Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Original Article
  • Published:

Overcoming the radioresistance of prostate cancer cells with a novel Bcl-2 inhibitor

Oncogene volume 26, pages 652–661 (2007)Cite this article

Abstract

Bcl-2 overexpression is an important mechanism underlying the aggressive behavior of prostate cancer cells and their resistance to radio- or chemotherapy. HA14-1, a recently discovered organic Bcl-2 inhibitor, potently induces apoptosis in various human cancer cells. Sequential exposure of radioresistant LNCaP (wild-type (wt) p53), LNCaP/Bcl-2 (wt p53) and PC3 (mutant p53) prostate cancer cells to a minimally cytotoxic concentration of 10 μ M HA14-1 for 1 h followed by 1–6 Gy gamma radiation, resulted in a highly synergistic (combination index <1.0) induction of cell death as determined by an apoptosis assay at 72 h, and a clonogenicity assay at 12 days, after the initial treatment. The reverse treatment sequence did not cause a synergistic induction of cell death. When compared to individual treatments, cell death induced by the combined treatment was associated with dramatically increased reactive oxygen species (ROS) generation, c-Jun N-terminal kinase (JNK) activation, Bcl-2 phosphorylation, cytochrome c release, caspase-3 activation and DNA fragmentation. Exposure to either 200 μg/ml of the antioxidant alpha-tocopherol or 10 μ M JNK inhibitor SP600125 before the combined treatment resulted in decreased activation of JNK and caspase-3 as well as decreased DNA fragmentation. However, treatment with the pancaspase inhibitor carbobenzoxyl-valyl-alanyl-aspartyl-[O-methyl]-fluoromethylketone before the combined treatment inhibited apoptosis without affecting JNK activation, and this inhibitory effect was enhanced in the presence of alpha-tocopherol or SP600125. Taken together, our results indicate that HA14-1 potently sensitizes radioresistant LNCaP and PC3 cells to gamma radiation, regardless of the status of p53. ROS and JNK are important early signals that trigger both caspase-dependent and -independent cell death pathways and contribute to the apoptotic synergy induced by the combined treatments.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
Figure 8

Similar content being viewed by others

Abbreviations

HA14-1:

ethyl 2-amino-6-bromo-4-(1-cyano-2-ethoxy-2-oxoethyl)-4H-chromene-3-carboxylate

ROS:

reactive oxygen species

JNK:

c-Jun NH2-terminal kinase

ER:

endoplasmic reticulum

SSB:

single-strand break

DSB:

double-strand break

DMSO:

dimethyl sulfoxide

H2DCFDA:

succinimidyl ester of dichlorodihydrofluorescein diacetate

z-VAD-fmk:

carbobenzoxyl-valyl-alanyl-aspartyl-[O-methyl]-fluoromethylketone

wt:

wild type

CI:

combination index

IP:

immunoprecipitation

References

  • Adams JM, Cory S . (1998). The Bcl-2 protein family: arbiters of cell survival. Science 281: 1322–1326.

    Article  CAS  PubMed  Google Scholar 

  • An J, Chen Y, Huang Z . (2004). Critical upstream signals of cytochrome c release induced by a novel Bcl-2 inhibitor. J Biol Chem 279: 19133–19140.

    Article  CAS  PubMed  Google Scholar 

  • Autorino R, Di Lorenzo G, Damiano R, De Placido S, D’Armiento M . (2003). Role of chemotherapy in hormone-refractory prostate cancer. Old issues, recent advances and new perspectives. Urol Int 70: 1–14.

    Article  CAS  PubMed  Google Scholar 

  • Bauer JJ, Sesterhenn IA, Mostofi FK, McLeod DG, Srivastava S, Moul JW . (1996). Elevated levels of apoptosis regulator proteins p53 and bcl-2 are independent prognostic biomarkers in surgically treated clinically localized prostate cancer. J Urol 156: 1511–1516.

    Article  CAS  PubMed  Google Scholar 

  • Belka C, Jendrossek V, Pruschy M, Vink S, Verheij M, Budach W . (2004). Apoptosis-modulating agents in combination with radiotherapy – current status and outlook. Int J Radiat Oncol Biol Phys 58: 542–554.

    Article  CAS  PubMed  Google Scholar 

  • Catz SD, Johnson JL . (2003). BCL-2 in prostate cancer: a minireview. Apoptosis 8: 29–37.

    Article  CAS  PubMed  Google Scholar 

  • Chaudhary KS, Abel PD, Stamp GW, Lalani E . (2001). Differential expression of cell death regulators in response to thapsigargin and adriamycin in Bcl-2 transfected DU145 prostatic cancer cells. J Pathol 193: 522–529.

    Article  CAS  PubMed  Google Scholar 

  • Chendil D, Ranga RS, Meigooni D, Sathishkumar S, Ahmed MM . (2004). Curcumin confers radiosensitizing effect in prostate cancer cell line PC-3. Oncogene 23: 1599–1607.

    Article  CAS  PubMed  Google Scholar 

  • Chou TC, Talalay P . (1984). Quantitative analysis of dose–effect relationships: the combined effects of multiple drugs or enzyme inhibitors. Adv Enzyme Regul 22: 27–55.

    Article  CAS  PubMed  Google Scholar 

  • Denmeade SR, Lin XS, Isaacs JT . (1996). Role of programmed (apoptotic) cell death during the progression and therapy for prostate cancer. Prostate 28: 251–265.

    Article  CAS  PubMed  Google Scholar 

  • Dewey WC, Miller HH, Leeper DB . (1971). Chromosomal aberrations and mortality of x-irradiated mammalian cells: emphasis on repair. Proc Natl Acad Sci USA 68: 667–671.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Fei P, El-Deiry WS . (2003). P53 and radiation responses. Oncogene 22: 5774–5783.

    Article  CAS  PubMed  Google Scholar 

  • Garzotto M, Haimovitz-Friedman A, Liao WC, White-Jones M, Huryk R, Heston WD et al. (1999). Reversal of radiation resistance in LNCaP cells by targeting apoptosis through ceramide synthase. Cancer Res 59: 5194–5201.

    CAS  PubMed  Google Scholar 

  • Hahn P, Baral E, Cheang M, Math MC, Kostyra J, Roelss R . (1996). Long-term outcome of radical radiation therapy for prostatic carcinoma: 1967–1987. Int J Radiat Oncol Biol Phys 34: 41–47.

    Article  CAS  PubMed  Google Scholar 

  • Hering FL, Lipay MV, Lipay MA, Rodrigues PR, Nesralah LJ, Srougi M . (2001). Comparison of positivity frequency of bcl-2 expression in prostate adenocarcinoma with low and high Gleason score. Sao Paulo Med J 119: 138–141.

    Article  CAS  PubMed  Google Scholar 

  • Higuchi Y . (2003). Chromosomal DNA fragmentation in apoptosis and necrosis induced by oxidative stress. Biochem Pharmacol 66: 1527–1535.

    Article  CAS  PubMed  Google Scholar 

  • Kimura K, Bowen C, Spiegel S, Gelmann EP . (1999). Tumor necrosis factor-alpha sensitizes prostate cancer cells to gamma-irradiation-induced apoptosis. Cancer Res 59: 1606–1614.

    CAS  PubMed  Google Scholar 

  • Kolesnick R, Fuks Z . (2003). Radiation and ceramide-induced apoptosis. Oncogene 22: 5897–5906.

    Article  CAS  PubMed  Google Scholar 

  • Krajewska M, Krajewski S, Epstein JI, Shabaik A, Sauvageot J, Song K et al. (1996). Immunohistochemical analysis of bcl-2, bax, bcl-X, and mcl-1 expression in prostate cancers. Am J Pathol 148: 1567–1576.

    CAS  PubMed  PubMed Central  Google Scholar 

  • Lei K, Davis RJ . (2003). JNK phosphorylation of Bim-related members of the Bcl2 family induces Bax-dependent apoptosis. Proc Natl Acad Sci USA 100: 2432–2437.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lickliter JD, Wood NJ, Johnson L, McHugh G, Tan J, Wood F et al. (2003). HA14-1 selectively induces apoptosis in Bcl-2-overexpressing leukemia/lymphoma cells, and enhances cytarabine-induced cell death. Leukemia 17: 2074–2080.

    Article  CAS  PubMed  Google Scholar 

  • Lin A, Dibling B . (2002). The true face of JNK activation in apoptosis. Aging Cell 1: 112–116.

    Article  CAS  PubMed  Google Scholar 

  • Lin X, Zhang F, Bradbury CM, Kaushal A, Li L, Spitz DR et al. (2003). 2-Deoxy-D-glucose-induced cytotoxicity and radiosensitization in tumor cells is mediated via disruptions in thiol metabolism. Cancer Res 63: 3413–3417.

    CAS  PubMed  Google Scholar 

  • Marchal S, Bezdetnaya L, Guillemin F . (2004). Modality of cell death induced by Foscan-based photodynamic treatment in human colon adenocarcinoma cell line HT29. Biochemistry (Moscow) 69: 45–49.

    Article  CAS  Google Scholar 

  • McConkey DJ, Greene G, Pettaway CA . (1996). Apoptosis resistance increases with metastatic potential in cells of the human LNCaP prostate carcinoma line. Cancer Res 56: 5594–5599.

    CAS  PubMed  Google Scholar 

  • Mikkelsen RB, Wardman P . (2003). Biological chemistry of reactive oxygen and nitrogen and radiation-induced signal transduction mechanisms. Oncogene 22: 5734–5754.

    Article  CAS  PubMed  Google Scholar 

  • Milella M, Estrov Z, Kornblau SM, Carter BZ, Konopleva M, Tari A et al. (2002). Synergistic induction of apoptosis by simultaneous disruption of the Bcl-2 and MEK/MAPK pathways in acute myelogenous leukemia. Blood 99: 3461–3464.

    Article  CAS  PubMed  Google Scholar 

  • Mishra KP . (2004). Cell membrane oxidative damage induced by gamma-radiation and apoptotic sensitivity. J Environ Pathol Toxicol Oncol 23: 61–66.

    Article  CAS  PubMed  Google Scholar 

  • Ortiz MA, Lopez-Hernandez FJ, Bayon Y, Pfahl M, Piedrafita FJ . (2001). Retinoid-related molecules induce cytochrome c release and apoptosis through activation of c-Jun NH(2)-terminal kinase/p38 mitogen-activated protein kinases. Cancer Res 61: 8504–8512.

    CAS  PubMed  Google Scholar 

  • Park J, Kim I, Oh YJ, Lee K, Han PL, Choi EJ . (1997). Activation of c-Jun N-terminal kinase antagonizes an anti-apoptotic action of Bcl-2. J Biol Chem 272: 16725–16728.

    Article  CAS  PubMed  Google Scholar 

  • Phillips DC, Allen K, Griffiths HR . (2002). Synthetic ceramides induce growth arrest or apoptosis by altering cellular redox status. Arch Biochem Biophys 407: 15–24.

    Article  CAS  PubMed  Google Scholar 

  • Raffo AJ, Perlman H, Chen MW, Day ML, Streitman JS, Buttyan R . (1995). Overexpression of bcl-2 protects prostate cancer cells from apoptosis in vitro and confers resistance to androgen depletion in vivo. Cancer Res 55: 4438–4445.

    CAS  PubMed  Google Scholar 

  • Reed JC, Zha H, Aime-Sempe C, Takayama S, Wang HG . (1996). Structure-function analysis of Bcl-2 family proteins. Regulators of programmed cell death. Adv Exp Med Biol 406: 99–112.

    Article  CAS  PubMed  Google Scholar 

  • Rosser CJ, Reyes AO, Vakar-Lopez F, Levy LB, Kuban DA, Hoover DC et al. (2003). Bcl-2 is significantly overexpressed in localized radio-recurrent prostate carcinoma, compared with localized radio-naive prostate carcinoma. Int J Radiat Oncol Biol Phys 56: 1–6.

    Article  CAS  PubMed  Google Scholar 

  • Scherr DS, Vaughan EDJ, Wei J, Chung M, Felsen D, Allbright R et al. (1999). BCL-2 and p53 expression in clinically localized prostate cancer predicts response to external beam radiotherapy. J Urol 162: 12–16.

    Article  CAS  PubMed  Google Scholar 

  • Scott SL, Higdon R, Beckett L, Shi XB, deVere White RW, Earle JD et al. (2002). BCL2 antisense reduces prostate cancer cell survival following irradiation. Cancer Biother Radiopharm 17: 647–656.

    Article  CAS  PubMed  Google Scholar 

  • Shaffer DR, Scher HI . (2003). Prostate cancer: a dynamic illness with shifting targets. Lancet Oncol 4: 407–414.

    Article  PubMed  Google Scholar 

  • Szostak MJ, Kyprianou N . (2000). Radiation-induced apoptosis: predictive and therapeutic significance in radiotherapy of prostate cancer (review). Oncol Rep 7: 699–706.

    CAS  PubMed  Google Scholar 

  • Tang DG, Porter AT . (1997). Target to apoptosis: a hopeful weapon for prostate cancer. Prostate 32: 284–293.

    Article  CAS  PubMed  Google Scholar 

  • Tepper AD, de Vries E, van Blitterswijk WJ, Borst J . (1999). Ordering of ceramide formation, caspase activation, and mitochondrial changes during CD95- and DNA damage-induced apoptosis. J Clin Invest 103: 971–978.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Todaro M, Catalano M, Di Liberto D, Patti M, Zerilli M, Di Gaudio F et al. (2002). High levels of exogenous C2-ceramide promote morphological and biochemical evidences of necrotic features in thyroid follicular cells. J Cell Biochem 86: 162–173.

    Article  CAS  PubMed  Google Scholar 

  • Tournier C, Hess P, Yang DD, Xu J, Turner TK, Nimnual A et al. (2000). Requirement of JNK for stress-induced activation of the cytochrome c-mediated death pathway. Science 288: 870–874.

    Article  CAS  PubMed  Google Scholar 

  • Tsuruta F, Sunayama J, Mori Y, Hattori S, Shimizu S, Tsujimoto Y et al. (2004). JNK promotes Bax translocation to mitochondria through phosphorylation of 14-3-3 proteins. EMBO J 23: 1889–1899.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Vivo C, Liu W, Broaddus VC . (2003). c-Jun N-terminal kinase contributes to apoptotic synergy induced by tumor necrosis factor-related apoptosis-inducing ligand plus DNA damage in chemoresistant, p53 inactive mesothelioma cells. J Biol Chem 278: 25461–25467.

    Article  CAS  PubMed  Google Scholar 

  • Wang JL, Liu D, Zhang ZJ, Shan S, Han X, Srinivasula SM et al. (2000). Structure-based discovery of an organic compound that binds Bcl-2 protein and induces apoptosis of tumor cells. Proc Natl Acad Sci USA 97: 7124–7129.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Xiao D, Choi S, Johnson DE, Vogel VG, Johnson CS, Trump DL et al. (2004). Diallyl trisulfide-induced apoptosis in human prostate cancer cells involves c-Jun N-terminal kinase and extracellular-signal regulated kinase-mediated phosphorylation of Bcl-2. Oncogene 23: 5594–5606.

    Article  CAS  PubMed  Google Scholar 

  • Yamaguchi H, Paranawithana SR, Lee MW, Huang Z, Bhalla KN, Wang HG . (2002). Epothilone B analogue (BMS-247550)-mediated cytotoxicity through induction of Bax conformational change in human breast cancer cells. Cancer Res 62: 466–471.

    CAS  PubMed  Google Scholar 

  • Yu N, Aramini JM, Germann MW, Huang Z . (2000). Reactions of salicylaldehydes with alkyl cyanoacetates on the surface of solid catalysts: syntheses of 4H-chromene derivatives. Tetrahedron Lett 41: 6993–6996.

    Article  CAS  Google Scholar 

  • Zhan Q, Kontny U, Iglesias M, Alamo IJ, Yu K, Hollander MC et al. (1999). Inhibitory effect of Bcl-2 on p53-mediated transactivation following genotoxic stress. Oncogene 18: 297–304.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This work was supported by grants from the American Cancer Society, the National Institutes of Health and the Sidney Kimmel Foundation for Cancer Research.

Author information

Authors and Affiliations

  1. Department of Biochemistry, University of Illinois at Urbana-Champaign, Urbana, IL, USA

    J An, A S Chervin & Z Huang

  2. Burnham Institute for Medical Research, La Jolla, CA, USA

    J An, A Nie & Z Huang

  3. Department of Molecular and Integrative Physiology, University of Illinois at Urbana-Champaign, Urbana, IL, USA

    H S Ducoff

Authors
  1. J An
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. A S Chervin
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. A Nie
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. H S Ducoff
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Z Huang
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to J An.

Rights and permissions

Reprints and permissions

About this article

Cite this article

An, J., Chervin, A., Nie, A. et al. Overcoming the radioresistance of prostate cancer cells with a novel Bcl-2 inhibitor. Oncogene 26, 652–661 (2007). https://doi.org/10.1038/sj.onc.1209830

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.onc.1209830

Keywords

This article is cited by

Search

Advanced search

Quick links