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P Workman

Publications and source records attributed to P Workman.

At least 163 records · Page 9Linked to original sources
PubMed1989

Effects of localised tumour hyperthermia on pimonidazole (Ro 03-8799) pharmacokinetics in mice.

We have investigated the effects of localised tumour hyperthermia (LTH; 43.5 degrees C x 30 min) on the acute toxicity and pharmacokinetics of the hypoxic cell sensitizer pimonidazole (Ro 03-8799) in mice. There were three treatment groups: unrestrained controls, sham-treated and LTH treated mice. LTH had minimal effects on the acute toxicity (LD50/7d) of pimonidazole with no significant difference between the three treatment groups. Pharmacokinetic studies were carried out at the maximum tolerated dose (MTD; approximately 60% LD50) of 437 micrograms g-1 i.v. in plasma, brain and tumour. Sham tumour treatment consistently increased plasma drug concentrations compared to unrestrained controls but had minimal effects on the elimination t1/2. The AUC0-infinitive was increased by 35% and the plasma clearance decreased by 26%. By contrast, LTH had minimal effects on these parameters compared to sham treatment. Brain pimonidazole concentrations were increased in restrained mice (sham and LTH treatments) compared to unrestrained controls, but average brain/plasma ratios were similar in all three groups at between 400 and 500%. Sham tumour treatment markedly reduced peak tumour pimonidazole concentrations compared to unrestrained controls giving a 29% lower AUC0-180min. Average tumour/plasma ratios were reduced from 236 to 129%. The most important finding was that LTH further reduced pimonidazole tumour concentrations, giving a 31% lower AUC0-180 min compared to sham treated tumours. Tumour/plasma ratios for pimonidazole were reduced by 41%. Plasma exposure to the pimonidazole N-oxide metabolite, Ro 31-0313, was unaltered by LTH. The markedly reduced drug concentrations in heated tumours may be a result of hyperthermia-stimulated bioreductive drug activation.

Animals↗
PubMed1988

Inhibition of intracellular esterases by antitumour chloroethylnitrosoureas. Measurement by flow cytometry and correlation with molecular carbamoylation activity.

Antitumour chloroethylnitrosoureas (Cnus) decompose in physiological conditions yielding alkylating species and organic isocyanates. While antitumour activity is mainly attributed to the alkylation of DNA, carbamoylation of intracellular proteins by isocyanates may also have pharmacological and toxicological relevance. We previously reported a novel dynamic flow cytoenzymological assay for esterase inhibition in intact murine cells by BCNU and related isocyanates, and proposed that this might form the basis of an assay for intracellular carbamoylation. We have now examined a wide range of Cnus, isocyanates, and alkylating agents for their ability to inhibit cellular esterases. BCNU, CCNU, their derived isocyanates, and the 4-OH metabolites of CCNU exhibited potent inhibitory activity (I50 values 5.5 x 10(-5)-7.3 x 10(-4) M). Chlorozotocin and GANU were relatively inactive (I50 much greater than 10(-2) M). ACNU, TCNU and the 2-OH metabolites of CCNU exhibited intermediate activity (I50 values 1.1 x 10(-3)-2.3 x 10(-2) M). Compounds not able to form isocyanates were essentially inactive. Poor membrane permeability was also implicated in the weak activity of chlorozotocin and GANU. There was overall a good correlation between esterase inhibition and chemical carbamoylating activity, but some particular differences were identified. We concluded that flow cytoenzymological assay appears to have the potential to provide useful measurement of intracellular protein carbamoylation by existing Cnus and novel derivatives, and also offers the advantage of cell subpopulation identification for in vivo evaluation of these agents.

Animals↗
PubMed1988

Pharmacokinetics and metabolism of the mixed-function hypoxic cell sensitizer prototype RSU 1069 in mice.

RSU 1069 is a leading compound in the class of mixed-function hypoxic cell sensitizers. Possessing an alkylating aziridine function as well as a nitro group, it represents an important prototype molecule for new sensitizer development. Using a novel HPLC assay for RSU 1069 and its metabolites with a cyanopropyl column, we studied the detailed pharmacokinetics and metabolism of this drug in mice. An i.v. dose of 100 mg kg-1 produced peak plasma concentrations of about 100 micrograms ml-1. Absorption was rapid after i.p. injection but peak plasma concentrations were some three- to fourfold lower, giving an i.p. bioavailability of 55%. The elimination t1/2 was route-dependent; e.g. after 50 mg kg-1 the t1/2 was 37.2 and 22.4 min for the i.v. and i.p. routes respectively (P less than 0.001). There was also an indication of dose-dependent kinetics, with a 37% increase in elimination t1/2 when the i.p. dose was doubled from 50 to 100 mg kg-1. Oral bioavailability was low. The volume of distribution was 0.65-1.31 ml g-1 at 50 mg kg-1, but tissue penetration was limited. Brain/plasma ratios ranged from 9.3% to 66.8%, while the mean steady-state tumour/plasma ratio was 28.4%, a value considerably less than the 80%-100% ratios occurring with the neutral 2-nitroimidazole misonidazole. About 18% and 8% of a dose were excreted as the parent drug and the ring-opened hydrolysis product (RSU 1137) in the 8 h urine, indicating the likelihood of extensive metabolism via aziridine-ring removal and nitroreduction. RSU 1137 was also detected in mouse plasma and tissues and, in contrast to the aziridine ring-intact parent compound, gave tumour/plasma ratios of 100%. These studies should provide a pharmacokinetic basis for the evaluation and development of improved mixed-function sensitizers.

Animals↗
PubMed1988

When can the infusion period be safely ignored in the estimation of pharmacokinetic parameters of drugs in humans?

We describe a simple and rapid method to determine the amount by which the area under the curve (AUC) is underestimated when a drug is given by i.v. infusion, but the infusion period is ignored and the post-infusion data are analysed as if they derive from a bolus injection. Charts are provided that allow the investigator to determine the approximate underestimation for drugs following the one- and two-compartment models, and hence to decide whether it is acceptable to ignore the infusion period in a given case (e.g., underestimation less than 5% or 10%). Equations are also provided that allow the exact underestimation to be calculated, together with the true value of the AUC.

Algorithms↗
PubMed1988

The multi-dose clinical tolerance and pharmacokinetics of the combined radiosensitizers, Ro 03-8799 (pimonidazole) and SR 2508 (etanidazole).

The hypoxic cell radiosensitizers Ro 03-8799 (pimonidazole) and SR 2508 (etanidazole) have differing physico-chemical properties and clinical toxicities. The former is basic, lipophilic and produces an acute but transient central nervous system syndrome; the latter is neutral, hydrophilic and causes cumulative peripheral neuropathy. We therefore investigated the possibility of combining these agents to achieve additive radiosensitization with no enhancement of toxicity, as demonstrated in a rodent tumor model. Following a single dose study which showed a lack of interaction with respect to both toxicity and pharmacokinetics, twenty-one patients have now completed simultaneous drug administration on an escalating, multiple dose schedule. There has been no adverse acute interaction up to 0.75 g/m2 Ro 03-8799 with 2 g/m2 SR 2508 for 15 doses. At this dose-level, however, all patients experienced peripheral neuropathy. There was no adverse pharmacokinetic interaction, or perturbation of plasma pharmacokinetics between initial and final infusions. Tumor concentrations were determined in 48 biopsy samples 0-60 min after administration. Mean values normalized to a dose of 0.75 g/m2 Ro 03-8799 plus 2 g/m2 SR 2508 were 33 micrograms/g Ro 03-8799 and 74 micrograms/g SR 2508. These would be expected to produce a single-dose sensitizer enhancement ratio of around 1.5. The combination is predicted to be around 6.8 times more active than misonidazole, and superior to any single agent tested to date. The current schedules are reaching the limits of clinical tolerance, and an attempt is now being made to define the optimal regimen for use in a randomized clinical trial of the combination.

Adult↗
PubMed1988

Additivity of radiosensitization by the combination of SR 2508 (etanidazole) and Ro 03-8799 (pimonidazole) in a murine tumor system.

The nitroimidazole radiosensitizers SR 2508 and Ro 03-8799 have different dose-limiting toxicities in man and hence can be used in combination. Such therapy will be beneficial only if their radiosensitizing properties are additive, which this study sought to determine using clinically relevant radiosensitizer concentrations in the EMT6 tumor in the flanks of BALB/c mice. 240 mg/kg of each drug gave tumor concentrations (+/- 2 se) 55 min after i.v. administration of the combination of 50.4 +/- 10.6 micrograms/g (236 +/- 50 nmol/g) for SR 2508 and 39.7 +/- 9.0 micrograms/g (137 +/- 31 nmol/g) for Ro 03-8799. The radiosensitization by both agents administered both singly and in combination at 240 mg/kg and singly at 480 mg/kg was measured, giving sensitizers 30 min before 20 Gy of 250 kV X rays. Tumor response was assayed by clonogenic cell survival. SER values (with 95% confidence limits) were 1.28 (1.20-1.37) for 240 mg/kg SR 2508, 1.20 (1.10-1.30) for 240 mg/kg Ro 03-8799, 1.46 (1.33-1.59) for 240 mg/kg of both drugs in combination, 1.46 (1.38-1.55) for 480 mg/kg SR 2508 and 1.46 (1.31-1.62) for 480 mg/kg Ro 03-8799. These data confirm the additivity of radiosensitization by the two drugs administered in combination.

Animals↗
PubMed1988

Hypoxic cell radiosensitizers in the treatment of high grade gliomas: a new direction using combined Ro 03-8799 (pimonidazole) and SR 2508 (etanidazole).

The hypoxic cell radiosensitizers Ro 03-8799 (pimonidazole) and SR 2508 (etanidazole) have been evaluated for their simultaneous penetration into human brain tumors and surrounding normal tissue. Thirteen patients received a dose of 1 g of each agent, infused over a 10 minute period during neurosurgery. Samples of glioma (20), brain (10) and cerebrospinal fluid (1) were obtained at a mean time (+/- SD) of 31 +/- 18 min from the end of infusion. A 24 hr plasma time course was measured in six patients. Nitroimidazole concentrations were determined by HPLC. For a mean dose of 0.55 g/m2 of each agent, the mean tumor concentrations (+/- SD) were 17.0 +/- 12.0 micrograms/g for Ro 03-8799 and 13.5 +/- 10.9 micrograms/g for SR 2508. The tumor/plasma ratios were 279 +/- 230% and 47 +/- 34% respectively. For adjacent 'normal' brain tissue, the radiosensitizer concentrations were 29.9 +/- 13.1 micrograms/g for Ro 03-8799, and 4.0 +/- 1.7 micrograms/g for SR 2508, and the brain/plasma ratios were 430 +/- 29% and 14 +/- 8% respectively. There was a significant trend towards increasing accumulation of both agents with time, in both tumor and normal brain. Concentrations in cerebrospinal fluid were very low. Plasma pharmacokinetics for Ro 03-8799 were similar to previous experience, but for SR 2508 the terminal half-life was greater in this series by a factor of 1.3. The results confirm that Ro 03-8799 is distributed widely in the central nervous system, and demonstrate that SR 2508 can achieve high tumor concentrations when the blood-brain barrier is compromised. The concentrations achieved with the combination are indicative of a significant advantage over metronidazole, misonidazole, or either agent alone, and normalized to the therapeutic dose of 0.75 g/m2 plus 2.0 g/m2 SR 2508 are consistent with those giving additive sensitization in an in vivo mouse tumor model.

Brain Neoplasms↗
PubMed1988

Polar fluorescein derivatives as improved substrate probes for flow cytoenzymological assay of cellular esterases.

Fluorescein esters are employed in assays of cell viability, membrane permeability and esterase activity. The ester most widely used, fluorescein diacetate (FDA), has the disadvantage of rapid cellular efflux of its hydrolysis product fluorescein. This is particularly problematic for flow cytoenzymology (FCE), where fluorescence is measured in individual cells allowing identification of subpopulations differing in esterase activity and/or membrane characteristics. We present a comparison of FDA with two potentially improved substrate probes for FCE, carboxyfluorescein diacetate (CFDA) and bis(carboxyethyl)-carboxyfluorescein-tetra acetoxy methyl ester (BCECF-AM). Substrates were characterized in terms of reaction and product efflux kinetics in EMT6 mouse mammary tumour cells, together with inhibition kinetics for the carbamoylating agent BCNU. Intact viable cells were analysed by FCE and spectrofluorimetry, and the latter was also used for cell sonicates and purified esterase. CFDA and BCECF-AM enter cells and are hydrolysed more slowly than FDA. CFDA and FDA hydrolyses obey Michaelis-Menten kinetics with Km values of around 19 and 2 microM, respectively, whereas BCECF-AM hydrolysis deviates from this classical behaviour. BCNU (5 X 10(-4) M) inhibits FDA and BCECF-AM hydrolyses by approximately 50%, compared to 30% for CFDA. CFDA may be partly hydrolysed by membrane-bound esterases. Efflux half-lives were 16 min, 94 min and greater than 2 h for products of FDA, CFDA and BCECF-AM, respectively. We conclude that BCECF-AM is the optimal substrate probe for FCE. This study emphasizes the need to optimize various parameters when selecting a substrate for flow cytoenzymological assay or when loading other reporter fluorochromes into cells via lipophilic esters.

Animals↗
PubMed1988

High dose BCNU chemotherapy with autologous bone marrow transplantation and full dose radiotherapy for grade IV astrocytoma.

In a series of 22 patients, high dose BCNU (800-1,000mg m-2) with autologous bone marrow transplantation was given as the first post-surgical treatment for grade IV astrocytoma and followed by full dose radiotherapy. When compared to historical experience and matched to control patients in national studies, there appeared to be a small prolongation of survival but no increase in the proportion of long survivors. Acute myelosuppression was mild but toxicity to lung and liver was substantial and limited further dose escalation. Late bone marrow failure was seen in 4 patients. Pharmacokinetic studies were performed and suggested that the late marrow failure was due to persistence of BCNU at the time of marrow return. Despite the suggestion of a prolongation of survival this approach is not routinely recommended and a randomised trial is probably not justified.

Adult↗
PubMed1987

Novel dynamic flow cytoenzymological determination of intracellular esterase inhibition by BCNU and related isocyanates.

We present a novel dynamic flow cytoenzymological demonstration of the potent inhibition by the antitumour chloroethylnitrosourea BCNU of the intracellular hydrolysis of fluorescein diacetate by esterases of viable, intact murine and human tumour cells in vitro. The BCNU metabonate chloroethyl isocyanate and the related compound n-butyl isocyanate were also potent inhibitors. I50 values were in the range 4.2 X 10(-5)-2.0 X 10(-4) M. Generally similar quantitative results were obtained for intact cells and sonicates by conventional spectrofluorimetry, and inhibition of purified porcine liver carboxyl esterase (EC 3.1.1.1) was demonstrated. Little or no inhibitory activity was seen with the alkylating agents methyl methane-sulphonate, melphalan and nitrogen mustard. The results are consistent with carbamoylation of the esterase molecules by isocyanates, and this may provide a basis for the flow cytometric determination of intracellular carbamoylation in discrete subpopulations of heterogeneous samples.

Animals↗
PubMed1987

Heat-stimulated nitroreductive bioactivation of the 2-nitroimidazole benznidazole in vitro.

Hyperthermia enhances nitroimidazole cytotoxicity, possibly through increased nitroreductive bioactivation. Using C3H/He mouse liver microsomes and KHT tumour homogenates, we have investigated the effects of temperature (33-44 degrees) on the anaerobic nitroreduction of benznidazole (BENZO) to its amine metabolite in vitro. Microsomal nitroreductase activity was unaltered after 2 hr anaerobic incubation at 37 and 41 degrees. However at 44 degrees and 47 degrees, inactivation occurred with half-lives of 68 and 17 min respectively. At 33 degrees microsomal reduction rates were 45% lower than at 37 degrees. Reduction rates were increased by 22% at 41 compared to 37 degrees, and by 0-54% depending on substrate concentration at 44 degrees. Microsomal amine formation followed Michaelis-Menten kinetics up to 41 degrees. The 4 degrees rise from 33 to 37 degrees increased the apparent Vmax by 45% (from 0.54 to 0.98 nmol min-1 mg-1 protein) with a further increase of 32% occurring at 41 degrees. Apparent Km values were unaltered. Deviation from Michaelis-Menten kinetics was seen for amine formation at 44 degrees. The kinetics of parent drug disappearance exhibited deviation from the Michaelis-Menten relationship at all temperatures studied. KHT tumour BENZO amine formation rates were also markedly increased at elevated temperatures, e.g. by 26% at 37 degrees compared to 33 degrees and by a further 35% from 42.5 to 57.4 pmol min-1 mg-1 protein over the range 37-41 degrees. In contrast to the microsomal results, tumour reduction rates were enhanced by an average of 54% (range, 26-79%) at 44 degrees compared to 37 degrees at low as well as high substrate concentrations. These results support the hypothesis that hyperthermia-enhanced nitroimidazole cytotoxicity may be a result of increased nitroreductive bioactivation.

Animals↗