Antiproliferative synergism of the allylamine SF 86-327 and ketoconazole on epimastigotes and amastigotes of Trypanosoma (Schizotrypanum) cruzi

Abstract

We have investigated the growth-inhibitory effects of two ergosterol biosynthesis inhibitors, the dioxolane imidazole ketoconazole and the allylamine SF 86-327, alone and in combination, on the proliferative stages of Trypanosoma (Schizotrypanum) cruzi, the causative agent of Chagas' disease. Proliferation of epimastigotes in liver infusion-tryptose medium at 28 degrees C was immediately arrested by any of these drugs at greater than or equal to 3 x 10(-5) M; cell lysis occurred 24 h later. Below that concentration, SF 86-327 at concentrations down to 1 x 10(-6) M stopped growth after 48 h. In contrast, ketoconazole slowed cell growth only moderately, but proliferation finally stopped and cell lysis occurred after 120 h at 3 x 10(-6) M. Synergistic effects could be observed when the two drugs were used in combination: the concentration of SF 86-327 required to reduce the cell growth to 25% of controls in 144 h was reduced 33-fold in the presence of 1 x 10(-6) M ketoconazole, which by itself reduced growth only by 30%. Amastigotes, proliferating in Vero cells at 37 degrees C, were much more susceptible to both drugs, but ketoconazole was definitely a more potent antiparasitic agent than the allylamine in this system: whereas the concentration of SF 86-327 required to reduce the number of infected cells to 50% of controls was 1 x 10(-7) M and that required to completely eradicate the parasite was 3 x 10(-6) M, for ketoconazole these concentrations were 1 x 10(-10) M and 1 x 10(-8) M, respectively. Again, strong synergistic effects were observed when the drugs were used in combination: the concentration of SF 86-327 required to reduce the number of infected cells to 50% of controls was 100-fold lower in the presence of 10(-11) M ketoconazole, which by itself had no effects on amastigote proliferation. The parasite was completely eradicated when the drugs were used in combination at concentrations as low as 10(-9) M. Synergy of the antiproliferative effects of the drugs on both froms of the parasite was further demonstrated by concave isobolograms. On the other hand, SF 86-327 at 10(-5) M had no effects on the proliferation of Vero cells, whereas ketoconazole at 10(-7) M reduced the proliferation of these cells by 50%.

Parasitological cure of Chagas disease: is it possible? Is it relevant?

“…Sterol biosynthesis inhibitors (SBI) have been found to be particularly useful, as endogenous sterols are essential for survival in these pathogenic organisms; these compounds are in most cases orally active, have broad spectrum of activity and very low toxicity (Ryder & Mieth 1992, Mercer 1993, Vanden Bossche 1995. T. cruzi has also an absolute requirement of specific endogenous sterols for cell viability and proliferation and is extremely sensitive to sterol biosynthesis inhibitors (SBI) in vitro (Docampo et al 1981, Beach et al 1986, Goad et al 1989, Urbina et al 1988, 1993, 1996b; thus, the sterol biosynthesis pathway in this organism is a valid chemotherapeutic target (Urbina 1997). Nevertheless, currently available SBI's, such as ketoconazole and itraconazole, have been shown to be unable to eradicate T. cruzi from experimentally infected animals or human patients (McCabe 1988, Moreira et al 1992, Brener et al 1993, although a recent report from Chile claims a high level (>50%) of parasitological cures in chronic patients treated with itraconazole (Apt et al 1998).…”

Structural Insights into Inhibition of Sterol 14α-Demethylase in the Human Pathogen Trypanosoma cruzi

“…In addition to blocking ergosterol production, antifungal azoles cause accu-mulation of toxic methylated sterol precursors, leading to pathogen growth arrest and cell death (16). The antiparasitic effect of antifungal azoles on TC has been observed by several investigators (17)(18)(19)(20)(21)(22); yet, only recently, the antifungal drug posaconazole, proven to be capable of producing a parasitological cure in a mouse model of the chronic stage of Chagas disease, has been reported to be entering clinical trials in June 2010 (23).…”

Potent Anti- Trypanosoma cruzi Activities of Oxidosqualene Cyclase Inhibitors

“…1). Terbinafine was shown to be synergistic with ketoconazole against cultures of T. cruzi (17,24). The polyene antifungal drug amphotericin B works by directly associating with ergosterol to disrupt the integrity of the cell membrane.…”