Abstract 2348: Low PU.1 expression not only attenuates neutrophil differentiation of AML cells but also increases resistance to cytotoxic therapies

AbstractASXL1 is one of the most frequently mutated genes in malignant myeloid diseases. In patients with myeloid malignancies, ASXL1 mutations are usually heterozygous frameshift or nonsense mutations leading to C-terminal truncation. Current disease models have predominantly total loss of ASXL1 or overexpressed C-terminal truncations. These models cannot fully recapitulate leukemogenesis and disease progression. We generated an endogenous C-terminal-truncated Asxl1 mutant in zebrafish that mimics human myeloid malignancies. At the embryonic stage, neutrophil differentiation was explicitly blocked. At 6 months, mutants initially exhibited a myelodysplastic syndrome-like phenotype with neutrophilic dysplasia. At 1 year, about 13% of mutants further acquired the phenotype of monocytosis, which mimics chronic myelomonocytic leukemia, or increased progenitors, which mimics acute myeloid leukemia. These features are comparable to myeloid malignancy progression in humans. Furthermore, transcriptome analysis, inhibitor treatment, and rescue assays indicated that asxl1-induced neutrophilic dysplasia was associated with reduced expression of bmi1a, a subunit of polycomb repressive complex 1 and a reported myeloid leukemia-associated gene. Our model demonstrated that neutrophilic dysplasia caused by asxl1 mutation is a foundation for the progression of myeloid malignancies, and illustrated a possible effect of the Asxl1-Bmi1a axis on regulating neutrophil development.

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Glycosphingolipid expression in acute nonlymphocytic leukemia: common expression of shiga toxin and parvovirus B19 receptors on early myeloblasts

Blood ◽

10.1182/blood-2002-03-0718 ◽

2003 ◽

Vol 101 (2) ◽

pp. 711-721 ◽

Cited By ~ 20

Author(s):

Laura L. W. Cooling ◽

De Sheng Zhang ◽

Stanley J. Naides ◽

Theodore A. W. Koerner

Keyword(s):

Shiga Toxin ◽

High Performance ◽

Cell Membranes ◽

Ex Vivo ◽

Parvovirus B19 ◽

Terminal Deoxynucleotidyl Transferase ◽

Acute Nonlymphocytic Leukemia ◽

Blood Neutrophils ◽

Neutrophil Differentiation ◽

Layer Chromatography

Glycosphingolipids (GSLs) are complex macromolecules on cell membranes that have been shown to play a role in neutrophil differentiation, activation, phagocytosis, and adhesion to both microorganisms and vascular endothelium. Because GSLs are often cryptic antigens on cell membranes, little is known regarding GSL expression in early myelopoiesis. To study the latter, myeloblasts were collected from patients with acute nonlymphocytic leukemia (ANLL) who required therapeutic leukocytopheresis for hyperleukocytosis. The neutral GSLs were isolated and identified by high-performance thin-layer chromatography (HPTLC), HPTLC immunostaining, gas chromatography, nuclear magnetic resonance, and fast atom bombardment–mass spectrometry. Like mature peripheral blood neutrophils, myeloblasts expressed glucosylceramide, lactosylceramide, and the neolacto-family GSLs, lactotriaosylceramide and neolactotetraosylceramide. Unlike neutrophils and chronic myeloid leukemia, most ANLL samples also expressed the globo-series GSLs, globotriaosylceramide and globotetraosylceramide. Globo GSL expression was strongly associated with a myeloblastic (ANLL M0-M2) and monoblastic phenotype (M5). A weak association was also noted with expression of either lymphoid (P < .10) or early hematopoietic markers (terminal deoxynucleotidyl transferase [TdT], CD34; P < .10). Globo-positive ANLL samples bound both shiga toxin and parvovirus B19 on HPTLC immunostaining. Based on these findings, we propose that neolacto and globo GSLs are expressed during early myeloid differentiation. Globotriaosylceramide expression on myeloblasts, and possibly myeloid stem cells, may have important implications for the use of shiga toxin as an ex vivo purging agent in autologous stem cell transplantation. Expression of globotetraosylceramide, the parvovirus B19 receptor, on myeloblasts may also explain the association between B19 infection, aplastic anemia, and chronic neutropenia of childhood.

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Dynamics of tumor growth and combination of anti-angiogenic and cytotoxic therapies

Physics in Medicine and Biology ◽

10.1088/0031-9155/52/13/001 ◽

2007 ◽

Vol 52 (13) ◽

pp. 3665-3677 ◽

Cited By ~ 41

Author(s):

M Kohandel ◽

M Kardar ◽

M Milosevic ◽

S Sivaloganathan

Keyword(s):

Tumor Growth ◽

Cytotoxic Therapies

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Correction: Differential Expression of Adenine Nucleotide Converting Enzymes in Mitochondrial Intermembrane Space: A Potential Role of Adenylate Kinase Isozyme 2 in Neutrophil Differentiation

PLoS ONE ◽

10.1371/journal.pone.0099247 ◽

2014 ◽

Vol 9 (5) ◽

pp. e99247

Author(s):

Keyword(s):

Differential Expression ◽

Adenylate Kinase ◽

Potential Role ◽

Adenine Nucleotide ◽

Intermembrane Space ◽

Mitochondrial Intermembrane Space ◽

Neutrophil Differentiation

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Congenital neutropenia

Hematology ◽

10.1182/asheducation-2009.1.344 ◽

2009 ◽

Vol 2009 (1) ◽

pp. 344-350 ◽

Cited By ~ 24

Author(s):

Christoph Klein

Keyword(s):

Ribosomal Proteins ◽

Mitochondrial Proteins ◽

Phenotypic Traits ◽

Genetic Defects ◽

Congenital Neutropenia ◽

Lysosomal Function ◽

Genes Encoding ◽

And Function ◽

Neutrophil Differentiation ◽

Remarkable Progress

Abstract Congenital neutropenia comprises a variety of genetically heterogeneous phenotypic traits. Molecular elucidation of the underlying genetic defects has yielded important insights into the physiology of neutrophil differentiation and function. Non-syndromic variants of congenital neutropenia are caused by mutations in ELA2, HAX1, GFI1, or WAS. Syndromic variants of congenital neutropenia may be due to mutations in genes controlling glucose metabolism (SLC37A4, G6PC3) or lysosomal function (LYST, RAB27A, ROBLD3/p14, AP3B1, VPS13B). Furthermore, defects in genes encoding ribosomal proteins (SBDS, RMRP) and mitochondrial proteins (AK2, TAZ) are associated with congenital neutropenia syndromes. Despite remarkable progress in the field, many patients with congenital neutropenia cannot yet definitively be classified by genetic terms. This review addresses diagnostic and therapeutic aspects of congenital neutropenia and covers recent molecular and pathophysiological insights of selected congenital neutropenia syndromes.

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Next-generation sequencing of adrenocortical carcinoma reveals new routes to targeted therapies

Journal of Clinical Pathology ◽

10.1136/jclinpath-2014-202514 ◽

2014 ◽

Vol 67 (11) ◽

pp. 968-973 ◽

Cited By ~ 33

Author(s):

J S Ross ◽

K Wang ◽

J V Rand ◽

L Gay ◽

M J Presta ◽

...

Keyword(s):

Next Generation Sequencing ◽

Adrenocortical Carcinoma ◽

Single Case ◽

Next Generation ◽

Illumina Hiseq ◽

Modest Improvement ◽

Aggressive Form ◽

Comprehensive Genomic Profiling ◽

Cytotoxic Therapies ◽

Generation Sequencing

AimsAdrenocortical carcinoma (ACC) carries a poor prognosis and current systemic cytotoxic therapies result in only modest improvement in overall survival. In this retrospective study, we performed a comprehensive genomic profiling of 29 consecutive ACC samples to identify potential targets of therapy not currently searched for in routine clinical practice.MethodsDNA from 29 ACC was sequenced to high, uniform coverage (Illumina HiSeq) and analysed for genomic alterations (GAs).ResultsAt least one GA was found in 22 (76%) ACC (mean 2.6 alterations per ACC). The most frequent GAs were in TP53 (34%), NF1 (14%), CDKN2A (14%), MEN1 (14%), CTNNB1 (10%) and ATM (10%). APC, CCND2, CDK4, DAXX, DNMT3A, KDM5C, LRP1B, MSH2 and RB1 were each altered in two cases (7%) and EGFR, ERBB4, KRAS, MDM2, NRAS, PDGFRB, PIK3CA, PTEN and PTCH1 were each altered in a single case (3%). In 17 (59%) of ACC, at least one GA was associated with an available therapeutic or a mechanism-based clinical trial.ConclusionsNext-generation sequencing can discover targets of therapy for relapsed and metastatic ACC and shows promise to improve outcomes for this aggressive form of cancer.

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IMMU-06. COMBINATORIAL IMMUNE CHECKPOINT INHIBITORS FOR TARGETED INTRATUMORAL DELIVERY IN GBM

Neuro-Oncology ◽

10.1093/neuonc/noab196.366 ◽

2021 ◽

Vol 23 (Supplement_6) ◽

pp. vi93-vi93

Author(s):

Stephanie Sanders ◽

Denise Herpai ◽

Waldemar Debinski

Keyword(s):

T Cells ◽

Tumor Cells ◽

Immune Checkpoint ◽

Immune Checkpoint Inhibitors ◽

Checkpoint Inhibitors ◽

Cell Activity ◽

Live Cells ◽

Normal Brain ◽

Cytotoxic Therapies ◽

Checkpoint Receptors

Abstract Glioblastoma (GBM) is an immunologically cold tumor. Using single cell sequencing of CD45+ cells we confirmed that T cells are present within GBM samples. These T cells are positive for exhaustion markers such as LAG3 and TIGIT, as well as CTLA4 and PD1 checkpoint receptors. Modulating T cell activity through use of immune checkpoint inhibitors (ICIs) has shown efficacy in the treatment of a variety of solid tumors, and the combination of anti-CTLA4 and anti-PD1 ICIs has shown increased efficacy over use of a single therapeutic. Additionally, targeting ICIs to the tumor cells may increase efficacy of this treatment. We therefore constructed a combinatorial ICI redirected to GBM via interleukin 13 receptor alpha 2 (IL13RA2), a receptor over-expressed on the majority of GBM cells but not normal brain. The first component of the construct, labeled with a histidine tag, targets CTLA4 while the second component, tagged with a StrepII tag, targets PD1. The tags added to the constructs will allow for purification of a combinatorial heterodimer simultaneously targeting PD1, CTLA4 and IL13RA2. We purified individual components via fast protein liquid chromatography (FPLC) using a proteinG column followed by a HisTrap or StrepTrap column. We obtained a recombinant, targeted multivalent ICI at > 95% purity. We found that these constructs are able to bind their target receptors via ELISA in which the Kd values ranged from picomolar to low nanomolar range. Additionally, our constructs bind their target on live cells by flow cytometry. We next designed a heterodimeric construct which can combinatorially target CTLA4 and PD1 while also directing the ICI therapy to GBM. These constructs in conjunction with other immune stimulants like cytotoxic therapies are intended to facilitate the interaction between T cells and GBM tumor cells directly in a tumor microenvironment.

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ATRA resolves the differentiation block in t(15;17) acute myeloid leukemia by restoring PU.1 expression

Blood ◽

10.1182/blood-2005-07-3068 ◽

2006 ◽

Vol 107 (8) ◽

pp. 3330-3338 ◽

Cited By ~ 139

Author(s):

Beatrice U. Mueller ◽

Thomas Pabst ◽

José Fos ◽

Vibor Petkovic ◽

Martin F. Fey ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Retinoic Acid ◽

Acute Promyelocytic Leukemia ◽

Myeloid Leukemia ◽

Retinoic Acid Receptor ◽

Promyelocytic Leukemia ◽

All Trans Retinoic Acid ◽

Conditional Expression ◽

Neutrophil Differentiation ◽

Acute Myeloid

Abstract Tightly regulated expression of the transcription factor PU.1 is crucial for normal hematopoiesis. PU.1 knockdown mice develop acute myeloid leukemia (AML), and PU.1 mutations have been observed in some populations of patients with AML. Here we found that conditional expression of promyelocytic leukemia-retinoic acid receptor α (PML-RARA), the protein encoded by the t(15;17) translocation found in acute promyelocytic leukemia (APL), suppressed PU.1 expression, while treatment of APL cell lines and primary cells with all-trans retinoic acid (ATRA) restored PU.1 expression and induced neutrophil differentiation. ATRA-induced activation was mediated by a region in the PU.1 promoter to which CEBPB and OCT-1 binding were induced. Finally, conditional expression of PU.1 in human APL cells was sufficient to trigger neutrophil differentiation, whereas reduction of PU.1 by small interfering RNA (siRNA) blocked ATRA-induced neutrophil differentiation. This is the first report to show that PU.1 is suppressed in acute promyelocytic leukemia, and that ATRA restores PU.1 expression in cells harboring t(15;17).

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