Abstract
BackgroundPhosphatidylinositol-4,5-bisphosphate 3-kinase, catalytic subunit alpha, PIK3CA, is one of the most frequently mutated genes in breast cancer, and the mutation status of PIK3CA has clinical relevance related to response to therapy.The aim of our study was to investigate the mutation status of PIK3CA gene and to evaluate the concordance between NGS and SGS for the most important hotspot regions in exon 9 and 20, to investigate additional hotspots outside of these exons using NGS, and to correlate the PIK3CA mutation status with the clinicopathological characteristics of the cohort.MethodsIn the current study, next-generation sequencing (NGS) and Sanger Sequencing (SGS) was used for the mutational analysis of PIK3CA in 186 breast carcinomas.ResultsAltogether, 64 tumors had PIK3CA mutations, 55 of these mutations occurred in exons 9 and 20. Out of these 55 mutations, 52 could also be detected by Sanger sequencing resulting in a concordance of 98.4 % between the two sequencing methods. The three mutations missed by SGS had low variant frequencies below 10 %. Additionally, 4.8 % of the tumors had mutations in exons 1, 4, 7, and 13 of PIK3CA that were not detected by SGS. PIK3CA mutation status was significantly associated with hormone receptor-positivity, HER2-negativity, tumor grade, and lymph node involvement. However, there was no statistically significant association between the PIK3CA mutation status and overall survival.ConclusionsBased on our study, NGS is recommended as follows: 1) for correctly assessing the mutation status of PIK3CA in breast cancer, especially for cases with low tumor content, 2) for the detection of subclonal mutations, and 3) for simultaneous mutation detection in multiple exons.Electronic supplementary materialThe online version of this article (doi:10.1186/s12907-015-0020-6) contains supplementary material, which is available to authorized users.
Highlights
Phosphatidylinositol-4,5-bisphosphate 3-kinase, catalytic subunit alpha, PIK3CA, is one of the most frequently mutated genes in breast cancer, and the mutation status of PIK3CA has clinical relevance related to response to therapy
Prevalence of different types of PIK3CA mutations using next-generation sequencing (NGS) Using NGS to sequence the PIK3CA gene in each of the 186 tissue samples identified a total of 64 tumors with exon mutations (34.4 %), which agreed with the 36 % PIK3CA mutation rate in breast cancer (BC) reported by the Cancer Genome Atlas (TCGA) [18, 19]
We found that PIK3CA was most frequently mutated in cases that were HR+ and HER2, which agrees with previously published data [51, 52]
Summary
Phosphatidylinositol-4,5-bisphosphate 3-kinase, catalytic subunit alpha, PIK3CA, is one of the most frequently mutated genes in breast cancer, and the mutation status of PIK3CA has clinical relevance related to response to therapy. Sanger sequencing (SGS) has been the gold standard for detecting DNA mutations. Somatic cancer mutations can be difficult to detect using SGS without performing microdissections because tumors are heterogeneous and often mixed with normal tissue. Targeted NGS, which involves the targeted enrichment of a set of DNA regions, is used for the parallel sequencing of amplicons derived from multiplex polymerase chain reaction (PCR) or other amplicon-based. Targeted NGS is more cost efficient than SGS [10]. This high-throughput technology is currently used with several platforms, including the Genome Analyzer/HiSeq/MiSeq (Illumina Solexa), the SOLiD System (Thermo Fisher Scientific), the Ion PGM/Ion Proton (Thermo Fisher Scientific), and the HeliScope Sequencer (Helicos BioSciences) [11, 12]
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