Abstract
Mutations at the p53 tumor suppressor gene locus are a frequent genetic alteration associated with human ovarian carcinoma. Little information exists regarding whether mutational events occur other than point mutations and large deletions, causing loss of heterozygosity. Small intragenic deletions and insertions in the p53 gene have been observed in various human neoplasias. We developed a multiplex polymerase chain reaction (MPCR) screening assay to amplify the complete p53 coding region from genomic DNA in a single step. Deletions and/or insertions were found in six out of 11 newly established ovarian carcinoma cell lines. MPCR detected deletions as small as 2bp, as confirmed by nucleotide sequence analysis. Most of the observed alterations (6/7) were homozygous or hemizygous. Structural aberrations of the p53 gene possibly leading to loss of p53 cell cycle control may be a consequence of a slipped-mispairing mechanism in rapid DNA replication during repetitious ovulation and wound repair of ovarian epithelial cells. MPCR may be a valuable tool for screening for possible p53 deletion and insertion mutations not only in ovarian cancer but also in other malignancies.
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References
Beenken SW, Karsenty G, Raycroft L, Lozano G (1991) An intron binding protein is required for transformation ability of p53. Nucleic Acids Res 19: 4747–4752.
Ben DY, Prideaux VR, Chow V, Benchimol S, Bernstein A (1988) Inactivation of the p53 oncogene by internal deletion or retroviral integration in erythroleukemic cell lines induced by Friend leukemia virus. Oncogene 3:179–185.
Beral V, Fraser P, Chilvers C (1978) Does pregnancy protect against ovarian cancer? Lancet 1:1083–1087.
Chamberlain JS, Gibbs RA, Ranier JE, Nguyen PN, Caskey CT (1988) Deletion screening of the Duchenne muscular dystrophy locus via multiplex DNA amplification. Nucleic Acids Res 16:111141–111156.
Eccles DM, Brett L, Lessells A, Gruber L, Lane D, Steel CM, Leonard RC (1992) Overexpression of the p53 protein and allele loss at 17p13 in ovarian carcinoma. Br J Cancer 65:40–44.
Fredrickson TN (1987) Ovarian tumors of the hen. Environ Health Perspect 73:35–51.
Frommentel CC de, Soussi T (1992) TP53 tumor suppressor gene: a model for investigating human mutagenesis. Genes Chromosom Cancer 4:1–15.
Godwin AK, Testa JR, Handel LM, Liu Z, Vanderveer LA, Tracey PA, Hamilton TC (1992) Spontaneous transformation of rat ovarian surface epithelial cells: association with cytogenetic changes and implications of repeated ovulation in the etiology of ovarian cancer. J Natl Cancer Inst 84:592–601.
Hartge P, Schiffman MH, Hoover R, McGowman L, Lesher L, Norris HJ (1989) A case-controlled study of epithelial ovarian cancer. Am J Obstet Gynecol 161:10–16.
Hartwell L (1992) Defects in a cell cycle checkpoint may be responsible for the genomic instability of cancer cells. Cell 71:543–546.
Hollstein M, Sidransky D, Vogelstein B, Harris CC (1991) p53 Mutations in human cancers. Science 253:49–53.
Jacobs IJ, Kohler MF, Wiseman RW, Marks JR, Whitaker R, Kerns BA, Humphrey P, Berchuck A, Ponder BA, Bast RCJ (1992) Clonal origin of epithelial ovarian carcinoma: analysis by loss of heterozygosity, p53 mutation, and X-chromosome inactivation. J Natl Cancer Inst 84:1793–1798.
Jego N, Thomas G, Hamelin R (1993) Short direct repeats flanking deletions, and duplicating insertions in p53 gene in human cancers. Oncogene 8:209–213.
Joly DJ, Lilienfeld AM, Diamond EL, Bross ID (1974) An epidemiologic study of the relationship of reproductive experience to cancer of the ovary. Am J Epidemiol 99:190–209.
Kastan MB, Zhan Q, el DW, Carrier F, Jacks T, Walsh WV, Plunkett BS, Vogelstein B, Fornace AJJ (1992) A mammalian cell cycle checkpoint pathway utilizing p53 and GADD45 is defective in ataxia-telangiectasia. Cell 71:587–597.
Kupryjanczyk J, Thor AD, Beauchamp R, Merritt V, Edgerton SM, Bell DA, Yandell DW (1993) p53 Gene mutations and protein accumulation in human ovarian cancer. Proc Natl Acad Sci USA 90:4961–4965.
Lingeman CH (1974) Etiology of cancer of the human ovary: a review. J Natl Cancer Inst 53:1603–1618.
Livingstone LR, White A, Sprouse J, Livanos E, Jacks T, Tlsty TD (1992) Altered cell cycle arrest and gene amplification potential accompany loss of wild-type p53. Cell 70:923–935.
Marks JR, Davidoff AM, Kerns BJ, Humphrey PA, Pence JC, Dodge RK, Clarke PD, Iglehart JD, Bast RJ, Berchuck A (1991) Overexpression and mutation of p53 in epithelial ovarian cancer. Cancer Res 51:2979–2984.
Mazars R, Pujol P, Maudelonde T, Jeanteur P, Theillet C (1991) p53 Mutations in ovarian cancer: a late event? Oncogene 6:1685–1690.
Milner BJ, Allan LA, Eccles DM, Kitchener HC, Leonard RCF, Kelly KF, Parkin DE, Haites NE (1993) p53 Mutation is a common genetic event in ovarian carcinoma. Cancer Res 53:2128–2132.
Moebus V, Gerharz CD, Press U, Moll R, Beck T, Mellin W, Pollow K, Knapstein PG, Kreienberg R (1992) Morphological, immunohistochemical and biochemical characterization of 6 newly established human ovarian carcinoma cell lines. Int J Cancer 52:76–84.
Mok C-H, Tsao S-W, Knapp RC, Fishbaugh PM, Lau CC (1992) Unifocal origin of advanced human epithelial ovarian cancers. Cancer Res 52:5119–5122.
Munroe DG, Rovinski B, Bernstein A, Benchimol S (1988) Loss of a highly conserved domain on p53 as a result of gene deletion during Friend virus-induced erythroleukemia. Oncogene 2:621–624.
Okamoto A, Sameshima Y, Yokoyama S, Terashima Y, Sugimura T, Terada M, Yokota J (1991) Frequent allelic losses and mutations of the p53 gene in human ovarian cancer. Cancer Res 51:5171–5176.
Rizzuto R, Nakase H, Darras B, Francke U, Fabrizi GM, Mengel T, Walsh F, Kadenbach B, DiMauro S, Schon EA (1989) A gene specifying subunit VIII of human cytochrome c oxidase is localized to chromosome 11 and is expressed in both muscle and non-muscle tissues. J Biol Chem 264:101595–101600.
Runnebaum IB, Nagarajan M, Bowman M, Soto D, Sukumar S (1991) Mutations in p53 as potential molecular markers for human breast cancer. Proc Natl Acad Sci USA 88:10657–10661.
Russell SEH, Hickey GI, Lowry WS, White P, Atkinson RJ (1990) Allele loss from chromosome 17 in ovarian cancer. Oncogene 5:1581–1583.
Sato T, Saito H, Morita R, Koi S, Lee JH, Nakamura Y (1991) Allelotype of human ovarian cancer. Cancer Res 51:5118–5122.
Soto D, Sukumar S (1992) Improved detection of mutations in the p53 gene in human tumors as single-stranded conformation polymorphs and double-stranded heteroduplex DNA. PCR Methods Appl 2:96–98.
Teneriello MG, Ebina M, Linnoila RI, Henry M, Nash JD, Park RC, Birrer MJ (1993) p53 and Ki-ras gene mutations in epithelial ovarian neoplasms. Cancer Res 53:3103–3108.
Tsao SW, Mok CH, Oike K, Muto M, Goodman HM, Sheets EE, Berkowitz RS, Knapp RC, Lau CC (1991) Involvement of p53 gene in the allelic deletion of chromosome 17p in human ovarian tumors. Anticancer Res 11:1975–1982.
Wilson JE (1958) Adeno-carcinomata in hens kept in a constant environment. Poult Sci 37:1253.
World Health Organization (1989) The World Health Organization collaborative study of neoplasia and steroid contraceptives: epithelial ovarian cancer and combined oral contraceptives. Int J Epidemiol 18:538–545.
Yaginuma Y, Westphal H (1992) Abnormal structure and expression of the p53 gene in human ovarian carcinoma cell lines. Cancer Res 52:4196–4199.
Yin Y, Tainsky MA, Bischoff FZ, Strong LC, Wahl GM (1992) Wild-type p53 restores cell cycle control and inhibits gene amplification in cells with mutant p53 alleles. Cell 70:937–948.
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Runnebaum, I.B., Tong, XW., Moebus, V. et al. Multiplex PCR screening detects small p53 deletions and insertions in human ovarian cancer cell lines. Hum Genet 93, 620–624 (1994). https://doi.org/10.1007/BF00201559
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DOI: https://doi.org/10.1007/BF00201559