Abstract | BACKGROUND: Philadelphia cells are human chronic myelogenous leukemia (CML) cells that contain the BCR/ABL oncogene (a fusion of the BCR and ABL genes). Selective eradication of these cells in vitro can be achieved by combined treatment with antisense phosphorothioate oligodeoxynucleotides ([S]ODNs) specifically targeted to this oncogene (bcr/abl [S]ODNs) and a suboptimal (for use as a single agent) dose of mafosfamide (the in vitro active form of cyclophosphamide). PURPOSE: We evaluated the ability of bcr/abl antisense [S]ODNs, alone or subsequent to treatment with a single injection of cyclophosphamide, to suppress the leukemic process induced in severe combined immunodeficient (SCID) mice by Philadelphia cells (i.e., primary CML- blast crisis [CML-BC] cells). In addition, we studied potential mechanisms that might explain the efficacy of the bcr/abl antisense [S]ODN- mafosfamide combination against Philadelphia cells in vitro. METHODS: The effects of treating leukemic mice with cyclophosphamide (25 mg/kg body weight; 25% of the dose required to eradicate evidence of leukemia in SCID mice) and/or bcr/abl antisense [S]ODNs were assessed by analysis of survival, by examination of bone marrow for the presence of leukemia cells (using a colony formation assay or using coupled reverse transcription and the polymerase chain reaction to screen for bcr/abl messenger RNA), and by examination of a variety of tissues for the presence of infiltrating leukemia cells. The induction of apoptosis (a cell death program) in vitro in primary CML-BC cells following treatment with bcr/abl antisense [S]ODNs plus or minus prior treatment with mafosfamide was monitored by use of a commercial assay. Relative cellular uptake of [S]ODNs by CML-BC cells treated in vitro with or without prior treatment with mafosfamide was determined by use of confocal microscopy and flow cytometry (for fluorescent [S]ODNs) or by use of blotting techniques that employed radioactively labeled probes (for extracted, unlabeled [S]ODNs). Levels of specific proteins in treated and untreated cells were determined by use of western blotting methods. Reported P values are two-sided. RESULTS: The disease process in leukemic mice was retarded substantially by combination treatment with cyclophosphamide and specific bcr/abl antisense [S]ODNs (P < .001, relative to treatment with specific antisense [S]ODNs alone, cyclophosphamide alone, or cyclophosphamide plus nonspecific [i.e., control] antisense [S]ODNs); 50% of the mice treated with cyclophosphamide and specific antisense [S]ODNs appeared to be cured of leukemia. The combination treatment was associated with increased induction of apoptosis. In addition, cellular uptake of bcr/abl antisense [S]ODNs appeared to be increased twofold to sixfold by prior treatment with mafosfamide. This increased uptake of [S]ODNs was associated with enhanced suppression of p210bcr/abl protein levels. CONCLUSIONS AND IMPLICATIONS: Combination therapy with antisense [S]ODNs targeted to specific oncogenes and less toxic doses of anticancer drugs may represent a rational strategy to purpose for the treatment of human leukemias.
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Authors | T Skorski, M Nieborowska-Skorska, P Wlodarski, D Perrotti, G Hoser, J Kawiak, M Majewski, L Christensen, R V Iozzo, B Calabretta |
Journal | Journal of the National Cancer Institute
(J Natl Cancer Inst)
Vol. 89
Issue 2
Pg. 124-33
(Jan 15 1997)
ISSN: 0027-8874 [Print] United States |
PMID | 8998181
(Publication Type: Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, P.H.S.)
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Chemical References |
- Antineoplastic Agents
- DNA Probes
- Oligonucleotides, Antisense
- Thionucleotides
- mafosfamide
- Cyclophosphamide
- Fusion Proteins, bcr-abl
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Topics |
- Animals
- Antineoplastic Agents
(therapeutic use)
- Apoptosis
(drug effects)
- Blotting, Western
- Bone Marrow
(drug effects)
- Bone Marrow Cells
- Cyclophosphamide
(analogs & derivatives, therapeutic use)
- DNA Probes
- Flow Cytometry
- Fusion Proteins, bcr-abl
(biosynthesis, drug effects, genetics)
- Gene Expression Regulation, Neoplastic
(drug effects)
- Humans
- Leukemia, Myelogenous, Chronic, BCR-ABL Positive
(drug therapy, metabolism)
- Mice
- Mice, SCID
- Oligonucleotides, Antisense
(therapeutic use)
- Philadelphia Chromosome
- Polymerase Chain Reaction
- Survival Analysis
- Thionucleotides
(therapeutic use)
- Treatment Outcome
- Tumor Cells, Cultured
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