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Biomedical subjects

D F Deen

Publications and source records attributed to D F Deen.

At least 109 records · Page 6Linked to original sources
PubMed1982

Decreased cytotoxicity of cis-diamminedichloroplatinum(II) by alpha-difluoromethylornithine depletion of polyamines in 9L rat brain tumor cells in vitro.

Before treatment with the cytotoxic drug cis-dichlorodiammineplatinum(II) (cis-platinum), 9L rat brain gliosarcoma cells grown in vitro were depleted of intracellular polyamines by alpha-difluoromethylornithine (DFMO; 10 mM, 48 hr), an enzyme-activated, irreversible inhibitor of the polyamine-synthetic enzyme ornithine decarboxylase. In contrast to studies that showed that the cytotoxicity of 1,3-bis (2-chloroethyl)-1-nitrosourea as measured by colony-forming efficiency is enhanced by DFMO depletion of intracellular polyamines, the cytotoxicity of cis-platinum was significantly decreased in polyamine-depleted 9L cells. At 10, 1, and 0.1% survival levels, the dose modification factors were 2.0 to 2.1. Addition of exogenous putrescine to polyamine-depleted 9L cells 24 hr before treatment with cis-platinum partially reversed this phenomenon. When 9L cells were treated either with DFMO and cis-platinum or with DFMO, putrescine, and cis-platinum for 1 hr, a time period that is too short for DFMO to affect intracellular polyamine levels, the cytotoxicity of cis-platinum was decreased, but to a significantly lesser extent than by the 48-hr DFMO pretreatment. Putrescine alone did not alter the cytotoxic effect of cis-platinum when administered either 24 or 1 hr before treatment. DFMO appears to affect cis-platinum cytotoxicity by two different mechanisms, the first mediated through polyamine depletion and the second through a direct interaction with cis-platinum.

Animals↗
PubMed1982

Quantitative dose-response relations for the cytotoxic activity of chloroethylnitrosoureas in cell culture.

A dose-response relation for the cytotoxic activity of chloroethylnitrosourea cancer chemotherapeutic agents in cell culture has been developed. Data for the activity of 1,3-bis(2-chloroethyl)-1-nitrosourea, 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea, 1-(2-chloroethyl)-3-(trans-4-methylcyclohexyl)-1-nitrosourea, and 1-(2-chloroethyl)-3-(piperidine-3,6-dion-3-yl)-1-nitrosourea against 9L rat brain tumor cells are presented. Cytotoxicity resulting from treatment schedules at different initial drug concentrations, exposure periods, and preincubation periods are correlated using a proposed dose function. Equations are presented that incorporate drug concentration, duration of exposure of cells, and the rate constant for conversion of the drug from an active intermediate into a single dose function. When compared in this way, all cell treatment schedules used were found to be equally effective in killing cells in culture. The cytotoxic activity of the four different chloroethyl-nitrosourea analogs were also found to be the same. These data demonstrate the application of quantitative dose-response relations to the activity of chloroethylnitrosoureas in cell culture and provide new insight into the mechanism of action and structure-activity relationships of these compounds.

Animals↗
PubMed1982

Factors that influence initiation and growth of 9L rat brain gliosarcoma multicellular spheroids.

This report details optimum conditions for initiating and growing 9L rat brain gliosarcoma multicellular spheroids. Best results were achieved when 0.5 to 2.0 x 10(6) cells were seeded initially into agarose (0.5 to 1.0%)-coated Petri dishes. Spheroids initiated in this manner appeared to grow in spinner culture at a rate slightly faster than spheroids initiated and grown in spinner culture. An optimum growth rate was obtained for individual spheroids grown in the well of cluster dishes coated with 0.5 to 0.75% agarose in growth medium containing 10% newborn calf serum and 0.9 g glucose per liter. Three methods for dissociation of spheroids at both room temperature and 37 degrees have been developed. The "race track" method of fractional dissociation produced a constant and similar number of cells for each disaggregation interval, and the plating efficiency of cells in each fraction was similar.

Animals↗
PubMed1981

Subhypothalamic grafts of human pituitary adenomas in total-body irradiated rats.

Human pituitary adenomas proliferate neither in cell culture nor in athymic "nude" mice. We propose that one or several of the humoral factors necessary for the growth of pituitary adenomas is missing in these experimental environments. The purpose of our experiments was to examine the possible influence of the hypothalamus in supporting cellular proliferation, and thus adenoma growth. Fragments from four human pituitary adenomas (three pituitary prolactinomas; one ACTH-secreting adenoma) were transplanted into the pituitary fossa of total-body irradiated, hypophysectomized rats. The rats were killed after two weeks and perfused with a mixture of formalin and India ink. Histologic examination of serial sagittal sections through the pituitary fossa and the adjacent brain showed: vascularization of the grafts from the pituitary stalk and from the scar tissue in the sphenoid bone; survival of some adenomas; and numerous mitoses in an ACTH-secreting specimen obtained from a patient who had Cushing's disease. We conclude from these experiments that as yet unidentified hypothalamus factors are essential for the growth of certain types of pituitary adenomas.

Adenoma↗
PubMed1981

The 9L rat brain tumor: description and application of an animal model.

Animal models allow determination of tumor response to anticancer agents under various experimental conditions. The chemically induced 9L rat brian tumor has been developed as both in vivo and in vitro models. Animal survival, clonogenic cell survival, and tumor growth delay provide means to measure the effectiveness of treatment modalities in this tumor model. Monolayer cultures, multicellular spheroid cultures, brain tumors, and flank tumors have been used to study the influence of different biological entities of the 9L model on the response to treatment with radiation and/or BCNU (1,3-bis (2-chloroethyl)-1-nitrosourea).

Animals↗
PubMed1981

Sensitization of 9L rat brain gliosarcoma cells to 1,3-bis(2-chloroethyl)-1-nitrosourea by alpha-difluoromethylornithine, an ornithine decarboxylase inhibitor.

alpha-Difluoromethylornithine, a known inhibitor of polyamine biosynthesis, significantly enhanced the cytotoxic effect of 1,3-bis(2-chloroethyl)-1-nitrosourea, a cell cycle-nonspecific agent, in 9L rat brain gliosarcoma cells in vitro. Administered as a single agent, alpha-difluoromethylornithine was not cytotoxic to 9L cells and, compared to untreated control cells, caused no perturbation of cell cycle kinetics. alpha-Difluoromethylornithine-induced depletion of intracellular polyamine levels appears to have caused the observed sensitization of 9L cells to 1,3-bis(2-chloroethyl)-1-nitrosourea. Restoration of intracellular polyamine levels by the addition of exogenous putrescine to treated 9L cells reversed this phenomenon.

Animals↗
PubMed1981

Cytokinetic and cytotoxic effects of antimetabolites on 9L rat brain tumor cells in vitro.

Treatment of exponentially growing 9L monolayer cells with graded concentrations of methotrexate or 5-fluorouracil (5-FUra) resulted in approximately 95% cell kill, corresponding to the percentage of cells in this system labeled by autoradiography (98%). Additional cell kill could be obtained if 5-FUra was administered at a high concentration (7.7 x 10(-4) M) and could be partly reversed in the presence of 8.2 x 10(-5) M uridine. When cell kill was measured as a function of time of exposure, a 3-hr exposure to 2.2 x 10(-4) M methotrexate or 7.7 x 10(-6) or 10(-4) M 5-FUra killed 30 to 50% of the cell population, which is equivalent to the percentage of the S-phase cells of the system. With both agents, additional cell kill was offset by division of unaffected G2 cells and a drug-induced block of G1 cells at the G1-S border, and it depended on long drug exposure. Nontoxic concentrations of 5-FUra could synchronize and increase the fraction of cells in the drug-sensitive S phase.

Animals↗
PubMed1981

Differential response to elutriated 9L cells to treatment with 1,3-bis(2-chloroethyl)-1-nitrosourea.

Earlier cytokinetic studies using 9L monolayer culture cells showed that cells began accumulating in the G2 phase (4C position in the DNA distribution) within 30 hr after administration of 1,3-bis(2-chloroethyl)-1-nitrosourea (3 micrograms/ml). Most of these cells reached the G2 phase at 48 hr and remained there for another 24 hr. Similar studies using 9L spheroid culture cells show that 9L cells also began to accumulate at the 4C DNA position 30 hr after administration of 1,3-bis(2-chloroethyl)-1-nitrosourea (3 micrograms/ml); however, only one-half of the cells accumulated in the 4C peak during the first 48 hr after treatment, while the other half remained at the 2C position. Those that remained at the 2C peak might correspond to the G0 cells that exist in 9L spheroids. When one-half the cells were in 2C and one-half were in 4C, the spheroids were dissociated using an enzyme cocktail and sorted using centrifugal elutriation. Cells from each elutriated fraction were analyzed for DNA content using flow cytometry and for viability using a colony-forming efficiency assay. Results indicated that in spheroids, 1,3-bis(2-chloroethyl)-1-nitrosourea (3 micrograms/ml) is more effective against noncycling cells and that repopulation is more vigorous in G2-arrested cells, which presumably were cycling at the time of treatment.

Animals↗
PubMed1981

Potentials and limitations of a clonogenic cell assay for human brain tumors.

Single cells from biopsies of ten malignant gliomas were treated with 1,3-bis (2-chloroethyl)-1-nitrosourea (BCNU) in vitro, and tumor cell survival was compared to patient response to nitrosourea therapy. The limitations of the methods and their interpretations and future prospects for clonogenic cell analyses are discussed.

Adult↗
PubMed1980

An analysis of the interaction between two nitrosourea compounds and X-radiation in rat brain tumour cells.

Experimental measurements have shown that both BCNU [1,3-bis(2-chloroethyl)-1-nitrosourea] and CCNU [1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea] are toxic in rat 9L brain tumour cells and also sensitize these cells to the action of ionizing radiation. The interaction of BCNU and CCNU with radiation has been interpreted using a recently developed extension of the molecular theory of cell survival. The experimental results are shown to be compatible with the mathematical equations predicted by the model and the analysis indicates that the sensitizing effect is caused by a synergistic interaction between sublethal damage caused by the nitrosourea compound and the radiation at the molecular level. The analysis of the dependence of the interaction on the time between nitrosourea treatment and radiation indicates that the optimal interaction occurs with a 5 hour interval.

Animals↗
PubMed1980

Analysis of clonogenic human brain tumour cells: preliminary results of tumour sensitivity testing with BCNU.

Biopsies from 6 patients with glioblastoma multiforme were disaggregated and single cells were treated in vitro with various concentrations of BCNU and plated for cell survival. One patient's cells were sensitive to BCNU in vitro; after a single dose of BCNU her brain scan reverted to normal and she was clinically well. Five tumours demonstrated resistance in vitro. Three of these tumours progressed during the first course of chemotherapy with a nitrosourea and the patients died at 2 1/2, 4 and 8 1/2 months after operation. Two patients who showed dramatic responses to radiation therapy were considered unchanged after the first course of nitrosourea therapy (although one demonstrated tumour enlargement on brain scan). The correlation of in vitro testing of tumour cell sensitivity with actual patient response is encouraging enough to warrant further work to determine whether such tests should weigh in decisions on patient therapy.

Aged↗
PubMed1980

Effect of feeder cell-released substances on the survival of clonogenic 9L cells after treatment with antimetabolites.

Exponentially growing monolayer cultures of 9L rat brain tumor cells were treated with either 5-fluorouracil or methotrexate. The surviving fraction of cells was determined by a colony formation assay. After 5-fluorouracil treatment, 2 to 5 X 10(5) feeder cells were required to maximize surviving fractions for each drug concentration and to generate a biphasic dose-response curve. If only 1 X 10(4) feeder cells were used, the dose-response curve was steep. Uridine added to the dishes containing 1 X 10(4) feeder cells restored the biphasic shape, while uridine and thymidine added to the dishes yielded the same curve obtained with 2 X 10(5) feeder cells. After methotrexate treatment, the surviving fraction of cells was dependent on feeder cell number when the medium in the dishes was supplemented with dialyzed fetal bovine serum, but it was not dependent on feeder cell number when nondialyzed fetal bovine serum was used. Biphasic dose-response curves were generated from methotrexate-treated cells plated in medium supplemented with either dialyzed or nondialyzed serum, but the drug was more toxic to cells plated in medium containing dialyzed serum. This additional toxicity could be reduced if either thymidine or N-5-formyltetrahydrofolate were added to the dishes and eliminated if 1 X 10(4) feeders were added. These results suggest that any cell culture system used to evaluate antimetabolites should be optimized for possible feeder cell and serum effects.

Animals↗