scholarly journals Role of Pten in leukemia stem cells

Oncotarget ◽  
2010 ◽  
Vol 1 (2) ◽  
pp. 156-160 ◽  
Author(s):  
Cong Peng ◽  
Yaoyu Chen ◽  
Dongguang Li ◽  
Shaoguang Li
Keyword(s):  
Stem Cells ◽  
Leukemia Stem Cells

Inhibition Of CXCR2 As a Therapeutic Strategy In AML and MDS

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 484-484 ◽  
Author(s):  
Carolina Schinke ◽  
Orsolya Giricz ◽  
Shanisha A. K. Gordon ◽  
Laura Barreyro ◽  
Tushar D. Bhagat ◽  
...  
Keyword(s):  
Stem Cells ◽  
Leukemic Cell ◽  
Leukemia Stem Cells ◽  
P Value ◽  
Healthy Controls ◽  
Functional Studies ◽  
Cxcr2 Expression ◽  
Cd34 Cells

Abstract Acute Myeloid Leukemia (AML) and Myelodysplastic syndrome (MDS) arise from accumulation of multiple stepwise genetic and epigenetic changes in hematopoietic stem cells (HSC) and/or committed progenitors. A series of transforming events can initially give rise to pre-leukemia stem cells (pre-LSC) as well as fully transformed leukemia stem cells (LSC), both of which need to be targeted in strategies aimed at curing these diseases. We conducted parallel transcriptional analysis of multiple, highly fractionated stem and progenitor populations in individual patients of MDS and AML (N=16) and identified candidate genes that are consistently dysregulated at multiple immature stem and progenitor cell stages. Interleukin 8 (IL8), was one of the most consistently overexpressed genes in MDS/AML Hematolpoetic Stem Cells (HSCs) and progenitors when compared to healthy control HSCs and progenitors. IL8 is a pro-inflammatory chemokine, which is able to activate multiple intracellular signaling pathways after binding to its surface receptor CXCR2. Even though increased IL8-CXCR2 signaling has been shown to promote angiogenesis, metastasis and chemotherapy resistance in many solid tumors, its role in AML and MDS is not well elucidated. We further analyzed gene expression profiles of CD34+ cells from 183 MDS patients and found significant increased expression of CXCR2 in MDS when compared to healthy controls (FDR<0.1). Most importantly, analysis of The Cancer Genome Atlas (TCGA) AML (n=200) dataset showed that CXCR2 expression was predictive of significantly adverse prognosis (log rank P value=0.0182; median survival of 245 days in cxcr2 high vs 607 days in cxcr2 low) in patients, further pointing to a critical role of IL8-CXCR2 signaling in AML/MDS. Next, we studied the functional role of IL8 and CXCR2 in AML. A panel of leukemic cell lines (THP-1, U937, KG-1, MOLM13, HL-60, K532) were screened for CXCR2 expression and revealed significantly higher expression when compared to healthy CD34+ control cells. SB-332235, a specific inhibitor of CXCR2 was used for functional studies. CXCR2 inhibition led to significant, (p<0.05) reduction in proliferation in all 6 cell lines tested and an effect was seen as early as 24 hrs of exposure. CXCR2 inhibition was found to lead to G0/G1 cell cycle arrest and trigged apoptosis in THP-1 and U937 cells (p-value 0.004 and 0.02 respectively). Incubation of primary AML/MDS bone marrow samples with SB-332235 similarly lead to significantly reduced proliferation at 24hrs, when compared to healthy CD34+ cells. Selective, and highly significant inhibition of leukemic cell growth was also seen in colony assays from primary MDS/AML samples (mean leukemic colonies in AML/MDS= 73 vs 313 in controls, P < 0.001). Interestingly, inhibition of CXCR2 in primary AML marrow samples led to induction of apoptosis in immature CD34+/CD38- cells when compared to healthy controls. Lastly, xenografting studies with THP-1 leukemic cells revealed that CXCR2 inhibitor treatment led to decreased leukemic burden and organ infiltration when compared to placebo controls in vivo. In summary we have found significantly increased expression of IL8 and its receptor CXCR2 in sorted HSCs and progenitors from AML and MDS patients. High CXCR2 expression was a marker of adverse prognosis in a large cohort of AML patients. Most importantly, in vitro and in vivo functional studies showed that CXCR2 is a potential therapeutic target in AML/MDS and is able to selectively target immature, LSC-enriched cell fractions in AML. Disclosures: No relevant conflicts of interest to declare.

Mutant p53 Drives the Development of Pre-Leukemic Hematopoietic Stem Cells through Modulating Epigenetic Regulators

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 307-307
Author(s):  
Sarah C Nabinger ◽  
Michihiro Kobayashi ◽  
Rui Gao ◽  
Sisi Chen ◽  
Chonghua Yao ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Bone Marrow Cells ◽  
Leukemia Stem Cells ◽  
Mutant P53 ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Tp53 Mutations ◽  
Marrow Cells

Abstract AML is thought to arise from leukemia stem cells (LSCs); however, recent evidence suggests that the transforming events may initially give rise to pre-leukemic hematopoietic stem cells (pre-leukemic HSCs), preceding the formation of fully transformed LSCs. Pre-leukemic HSCs have been shown to contribute to normal blood development and harbor a selective growth advantage compared to normal HSCs. Pre-leukemic HSCs can acquire subsequent mutations, and once differentiation capacity is impaired, leukemia emerges. Recently, acquired somatic TP53 mutations, including p53R248W and p53R273H, were identified in healthy individuals as well as AML patients, suggesting that TP53 mutations may be early events in the pathogenesis of AML. We found that p53R248W HSCs showed a multi-lineage repopulation advantage over WT HSCs in transplantation experiments, demonstrating that mutant p53 confers a pre-leukemic phenotype in murine HSCs. Although TP53 mutations are limited in AML, TP53 mutations do co-exist with mutations of epigenetic regulator, ASXL-1, or receptor tyrosine kinase, FLT3, in AML. Mutations in Asxl-1 are present in ~10-30% of patients with myeloid malignancies and confer poor prognosis. Loss of Asxl-1 in the hematopoietic compartment leads to a myelodysplastic-like syndrome in mice and reduced stem cell self-renewal. Internal tandem duplications in Flt3 (Flt3-ITD) occur in ~30% of AML patients and are associated with adverse clinical outcome. Flt3-ITD-positive mice develop a myeloproliferative neoplasm (MPN) and HSCs expressing Flt3-ITD have decreased self-renewal capabilities. We hypothesize that mutant p53 drives the development of pre-leukemic HSCs with enhanced self-renewal capability, allowing clonal expansion and subsequent acquisition of Asxl-1 or Flt3 mutations leading to the formation of fully transformed leukemia stem cells. To define the role of mutant p53 in Asxl-1+/- HSCs, we generated p53R248W/+ Asxl-1+/- mice and performed in vitro serial replating assays as well as in vivo competitivebone marrow transplantation experiments. We found that p53R248W significantly enhanced the serial replating ability of Asxl-1-deficient bone marrow cells. Interestingly, while bone marrow from Asxl-1+/- mice had very poor engraftment compared to wild type bone marrow cells 16 weeks post-transplantation, the expression of p53R248W in Asxl-1+/- bone marrow rescued the defect. To examine the role of mutant p53 in Flt3-ITD-positive HSCs, we generated p53R248W/+ Flt3ITD/+ mice. We found that p53R248W enhanced the replating ability of Flt3ITD/+ bone marrow cells. Despite the fact that Flt3ITD/+ bone marrow cells displayed decreased repopulating ability compared to wild type cells 16 weeks post-transplant, expression of p53R248W in Flt3ITD/+ cells rescued the defect. We are monitoring leukemia development in primary and secondary transplant recipients as well as in de novo p53R248W/+ Asxl-1+/- and p53R248W/+ Flt3ITD/+ animals and predict that mutant p53 may cooperate with Asxl-1 deficiency or Flt3-ITD in the formation of LSCs to accelerate leukemia development in Asxl-1 deficient or Flt-ITD-positive neoplasms. Mechanistically, dysregulated epigenetic control underlies the pathogenesis of AML and we discovered that mutant p53 regulates epigenetic regulators, including Ezh1, Ezh2, Kdm2a, and Setd2, in HSCs. H3K27me3 is catalyzed by EZH1 or EZH2 of the Polycomb repressing complex 2 (PRC2). Both Ezh1 and Ezh2 are important for HSC self-renewal. SETD2 is a histone H3K36 methyltransferase and mutations in SETD2 have been identified in 6% of patients with AML. SETD2 deficiency resulted in a global loss of H3K36me3 and increased self-renewal capability of leukemia stem cells. We found that there were increased levels of H3K27me3 and decreased levels of H3K36me3 in p53R248W/+ HSCs compared to that of the WT HSCs. In ChIP experiments, we found that p53R248W, but not WT p53, was associated with the promoter region of Ezh2 in mouse myeloid progenitor cells, suggesting that p53R248W may directly activate Ezh2 expression in hematopoietic cells. Given that Asxl-1 has been shown to regulate H3K27me3 in HSCs, the synergy between mutant p53 and Asxl-1 deficiency on LSC self-renewal could be due to changes in histone modifications. Overall, we demonstrate that mutant p53 promotes the development of pre-leukemic HSCs by a novel mechanism involving dysregulation of the epigenetic pathways. Disclosures No relevant conflicts of interest to declare.

scholarly journals The telomere complex and the origin of the cancer stem cell

2021 ◽  
Vol 9 (1) ◽  
Author(s):  
A. Torres-Montaner
Keyword(s):  
Stem Cells ◽  
Malignant Transformation ◽  
Developmental Stages ◽  
Telomere Maintenance ◽  
Leukemia Stem Cells ◽  
Self Renewal ◽  
Telomere Attrition ◽  
Oncogenic Pathways ◽  
Tumor Protection

AbstractExquisite regulation of telomere length is essential for the preservation of the lifetime function and self-renewal of stem cells. However, multiple oncogenic pathways converge on induction of telomere attrition or telomerase overexpression and these events can by themselves trigger malignant transformation. Activation of NFκB, the outcome of telomere complex damage, is present in leukemia stem cells but absent in normal stem cells and can activate DOT1L which has been linked to MLL-fusion leukemias. Tumors that arise from cells of early and late developmental stages appear to follow two different oncogenic routes in which the role of telomere and telomerase signaling might be differentially involved. In contrast, direct malignant transformation of stem cells appears to be extremely rare. This suggests an inherent resistance of stem cells to cancer transformation which could be linked to a stem cell’specific mechanism of telomere maintenance. However, tumor protection of normal stem cells could also be conferred by cell extrinsic mechanisms.

scholarly journals Mir-125b Regulates the Self-Renewal of Acute Myeloid Leukemia Stem Cells through PTPN18 and GSK3

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 16-17
Author(s):  
Qiang Liu ◽  
Olga I. Gan ◽  
Gabriela Krivdova ◽  
Aaron Trotman-Grant ◽  
Stephanie M. Dobson ◽  
...  
Keyword(s):  
Stem Cells ◽  
Acute Myeloid Leukemia ◽  
Tyrosine Phosphorylation ◽  
Myeloid Leukemia ◽  
The Self ◽  
Lower Percentage ◽  
Leukemia Stem Cells ◽  
Self Renewal ◽  
Acute Myeloid

Acute myeloid leukemia (AML) is an aggressive hematologic malignancy with poor survival, especially in older patients. Despite high remission rates after chemotherapy, relapse and death are frequent due to persistence of leukemia stem cells (LSCs), which possess properties linked to therapy resistance. Thus, there is an urgent need for a deeper understanding of the unique properties of LSCs. MicroRNAs (miRNAs) are non-coding RNAs that decrease expression of their target mRNAs by post-translational silencing. miRNA profiling of human AML samples fractionated based on LSC activity revealed that miR-125b is expressed at significantly higher levels on cell fractions enriched in LSCs. To evaluate the role of miR-125b in LSCs, expression of miR-125b was enforced in a hierarchical AML model cell line (OCI-AML-8227). miR-125b overexpression (OE) resulted in a significantly lower percentage of CD14+CD15+ differentiated myeloblasts (Figure 1A) and enhanced clonogenic potential in vitro (Figure 1B). Xenotransplantation of four AML patient samples with miR-125b OE revealed a significant increase in the proportion of CD117+ cells, a marker of hematopoietic and leukemic progenitors (Figure 1C). Secondary transplantation of cells harvested from primary engrafted mice at limiting dilution demonstrated a marked increase in LSC frequency with miR-125b OE compared to controls for the two AML samples tested (Figure 1D). Together, these data strongly suggest that miR-125b enhances the self-renewal of LSCs. To investigate the mechanisms by which miR-125b enhances self-renewal, proteomic analysis of miR-125b-OE Ba/F3 cells as well as in silico target prediction were performed and identified PTPN18 as a top putative target for miR-125b. PTPN18 is a tyrosine phosphatase that has been reported to dephosphorylate auto-phosphorylated kinases such as Her2 and Abl to prevent their activation. To evaluate whether PTPN18 OE can rescue the effects miR-125b on LSCs, we carried out transduction of an AML patient sample with control, miR-125b OE, PTPN18 OE, or both miR-125b and PTPN18 OE vectors followed by xenotransplantation. Similar to previous findings, miR-125b OE alone significantly reduced the frequency of CD11b+CD15+ differentiated myeloblasts. Co-transduction of miR-125b/PTPN18 OE vectors resulted in generation of significantly more CD11b+CD15+ cells compared to miR-125b OE alone (Figure 1E), suggesting that suppression of PTPN18 contributes to miR-125b-mediated enhancement of LSC self-renewal. To identify putative phosphotyrosines that might be altered through the miR-125b-PTPN18 signalling axis, we performed immunoprecipitation of phosphotyrosines followed by mass spectrometry in miR-125b-OE Ba/F3 cells and identified increased GSK3 tyrosine phosphorylation as a top target. Additionally, miR-125b OE was confirmed to enhance GSK3 tyrosine phosphorylation, whereas PTPN18 OE reduced it (Figure 1F), together strongly suggesting that miR-125b could enhance tyrosine phosphorylation of GSK3 by silencing PTPN18. GSK3A and GSK3B (GSK3A/B) are paralogous genes that share a high degree of sequence homology and belong to the glycogen synthase kinase 3 (GSK3) family. Tyrosine phosphorylation activates the kinase activity of GSK3, whereas serine phosphorylation inactivates it. We recently identified GSK inhibitors as top candidates targeting LSCs in a stemness-based drug screen using OCI-AML-8227 cells (data not shown). Treatment of OCI-AML-8227 cells with two selective inhibitors of GSK3 selectively reduced the proportion of CD34+ cells while concomitantly increasing expression of myeloid markers CD14 and CD15 (Figure 1G). Overall, our results support an important functional role for PTPN18 and GSK3 in LSC function, and present a potential novel therapeutic target against LSCs. This study highlights the importance of understanding the role of miRNAs and may identify a new druggable vulnerability in LSCs that could lead to the development of new treatment options for AML patients. Figure 1 Disclosures Dick: Bristol-Myers Squibb/Celgene: Research Funding. Wang:Trilium Therapeutics: Patents & Royalties.

Targeting of MDS and AML Stem Cells Via Inhibition of STAT3 By Pyrimethamine

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 3602-3602 ◽  
Author(s):  
Aditi Shastri ◽  
Carolina Schinke ◽  
Asya Varshavsky Yanovsky ◽  
Tushar D. Bhagat ◽  
Orsolya Giricz ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Lines ◽  
Critical Role ◽  
Sh2 Domain ◽  
Leukemia Stem Cells ◽  
P Value ◽  
High Avidity ◽  
Healthy Controls ◽  
Hematopoietic Stem

Abstract Acute Myeloid Leukemia (AML) and Myelodysplastic syndrome (MDS) arise from accumulation of multiple stepwise genetic and epigenetic changes in hematopoietic stem cells (HSC) and/or committed progenitors. A series of transforming events can initially give rise to pre-leukemia stem cells (pre-LSC) as well as fully transformed leukemia stem cells (LSC), both of which need to be targeted in strategies aimed at curing these diseases. We conducted parallel transcriptional and epigenetic analysis of highly fractionated stem and progenitor populations in individual patients of MDS and identified STAT3 upregulation in MDS HSCs. qRTPCR in an independent set of sorted MDS/AML HSCs (Lineage-negative, CD34+, CD38-) confirmed the significant increase in STAT3 expression in 60% of cases when compared to healthy controls. We further analyzed gene expression profiles of CD34+ cells from 183 MDS patients and found significant increased expression of STAT3 in MDS when compared to healthy controls (FDR<0.1) and importantly, found that increased STAT3 expression was predictive of significantly adverse prognosis (log rank P value < 0.01, median survival of 2.6 years compared to 5.8 years for group with lower STAT3) in patients. This further points to a critical role of STAT3 signaling in AML/MDS pathogenesis and progression. Next, we studied the functional role of STAT3 by using Pyrimethamine as a clinically relevant inhibitor. Pyrimethamine is an FDA approved antifolate compound that was found to be a specific inhibitor of STAT3 activity in an initial screen of 1120 compounds. 3D modeling studies reveal that Pyrimethamine can occupy the SH2 domain of STAT3 protein with high avidity. It shows half-maximal activity (EC50) for STAT3 inhibition at approximately 1.5 μM, well within the plasma concentrations found with clinical use. We assessed the effect of Pyrimethamine on cellular proliferation of multiple leukemic cell lines (KG-1, KT-1 and CMK), and observed that these were significantly inhibited at 120 hours of exposure in a dose dependent fashion (t test, p value <0.004, Mean + SD of 3 experiments). Pyrimethamine was also able to induce significant apoptosis in AML cell lines at 72 hours at a 10 μM concentration. (P Value <0.05) In summary, we have found significant demethylation and increased expression of STAT3 in sorted HSCs from AML and MDS patients. High STAT3 expression is a marker of adverse prognosis in a large cohort of MDS patients. In vitro studies show that Pyrimethamine can inhibit STAT3 activation and is able to inhibit proliferation by inducing apoptosis in leukemic cells. A phase II study evaluating Pyrimethamine for the treatment of intermediate/high-risk MDS patients that have relapsed or are refractory to azanucleoside therapy has been initiated. Disclosures Brown: Sanofi, Onyx, Vertex, Novartis, Boehringer, GSK, Roche/Genentech, Emergent, Morphosys, Celgene, Janssen, Pharmacyclics, Gilead: Consultancy.

SETD2, a H3K36 Lysine Methyltransferase, Is Essential for Adult Normal Hematopoiesis and Leukemia Stem Cells

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1055-1055
Author(s):  
Yile Zhou ◽  
Yunzhu Dong ◽  
Jiachen Bu ◽  
Xiaomei Yan ◽  
Yoshihiro Hayashi ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Conflicts Of Interest ◽  
Conditional Knockout ◽  
The Other ◽  
Leukemia Stem Cells ◽  
Transcriptional Networks ◽  
Loss Of Function ◽  
Hematopoietic Stem

Abstract Hematopoietic stem cells (HSCs) are characterized by their capability for self-renewal and multi-potency. Hematopoiesis is dynamically controlled by the interplay between epigenetic and transcriptional networks. Dysregulation of these networks can lead to unfitness of hematopoiesis, cell transformation, and hematological diseases. The human SETD2 gene was originally isolated from HSCs and progenitors. SETD2 is a histone methyltransferase, which specifically catalyzes tri-methylation of histone 3 lysine 36 (H3K36me3). SETD2 functions as a tumor suppressor, as loss-of-function mutations have been identified in many cancers. However, the role of SETD2 in hematopoiesis has not been fully understood. To assess the function of Setd2 in hematopoiesis, we generated three Setd2 mouse alleles with Crispr/CAS9 technology; Setd2F2478/WT knock-in, Setd2Exon6-Δ/WT, and Setd2-Exon6flox/flox/Mx1-Cre conditional knockout alleles, as homozygous Setd2 mutation showed embryonic lethality. Setd2-F2478 point mutation, which is located in the SRI domain, can express SETD2 mutant protein but completely lose the interaction with RNA pol II. Setd2Exon6-Δ/WT allele results in a frame shift and nonsense mediated decay of Setd2 mRNA and protein. After induction of excision with pIpC injection, Setd2-exon6flox/flox/Mx1-Cre+ (Setd2Exon6-Δ/Δ) mice showed severe anemia, increased platelet count, and a reduction in bone marrow (BM) cellularity compared to wild-type (WT) mice, while Setd2F2478/WT and Setd2Exon6-Δ/WT mice did not show any obvious hematological changes. The Lin- Sca-1+ c-Kit+ (LSK) population in Setd2Exon6-Δ/Δ mice was 2.5-fold decreased compared to those in WT, while the LSK populations in Setd2F2478/WT and Setd2Exon6-Δ/WT mice were comparable with those in WT. Interestingly, all three of these Setd2 mutant alleles showed a higher frequency of Lin- Sca-1- c-Kit+ (LK) cells in the BM. In the LK populations, we found an increased CMP population in Setd2F2478/WT and Setd2Exon6-Δ/WT mice; of note, the CMP population in the Setd2Exon6-Δ/Δ mice had disappeared while the MEP population expanded with higher expression of CD16/32. Next, to assess the function of the HSPCs, we performed CFU assays and competitive bone marrow transplantations (CBMT). Consistent with our phenotypic findings, the number of colonies derived from Setd2F2478/WT and Setd2Exon6-Δ/WT BM cells was increased in the first two passages, while the number of colonies derived from Setd2Exon6-Δ/Δ mice was significantly decreased. In CBMT, we found that mice transplanted with Setd2Exon6-Δ/Δ BM cells showed anemia and an impaired BM reconstitution, compared to the control (p = 0.0002). On the other hand, the Setd2F2478/WT and Setd2Exon6-Δ/WT models showed comparable capabilities of BM reconstitution. Taken together, these results suggest that Setd2 has an essential role in the maintenance of adult hematopoiesis. SETD2 mutations (mainly one allele mutation) have been frequently identified in acute leukemia, especially in about 22% of MLL leukemia. To understand the role of SETD2 in leukemic stem cells, Setd2 mutant mice were bred with the Mll-AF9 knock-in mouse. The Mll-AF9/ Setd2F2478/WT and Mll-AF9/ Setd2Exon6-Δ/WT mice showed higher frequencies of LK and LSK populations compared to Mll-AF9 mice, indicating that Setd2 mutations may increase the stemness of leukemia stem cells (LSCs). The cells derived from Mll-AF9/ Setd2F2478/WT and Mll-AF9/ Setd2Exon6-Δ/WT mice resulted in a significantly higher yield of colonies and growth advantage in serial replating CFU assay compared to the cells derived from Mll-AF9 mice. After BMT of equal numbers of cells from Mll-Af9 or Mll-AF9/ Setd2F2478/WT mice into recipient mice, the Mll-AF9/ Setd2F2478/WTBMT mice developed leukemia with significantly shortened latencies compared with MLL-Af9 BMT mice. In conclusion, our data suggests that Setd2 plays an important role in maintaining normal HSPCs. Half the doses of Setd2 can still maintain the normal hematopoiesis while a total loss of Setd2 leads to a failure of hematopoiesis. In leukemia, heterozygous mutants of Setd2 can accelerate leukemogenesis by expanding LSCs. Whether the remaining WT allele is required for leukemia maintenance is unclear. Further reduction of Setd2 levels, or complete deletion of the other WT allele, may diminish SETD2-mutated leukemia. Such tumor vulnerability can be explored as a therapeutic strategy. Disclosures No relevant conflicts of interest to declare.

scholarly journals Myc-Miz-1 signaling promotes self-renewal of leukemia stem cells by repressing Cebpα and Cebpδ

Blood ◽  
2020 ◽  
Author(s):  
Lei Zhang ◽  
Jing Li ◽  
Hui Xu ◽  
Xianyu Shao ◽  
Li Fu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Leukemia Stem Cells ◽  
Mutant Form ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Reduced Frequency ◽  
Almost All

c-Myc (Myc hereafter) is found to be deregulated and/or amplified in most acute myeloid leukemias (AML). Almost all AML cells are dependent upon Myc for their proliferation and survival. Thus Myc has been proposed as a critical anti-AML target. Myc has Max-mediated trans-activational and Miz1-mediated trans-repressional activities. The role of Myc-Max-mediated trans-activation in the pathogenesis of AML has been well-studied; however the role of Myc-Miz1-mediated trans-repression in AML is still somewhat obscure. MycV394D is a mutant form of Myc which lacks trans-repressional activity due to a defect in its ability to interact with Miz1. We found that, compared to Myc, the oncogenic function of MycV394D is significantly impaired. The AML/myeloproliferative disorder which develops in mice receiving MycV394D-transduced hematopoietic stem/progenitor cells (HSPCs) is significantly delayed compared to mice receiving Myc-transduced HSPCs. Using a murine MLL-AF9 AML model, we found that AML cells expressing MycV394D (intrinsic Myc deleted) are partially differentiated and show reductions in both colony-forming ability in vitro and leukemogenic capacity in vivo. The reduced frequency of leukemia stem cells (LSCs) among MycV394D-AML cells and their reduced leukemogenic capacity during serial transplantation suggest that Myc-Miz1 interaction is required for the self-renewal of LSCs. In addition, we found that MycV394D-AML cells are more sensitive to chemotherapy than are Myc-AML cells. Mechanistically, we found that the Myc represses Miz1-mediated expression of Cebpα and Cebpδ, thus playing an important role in the pathogenesis of AML by maintaining the undifferentiated state and self-renewal capacity of LSCs.

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