title: A new rapid and sensitive assay for detecting the T315I BCR-ABL kinase domain mutation in chronic myeloid leukaemia.
abstract: A significant minority of chronic myeloid leukaemia patients eventually develop resistance to imatinib, often as a result of point mutations within the BCR-ABL kinase domain. Second-line tyrosine kinase inhibitors (TKIs) are effective against mutations that confer imatinib resistance; however, the T315I BCR-ABL mutant has proved resistant to all available TKIs. An assay facilitating early identification of BCR-ABL(T315I) would therefore aid in identifying high-risk patients who may benefit from alternative therapy. This report describes the development of a sensitive T315I mutation detection methodology based on real-time PCR with self-probing fluorescent primers. The technique demonstrated complete concordance with direct sequencing, correctly identifying 34 T315I-positive samples from a total of 61 samples screened. In a limiting dilution assay, the mutated clone was detectable to a level of 1% of total cells. The data show that Scorpions PCR enables rapid screening for BCR-ABL(T315I) in chronic myeloid leukaemia patients and is appropriate for use in a clinical setting.
authors.affiliation: Department of Haematology, Imperial College London, London, UK.
authors.forename: J S
authors.initials: JS
authors.lastname: Khorashad
authors.forename: N
authors.initials: N
authors.lastname: Thelwell
authors.forename: D
authors.initials: D
authors.lastname: Milojkovic
authors.forename: D
authors.initials: D
authors.lastname: Marin
authors.forename: J A
authors.initials: JA
authors.lastname: Watson
authors.forename: J M
authors.initials: JM
authors.lastname: Goldman
authors.forename: J F
authors.initials: JF
authors.lastname: Apperley
authors.forename: L
authors.initials: L
authors.lastname: Foroni
authors.forename: A G
authors.initials: AG
authors.lastname: Reid
mesh_terms: D018076:DNA, Complementary / Q000235:genetics; D004273:DNA, Neoplasm / Q000235:genetics; D016044:Fusion Proteins, bcr-abl / Q000235:genetics*; D006801:Humans; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000175:diagnosis* / Q000235:genetics; D017354:Point Mutation; D016133:Polymerase Chain Reaction / Q000379:methods; D012189:Retrospective Studies; D012680:Sensitivity and Specificity
chemical_list: D018076:DNA, Complementary; D004273:DNA, Neoplasm; D016044:Fusion Proteins, bcr-abl