Multiplex Detection of Mutations in Myeloproliferative Disorder and Leukemia Pati

DESCRIPTION (provided by applicant): The overall objective for this project is to develop a diagnostic assay for rapid detection of major mutations in patients with acute myeloid leukemia (AML), non-BCR/ABL chronic myelogenous leukemia (CML) and myeloproliferative disorders (MPDs). We propose to develop a single tube multiplex assay which uses high fidelity PCR for amplification of target mutations and hybridization of Luminex bead-tagged probes for detection of specific sequences. We have successfully developed highly multiplexed DNA- and RNA-based assays using this technology. The detection of point mutations in a background of wild-type transcripts will provide substantial benefits for diagnostic and prognostic testing and clinical management of these diseases. According to the American Cancer Society, 34,800 new cases of all types of leukemia were reported in the US in 2005. Leukemia is the number one cause of deaths from cancer in children and young adults under age 20. Although myeloid malignancies have been traditionally classified by their morphological and cytogenetic features, mutations that are undetectable at a chromosomal level have been shown to be correlated with distinct prognosis in leukemia and MPDs. As the molecular pathogenesis for these diseases is becoming evident, and mutation-specific inhibitors become available, classification will need to include molecular identification. It is our goal to develop a comprehensive assay system which 1) rapidly detects aberrant sequences for resolving ambiguous diagnosis of AML, non-BCR/ABL CML and MPDs, and 2) is predictive of therapeutic outcomes and thus useful in selecting between different treatment options. The availability of this assay for leukemia and MPD mutations should greatly improve the clinical decisions necessary for effective treatment of leukemia and MPD patients while improving clinical laboratory workflow and reducing costs. The Specific Aims are: 1) Develop a rapid single-tube assay for the simultaneous detection of major mutations in patients with AML and CML/MPDs. 2) Develop comprehensive controls for the clinical evaluation of the assay. In Phase II, we will evaluate the diagnostic value of the assay against a large number of clinical samples in a diagnostic laboratory setting. We will develop software for analyzing the assay results and to assist in clinical interpretation of the results. The final outcome is to launch mutation-specific Analyte Specific Reagents (primers, probes) and apply for FDA approval as an In Vitro Diagnostic Assay. The availability of this assay for leukemia and MPD mutations should greatly improve the clinical decisions necessary for effective treatment of leukemia and MPD patients while improving clinical laboratory workflow and reducing costs. The overall goal of this proposal is to develop an assay to detect mutations that are important for determining treatment of myeloproliferative disorders and leukemia. This liquid bead array format of this assay will provide efficiency and cost benefits to diagnostic laboratories and provide crucial information to physicians and patients for use in making treatment decisions.