ABSTRACT
The nitric oxide (NO) prodrug JS-K, a promising anti-cancer agent, consists of a diazeniumdiolate group necessary for the release of NO as well as an arylating ring. In this study, we research the mechanism by which JS-K kills a murine erythroleukemia cell line and determine the roles of NO and arylation in the process. Our studies indicate that JS-K inhibits the PI 3-kinase/Akt and MAP kinase pathways. This correlates with the activation of the tumor suppressor FoxO3a and increased expression of various caspases, leading to apoptosis. The arylating capability of JS-K appears to be sufficient for inducing these biological effects. Overall, these data suggest that JS-K kills tumor cells by arylating and inactivating signaling molecules that block the activation of a tumor suppressor.
Subject(s)
Azo Compounds/pharmacology , Cytotoxins/pharmacology , Gene Expression Regulation, Leukemic/drug effects , Nitric Oxide Donors/pharmacology , Piperazines/pharmacology , Prodrugs/pharmacology , Animals , Caspases/genetics , Caspases/metabolism , Cell Line, Tumor , Forkhead Box Protein O3 , Forkhead Transcription Factors/agonists , Forkhead Transcription Factors/genetics , Forkhead Transcription Factors/metabolism , Mice , Mitogen-Activated Protein Kinases/antagonists & inhibitors , Mitogen-Activated Protein Kinases/genetics , Mitogen-Activated Protein Kinases/metabolism , Phosphatidylinositol 3-Kinases/genetics , Phosphatidylinositol 3-Kinases/metabolism , Phosphoinositide-3 Kinase Inhibitors , Proto-Oncogene Proteins c-akt/antagonists & inhibitors , Proto-Oncogene Proteins c-akt/genetics , Proto-Oncogene Proteins c-akt/metabolism , Signal TransductionABSTRACT
Lack of suitable mouse models for central nervous system (CNS)-associated leukemias has hindered mechanism-guided development of therapeutics. By transplanting retrovirus-transformed mouse erythroleukemia cells into syngeneic mice, we developed a new animal model of meningeal leukemia associated with rapid paralysis. Necropsy revealed massive proliferation of the leukemic cells in the bone marrow (BM) followed by pathological angiogenesis and invasion of the leukemic cells into the meninges of the CNS. Further analysis demonstrated that the erythroleukemia cells secreted high levels of VEGF and preferentially adhered in vitro to fibronectin. This unique animal model for meningeal leukemia should facilitate studies of engraftment and proliferation of leukemic cells in the BM and their invasion of the CNS as well as pre-clinical evaluation of experimental therapeutics for CNS-associated leukemias.
Subject(s)
Biomarkers, Tumor/metabolism , Central Nervous System Neoplasms/pathology , Disease Models, Animal , Leukemia, Erythroblastic, Acute/physiopathology , Leukemia, Experimental/pathology , Meningeal Neoplasms/pathology , Retroviridae/genetics , Animals , Biomarkers, Tumor/genetics , Blotting, Western , Cell Adhesion , Cell Proliferation , Central Nervous System Neoplasms/blood supply , Central Nervous System Neoplasms/etiology , Enzyme-Linked Immunosorbent Assay , Fibronectins/metabolism , Gene Expression Profiling , Integrin alpha5beta1/metabolism , Leukemia, Experimental/etiology , Meningeal Neoplasms/blood supply , Meningeal Neoplasms/etiology , Mice , Mice, Inbred BALB C , Oligonucleotide Array Sequence Analysis , Vascular Endothelial Growth Factor A/metabolismABSTRACT
The cell lines of the NCI-60 panel represent different cancer types and have been widely utilized for drug screening and molecular target identification. Screening these cell lines for envelope proteins or gene sequences related to xenotropic murine leukemia viruses (X-MLVs) revealed that one cell line, EKVX, was a candidate for production of an infectious gammaretrovirus. The presence of a retrovirus infectious to human cells was confirmed by the cell-free transmission of infection to the human prostate cancer cell line LNCaP. Amplification and sequencing of additional proviral sequences from EKVX confirmed a high degree of similarity to X-MLV. The cell line EKVX was established following passage of the original tumor cells through nude mice, providing a possible source of the X-MLV found in the EKVX cells.
Subject(s)
Adenocarcinoma/virology , Leukemia Virus, Murine/metabolism , Lung Neoplasms/virology , Animals , Base Sequence , Cell Line, Tumor , Genes, Viral/genetics , Humans , Immunoblotting , Leukemia Virus, Murine/genetics , Leukemia, Experimental/virology , Mice , Mice, Nude/virology , Molecular Sequence Data , Polymerase Chain Reaction , Retroviridae Infections/virology , Sequence Alignment , Tumor Virus Infections/virology , Viral Envelope Proteins/geneticsABSTRACT
Chronic fatigue syndrome (CFS) is a debilitating disease of unknown etiology that is estimated to affect 17 million people worldwide. Studying peripheral blood mononuclear cells (PBMCs) from CFS patients, we identified DNA from a human gammaretrovirus, xenotropic murine leukemia virus-related virus (XMRV), in 68 of 101 patients (67%) as compared to 8 of 218 (3.7%) healthy controls. Cell culture experiments revealed that patient-derived XMRV is infectious and that both cell-associated and cell-free transmission of the virus are possible. Secondary viral infections were established in uninfected primary lymphocytes and indicator cell lines after their exposure to activated PBMCs, B cells, T cells, or plasma derived from CFS patients. These findings raise the possibility that XMRV may be a contributing factor in the pathogenesis of CFS.
Subject(s)
Fatigue Syndrome, Chronic/virology , Gammaretrovirus/isolation & purification , Leukocytes, Mononuclear/virology , Retroviridae Infections/virology , Tumor Virus Infections/virology , Animals , Antibodies, Viral/blood , B-Lymphocytes/immunology , B-Lymphocytes/virology , Base Sequence , Cell Line , Cell Line, Tumor , Coculture Techniques , DNA/genetics , Gammaretrovirus/genetics , Gammaretrovirus/immunology , Gammaretrovirus/physiology , Gene Products, env/analysis , Gene Products, gag/analysis , Genome, Viral , Humans , Lymphocyte Activation , Male , Mice , Molecular Sequence Data , Prostatic Neoplasms/virology , Retroviridae Infections/epidemiology , Retroviridae Infections/transmission , T-Lymphocytes/immunology , T-Lymphocytes/virology , Tumor Virus Infections/epidemiology , Tumor Virus Infections/transmissionABSTRACT
PVC-211 murine leukemia virus (MuLV) is a neuropathogenic variant of Friend MuLV (F-MuLV) which causes a rapidly progressive spongiform neurodegenerative disease in rodents. The primary target of PVC-211 MuLV infection in the brain is the brain capillary endothelial cell (BCEC), which is resistant to F-MuLV infection. Previous studies have shown that changes in the envelope gene of PVC-211 MuLV confer BCEC tropism to the virus. However, little is known about how infection of BCECs by PVC-211 MuLV induces neurological disease. Previous results suggest that nitric oxide (NO), which has been implicated as a potential neurotoxin, is involved in PVC-211 MuLV-induced neurodegeneration. In this study, we show that expression of inducible nitric oxide synthase (iNOS), which produces NO from L-arginine, is induced in BCECs from PVC-211 MuLV-infected rats. Furthermore, elevated levels of a 32-kDa cellular protein modified by 3-nitrotyrosine, which is a hallmark of NO production, were observed in virus-infected BCECs. BCECs from rats infected with BCEC-tropic but nonneuropathogenic PVF-e5 MuLV, which is a chimeric virus between PVC-211 MuLV and F-MuLV, fail to induce either iNOS expression or elevation of tyrosine nitration of a 32-kDa protein. These results suggest that expression of iNOS and nitration of tyrosine residues of a 32-kDa protein in PVC-211 MuLV-infected BCECs may play an important role in neurological disease induction.
