Extensive analysis of the T315I substitution and detection of additional ABL mutations in progenitors and primitive stem cell compartment in a patient with tyrosine kinase inhibitor-resistant chronic myeloid leukemia

Abstract

Tyrosine kinase inhibitors (TKIs) have revolutionized the treatment of chronic myeloid leukemia (CML). However, resistance is occasionally observed, mainly due to mutations within the BCR-ABL kinase domain. The T315I substitution confers complete resistance to all TKIs commonly used in clinical practice. To date, the hierarchical level of stem cells in which this mutation initially appears has not been studied. The aim of this study was to evaluate the behavior of T315I mutated cells and to study the presence of potential additional mutations in progenitors and stem cells from a patient with CML. A comprehensive analysis of BCR-ABL(315I) mRNA expressing cells in mononuclear cells, in CD34+ cell populations, and in their primitive long-term culture-initiating cell (LTC-IC) derived progenitors was performed. We show that the T315I substitution arises in primitive Ph1 stem cells without altering their myeloid and erythroid terminal differentiation potential. Leukemic cells expressing a T315I mutated BCR-ABL display a progressive decline in LTC-IC assays as described for non-mutated CML cells at diagnosis. Finally, in our experiments, additional non-315 ABL mutations were identified in hematopoietic stem cell colonies. This observation is suggestive of genetic instability affecting CML progenitors.

BCR-ABL expression in leukemic progenitors and primitive stem cells of patients with chronic myeloid leukemia

“…The analysis by Chomel and colleagues on hundreds of single residual CML stem and progenitor cell clones, derived from precious marrow samples of yearlong-followed CML patients deserves respect-also for the elaborate translational approach. 1 In keeping with our referenced finding, 2 Chomel et al demonstrate that persisting CML stem cells (in vitro referred to as long-term culture initiating cells, LTC-IC), but also committed precursor cells express substantially less BCR-ABL mRNA than their imatinib-resistant counterparts. These results can be interpreted as follows: high-level BCR-ABL expression is incompatible with persistence under imatinib, whereas low BCR-ABL levels contribute to intrinsic BCR-ABL kinase inhibitor resistance.…”

“…1 For example, oncogene-initiated signal flux must exceed a critical threshold to trigger cell intrinsic tumor suppressive responses via activation of Arf/p53. 2,3 Hence, lowlevel oncogenic BCR-ABL may not suffice to induce oncogene addiction in the first place. As a consequence, even potent BCR-ABL inhibition by imatinib at the stem cell level 4 would not generate a genetic pressure toward mutating BCR-ABL in persistent residual CML cells, because the resolution of this pressure (eg, by kinase mutations) would not result in a growth/survival advantage.…”

Getting to the root of the stem cell in mutated chronic myeloid leukemia

“…The point mutations and genetic instability seen in the patient analyzed by Chomel et al [7] are well described, and several pathways likely contribute to this finding [4,11,12]. Stoklosa et al demonstrated abnormal mismatch repair (MMR) and MMRdependent apoptosis by exposing BCR-ABL-positive cell lines, CD34þ CML cell lines, and CD34þ cells from normal bone marrow to O 6 -methyl-N'-nitro-Nnitrosoguanidine (MNNG) [6].…”

“…In this issue of Leukemia and Lymphoma, Chomel and colleagues [7] present data analyzing when the T315I mutation appears in stem cell differentiation and the interaction of three distinct cell populations: the BCR-ABL T315I -containing cells, the BCR-ABL wt -containing cells, and the non-BCR-ABLcontaining cell populations in vitro. Specifically, they used blood and bone marrow samples from a patient with an 8-year history of chronic phase CML, treated with interferon/cytarabine, imatinib, and dasatinib, with the eventual development of BCR-ABL T315I and TKI resistance.…”

Chronic Myeloid Leukemia Stem Cells: Lessons from a Lead Paradigm

“…They do not display any efficacy however, against leukemic cells harbouring T315I mutation (8). Importantly, ABLkinase mutations including T315 can occur in primitive hematopoietic stem cells suggesting that genetic instability inherent to BCR-ABL expressing cells are operational in these primitive cells (9).…”