Transcription factor IKZF1 (IKAROS) acts as a critical regulator of lymphoid differentiation and is frequently deleted or mutated in B-cell precursor acute lymphoblastic leukemia.
gene defects are ...associated with inferior treatment outcome in both childhood and adult B-cell precursor acute lymphoblastic leukemia and occur in more than 70% of
-positive and
-like cases of acute lymphoblastic leukemia. Over the past few years, much has been learned about the tumor suppressive function of IKZF1 during leukemia development and the molecular pathways that relate to its impact on treatment outcome. In this review, we provide a concise overview on the role of IKZF1 during normal lymphopoiesis and the pathways that contribute to leukemia pathogenesis as a consequence of altered IKZF1 function. Furthermore, we discuss different mechanisms by which
alterations impose therapy resistance on leukemic cells, including enhanced cell adhesion and modulation of glucocorticoid response.
Deletions and mutations affecting lymphoid transcription factor IKZF1 (IKAROS) are associated with an increased relapse risk and poor outcome in B-cell precursor acute lymphoblastic leukemia. ...However, additional genetic events may either enhance or negate the effects of
deletions on prognosis. In a large discovery cohort of 533 childhood B-cell precursor acute lymphoblastic leukemia patients, we observed that single-copy losses of
were significantly enriched in
-deleted B-cell precursor acute lymphoblastic leukemia (
=0.007). While
deletions alone had no impact on prognosis, the combined presence of
and
deletions was associated with a significantly lower 5-year event-free survival (
=0.0003) and a higher 5-year cumulative incidence of relapse (
=0.005), when compared with
-deleted cases without
aberrations. In contrast, other copy number losses commonly observed in B-cell precursor acute lymphoblastic leukemia, such as
or
, did not affect the outcome of
-deleted acute lymphoblastic leukemia patients. To establish whether the combined loss of IKZF1 and BTG1 function cooperate in leukemogenesis,
-deficient mice were crossed onto an
heterozygous background. We observed that loss of
increased the tumor incidence of
mice in a dose-dependent manner. Moreover, murine B cells deficient for
and
displayed increased resistance to glucocorticoids, but not to other chemotherapeutic drugs. Together, our results identify
as a tumor suppressor in leukemia that, when deleted, strongly enhances the risk of relapse in
-deleted B-cell precursor acute lymphoblastic leukemia, and augments the glucocorticoid resistance phenotype mediated by the loss of IKZF1 function.
Although long-term survival in pediatric acute lymphoblastic leukemia (ALL) currently exceeds 90%, some subgroups, defined by specific genomic aberrations, respond poorly to treatment. We previously ...reported that leukemias harboring deletions or mutations affecting the B-cell transcription factor
IKZF1
exhibit a tumor cell intrinsic resistance to glucocorticoids (GCs), one of the cornerstone drugs used in the treatment of ALL. Here, we identified increased activation of both AKT and ERK signaling pathways as drivers of GC resistance in
IKZF1-
deficient leukemic cells. Indeed, combined pharmacological inhibition of AKT and ERK signaling effectively reversed GC resistance in
IKZF1
-deficient leukemias. As inhibitors for both pathways are under clinical investigation, their combined use may enhance the efficacy of prednisolone-based therapy in this high-risk patient group.
In acute myeloid leukaemia, long-term survival is poor as most patients relapse despite achieving remission. Historically, the failure of therapy has been thought to be due to mutations that produce ...drug resistance, possibly arising as a consequence of the mutagenic properties of chemotherapy drugs. However, other lines of evidence have pointed to the pre-existence of drug-resistant cells. For example, deep sequencing of paired diagnosis and relapse acute myeloid leukaemia samples has provided direct evidence that relapse in some cases is generated from minor genetic subclones present at diagnosis that survive chemotherapy, suggesting that resistant cells are generated by evolutionary processes before treatment and are selected by therapy. Nevertheless, the mechanisms of therapy failure and capacity for leukaemic regeneration remain obscure, as sequence analysis alone does not provide insight into the cell types that are fated to drive relapse. Although leukaemia stem cells have been linked to relapse owing to their dormancy and self-renewal properties, and leukaemia stem cell gene expression signatures are highly predictive of therapy failure, experimental studies have been primarily correlative and a role for leukaemia stem cells in acute myeloid leukaemia relapse has not been directly proved. Here, through combined genetic and functional analysis of purified subpopulations and xenografts from paired diagnosis/relapse samples, we identify therapy-resistant cells already present at diagnosis and two major patterns of relapse. In some cases, relapse originated from rare leukaemia stem cells with a haematopoietic stem/progenitor cell phenotype, while in other instances relapse developed from larger subclones of immunophenotypically committed leukaemia cells that retained strong stemness transcriptional signatures. The identification of distinct patterns of relapse should lead to improved methods for disease management and monitoring in acute myeloid leukaemia. Moreover, the shared functional and transcriptional stemness properties that underlie both cellular origins of relapse emphasize the importance of developing new therapeutic approaches that target stemness to prevent relapse.
In acute myeloid leukaemia (AML), the cell of origin, nature and biological consequences of initiating lesions, and order of subsequent mutations remain poorly understood, as AML is typically ...diagnosed without observation of a pre-leukaemic phase. Here, highly purified haematopoietic stem cells (HSCs), progenitor and mature cell fractions from the blood of AML patients were found to contain recurrent DNMT3A mutations (DNMT3A(mut)) at high allele frequency, but without coincident NPM1 mutations (NPM1c) present in AML blasts. DNMT3A(mut)-bearing HSCs showed a multilineage repopulation advantage over non-mutated HSCs in xenografts, establishing their identity as pre-leukaemic HSCs. Pre-leukaemic HSCs were found in remission samples, indicating that they survive chemotherapy. Therefore DNMT3A(mut) arises early in AML evolution, probably in HSCs, leading to a clonally expanded pool of pre-leukaemic HSCs from which AML evolves. Our findings provide a paradigm for the detection and treatment of pre-leukaemic clones before the acquisition of additional genetic lesions engenders greater therapeutic resistance.
To investigate miRNA function in human acute myeloid leukemia (AML) stem cells (LSC), we generated a prognostic LSC-associated miRNA signature derived from functionally validated subpopulations of ...AML samples. For one signature miRNA, miR-126, high bioactivity aggregated all in vivo patient sample LSC activity into a single sorted population, tightly coupling miR-126 expression to LSC function. Through functional studies, miR-126 was found to restrain cell cycle progression, prevent differentiation, and increase self-renewal of primary LSC in vivo. Compared with prior results showing miR-126 regulation of normal hematopoietic stem cell (HSC) cycling, these functional stem effects are opposite between LSC and HSC. Combined transcriptome and proteome analysis demonstrates that miR-126 targets the PI3K/AKT/MTOR signaling pathway, preserving LSC quiescence and promoting chemotherapy resistance.
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•Clinical outcome in AML correlates with LSC-associated miRNA expression•miR-126 targets multiple components of the PI3K/AKT/MTOR signaling pathway•miR-126 promotes chemotherapy resistance by preserving LSC in a quiescent state•miR-126 governs opposing self-renewal outcomes in normal and malignant stem cells
Lechman et al. show that miR-126 targets the PI3K/AKT/MTOR signaling pathway to preserve quiescence, increase self-renewal, and promote chemotherapy resistance of acute myeloid leukemia stem cells (LSC). Reducing the miR-126 level impairs LSC maintenance in contrast to expanding normal hematopoietic stem cells.
Deletions and mutations affecting transcription factor IKZF1 are associated with increased relapse risk and poor outcome in B cell precursor acute lymphoblastic leukemia (BCP-ALL). However, ...additional genetic events may either enhance or negate the effects of IKZF1 on prognosis. We observed that deletions of the gene encoding the transcriptional coregulator BTG1 frequently co-occured with loss of IKZF1 function, suggesting a synergistic role for these events during leukemia development or progression. Targeted deletion of Btg1 predisposed both Btg1+/- and Btg1-/- mice to T cell malignancies, similar to what has been observed in Ikzf1 heterozygous knockout animals. Hence, while somatic single single-copy losses of either BTG1 and IKZF1 in the patient are predominantly found in BCP-ALL, targeted deletion of these genes in the mouse gives rise to T cell malignancies.
To establish whether loss of BTG1 function affected the tumor suppressive role of IKZF1, the Btg1 knockout allele was crossed onto mice heterozygous for a loss-of-function Ikzf1 allele. Leukemia penetrance in these compound mice increased in a Btg1 dose-dependent manner. These leukemias were characterized by clonal TCRb rearrangement and aggressive infiltration into secondary organs, indicating synergistic roles for these tumor suppressors during mouse leukemia development.
To investigate the effects of combined IKZF1/BTG1 loss in human BCP-ALL, we examined a large pediatric cohort of BCP-ALL cases, and found that the combined presence of BTG1 and IKZF1 deletions was associated with a markedly higher incidence of relapse, relative to IKZF1-deleted cases without BTG1 aberrations. Similar to BTG1 copy number losses, deletions in EBF1, PAX5, RB1 and CDKN2A/B appeared to be selectively enriched in IKZF1 deleted ALL. However, in contrast to BTG1, none of these other copy number alterations affected relapse incidence or outcome in this patient group. In conclusion, our data demonstrate synergy between the tumor suppressors BTG1 and IKZF1 during mouse leukemia development while the combined (single copy) loss of these two tumor suppressors identifies a patient group with an extremely poor outcome.
Event free survival in a cohort of 514 children newly diagnosed with BCP-ALL, divided into four categories based on IKZF1 and BTG1 deletion status.
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Pieters:Eusa Pharma: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees.
Treatment of relapsed acute myeloid leukemia (AML) results in lower complete remission rates compared to treatment of AML at diagnosis. Immunophenotypic changes in leukemic blasts are common from ...diagnosis to relapse (Baer et al. Blood. 2001), suggesting that the underlying biology of AML changes with disease progression. A better understanding of the biologic properties of AML cells at different stages of disease will facilitate the development of biomarker tools and more effective therapies. We therefore studied paired diagnostic/relapse samples obtained from AML patients. Cells from 11 paired samples were transplanted into immune deficient mice (NOD.SCID IL2Rg null) over a range of cell doses. In 10 of 11 patients, a leukemic graft could be generated after transplantation of lower cell doses from the relapse sample compared to the paired diagnostic sample. By limiting dilution analysis, leukemia stem cell (LSC) frequency was higher in relapse samples (1 in 5.8×102 to 1 in 2.4×106, median 1 in 2.0×103) compared to diagnostic samples (1 in 5.0×103 to 1 in 6.1×106, median 1 in 5.5×104); the fold increase in LSC frequency ranged from 2.2 to 745 (median 8.6). Multiparameter flow cytometric analysis carried out on 13 paired diagnostic/relapse samples demonstrated an increase in 2 known stem cell markers, CD34 and CD117, from diagnosis to relapse: CD34 was gained or increased at relapse in 7/13 (54%) of paired samples, while CD117 was gained or increased at relapse in 9/13 (69%) of paired samples. We plan to take a comprehensive approach to examine the surface marker expression of paired diagnostic/relapse samples by a high throughput flow cytometric screen (HTS) of both bulk and LSC-containing AML populations to identify markers that are altered at relapse. As a first step, we have performed HTS of 373 surface markers on 10 AML patient samples, including a relapse sample from a diagnostic/relapse pair. A significant proportion of markers (155/373, 42%) were expressed on less that 5% of cells in all 10 AML samples analyzed. We will therefore focus on the remaining markers in our HTS analysis of paired samples. Surface markers that are differentially expressed from diagnosis to relapse will be further characterized in order to gain insight into disease progression and identify potential therapeutic targets.
No relevant conflicts of interest to declare.
In acute myeloid leukemia (AML), the cell of origin, nature and biological consequences of initiating lesions and order of subsequent mutations remain poorly understood, as AML is typically diagnosed ...without observation of a pre-leukemic phase. Here, highly purified hematopoietic stem cells (HSC), progenitor and mature cell fractions from the blood of AML patients were found to contain recurrent
DNMT3a
mutations (
DNMT3a
mut
) at high allele frequency, but without coincident
NPM1
mutations (
NPM1c
) present in AML blasts.
DNMT3a
mut
-bearing HSC exhibited multilineage repopulation advantage over non-mutated HSC in xenografts, establishing their identity as pre-leukemic-HSC (preL-HSC). preL-HSC were found in remission samples indicating that they survive chemotherapy. Thus
DNMT3a
mut
arises early in AML evolution, likely in HSC, leading to a clonally expanded pool of preL-HSC from which AML evolves. Our findings provide a paradigm for the detection and treatment of pre-leukemic clones before the acquisition of additional genetic lesions engenders greater therapeutic resistance.
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