Multiplexed assays for detection of mutations in PIK3CA

Ruth E. Board, Nicola J. Thelwell, Paul F. Ravetto, Stephen Little, Malcolm Ranson, Caroline Dive, Andrew Hughes, David Whitcombe

Research output: Contribution to journalArticlepeer-review


BACKGROUND: Mutations in the PIK3CA gene (phosphoinositide-3-kinase, catalytic, alpha polypeptide) have recently been described in a number of cancers, and their detection is currently limited because of the low sensitivity of conventional sequencing techniques. METHODS: We combined Amplification Refractory Mutation System (ARMS™; AstraZeneca) allele-specific PCR and Scorpions™ (DxS) to develop assays for tumor-borne PIK3CA mutations and used real-time PCR to develop high-throughput multiplexed assays for the most commonly reported PIK3CA mutants (H1047L, H1047R, E542K, E545K). RESULTS: These assays were more sensitive than sequencing and could detect 5 copies of mutant DNA in proportions as low as 0.1% of the total DNA. We assayed DNA extracted from human tumors and detected PIK3CA mutation frequencies of 10.2% in colorectal cancer, 38.7% in breast cancer, 1.9% in lung cancer, and 2.9% in melanoma. In contrast, sequencing detected only 53% of the mutations detected by our assay. CONCLUSIONS: Multiplexed assays, which can easily be applied to clinical samples, have been developed for the detection of PIK3CA mutations.
Original languageEnglish
Pages (from-to)757-760
Number of pages3
JournalClinical Chemistry
Issue number4
Publication statusPublished - 1 Apr 2008


  • genetics: 1-Phosphatidylinositol 3-Kinase
  • genetics: Breast Neoplasms
  • genetics: Colorectal Neoplasms
  • Female
  • Humans
  • genetics: Lung Neoplasms
  • genetics: Melanoma
  • Mutation
  • methods: Polymerase Chain Reaction
  • Sensitivity and Specificity
  • genetics: Tumor Markers, Biological


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