Gerald D. Bottoms

Phase I trial of piroxicam in 62 dogs bearing naturally occurring tumors

SummaryPiroxicam, a nonsteroidal antiinflammatory drug, was given to 62 dogs bearing naturally occurring tumors in a phase I clinical trial. Dose escalation was performed, with oral doses ranging from 0.5 mg/kg every 48 h (q48h) to 1.5 mg/kg q48h being tested. Dose-limiting gastromestinal irritation/ulceration occurred in all four animals that received 1.5 mg/kg q48h. The maximum tolerated dose was 1 mg/kg q48h. Subclinical renal papillary necrosis occurred in two dogs (initial dosages, 1 and 1.5 mg/kg q48h, respectively). Following dose escalation, an additional group of dogs was treated with 0.3 mg/kg piroxicam q24h per os, the accepted canine dosage prior to this trial. Inclusion of this treatment group enabled evaluation of the toxicity of and tumor response to a daily dosage regimen. No complete remissions occurred in this trial. Partial remission was documented in three of ten dogs exhibiting transitional-cell carcinoma, in three of five animals bearing squamous-cell carcinoma, in one of three dogs displaying mammary adenocarcinoma, and in the one dog that exhibited a transmissible venereal tumor. The results of this study support the additional evaluation of piroxicam in a phase II clinical trial in dogs bearing naturally occurring tumors.

Effects of Corticosterone on Phosphorylation of Rat Liver Nuclear Proteins in Vitro

Summary Glucocorticoids are known to bind to liver cytosol proteins and alter nuclear RNA synthesis. Nuclear proteins are believed to assist in the regulation of nuclear RNA synthesis. The work reported here tested the possibilities that one of the very early effects of corticosterone treatment in rats consists of altered phosphorylation of nuclear proteins and that cytosol steroid-receptor complexes are necessary for altered phosphorylation of nuclear proteins. It was found that phosphorylation of proteins in liver nuclei isolated from rats previously injected with corticosterone was significantly (p < 0.05) greater than control. Corticosterone in vitro had no effect when incubated directly with nuclei, but phosphorylation was significantly (p < 0.05) increased in nuclei previously incubated with rat liver cytosol containing 10−7 M corticosterone. Corticosterone treatment increased phosphorylation in histones and nuclear acidic proteins. Although phosphorylation of histones was generally increased, no selective phosphorylation of any specific histone fraction was observed. In contrast, corticosterone administration resulted in a significant selective increase in the phosphorylation of a nuclear acidic protein fraction. This work was supported by ACS BC-37 and NSF GB-29297 grants. The authors appreciate the technical assistance of Curt Lundgren and Rosemary Jonassen.

Distribution of Radioactivity in Different Tissues of Pigs After 3H-Hydrocortisone Injection

Summary The concentration of radioactivity in different tissues of Yorkshire pigs 45 min after an iv injection of 3H-hydrocortisone was determined. Also, chromatographic separation of the tissue supernatant fractions was conducted. Only the liver was capable of concentrating radioactivity from the blood. Radioactivity in the brain, thymus, and heart was less than in the plasma, while activity in the spleen, skeletal muscle, testes, and pituitary was the same as plasma. Molecular-sieve chromatography of the supernatant fractions from the liver, spleen, and thymus on the Sephadex G-100 indicated part of the radioactivity was present in the macromolecular fractions excluded from the gel. It is possible that a relationship between the intracellular binding of the radioactive steroid and its physiological actions may exist. The authors express sincere appreciation to Jennifer Couts for technical assistance during the course of this investigation.

Effects of cortisol-protein complexes on RNA polymerase activity in isolated pig liver and thymus nuclei.

Summary Cortisol is known to alter RNA polymerase activity in liver and thymus. It also binds to cytosol proteins in several tissues. These experiments tested the possibility that cytosol steroid-protein complexes could alter the RNA polymerase activity of homologous nuclei. Steroid-protein complexes were prepared by incubating liver and thymus minces with 10-7 M 3H-cortisol at 37°. The partially purified steroid-protein complexes were obtained by fractionation of the cytosol on Sephadex G-100. Steroid-protein complexes were incubated with intact homologous nuclei and a nuclear sediment was prepared by hypotonic lysis of nuclei and centrifugation. RNA polymerase activity was determined in the nuclear sediment. It was found that a liver cytosol steroid-protein complex significantly (p < .01) increased RNA polymerase activity when compared to free cortisol or buffer. Cytosol proteins from adrenalectomized pigs had no effect on nuclear RNA polymerase. A steroid-protein complex in the thymus significantly decreased RNA polymerase activity in thymus nuclei when compared to buffer, but the difference was no greater than that observed for free cortisol. This indicates that an association of cortisol with liver cytosol proteins may be an early event in the action of cortisol. This work was supported by a grant from the National Science Foundation (GB-29297). The authors appreciate the technical assistance of Mrs. Lois Jaques.

Intranuclear Binding of Hydrocortisone In Liver and Thymus Tissues

Summary Intranuclear uptake and binding of hydrocortisone was determined in several tissues of the rat. The first experiment was a time-uptake study designed to determine the relative distribution of 3H-hydrocortisone between the cytosol and nuclear fractions of liver and hypothalamus tissue. Maximum uptake was observed in each tissue as early as 1 min after the addition of the isotope. This was followed by a rapid migration of the radioactivity into the nucleus. Maximum nuclear uptake had occurred by 20 min. The increased nuclear uptake was accompanied by a decreased cytoplasmic radioactivity. The second experiment was designed to determine if pretreatment of rats. with hydrocortisone increased specific nuclear binding in the cerebral cortex, heart, liver, and thymus. These tissues were selected because the liver and thymus are known to respond greatly to glucocorticoids, whereas, the cerebral cortex and heart have now shown such a response. It was observed that pretreatment with hydrocortisone increased nuclear binding in the liver and thymus, but hydrocortisone treatment had no effect on the cortex or the heart. The inability of nonglucocorticoids to drive receptors into the nucleus was apparent since pretreatment with either aldosterone or testosterone did not increase the number of specific nuclear binding sites in either the liver or the thymus. The nuclear receptors in the liver and thymus were shown to be specific for hydrocortisone by the ability of non-radioactive hydrocortisone to compete for the receptor sites, whereas, aldosterone or testosterone offered no competition.