A.I. Minchinton

The clinical testing of Ro 03-8799—Pharmacokinetics, toxicology, tissue and tumor concentrations

Ro 03-8799, a lipophilic nitroimidazole with a basic side chain, has now been administered intravenously to 69 patients. The elimination half-life in plasma was 5.1 hr and the plasma concentration at 30 min was 14.8 micrograms/ml standardized to a dose of 1 g per square meter of surface area. Immediate symptoms of malaise, heat, sweating and disorientation limit the amount of the drug which may be given on any one occasion. However, a dose of 750 mg per square meter of surface area may be given combined with daily radiotherapy. Our data suggest that when given with a 20 fraction course of radiotherapy, sensitization of hypoxic cells may be achieved equal to a 10-fold increase in the dose of misonidazole above that presently permitted.

Glutathione depletion in tissues after administration of buthionine sulphoximine

Buthionine sulphoximine (BSO) an inhibitor of glutathione (GSH) biosynthesis, was administered to mice in single and repeated doses of 0.5, 1 and 5 mmol kg-1 (i.p.). The resultant pattern of GSH depletion was studied in liver, kidney, skeletal muscle and three types of murine tumor. Liver and kidney exhibited a rapid depletion to GSH levels of ca. 20% of controls after single doses of 1-5 mmol kg-1 BSO. Muscle was depleted to a similar level, but at a slower rate after a single dose. All three tumors required repeated administration of BSO over several days to obtain a similar degree of depletion to that shown in the other tissues.

A comparison of the tumour concentrations obtainable with misonidazole and Ro 03-8799.

Both Ro 03-8799 and misonidazole were administered to six patients before tumour sampling. Using similar doses, tumour concentrations of Ro 03-8799 were approximately double those of misonidazole. Examination of the data with regard to the relative sensitising efficiencies of these two compounds gives us, as a conservative estimate, a factor of 1.6 in favour of Ro 03-8799. From these observations it is predicted that, when given with radiotherapy in a 25-fraction course Ro 03-8799, is likely to be at least five times more effective as a hypoxic cell sensitiser than misonidazole.

Clinical experience with nitroimidazoles as radiosensitizers

Abstract Clinical experience at Mount Vernon now includes 248 patients who have been given misonidazole (Ro 07–0582) and 57 patients who have been given desmethylmisonidazole (Ro 05–9963). Work with Ro 03–8799 is now beginning. The promise of chemical agents to improve the results of the radiotherapy is dependent upon effective levels of radiosensitization being achieved in human tumors throughout a course of radiotherapy. We have shown that desmethylmisonidazole is well absorbed when administered orally and that good concentrations may be achieved in human tumors. A shorter half-life in plasma and much lower CSF concentrations than with misonidazole appeared to confirm the promise of reduced neurotoxicity suggested by laboratory study. Our recent work shows: (i) in an updating of the data with misonidazole a highly significant correlation of the area under the curve relating plasma concentration with time (AUC) with the incidence of neurotoxicity exists (P

Measurements of glutathione and other thiols in cells and tissues: A simplified procedure based on the HPLC separation of monobromobimane derivatives of thiols

Although there is intense interest in the role of thiols in controlling the efficiency of radiosensitizers, and in developing thiols (or pro-drugs liberating thiols) as radioprotectors, there is little information regarding the concentration of specific thiols in cells, tumors and normal tissues. Details are presented of a modified procedure using the thiol binding agent monobromobimane with separation using paired-ion reverse phase high performance liquid chromatography (HPLC). This method has been extended to include measurements of the radiosensitizer misonidazole and its desmethylated metabolite Ro 05-9963 in tissues.

Concentrations achieved in human tumors after administration of misonidazole, SR-2508 and Ro 03-8799☆

Misonidazole, SR-2508 and Ro 03-8799 have been given in sequence to patients before tumor sampling. Tumor concentrations of the three drugs have been measured and it has been possible to make prediction as to the likely advantage of the newer drugs over misonidazole. Based upon the three cases described here and 13 others given two drug combination, we suggest that in multifraction radiotherapy, both are likely to prove more than five times more efficient than misonidazole.

Enhancement of misonidazole radiosensitization by buthionine sulphoximine

The influence of glutathione (GSH) depletion on the radiation response and on the radiosensitizing efficiency of misonidazole (miso) has been studied in two types of mouse tumour and in mouse skin. Buthionine sulphoximine (BSO) has been administered in a variety of regimes, leading to a maximal depletion of GSH to 37% of control values in one tumour (CA MT) and 61% in the other (SA FA). Pretreatment with BSO did not alter the radiosensitivity of either tumour when treated with X-rays. It had a slight effect on the sensitizing efficiency of miso, corresponding to a factor less than three, which was detectable only at the highest X-ray doses used. No enhancement of miso efficiency was seen with 5 daily fractions. Prolonged administration of BSO resulted in a slight radiosensitization of mouse skin. When combined with miso the effect was very small and was only detectable at high X-ray doses. BSO however produced a marked enhancement of the acute toxicity of miso, as judged by lethality after large single doses.

A report on misonidazole in a randomized trial in locally advanced head and neck cancer

A prospective randomized trial is reported involving 97 patients with locally advanced cancer of the head and neck. Using six large fractions of radiation (3600 rad in 17 days) the addition of misonidazole (2.0gm per m2 body surface) with each fraction did not increase the local control rate at one year. It is thought that this is probably because of an inadequate tumor concentration of the drug.

Reduction of nitroimidazoles in model chemical and biological systems

Some chemical and biological properties of intermediates obtained during reduction of nitroimidazoles are discussed. These include: rate data for the decay of the nitro radical-anion, stoichiometry and absorption spectra for reduction via the radical-anion or using dithionite, stoichiometry with other reducing agents, and rate of reduction by xanthine/xanthine oxidase. Increased radiosensitization by misonidazole is seen upon prolonged pre-irradiation incubation using E. coli, enabling demonstration that a freely-diffusable metabolite is responsible for this effect. Preliminary experiments designed to extend studies of the radiobiological properties of extracellularly-added metabolites to mammalian cells and the use of liver perfusion to generate metabolites are described.

In vivo testing of a 2-nitroimidazole radiosensitizer (Ro 03–8799) using repeated administration

The radiosensitizing efficiency of a novel 2-nitroimidazole (Ro 03-8799) has been compared with that of misonidazole. Each compound was assessed by constructing dose response curves for regrowth delay using a range of drug doses. The concentration of each compound was measured in tumor homogenates with high performance liquid chromatography (HPLC). When compared on the basis of administered dose the new compound was no more efficient than misonidazole. Comparison on the basis of measured tumor concentrations showed that Ro 03-8799 was 3--4 times more efficient than misonidazole, but only at very high drug levels. Previous in vitro studies had shown a constant 10-fold difference in potency. In order to eliminate possible artifacts caused by the short half life in mice, repeated injections of Ro 03-8799 were used to maintain a constant tumor concentration for two hours before irradiation. No increase in effectiveness was observed with prolonged exposure. This is a charged compound whose distribution is pH dependent; gross tumor levels should therefore be interpreted with caution.