Abstract
Tyrosine kinase inhibitors (TKIs) have profoundly changed the natural history of chronic myeloid leukemia (CML). However, acquired resistance to imatinib, dasatinib or nilotinib (1(st) and 2(nd) generation TKIs), due in part to BCR-ABL1 kinase mutations, has been largely described. These drugs are ineffective on the T315I gatekeeper substitution, which remains sensitive to 3(rd) generation TKI ponatinib. It has recently been suggested that the hematopoietic niche could protect leukemic cells from targeted therapy. In order to investigate the role of a stromal niche in mutation-related resistance, we developed a niche-based cell mutagenesis assay. For this purpose, ENU (N-ethyl-N-nitrosourea)-exposed UT-7 cells expressing non-mutated or T315I-mutated BCR-ABL1 were cultured with or without murine MS-5 stromal cells and in the presence of imatinib, dasatinib, nilotinib, or ponatinib. In the assays relative to 1(st) and 2(nd) generation TKIs, which were performed on non-mutated BCR-ABL1 cells, our data highlighted the increasing efficacy of the latter, but did not reveal any substantial effect of the niche. In ponatinib assays performed on both non-mutated and T315I-mutated BCR-ABL1 cells, an increased number of resistant clones were observed in the presence of MS-5. Present data suggested that T315I mutants need either compound mutations (e.g. E255K/T315I) or a stromal niche to escape from ponatinib. Using array-comparative genomic hybridization experiments, we found an increased number of variations (involving some recurrent chromosome regions) in clones cultured on MS-5 feeder. Overall, our study suggests that the hematopoietic niche could play a crucial role in conferring resistance to ponatinib, by providing survival signals and favoring genetic instability.
Highlights
Chronic myeloid leukemia (CML) is a paradigm of leukemogenesis initiated by the appearance of the Philadelphia (Ph1) chromosome in a primitive hematopoietic stem cell (HSC) [1]
Mutagenesis assay in UT-7 cells and Tyrosine kinase inhibitors (TKIs) selection
Our assay was based on human UT-7 cell lines, mutagenized with ENU (N-ethyl-N-nitrosourea) and cultured with or without murine MS-5 stromal cells in the presence of imatinib, dasatinib, nilotinib, or ponatinib (Supplementary Fig. S1)
Summary
Chronic myeloid leukemia (CML) is a paradigm of leukemogenesis initiated by the appearance of the Philadelphia (Ph1) chromosome in a primitive hematopoietic stem cell (HSC) [1]. The ability to target this tyrosine kinase protein by the use of small inhibitors is challenging since BCR-ABL1 activates a plethora of signaling pathways [3]. In this context, imatinib, which showed selective inhibitory activity with regard to BCRABL1, was the first TKI (tyrosine kinase inhibitor) developed and tested successfully in patients to become the standard front-line treatment of chronic phase CML [4,5]. Up to 20-30% of patients develop resistance towards imatinib This phenomenon can be either oncogene-dependent (e.g. BCR-ABL1 amplification or mutations), or –independent (e.g. activation of SRC kinase families) [6]
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