Analysis of mutant P53 protein in osteosarcomas and other malignant and benign lesions of bone

  • Yoshimichi Ueda
  • Barbara Dockhorn-Dworniczak
  • Sebastian Blasius
  • Walter Mellin
  • Paul Wuisman
  • Werner Böcker
  • Albert Roessner
Original Papers Clinical Oncology

Summary

Alterations of tumour suppressor genes are considered crucial steps in the development of human cancers. Expressions of p53 protein, a product of the tumour suppressor gene altered most commonly in human cancers examined so far, were investigated immunohistochemically in 18 osteosarcomas and 40 other malignant and benign lesions of bone. A monoclonal antibody clone PAb240, which recognizes a common conformational epitope of mutant p53 proteins, stained nuclei of tumour cells in 12 of 18 osteosarcomas (67%). Six tumours (33%) particularly showed positive immunoreactions in more than half of the tumour cells. PAb240 also stained tumour cells in a small number of other malignant bone tumours, such as malignant fibrous histiocytoma, chondrosarcoma, and Ewing's sarcomas. Furthermore, a small number of cells of giant-cell tumours were positively stained. In contrast, PAb240 was completely negative in 21 benign bone tumours and reactive lesions examined. Another monoclonal antibody clone PAb1801, which reacts with both wild- and mutant-type p53 protein, reacted in nuclei of tumour cells of 7 osteosarcomas (39%). Most of those also reacted with PAb240. PAb1801 was expressed much more frequently in other malignant bone tumours and giant-cell tumours. In addition, PAb1801 showed intranuclear positive reactions in tumour cells of a benign chondroblastoma, and reactive cells such as actively proliferating preosteoblasts in a myositis ossificans and osteoclast-like giant cells in a giantcell tumour. The immunoelectron-microscopic observation that p53 protein was localized in euchromatic areas of nuclei of osteosarcoma cells supported the specificity of immunoreaction for p53 protein, indicating an active role of p53 protein in the regulation of DNA synthesis and transcription. These findings suggest that point mutation of the p53 gene is frequently involved in the development of osteosarcomas. PAb240 may be a useful tool not only in screening point mutations of the p53 gene in osteosarcomas but also in the differential diagnosis between osteosarcomas and reactive bone-forming lesions. Expressions of mutant p53 protein were not correlated with any clinical or pathological factors examined, although the results should be confirmed in studies of a large number of osteosarcomas.

Key words

p53 protein Point mutation Osteosarcoma Bone tumour Immunohistochemistry Immunoelectron microscopy 

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References

  1. Araki N, Uchida A, Kimura T, Yoshikawa H, Aoki Y, Ueda T, Takai S, Miki T, Ono K (1990) Involvement of the retinoblastoma gene in primary osteosarcomas and other bone and soft tissue tumors. Clin Orthop 270:271–277Google Scholar
  2. Baker SJ, Fearon ER, Nigro JM, Hamilton SR, Preisinger AC, Jessup JM, van Tuinen P, Ledbetter DF, Barker DF, Nakamura Y, White R, Vogelstein B (1989) Chromosome 17 deletions and p53 gene mutations in colorectal carcinomas. Science 244:217–221Google Scholar
  3. Banks L, Matlashewski G, Crawford L, (1986) Isolation of human p53 specific monoclonal antibodies and their use in the studies of human p53 expression. Eur J Biochem 159:529–534Google Scholar
  4. Bloom W, Fawcett DW (1975) A text book of histology 10th edn. Saunders, PhiladelphiaGoogle Scholar
  5. Chiba I, Takahashi T, Nau MM, D'Amico D, Curier DT, Mitsudomi T, Buchhagen DL, Carbone D, Piantadosi S, Koga H, Reissmann PT, Slamon DJ, Holmes EC, Minna JD (1990) Mutations in the p53 gene are frequent in primary, resected non-small cell cancer. Oncogene 5:1603–1610Google Scholar
  6. Cordell JL, Falini B, Erber WN, Ghosh AK, Abdulaziz M, MacDonald S, Pulford KAF, Stein H, Mason DY (1984) Immunoenzymatic labeling of monoclonal antibodies using immune complexes of alkaline phosphatase and monoclonal anti-alkaline phosphatases (APAAP) complexes. J Histochem Cytochem 32:219–229Google Scholar
  7. Cossman J, Schlegel R (1991) p53 in the diagnosis of human neoplasia. JNCI 83:980–981Google Scholar
  8. Dahlin DC, Unni KK (1986) Bone tumours. General aspects and data on 8542 cases, 4th edn. Thomas, Springfield IllGoogle Scholar
  9. Eliyasu D, Michalovitz D, Eliyasu S, Pinhasi-Kimhi O, Oren M (1989) Wild-type p53 can inhibit oncogene-mediated focus formation. Proc Natl Acad Sci USA 86:8763–8767Google Scholar
  10. Felix CA, Kappel CC, Mitsudomi T, Nau MM, Tsokos M, Crouch GD, Nisen PD, Winick NJ, Helman LJ (1992) Frequency and diversity of p53 mutations in childhood rabdomyosarcoma. Cancer Res 52:2243–2247Google Scholar
  11. Finlay CA, Hinds PW, Levine AJ (1989) The p53 proto-oncogene can act as a suppressor of transformation. Cell 57:1083–1093Google Scholar
  12. Frankel RH, Bayona W, Koslow M, Newcomb EW (1992) p53 mutations in human malignant gliomas: comparison of loss of heterozygosity with mutation frequency. Cancer Res 52:1427–1433Google Scholar
  13. Friend SH, Bernard R, Rogeli S, Weinberg RA, Rapaport JM, Albert DM, Drija TP (1986) A human DNA segment with properties of the gene that predisposes to retinoblastoma and osteosarcoma. Nature 323:643–646Google Scholar
  14. Gannon JV, Greaves R, Iggo R, Lane P (1990) Activating mutations in p53 produce a common conformational effect. A monoclonal antibody specific for mutation for the mutant form. EMBO J 9:1595–1602Google Scholar
  15. Hall PA, Ray A, Lemoine NR, Midgley CA, Kransz T, Lane DP (1991) p53 immunostaining as a marker of malignant disease in diagnostic cytopathology. Lancet 338:513–514Google Scholar
  16. Hollstein M, Sidransky D, Vogelstein B, Harris CC (1991) p53 mutations in human cancers. Science 253:49–53Google Scholar
  17. Iggo R, Gatter K, Bartek J, Lane D, Harris AL (1990) Increased expression of mutant forms of p53 oncogene in primary lung cancer. Lancet 335:675–679Google Scholar
  18. Kohler MF, Berchuck A, Davidoff AM, Humphrey PA, Dodge RK, Igelhart JD, Soper JT, Clarke-Pearson DL, Bast RC, Marks JR (1992) Overexpression and mutation of p53 in endometrial carcinoma. Cancer Res 52:1622–1627Google Scholar
  19. Ladanyi M, Traganos F, Huvos AG (1989) Benign metastasizing giant cell tumors of bone. A DNA flow cytometric study. Cancer 64:1521–1526Google Scholar
  20. Lane DP, Crawford LV (1979) T antigen is bound to a host protein in SV40-transformed cells. Nature 278:261–263Google Scholar
  21. Levine AJ, Momand J, Finlay CA (1991) The p53 tumour suppressor gene. Nature 351:453–456Google Scholar
  22. Linzer DIH, Levine AJ (1979) Characterization of a 54K dalton cellular SV40 tumour antigen present in SV40 transformed cells and uninfected embryonal carcinoma cells. Cell 17:43–52Google Scholar
  23. Look AT, Douglass EC, Meyer WH (1988) Clinical importance of neardiploid tumor stem lines in patients with osteosarcoma of an extremity. N Engl J Med 318:1567–1572Google Scholar
  24. Mankin HJ, Gebhardt MC, Springfield DS, Litwak GJ, Kusazaki K, Rosenberg AE (1991) Flow cytometric studies of human osteosarcoma. Clin Orthop 270:169–180Google Scholar
  25. Marks JR, Davidoff AM, Humphrey PA, Pence JC, Dodge RK, Clarke-Pearson DL, Iglehart JD, Bast RC, Berchuck A (1991) Overexpression and mutation of p53 in epithelial ovarian cancer. Cancer Res 51:2979–2984Google Scholar
  26. Masuda H, Miller C, Koeffler HP, Battifora H, Cline MJ (1987) Rearrangement of the p53 gene in human osteogenic sarcomas. Proc Natl Acad Sci USA 84:7716–7719Google Scholar
  27. McBride OW, Merry D, Givol D (1986) The gene for human p53 cellular tumor antigen is located on chromosome 17 short arm (17p13). Proc Natl Acad Sci USA 83:130–134Google Scholar
  28. Miller CW, Aslo A, Tsay C, Slamon D, Ishizaki K, Toguchida J, Yamamuro T, Lampkin B, Koeffler HP (1990) Frequency and structure of p53 rearrangements in human osteosarcoma. Cancer Res 50:7950–7954Google Scholar
  29. Nigro JM, Baker SJ, Preisinger AC, Jessup JM, Hostetter R, Cleary K, Bigner SH, Davidson N, Baylin S, Devilee P, Glover T, Collins FS, Weston A, Modali R, Harris CC, Vogelstein B (1989) Mutations in the p53 gene occur in diverse human tumour types. Nature 342:705–708Google Scholar
  30. Porter PL, Gown AM, Kramp SG, Coltrera MD (1992) Widespread p53 overexpression in human malignant tumors. An immunohistochemical study using methacarn-fixed, embedded tissue. Am J Pathol 140:145–153Google Scholar
  31. Purdie CA, O'Grady J, Piris J, Wyllie AH, Bird CC (1991) p53 expression in colorectal tumors. Am J Pathol 138:807–813Google Scholar
  32. Rodrigues NR, Rowan A, Smith MEF, Kerr IB, Bodmer WF, Gannon JV, Lane DP (1990) p53 mutations in colorectal cancer. Proc Natl Acad Sci USA 87:7555–7559Google Scholar
  33. Rosen G, Caparros B, Huvos AG, Kosloff C, Nirenberg A, Cacavio A, Marcove RC, Lane JM, Mehta B, Urban C (1982) Preoperative chemotherapy for osteogenic sarcoma: selection of postoperative adjuvant chemotherapy based on the response of the primary tumor to preoperative chemotherapy. Cancer 49:1221–1230Google Scholar
  34. Salzer-Kuntschik M, Delling D, Beron G, Sigmund R (1983) Morphological grades of regression on osteosarcoma after polychemotherapy — Study COSS 80. J Cancer Res Clin Oncol 106:21–24Google Scholar
  35. Takahashi T, Nau MM, Chiba I, Birrer MJ, Rosenberg RK, Vinocour M, Levitt M, Pass H, Gazdar AF, Minna JD (1989) p53 a frequent target for genetic abnormalities in lung cancer. Science 246:491–494Google Scholar
  36. Toguchida J, Ishizaki K, Sasaki MS, Ikenaga M, Sugimoto M, Kotoura Y, Yamamuro T (1988) Chromosomal reorganization for the expression of recessive mutatioin of retinoblastoma susceptibility gene in the development of osteosarcoma. Cancer Res 48:3939–3943Google Scholar
  37. Toguchida J, Ishizaki K, Nakamura Y, Sasaki MS, Ikenaga M, Kato M, Sugimoto M, Kotoura Y, Yamamuro T (1989) Assignment of common allele loss of osteosarcoma to the subregion 17p13. Cancer Res 49:6247–6251Google Scholar
  38. Walker RA, Dearing SJ, Lane DP, Varley JM (1991) Expression of p53 protein in infiltrating and in situ breast carcinomas. J Pathol 165:203–211Google Scholar
  39. Wynford-Thomas D (1991) Oncogenes and anti-oncogenes; the molecular basis of tumour behaviour. J Pathol 165:187–201Google Scholar
  40. Wynford-Thomas D (1992) p53 in tumour pathology: can we trust immunocytochemistry? J Pathol 166:329–330Google Scholar
  41. Yamaguchi T, Toguchida J, Yamamuro T, Kotoura Y, Takada N, Kawaguchi N, Kaneko Y, Nakamura Y, Sasaki MS (1992) Allelotype analysis in osteosarcoma — frequent allele loss on 3q, 17p, and 18q. Cancer Res 52:2419–2423Google Scholar

Copyright information

© Springer-Verlag 1993

Authors and Affiliations

  • Yoshimichi Ueda
    • 1
  • Barbara Dockhorn-Dworniczak
    • 1
  • Sebastian Blasius
    • 1
  • Walter Mellin
    • 1
  • Paul Wuisman
    • 2
  • Werner Böcker
    • 1
  • Albert Roessner
    • 1
  1. 1.Gerhard-Domagk Institute of PathologyUniversity of MünsterMünsterFederal Republic of Germany
  2. 2.Department of OrthopaedicsUniversity of MünsterMünsterFederal Republic of Germany

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