scholarly journals Human malignancy-associated nucleolar antigen as a marker for tumor cells in patients with acute leukemia

Blood ◽  
1984 ◽  
Vol 63 (3) ◽  
pp. 676-683
Author(s):  
FM Davis ◽  
WN Hittelman ◽  
KB McCredie ◽  
MJ Keating ◽  
L Vellekoop ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Leukemia ◽  
Leukemia Cell ◽  
Leukemic Cells ◽  
Lower Percentage ◽  
Differentiated Cells ◽  
Human Malignancy ◽  
Blast Cells

Tumor burden in adult patients with acute leukemia is assessed using the percentage of blast cells in the bone marrow or blood. It is clear, however, that not all blast cells are leukemic cells, especially during rapid marrow regeneration. Similarly, some leukemia cell lines have been shown to differentiate in vitro, and the same process also occurs in vivo. Therefore, the leukemic burden may be due to more differentiated cells as well as to blast cells. The purpose of this study was to investigate whether the human malignancy-associated nucleolar antigen (HMNA) could be used as a marker for leukemic cells and to examine its potential as a diagnostic tool. The proportion of HMNA-positive cells in the bone marrow of patients with acute leukemia was determined by indirect immunofluorescence with antibodies to HMNA and was compared with the differential counts routinely made in the clinic laboratory. The percentages of HMNA-positive cells among the nucleated cells in the marrow of 72 patients with clinical evidence of leukemia were significantly higher (range 9%-98%, median 83%) than those observed for nonleukemic individuals (range less than 0.05%-2.5%, median 1%) or for fractions of marrow cells from normal volunteers enriched for normal early progenitor cells (less than or equal to 2%). Patients with leukemia in remission had a lower percentage of HMNA- positive cells (range 0%-83%, median 3%). The percentage of HMNA- positive cells increased as patients approached relapse. Although the percentage of HMNA-positive cells was related to the percentage of blast cells in the bone marrow of the patients with leukemia, some partially differentiated cells were also HMNA-positive in some specimens, and some blastic cells were HMNA-negative in other specimens. These studies indicate the potential usefulness of HMNA as a marker for leukemic cells.

scholarly journals Human malignancy-associated nucleolar antigen as a marker for tumor cells in patients with acute leukemia

Blood ◽  
1984 ◽  
Vol 63 (3) ◽  
pp. 676-683 ◽  
Author(s):  
FM Davis ◽  
WN Hittelman ◽  
KB McCredie ◽  
MJ Keating ◽  
L Vellekoop ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Leukemia ◽  
Leukemia Cell ◽  
Leukemic Cells ◽  
Lower Percentage ◽  
Differentiated Cells ◽  
Human Malignancy ◽  
Blast Cells

Abstract Tumor burden in adult patients with acute leukemia is assessed using the percentage of blast cells in the bone marrow or blood. It is clear, however, that not all blast cells are leukemic cells, especially during rapid marrow regeneration. Similarly, some leukemia cell lines have been shown to differentiate in vitro, and the same process also occurs in vivo. Therefore, the leukemic burden may be due to more differentiated cells as well as to blast cells. The purpose of this study was to investigate whether the human malignancy-associated nucleolar antigen (HMNA) could be used as a marker for leukemic cells and to examine its potential as a diagnostic tool. The proportion of HMNA-positive cells in the bone marrow of patients with acute leukemia was determined by indirect immunofluorescence with antibodies to HMNA and was compared with the differential counts routinely made in the clinic laboratory. The percentages of HMNA-positive cells among the nucleated cells in the marrow of 72 patients with clinical evidence of leukemia were significantly higher (range 9%-98%, median 83%) than those observed for nonleukemic individuals (range less than 0.05%-2.5%, median 1%) or for fractions of marrow cells from normal volunteers enriched for normal early progenitor cells (less than or equal to 2%). Patients with leukemia in remission had a lower percentage of HMNA- positive cells (range 0%-83%, median 3%). The percentage of HMNA- positive cells increased as patients approached relapse. Although the percentage of HMNA-positive cells was related to the percentage of blast cells in the bone marrow of the patients with leukemia, some partially differentiated cells were also HMNA-positive in some specimens, and some blastic cells were HMNA-negative in other specimens. These studies indicate the potential usefulness of HMNA as a marker for leukemic cells.

Massive Mobilization of AML Cells into Circulation by Disruption of Leukemia/Stroma Cell Interactions Using CXCR4 Antagonist AMD3100: First Evidence in Patients and Potential for Abolishing Bone Marrow Microenvironment-Mediated Resistance.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 568-568 ◽  
Author(s):  
Michael Andreeff ◽  
Sergej Konoplev ◽  
Rui-Yu Wang ◽  
Zhihong Zeng ◽  
Teresa McQueen ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Leukemia Cell ◽  
Surface Expression ◽  
Bone Marrow Microenvironment ◽  
Leukemic Cells ◽  
Fish Analysis ◽  
Cxcr4 Antagonist ◽  
Induced Apoptosis

Abstract The chemokine receptor CXCR4 is critically involved in migration of hematopoietic cells to the stromal derived factor (SDF-1α)-producing bone marrow microenvironment. CXCR4 is regulated in part by mutant FLT3 signaling, but in a series of 122 AML samples with diploid karyotype and lack of FLT3 mutation (ITD), high CXCR4 expression negatively correlated with DFS and OS (p=0.03 and p=0.04, respectively), after multivariate analysis (Konoplev, ASH 2006). We hypothesized that inhibition of SDF-1α-/CXCR4 interactions would result in mobilization of leukemic blasts from the bone marrow into circulation. The in vivo effect of the CXCR4 antagonist AMD3100 was studied in three patients with AML, who had insufficient mobilization of CD34+ cells for autologous stem cell transplantation with G-CSF and/or cytoxan. The combination of G-CSF (10 μg/kg QD) and AMD3100 (240 μg/kg QD SC starting on d4 and repeated for 3–4 days) resulted in massive mobilization of leukemic cells into the circulation in a time-dependent fashion, as determined by flow cytometry and interphase FISH analysis of their respective cytogenetic abnormalities. Patient # Cytogenetics % (+) cells % (+) cells Apheresis FCM Day 2 Day 4/5 CD34x106/kg 1 Trisomy 21 22.6 57.0 FCM CD7/33 22.0 2 Trisomy 9 28.6 68.6 Inv 16 29.0 75.8 4.8 FCM CD13/33 74.0 3 Mono 17 40.4 53.4 5q31 37.5 49.6 8.7 FCM CD13/33 50.0 We and others have previously demonstrated that stroma/leukemia interactions mediate protection of leukemic cells from chemotherapy-induced apoptosis (Konopleva et al, Leukemia2002:1713). We then tested the hypothesis that CXCR4 inhibition would result in increased sensitivity to chemotherapy, using AMD3465, the second generation small-molecule CXCR4 inhibitor with greater potency than AMD3100. Results demonstrate inhibition of surface expression of CXCR4 and of SDF-1α-, and stroma(MS-5)-induced migration of AML cells. In vitro co-culture systems with stromal cells significantly protected leukemic cells (p < 0.01), while AMD3465 decreased stroma-mediated protection from AraC and Busulfan apoptosis and downregulated AKT signaling in AML cells. In a murine model of luciferase labeled Baf-FLT3ITD leukemias, AMD3465 induced massive dissemination of leukemia, which was abrogated by treatment with Sorafenib, a potent FLT3ITD inhibitor (Zhang, ASH 2006). Taken together, our data suggest that SDF-1α/CXCR4 interactions contribute to the resistance of leukemic cells to chemotherapy-induced apoptosis. Disruption of these interactions by CXCR4 inhibition results in leukemia dissemination and chemosensitization. Our results in leukemia patients provide first in man proof-of principle for a novel strategy of targeting the leukemia cell/bone marrow microenvironment interactions. A clinical trial testing this concept in patients with AML is under development.

scholarly journals In vivo control of differentiation of myeloid leukemic cells by recombinant granulocyte-macrophage colony-stimulating factor and interleukin 3

Blood ◽  
1988 ◽  
Vol 71 (2) ◽  
pp. 375-382 ◽  
Author(s):  
J Lotem ◽  
L Sachs
Keyword(s):  
Therapeutic Potential ◽  
Regulatory Proteins ◽  
Leukemic Cells ◽  
Cell Multiplication ◽  
Interleukin 3 ◽  
Gm Csf ◽  
Blast Cells ◽  
Macrophage Colony

The normal myeloid hematopoietic regulatory proteins include one class of proteins that induces viability and multiplication of normal myeloid precursor cells to form colonies (colony-stimulating factors [CSF] and interleukin 3 [IL-3], macrophage and granulocyte inducing proteins, type 7 [MGI-1]) and another class (called MGI-2) that induces differentiation of normal myeloid precursors without inducing cell multiplication. Different clones of myeloid leukemic cells can differ in their response to these regulatory proteins. One type of leukemic clone can be differentiated in vitro to mature cells by incubating with the growth-inducing proteins granulocyte-macrophage (GM) CSF or IL-3, and another type of clone can be differentiated in vitro to mature cells by the differentiation-inducing protein MGI-2. We have now studied the ability of different myeloid regulatory proteins to induce the in vivo differentiation of these different types of mouse myeloid leukemic clones in normal and cyclophosphamide-treated mice. The results show that in both types of mice (a) the in vitro GM-CSF- and IL- 3-sensitive leukemic cells were induced to differentiate to mature cells in vivo in mice injected with pure recombinant GM-CSF and IL-3 but not with G-CSF, M-CSF, or MGI-2; (b) the in vitro MGI-2-sensitive leukemic cells differentiated in vivo by injection of MGI-2 and also, presumably indirectly, by GM-CSF and IL-3 but not by M-CSF or G-CSF; (c) in vivo induced differentiation of the leukemic cells was associated with a 20- to 60-fold decrease in the number of blast cells; and (d) all the injected myeloid regulatory proteins stimulated the normal myelopoietic system. Different normal myeloid regulatory proteins can thus induce in vivo terminal differentiation of leukemic cells, and it is suggested that these proteins can have a therapeutic potential for myeloid leukemia in addition to their therapeutic potential in stimulating normal hematopoiesis.

Modeling the human 8p11-myeloproliferative syndrome in immunodeficient mice

Blood ◽  
2010 ◽  
Vol 116 (12) ◽  
pp. 2103-2111 ◽  
Author(s):  
Helena Ågerstam ◽  
Marcus Järås ◽  
Anna Andersson ◽  
Petra Johnels ◽  
Nils Hansen ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Leukemia ◽  
Tyrosine Kinases ◽  
Cellular Proliferation ◽  
Myeloid Cell ◽  
Myeloproliferative Disorder ◽  
Immunodeficient Mice ◽  
Myeloproliferative Syndrome

Abstract The 8p11 myeloproliferative syndrome (EMS), also referred to as stem cell leukemia/lymphoma, is a chronic myeloproliferative disorder that rapidly progresses into acute leukemia. Molecularly, EMS is characterized by fusion of various partner genes to the FGFR1 gene, resulting in constitutive activation of the tyrosine kinases in FGFR1. To date, no previous study has addressed the functional consequences of ectopic FGFR1 expression in the potentially most relevant cellular context, that of normal primary human hematopoietic cells. Herein, we report that expression of ZMYM2/FGFR1 (previously known as ZNF198/FGFR1) or BCR/FGFR1 in normal human CD34+ cells from umbilical-cord blood leads to increased cellular proliferation and differentiation toward the erythroid lineage in vitro. In immunodeficient mice, expression of ZMYM2/FGFR1 or BCR/FGFR1 in human cells induces several features of human EMS, including expansion of several myeloid cell lineages and accumulation of blasts in bone marrow. Moreover, bone marrow fibrosis together with increased extramedullary hematopoiesis is observed. This study suggests that FGFR1 fusion oncogenes, by themselves, are capable of initiating an EMS-like disorder, and provides the first humanized model of a myeloproliferative disorder transforming into acute leukemia in mice. The established in vivo EMS model should provide a valuable tool for future studies of this disorder.

scholarly journals Curcumin and Its Analogue Induce Apoptosis in Leukemia Cells and Have Additive Effects with Bortezomib in Cellular and Xenograft Models

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
L. I. Nagy ◽  
L. Z. Fehér ◽  
G. J. Szebeni ◽  
M. Gyuris ◽  
P. Sipos ◽  
...  
Keyword(s):  
Leukemia Cell ◽  
Leukemic Cells ◽  
Leukemia Cell Line ◽  
Leukemia Cells ◽  
Great Promise ◽  
Human Leukemia ◽  
Human Leukemia Cell Line ◽  
Apoptotic Effects

Combination therapy of bortezomib with other chemotherapeutics is an emerging treatment strategy. Since both curcumin and bortezomib inhibit NF-κB, we tested the effects of their combination on leukemia cells. To improve potency, a novel Mannich-type curcumin derivative, C-150, was synthesized. Curcumin and its analogue showed potent antiproliferative and apoptotic effects on the human leukemia cell line, HL60, with different potency but similar additive properties with bortezomib. Additive antiproliferative effects were correlated well with LPS-induced NF-κB inhibition results. Gene expression data on cell cycle and apoptosis related genes, obtained by high-throughput QPCR, showed that curcumin and its analogue act through similar signaling pathways. In correlation with in vitro results similar additive effect could be obsereved in SCID mice inoculated systemically with HL60 cells. C-150 in a liposomal formulation given intravenously in combination with bortezomib was more efficient than either of the drugs alone. As our novel curcumin analogue exerted anticancer effects in leukemic cells at submicromolar concentration in vitro and at 3 mg/kg dose in vivo, which was potentiated by bortezomib, it holds a great promise as a future therapeutic agent in the treatment of leukemia alone or in combination.

The Bone Marrow Niche of Patients with Acute Lymphoblastic Leukemia Produces No Increased Asparagine Levels In Vivo That May Lead to Clinical Asparaginase Resistance

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1505-1505
Author(s):  
Wing H. Tong ◽  
Rob Pieters ◽  
Wim C.J. Hop ◽  
Claudia Lanvers-Kaminsky ◽  
Joachim Boos ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Aspartic Acid ◽  
Peripheral Blood ◽  
Lymphoblastic Leukemia ◽  
Mesenchymal Cells ◽  
Leukemic Cells ◽  
Significant Difference ◽  
Trough Levels

Abstract Abstract 1505 Asparaginase is an essential component of combination chemotherapy of acute lymphoblastic leukemia (ALL). Asparaginase breaks down asparagine into aspartic acid and ammonia. Because asparagine is necessary for protein synthesis, its depletion leads to cell death. Recently, it has been suggested that mesenchymal cells in the bone marrow may produce asparagine and form ‘protective niches’ for leukemic cells. In vitro, this led to high levels of asparagine and asparaginase resistance of the ALL cells (Iwamoto et al. (J Clin Invest. 2007)). However, it is unknown if this holds true for the clinical in vivo situation. The aim of our study is to analyse whether mesenchymal cells or other cells in the bone marrow indeed produce significant amounts of asparagine in vivo that may lead to clinical asparaginase resistance. Ten de novo ALL patients were enrolled in this study. All children received induction chemotherapy according to protocol 1-A and 1-B of the Dutch Childhood Oncology Group (DCOG) ALL-10 protocol. Asparaginase levels and amino acid levels (asparagine, aspartic acid, glutamine and glutamic acid) were measured in bone marrow (BM) and peripheral blood at diagnosis (day 1), days 15, 33 and 79. On days that asparaginase was administered (days 15 and 33) it was ensured that study material was obtained before the E-coli L-asparaginase infusions. Changes over time of asparaginase trough levels in BM and peripheral blood were evaluated using Mixed models ANOVA. The amino acids levels in 0.5 ml BM, 3 ml BM and peripheral blood at days 15 and 33 were also compared using Mixed models ANOVA. All these analyses were done after log transformation of measured values to get approximate normal distributions. A two-sided p-value < 0.05 was considered statistically significant. The asparaginase levels were all below detection limit (< 5 IU/L) in BM and peripheral blood at days 1 and 79. In both compartments, the median asparaginase trough levels were not significantly different at days 15 and 33. At diagnosis, no significant difference in asparagine level between 3 ml BM and peripheral blood was found (median: 44.5 μM (range 20.6–59.6 μM) and 43.9 μM (range 18.4 –58.5 μM), respectively). However, the median level of aspartic acid at diagnosis in 3 ml BM (19.2 μM; range 6.2–52.6 μM) was significantly higher as compared to median level of peripheral blood (5.7 μM; range 2.4–10.1 μM) (p=0.002). The aspartic acid levels were also higher in BM compared to peripheral blood at days 15 and 33 (both p=0.001) and at day 79 (p=0.002). Aspartic acid levels were significantly higher in 0.5 ml versus 3 ml BM (p=0.001) and this difference was also found when comparing 0.5 ml BM versus peripheral blood (p<0.001) suggesting dilution with peripheral blood when taking higher volumes of ‘bone marrow’. Asparagine levels were all below the lower limit of quantification (LLQ < 0.2 μM) in both BM and blood during asparaginase treatment at days 15 and 33. At day 79, no significant difference in asparagine levels between BM (37.7 μM; range 33.4–50.3 μM) and peripheral blood (38.9 μM; range 25.7 –51.3 μM) was seen. During the time course of asparaginase infusions, the glutamine and glutamic acid levels did not change significantly. In conclusion, we demonstrate higher aspartic acid levels in bone marrow compared to peripheral blood. The higher aspartic acid levels are detected at diagnosis, during asparaginase therapy at days 15 and 33, and also at day 79 at complete remission, showing that these do not originate from leukemic cells nor from asparagine breakdown by asparaginase but from cells in the microenvironment of the bone marrow. However, there is no increased asparagine synthesis in vivo in the bone marrow of ALL patients. Therefore, increased asparagine synthesis by mesenchymal cells may be of relevance for resistance to asparaginase of leukemic cells in vitro but not in vivo. Disclosures: No relevant conflicts of interest to declare.

Identification of Non-Cardiotoxic hERG1 Blockers to Overcome Chemoresistance in Acute Lymphoblastic Leukemias

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1506-1506
Author(s):  
Marika Masselli ◽  
Serena Pillozzi ◽  
Massimo D'Amico ◽  
Luca Gasparoli ◽  
Olivia Crociani ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mouse Model ◽  
Map Kinases ◽  
Conflicts Of Interest ◽  
Lymphoblastic Leukemia ◽  
Significant Proportion ◽  
Leukemic Cells ◽  
K Channel

Abstract Abstract 1506 Although cure rates for children with acute lymphoblastic leukemia (ALL), the most common pediatric malignancy, have markedly improved over the last two decades, chemotherapy resistance remains a major obstacle to successful treatment in a significant proportion of patients (Pui CH et al. N Engl J Med., 360:2730–2741, 2009). Increasing evidence indicates that bone marrow mesenchymal cells (MSCs) contribute to generate drug resistance in leukemic cells (Konopleva M et al., Leukemia, 16:1713–1724, 2002). We contributed to this topic, describing a novel mechanism through which MSCs protect leukemic cells from chemotherapy (Pillozzi S. et al., Blood, 117:902–914, 2011.). This protection depends on the formation of a macromolecular membrane complex, on the plasma membrane of leukemic cells, the major players being i) the human ether-a-gò-gò-related gene 1 (hERG1) K+ channel, ii) the β1integrin subunit and iii) the SDF-1α receptor CXCR4. In leukemic blasts, the formation of this protein complex activates both the ERK 1/2 MAP kinases and the PI3K/Akt signalling pathways triggering antiapoptotic effects. hERG1 exerts a pivotal role in the complex, as clearly indicated by the effect of hERG1 inhibitors to abrogate MSCs protection against chemotherapeutic drugs. Indeed, E4031, a class III antiarrhythmic that specifically blocks hERG1, enhances the cytotoxicity of drugs commonly used to treat leukemia, both in vitro and in vivo. The latter was tested in a human ALL mouse model, consisting of NOD/SCID mice injected with REH cells, which are relatively resistant to corticosteroids. Mice were treated for 2 weeks with dexamethasone, E4031, or both. Treatment with dexamethasone and E4031 in combination nearly abolished bone marrow engraftment while producing marked apoptosis, and strongly reducing the proportion of leukemic cells in peripheral blood and leukemia infiltration of extramedullary sites. These effects were significantly superior to those obtained by treatment with either dexamethasone alone or E4031 alone. This model corroborated the idea that hERG1 blockers significantly increase the rate of leukemic cell apoptosis in bone marrow and reduced leukemic infiltration of peripheral organs. From a therapeutic viewpoint, to develop a pharmacological strategy based on hERG1 targeting we must consider to circumvent the side effects exerted by hERG1 blockers. Indeed, hERG1 blockers are known to retard the cardiac repolarization, thus lengthening the electrocardiographic QT interval, an effect that in some cases leads to life threatening ventricular arrhythmias (torsades de points). On the whole, it is mandatory to design and test non-cardiotoxic hERG1 blockers as a new strategy to overcome chemoresistance in ALL. On these bases, we tested compounds with potent anti-hERG1 effects, besides E4031, but devoid of cardiotoxicity (e.g. non-torsadogenic hERG1 blockers). Such compounds comprise erythromycin, sertindole and CD160130 (a newly developed drug by BlackSwanPharma GmbH, Leipzig, Germany). We found that such compounds exert a strong anti-leukemic activity both in vitro and in vivo, in the ALL mouse model described above. This is the first study describing the chemotherapeutic effects of non-torsadogenic hERG1 blockers in mouse models of human ALL. This work was supported by grants from the Associazione Genitori contro le Leucemie e Tumori Infantili Noi per Voi, Associazione Italiana per la Ricerca sul Cancro (AIRC) and Istituto Toscano Tumori. Disclosures: No relevant conflicts of interest to declare.

ARC Is Regulated by MAPK and PI3K and Confers Drug Resistance and Survival Advantage to AML in Vitro and in Vivo

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 893-893
Author(s):  
Po Yee Mak ◽  
Duncan H Mak ◽  
Yuexi Shi ◽  
Vivian Ruvolo ◽  
Rodrigo Jacamo ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Leukemia Cell ◽  
Leukemia Cells ◽  
Apoptosis Resistance ◽  
Control Mscs ◽  
Extrinsic Apoptosis ◽  
Induced Apoptosis ◽  
And Control

Abstract Abstract 893 ARC (Apoptosis repressor with caspase recruitment domain) is a unique antiapoptotic protein that has been shown to suppress the activation of both intrinsic and extrinsic apoptosis. We previously reported that ARC is one of the most potent adverse prognostic factors in AML and that high ARC protein expression predicted shorter survival and poor clinical outcome in patients with AML (Carter BZ et al., Blood 2011). Here we report how ARC is regulated and its role in inhibition of AML apoptosis and in cell survival. We provide evidence that ARC expression is regulated by MAPK and PI3K signaling. Inhibition of MAPK and PI3K pathways decreased ARC mRNA and protein levels in AML cells. ARC expression in AML cells is upregulated in co-cultures with bone marrow-derived mesenchymal stromal cells (MSCs) and the upregulation is suppressed in the presence of MAPK or PI3K inhibitors. To investigate the role of ARC in apoptosis resistance in AML, we generated stable ARC overexpressing (O/E) KG-1 and stable ARC knock down (K/D) OCI-AML3 and Molm13 cells and treated them with Ara-C and agents selectively inducing intrinsic (ABT-737) or extrinsic (TRAIL) apoptosis. We found that ARC O/E cells are more resistant and ARC K/D cells more sensitive to Ara-C, ABT-737, and TRAIL-induced apoptosis: EC50s of Ara-C, ABT-737, or TRAIL treatment at 48 hours for ARC O/E KG-1 and control cells were 1.5 ± 0.1 μM vs. 83.5 ± 4.6 nM, 2.2 ± 0.2 μM vs. 60.2 ± 3.1 nM, or 0.97 ± 0.03 μg/mL vs. 0.17 ± 0.08 μg/mL, respectively and for ARC K/D OCI-AML3 and control cells were 0.33 ± 0.02 μM vs. 3.4 ± 0.2 μM, 0.24 ± 0.01 μM vs. 1.3 ± 0.1 μM, or 0.13 ± 0.09 μg/mL vs. 0.36 ± 0.03 μg/mL, respectively. Bone marrow microenvironment is known to play critical roles in AML disease progression and in protecting leukemia cells from various therapeutic agent-induced apoptosis. Leukemia cells were co-cultured with MSCs in vitro study to mimic the in vivo condition. ARC was found to be highly expressed in MSCs and stable ARC K/D MSCs were generated. AML cell lines and primary patient samples were co-cultured with ARC K/D or control MSCs and treated with Ara-C, ABT-737, or TRAIL. Interestingly, ARC K/D MSCs lost their protective activity for leukemia cells treated with these agents. EC50s for OCI-AML3 cells co-cultured with ARC K/D or control MSCs for 48 hours treated with Ara-C, ABT-737, or TRAIL were 1.0 ± 0.04 μM vs. 4.5 ± 0.2 μM, 0.15 ± 0.06 μM vs. 0.53 ± 0.02 μM, or 1.4 ± 0.8 μg/mL vs. 8.1 ± 0.3 μg/mL, respectively. In addition, ARC O/E KG-1 cells grew faster and ARC K/D OCI-AML3 and Molm13 cells and ARC K/D MSCs grew slower than their respective controls. We then injected KG-1 cells into mice and found that NOD-SCID mice harboring ARC O/E KG-1 had significantly shorter survival than mice injected with the vector control KG-1 (median 84 vs. 111 days) as shown in the figure. Collectively, results demonstrate that ARC plays critical roles in AML. ARC is regulated by MSCs through various signaling pathways in AML cells, protects leukemia cells from apoptosis induced by chemotherapy and by agents selectively inducing intrinsic and extrinsic apoptosis. ARC regulates leukemia cell growth in vitro and in vivo. The results suggest that ARC is a potential target for AML therapy. In addition, targeting ARC in MSCs suppresses microenvironmental protection of AML cells. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Proliferation and differentiation of human myeloid leukemic cells in immunodeficient mice: electron microscopy and cytochemistry

Blood ◽  
1984 ◽  
Vol 63 (5) ◽  
pp. 1015-1022 ◽  
Author(s):  
EA Machado ◽  
DA Gerard ◽  
CB Lozzio ◽  
BB Lozzio ◽  
JR Mitchell ◽  
...  
Keyword(s):  
Nude Mice ◽  
Leukemia Cell ◽  
Leukemic Cells ◽  
Leukemia Cell Line ◽  
Leukemia Cells ◽  
Human Leukemia ◽  
Immunodeficient Mice ◽  
Human Leukemia Cells

Abstract To study the influence of a biologic environment on cultured human leukemia cells, KG-1, KG-1a, and HL-60 cells were inoculated subcutaneously into newborn nude mice. The cells developed myelosarcomas at the site of inoculation and in lungs and kidneys. KG-1 and HL-60 myelosarcomas were successfully passaged through adult nude mice, whereas KG-1a tumors proliferated only after transplantation into newborn hosts. The human nature of the cells forming myelosarcomas in mice was assessed by chromosomal analyses and detection of cross- reactivity with an antibody to the human leukemia cell line K562. We undertook electron microscopic and cytochemical examinations of the cells proliferating in vitro and in the mice. The granules of KG-1 cells in vivo did not react for acid phosphatase, as observed in vitro, and the HL-60 cells proliferating in mice lost the perinuclear myeloperoxidase (MPO) demonstrated in cultured cells. Although the influence of an in vivo selection of cell subpopulations cannot be ruled out, the enzymatic changes are compatible with induced cell differentiation. Conclusive evidence of differentiation in vivo was observed in the KG-1a cell subline. The undifferentiated KG-1a blasts developed cytoplasmic granules and synthesized MPO during proliferation in vivo. These observations indicate that human leukemia cells from established cell lines proliferate in nude mice and may acquire new differentiated properties in response to the in vivo environment.

scholarly journals Leukemia-Associated HSC Vascular Niche Is Negatively Regulated By PERK of Unfolded Protein Response (UPR)

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 2486-2486
Author(s):  
Lan Zhou ◽  
Cui Liu ◽  
Stanley A Adoro ◽  
Lechuang Chen ◽  
Diana Ramirez ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Endothelial Cells ◽  
Er Stress ◽  
Leukemia Cell ◽  
Unfolded Protein ◽  
Vascular Niche ◽  
Activating Transcription Factor ◽  
Protein Response

T cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematological malignancy derived from early T cell progenitors. Diffuse infiltration of the bone marrow by T-ALL is associated with worse prognosis. We previously reported that actively proliferating leukemia cells inhibit normal hematopoietic stem and progenitor cell (HSPC) proliferation and homing to the perivascular region. We found that aberrant Notch activation in the stroma plays an important role in negatively regulating the expression of CXLC12 on osteoblasts and their differentiation. However, the underlying molecular mechanism that leads to the suppression of hematopoiesis and decreased HSPC in the vascular niche is unclear. It has been demonstrated that rapid cellular proliferation associated with oncogenic activity such as MYC in T-ALL leads to a global increase in protein synthesis and an increase in misfolded/unfolded polypeptides in the endoplasmic reticulum (ER), referred to as unfolded protein response (UPR) or ER stress. Elevated ER stress leads to activation of at least three types of ER stress transducers through the release of inhibitory binding by glucose-regulated chaperone protein (GRP78/BIP): the protein kinase RNA-like ER kinase (PERK), the inositol-requiring enzyme 1 (IRE1), and the activating transcription factor 6 (ATF6). Activation of PERK phosphorylates eIF2 to repress global translation with the exception of a small number of proteins including ATF4 (activating transcription factor-4). ATF4 regulates genes involved in restoring ER homeostasis and genes in apoptosis. Here, we studied the role of UPR in the regulation of HSC niche function in the setting of T-ALL progression. Using in vitro assays in which T-ALL leukemia cells driven by activated Notch1 (ICN1) were co-cultured with endothelial cells (MILE SVEN 1, MS1), and in vivo ICN1-driven T-ALL model, we found that PERK-eIF2a-ATF4 pathway was activated in both MS1 cells and BM endothelial cells isolated from T-ALL mice, while IRE1 and ATF6 pathways were only mildly altered. The activation of PERK was accompanied with the increased expression of Jagged1 and suppressed expression of CXCL12 in both cultured endothelial cells and bone marrow endothelial cells from leukemia mice. PERK inhibitor (GSK2606414) treatment of co-cultured cells largely restored CXCL12 expression, which was also negatively regulated by Jagged1, and accelerated the leukemia cell apoptosis as indicated by the enhanced annexin staining. These findings suggest that PERK is the upstream regulator of Jagged1 and CXCL12 in the endothelial cells; however, the function of cell-autonomous PERK on leukemia cell survival needs to be further clarified. To understand the role of PERK in bone marrow endothelium during leukemia development in vivo, we examined T-ALL leukemia progression and its effect on vascular niche function in VE-CadherinERT2/PERKF/F mice in which Perk was specifically deleted in endothelial cells. Consistent with in vitro findings, T-ALL development induced endothelial PERK-eIF2a-ATF4 activation, while up-regulated Jagged1 and down-regulated CXCL12 were also identified in isolated BM endothelial cells. Compared to the wild type mice, VE-CadherinERT2/PERKF/F mice showed attenuated leukemia progression, increased HSPC (Lin-Sca-1+c-kit+) frequency, and improved survival. Taken together, our findings suggest that PERK activation in BM endothelial cells is a key regulator of the leukemia vascular niche to promote leukemia progression and to suppress normal hematopoiesis. Therefore, targeting PERK may offer an effective strategy in restoring normal HSPC homeostasis and limiting leukemia progression. Disclosures No relevant conflicts of interest to declare.

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