scholarly journals In VivoMurine Model of Leukemia Cell-Induced Spinal Bone Destruction

2017 ◽  
Vol 2017 ◽  
pp. 1-6 ◽  
Author(s):  
Jia-Jie Chen ◽  
Wei Zhou ◽  
Nan Cai ◽  
Gang Chang
Keyword(s):  
Negative Impact ◽  
Leukemia Cell ◽  
Bone Destruction ◽  
Leukemic Cells ◽  
Inflammatory Factors ◽  
Bone Lesions ◽  
Nonobese Diabetic ◽  
Unbearable Pain ◽  
Hind Limb Paralysis

Osteolytic bone lesions can be a consequence of leukemic bone infiltration or focal bone destruction by inflammatory factors released from leukemic cells. Destructive bone lesions have a negative impact on the quality of life of leukemia patients, causing unbearable pain and, in some cases, limb paralysis. However, the mechanism, by which leukemic cells produce destructive bone lesions, and the effect of therapeutics on osteolytic lesions have not been fully elucidated yet and, thus, stand to benefit from anin vivomodel. To that end, HL-60 cells were transformed by retrovirus-mediated constitutively active (CA) STAT5 expression and injected into nonobese diabetic (NOD)/SCID mice via the tail vein. After three weeks, lumbar spines were subjected to histocytometric analysis. Xenograft mice developed hind limb paralysis in 2-3 weeks, which was consistent with the consequences of spinal bone destruction by extramedullary invasion of leukemia cells. Thein vivomodel will improve the understanding and treatment of osteolytic bone lesions caused by myeloid leukemic cells.

scholarly journals Stearoyl-CoA Desaturase (SCD) Enhances Central Nervous System Leukemia

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 389-389
Author(s):  
Angela Maria Savino ◽  
Orianne Olivares ◽  
Shani Barel ◽  
Lev Yakimov ◽  
Ifat Geron ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Hind Limb ◽  
Leukemia Cell ◽  
Leukemic Cells ◽  
Nsg Mice ◽  
Cns Relapse ◽  
Hind Limb Paralysis

Abstract Background: Central nervous system (CNS) involvement by acute lymphoblastic leukemia (ALL) is a major clinical concern. Leukemic cells can be found in the CNS at diagnosis (1-2%) or, more frequently, at relapse (30%). Very little is known about the pathogenesis and therefore there are no targeted therapies. Prophylactic CNS-directed conventional intrathecal chemotherapy or irradiation are required for relapse-free survival. However, they are associated with substantial rates of short and long term toxicity. Therefore, elucidation of molecular mechanisms and pathways mediating leukemia-cell entry and survival in the CNS is needed to develop alternative CNS-directed treatment strategies. Previous studies showed an increased expression of Stearoyl-CoA desaturase (SCD), a key enzyme of the de novo fatty acid synthesis pathway, in B cell precursor (BCP) ALL cells isolated from cerebrospinal fluid (CSF) of patients at the time of CNS relapse. A small SCD positive population was detected in the bone marrow (BM) at leukemia diagnosis in patients who later developed isolated CNS relapse, defining a potential biomarker for CNS relapse. It is unknown, however, if SCD has a functional role in CNS leukemia. Aim: To examine the hypothesis that increased expression of SCD enhances trafficking and survival of human B-ALL cells in the CNS Methods: We analyzed leukemia-cell entry into the CNS using xenografts of human BCP-ALL cell lines. Microarray profile of cells isolated from CNS and BM of transplanted mice was performed. Cell lines were transduced to overexpress human SCD and evaluated in vitro for proliferation kinetics and metabolic SCD activity. In vivo, SCD overexpressing cells were transplanted in NSG mice,sacrificed upon the first symptoms of CNS involvement, e.g. hind limb paralysis. BM, spleen and meninges were collected and analyzed to check human engraftment by FACS. The tumor load was expressed as total amount of leukemic cells in each organ. Competition assays were performed by transplanting SCD overexpressing and WT cells in the same mouse in a 1:1 ratio. Results: BCP-ALL cells transplanted into NSG mice faithfully recapitulated pathological features of meningeal infiltration seen in patients with ALL. Gene expression analysis of cells collected from BM and meninges of leukemic mice revealed up-regulation of the genes belonging to the signaling pathway of sterol regulatory element binding proteins (SREBPs) in ALL cells isolated from the CNS. SCD, whose transcription is controlled by the SREBP family, was significantly upregulated. SCD overexpression did not alter proliferation in vitro. Since SCD introduces a double bond in Stearoyl-CoA, its activity was measured as the ratio of unsaturated/saturated fatty acids in the cells. That ratio was increased in SCD overexpressing cells in vitro, confirming the functionality of the enzyme. In vivo, mice transplanted with SCD overexpressing cells led to a faster onset of CNS disease manifested by a clinical phenotype of earlier hind limb paralysis compared to control and significant increased number of leukemic cells in the CNS (Figure 1A).SCD overexpression also induced CNS engraftment of another B-ALL cell line, REH, which is not naturally prone to invade the central nervous system. Mice transplanted with SCD overexpressing REH cells showed the same phenotype of earlier hind limb paralysis and accumulation of leukemic cells in the CNS as the CNS-prone 018z cells, while WT REH did not show any CNS engraftment but comparable tumor load in BM and spleen (Figure1B). To reproduce the clonal heterogeneity in SCD expression observed previously in patients' BM, we performed a competition assay transplanting SCD overexpressing cells and control cells, expressing different fluorochromes, in the same mouse in a 1:1 ratio. In the CNS, the ratio between SCD overexpressing and WT cells ranged from 2 to 20 fold. This effect was unique to the CNS and not reproducible in the other hematopoietic organs where the 1:1 ratio was maintained (Figure 1C). Moreover, SCD overexpression sensitized leukemic cells to mTOR inhibitors, suggesting a potential therapeutic option Conclusion: SCD has a role in homing and survival of leukemic cells in the CNS and may be used as early predictor of CNS relapse. This study reveals a role for SCD and fatty acid metabolism in the pathogenesis of CNS leukemia suggesting that this pathway maybe targeted for specific therapy of this devastating disease. Figure 1. Figure 1. Disclosures Halsey: Jazz Pharmaceuticals: Honoraria, Other: Support for conference attendance.

scholarly journals Hypoxia-induced CREB cooperates MMSET to modify chromatin and promote DKK1 expression in multiple myeloma

Oncogene ◽  
2021 ◽  
Author(s):  
Yinyin Xu ◽  
Jing Guo ◽  
Jing Liu ◽  
Ying Xie ◽  
Xin Li ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Combined Treatment ◽  
Hypoxia Inducible Factor ◽  
Bone Destruction ◽  
Bone Lesions ◽  
Myeloma Cells ◽  
Downstream Target ◽  
Dkk1 Expression

AbstractMyeloma cells produce excessive levels of dickkopf-1 (DKK1), which mediates the inhibition of Wnt signaling in osteoblasts, leading to multiple myeloma (MM) bone disease. Nevertheless, the precise mechanisms underlying DKK1 overexpression in myeloma remain incompletely understood. Herein, we provide evidence that hypoxia promotes DKK1 expression in myeloma cells. Under hypoxic conditions, p38 kinase phosphorylated cAMP-responsive element-binding protein (CREB) and drove its nuclear import to activate DKK1 transcription. In addition, high levels of DKK1 were associated with the presence of focal bone lesions in patients with t(4;14) MM, overexpressing the histone methyltransferase MMSET, which was identified as a downstream target gene of hypoxia-inducible factor (HIF)-1α. Furthermore, we found that CREB could recruit MMSET, leading to the stabilization of HIF-1α protein and the increased dimethylation of histone H3 at lysine 36 on the DKK1 promoter. Knockdown of CREB in myeloma cells alleviated the suppression of osteoblastogenesis by myeloma-secreted DKK1 in vitro. Combined treatment with a CREB inhibitor and the hypoxia-activated prodrug TH-302 (evofosfamide) significantly reduced MM-induced bone destruction in vivo. Taken together, our findings reveal that hypoxia and a cytogenetic abnormality regulate DKK1 expression in myeloma cells, and provide an additional rationale for the development of therapeutic strategies that interrupt DKK1 to cure MM.

scholarly journals E4F1 deficiency results in oxidative stress–mediated cell death of leukemic cells

2011 ◽  
Vol 208 (7) ◽  
pp. 1403-1417 ◽  
Author(s):  
Elodie Hatchi ◽  
Genevieve Rodier ◽  
Matthieu Lacroix ◽  
Julie Caramel ◽  
Olivier Kirsh ◽  
...  
Keyword(s):  
Cell Death ◽  
Cell Lines ◽  
Myeloid Leukemia ◽  
Tumor Regression ◽  
Fetal Liver ◽  
Leukemia Cell ◽  
Leukemic Cells ◽  
Viral Oncoprotein ◽  
Mitochondrial Defects

The multifunctional E4F1 protein was originally discovered as a target of the E1A viral oncoprotein. Growing evidence indicates that E4F1 is involved in key signaling pathways commonly deregulated during cell transformation. In this study, we investigate the influence of E4F1 on tumorigenesis. Wild-type mice injected with fetal liver cells from mice lacking CDKN2A, the gene encoding Ink4a/Arf, developed histiocytic sarcomas (HSs), a tumor originating from the monocytic/macrophagic lineage. Cre-mediated deletion of E4F1 resulted in the death of HS cells and tumor regression in vivo and extended the lifespan of recipient animals. In murine and human HS cell lines, E4F1 inactivation resulted in mitochondrial defects and increased production of reactive oxygen species (ROS) that triggered massive cell death. Notably, these defects of E4F1 depletion were observed in HS cells but not healthy primary macrophages. Short hairpin RNA–mediated depletion of E4F1 induced mitochondrial defects and ROS-mediated death in several human myeloid leukemia cell lines. E4F1 protein is overexpressed in a large subset of human acute myeloid leukemia samples. Together, these data reveal a role for E4F1 in the survival of myeloid leukemic cells and support the notion that targeting E4F1 activities might have therapeutic interest.

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.

scholarly journals Identification of Novel Lncrnas That Predict Survival in AML Patients and Modulate Leukemic Cells

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 3909-3909
Author(s):  
Sarah Grasedieck ◽  
Christoph Rueß ◽  
Nicole Pochert ◽  
Susanne Lux ◽  
Adrian Schwarzer ◽  
...  
Keyword(s):  
Conflicts Of Interest ◽  
Negative Impact ◽  
Strong Association ◽  
The Cancer Genome Atlas ◽  
High Expression ◽  
Leukemic Cells ◽  
Monocytic Differentiation ◽  
Granulocytic Differentiation

Abstract Long noncoding RNAs (lncRNAs) have complex, mainly chromatin-associated functions and their expression is highly coordinated and cell-type specific. Based on their tight regulation in normal differentiation, we set out to investigate whether lncRNAs are dysregulated in diseases where differentiation is impaired, such as in acute myeloid leukemia (AML). To identify lncRNAs that are essential for both normal hematopoiesis as well as AML maintenance, we sequenced the long polyA- and non-polyA-tagged transcriptome from successive stages of human myelopoiesis (myeloblasts, promyelocytes, metamyelocytes, and neutrophils) isolated from bone marrow of healthy donors (n=3). Applying a high-dimensional data portraying approach (OposSOM, Löffler-Wirth et al., BMC Bioinformatics, 2015), we identified functional expression modules of lncRNAs that are either positively or negatively associated with myeloid lineage commitment in our dataset. Seven out of the top15 differentiation-associated lncRNAs exhibit significant prognostic relevance in overall and event-free survival analyses of independent AML patient datasets and improve the predictive power of the current prognosis standards (cytogenetic risk/age/TP53-status). In particular, a combination of 3 transcripts, PROMYS (Promoter of Myelopoiesis, annotated as uncharacterized ncRNA LOC107985167), ANTAMY (Antagonist of Myelopoiesis, uncharacterized ncRNA LOC101927745) and LINC00677, outperformed the recently reported prognostic benefit of the LSC17high score (Ng et al, Nature, 2016) by a factor of Ø 22.7 based on concordance index score increase (Ø 4.8% vs. 0.21%). All three lncRNAs are highly conserved, expressed in 10 tested human AML cell lines as well as significantly differentially expressed in distinct cytogenetic patient subgroups of The Cancer Genome Atlas (TCGA) LAML cohort (n=171). PROMYS is downregulated in t(15;17) and t(8;21) cases, supporting its strong association with worse OS in the TCGA-LAML dataset (p=0.0001). In contrast, ANTAMY shows high expression in AML with t(8;21), and LINC00677 in NPM1+/FLT3- mutated AML patient samples with normal karyotype (CN-AML) and in core Binding factor (CBF) AMLs. Accordingly, high expression levels of both lncRNAs associate with a significantly better OS in the TCGA LAML dataset (p=0.01 and 0.02, respectively). To investigate their function in vitro, we knocked out each lncRNA individually in the human OCI/AML-5 AML cell line using CRISPR/Cas9. Loss of ANTAMY impaired proliferation (p=0.04) and increased both monocytic differentiation upon treatment with 2-0-tetradecanoylphorbol-13-acetate (TPA) (p=0.0001) and granulocytic differentiation with all-trans retinoic acid (ATRA) (p=0.0002) compared to the empty vector control. Loss of LINC00677 in OCI/AML-5 cells specifically increased granulocytic differentiation through ATRA (p=0.0002). In contrast, inactivation of PROMYS led to reduced differentiation induced by ATRA (p=0.00004) and TPA (p=0.002). Furthermore, we found that PROMYS is involved in the regulation of the Macrophage colony-stimulating factor 1 (CSF1), which is deregulated in ATRA- and TPA-induced differentiation in PROMYS knockout but not in control cells (p<0.002 and <0.00002, respectively), explaining its negative impact on differentiation. Through screening of human myelopoiesis, we identified three unexplored lncRNAs: LINC00677, PROMYS, and ANTAMY, which play a role in myeloid differentiation and have an impact on patient prognosis. Our in vitro findings confirm that ANTAMY, LINC00677, and PROMYS are active modulators of leukemic cells, which influence their proliferation, morphology, myeloid marker expression as well as apoptosis rate. These transcripts and their interaction partners add an additional layer of regulation to the understanding of differentiation and might represent previously unknown vulnerabilities of AML cells, which warrants their further investigation in vivo. 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.

RANK-RANKL Mediated Bone Destruction in B-Cell Acute Lymphoblastic Leukemia

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 908-908 ◽  
Author(s):  
Sujeetha A. Rajakumar ◽  
Eniko Papp ◽  
Ildiko Grandal ◽  
Daniele Merico ◽  
Careesa C. Liu ◽  
...  
Keyword(s):  
Bone Loss ◽  
B Cell ◽  
Lymphoblastic Leukemia ◽  
Leukemia Cell ◽  
Bone Destruction ◽  
Leukemic Cells ◽  
Leukemia Cells ◽  
Cell Acute Lymphoblastic Leukemia

Abstract Survival rates for pediatric B-Cell Acute Lymphoblastic Leukemia (B-ALL) have improved dramatically over the past 40 years approaching a current long-term survival rate of 85%. However childhood B-ALL patients continue to confront co-morbidities and their long-term consequences. For example, osteopenia and osteoporosis associated fractures are a common complication of pediatric leukemia at diagnosis, during treatment and in long-term B-ALL survivors. The STeroid-associated Osteoporosis in the Pediatric Population (STOPP) study reported that at ALL diagnosis, 16% of children and adolescents present with bone pain, vertebral compression and low vertebral Bone Mineral Density (BMD) scores, with the greatest incidence of vertebral fractures (VF) seen in the first year following diagnosis (J Clin Endocrinol Metab. 2015, 100:3408-17). Glucocorticoid treatment further elevated fracture risk in this population. These data underscore the need to identify molecular mechanism by which leukemic cells contribute to bone loss, and provide targeted therapies to limit these effects. Our laboratory previously showed that Rag2-/- p53-/- Prkdcscid/scid triple mutant (TM) and p53-/- Prkdcscid/scid double mutant (DM) mice develop spontaneous B-ALL, but only TM animals exhibit dissemination of leukemic blasts to the leptomeninges of the CNS, a poor prognosis feature observed in pediatric and adult ALL patients. We observed that TM leukemic mice also displayed fragile vertebral bones. Using comparative transcriptome analysis, we found that RANKL (Receptor Activator of the Nuclear factor-kB Ligand), a Tumor Necrosis Factor (TNF) superfamily member ligand and a key regulator of B cell and osteoclast differentiation, was expressed at greater levels in TM compared to the DM leukemia cells. RANKL binds to its receptor RANK, which is expressed in osteoclast precursor cells. RANK-RANKL interaction induces signaling in the osteoclast precursors and drives their differentiation into mature bone resorbing osteoclasts (Proc. Natl. Acad. Sci. 1999, 96:3540-3545). Upon adoptive leukemia cell transfer into immune deficient mice, RANKL+ TM but not DM cells caused decreased vertebral trabecular bone density in the recipients. Treatment with the recombinant RANKL antagonist protein Osteoprotegerin (OPG-Fc) inhibited the growth and dissemination of RANKL+TM leukemic cells and attenuated bone destruction in the recipient mice. These data suggested that TM mouse leukemia cells cause bone loss in the absence of glucocorticoid or other chemotherapy agents. We then examined the potential role of RANKL in osteoporosis associated with human B-ALL. RANKL mRNA was expressed by a majority of primary human adult and pediatric B-ALL. To determine whether primary patient B-ALL can cause bone loss, we transplanted RANKL+ human B-ALL samples of multiple cytogenetic high-risk subgroups (Complex, hypo-diploid and Mixed Lineage Leukemia (MLL) rearranged) into NOD.SCID.gC-/-(NSG) recipient mice. Micro-CT imaging and bone density measures in the xenotransplant recipients revealed extensive vertebral trabecular bone destruction. Immuno-histological analysis of the human B-ALL recipient mice demonstrated extensive osteoporotic damage of the long bones and marked RANKL protein expression in the long bones of mice harboring extensive human B-ALL cell burden compared to NSG control mice. To determine whether RANKL-RANK interaction was required for the B-ALL mediated bone destruction, cohorts of NSG mice engrafted with human B-ALL were treated with recombinant OPG-Fc compared to a matched Fc control protein. OPG-Fc treatment did not attenuate leukemia cell expansion and bone marrow burden, but despite bulky disease, the treatment conferred robust protection from bone destruction suggesting that RANKL was a critical mediator of this clinical complication. Our data demonstrate a central role of the RANK-RANKL axis in B-ALL-mediated bone disease and identify an actionable therapeutic target to reduce acute and long-term morbidity. Denosumab, an anti-RANKL antibody has been approved for the treatment of bone metastasis by solid tumors and for post-menopausal osteoporosis. Our pre-clinical studies suggest that Denosumab and other agents that inhibit the RANK-RANKL pathway may be efficacious in patients with B-ALL associated bone degeneration. Disclosures No relevant conflicts of interest to declare.

scholarly journals Eltrombopag inhibits the proliferation of leukemia cells via reduction of intracellular iron and induction of differentiation

Blood ◽  
2012 ◽  
Vol 120 (2) ◽  
pp. 386-394 ◽  
Author(s):  
Michael Roth ◽  
Britta Will ◽  
Guillermo Simkin ◽  
Swathi Narayanagari ◽  
Laura Barreyro ◽  
...  
Keyword(s):  
Leukemia Cell ◽  
Leukemic Cells ◽  
Leukemia Cells ◽  
Dependent Manner ◽  
Intracellular Iron ◽  
Division Rate ◽  
Cell Division Rate ◽  
Antileukemic Effect ◽  
Species Specific

Abstract Eltrombopag (EP) is a small-molecule, nonpeptide thrombopoietin receptor (TPO-R) agonist that has been approved recently for the treatment of thrombocytopenia in patients with chronic immune thrombocytopenic purpura. Prior studies have shown that EP stimulates megakaryopoiesis in BM cells from patients with acute myeloid leukemia and myelodysplastic syndrome, and the results also suggested that it may inhibit leukemia cell growth. In the present study, we studied the effects of EP on leukemia cell proliferation and the mechanism of its antiproliferative effects. We found that EP leads to a decreased cell division rate, a block in G1 phase of cell cycle, and increased differentiation in human and murine leukemia cells. Because EP is species specific in that it can only bind TPO-R in human and primate cells, these findings further suggested that the antileukemic effect is independent of TPO-R. We found that treatment with EP leads to a reduction in free intracellular iron in leukemic cells in a dose-dependent manner. Experimental increase of intracellular iron abrogated the antiproliferative and differentiation-inducing effects of EP, demonstrating that its antileukemic effects are mediated through modulation of intracellular iron content. Finally, determination of EP's antileukemic activity in vivo demonstrated its ability to prolong survival in 2 mouse models of leukemia.

scholarly journals Clinical MDR1 inhibitors enhance Smac-mimetic bioavailability to kill murine LSCs and improve survival in AML models

2020 ◽  
Vol 4 (20) ◽  
pp. 5062-5077
Author(s):  
Emma Morrish ◽  
Anthony Copeland ◽  
Donia M. Moujalled ◽  
Jason A. Powell ◽  
Natasha Silke ◽  
...  
Keyword(s):  
Leukemia Cell ◽  
Leukemic Cells ◽  
Human Leukemia ◽  
Combination Strategy ◽  
Multidrug Resistance Protein 1 ◽  
Leukemia Cell Lines ◽  
Therapeutic Opportunity ◽  
Smac Mimetic ◽  
Approved Drugs

Abstract The specific targeting of inhibitor of apoptosis (IAP) proteins by Smac-mimetic (SM) drugs, such as birinapant, has been tested in clinical trials of acute myeloid leukemia (AML) and certain solid cancers. Despite their promising safety profile, SMs have had variable and limited success. Using a library of more than 5700 bioactive compounds, we screened for approaches that could sensitize AML cells to birinapant and identified multidrug resistance protein 1 inhibitors (MDR1i) as a class of clinically approved drugs that can enhance the efficacy of SM therapy. Genetic or pharmacological inhibition of MDR1 increased intracellular levels of birinapant and sensitized AML cells from leukemia murine models, human leukemia cell lines, and primary AML samples to killing by birinapant. The combination of clinical MDR1 and IAP inhibitors was well tolerated in vivo and more effective against leukemic cells, compared with normal hematopoietic progenitors. Importantly, birinapant combined with third-generation MDR1i effectively killed murine leukemic stem cells (LSCs) and prolonged survival of AML-burdened mice, suggesting a therapeutic opportunity for AML. This study identified a drug combination strategy that, by efficiently killing LSCs, may have the potential to improve outcomes in patients with AML.

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.

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