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. 1999 Oct;81(4):592–599. doi: 10.1038/sj.bjc.6690735

Expression of caspases 3, 6 and 8 is increased in parallel with apoptosis and histological aggressiveness of the breast lesion

M Vakkala 1, P Pääkkö 1, Y Soini 1
PMCID: PMC2362889  PMID: 10574243

Abstract

The aim of this investigation was to study the expression of caspases 3, 6 and 8 and their association to apoptosis in preneoplastic and neoplastic lesions of the breast. The material consisted of nine benign breast epithelial hyperplasias, 15 atypical hyperplasias, 74 in situ and 82 invasive carcinomas. The extent of apoptosis was assessed by the TUNEL method and caspase 3, 6 and 8 expression by immunohistochemistry with specific antibodies. Increased caspase 3 immunopositivity, as compared to staining of normal breast ductal epithelium, was seen in 22% of benign epithelial hyperplasias, 25% of atypical hyperplasias, 58% of in situ carcinomas and 90% of invasive carcinomas. The corresponding percentages for caspase 6 and 8 were 11%, 25%, 60%, 87% and 22%, 57%, 84%, 83% respectively. In high-grade in situ lesions there were significantly more cases with strong caspase 3, 6 and 8 immunoreactivity than in low- and intermediate-grade lesions (P = 0.0045, P = 0.049 and P = 0.0001 respectively). In invasive carcinomas, however, no association between a high tumour grade and caspase 3, 6 or 8 expression was found (P = 0.27, P = 0.26 and P = 0.69 respectively). The mean apoptotic index was 0.14 ± 0.14% in benign epithelial hyperplasias, 0.17 ± 0.12% in atypical hyperplasias, 0.61 ± 0.88% in in situ carcinomas and 0.94 ± 1.21% in invasive carcinomas. In all cases strong caspase 3, 6 and 8 positivity was significantly associated with the extent of apoptosis (P < 0.001, P = 0.015 and P = 0.050 respectively). The results show that synthesis of caspases 3, 6 and 8 is up-regulated in neoplastic breast epithelial cells in parallel to the increase in the apoptotic index and progression of the breast lesions. © 1999 Cancer Research Campaign

Keywords: apoptosis, caspase, breast, carcinoma

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Selected References

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  1. Alnemri E. S., Livingston D. J., Nicholson D. W., Salvesen G., Thornberry N. A., Wong W. W., Yuan J. Human ICE/CED-3 protease nomenclature. Cell. 1996 Oct 18;87(2):171–171. doi: 10.1016/s0092-8674(00)81334-3. [DOI] [PubMed] [Google Scholar]
  2. Barge R. M., Willemze R., Vandenabeele P., Fiers W., Beyaert R. Differential involvement of caspases in apoptosis of myeloid leukemic cells induced by chemotherapy versus growth factor withdrawal. FEBS Lett. 1997 Jun 9;409(2):207–210. doi: 10.1016/s0014-5793(97)00507-3. [DOI] [PubMed] [Google Scholar]
  3. Barnes R., Masood S. Potential value of hormone receptor assay in carcinoma in situ of breast. Am J Clin Pathol. 1990 Nov;94(5):533–537. doi: 10.1093/ajcp/94.5.533. [DOI] [PubMed] [Google Scholar]
  4. Brancolini C., Lazarevic D., Rodriguez J., Schneider C. Dismantling cell-cell contacts during apoptosis is coupled to a caspase-dependent proteolytic cleavage of beta-catenin. J Cell Biol. 1997 Nov 3;139(3):759–771. doi: 10.1083/jcb.139.3.759. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bur M. E., Zimarowski M. J., Schnitt S. J., Baker S., Lew R. Estrogen receptor immunohistochemistry in carcinoma in situ of the breast. Cancer. 1992 Mar 1;69(5):1174–1181. doi: 10.1002/cncr.2820690518. [DOI] [PubMed] [Google Scholar]
  6. Caulín C., Salvesen G. S., Oshima R. G. Caspase cleavage of keratin 18 and reorganization of intermediate filaments during epithelial cell apoptosis. J Cell Biol. 1997 Sep 22;138(6):1379–1394. doi: 10.1083/jcb.138.6.1379. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chen L., Marechal V., Moreau J., Levine A. J., Chen J. Proteolytic cleavage of the mdm2 oncoprotein during apoptosis. J Biol Chem. 1997 Sep 5;272(36):22966–22973. doi: 10.1074/jbc.272.36.22966. [DOI] [PubMed] [Google Scholar]
  8. Chhanabhai M., Krajewski S., Krajewska M., Wang H. G., Reed J. C., Gascoyne R. D. Immunohistochemical analysis of interleukin-1beta-converting enzyme/Ced-3 family protease, CPP32/Yama/Caspase-3, in Hodgkin's disease. Blood. 1997 Sep 15;90(6):2451–2455. [PubMed] [Google Scholar]
  9. Clem R. J., Cheng E. H., Karp C. L., Kirsch D. G., Ueno K., Takahashi A., Kastan M. B., Griffin D. E., Earnshaw W. C., Veliuona M. A. Modulation of cell death by Bcl-XL through caspase interaction. Proc Natl Acad Sci U S A. 1998 Jan 20;95(2):554–559. doi: 10.1073/pnas.95.2.554. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Droin N., Dubrez L., Eymin B., Renvoizé C., Bréard J., Dimanche-Boitrel M. T., Solary E. Upregulation of CASP genes in human tumor cells undergoing etoposide-induced apoptosis. Oncogene. 1998 Jun 4;16(22):2885–2894. doi: 10.1038/sj.onc.1201821. [DOI] [PubMed] [Google Scholar]
  11. Elston C. W., Ellis I. O. Pathological prognostic factors in breast cancer. I. The value of histological grade in breast cancer: experience from a large study with long-term follow-up. Histopathology. 1991 Nov;19(5):403–410. doi: 10.1111/j.1365-2559.1991.tb00229.x. [DOI] [PubMed] [Google Scholar]
  12. Enari M., Sakahira H., Yokoyama H., Okawa K., Iwamatsu A., Nagata S. A caspase-activated DNase that degrades DNA during apoptosis, and its inhibitor ICAD. Nature. 1998 Jan 1;391(6662):43–50. doi: 10.1038/34112. [DOI] [PubMed] [Google Scholar]
  13. Faleiro L., Kobayashi R., Fearnhead H., Lazebnik Y. Multiple species of CPP32 and Mch2 are the major active caspases present in apoptotic cells. EMBO J. 1997 May 1;16(9):2271–2281. doi: 10.1093/emboj/16.9.2271. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Fraser A., Evan G. A license to kill. Cell. 1996 Jun 14;85(6):781–784. doi: 10.1016/s0092-8674(00)81005-3. [DOI] [PubMed] [Google Scholar]
  15. Fujita N., Tsuruo T. Involvement of Bcl-2 cleavage in the acceleration of VP-16-induced U937 cell apoptosis. Biochem Biophys Res Commun. 1998 May 19;246(2):484–488. doi: 10.1006/bbrc.1998.8587. [DOI] [PubMed] [Google Scholar]
  16. Harvey N. L., Butt A. J., Kumar S. Functional activation of Nedd2/ICH-1 (caspase-2) is an early process in apoptosis. J Biol Chem. 1997 May 16;272(20):13134–13139. doi: 10.1074/jbc.272.20.13134. [DOI] [PubMed] [Google Scholar]
  17. Helin H. J., Helle M. J., Kallioniemi O. P., Isola J. J. Immunohistochemical determination of estrogen and progesterone receptors in human breast carcinoma. Correlation with histopathology and DNA flow cytometry. Cancer. 1989 May 1;63(9):1761–1767. doi: 10.1002/1097-0142(19900501)63:9<1761::aid-cncr2820630918>3.0.co;2-3. [DOI] [PubMed] [Google Scholar]
  18. Holland R., Hendriks J. H., Vebeek A. L., Mravunac M., Schuurmans Stekhoven J. H. Extent, distribution, and mammographic/histological correlations of breast ductal carcinoma in situ. Lancet. 1990 Mar 3;335(8688):519–522. doi: 10.1016/0140-6736(90)90747-s. [DOI] [PubMed] [Google Scholar]
  19. Jänicke R. U., Sprengart M. L., Wati M. R., Porter A. G. Caspase-3 is required for DNA fragmentation and morphological changes associated with apoptosis. J Biol Chem. 1998 Apr 17;273(16):9357–9360. doi: 10.1074/jbc.273.16.9357. [DOI] [PubMed] [Google Scholar]
  20. Kerr J. F., Winterford C. M., Harmon B. V. Apoptosis. Its significance in cancer and cancer therapy. Cancer. 1994 Apr 15;73(8):2013–2026. doi: 10.1002/1097-0142(19940415)73:8<2013::aid-cncr2820730802>3.0.co;2-j. [DOI] [PubMed] [Google Scholar]
  21. Kluck R. M., Bossy-Wetzel E., Green D. R., Newmeyer D. D. The release of cytochrome c from mitochondria: a primary site for Bcl-2 regulation of apoptosis. Science. 1997 Feb 21;275(5303):1132–1136. doi: 10.1126/science.275.5303.1132. [DOI] [PubMed] [Google Scholar]
  22. Krajewska M., Wang H. G., Krajewski S., Zapata J. M., Shabaik A., Gascoyne R., Reed J. C. Immunohistochemical analysis of in vivo patterns of expression of CPP32 (Caspase-3), a cell death protease. Cancer Res. 1997 Apr 15;57(8):1605–1613. [PubMed] [Google Scholar]
  23. Krajewski S., Gascoyne R. D., Zapata J. M., Krajewska M., Kitada S., Chhanabhai M., Horsman D., Berean K., Piro L. D., Fugier-Vivier I. Immunolocalization of the ICE/Ced-3-family protease, CPP32 (Caspase-3), in non-Hodgkin's lymphomas, chronic lymphocytic leukemias, and reactive lymph nodes. Blood. 1997 May 15;89(10):3817–3825. [PubMed] [Google Scholar]
  24. Krajewski S., Krajewska M., Shabaik A., Miyashita T., Wang H. G., Reed J. C. Immunohistochemical determination of in vivo distribution of Bax, a dominant inhibitor of Bcl-2. Am J Pathol. 1994 Dec;145(6):1323–1336. [PMC free article] [PubMed] [Google Scholar]
  25. Kroemer G. The proto-oncogene Bcl-2 and its role in regulating apoptosis. Nat Med. 1997 Jun;3(6):614–620. doi: 10.1038/nm0697-614. [DOI] [PubMed] [Google Scholar]
  26. Li P., Nijhawan D., Budihardjo I., Srinivasula S. M., Ahmad M., Alnemri E. S., Wang X. Cytochrome c and dATP-dependent formation of Apaf-1/caspase-9 complex initiates an apoptotic protease cascade. Cell. 1997 Nov 14;91(4):479–489. doi: 10.1016/s0092-8674(00)80434-1. [DOI] [PubMed] [Google Scholar]
  27. Lipponen P., Aaltomaa S., Kosma V. M., Syrjänen K. Apoptosis in breast cancer as related to histopathological characteristics and prognosis. Eur J Cancer. 1994;30A(14):2068–2073. doi: 10.1016/0959-8049(94)00342-3. [DOI] [PubMed] [Google Scholar]
  28. Liu X., Kim C. N., Yang J., Jemmerson R., Wang X. Induction of apoptotic program in cell-free extracts: requirement for dATP and cytochrome c. Cell. 1996 Jul 12;86(1):147–157. doi: 10.1016/s0092-8674(00)80085-9. [DOI] [PubMed] [Google Scholar]
  29. Mack L., Kerkvliet N., Doig G., O'Malley F. P. Relationship of a new histological categorization of ductal carcinoma in situ of the breast with size and the immunohistochemical expression of p53, c-erb B2, bcl-2, and ki-67. Hum Pathol. 1997 Aug;28(8):974–979. doi: 10.1016/s0046-8177(97)90014-9. [DOI] [PubMed] [Google Scholar]
  30. Manon S., Chaudhuri B., Guérin M. Release of cytochrome c and decrease of cytochrome c oxidase in Bax-expressing yeast cells, and prevention of these effects by coexpression of Bcl-xL. FEBS Lett. 1997 Sep 22;415(1):29–32. doi: 10.1016/s0014-5793(97)01087-9. [DOI] [PubMed] [Google Scholar]
  31. Martins L. M., Kottke T., Mesner P. W., Basi G. S., Sinha S., Frigon N., Jr, Tatar E., Tung J. S., Bryant K., Takahashi A. Activation of multiple interleukin-1beta converting enzyme homologues in cytosol and nuclei of HL-60 cells during etoposide-induced apoptosis. J Biol Chem. 1997 Mar 14;272(11):7421–7430. doi: 10.1074/jbc.272.11.7421. [DOI] [PubMed] [Google Scholar]
  32. Minn A. J., Vélez P., Schendel S. L., Liang H., Muchmore S. W., Fesik S. W., Fill M., Thompson C. B. Bcl-x(L) forms an ion channel in synthetic lipid membranes. Nature. 1997 Jan 23;385(6614):353–357. doi: 10.1038/385353a0. [DOI] [PubMed] [Google Scholar]
  33. Mustonen M., Raunio H., Päkkö P., Soini Y. The extent of apoptosis is inversely associated with bcl-2 expression in premalignant and malignant breast lesions. Histopathology. 1997 Oct;31(4):347–354. doi: 10.1046/j.1365-2559.1997.2710877.x. [DOI] [PubMed] [Google Scholar]
  34. Muzio M., Chinnaiyan A. M., Kischkel F. C., O'Rourke K., Shevchenko A., Ni J., Scaffidi C., Bretz J. D., Zhang M., Gentz R. FLICE, a novel FADD-homologous ICE/CED-3-like protease, is recruited to the CD95 (Fas/APO-1) death--inducing signaling complex. Cell. 1996 Jun 14;85(6):817–827. doi: 10.1016/s0092-8674(00)81266-0. [DOI] [PubMed] [Google Scholar]
  35. Nagata S. Apoptosis by death factor. Cell. 1997 Feb 7;88(3):355–365. doi: 10.1016/s0092-8674(00)81874-7. [DOI] [PubMed] [Google Scholar]
  36. O'Malley F. P., Vnencak-Jones C. L., Dupont W. D., Parl F., Manning S., Page D. L. p53 mutations are confined to the comedo type ductal carcinoma in situ of the breast. Immunohistochemical and sequencing data. Lab Invest. 1994 Jul;71(1):67–72. [PubMed] [Google Scholar]
  37. Patel T., Gores G. J., Kaufmann S. H. The role of proteases during apoptosis. FASEB J. 1996 Apr;10(5):587–597. doi: 10.1096/fasebj.10.5.8621058. [DOI] [PubMed] [Google Scholar]
  38. Rao L., Perez D., White E. Lamin proteolysis facilitates nuclear events during apoptosis. J Cell Biol. 1996 Dec;135(6 Pt 1):1441–1455. doi: 10.1083/jcb.135.6.1441. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Soini Y., Päkkö P., Lehto V. P. Histopathological evaluation of apoptosis in cancer. Am J Pathol. 1998 Oct;153(4):1041–1053. doi: 10.1016/S0002-9440(10)65649-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Soini Y., Virkajärvi N., Lehto V. P., Päkkö P. Hepatocellular carcinomas with a high proliferation index and a low degree of apoptosis and necrosis are associated with a shortened survival. Br J Cancer. 1996 May;73(9):1025–1030. doi: 10.1038/bjc.1996.199. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Tan X., Martin S. J., Green D. R., Wang J. Y. Degradation of retinoblastoma protein in tumor necrosis factor- and CD95-induced cell death. J Biol Chem. 1997 Apr 11;272(15):9613–9616. doi: 10.1074/jbc.272.15.9613. [DOI] [PubMed] [Google Scholar]
  42. Thornberry N. A., Lazebnik Y. Caspases: enemies within. Science. 1998 Aug 28;281(5381):1312–1316. doi: 10.1126/science.281.5381.1312. [DOI] [PubMed] [Google Scholar]
  43. Thornberry N. A., Peterson E. P., Zhao J. J., Howard A. D., Griffin P. R., Chapman K. T. Inactivation of interleukin-1 beta converting enzyme by peptide (acyloxy)methyl ketones. Biochemistry. 1994 Apr 5;33(13):3934–3940. doi: 10.1021/bi00179a020. [DOI] [PubMed] [Google Scholar]
  44. Törmänen U., Eerola A. K., Rainio P., Vähäkangas K., Soini Y., Sormunen R., Bloigu R., Lehto V. P., Päkkö P. Enhanced apoptosis predicts shortened survival in non-small cell lung carcinoma. Cancer Res. 1995 Dec 1;55(23):5595–5602. [PubMed] [Google Scholar]
  45. Virkajärvi N., Päkkö P., Soini Y. Apoptotic index and apoptosis influencing proteins bcl-2, mcl-1, bax and caspases 3, 6 and 8 in pancreatic carcinoma. Histopathology. 1998 Nov;33(5):432–439. doi: 10.1046/j.1365-2559.1998.00553.x. [DOI] [PubMed] [Google Scholar]
  46. Wang T. T., Phang J. M. Effects of estrogen on apoptotic pathways in human breast cancer cell line MCF-7. Cancer Res. 1995 Jun 15;55(12):2487–2489. [PubMed] [Google Scholar]
  47. Yang E., Korsmeyer S. J. Molecular thanatopsis: a discourse on the BCL2 family and cell death. Blood. 1996 Jul 15;88(2):386–401. [PubMed] [Google Scholar]
  48. Yang J., Liu X., Bhalla K., Kim C. N., Ibrado A. M., Cai J., Peng T. I., Jones D. P., Wang X. Prevention of apoptosis by Bcl-2: release of cytochrome c from mitochondria blocked. Science. 1997 Feb 21;275(5303):1129–1132. doi: 10.1126/science.275.5303.1129. [DOI] [PubMed] [Google Scholar]
  49. van de Vijver M. J., Peterse J. L., Mooi W. J., Wisman P., Lomans J., Dalesio O., Nusse R. Neu-protein overexpression in breast cancer. Association with comedo-type ductal carcinoma in situ and limited prognostic value in stage II breast cancer. N Engl J Med. 1988 Nov 10;319(19):1239–1245. doi: 10.1056/NEJM198811103191902. [DOI] [PubMed] [Google Scholar]

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