Implication of mammalian ribosomal protein S3 in the processing of DNA damage.

A human apurinic/apyrimidinic endonuclease activity, called AP endonuclease I, is missing from or altered specifically in cells cultured from Xeroderma pigmentosum group-D individuals (XP-D cells) (Kuhnlein, U., Lee, B., Penhoet, E. E., and Linn, S. (1978) Nucleic Acids Res. 5,951-960). We have now observed that another nuclease activity, UV endonuclease III, is similarly not detected in XP-D cells and is inseparable from the AP endonuclease I activity. This activity preferentially cleaves the phosphodiester backbone of heavily ultraviolet-irradiated DNA at unknown lesions as well as at one of the phosphodiester bonds within a cyclobutane pyrimidine dimer. The nuclease activities have been purified from mouse cells to yield a peptide of M(r) = 32,000, whose sequence indicates identity with ribosomal protein S3. The nuclease activities all cross-react with immunopurified antibody directed against authentic rat ribosomal protein S3, and, upon expression in Escherichia coli of a cloned rat cDNA for ribosomal protein S3, each of the activities was recovered and was indistinguishable from those of the mammalian UV endonuclease III. Moreover, the protein expressed in E. coli and its activities cross-react with the rat protein antibody. Ribosomal protein S3 contains a potential nuclear localization signal, and the protein isolated as a nuclease also has a glycosylation pattern consistent with a nuclear localization as determined by lectin binding. The unexpected role of a ribosomal protein in DNA damage processing and the unexplained inability to detect the nuclease activities in extracts from XP-D cells are discussed.

Cytoplasmic accumulation of ditercalinium in rat hepatocytes and induction of mitochondrial damage.

Ditercalinium (NSC 335153), a 7H-pyridocarbazole dimer, is a bis-intercalating agent whose mechanism of action differs from that of other mono-intercalating compounds, such as ellipticine derivatives. After a Phase I clinical trial, where irreversible hepatotoxicity was the dose-limiting side-effect, we have reinvestigated the disposition of ditercalinium in rats after i.v. administration. Tissue distribution of the tritiumlabeled drug was studied during 5 weeks. The drug distributed very quickly into tissues, and accumulated mostly in liver and kidneys. A much slower clearance followed, with a half-life greater than 7 days. The present study raises the question of whether ditercalinium, a biscationic lipophilic agent, exerts a direct mitochondria interaction both in vitro and in vivo. Fluorescence microscopy on rat tissue cryosections, after drug administration, showed that the major cellular site of drug accumulation corresponds to mitochondria. The mitochondrial probe 3,3'-diethyloxadicarbocyanine confirmed the mitochondrial localization of ditercalinium. An identical fluorescent pattern was found in cultured rat hepatocytes. These fluorescent granulation patterns suggest mitochondrial damage. To further study cell alterations, ultrastructural changes in rat liver and kidneys were observed with selective mitochondrial damage while nuclei remained apparently normal. The same observations were made in rat hepatocyte cultures. Therefore, the accumulation of ditercalinium in tissues and, more particularly, in mitochondrial probably plays an important role in ditercalinium-induced toxicity.

Human antibodies to the antineoplastic drug elliptinium: characterization and structure-activity relationships.

Immune-mediated hemolytic disease is a phenomenon rarely encountered with cancer chemotherapeutic agents. Elliptinium, a tetracyclic ammonium compound used in breast and kidney cancer, can induce antibodies that may result in clinical hemolysis. This study reports the characterization of the elliptinium haptenic determinant by use of two different methodologies: a hemagglutination test and a radioimmunoassay. Binding of 12 analogues or derivatives of elliptinium was also studied. Good correlation between the two methods was obtained, indicating that the determinant is most likely located on the quaternary ammonium-containing ring. Furthermore, the hydrophilicity of the drug appears to be an important factor in the antibody reaction. The mechanism of the binding of elliptinium to its antibodies is discussed.

Benzodiazepine receptors on human blood platelets.

Binding studies conducted on membrane preparation from human platelets using (3H) Ro5-4864 and (3H) diazepam showed specific and saturable binding. Scatchard analysis revealed a single class of binding sites with KD = 10.8 +/- 0.9 nM and Bmax = 775 +/- 105 fmol/mg protein for (3H) Ro5-4864 and KD = 10.5 +/- 1.1 nM and Bmax = 133 +/- 19 fmol/mg for (3H) diazepam. We were unable to detect any GABA binding site on crude membrane preparation, nor did GABA enhance the binding of (3H) Ro5-4864 or (3H) diazepam. This suggests that benzodiazepine receptors are uncoupled to GABA system on human platelets. Ro15-1788, a specific antagonist for "central type" benzodiazepine (BDZ) binding sites was inactive in displacing (3H) Ro5-4864 from membrane receptors, while PK 11195 (a specific ligand for the "peripheral type" receptor) was the most potent of the drugs tested in inhibiting (3H) Ro5-4864 binding. These results indicate that human blood platelets bear "peripheral-type" BDZ receptor. Moreover, we could not detect any (3H) propyl beta carboline specific binding on platelet membranes. Results on benzodiazepine receptors on human circulating lymphocytes are also reported and similarity in pharmacological properties with platelet benzodiazepine receptors is suggested.

[Serum concentrations and factors affecting the blood bioavailability of chloramphenicol in bovine (author's transl)]. / Concentrations sériques et biodisponibilité sanguine du chloramphénicol chez les bovins.

Serum concentrations and factors affecting the blood bioavailability of chloramphenicol in bovineThe authors have compared the serum concentrations and the factors affecting blood bioavailability of chloramphenicol after intramuscular administration of canadian commercial preparations containing 500 mg/mL of antibiotic. The animals (dairy cows and heifers) received each drug (20 mg/kg) in one or two injection sites. The serum samples, analysed by colorimetric or microbiological methods, showed that considerable differences in concentration exist between the two methods. The evolution of biodisponibility factors proved identical in both cases. It appears that therapeutic levels of chloramphenicol are reached only by drug A for four to five hours. The usual dosage (2-10 mg/kg), by intramuscular route, is not sufficient to attain these active concentrations using the other drugs. However, the important variability obtained during the experiment and reflected in the standard deviation values, has not proved that drug A has a better bioavailability based on the criteria of the only microbiological analysis.