Targeted detection of mutations associated with imatinib-resistance. Strategies to increase sensitivity and specificity
AuthorJohansen, Linda Eilen
Background: The main cause to chronic myeloid leukemia (CML) is a translocation between chromosome 9 and 22, resulting in the Philadelphia chromosome, containing a BCR-ABL fusion-gene that encodes a constitutively active tyrosine kinase. Imatinib, a selective inhibitor of the tyrosine kinase, is the first-line treatment for CML-patients. However, drug-resistance is an increasing problem, and the most important reasons for this are point mutations within the BCR-ABL kinase domain. More than 40 such mutations are described, and because the mutations can have a great impact on clinical decision-making, early detection is important. With new tyrosine kinases developing, the resistance problem seems to be diminished, as some of the new drugs are less prone to resistance. Only the important T3151-mutation seems to remain a problem, which is why this particular mutation should be in focus for developing a more sensitive detection method. Materials and methods: Plasmids and patient-samples containing wild-type and mutated BCR-ABL sequences were used in an effort to design a sensitive and specific, biased assay for detection of the T3151-mutation in BCR-ABL alleles. By comparing a single-step versus a nested approach, the amplification of long versus short templates, TaqMan versus SYBR Green Master Mixes, and ASO versus LNA and ARMS-primers, a patient-test for detection of the T3151-mutation was developed. Results: A nested PCR approach using SYBR Green Master Mix and BCR-ABL-specific primer pair with a LNA-nucleotide incorporated in the mutation-specific reverse primer provided the greatest improvements in sensitivity and specificity for detection of the T3151-mutation in samples from CML-patients.
PublisherUniversitetet i Tromsø
University of Tromsø
The following license file are associated with this item:
Showing items related by title, author, creator and subject.
Prognostic Impacts of Angiopoietins in NSCLC Tumor Cells and Stroma : VEGF-A Impact Is Strongly Associated with Ang-2 Andersen, Sigve; Dønnem, Tom; Al-Shibli, Khalid Ibrahim; Al-Saad, Samer; Stenvold, Helge; Busund, Lill-Tove; Bremnes, Roy M. (Journal article; Tidsskriftartikkel; Peer reviewed, 2011)Angiopoietins and their receptor Tie-2 are, in concert with VEGF-A, key mediators in angiogenesis. This study evaluates the prognostic impact of all known human angiopoietins (Ang-1, Ang-2 and Ang-4) and their receptor Tie-2, as well as their relation to the prognostic expression of VEGF-A. 335 unselected stage I-IIIA NSCLC-patients were included and tissue samples of respective tumor cells and ...
Humant papillomavirus : en litteraturstudie om HPV, dets relasjon til cancer og tiltak mot videre spredning av virus Gabrielsen, Endre (Master thesis; Mastergradsoppgave, 2012-06-01)I 1983 oppdaget zur Hausen sammenhengen mellom Humant Papillomavirus (HPV) og livmorhalskreft. På denne tiden visste man ikke at det var HPV som var årsaken til at Helaceller kunne leve in vitro. Ny forskning relaterer HPV til en rekke andre cancertyper. En stor andel anal-, oropharyngeal-, penis-, vaginal-, og vulvacancer skyldes HPV. Det er også påvist HPV i tumorvev fra øsofagus, larynx, lunge, ...
Hoem, Gry (Master thesis; Mastergradsoppgave, 2012-06)Fragile X syndrome (FXS) is caused by an expanded CGG repeat (>200 repeats) in the 5’ un-translated portion of the fragile mental retardation 1 gene (FMR1) leading to deficiency or absence of the FMR1 protein (FMRP). FMRP is an RNA carrier protein that controls the translation of a number of other genes that regulate synaptic development and plasticity. Autism occurs in approximately 30% of FXS ...