In our previous study we showed that constant levels of BCR-ABL gene expression ≥ 0.1% (IS) in minimally 3 consecutive samples were associated with the mutation development and disease progression in small proportion of patients with chronic myeloid leukemia (CML) in complete cytogenetic remission after the imatinib treatment (Polakova KM et al. Exp Hem 2010;38:20). Here we aimed to reveal whereas the BCR-ABL transcript stable levels below the 0.1% but higher than 0.01% for minimally 9 months may be attributed to the resistance development.
From 212 patients with major molecular response (MMoR) after the imatinib (n=169) and dasatinib (n= 43) treatment in standard dose, 10 of them (9 pts on imatinib and 1 pt on dasatinib) reached a plateau of BCR-ABL gene expression at levels between 0.01 and 0.1% (IS) by median 26 months (range 5–46) and remained unchanged for a median of next 22 months (range 9–41). High resolution melt curve analysis (HRM; Polakova KM et al. Leuk Res 2008;32:1236) was applied to analyze mutations in BCR-ABL kinase domain because of higher sensitivity in comparison to direct sequencing. The last 2–3 consecutive samples of peripheral blood total leukocytes of all 10 patients were analyzed. Good quality sequencing is dependent on the amount of total BCR-ABL transcript. The minimum for optimal direct sequencing performance is 0.1% (Polakova KM et al. Exp Hem 2010;38:20). Direct sequencing was in this study however possible applied in some low level BCR-ABL samples.
The HRM positivity was found within the region spanning the P-loop in 2 patients and within the region of the activation loop in 1 patient. Two patients on imatinib lost MMoR with the significant BCR-ABL rise over the 0.1% and sequencing confirmed presence of M244V (100%) and M351T (100%) in 2 and 7 consecutive samples, respectively. In patient with M244V mutation, sequencing revealed the development of other mutation F359V during the next 10 months of follow-up with BCR-ABL transcript level rise from 0.6% to 2% in 3 subsequent samples, i.e. 9 months. The F359V mutant clone seemed to have proliferation advantage over the M244V. The patient on dasatinib has not shown any rise of BCR-ABL transcript up to now, i.e. 5 months after positive HRM. However in two out of the last three consecutive HRM positive samples P-loop mutation E255K (100%) was detected by direct sequencing.
Based on our results it seems that BCR-ABL transcript level fluctuation at values within the range of 0.01% to 0.1% may be associated with mutation development and the MMoR lost in a number of CML patients after the tyrosine kinase inhibitor treatment.
Supported by MZOUHKT2005, MSM0021622430, research grant from Bristol-Myers Squibb and CELL the Czech Leukemia Study Group for Life.
Machova Polakova:Bristol-Myers Squibb: Research Funding.
Asterisk with author names denotes non-ASH members.