Konference: 2006 48th ASH Annual Meeting - účast ČR
Kategorie: Maligní lymfomy a leukémie
Téma: Publikace ve sborníku
Číslo abstraktu: 4906
Autoři: MUDr. Soňa Peková, Ph.D.; Rudolf Hoffmann; MUDr. Pavel Jindra, Ph.D.; MUDr. Ludmila Nováková; doc. MUDr. Tomáš Kozák, Ph.D., MBA; MUDr. Věra Vozobulová; MUDr. Miroslava Schűtzová; Jozef Balasz; Lucie Tomsikova; Doc.MUDr. Miroslav Průcha, Ph.D.
The incidence of V617 in PV is high, reaching 96% of cases; in ET or IMF, the mutation is less frequent. Nevertheless, the percentage of JAK2 V617F-positive patients differs greatly among studies, depending on the detection technique used. Sequencing analysis, as one of the options, can underrate the number of positive cases, as its sensitivity in authentic samples with varying proportion of mutant granulocytes is unsatisfactory. The nucleotide context of the V617F site is complicated (direct repeat TGTG/TT), and thus prone to sequencing artifacts.
For specific and sensitive detection of V617F mutation in JAK2 gene, we have developed a novel allelic discrimination assay, based on the usage of Real-Time PCR technology and LNA-modified fluorescently labeled probes (Locked Nucleic Acids). This technology is distinguished by 100% discrimination efficiency, and sensitivity detection rate reaching 10% of mutant granulocytes in an authentic sample. LNA-modified probes are extremely powerful tool for precise genotyping, since they are short (12-mers), and due to the interspersed LNA nucleotides, melting temperature (Tm) of the probe/target hybrid is high. Hence, the LNA-based genotyping technology combines the desired features of high sensitivity and absolute specificity.
Herein, we present an analysis of JAK2 V617F detection in our group of 157 consecutive patients with the diagnosis of MPD or suspected MPD. Using our novel LNA-based Real-Time PCR genotyping technique, we were able to identify varying proportions of the mutant JAK2 allele in 87 cases (55.4%); in the remaining 70 patients the mutation was not detected.
The LNA-based Real-Time PCR technique is extremely precise and efficient in identifying even subtle proportion of JAK2 V617F mutant granulocytes, which in cases of diagnostic uncertainties on the origin of the polycythemia might be of great importance.
Datum přednesení příspěvku: 9. 12. 2006