Profiling of Metabolic Differences between Hematopoietic Stem Cells and Acute/Chronic Myeloid Leukemia

Although many studies have been conducted on leukemia, only a few have analyzed the metabolomic profiles of various leukemic cells. In this study, the metabolomes of THP-1, U937, KG-1 (acute myelogenous leukemia, AML), K562 (chronic myelogenous leukemia, CML), and cord blood-derived CD34-positive hematopoietic stem cells (HSC) were analyzed using gas chromatography-mass spectrometry, and specific metabolic alterations were found using multivariate statistical analysis. Compared to HSCs, leukemia cell metabolomes were found to have significant alterations, among which three were related to amino acids, three to sugars, and five to fatty acids. Compared to CML, four metabolomes were observed specifically in AML. Given that overall more metabolites are present in leukemia cells than in HSCs, we observed that the activation of glycolysis and oxidative phosphorylation (OXPHOS) metabolism facilitated the incidence of leukemia and the proliferation of leukemic cells. Analysis of metabolome profiles specifically present in HSCs and leukemia cells greatly increases our basic understanding of cellular metabolic characteristics, which is valuable fundamental knowledge for developing novel anticancer drugs targeting leukemia metabolism.

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Immunoglobulin Superfamily Member 8 Is Indispensable for Myeloid Leukemia Via Wnt/β-Catenin Signaling Pathway

Blood ◽

10.1182/blood-2020-136045 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 23-24

Author(s):

Koji Jimbo ◽

Takaaki Konuma ◽

Takahiro Ito ◽

Yaeko Nakajima-Takagi ◽

Atsushi Iwama ◽

...

Keyword(s):

Stem Cells ◽

Myeloid Leukemia ◽

Research Funding ◽

Leukemia Cell ◽

Myelogenous Leukemia ◽

Leukemic Cells ◽

Leukemia Cells ◽

Immunoglobulin Superfamily ◽

Hematopoietic Stem

Immunoglobulin superfamily member 8 (IGSF8, also known as EWI-2, PGRL, and CD316), is a cell surface protein containing 4 immunoglobulin domains. IGSF8 directly binds to the tetraspanin molecules, CD9 and CD81, and modulates cell adhesion, migration, and growth. Previous studies demonstrated that IGSF8 was associated with prognosis and metastasis in several solid tumors. However, the role of IGSF8 in normal hematopoiesis and myeloid leukemia is still unclear. First, we examined the expression levels of Igsf8 in various hematopoietic fraction of wild-type murine bone marrow cells, and found that Igsf8 is expressed in all hematopoietic lineages. To investigate hematopoietic functions of Igsf8, we generated hematopoietic cells specific Igsf8 deleted mice (Igsf8fl/fl; Vav-Cre) and tamoxifen induced Igsf8 deleted mice (Igsf8fl/fl; Rosa26-CreERT). Igsf8fl/fl, Vav-Cre (denoted as Igsf8-/-) mice represented normal maturation. Deletion of Igsf8 did not significantly affect adult hematopoiesis in peripheral blood and bone marrow. Igsf8-/- long-term hematopoietic stem cells (LT-HSCs: CD34- Flk2- c-Kit+ Sca-1+ Lineage- cells) reduced colony forming ability in vitro, and serial competitive transplantation assay showed comparable donor chimerism by 3 months, but led to decrease Igsf8-/- donor chimerism at 4 months and those after second transplantation in vivo. These results suggest that Igsf8 does not affect the adult hematopoiesis, but it can affect their proliferative and reconstitutive capacity of HSCs. To investigate the effects of Igsf8 on myeloid leukemia, we generated MLL-AF9 and NRASG12V-driven acute myelogenous leukemia (AML), or BCR-ABL and NUP98-HOXA9-driven blast crisis of chronic myelogenous leukemia (CML-BC) mice models. Igsf8-/- led to a dramatic reduction in the number of leukemic colonies formed in vitro (Figure 1A). Igsf8-/- leukemia mice showed significantly longer survival in vivo (Figure 1B). This effect was also observed by eliminating Igsf8 expression after leukemia establishment using conditionally deletion. Igsf8-/- AML cells showed decreased S phase fraction. Igsf8-/- leukemia stem cells (LSCs: c-Kit+ Lineage- cells) triggered an increment of the apoptosis, which contribute to significantly lower proportion of LSCs in spleen of Igsf8-/- leukemic mice. Given that Igsf8-/- did not affect homing ability of leukemia cells, these results indicate that Igsf8 is required for propagation of myeloid leukemia and maintenance of LSC. To understand the Igsf8-mediated regulation of myeloid leukemia, we conducted RNA sequencing analysis of LT-HSCs, and LSCs of AML and CML-BC. Gene set enrichment analysis exhibited increase apoptosis related genes and decrease Wnt/β-catenin related genes in Igsf8-/- leukemic cells, but not in LT-HSCs (Figure 1C). Increment of pro-apoptosis genes, and decrement of anti-apoptosis genes and Wnt/β-catenin target genes in Igsf8-/- AML stem cells were validated in quantitative polymerase chain reaction analysis. Further, expression levels of β-catenin protein in Igsf8-/- leukemic cells were significantly lower compared to Igsf8+/+ leukemic cells, but not in normal hematopoietic stem and progenitor cells (Figure 1D). These results suggest that Igsf8 might be critical for myeloid leukemia maintenance via Wnt/β-catenin signaling pathway. Then, we investigated the effects of IGSF8 on human myeloid leukemia. We confirmed IGSF8 expression in several human myeloid leukemia cell line and primary patient-derived leukemia cells. Knockdown of IGSF8 by small hairpin RNA in myeloid leukemia cell lines (THP-1, MV4-11, SKM-1, and K562) and primary patient-derived AML cells exhibited reduced numbers of colony forming cells in vitro. Knockdown of IGSF8 also caused decrease expression of β-CATENIN in AML cell lines. These results indicate that IGSF8 is also required for propagation of human myeloid leukemia cells. Taken together, our present study reveals that Igsf8 is indispensable for myeloid leukemia, but not adult hematopoiesis, suggesting that IGSF8 inhibition should be considered for targeting myeloid leukemia. Disclosures Jimbo: Japan Society for the Promotion of Science: Research Funding. Konuma:SGH Foundation: Research Funding; The Japanese Society of Hematology: Research Funding; Institute for Frontier Life and Medical Sciences, Kyoto University: Research Funding. Ito:Institute for Frontier Life and Medical Sciences, Kyoto University: Research Funding.

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Remodeling of Bone Marrow Niches and Roles of Exosomes in Leukemia

International Journal of Molecular Sciences ◽

10.3390/ijms22041881 ◽

2021 ◽

Vol 22 (4) ◽

pp. 1881

Author(s):

Takanori Yamaguchi ◽

Eiji Kawamoto ◽

Arong Gaowa ◽

Eun Jeong Park ◽

Motomu Shimaoka

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Hematopoietic Stem Cells ◽

Chemotherapy Resistance ◽

Leukemia Cell ◽

Cell Communication ◽

Leukemia Cells ◽

Hematopoietic Stem ◽

Functional Changes ◽

Bone Marrow Niches

Leukemia is a hematological malignancy that originates from hematopoietic stem cells in the bone marrow. Significant progress has made in understanding its pathogensis and in establishing chemotherapy and hematopoietic stem cell transplantation therapy (HSCT). However, while the successive development of new therapies, such as molecular-targeted therapy and immunotherapy, have resulted in remarkable advances, the fact remains that some patients still cannot be saved, and resistance to treatment and relapse are still problems that need to be solved in leukemia patients. The bone marrow (BM) niche is a microenvironment that includes hematopoietic stem cells and their supporting cells. Leukemia cells interact with bone marrow niches and modulate them, not only inducing molecular and functional changes but also switching to niches favored by leukemia cells. The latter are closely associated with leukemia progression, suppression of normal hematopoiesis, and chemotherapy resistance, which is precisely the area of ongoing study. Exosomes play an important role in cell-to-cell communication, not only with cells in close proximity but also with those more distant due to the nature of exosomal circulation via body fluids. In leukemia, exosomes play important roles in leukemogenesis, disease progression, and organ invasion, and their usefulness in the diagnosis and treatment of leukemia has recently been reported. The interaction between leukemia cell-derived exosomes and the BM microenvironment has received particular attention. Their interaction is believed to play a very important role; in addition to their diagnostic value, exosomes could serve as a marker for monitoring treatment efficacy and as an aid in overcoming drug resistance, among the many problems in leukemia patients that have yet to be overcome. In this paper, we will review bone marrow niches in leukemia, findings on leukemia-derived exosomes, and exosome-induced changes in bone marrow niches.

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Pioglitazone inhibits the growth of human leukemia cell lines and primary leukemia cells while sparing normal hematopoietic stem cells

International Journal of Oncology ◽

10.3892/ijo.29.2.437 ◽

2006 ◽

Cited By ~ 3

Author(s):

Minoru Saiki ◽

Yoshihiro Hatta ◽

Tetsuo Yamazaki ◽

Takeyoshi Itoh ◽

Yuko Enomoto ◽

...

Keyword(s):

Stem Cells ◽

Hematopoietic Stem Cells ◽

Cell Lines ◽

Leukemia Cell ◽

Leukemia Cells ◽

Human Leukemia ◽

Hematopoietic Stem ◽

Human Leukemia Cell ◽

Leukemia Cell Lines ◽

Primary Leukemia

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Indirubin-3'-Monoxime Induces Apoptosis and Autophagy In Acute Lymphoblastic Leukemia Cells and Chronic Myelogenous Leukemia Cells, Exhibits Limited Cytotoxicity In CD34+ Hematopoietic Stem Cells, Lymphocytes and Granulocytes

Blood ◽

10.1182/blood.v116.21.3984.3984 ◽

2010 ◽

Vol 116 (21) ◽

pp. 3984-3984

Author(s):

Ming-Yang Lee ◽

Yi-Wen Liu ◽

Ming-Ho Chen ◽

Jing-Jing Chuang

Keyword(s):

Stem Cells ◽

Cell Death ◽

Acute Lymphoblastic Leukemia ◽

Hematopoietic Stem Cells ◽

Cell Survival ◽

Cytotoxic Effect ◽

Lymphoblastic Leukemia ◽

Myelogenous Leukemia ◽

Leukemia Cells ◽

Hematopoietic Stem

Abstract Abstract 3984 The rates of treatment-related mortality (TRM) and relapse are unacceptably high in adults undergoing antileukemia treatments for acute lymphoblastic leukemia (ALL). So far has no better therapy with low side effects to improve long-term survival in these patients. Indirubin, a Chinese translational anti-chronic myelogenous leukemia (CML) agent, is able to induce cellular apoptosis. However, until now the functional action of IO on ALL remains still unknown. Therefore, here we investigated and compared the cytotoxic efficacy and action of indirubin-3'-monoxime (IO) in JM1 (ALL cell) and K562 (CML cell). ALL and CML cells were treated with a series of IO dose for 24 and 48h, and cell survival was determined by WST-1 assay. ALL and CML cells were shown to be similar susceptible to IO cytotoxicity. In order to clear in which way of cell death induced by IO, we performed analyses for apoptosis, necrosis and autophagy, respectively. After IO treatment, both ALL and CML cells were arrested in the G2/M cell cycle phase. In addition, an increase of sub-G1 proportion was caused. We found also increasing of caspase-3 activation and formation of cleaved PARP in a dose dependent manner. These were associated with the form of apoptosis. However, the caspase inhibitor Z-VAD-FMK only could partially prevent cell death in ALL and CML cells. When further analyzing the necrotic phenomenon through measuring LDH release, the result clearly showed that LDH release was not remarkable after cell treatment with high dose of IO. Besides, we observed surprisingly in Western blot the increasing expression of microtubule-associated protein light chain 3-II (LC3-II), which generally correlates to formation of autophagosomes. Because better antileukemic drug should not induce toxicity in normal blood cells as much as possible, so the cytotoxic effect of IO in CD34+ hematopoietic stem cells, lymphocytes and granulocytes was analyzed. Excitingly, results showed that IO could not affect cell viability of granulocytes, and IO cytotoxicity in lymphocytes was only marginal. If CD34+ hematopoietic stem cells were treated with IO, the rate of cell survival and their ability of differentiation were almost identical in contrast with non-treated control. Apparently, these data indicated that IO possesses the capability to induce apoptosis and autophagy in both CML and ALL cells. The most important is that the IO hardly influences the cell survival and the differentiation of CD34+ hematopoietic stem cells, the cytotoxic effect in granulocytes and lymphocytes is only limited. In conclusion, IO can be considered as a potential agent for clinical anti- ALL treatment. Disclosures: No relevant conflicts of interest to declare.

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USP15 Deubiquitinase Safeguards Hematopoiesis and Genome Integrity in Hematopoietic Stem Cells and Leukemia Cells

Cell Reports ◽

10.1016/j.celrep.2020.108533 ◽

2020 ◽

Vol 33 (13) ◽

pp. 108533

Author(s):

Paul van den Berk ◽

Cesare Lancini ◽

Carlos Company ◽

Michela Serresi ◽

Maria Pilar Sanchez-Bailon ◽

...

Keyword(s):

Stem Cells ◽

Hematopoietic Stem Cells ◽

Leukemia Cells ◽

Genome Integrity ◽

Hematopoietic Stem

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Aurora-A kinase: a novel target of cellular immunotherapy for leukemia

Blood ◽

10.1182/blood-2008-06-164889 ◽

2009 ◽

Vol 113 (1) ◽

pp. 66-74 ◽

Cited By ~ 52

Author(s):

Toshiki Ochi ◽

Hiroshi Fujiwara ◽

Koichiro Suemori ◽

Taichi Azuma ◽

Yoshihiro Yakushijin ◽

...

Keyword(s):

Cell Lines ◽

Leukemia Cell ◽

Myelogenous Leukemia ◽

Leukemia Cells ◽

Cellular Immunotherapy ◽

Aurora A Kinase ◽

Aurora A ◽

Hematopoietic Stem ◽

Specific Ctls ◽

Leukemia Cell Lines

Abstract Aurora-A kinase (Aur-A) is a member of the serine/threonine kinase family that regulates the cell division process, and has recently been implicated in tumorigenesis. In this study, we identified an antigenic 9–amino-acid epitope (Aur-A207-215: YLILEYAPL) derived from Aur-A capable of generating leukemia-reactive cytotoxic T lymphocytes (CTLs) in the context of HLA-A*0201. The synthetic peptide of this epitope appeared to be capable of binding to HLA-A*2402 as well as HLA-A*0201 molecules. Leukemia cell lines and freshly isolated leukemia cells, particularly chronic myelogenous leukemia (CML) cells, appeared to express Aur-A abundantly. Aur-A–specific CTLs were able to lyse human leukemia cell lines and freshly isolated leukemia cells, but not normal cells, in an HLA-A*0201–restricted manner. Importantly, Aur-A–specific CTLs were able to lyse CD34+ CML progenitor cells but did not show any cytotoxicity against normal CD34+ hematopoietic stem cells. The tetramer assay revealed that the Aur-A207-215 epitope–specific CTL precursors are present in peripheral blood of HLA-A*0201–positive and HLA-A*2402–positive patients with leukemia, but not in healthy individuals. Our results indicate that cellular immunotherapy targeting Aur-A is a promising strategy for treatment of leukemia.

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Evidence for malignant transformation in acute myeloid leukemia at the level of early hematopoietic stem cells by cytogenetic analysis of CD34+ subpopulations

Blood ◽

10.1182/blood.v86.8.2906.bloodjournal8682906 ◽

1995 ◽

Vol 86 (8) ◽

pp. 2906-2912 ◽

Cited By ~ 2

Author(s):

D Haase ◽

M Feuring-Buske ◽

S Konemann ◽

C Fonatsch ◽

C Troff ◽

...

Keyword(s):

Stem Cells ◽

Acute Myeloid Leukemia ◽

Hematopoietic Stem Cells ◽

Malignant Transformation ◽

Cell Sorting ◽

Myeloid Leukemia ◽

De Novo ◽

Leukemic Cells ◽

Hematopoietic Stem ◽

Acute Myeloid

Acute myeloid leukemia (AML) is a heterogenous disease according to morphology, immunophenotype, and genetics. The retained capacity of differentiation is the basis for the phenotypic classification of the bulk population of leukemic blasts and the identification of distinct subpopulations. Within the hierarchy of hematopoietic development and differentiation it is still unknown at which stage the malignant transformation occurs. It was our aim to analyze the potential involvement of cells with the immunophenotype of pluripotent stem cells in the leukemic process by the use of cytogenetic and cell sorting techniques. Cytogenetic analyses of bone marrow aspirates were performed in 13 patients with AML (11 de novo and 2 secondary) and showed karyotype abnormalities in 10 cases [2q+, +4, 6p, t(6:9), 7, +8 in 1 patient each and inv(16) in 4 patients each]. Aliquots of the samples were fractionated by fluorescence-activated cell sorting of CD34+ cells. Two subpopulations, CD34+/CD38-(early hematopoietic stem cells) and CD34+/CD38+ (more mature progenitor cells), were screened for karyotype aberations as a marker for leukemic cells. Clonal abnormalities and evaluable metaphases were found in 8 highly purified CD34+/CD38-populations and in 9 of the CD34+/CD38-specimens, respectively. In the majority of cases (CD34+/CD38-, 6 of 8 informative samples; CD34+/CD38+, 5 of 9 informative samples), the highly purified CD34+ specimens also contained cytogenetically normal cells. Secondary, progression-associated chromosomal changes (+8, 12) were identified in the CD34+/CD38-cells of 2 patients. We conclude that clonal karyotypic abnormalities are frequently found in the stem cell-like (CD34+/CD38-) and more mature (CD34+/CD38+) populations of patients with AML, irrespective of the phenotype of the bulk population of leukemic blasts and of the primary or secondary character of the leukemia. Our data suggest that, in AML, malignant transformation as well as disease progression may occur at the level of CD34+/CD38-cells with multilineage potential.

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A Novel, Myeloid Transcription Factor, C/EBPε, Is Upregulated During Granulocytic, But Not Monocytic, Differentiation

Blood ◽

10.1182/blood.v90.7.2591.2591_2591_2600 ◽

1997 ◽

Vol 90 (7) ◽

pp. 2591-2600 ◽

Cited By ~ 9

Author(s):

Roberta Morosetti ◽

Dorothy J. Park ◽

Alexey M. Chumakov ◽

Isabelle Grillier ◽

Masaaki Shiohara ◽

...

Keyword(s):

Stem Cells ◽

Transcription Factor ◽

Hematopoietic Stem Cells ◽

Cell Line ◽

Cell Lines ◽

Myeloid Leukemia ◽

Leukemia Cell ◽

Rt Pcr ◽

Hematopoietic Stem ◽

Nb4 Cells

Human C/EBPε is a newly cloned CCAAT/enhancer-binding transcription factor. Initial studies indicated it may be an important regulator of human myelopoiesis. To elucidate the range of expression of C/EBPε, we used reverse transcription-polymerase chain reaction (RT-PCR) analysis and examined its expression in 28 hematopoietic and 14 nonhematopoietic cell lines, 16 fresh myeloid leukemia samples, and normal human hematopoietic stem cells and their mature progeny. Prominent expression of C/EBPε mRNA occurred in the late myeloblastic and promyelocytic cell lines (NB4, HL60, GFD8), the myelomonoblastic cell lines (U937 and THP-1), the early myeloblast cell lines (ML1, KCL22, MDS92), and the T-cell lymphoblastic leukemia cell lines CEM and HSB-2. For the acute promyelocytic leukemia cell line NB4, C/EBPε was the only C/EBP family member that was easily detected by RT-PCR. No C/EBPε mRNA was found in erythroid, megakaryocyte, basophil, B lymphoid, or nonhematopoietic cell lines. Most acute myeloid leukemia samples (11 of 12) from patients expressed C/EBPε. Northern blot and RT-PCR analyses showed that C/EBPε mRNA decreased when the HL60 and KG-1 myeloblast cell lines were induced to differentiate toward macrophages. Similarly, Western blot analysis showed that expression of C/EBPε protein was either unchanged or decreased slightly as the promyelocytic cell line NB4 differentiated down the macrophage-like pathway after treatment with a potent vitamin D3 analog (KH1060). In contrast, C/EBPε protein levels increased dramatically as NB4 cells were induced to differentiate down the granulocytic pathway after exposure to 9-cis retinoic acid. Furthermore, very early, normal hematopoietic stem cells (CD34+/CD38−), purified from humans had very weak expression of C/EBPε mRNA, but levels increased as these cells differentiated towards granulocytes. Likewise, purified granulocytes appeared to express higher levels of C/EBPε mRNA than purified macrophages. Addition of phosphothiolated antisense, but not sense oligonucleotides to C/EBPε, decreased clonal growth of HL-60 and NB4 cells by about 50% compared with control cultures. Taken together, our results indicate that expression of C/EBPε is restricted to hematopoietic tissues, especially myeloid cells as they differentiate towards granulocytes and inhibition of its expression in HL-60 and NB4 myeloblasts and promyelocytes decreased their proliferative capacity. Therefore, this transcriptional factor may play an important role in the process of normal myeloid development.

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Differential expression of a novel C-terminally truncated splice form of SMAD5 in hematopoietic stem cells and leukemia

Blood ◽

10.1182/blood.v95.12.3945 ◽

2000 ◽

Vol 95 (12) ◽

pp. 3945-3950 ◽

Cited By ~ 15

Author(s):

Yunfang Jiang ◽

Hong Liang ◽

Wei Guo ◽

Lazar V. Kottickal ◽

Lalitha Nagarajan

Keyword(s):

Stem Cells ◽

Hematopoietic Stem Cells ◽

Transforming Growth Factor ◽

Receptor Activation ◽

Myelogenous Leukemia ◽

Blood Leukocytes ◽

Hematopoietic Stem ◽

Acid Protein ◽

Acute Myelogenous ◽

Inactivating Mutations

Abstract SMADs are evolutionarily conserved transducers of the differentiation and growth arrest signals from the transforming growth factor/BMP (TGF/BMP) family of ligands. Upon receptor activation, the ligand-restricted SMADs1–35 are phosphorylated in the C-terminal MH2 domain and recruit the common subunit SMAD4/DPC-4 gene to the nucleus to mediate target gene expression. Frequent inactivating mutations of SMAD4, or less common somatic mutations ofSMAD2 seen in solid tumors, suggest that these genes have a suppressor function. However, there have been no identified mutations of SMAD5, although the gene localizes to the critical region of loss in chromosome 5q31.1 (chromosome 5, long arm, region 3, band 1, subband 1) in myelodysplasia (MDS) and acute myelogenous leukemia (AML). A ubiquitously expressed novel isoform,SMAD5β, encodes a 351 amino acid protein with a truncated MH2 domain and a unique C-terminal tail of 18 amino acids, which may be the functional equivalent of inactivating mutations. The levels of SMAD5β transcripts are higher in the undifferentiated CD34+ hematopoietic stem cells than in the terminally differentiated peripheral blood leukocytes, thereby implicating the β form in stem cell homeostasis. Yeast 2-hybrid interaction assays reveal the lack of physical interactions between SMAD5β and SMAD5 or SMAD4. The expression ofSMAD5β may represent a novel mechanism to protect pluripotent stem cells and malignant cells from the growth inhibitory and differentiation signals of BMPs.

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