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Molecular Biology and Genetics |
Department of Pathology [W-H. W., M. F. P.], The Norris Comprehensive Cancer Center [L. B., J. S-H., M. F. P.], and Department of Preventive Medicine [L. B., J. S-H.], University of Southern California School of Medicine, Los Angeles, California 90033; Affymetrix, Inc., Santa Clara, California 95051 [J. L.]; and Gene Logic, Inc., Gaithersburg, Maryland 20878 [Y. B-B.]
As the rate of gene discovery accelerates, more efficient methods are needed to analyze genes in human tissues. To assess the efficiency, sensitivity, and specificity of different methods, alterations of TP53 were independently evaluated in 108 ovarian tumors by conventional DNA sequence analysis and oligonucleotide microarray (p53 GeneChip). All mutations identified by oligonucleotide microarray and all disagreements with conventional gel-based DNA sequence analysis were confirmed by re-analysis with manual and automated dideoxy DNA sequencing. A total of 77 ovarian cancers were identified as having TP53 mutations by one of the two approaches, 71 by microarray and 63 by gel-based DNA sequence analysis. The same mutation was identified in 57 ovarian cancers, and the same wild type TP53 sequence was observed in 31 ovarian cancers by both methods, for a concordance rate of 81%. Among the mutation analyses discordant by these methods for TP53 sequence were 14 cases identified as mutated by microarray but not by conventional DNA sequence analysis and 6 cases identified as mutated by conventional DNA sequence analysis but not by microarray. Overall, the oligonucleotide microarray demonstrated a 94% accuracy rate, a 92% sensitivity, and an 100% specificity. Conventional DNA sequence analysis demonstrated an 87% accuracy rate, 82% sensitivity, and a 100% specificity. Patients with TP53 mutations had significantly shorter overall survival than those with no mutation (P = 0.02). Women with mutations in loop2, loop3, or the loop-sheet-helix domain had shorter survival than women with other mutations or women with no mutations (P = 0.01). Although further refinement would be helpful to improve the detection of certain types of TP53 alterations, oligonucleotide microarrays were shown to be a powerful and effective tool for TP53 mutation detection.
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