The fluoroquinolone antibiotics are structurally related to nalidixic acid. Their primary antibacterial action appears to be mainly due to inhibition of DNA gyrase (DNA topoisomerase II). We determined the activity of several fluoroquinolones in vitro against two strains of Plasmodiumfalciparum, FCC, (chloroquine susceptible) and VNS (chloroquine resistant).[3lH]hypoxanthine incorporation by malarial parasites was determined at 48 and 96 h. The molarity at which each agent caused a 50% decrease in the incorporation of [3H]hypoxanthine compared with that of drug-free controls was defined as the 50% inhibitory concentration. The fluoroquinolones evaluated were amifloxacin, ciprofloxacin, enoxacin, norfloxacin, ofloxacin, and pefloxacin. Other DNA gyrase inhibitors tested were nalidixic acid, oxolinic acid, novobiocin, and coumermycin Al. Among the fluoroquinolones, ciprofloxacin had the lowest 50% inhibitory concentrations at 48 h against both chloroquinesusceptible and -resistant strains of P. falciparum, (0.26 ± 0.08) x 10-4 and (0.38 ± 0.15) x 10-4 M, respectively (mean + standard deviation). Enoxacin had the lowest 50% inhibitory concentrations against FCC1 and VNS at 96 h, 0.23 x 10-5 and (0.06 ± 0.04) x 10-5 M, respectively. With the VNS strain, fractional inhibitory concentration indexes for the combination of ciprofloxacin and tetracycline were calculated at 48 and 96 h to be 0.93 and 0.79, respectively, indicating modest additive effects. The combination of novobiocin with ciprofloxacin showed indifference in the same system. The antimalarial effects of some fluoroquinolones occur at achievable serum concentrations. Whether inhibition of DNA gyrase contributes to the antimalarial activity of the fluoroquinolones is unknown at present.

In Trypanosoma brucei gambiense, the Ca2+ binding protein calmodulin is encoded by three identical tandemly repeated genes. The transcripts of these genes consist of several RNA species similar in size. A 35-nucleotide spliced leader sequence is present at the 5' end of each mRNA but is not encoded by DNA contiguous to these genes. We have identified two different sites for the fusion of the leader to the mRNA. These results strongly support the idea that a novel, possibly discontinuous, transcription mechanism is used by these parasites.African trypanosomes can change their antigenic identity by successively expressing variant surface glycoproteins (VSGs). This variation allows the trypanosome to escape the host immune response, resulting in relapsing parasitemia. The study of nuclear gene expression in these parasitic protozoa has been limited almost exclusively to the mechanism of antigenic variation (reviewed in ref. The library was screened by the method of Benton and Davis (9) using the nick-translated electric eel calmodulin cDNA pCM109 as a probe. Baked nitrocellulose filters were preincubated for 2-6 hr at 37°C in 50% deionized formamide containing 5 x NaCl/Na Cit (lx NaCl/Na Cit = 0.15 M NaCl/0.015 M Na citrate, pH 7), 0.1% bovine serum albumin, 0.1% polyvinylpyrrolidone, 0.1% Ficoll, and 0.2% NaDodSO4. Hybridization with labeled DNA was done in the same solution for 48 hr at 37°C. Filters were then washed three times at room temperature for 30 min in 2x NaCl/Na Cit containing 0.2% NaDodSO4. Primer Extension. The synthetic oligonucleotide (5' T-C-A-A-G-T-G-G-A-T-G-T-T-A-C 3') was 5'-end-labeled with[y_32P]ATP and T4 polynucleotide kinase and was annealed for 2-4 hr at 65°C with 2 Mg of poly(A)+ RNA in 10 ,lI of 10 mM Pipes, pH 6.4/0.4 M KCl. At the end of the hybridization, the reaction mixture was diluted to 0.1 M KCl and adjusted to 50 mM Tris HCl (pH 8.0), 50 mM MgCl2, 10 mM dithiothreitol and 0.5 mM dATP, dGTP, dCTP, and TTP. Avian myeloblastosis virus reverse transcriptase (5-10 units) was added, and the incubation was continued at 43°C for 1 hr. The sample was extracted with phenol, precipitated with ethanol, and analyzed by electrophoresis on 8% polyacrylamide/7 M urea gels. For DNA sequence analysis, the extension products were cut out, extracted in 0.

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Several 1,2,2-tris(sulfonyl)hydrazines, conceived as prodrugs of 1,2-bis(sulfonyl)hydrazines, were synthesized and evaluated for antineoplastic and trypanocidal activities in mice. 1-Methyl-1,2,2-tris(methylsulfonyl)hydrazine emerged as an extremely efficacious antitrypanosomal agent, whereas 1-(2-chloroethyl)-1,2,2-tris(methylsulfonyl)hydrazine was inactive. In contrast, 1-(2-chloroethyl)-1,2,2-tris(methylsulfonyl)hydrazine displayed potent antineoplastic activity, producing several 60-day "cures" of mice bearing leukemia L1210, leukemia P388, or Sarcoma 180. Furthermore, the fact that the tris(sulfonyl) derivatives will not generate isocyanates, which contribute to the host toxicity of nitrosoureas like 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU), makes them agents of significant promise in trypanosomal and cancer chemotherapy.

Sensitivity to pentamidine of bloodstream forms and culture forms of Trypanosoma brucei brucei, strains of this subspecies, and strains of T. brucei rhodesiense characteristically differs in vitro. Analyses of transport parameters for pentamidine uptake in these organisms show differences that correspond with drug sensitivity. Long slender bloodstream forms of T. b. brucei have a high affinity for the drug and high rates of uptake at indicated by Km and Vmax values for [3H]pentamidine transport. Although pentamidine and stilbamidine resistance is associated with dyskinetoplasty, this condition does not itself confer resistance to pentamidine nor does it affect pentamidine transport. However, drug-resistant strains show lower rates for pentamidine transport as does T. b. rhodesiense, which is characteristically less sensitive to the drug. Of all the forms and strains studied, procyclic trypomastigotes were least sensitive to pentamidine and had a remarkable ability to exclude the drug.