phosphorus-radioisotopes and Plasmacytoma

Based on the specificity of the Watson-Crick base pairing formation, antisense deoxyoligonucleotides have been used to inhibit the expression of oncogenes in various cancer cells. Activation of an oncogene by means of amplification leads to an increased, detectable amount of the mRNA transcript in the cytoplasm. The aim of this study was to demonstrate that cells which are expressing a particular mRNA transcript do preferentially and specifically retain the antisense probe targeting that mRNA. Using a mouse plasmacytoma cell line (MOPC315) which produces high levels of IgA heavy chain mRNA, a control mouse pre B cell line (7OZ/3B), a human mammary cell line (MCF7) which expresses the erbB2 or neu oncogene, MOPC315 cells as neu-negative controls, and antisense DNA oligonucleotides complementary to the 5' region of the mRNAs and the sense sequence, we have shown that there is a preferential, specific retention of the IgA and neu antisense sequence in MOPC315 and MCF7 cells, respectively. We have further demonstrated that this retention is time and concentration dependent with a maximum at 24 h. We conclude that cancer cells which express a particular oncogene are suitable targets for radiolabeled antisense deoxyoligonucleotides directed toward the oncogene transcript. This work and recent developments in the antisense field lead to the expectation of a new class of radiopharmaceuticals with unique specificity.

Topics: Adenosine Triphosphate; Animals; Antisense Elements (Genetics); Blotting, Northern; Breast Neoplasms; Female; Genes, erbB-2; Humans; Immunoglobulin A; Immunoglobulin Heavy Chains; In Vitro Techniques; Mice; Phosphorus Radioisotopes; Plasmacytoma; RNA, Messenger; RNA, Neoplasm; Time Factors; Transcription, Genetic; Tumor Cells, Cultured

Highly purified, but not homogeneous, samples of helix-destabilizing protein 1 from mouse myeloma contain a novel oligonucleotide-releasing DNA exonuclease. This enzyme was separated from helix-destabilizing protein 1 and obtained in highly purified form. A polypeptide of Mr 41 000 is a main constituent of the purified enzyme, and this polypeptide comigrated with the exonuclease activity during the final step of the purification, Sephacryl S-200 gel filtration where the enzyme had a native Mr of 40 000. Overall purification of enzyme activity was greater than 20 000-fold. This exonuclease releases 5'-oligonucleotides in a limited processive manner in both the 5'----3' and 3'----5' directions. Activity of the enzyme is resistant to 1 mM N-ethylmaleimide, requires a divalent cation, has an alkaline pH optimum, and degrades single-stranded DNA much faster than double-stranded DNA or RNA. The predominant oligonucleotide product with uniformly labeled substrates is (pdN)2. With 3' end labeled substrates, greater than 95% of the labeled products are (pdN)4 and (pdN)5; with 5' end labeled substrates, the main labeled product is (pdA)2. The rate of product release from 3' and 5' end labeled substrates is nearly identical at 37 degrees C. A model of the action of this enzyme and a comparison with a human placenta exonuclease [Doniger, J., & Grossman, L. (1976) J. Biol. Chem. 251, 4579-4587] are discussed.

Topics: Animals; Cell Line; DNA-Binding Proteins; Exodeoxyribonucleases; Kinetics; Mice; Molecular Weight; Phosphorus Radioisotopes; Plasmacytoma; Substrate Specificity

Topics: Cell Division; Cell Fractionation; Cell Line; Cells, Cultured; Centrifugation, Density Gradient; Cytoplasm; Electrophoresis, Polyacrylamide Gel; Nucleotides; Phosphorus Radioisotopes; Plasmacytoma; Polyribosomes; RNA, Ribosomal; Time Factors

Topics: Animals; Carbon Radioisotopes; Centrifugation, Density Gradient; Chromatography, Affinity; Chromatography, DEAE-Cellulose; Deuterium; Electrophoresis, Polyacrylamide Gel; Methods; Mice; Mice, Inbred BALB C; Neoplasm Proteins; Phosphorus Radioisotopes; Plasmacytoma; Protein Kinases; Ribosomes; RNA, Neoplasm; Tritium

Topics: Animals; Base Sequence; Binding Sites; Cell Line; Cell Nucleus; Centrifugation, Density Gradient; Chromatography, Ion Exchange; Deoxyribonucleotides; DNA Nucleotidyltransferases; DNA, Neoplasm; Escherichia coli; Guanine Nucleotides; Kinetics; Mice; Mice, Inbred BALB C; Nucleic Acid Conformation; Nucleic Acid Denaturation; Nucleic Acid Hybridization; Nucleic Acid Renaturation; Phosphorus Radioisotopes; Plasmacytoma; Pronase; RNA-Directed DNA Polymerase; RNA, Messenger; RNA, Neoplasm; Spectrophotometry, Ultraviolet

Topics: Animals; Base Sequence; Cell Line; Cytosol; Methionine; Mice; Nucleic Acid Conformation; Phosphorus Radioisotopes; Plasmacytoma; Rabbits; Ribonucleotides; RNA, Neoplasm; RNA, Transfer; Species Specificity; Valine

Topics: Animals; Autoradiography; Cells, Cultured; Centrifugation, Density Gradient; Cytoplasm; Electrophoresis, Polyacrylamide Gel; Immunoglobulin G; In Vitro Techniques; Membranes; Methods; Mice; Microsomes; Mitochondria; Nucleoproteins; Phosphorus Radioisotopes; Plasmacytoma; Puromycin; Ribonucleases; Ribosomes; RNA; RNA, Messenger; Ultracentrifugation

Topics: Amino Acids; Aminoacylation; Animals; Binding Sites; Carbon Radioisotopes; Cell Line; Cell Membrane; Centrifugation, Density Gradient; Detergents; Kinetics; Methods; Mice; Mice, Inbred BALB C; Microsomes; Neoplasm Proteins; Nitrogen; Peptide Chain Initiation, Translational; Peptide Initiation Factors; Phosphorus Radioisotopes; Plasmacytoma; Polyribosomes; Protein Biosynthesis; Ribosomes; Spectrophotometry, Ultraviolet; Time Factors

Topics: Amino Acid Sequence; Animals; Base Sequence; Cell Line; Centrifugation, Density Gradient; DNA; Electrophoresis, Polyacrylamide Gel; Genetic Code; Hydrolysis; Immunoglobulin Fragments; Immunoglobulins; Mice; Myeloma Proteins; Oligonucleotides; Phosphorus Radioisotopes; Plasmacytoma; Protein Biosynthesis; Ribonucleases; RNA, Messenger; Transcription, Genetic

Topics: Animals; Binding Sites; Cell Nucleus; Centrifugation, Density Gradient; DNA; Embryo, Mammalian; Female; Fibroblasts; Immunoglobulin Fragments; Liver; Mice; Mice, Inbred BALB C; Myeloma Proteins; Nucleic Acid Conformation; Nucleic Acid Hybridization; Pancreas; Phosphorus Radioisotopes; Plasmacytoma; Polyribosomes; Pregnancy; Ribonucleases; RNA, Messenger; RNA, Neoplasm; RNA, Ribosomal; Tritium; Uridine

Topics: Multiple Myeloma; Neoplasms, Plasma Cell; Phosphorus; Phosphorus Radioisotopes; Phosphorus, Dietary; Plasma Cells; Plasmacytoma; Radioactivity