High frequency of NPM1 gene mutations in acute myeloid leukemia with prominent nuclear invaginations ("cuplike" nuclei).

sive disappearance of the V379I BCR-ABL allele and its replacement by a novel BCR-ABL allele (Figure 1, lanes 4-6, band c). Direct sequencing further characterized a double mutation V379I and T315I (Figure S1, available at the Blood website; see the Supplemental Figure link at the top of the online letter). It is noteworthy that the 2 mutations were present on the same allele, since only 1 band was seen in DGGE after 10 months of treatment (Figure 1, lane 6) and the V379I and T315I mutations were observed on the same electropherogram without detection of the nonmutated sequence. As the BCR-ABL/ABL ratio increased to 35% after 10 months of dasatinib therapy and only hematopoietic cells carrying the 2 mutations were detected, the treatment was interrupted. Our observation confirms that targeted therapy of CML can lead to a clonal selection of BCR-ABL mutants. In our patient, IM therapy induced the selection of V379I mutants, then dasatinib selected among these IM-resistant cells those carrying the T315I mutation. These data highlight a problem inherent to targeted therapy that could directly induce a resistance phenomenon by means of selection of pre-existing mutants.6 The utility and feasibility of a sensitive and accurate mutation analysis before IM therapy has recently been studied, but even for the T315I mutant such an approach remains controversial.7 Moreover, whatever the methodology used, it seems improbable to detect any rare preexisting mutant stem cells, especially the quiescent ones. Komarova and Wodarz8 suggested that the combination of 3 targeting drugs could overcome the problem of resistance in CML. Current protocols allow only the use of tyrosine kinase inhibitors as single agents. Consequently, when a resistance is observed, mutation screening within the BCR-ABL kinase domain is mandatory. In conclusion, our study demonstrates a sequential emergence of ABL kinase domain mutations in a patient treated with anti–tyrosine kinase agents and highlights the importance of a mutation follow-up for therapeutic decisions.