Skip to main content

Advertisement

SpringerLink
Log in

Efficacy of Hsp90 inhibition for induction of apoptosis and inhibition of growth in cervical carcinoma cells in vitro and in vivo

  • Original Article
  • Published:
Cancer Chemotherapy and Pharmacology Aims and scope Submit manuscript

Abstract

Purpose

Heat shock protein 90 (Hsp90) is a conserved chaperone involved in crucial signaling events in normal and malignant cells. Previous research suggests that tumor cells are particularly dependent on Hsp90 for survival as well as malignant progression. Hsp90 inhibitors which are derivates of the natural compound geldanamycin, such as the orally bioavailable 17-(dimethylaminoethylamino)-17-demethoxygeldanamycin (17-DMAG), are currently being tested in clinical trials and small molecule inhibitors are in development. In this study we investigated the response of a panel of cervical carcinoma cell lines in vitro and in vivo to determine potential factors that might influence the sensitivity towards Hsp90 inhibition.

Methods

Cell viability, proliferation and drug-induced changes on Hsp90 chaperoned “client” factors were examined with focus on G2/M cell cycle regulators, and a comparison with immortalized and normal keratinocytes was performed. ME180 and CaSki cells were grown as subcutaneous xenografts in mice treated with 6–10 mg/kg 17-DMAG by oral gavage 2×/day on a chronic schedule. Tissue concentrations of 17-DMAG were measured by high performance liquid chromatography.

Results

Cell death during abnormal mitosis was observed within 48 h after treatment start. ME180 and CaSki showed more cell death at this time point than SiHa and HeLa, and higher levels of pre-treatment Akt activity. IC50 values ranged between 17 and 37 nanoM geldanamycin (MTS). Keratinocytes were at least as sensitive as carcinoma cells. All cell lines responded with an increase of the G2/M fraction. Despite in vitro effectiveness and tissue concentrations of 1 µM, only a limited tumor growth reduction was observed with 17-DMAG given close to the maximum tolerated dose level. Lower levels of Hsp90 protein, a lower Akt activity and signs of tissue hypoxia were observed in xenografts compared to cell cultures.

Conclusions

We show here that Hsp90 inhibition effectively induces apoptosis and growth arrest in cervical carcinoma cells in vitro. Mitotic catastrophe was identified as one mechanism of cell death. In contrast, a limited efficacy of 17-DMAG was observed in subcutaneous xenograft models. Induction of a heat shock response has previously been implicated in resistance towards Hsp90 inhibition. Additional factors might be (1) an altered abundance and/or activity of primary (Hsp90) and secondary (e.g., Akt) target(s), (2) a narrow therapeutic range of 17-DMAG by oral application and (3) response-modifying factors within the tumor environment. The further development of synthetic Hsp90 inhibitors with increased therapeutic window is warranted.

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
Fig. 6

Similar content being viewed by others

References

  1. Pratt WB, Toft DO (2003) Regulation of signaling protein function and trafficking by the hsp90/hsp70-based chaperone machinery. Exp Biol Med (Maywood) 228:111–133

    CAS  Google Scholar 

  2. Kamal A, Thao L, Sensintaffar J, Zhang L, Boehm MF, Fritz LC, Burrows FJ (2003) A high-affinity conformation of Hsp90 confers tumour selectivity on Hsp90 inhibitors. Nature 425:407–410

    Article  PubMed  CAS  Google Scholar 

  3. BeBoer C, Dietz A (1976) The description and antibiotic production of Streptomyces hygroscopicus var. Geldanus. J Antibiotics (Tokyo) 29:1182–1188

    CAS  Google Scholar 

  4. Whitesell L, Mimnaugh EG, De Costa B, Myers CE, Neckers LM (1994) Inhibition of heat shock protein HSP90-pp60v-src heteroprotein complex formation by benzoquinone ansamycins: essential role for stress proteins in oncogenic transformation. Proc Natl Acad Sci USA 91:8324–8328

    Article  PubMed  CAS  Google Scholar 

  5. Dutta R, Inouye M (2000) GHKL, an emergent ATPase/kinase superfamily. Trends Biochem Sci 25:24–28

    Article  PubMed  CAS  Google Scholar 

  6. Pacey S, Banerji U, Judson I, Workman P (2006) Hsp90 inhibitors in the clinic. Handbook Exp Pharmacol 172:331–358

    CAS  Google Scholar 

  7. Egorin MJ, Lagattuta TF, Hamburger DR, Covey JM, White KD, Musser SM, Eiseman JL (2002) Pharmacokinetics, tissue distribution, and metabolism of 17-(dimethylaminoethylamino)-17-demethoxygeldanamycin (NSC 707545) in CD2F1 mice and Fischer 344 rats. Cancer Chemother Pharmacol 49:7–19

    Article  PubMed  CAS  Google Scholar 

  8. Eiseman JL, Lan J, Lagattuta TF, Hamburger DR, Joseph E, Covey JM, Egorin MJ (2005) Pharmacokinetics and pharmacodynamics of 17-demethoxy 17-[[(2-dimethylamino)ethyl]amino]geldanamycin (17DMAG, NSC 707545) in C.B-17 SCID mice bearing MDA-MB-231 human breast cancer xenografts. Cancer Chemother Pharmacol 55:21–32

    Article  PubMed  CAS  Google Scholar 

  9. Hollingshead M, Alley M, Burger AM, Borgel S, Pacula-Cox C, Fiebig HH, Sausville EA (2005) In vivo antitumor efficacy of 17-DMAG (17-dimethylaminoethylamino-17-demethoxygeldanamycin hydrochloride), a water-soluble geldanamycin derivative. Cancer Chemother Pharmacol 56:115–125

    Article  PubMed  CAS  Google Scholar 

  10. Glaze ER, Lambert AL, Smith AC, Page JG, Johnson WD, McCormick DL, Brown AP, Levine BS, Covey JM, Egorin MJ, Eiseman JL, Holleran JL, Sausville EA, Tomaszewski JE (2005) Preclinical toxicity of a geldanamycin analog, 17-(dimethylaminoethylamino)-17-demethoxygeldanamycin (17-DMAG), in rats and dogs: potential clinical relevance. Cancer Chemother Pharmacol 56:637–647

    Article  PubMed  CAS  Google Scholar 

  11. Shadad FN, Ramanathan RK (2006) 17-dimethylaminoethylamino-17-demethoxygeldanamycin in patients with advanced-stage solid tumors and lymphoma: a phase I study. Clin Lymphoma Myeloma 6:500–501

    Article  PubMed  CAS  Google Scholar 

  12. Zhang L, Fan J, Vu K, Hong K, Le Brazidec JY, Shi J, Biamonte M, Busch DJ, Lough RE, Grecko R, Ran Y, Sensintaffar JL, Kamal A, Lundgren K, Burrows FJ, Mansfield R, Timony GA, Ulm EH, Kasibhatla SR, Boehm MF (2006) 7′-substituted benzothiazolothio- and pyridinothiazolothio-purines as potent heat shock protein 90 inhibitors. J Med Chem 49:5352–5362

    Article  PubMed  CAS  Google Scholar 

  13. Parkin DM, Bray F (2006) Chapter 2: The burden of HPV-related cancers. Vaccine 24(Suppl 3):S11–S25

    Article  PubMed  Google Scholar 

  14. Walboomers JM, Jacobs MV, Manos MM, Bosch FX, Kummer JA, Shah KV, Snijders PJ, Peto J, Meijer CJ, Munoz N (1999) Human papillomavirus is a necessary cause of invasive cervical cancer worldwide. J Pathol 189:12–19

    Article  PubMed  CAS  Google Scholar 

  15. Soutter WP, Sasieni P, Panoskaltsis T (2006) Long-term risk of invasive cervical cancer after treatment of squamous cervical intraepithelial neoplasia. Int J Cancer 118:2048–2055

    Article  PubMed  CAS  Google Scholar 

  16. Yuan C, Wang P, Lai C, Tsu E, Yen M, Ng H (1999) Recurrence and survival analyses of 1,115 cervical cancer patients treated with radical hysterectomy. Gynecol Obstet Invest 47:127–132

    Article  PubMed  CAS  Google Scholar 

  17. Whitesell L, Lindquist SL (2005) HSP90 and the chaperoning of cancer. Nat Rev Cancer 5: 761–772

    Article  PubMed  CAS  Google Scholar 

  18. Bagatell R, Paine-Murrieta GD, Taylor CW, Pulcini EJ, Akinaga S, Benjamin IJ, Whitesell L (2000) Induction of a heat shock factor 1-dependent stress response alters the cytotoxic activity of hsp90-binding agents. Clin Cancer Res 6:3312–3318

    PubMed  CAS  Google Scholar 

  19. Guo F, Rocha K, Bali P, Pranpat M, Fiskus W, Boyapalle S, Kumaraswamy S, Balasis M, Greedy B, Armitage ES, Lawrence N, Bhalla K (2005) Abrogation of heat shock protein 70 induction as a strategy to increase antileukemia activity of heat shock protein 90 inhibitor 17-allylamino-demethoxy geldanamycin. Cancer Res 65:10536–10544

    Article  PubMed  CAS  Google Scholar 

  20. McCollum AK, Teneyck CJ, Sauer BM, Toft DO, Erlichman C (2006) Up-regulation of heat shock protein 27 induces resistance to 17-allylamino-demethoxygeldanamycin through a glutathione-mediated mechanism. Cancer Res 66:10967–10975

    Article  PubMed  CAS  Google Scholar 

  21. Schwock J, Geddie WR, Hedley DW (2005) Analysis of hypoxia-inducible factor-1alpha accumulation and cell cycle in geldanamycin-treated human cervical carcinoma cells by laser scanning cytometry. Cytometry A 68:59–70

    PubMed  Google Scholar 

  22. Pham NA, Gal MR, Bagshaw RD, Mohr AJ, Chue B, Richardson T, Callahan JW (2005) A comparative study of cytoplasmic granules imaged by the real-time microscope, Nile Red and Filipin in fibroblasts from patients with lipid storage diseases. J Inherit Metab Dis 28:991–1004

    Article  PubMed  Google Scholar 

  23. Xie XY, Robb D, Chow S, Hedley DW (1995) Discordant P-glycoprotein antigen expression and transport function in acute myeloid leukemia. Leukemia 9:1882–1887

    PubMed  CAS  Google Scholar 

  24. Burrows F, Zhang H, Kamal A (2004) Hsp90 activation and cell cycle regulation. Cell Cycle, 3:1530–1536

    PubMed  CAS  Google Scholar 

  25. de Carcer G (2004) Heat shock protein 90 regulates the metaphase–anaphase transition in a polo-like kinase-dependent manner. Cancer Res 64:5106–5112

    Article  PubMed  Google Scholar 

  26. Okumura E, Fukuhara T, Yoshida H, Hanada Si S, Kozutsumi R, Mori M, Tachibana K, Kishimoto T (2002) Akt inhibits Myt1 in the signalling pathway that leads to meiotic G2/M-phase transition. Nat Cell Biol 4:111–116

    Article  PubMed  CAS  Google Scholar 

  27. Ibrahim NO, Hahn T, Franke C, Stiehl DP, Wirthner R, Wenger RH, Katschinski DM (2005) Induction of the hypoxia-inducible factor system by low levels of heat shock protein 90 inhibitors. Cancer Res 65:11094–11100

    Article  PubMed  CAS  Google Scholar 

  28. Hockel M, Dornhofer N (2005) The hydra phenomenon of cancer: why tumors recur locally after microscopically complete resection. Cancer Res 65:2997–3002

    PubMed  Google Scholar 

  29. Castle PE, Ashfaq R, Ansari F, Muller CY (2005) Immunohistochemical evaluation of heat shock proteins in normal and preinvasive lesions of the cervix. Cancer Lett 229:245–252

    Article  PubMed  CAS  Google Scholar 

  30. Nomura M, Nomura N, Newcomb EW, Lukyanov Y, Tamasdan C, Zagzag D (2004) Geldanamycin induces mitotic catastrophe and subsequent apoptosis in human glioma cells. J Cell Physiol 201:374–384

    Article  PubMed  CAS  Google Scholar 

  31. Basto R, Gergely F, Draviam VM, Ohkura H, Liley K, Raff JW (2007) Hsp90 is required to localise cyclin B and Msps/ch-TOG to the mitotic spindle in Drosophila and humans. J Cell Sci 120:1278–1287

    Article  PubMed  CAS  Google Scholar 

  32. Franke TF, Hornik CP, Segev L, Shostak GA, Sugimoto C (2003) PI3K/Akt and apoptosis: size matters. Oncogene 22:8983–8998

    Article  PubMed  CAS  Google Scholar 

  33. Sato S, Fujita N, Tsuruo T (2000) Modulation of Akt kinase activity by binding to Hsp90. Proc Natl Acad Sci USA 97:10832–10837

    Article  PubMed  CAS  Google Scholar 

  34. Meares GP, Zmijewska AA, Jope RS (2004) Heat shock protein-90 dampens and directs signaling stimulated by insulin-like growth factor-1 and insulin. FEBS Lett 574:181–186

    Article  PubMed  CAS  Google Scholar 

  35. Sanderson S, Valenti M, Gowan S, Patterson L, Ahmad Z, Workman P, Eccles SA (2006) Benzoquinone ansamycin heat shock protein 90 inhibitors modulate multiple functions required for tumor angiogenesis. Mol Cancer Ther 5:522–532

    Article  PubMed  CAS  Google Scholar 

  36. Bisht KS, Bradbury CM, Mattson D, Kaushal A, Sowers A, Markovina S, Ortiz KL, Sieck LK, Isaacs JS, Brechbiel MW, Mitchell JB, Neckers LM, Gius D (2003) Geldanamycin and 17-allylamino-17-demethoxygeldanamycin potentiate the in vitro and in vivo radiation response of cervical tumor cells via the heat shock protein 90-mediated intracellular signaling and cytotoxicity. Cancer Res 63:8984–8995

    PubMed  CAS  Google Scholar 

  37. Dote H, Burgan WE, Camphausen K, Tofilon PJ (2006) Inhibition of hsp90 compromises the DNA damage response to radiation. Cancer Res 66:9211–9220

    Article  PubMed  CAS  Google Scholar 

  38. Sharp S, Workman P (2006) Inhibitors of the HSP90 molecular chaperone: current status. Adv Cancer Res 95:323–348

    Article  PubMed  CAS  Google Scholar 

  39. Chaudhury S, Welch TR, Blagg BS (2006) Hsp90 as a target for drug development. ChemMedChem 1:1331–1340

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

17-DMAG (NSC 707545) from Kosan Biosciences was supplied by the Cancer Therapy Evaluation Program of the National Cancer Institute (Rockville, MD). We are indebted to the research team of Richardson Technologies, Inc., particularly Bruno Chue, for their support with the live-cell imaging studies and data processing. We also thank Wen-Jiang Zhang and Dr. Xueyu Chen (Ontario Cancer Institute) for the HPLC measurements performed for 17-DMAG in xenograft samples and Melania Pintilie for help with the statistical analysis of the animal data. This study was supported by the Terry Fox Program Project Grant of the National Cancer Institute of Canada. J.S. was supported by the CIHR Training Program on Clinician Scientists in Molecular Oncologic Pathology (STP-53912).

Author information

Authors and Affiliations

  1. Department of Laboratory Medicine and Pathobiology, University of Toronto, 610 University Ave., 5th Floor, Rm 203, M5G 2M9, Toronto, ON, Canada

    Jörg Schwock & David W. Hedley

  2. Department of Medical Biophysics, University of Toronto, 610 University Ave., 5th Floor, Rm 203, M5G 2M9, Toronto, ON, Canada

    Nhu-An Pham & David W. Hedley

  3. Division of Applied Molecular Oncology, Princess Margaret Hospital, Ontario Cancer Institute, University of Toronto, 610 University Ave., 5th Floor, Rm 203, M5G 2M9, Toronto, ON, Canada

    Mary P. Cao & David W. Hedley

Authors
  1. Jörg Schwock
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Nhu-An Pham
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Mary P. Cao
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. David W. Hedley
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to David W. Hedley.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Schwock, J., Pham, NA., Cao, M.P. et al. Efficacy of Hsp90 inhibition for induction of apoptosis and inhibition of growth in cervical carcinoma cells in vitro and in vivo. Cancer Chemother Pharmacol 61, 669–681 (2008). https://doi.org/10.1007/s00280-007-0522-8

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00280-007-0522-8

Keywords

Search

Navigation