Studies on Isometamidium: The Effect of Isometamidium, Homidium and Pyrithidium on the Infectivity of Trypanosomas for Mice

Abstract

During the course of an investigation into the antitrypanosomal activity of isometamidium, some phenanthridinium compounds were administered to mice infected with Trypanosoma rhodesiense or T. congolense; trypanosomes from these mice were subinoculated into fresh mice at various times after treatment, and the effect of the compounds on infectivity was noted. In vivo/in vivo experiments of this kind have been carried out in the past by Lock (1950) with dimidium bromide and T. congolense and by Ormerod (1951) with antrycide and T. equiperdum. These authors also carried out in vitro/in vivo experiments similar to those described below, as did Hawking (1939) with suramin and T. rhodesiense. But so far no one seems to have compared the activities of compounds against different species of trypanosome by either of these methods. METHODSThe Lugala II strain of T. rhodesiense was used. This was isolated in Uganda in 1955 and received by us from EATRO in 1958. Since then it has been passaged by blood inoculation through mice, which it kills in 3 to 4 days. The strain of T. congolense was an old laboratory one which produces an acute or subacute infection in mice, killing them in 7 to 20 days (Brown, Hill & Holland, 1961).Mice were infected intraperitoneally with approximately 750,000 trypanosomes each in 0.2 ml. of citrate-saline, as in a normal therapeutic experiment. They were treated with homidium, isometamidium or pyrithidium (Prothidium) 1 or 2 days later, when there were 1 to 10 trypanosomes per high power field in the peripheral blood-stream. One mouse was left untreated, as a control, in each experiment. At 2, 24 or 48 hr after treatment blood was taken from the tail of the treated mice with a syringe and a No. 18 hypodermic needle, diluted with citrate-saline and injected intraperitoneally into previously untreated and uninfected mice. Three such mice were subinoculated from each donor. Bloods from different donors were not mixed. Three control mice were similarly infected from the untreated control donor referred to above.Beginning 4 to 5 days after subinoculation, the peripheral blood of the donor and subinoculated mice was examined three times a week for up to 28 days after subinoculation. Failure to detect trypanosomes during this period in a donor was taken as an indication that it had been cured, or in a subinoculated mouse as an indication that the trypanosomes in the inoculum had been noninfective due to their exposure to the drug before subinoculation.Some preliminary experiments with T. congolense and isometamidium failed to reveal any difference between the effects on infectivity of the intravenous and subcutaneous administration of the drug, so the subcutaneous route was used in the later work with both parasites and all three compounds.

Ethidium bromide: pharmacokinetics and efficacy against trypanosme infections in rabbits and calves

“…The highest levels of ethidium detected in blood and tissue fluid (120-180 ng/ml) following i.m. injection of the recommended curative dose (1 mg/kg) are maintained for a short period and are approximately 2 orders of magnitude less than trypanocidal levels observed in the in vitro studies of Newton (1957) and Hill (1965). Newton showed the growth inhibitory concentration of ethidium for trypanosomatid flagellates cultured in vitro to be 10-20 fig/ml and Hill showed that the ethidium concentration needed to render T. congolense uninfective to mice was 10 /ig/val for 6 h. The maximum blood concentration detected in the current work is much closer to the value of 312 ng/ml required to inhibit the motility of T. brucei in vitro.…”

“…Many instances of ethidiumresistant trypanosome strains have been reported (for example, Williamson, 1970;MacLennan & Na'lsa, 1970;Gray & Roberts, 1971) but the failure of ethidium to cure T. congolense or T. brucei infections in rabbits under the experimental conditions reported in this paper was not due to drug resistance, and extrapolation from the ethidium levels found to be trypanocidal in vitro may be misleading. The work of Newton (1957), Hill (1965) and Goodwin & Tierney (1977) all used constant ethidium concentrations in vitro. While the maximum blood ethidium concentrations present in infected rabbits are clearly sufficient to kill many of the trypanosomes present, it is equally clear that some survive.…”

The pharmacology of isometamidium

“…rhodesiense at concentrations of up to 0.1 mg/ml, probably due to the in vitro system used. Hill (1965) found that the drug was active against T. congoleme in vitro but the activity was less than that observed in vivo. This may be attributed to variations in the parasite metabohn under in vztro and in vivo conditions.…”