710 In vivo assessment of radiation induced apoptosis in follicular lymphoma patients by 99mTc-Annexin V scintigraphy

The key factors leading to transformed follicular lymphoma (t-FL) include the aberrations of epigenetic modifiers as early and driving events, especially mutations in the gene encoding for histone acetyltransferase. Therefore, reversal of this phenomenon by histone deacetylase (HDAC) inhibitors is essential for the development of new treatment strategies in t-FL. Several t-FL cell lines were treated with various doses of chidamide and subjected to cell proliferation, apoptosis and cell cycle analyses with CCK-8 assay, Annexin V/PI assay and flow cytometry, respectively. Chidamide dose-dependently inhibited cell proliferation, caused G0/G1 cycle arrest and triggered apoptosis in t-FL cells. In addition, the effects of chidamide on tumor growth were evaluated in vivo in xenograft models. RNA-seq analysis revealed gene expression alterations involving the PI3K-AKT signaling pathway might account for the mechanism underlying the antitumor activity of chidamide as a single agent in t-FL. These findings provide a basis for further clinical exploration of chidamide as a promising treatment for FL.

Download Full-text

In Vitro and In Vivo Detection of Drug-induced Apoptosis Using Annexin V-conjugated Ultrasmall Superparamagnetic Iron Oxide (USPIO): A Pilot Study

Magnetic Resonance in Medical Sciences ◽

10.2463/mrms.mp.2017-0157 ◽

2019 ◽

Vol 18 (2) ◽

pp. 142-149 ◽

Cited By ~ 1

Author(s):

Akihiro Nishie ◽

Osamu Togao ◽

Chihiro Tamura ◽

Mayumi Yamato ◽

Kazuhiro Ichikawa ◽

...

Keyword(s):

Iron Oxide ◽

Pilot Study ◽

Annexin V ◽

Superparamagnetic Iron Oxide ◽

Drug Induced ◽

Ultrasmall Superparamagnetic Iron Oxide ◽

Induced Apoptosis

Download Full-text

A Higher Frequency Administration of the Nontoxic Cycloartane-Type Triterpene Argentatin A Improved Its Anti-Tumor Activity

Molecules ◽

10.3390/molecules25081780 ◽

2020 ◽

Vol 25 (8) ◽

pp. 1780 ◽

Cited By ~ 1

Author(s):

Zaira Tavarez-Santamaría ◽

Nadia J. Jacobo-Herrera ◽

Leticia Rocha-Zavaleta ◽

Alejandro Zentella-Dehesa ◽

Beatriz del Carmen Couder-García ◽

...

Keyword(s):

Waste Product ◽

Industrial Process ◽

Annexin V ◽

Control Group ◽

Healthy Control ◽

Hct 116 ◽

Hct 116 Cells ◽

Induced Apoptosis

Parthenium argentatum (Gray), commonly known as guayule, has been used to obtain natural rubber since the beginning of the 20th century. Additionally, the so called “resin” is a waste product derived from the industrial process. The cycloartane-type triterpene Argentatin A (AA) is one of the main constituents of the industrial waste resin. In this study we evaluated the AA anticancer activity both in vitro and in vivo in the HCT116 colon cancer cells. The apoptosis promotion of AA was assessed by the annexin V/propidium iodide (PI) assay. The senescence was evaluated for SA-β-galactosidase, and PCNA was used as a marker of proliferation. Its antitumor activity was evaluated using a xenograft mouse model. The results indicated that AA-induced apoptosis in HCT-116 cells and was positively stained for SA-β-galactosidase. In the xenografted mice test, the administration of AA at the dose of 250 mg/kg three times a week for 21 days reduced tumor growth by 78.1%. A comparable tumor reduction was achieved with cisplatin at the dose of 2 mg/kg administered three times a week for 21 days. However, nude mice treated with AA did not lose weight, as they did remarkably when treated with cisplatin. Furthermore, the animals treated with AA showed similar blood profiles as the healthy control group. These data indicate the low toxicity of AA compared to that shown by cisplatin.

Download Full-text

A Novel High-Throughput Technique for Identifying Monoclonal Antibodies capable of Death Receptor Induced Apoptosis

Journal of Cell Death ◽

10.4137/jcd.s3660 ◽

2009 ◽

Vol 2 ◽

pp. JCD.S3660

Author(s):

Hang Fai Kwok ◽

Julie A. Gormley ◽

Christopher J. Scott ◽

James A. Johnston ◽

Shane A. Olwill

Keyword(s):

Cell Death ◽

High Throughput ◽

High Throughput Screening ◽

False Negative ◽

Death Receptor ◽

Annexin V ◽

Fas Receptor ◽

Induced Apoptosis

The study of death receptor family induced apoptosis has gained momentum in recent years with the knowledge that therapeutic antibodies targeting DR4 and DR5 (death receptor's 4 and 5) have proved efficacious in multiple clinical trials. The therapeutic rationale is based on targeting and amplifying a tumour tissues normal cell death programme (apoptosis). While advances in the targeting of DR4 and DR5 have been successful the search for an agonistic antibody to another family member, the Fas receptor, has proven more elusive. This is partly due to the differing in vitro and in vivo characteristics of individual antibodies. In order to induce Fas targeted cell death an antibody must be capable of binding to and trimerising the receptor. It has been shown that antibodies capable of performing this function in vivo, with the assistance of tumour associated cells, do not always induce apoptosis in vitro. As a result the use of current methodologies to detect functional antibodies in vitro may have dismissed potential therapeutic candidates ('false negative'). Here we report a novel high throughput screening technique which artificially cross-links antibodies bound to the Fas receptor. By combining this process with Annexin-V and Prodidium Iodide (PI) staining we can select for antibodies which have the potential to induce apoptosis in vivo.

Download Full-text

Anti-apoptotic effects of 3,5,3′-tri-iodothyronine in mouse hepatocytes

Journal of Endocrinology ◽

10.1677/joe.1.07061 ◽

2006 ◽

Vol 191 (2) ◽

pp. 447-458 ◽

Cited By ~ 25

Author(s):

O A Sukocheva ◽

D O Carpenter

Keyword(s):

Early Stage ◽

Dose Range ◽

Annexin V ◽

Intracellular Camp ◽

Camp Content ◽

Mouse Hepatocytes ◽

Induced Apoptosis ◽

Apoptotic Effects ◽

Intracellular Alkalinization

The present study demonstrates that 3,5,3′-tri-iodothyronine (T3) in physiological dose range inhibits tumor necrosis factor α(TNFα)/Fas-induced apoptosis in mouse hepatocytes. T3 pretreatment prevented Fas-induced early stage of apoptosis signs assessed by flow cytometry analysis of the annexin V positive cell population. T3 attenuated TNFα/Fas-induced cleavage of caspase-8 and DNA fragmentation. We found that T3 exerted its anti-apoptotic effects by mobilization of several non-genomic mechanisms independent of transcriptional activity. Inhibition of protein kinase A (PKA), extracellular signal-regulated kinase (ERK), and Na+/H+ exchanger blocked T3-dependent anti-apoptotic effects indicating an involvement of these intracellular targets into T3-induced signaling cascade. Furthermore, physiological concentrations of T3, but not reverse T3, caused increases in intracellular cAMP content and activated PKA. T3 markedly induced phosphorylation of ERK. We also detected T3-dependent intracellular alkalinization that abolished TNFα-induced acidification. PKA inhibitor KT-5720 blocked T3-induced activation of ERK and intracellular alkalinization confirming the upstream position of PKA signaling. We further detected that hepatocytes from hypothyroid mice are more sensitive to TNFα/Fas-induced apoptosis than euthyroid animals in vivo. Together, these findings imply that T3 triggers PKA- and ERK-regulated intracellular pathways capable of driving and ensuring hepatocytes survival in the presence of death receptor ligand-induced damage under chronic inflammatory conditions.

Download Full-text

NPI-2358, a Novel Vascular Disrupting Agent Blocks Growth and Angiogenesis in Multiple Myeloma Cells.

Blood ◽

10.1182/blood.v114.22.3842.3842 ◽

2009 ◽

Vol 114 (22) ◽

pp. 3842-3842

Author(s):

Dharminder Chauhan ◽

Ajita V. Singh ◽

Madhavi Bandi ◽

Noopur Raje ◽

Robert L Schlossman ◽

...

Keyword(s):

Cell Lines ◽

Annexin V ◽

Vascular Endothelial ◽

Caspase 9 ◽

Migration Assays ◽

Induced Apoptosis ◽

Rpmi 8226 ◽

Anti Tumor Activity

Abstract Abstract 3842 Poster Board III-778 Background and Rationale Vascular disrupting agents (VDAs) act via selectively disrupting established tumor vasculature and have shown remarkable clinical success as anti-cancer therapies. NPI-2358 is a novel VDA with a distinct structure and mechanism of action from other available VDAs. NPI-2358 binds to the colchicine-binding site of beta-tubulin preventing polymerization and disrupting the cytoplasmic microtubule network, thereby causing loss of vascular endothelial cytoskeletal function, and inducing cytotoxicity in cancer cells. Here, we examined the anti-angiogenic and anti-tumor activity of NPI-2358 in multiple myeloma (MM) cells using both in vitro and in vivo model systems. Material and Methods We utilized MM.1S, MM.1R, RPMI-8226, U266, and INA-6 human MM cell lines, as well as purified tumor cells from MM patients relapsing after prior anti-MM therapies. Cell viability/apoptosis assays were performed using MTT, trypan blue exclusion, and Annexin V/PI staining. Angiogenesis was measured in vitro using Matrigel capillary-like tube structure formation assays: Since human vascular endothelial cells (HUVECs) plated onto Matrigel differentiate and form capillary-like tube structures similar to in vivo neovascularization, this assay measures anti-angiogenic effects of drugs/agents. Migration assays were performed using transwell insert assays. Immunoblot analysis was performed using antibodies to caspase-8, caspase-9, caspase-3, PARP, Bcl-2, Bax, pJNK and GAPDH. Statistical significance was determined using a Student t test. Results Treatment of MM.1S, RPMI-8226, MM.1R, INA-6, and KMS-12BM with NPI-2358 for 24h induces a dose-dependent significant (P < 0.005) decrease in viability of all cell lines (IC50 range: 5-8 nM; n=3). To determine whether NPI-2358-induced decrease in viability is due to apoptosis, MM cell lines were treated with NPI-2358 for 24h; harvested, and analyzed for apoptosis using Annexin V/PI staining. A significant increase in NPI-2358-induced apoptosis was observed in all MM cell lines (% Annexin V+/PI- apoptotic cells: MM.1S, 48 ± 2.3%; MM.1R, 46.6 ± 3.1%; RPMI-8226, 61.7 ± 4.5%; and INA-6, 59.9 ± 3.2%; P < 0.05; n=3). Importantly, NPI-2358 decreased viability of freshly isolated MM cells from patients (IC50 range: 3-7 nM; P < 0.005), without affecting the viability of normal peripheral blood mononuclear cells, suggesting specific anti-MM activity and a favorable therapeutic index for NPI-2358. Examination of in vitro angiogenesis using capillary-like tube structure formation assay showed that even low doses of NPI-2358 (7 nM treatment for 12h; IC50: 20 nM at 24h) significantly decreased tubule formation in HUVECs (70-80% decrease; P < 0.05). Transwell insert assays showed a marked reduction in serum-dependent migration of NPI-2358-treated MM cells (42 ± 2.1% inhibition in NPI-2358-treated vs. control; P < 0.05). NPI-2358 at the concentrations tested (5 nM for 12h) in the migration assays did not affect survival of MM cells (> 95% viable cells). A similar anti-migration activity of NPI-2358 was noted against HUVEC cells (48 ± 1.7% decrease in migration; P < 0.05). Mechanistic studies showed that NPI-2358-induced apoptosis was associated with activation of caspase-8, caspase-9, caspase-3 and PARP. Importantly, treatment of MM.1S cells with NPI-2358 (5 nM) triggered phosphorylation of c-Jun amino-terminal kinase (JNK), a classical stress response protein, without affecting Bcl-2 family members Bax and Bcl-2. Blockade of JNK using dominant negative strategy markedly abrogated NPI-2358-induced apoptosis. Conclusion Our preclinical data provide evidence for remarkable anti-angiogenic and anti-tumor activity of NPI-2358 against MM cells, without significant toxicity in normal cells. Ongoing studies are examining in vivo anti-MM activity of NPI-2358 in animal models. Importantly, a Phase-1 study of NPI-2358 as a single agent in patients with advanced malignancies (lung, prostrate and colon cancer) has already established a favorable pharmacokinetic, pharmacodynamic and safety profile; and, a Phase-2 study of the combination of NPI-2358 and docetaxel in non-small cell lung cancer showed encouraging safety, pharmacokinetic and activity data. These findings, coupled with our preclinical studies, provide the framework for the development of NPI-2358-based novel therapies to improve patient outcome in MM. Disclosures: Chauhan: Nereus Pharmaceuticals, Inc: Consultancy. Lloyd:Nereus Pharmaceuticals, In: Employment. Palladino:Nereus Pharmaceuticals, Inc: Employment. Anderson:Nereus Pharmaceuticals, Inc: Consultancy.

Download Full-text

DR5 Knockout Mice Are Compromised in Radiation-Induced Apoptosis

Molecular and Cellular Biology ◽

10.1128/mcb.25.5.2000-2013.2005 ◽

2005 ◽

Vol 25 (5) ◽

pp. 2000-2013 ◽

Cited By ~ 77

Author(s):

Niklas Finnberg ◽

Joshua J. Gruber ◽

Peiwen Fei ◽

Dorothea Rudolph ◽

Anka Bric ◽

...

Keyword(s):

Cell Death ◽

Null Mouse ◽

Organ Specific ◽

Radiation Induced ◽

Induced Apoptosis ◽

The Brain ◽

Necrosis Factor

ABSTRACT DR5 (also called TRAIL receptor 2 and KILLER) is an apoptosis-inducing membrane receptor for tumor necrosis factor-related apoptosis-inducing ligand (also called TRAIL and Apo2 ligand). DR5 is a transcriptional target of p53, and its overexpression induces cell death in vitro. However, the in vivo biology of DR5 has remained largely unexplored. To better understand the role of DR5 in development and in adult tissues, we have created a knockout mouse lacking DR5. This mouse is viable and develops normally with the exception of having an enlarged thymus. We show that DR5 is not expressed in developing embryos but is present in the decidua and chorion early in development. DR5-null mouse embryo fibroblasts expressing E1A are resistant to treatment with TRAIL, suggesting that DR5 may be the primary proapoptotic receptor for TRAIL in the mouse. When exposed to ionizing radiation, DR5-null tissues exhibit reduced amounts of apoptosis compared to wild-type thymus, spleen, Peyer's patches, and the white matter of the brain. In the ileum, colon, and stomach, DR5 deficiency was associated with a subtle phenotype of radiation-induced cell death. These results indicate that DR5 has a limited role during embryogenesis and early stages of development but plays an organ-specific role in the response to DNA-damaging stimuli.

Download Full-text