J Antimicrob Chemother. 2008 Feb 19 [Epub ahead of print]
A new combined flow-cytometry-based assay reveals excellent activity against Toxoplasma gondii and low toxicity of new bisphosphonates in vitro and in vivo
Shubar HM, Mayer JP, Hopfenmüller W, Liesenfeld O
Institut für Mikrobiologie und Hygiene, Charité Universtätsmedizin Berlin, Campus Benjamin Franklin, Hindenburgdamm 27, 12203 Berlin, Germany.
Objectives The aim of this study was to investigate the antiparasitic activity and toxicity of bisphosphonates using a new combined flow cytometry assay. Methods Using Toxoplasma gondii tachyzoites carrying the green-fluorescent protein (GFP), we established a new flow cytometry assay combining testing of in vitro and in vivo activity plus toxicity of newly synthesized bisphosphonates against T. gondii. Toxicity as determined by this assay was compared with toxicity as determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test. Results In vivo, therapeutic efficacy was 100% for bisphosphonates 2F, 3B, 18A, 22A and 30B at 490, 1000, 512, 44.05 and 47.6 microM concentrations, respectively. Toxicity at 100% inhibitory concentrations was 20% for 2F and 3B, 60% for 22A and 30B, and 75% for 18A. In vitro, 6 (91A, 203A, 200C, 210A, 204A and 282A) of 15 newly synthesized bisphosphonates (12 nitrogen-containing and 3 n-alkyl) inhibited parasite replication by >50% at a concentration of 100 microM. Whereas substances 91A and 282A (high efficacy) showed moderate and low toxicity (cell viability between 70% and 100%), respectively, toxicities of 203A, 200C, 210A and 204A were 70%, 65%, 80% and 70%, respectively, as determined by flow cytometry. Compounds 290A, 218A, 214A, 266A and 219A inhibited parasite replication by between 20% and 50% at a concentration of 100 microM. Conclusions Newly synthesized bisphosphonates 2F, 3B, 91A and 282A showed excellent therapeutic activity and low toxicity. These antiparasitic drugs may therefore be promising compounds for use in patients with acute and reactivated toxoplasmosis. The new flow cytometry assay allowed simultaneous determination of therapeutic efficacy and toxicity.
PMID: 18285314 [PubMed - as supplied by publisher]