ascorbic-acid and Agranulocytosis

Topics: Aging; Agranulocytosis; Anemia; Animals; Ascorbic Acid; Cicatrix; Collagen; Contracture; Drainage; Endothelium; Fibroblasts; Foreign Bodies; Granulation Tissue; Hematoma; Humans; Neutrophils; Protein Deficiency; Recurrence; Skin; Steroids; Stress, Mechanical; Surgical Procedures, Operative; Wound Healing; Wound Infection

Topics: Adrenal Cortex Hormones; Adrenocorticotropic Hormone; Agranulocytosis; Anemia, Hemolytic; Anemia, Hemolytic, Autoimmune; Ascorbic Acid; Autoantibodies; Autoimmune Diseases; Blood Transfusion; Chloroquine; Colitis; Colitis, Ulcerative; Drug Hypersensitivity; Drug Therapy; Hemoglobinuria; Hemoglobinuria, Paroxysmal; Humans; Internal Medicine; Leukopenia; Lupus Erythematosus, Systemic; Neutrophils; Purpura; Purpura, Thrombocytopenic; Purpura, Thrombotic Thrombocytopenic; Splenectomy; Thrombocytopenia; Thyroiditis; Toxicology; Vitamins

The use of clozapine is limited by a relatively high incidence of drug-induced agranulocytosis. Clozapine is oxidized by bone marrow cells to a reactive nitrenium ion. Although many idiosyncratic drug reactions are immune-mediated, the fact that patients with a history of clozapine-induced agranulocytosis do not immediately develop agranulocytosis on rechallenge suggests that some other factor may be responsible for the idiosyncratic nature of this reaction. The reactive nitrenium ion is very rapidly reduced back to clozapine by vitamin C, and many schizophrenic patients are vitamin C deficient. We set out to test the hypothesis that vitamin C deficiency is a major risk factor for clozapine-induced agranulocytosis using a vitamin C deficient guinea pig model. Although the vitamin C deficient guinea pigs did not develop agranulocytosis, the amount of clozapine covalent binding in these animals was less than we had previously observed in samples from rats and humans. Therefore, we studied ODS rats that also cannot synthesize vitamin C. Vitamin C deficient ODS rats also did not develop agranulocytosis, and furthermore, although covalent binding in the bone marrow was greater than that in the guinea pig, it was not increased in the vitamin C deficient ODS rats relative to ODS rats that had adequate vitamin C in their diet. Therefore, it is very unlikely that vitamin C deficiency is a major risk factor for clozapine-induced agranulocytosis.

Topics: Agranulocytosis; Animals; Antipsychotic Agents; Ascorbic Acid; Ascorbic Acid Deficiency; Clozapine; Female; Guinea Pigs; Leukocyte Count; Rats; Risk Factors

Aminoglutethimide (AG) is a first-generation aromatase inhibitor used for estrogen-dependent breast cancer. Unfortunately, its use has also been associated with agranulocytosis. We have investigated the metabolism of AG by myeloperoxidase (MPO) and the formation of an MPO protein free radical. We hypothesized that AG oxidation by MPO/H2O2 would produce an AG cation radical that, in the absence of a biochemical reductant, would lead to the oxidation of MPO. We utilized a novel anti-DMPO antibody to detect DMPO (5,5-dimethyl-1-pyrroline N-oxide) covalently bound to protein, which forms only by the reaction of DMPO with a protein free radical. We found that AG metabolism by MPO/H2O2 induced the formation of DMPO-MPO, which was inhibited by MPO inhibitors and ascorbate. Glutethimide, a congener of AG that lacks the aromatic amine, did not cause DMPO-MPO formation, indicating the necessity of oxidation of the aniline moiety in AG. When analyzed by electron spin resonance spectroscopy, we detected a phenyl radical adduct, derived from AG, which may be involved in the free radical formation on MPO. Furthermore, we also found protein-DMPO adducts in MPO-containing, intact human promyelocytic leukemia cells (HL-60). MPO was affinity-purified from HL-60 cells treated with AG/H2O2 and was found to contain DMPO. These findings were also supported by the detection of protein free radicals with electron spin resonance in the cellular cytosolic lysate. The formation of an MPO protein free radical is believed to be mediated by one of two free radical drug metabolites of AG, one of which was characterized by spin trapping with 2-methyl-2-nitrosopropane. These results are the first demonstration of MPO free-radical detection by the anti-DMPO antibody that results from drug oxidation. We propose that drug-dependent free radical formation on MPO may play a role in the origin of agranulocytosis.

Topics: Adenosine Triphosphate; Agranulocytosis; Aminoglutethimide; Aniline Compounds; Aromatase Inhibitors; Ascorbic Acid; Blotting, Western; Chromatography, Affinity; Cyclic N-Oxides; Dose-Response Relationship, Drug; Electron Spin Resonance Spectroscopy; Enzyme-Linked Immunosorbent Assay; Free Radicals; Glucose; Glutethimide; HL-60 Cells; Humans; Hydrogen Peroxide; Nitrogen Oxides; Nitroso Compounds; Peroxidase; Spectrophotometry

The use of clozapine, a unique antipsychotic drug, has been restricted due to a 1-2% incidence of drug-induced agranulocytosis. Metabolic activation of clozapine in neutrophils or stem cells could be the molecular mechanism underlying this side effect. Clozapine oxidation by human myeloperoxidase and horseradish peroxidase was evident from the disappearance of the UV absorbance at 290 nm. High performance liquid chromatography analysis revealed the formation of at least four radioactive peaks as a result of clozapine metabolism, including radioactivity coeluting with the protein. The tight association of radioactivity with the enzymatic protein was metabolism-dependent. This protein binding, which correlates with the total metabolism of clozapine, was reduced in the presence of glutathione and was absent in the presence of ascorbate. Similarly, in the presence of both reducing agents, the metabolite peaks in the high performance liquid chromatography radiogram, which are not associated with protein, disappeared. In contrast, in the presence of glutathione, two additional metabolites were found that could be isolated and identified by NMR and mass spectroscopy as clozapine glutathionyl adducts. Evidence for one-electron transfer reactions or the intermediate formation of a clozapine radical during the peroxidase-mediated metabolism of clozapine stems from the observation of thiyl and ascorbyl radicals in the presence of glutathione and ascorbate, respectively. The ascorbyl radical was detected by direct ESR spectroscopy in a peroxidase system. Its steady state concentration was significantly increased in the presence of clozapine. Glutathionyl radical formation was demonstrated by radical trapping with 5,5-dimethyl-1-pyrroline N-oxide in a peroxidase system. Again, the radical adduct concentration was significantly increased in the presence of clozapine. Similarly, when oxygen consumption was measured in peroxidase systems in the presence of glutathione or NADPH, the rate of oxygen uptake was markedly enhanced upon addition of clozapine. Thus, the data support the possibility of clozapine activation to free radical metabolites, which may cause oxidative stress or lead to adduct formation. Further, it can be concluded from these data that radical scavengers such as ascorbic acid, when coadministered with clozapine to patients, may reduce oxidative stress and protein adduct formation.

Topics: Agranulocytosis; Ascorbic Acid; Clozapine; Free Radicals; Glutathione; Horseradish Peroxidase; Humans; Peroxidase

Topics: Adult; Agranulocytosis; Ascorbic Acid; Bone Marrow Examination; Female; Humans; Infectious Mononucleosis; Serologic Tests

Topics: Adrenal Cortex Hormones; Agranulocytosis; Anti-Bacterial Agents; Ascorbic Acid; Blood Transfusion; Drug Hypersensitivity; gamma-Globulins; Humans; Morphinans

Topics: Adrenal Glands; Adrenocorticotropic Hormone; Agranulocytosis; Ascorbic Acid; Carbohydrate Metabolism; Eosinophils; Leukocyte Count