scholarly journals Pharmacogenomics of intracellular methotrexate polyglutamates in patients’ leukemia cells in vivo

2020 ◽  
Vol 130 (12) ◽  
pp. 6600-6615
Author(s):  
Elixabet Lopez-Lopez ◽  
Robert J. Autry ◽  
Colton Smith ◽  
Wenjian Yang ◽  
Steven W. Paugh ◽  
...  
Keyword(s):  
Leukemia Cells ◽  
Methotrexate Polyglutamates

scholarly journals New insights into methotrexate accumulation in leukemia cells in vivo

2021 ◽  
Vol 8 (1) ◽  
pp. 1865086
Author(s):  
Elixabet Lopez-Lopez ◽  
William E. Evans
Keyword(s):  
Leukemia Cells

scholarly journals Proliferation of human malignant hematopoietic cells in immunodeficient mice: suppression by antibody to pluripotent K-562 leukemia cells involves direct cytolysis and effector cells

Blood ◽  
1983 ◽  
Vol 61 (6) ◽  
pp. 1045-1053 ◽  
Author(s):  
BB Lozzio ◽  
EA Machado ◽  
J Mitchell ◽  
CB Lozzio ◽  
CJ Wust ◽  
...  
Keyword(s):  
Gamma Globulin ◽  
Hematopoietic Cells ◽  
Leukemia Cells ◽  
Control Group ◽  
Fab Fragment ◽  
Lymphoma Cells ◽  
Effector Cells ◽  
Immunodeficient Mice ◽  
Immune Gamma Globulin

Abstract Six human hematopoetic cell lines were successfully heterotransplanted into athymic (nude) and asplenic-athymic (lasat) neonatal mice. The tumors arising from leukemia and lymphoma cells could then be serially transplanted into adult nude mice. Seven days after the fourth serial mouse passage, each mouse was treated with goat immune gamma globulin against K-562 cells. One control group was treated similarly, but with nonimmune (normal) gamma globulin, while another control group was not treated. The goat gamma globulin was not toxic for nude and lasat mice, and the immune, but not the normal, gamma globulin suppressed local subcutaneous growth of myelosarcomas, lymphosarcomas, and Burkitt lymphoma cells. On the other hand, the growth of lung, breast, and prostatic carcinomas and a melanoma of human origin were not altered by the immune gamma globulin. Since suppression of cell growth occurred equally well in decomplemented mice, a complement-mediated cytotoxicity apparently cannot be considered as responsible for the abrogation. The Fab fragment of the immunoglobulin did not suppress the growth of the myelosarcomas. We conclude that antibody suppression of the in vivo proliferation was specific for malignant hematopoietic cells and that the Fc portion of IgG is necessary for in vivo cytolysis of leukemia cells. The most probable mechanisms are direct antibody cytolysis and antibody-dependent macrophage-mediated cytotoxicity.

scholarly journals In Vivo Imaging Enables High Resolution Preclinical Trials on Patients’ Leukemia Cells Growing in Mice

PLoS ONE ◽  
2012 ◽  
Vol 7 (12) ◽  
pp. e52798 ◽  
Author(s):  
Nadia Terziyska ◽  
Catarina Castro Alves ◽  
Volker Groiss ◽  
Katja Schneider ◽  
Katarina Farkasova ◽  
...  
Keyword(s):  
High Resolution ◽  
In Vivo Imaging ◽  
Leukemia Cells ◽  
Preclinical Trials

Involvement of oxygen radicals in the differentiation of rat myelomonocytic leukemia cells in vitro and in vivo

1994 ◽  
Vol 18 (3) ◽  
pp. 199-203 ◽  
Author(s):  
Masaobu Kobayashi ◽  
Jun Nishihara ◽  
Yoshihiro Fujii ◽  
Hiroshi Maeda ◽  
Masuo Hosokawa ◽  
...  
Keyword(s):  
Oxygen Radicals ◽  
Leukemia Cells ◽  
Myelomonocytic Leukemia

scholarly journals In Vivo RNA Interference Screening Identifies a Leukemia-Specific Dependence on Integrin Beta 3 Signaling

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 758-758
Author(s):  
◽  
Fatima Al-Shahrour ◽  
Kimberly A. Hartwell ◽  
Lisa P Chu ◽  
Jaras Marcus ◽  
...  
Keyword(s):  
Rna Interference ◽  
Cell Growth ◽  
Leukemia Cell ◽  
Leukemia Cells ◽  
Growth And Survival ◽  
Integrin Beta 3 ◽  
Shrna Screen ◽  
Primary Leukemia

Abstract Abstract 758 Primary leukemia stem cells (LSCs) reside in an in vivo microenvironment that supports the growth and survival of malignant cells. Despite the increasing understanding of the importance of niche interactions and primary cell biology in leukemia, many studies continue to focus on cell autonomous processes in artificial model systems. The majority of strategies to-date that attempt to define therapeutic targets in leukemia have relied on screening cell lines in culture; new strategies should incorporate the use of primary disease within a physiologic niche. Using a primary murine MLL-AF9 acute myeloid leukemia (AML) model highly enriched for LSCs, we performed an in vivo short hairpin RNA (shRNA) screen to identify novel genes that are essential for leukemia growth and survival. LSCs infected with pools of shRNA lentivirus were transplanted and grown in recipient mice for 2 weeks, after which bone marrow and spleen cells were isolated. Massively parallel sequencing of infected LSCs isolated before and after transplant was used to quantify the changes in shRNA representation over time. Our in vivo screens were highly sensitive, robust, and reproducible and identified a number of positive controls including genes required for MLL-AF9 transformation (Ctnnb1, Mef2c, Ccna1), genes universally required for cell survival (Ube2j2, Utp18), and genes required in other AML models (Myb, Pbx1, Hmgb3). In our primary and validation screens, multiple shRNAs targeting Integrin Beta 3 (Itgb3) were consistently depleted by more than 20-fold over two weeks in vivo. Follow up studies using RNA interference (RNAi) and Itgb3−/− mice identified Itgb3 as essential for murine leukemia cells growth and transformation in vivo, and loss of Itgb3 conferred a statistically significant survival advantage to recipient mice. Importantly, neither Itgb3 knockdown or genetic loss impaired normal hematopoietic stem and progenitor cell (HSPC) function in 16 week multilineage reconstitution assays. We further identified Itgav as the heterodimeric partner of Itgb3 in our model, and found that knockdown of Itgav inhibited leukemia cell growth in vivo. Consistent the therapeutic aims or our study, flow cytometry on primary human AML samples revealed ITGAV/ITGB3 heterodimer expression. To functionally assess the importance of gene expression in a human system, we performed another RNAi screen on M9 leukemia cells, primary human cord blood CD34+ cells transduced with MLL-ENL that are capable of growing in vitro or in a xenotransplant model in vivo. We found that ITGB3 loss inhibited M9 cell growth in vivo, but not in vitro, consistent with the importance of ITGB3 in a physiologic microenvironment. We explored the signaling pathways downstream of Itgb3 using an additional in vivo, unbiased shRNA screen and identified Syk as a critical mediator of Itgb3 activity in leukemia. Syk knockdown by RNAi inhibited leukemia cell growth in vivo; downregulation of Itgb3 expression resulted in decreased levels of Syk phosphorylation; and expression of an activated form of Syk, TEL-SYK, rescued the effects of Itgb3 knockdown on leukemia cell growth in vivo. To understand cellular processes controlled by Itgb3, we performed gene expression studies and found that, in leukemia cells, Itgb3 knockdown induced differentiation and inhibited multiple previously published LSC transcriptional programs. We confirmed these results using primary leukemia cell histology and a model system of leukemia differentiation. Finally, addition of a small molecule Syk inhibitor, R406, to primary cells co-cultured with bone marrow stroma caused a dose-dependent decrease in leukemia cell growth. Our results establish the significance of the Itgb3 signaling pathway, including Syk, as a potential therapeutic target in AML, and demonstrate the utility of in vivo RNA interference screens. Disclosures: Armstrong: Epizyme: Consultancy.

scholarly journals Isoform Selective HDAC8 Inhibitor (OCH3) Shows Potency in Selectively Targeting Primary AML Cells

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 2944-2944
Author(s):  
Amal Mechaal ◽  
Amudha Ganapathy ◽  
Dolores Mahmud ◽  
Taha Y Taha ◽  
Rajeev Ranjan ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
Ex Vivo ◽  
Vehicle Control ◽  
Leukemia Cells ◽  
Apoptotic Cells ◽  
Histone H4 ◽  
Inhibitory Effects ◽  
Growth Inhibitory ◽  
Cd34 Cells

Abstract The treatment outcomes for patients diagnosed with acute myeloid leukemia (AML) are still dismal. Recent advances in understanding AML indicate that the lack of efficacy is primarily due to non-specificity of currently used chemotherapeutics targeting both leukemic stem/progenitor cells (LSC) and normal hematopoietic stem cells (HSC). Thus, a critical barrier is the identification of innovative therapies that selectively target LSC. Histone deacetylase 8 (HDAC8) has been shown to enhance p53 protein deacetylation, which results in inactivation of p53, promoting LSC survival. We hypothesize that enzymatic/non-enzymatic role of HDAC8 is critical for LSC survival but not for HSCs. Then, we characterized our two tetrahydroisoquinoline (TIQ)-based selective HDAC8 inhibitors (HDAC8i) BIP and OCH3 for growth inhibition, apoptosis, activation of caspase 3, integrity of mitochondrial membrane potential (MMP), and acetylation of histone H4 in human leukemia cell lines. The growth inhibitory effects observed in cell lines were validated using bone marrow (BM) or peripheral blood (PB) cells from AML patients. Colony forming cell (CFC) assays were performed using AML BM/PB cells treated with OCH3 or BIP. OCH3 and BIP were also tested for hematotoxicity using normal CB CD34+ cells. Furthermore, we compared class I HDAC isoform engagement in human normal cord blood (CB) CD34+ cells and in SET-2 leukemia cells using our novel photoreactive probe TH1143. In CD34+ cells, TH1143 had higher level of engagement for HDAC1 and 2, whereas engagement of HDAC3 and 8 was minimal. In SET-2 cells, HDAC3 and HDAC8 displayed relatively higher engagement with TH1143 indicating HDAC engagement is likely cell type specific. The biological efficacies of OCH3 at 50uM and BIP at 25uM were noted to exert >50% growth inhibition in KG1 and in K562 leukemia cells. Both OCH3 and BIP significantly increased the number of apoptotic cells and there was an enhanced active caspase-3 activity. Furthermore, OCH3 and BIP treated cells displayed lower red/green ratio in comparison to control, indicative of poor MMP and depolarization to induce apoptosis (Table 1.a). OCH3 and BIP were further validated by using BM/PB cells from AML patients showing growth inhibition. This was also accompanied by increase in apoptotic cells by OCH3 and BIP. In contrast to BIP, OCH3 spared CB CD34+ cells as demonstrated by notably lower growth inhibition, apoptotic cells vs control when compared with primary AML cells from patients. Both OCH3 and BIP displayed minimal inhibition of CFU growth in CD34+ cells. However, HDAC8i induced significant CFU growth inhibition in primary AML samples suggesting that HDAC8i spares normal CFU progenitors but not leukemia progenitors (Table 1.b). Notably, both BIP and OCH3 lack ability to exert acetylation of histone H4, unlike broad spectrum HDAC inhibitor TSA (MFI with OCH3=0.96±0.03, BIP=0.77±0, TSA =1.63±0.15) which is consistent with isoform selectivity of OCH3 and BIP. The leukemia growth inhibitory effects at LSC level was demonstrated using ex vivo OCH3 treated AML patient derived BM/PB cells transplantation in humanized immunodeficient NSGS mice. After 10 to 12 weeks of transplantation mice receiving untreated AML cells had 7.73±2.18% while with OCH3 treatment mice had 4.84±1.37% human CD34+ leukemia cells, a 38% reduction in CD34+ leukemia cells, despite only a single ex vivo exposure to OCH3. Furthermore, in a second model, NSGS humanized mice were transplanted (IV) with primary leukemia cells from AML patients and after 4 weeks injected (IP) with OCH3 or vehicle control. After 12 weeks of transplantation in this second model human primary AML cell burden was 5.74±1.31% (OCH3) and 18.13±12.76% (vehicle control), while mice transplanted with normal CD34+ cells treated similarly with OCH3 or vehicle control displayed no detectable inhibition of human myeloid cell chimerism (OCH3:12.28 ± 3.31% vs vehicle control: 17.92±11.96%). Taken together, our data indicate that HDAC8 isoform inhibitor, OCH3 displayed significant inhibition of primary AML patient derived leukemia cells growth in vitro and in vivo in contrast to normal CD34+ cells. Selective inhibition of HDAC8 is sufficient to cause growth inhibition in primary AML progenitors including LSCs in vivo while sparing normal HSCs thus offer opportunities for further development of HDAC8i as new experimental therapeutics in AML. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

scholarly journals Natural killer-mediated lysis of normal and malignant target cells, and its regulation by monocytes.

1985 ◽  
Vol 162 (2) ◽  
pp. 472-486 ◽  
Author(s):  
K Oshimi ◽  
Y Oshimi ◽  
M Satake ◽  
H Mizoguchi
Keyword(s):  
Nk Cells ◽  
Natural Killer ◽  
B Lymphocytes ◽  
Leukemia Cells ◽  
Target Cells ◽  
Large Granular Lymphocytes ◽  
Percoll Density Gradient ◽  
Granular Lymphocytes ◽  
Density Gradient Sedimentation

After depletion of monocytes, natural killer (NK) cells were partially purified from peripheral blood by Percoll density gradient sedimentation. The NK cells were then cultured for 1 d and assayed for their cytotoxicity against various types of normal and malignant target cells. All types of target cells tested were found to be susceptible to NK cells. The susceptible targets were autologous T and B lymphocytes, mitogen-induced T and B blasts, monocytes, large granular lymphocytes, autologous or allogeneic lymphoma and leukemia cells isolated from patients, and cultured cell lines, including those resistant to interferon-activated lymphocytes. Such a broad spectrum of cytotoxicity was demonstrated in 1 d of culture, and freshly prepared NK cells were not cytotoxic, or, if anything, were less cytotoxic. Monocytes and their supernatants, added throughout the course of culture, markedly inhibited the development of their cytotoxicity. These results may suggest that, although NK cells having ability to lyse autologous normal and malignant target cells are present in vivo, their lytic activity is regulated by coexisting monocytes.

scholarly journals Acute lymphoblastic leukemia cells that survive combination chemotherapy in vivo remain sensitive to allogeneic immune effects

2011 ◽  
Vol 35 (6) ◽  
pp. 800-807 ◽  
Author(s):  
Johan Jansson ◽  
Yu-Chiao Hsu ◽  
Igor I. Kuzin ◽  
Andrew Campbell ◽  
Craig A. Mullen
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Combination Chemotherapy ◽  
Lymphoblastic Leukemia ◽  
Leukemia Cells
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