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Molecular Diagnostics and Genetics |
1 Department of Pathology and Laboratory Medicine, Mount Sinai Hospital, University of Toronto, Toronto, Ontario, Canada.2 Department of Epidemiology and Population Health, Albert Einstein College of Medicine, Bronx, NY.
aAddress correspondence to this author at: Department of Pathology and Laboratory Medicine, Mount Sinai Hospital, 600 University Ave. Suite 600, Toronto, Ontario M5G 1X5, Canada. Fax 416-586-8628; e-mail rkandel{at}mtsinai.on.ca.
Background: Routine tissue processing has generated banks of paraffin-embedded tissue that could be used in retrospective cohort studies to study the molecular changes that occur during cancer development. The purpose of this study was to determine whether a p53 microarray could be used to sequence the p53 gene in DNA extracted from formalin-fixed, paraffin-embedded (FFPE) tissues.
Methods: DNA was extracted from 70 FFPE breast cancer tissue specimens. p53 was sequenced with an oligonucleotide microarray (p53 GeneChip®; Affymetrix), and the results were compared with the results obtained from direct sequencing.
Results: DNA was extracted from 62 of 70 cases. We identified 26 mutations in 24 of the 62 cases by the p53 GeneChip. No polymorphisms were detected, and exon 4 could not be evaluated in 20 cases. There were 43 genetic alterations detected by direct sequencing in 35 of the 62 cases. These consisted of 26 polymorphisms and 17 mutations in exons or splice sites. Fifteen mutations were identified by both methods. Direct sequencing detected significantly more gene alterations (43 of 54) in DNA extracted from FFPE tissue than the p53 GeneChip (26 of 54; P = 0.018). However, if the changes in exon 4 were eliminated from this comparison, the p53 GeneChip detected 26 of 27 mutations compared with direct sequencing, which identified 16 of 27 mutations. (P = 0.016).
Conclusions: A combination of oligonucleotide microarray and direct sequencing may be necessary to accurately identify p53 gene alterations in FFPE breast cancer. The p53 GeneChip cannot be used to detect exon 4 polymorphisms (codon 72) in FFPE breast cancer tissue.
The following articles in journals at HighWire Press have cited this article:
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  D. S.P. Tan, M. B.K. Lambros, S. Rayter, R. Natrajan, R. Vatcheva, Q. Gao, C. Marchio, F. C. Geyer, K. Savage, S. Parry, et al. PPM1D Is a Potential Therapeutic Target in Ovarian Clear Cell Carcinomas Clin. Cancer Res., April 1, 2009; 15(7): 2269 - 2280. [Abstract] [Full Text] [PDF]
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 J Peltonen, J A Welsh, and K H Vahakangas Is there a role for PCR-SSCP among the methods for missense mutation detection of TP53 gene? Human and Experimental Toxicology, January 1, 2007; 26(1): 9 - 18. [Abstract] [PDF]
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 T. E. Rohan, S.-Q. Li, R. Hartwick, and R. A. Kandel p53 Alterations and Protein Accumulation in Benign Breast Tissue and Breast Cancer Risk: A Cohort Study. Cancer Epidemiol. Biomarkers Prev., July 1, 2006; 15(7): 1316 - 1323. [Abstract] [Full Text] [PDF]
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M. Tennis, S. Krishnan, M. Bonner, C. B. Ambrosone, J. E. Vena, K. Moysich, H. Swede, S. McCann, P. Hall, P. G. Shields, et al. p53 Mutation Analysis in Breast Tumors by a DNA Microarray Method Cancer Epidemiol. Biomarkers Prev., January 1, 2006; 15(1): 80 - 85. [Abstract] [Full Text] [PDF]
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 S. J. Oh, J. Ju, B. C. Kim, E. Ko, B. J. Hong, J.-G. Park, J. W. Park, and K. Y. Choi DNA microarrays on a dendron-modified surface improve significantly the detection of single nucleotide variations in the p53 gene Nucleic Acids Res., June 6, 2005; 33(10): e90 - e90. [Abstract] [Full Text] [PDF]
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Y. Hu, M. P. McDermott, and S. A. Ahrendt The p53 Codon 72 Proline Allele Is Associated with p53 Gene Mutations in Non-Small Cell Lung Cancer Clin. Cancer Res., April 1, 2005; 11(7): 2502 - 2509. [Abstract] [Full Text] [PDF]
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 A. C.P. Allen, F. A. Chiafari, R. Kandel, and T. Rohan Additional Data for Oligonucleotide Arrays of the p53 Gene in DNA from Formalin-Fixed, Paraffin-Embedded Tissue * Drs. Kandel and Rohan respond: Clin. Chem., December 1, 2004; 50(12): 2461 - 2462. [Full Text] [PDF]
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M. Maekawa, T. Nagaoka, T. Taniguchi, H. Higashi, H. Sugimura, K. Sugano, H. Yonekawa, T. Satoh, T. Horii, N. Shirai, et al. Three-Dimensional Microarray Compared with PCR-Single-Strand Conformation Polymorphism Analysis/DNA Sequencing for Mutation Analysis of K-ras Codons 12 and 13 Clin. Chem., August 1, 2004; 50(8): 1322 - 1327. [Abstract] [Full Text] [PDF]
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