Multiplex high-throughput gene mutation analysis in acute myeloid leukemia

Jennifer Dunlap, Carol Beadling, Andrea Warrick, Tanaya Neff, William Fleming, Marc Loriaux, Michael C. Heinrich, Tibor Kovacsovics, Katalin Kelemen, Nicky Leeborg, Ken Gatter, Rita Braziel, Richard Press, Christopher L. Corless, Guang Fan

Research output: Contribution to journalArticlepeer-review

16 Scopus citations


Classification of acute myeloid leukemia increasingly depends on genetic analysis. However, the number of known mutations in acute myeloid leukemia is expanding rapidly. Therefore, we tested a high-throughput screening method for acute myeloid leukemia mutation analysis using a multiplex mass spectrometry-based approach. To our knowledge, this is the first reported application of this approach to genotype leukemias in a clinical setting. One hundred seven acute myeloid leukemia cases were screened for mutations using a panel that covers 344 point mutations across 31 genes known to be associated with leukemia. The analysis was performed by multiplex polymerase chain reaction for mutations in genes of interest followed by primer extension reactions. Products were analyzed on a Sequenom MassARRAY system (San Diego, CA). The multiplex panel yielded mutations in 58% of acute myeloid leukemia cases with normal cytogenetics and 21% of cases with abnormal cytogenetics. Cytogenetics and routine polymerase chain reaction-based screening of NPM1, CEBPA, FLT3-ITD, and KIT was also performed on a subset of cases. When combined with the results of these standard polymerase chain reaction-based tests, the mutation frequency reached 78% in cases with normal cytogenetics. Of these, 42% harbored multiple mutations primarily involving NPM1 with NRAS, KRAS, CEBPA, PTPN11, IDH1, or FLT3. In contrast, cases with abnormal cytogenetics rarely harbored more than 1 mutation (1.5%), suggesting different underlying biology. This study demonstrates the feasibility and utility of broad-based mutation profiling of acute myeloid leukemia in a clinical setting. This approach will be helpful in defining prognostic subgroups of acute myeloid leukemia and contribute to the selection of patients for enrollment into trials with novel inhibitors.

Original languageEnglish (US)
Pages (from-to)2167-2176
Number of pages10
JournalHuman Pathology
Issue number12
StatePublished - Dec 2012


  • Acute myeloid leukemia
  • Gene mutation
  • Genetics
  • Molecular

ASJC Scopus subject areas

  • Pathology and Forensic Medicine


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