brit. J. Cancer 89 (2003) 2244–2248.
Lo Muzio, L., Campisi, G., Giovanelli, L., Ammatuna, P., Greco, I., Staibano, S., Pannone, G., De Rosa, G., Di Liberto, C. and D’Angelo, M. HPV DNA and survivin expression in epithelial oral carcinogenesis: a relationship? Oral Oncol. 40 (2004) 736–741.
Tanaka, C., Uzawa, K., Shibahara, T., Yokoe, H., Noma, H. and Tanzawa, H. Expression of an inhibitor of apoptosis, survivin, in oral carcinogenesis. J. Dent. Res. 82 (2003) 607–811.
Lin, C.Y., Hung, H.C., Kuo, H.C., Chiang, C.P. and Kuo, M.Y. Survivin expression predicts poorer prognosis in patients with areca quid chewingrelated oral squamous cell carcinoma in Taiwan. Oral Oncol. 41 (2005) 645–654.
Bao, R., Connolly, D.C., Murphy, M., Green, J., Weinstein, J.K., Pisarcik, D.A. and Hamilton, T. Activation of cancer-specific gene expression by the survivin promoter. J. Natl. Cancer Inst. 94 (2002) 522–528.
Chiou, S., Jones, M.K. and Tarnawski, A.S. Survivin- an anti-apoptosis protein: its biological roles and implications for cancer and beyond. Med. Sci. Monit. 9 (2003) P143–P147.
Chen, J.S., Liu, J.C., Shen, L., Rau, K.M., Kuo, H.P., Li, Y.M., Shi, D., Lee, Y.C., Chang, K.J. and Hung, M.C. Cancer-specific activation of the survivin promoter and its potential use in gene therapy. Cancer Gene Ther. 11 (2004) 740–747.
Scully, C. Oral precancer: preventive and medical approaches to management. Eur. J. Cancer B Oral Oncol. 31(B) (1995) 16–26.
Schepman, K., der Meij, E., Smeele, L. and der Waal, I. Concomitant leukoplakia in patients with oral squamous cell carcinoma. Oral. Dis. 5 (1999) 206–209.
Choi, S. and Myers, J.N. Molecular pathogenesis of oral squamous cell carcinoma: implications for therapy. J. Dent. Res. 87 (2007) 14–32.
Lo Muzio, L., Staibano, S., Pannone, G., Mignona, M.D., Mariggio, A., Salvatore, G., Chieffi, P., Tramontano, D., De Rosa, G. and Altieri D.C. Expression of the apoptosis inhibitor survivin in aggressive squamous cell carcinoma. Exp. Mol. Pathol. 70 (2001) 249–254.
Lo Muzio, L. Farina, A., Rubini, C., Pezzetti, F., Stabellini, G., Laino, G., Santarelli, A., Pannone, G., Bufo, P., de Lillo, A. and Carinci, F. Survivin as prognostic factor in squamous cell carcinoma of the oral cavity. Cancer Lett. 225 (2005) 27–33.
Harras, A., Edwards, B.K., Blot, W.J. and Ries, L.A. Cancer rates and risks. National Institutes of Health, Bethesda, MD. NIH Publication (1996) No. 96–691.
Silverman, S. Jr. Oral cancer: complication of therapy. Oral Surg. Oral Med. Oral Pathol. Oral Radiol. Endod. 88 (1999) 122–126.
Shillitoe, E.J. Gene therapy for oral cancer: recent progress in research. Oral Oncol. 34 (1998) 157–160.
Xi, S. and Grandis, J.R. Gene therapy for the treatment of oral squamous cell carcinoma. J. Dental. Res. 82 (2003) 11–16.
Ladeinde, A.L., Ogunlewe, M.O., Adeyemo, W.L. and Bamgbose, B.O. Gene therapy in the management of oral cancer: a review of recent documents. Niger. Postgrad. Med. J. 12 (2005) 18–22.
Gibbs J.B. Mechanism based target identification and drug discovery in cancer research. Science 287 (2000) 1969–1973.
Hart, I.R. Tissue specific promoters in targeting systemically delivered gene therapy. Semin. Oncol. 23 (1996) 154–158.
O’Malley, B.W., Cope, K.A., Chen, S.-H., Li, D., Schwartz, M.R. and Woo, S.L.C. Combination gene therapy for oral cancer in a murine model. Cancer Res. 56 (1996) 1737–1741.
O’Malley, B.W., Cope, K.A., Johnson, C.S. and Schwartz, M.R. A new immunocompetent murine model for oral cancer. Arch. Otolaryngol. Head Neck Surg. 123 (1997) 20–24.
Yen, A., Overlid, N., Li, F., Düzgüneş N. and Konopka, K. Survivin promoter-driven gene expression in human oral cancer cells. The 85th General Session of the International Association for Dental Research and 36th Annual Meeting of the American Association for Dental Research, New Orleans, LA, 2007, J. Dent. Res. 86 (Special issue A) Abstract No. 0758, Seq. #98.
Spain, C., Overlid, N., Li, F., Düzgüneş, N. and Konopka, K. Murine survivin promoter-driven gene expression in cancer and non-tumor cells. International Association for Dental Research and 36th Annual Meeting of the American Association for Dental Research, New Orleans, LA, 2007, J. Dent. Res. 86 (Special issue A) Abstract No. 2394, Seq. #240.
Li, F. and Altieri, D.C. Transcriptional analysis of human survivin gene expression. Biochem. J. 344 (1999) 305–311.
Yang, L., Cao, Z., Li, F., Post, D.E., Van Meir, E.G., Zhong, H. and Wood, W.C. Tumor-specific gene expression using the survivin promoter is further increased by hypoxia. Gene Ther. 11 (2004) 1215–1223.
Zhu, Z.B., Makhija, S.K., Lu, B., Wang, M., Kaliberova, L., Liu, B., Rivera, A.A., Nettelbeck, D.M., Mahasreshti, P.J., Leath III, C.A., Barker, S., Yamaoto, M., Li, F., Alvarez, R.D. and Curiel D.T. Transcriptional targeting of tumors with a novel tumor-specific survivin promoter. Cancer Gene Ther. 11 (2004) 256–262.
Matsumoto, K., Horikoshi, M., Rikimaru, K. and Enomoto, S. A study of an in vitro model for invasion of oral squamous cell carcinoma. J. Oral Pathol. Med. 18 (1989) 498–501.
Prime, S.S., Nixon, S.V.R., Crane, I.J., Stone, A., Matthews, J.B., Maitland, N.J., Remnant, L., Powell, S.K., Game, S.M. and Scully, C. The behaviour of human oral squamous cell carcinoma in cell culture. J. Pathol. 160 (1990) 259–269.
Eagle, H. Propagation in a fluid medium of a human epidermoid carcinoma strain KB. Proc. Soc. Exp. Biol. Med. 89 (1955) 362–364.
Gilchrist, E.P., Moyer M.P., Shillitoe, E.J., Clare, N. and Murrah, V.A. Establishment of a human polyclonal oral epithelial cell line. Oral Surg. Oral Med Oral Pathol. Oral Radiol. Endod. 90 (2000) 340–347.
Stampfer, M.R. and Bartley, J.C. Induction of transformation and continuous cell lines from normal human mammary epithelial cells after exposure to benzeno(a)pyrene. Proc. Nat. Acad. Sci. 82 (1985) 2394–2398.
Walen, K.H. and Stampfer, M.R. Chromosome analyses of human mammary epithelial cells at stages of chemical-induced transformation progression to immortality. Cancer Genet. Cytogenet. 37 (1989) 249–261.
Fu, K.K., Rayner, P.A. and Lam, K.N. Modification of the effects of continuous low dose irradiation by concurrent chemotherapy infusion. Int. J. Radiat. Oncol. Biol. Phys. 10 (1984) 1473–1478.
Jainchill, J.L., Aaronson, S.A., and Todaro, G.J. Murine sarcoma and leukemia viruses: assay using clonal lines of contact-inhibited mouse cells. J. Virol. 4 (1969) 549–553.
Owens, R.B., Smith, H.S. and Hackett, A.J. Epithelial cell cultures from normal glandular tissue of mice. J. Natl. Cancer Inst. 53 (1974) 261–269.
Konopka, K. Fallah, B., Monzon-Duller, J., Overlid, N. and Düzgüneş, N. Serum-resistant gene transfer to oral cancer cells by Metafectene and GeneJammer: Application to HSV-tk/ganciclovir-mediated cytotoxicity. Cell. Mol. Biol. Lett. 10 (2005) 455–470.
Fields, R.D. and Lancaster, M.V. Dual-attribute continuous monitoring of cell proliferation/cytotoxicity. Am. Biotechnol. Lab. 11 (1993) 48–50.
Konopka, K., Pretzer, E., Felgner, P.L. and Düzgüneş, N. Human immunodeficiency virus type-1 (HIV-1) infection increases the sensitivity of macrophages and THP-1 cells to cytotoxicity by cationic liposomes. Biochim. Biophys. Acta 1312 (1996) 186-196.
Konopka, K., Overlid, N., Nagaraj, A.C. and Düzgüneş, N. Serum decreases the size of Metafectene- and GeneJammer-DNA complexes but does not affect significantly their transfection activity in SCCVII squamous cell carcinoma cells. Cell. Mol. Biol. Lett. 11 (2006) 171–190.
Bandyopadhyay, A., Cibull, M.L. and Sun, L. Isolation and characterization of a spontaneously transformed mouse mammary epithelial cell line in culture. Carcinogenesis 19 (1998) 1907–1911.
Zhu, Z.B., Makhija, S.K., Lu, B., Wang, M., Wang, S., Takayama, K., Siegal, G.P., Reynolds, P.N. and Curiel D.T. Targeting mesothelioma using an infectivity enhanced survivin-conditionally replicative adenoviruses. J. Thorac. Oncol. 1 (2006) 701–711.
Li, B., Liu, X., Fan, J., Qi, R., Bo, L., Gu, J., Qian, Q., Qian, C. and Liu, X. A survivin-mediated oncolytic adenovirus induces non-apoptotic cell death in lung cancer cells and shows antitumoral potential in vivo. J. Gene Med. 8 (2006) 1232–1242.
Ulasov, I.V., Zhu, Z.B., Tyler, M.A., Han, Y., Rivera, A.A., Khramtsov, A., Curiel, D.T. and Lesniak, M.S. Survivin-driven and fiber-modified oncolytic adenovirus exhibits potent antitumor activity in established intracranial glioma. Hum. Gene Ther. 18 (2007) 589–602.
Kamizono, J., Nagano, S., Murofushi, Y., Komiya, S., Fujiwara, H., Matsuishi, T. and Kosai, K. Survivin-responsive conditionally replicating adenovirus exhibits cancer-specific and efficient viral replication. Cancer Res. 65 (2005) 5284–5291.
Xu, Y., Fang, F., Ludewig, G., Jones, G. and Jones, D. A mutation found in the promoter region of the human survivin gene is correlated to overexpression of survivin in cancer cells. DNA Cell Biol. 23 (2004) 527–537.
Jang, J.S., Kim, K.M., Kang, K.H., Choi, J.E., Lee, W.K., Kim, C.H., Kang, Y.M., Kam, S., Kim, I.S., Jun, J.E. and Park, J.Y. Polymorphisms in the survivin gene and the risk of lung cancer. Lung Cancer 60 (2008) 31–39.
Borbely, A.A., Murvai, M., Szarka, K., Konya, J., Gergely, L., Hernadi, Z. and Veress, G. Survivin promoter polymorphism and cervical carcinogenesis. J. Clin. Pathol. 60 (2007) 303–306.
Kappler, M., Kotzsch, M., Bartel, F., Füssel, S., Lautenschläger, C., Schmidt, U., Würl, P., Bache, M., Schmidt, H., Taubert, H. and Meye, A. Elevated expression level of survivin protein in soft-tissue sarcomas is a strong independent predictor of survival. Clin. Cancer Res. 9 (2003) 1098–1104.
About this article
- Metafectene PRO
- Survivin promoter
- Non-cancer cells
- CMV promoter
- Oral squamous cell carcinoma cells