Regular Article

British Journal of Cancer (2001) 85, 115–121. doi:10.1054/bjoc.2001.1850 www.bjcancer.com
Published online 3 July 2001

Induction of apoptosis in prostate carcinoma cells by BH3 peptides which inhibit Bak/Bcl-2 interactions

N M Finnegan1, J F Curtin1, G Prevost2, B Morgan3 and T G Cotter1

  1. 1Dept of Biochemistry, University College Cork, Lee Maltings, Prospect Row, Cork, Ireland
  2. 2Beaufour Ipsen, Institut Henri Beaufour, 5 Avenue du Canada, Les Ulis Cedex, 91966, France
  3. 3Biomeasure Inc., 27 Maple St., Milford, MA

Received 27 November 2000; Revised 26 March 2001; Accepted 29 March 2001.

Top

Abstract

Interactions between proteins of the Bcl-2 family play an important role in the regulation of apoptosis. Anti-apoptotic family members can heterodimerize with pro-apoptotic family members and antagonize their function, thus protecting against death. In cells protected from death by overexpression of Bcl-2 much of the Bax is present in Bax/Bcl-2 hetero-multimers and its death signal is blocked as it cannot homodimerize. This led us to use the Bcl-2/Bax heterodimer as a target for new compounds which may provide a therapy particularly suited to tumour cells for which resistance to conventional therapy is associated with elevated expression of Bcl-2. We assessed whether apoptosis could be induced in prostate tumour cells by blocking this heterodimerization with synthetic peptide sequences derived from the BH3 domain of pro-apoptotic Bcl-2 family members. Prostate cells were found to undergo up to 40% apoptosis 48 h following the introduction of synthetic peptides from the BH3 domains of Bax and Bak. The caspase inhibitor z-VAD.fmk provided protection against apoptosis mediated by these peptides. Immunoprecipitation studies revealed that introduction of peptides derived from the BH3 regions of Bak and Bax into cells blocked Bak/Bcl-2 heterodimerization. These data suggest that by blocking the dimerization through which Bcl-2 would normally inhibit apoptosis the apoptotic pathway driven by Bak was re-opened. © 2001 Cancer Research Campaign http://www.bjcancer.com

Keywords:

prostate, apoptosis, BH3, Bcl-2, peptide

Top

References

  1. Adams JM and Cory S (1998) The Bcl-2 protein family: Arbiters of cell survival. Science 281: 1322–1326 | Article | PubMed | ISI | ChemPort |
  2. Allen RT, Cluck MW and Agrawal DK (1998) Mechanisms controlling cellular suicide: role of Bcl-2 and caspases. Cell Mol Life Sci 54: 427–445 | Article | PubMed | ISI | ChemPort |
  3. Berchem GJ, Bosseler M, Sugars LY, Voeller HJ, Zeitlin S and Gellmann EP (1995) Androgens induce resistance to Bcl-2-mediated apoptosis in LNCaP prostate cancer cells. Cancer Res 55: 735–738 | PubMed |
  4. Chittenden T, Flemmington C, Houghton AB, Ebb RG, Gallo GL, Elangovan B, Chinnadurai G and Lutz RJ (1995) A conserved domain in Bak, distinct from BH1 and BH2, mediates cell death and protein binding function. EMBO J 14: 5589–5596 | PubMed | ISI | ChemPort |
  5. Colombel M, Symmans F, Gil S, O'Toole KM, Chopin D, Benson M, Olsson CA, Korsmeyer S and Buttyan R (1993) Detection of the apoptosis-suppressing oncoprotein bcl-2 in hormone refractory human prostate cancers. Am J Pathol 143: 390–400 | PubMed | ISI | ChemPort |
  6. Diaz JL, Oltersdorf T, Horne W, McConnell M, Wilson G, Weeks S, Garcia T and Fritz CF (1997) A common binding site mediates heterodimerization and homodimerization of Bcl-2 family members. J Biol Chem 272, 17 11350–11355 | Article | PubMed | ISI | ChemPort |
  7. Haldar S, Chintapalli J and Croce CM (1996) Taxol induces Bcl-2 phosphorylation and death in prostate cancer cells. Cancer Res 56: 1253–1255 | PubMed | ISI | ChemPort |
  8. Han JP, Sabbatini P, Perez D, Rao L, Modha D and White E (1996) The E1B 19K protein blocks apoptosis by interacting with and inhibiting the p53-inducible and death-promoting Bax protein. Genes Dev 10: 461–477 | PubMed | ISI | ChemPort |
  9. Hirotani M, Zhang Y, Fujita N, Naito M and Tsuruo T (1999) NH2-terminal BH4 domain of Bcl-2 is functional for heterodimerization with Bax and inhibition of apoptosis. J Biol Chem 274: 20415–20420 | Article | PubMed | ChemPort |
  10. Jurgensmeier JM, Xie Z, Deveraux Q, Ellerby L, Bredesen D and Reed JC (1998) Bax directly induces release of cytochrome c from isolated mitochondria. Proc Natl Acad Sci USA 95: 4997–5002 | Article | PubMed | ISI | ChemPort |
  11. Krajewska M, Krajewski S, Epstein JI, Shabaik A, Sauvageot J, Song K, Kitada S and Reed JC (1996) Immunohistochemical analysis of bcl-2, bax, bcl-xL, and mcl-1 expression on prostate cancers. Am J Pathol 148: 1567–1576 | PubMed | ISI | ChemPort |
  12. Kreis W (1995) Current chemotherapy and future directions in research for the treatment of advanced hormone-refractory prostate cancer. Cancer Invest 13: 296–312 | PubMed |
  13. Kroemer G (1997) The proto-oncogene Bcl-2 and its role in regulating apoptosis. Nature Med 3(6): 614–620 | Article |
  14. Mackey TJ, Borkowski A, Amin P, Jacobs SC and Kyprianou N (1998) bcl-2/bax ratio as a predictive marker for therapeutic response to radiotherapy in patients with prostate cancer. Urology 52(65): 1085–1090 | Article | PubMed | ISI | ChemPort |
  15. McDonnell TJ, Troncoso P, Brisbay SM, Logothetis C, Chung LW, Hsieh JT, Tu SM and Campbell ML (1992) Expression of the proto-oncogene Bcl-2 in the prostate and its association with emergence of androgen-independent prostate cancer. Cancer Res 52: 6940–6944 | PubMed | ISI | ChemPort |
  16. Mundle SD, Gregory SA, Preislert HD and Raza A (1996) Enzymatic programming of apoptotic cell death. Pathiobiol 64: 161–170
  17. Oltvai ZN, Milliman CL and Korsmeyer SJ (1993) Bcl-2 heterodimerizes in-vivo with a conserved homolog, Bax, that accelerates programmed cell death. Cell 74: 609–619 | Article | PubMed | ISI | ChemPort |
  18. Orth K and Dixit VM (1997) Bik and Bak induce apoptosis downstream of CrmA but upstream of inhibitor of apoptosis. J Biol Chem 272: 8841–8844 | Article | PubMed | ISI | ChemPort |
  19. Otter I, Conus S, Ravn U, Rager M, Olivier R, Monney L, Fabbro D and Borner C (1998) The binding properties and biological activities of Bcl-2 and Bax in cells exposed to apoptotic stimuli. J Biol Chem 273: 6110–6120 | Article | PubMed | ISI | ChemPort |
  20. Pan G, O'Rourke K and Dixit VM (1998) Caspase-9, Bcl-XL, and Apaf-1 form a terniary complex. J Biol Chem 273: 5841–5845 | Article | PubMed | ISI | ChemPort |
  21. Rokhlin OW, Bishop GA, Hostager BS, Waldschmidt TJ, Sidorenko SP, Pavloff N, Kiefer MC, Umansky SR, Glover RA and Cohen MB (1997) Fas-mediated apoptosis in human prostatic carcinoma cell lines. Cancer Res 57: 1758–1768 | PubMed | ISI | ChemPort |
  22. Tang DG and Porter AT (1997) Target to apoptosis: a hopeful weapon for prostate cancer. Prostate 32: 284–293 | Article | PubMed | ISI | ChemPort |
  23. Yang E and Korsmeyer SJ (1996) Molecular thanatopsis: a discourse on the Bcl-2 family and cell death. Blood 88: 386–401 | PubMed | ISI | ChemPort |
  24. Yang E, Zha J, Jockel J, Boise LH, Thompson CB and Korsmeyer SJ (1995) Bad, a heterodimeric partner for Bcl-xLand Bcl-2, displaces Bax and promotes cell death. Cell 80: 285–291 | Article | PubMed | ISI | ChemPort |
  25. Yin XM, Oltvai ZN and Korsmeyer SJ (1994) BH1 and BH2 domains of Bcl-2 are required for inhibition of apoptosis and heterodimerization with Bax. Nature 369: 321–323 | Article | PubMed | ISI | ChemPort |
  26. Zha H, Aime-Sempe C, Sato T and Reed JC (1996) Proapoptotic protein Bax heterodimerises with Bcl-2 and homodimerizes with Bax via a novel domain (BH3) distinct from BH1 and BH2. J Biol Chem 271: 7440–7444 | Article | PubMed | ISI | ChemPort |
  27. Zhou XM, Liu Y, Payne G, Lutz RJ and Chittenden T (2000) Growth factors inactivate the cell death promoter BAD by phosphorylation of its BH3 domain on Ser155. J Biol Chem 275: 25046–25051 | Article | PubMed | ISI | ChemPort |
  28. Zhu W, Cowie A, Wasfy GW, Penn LZ, Leber B and Andrews DW (1996) Bcl-2 mutants with restricted subcellular location reveal spatially distinct pathways for apoptosis in different cell types. EMBO J 15: 4130–4141 | PubMed | ISI | ChemPort |