Antiparasitic Effect of Stilbene and Terphenyl Compounds against Trypanosoma cruzi Parasites.

BACKGROUND: Chagas disease, also known as American trypanosomiasis, is a potentially life-threatening illness caused by the protozoan parasite Trypanosoma cruzi. No progress in the treatment of this pathology has been made since Nifurtimox was introduced more than fifty years ago, and this drug is considered very aggressive and may cause several adverse effects. This drug currently has severe limitations, including a high frequency of undesirable side effects and limited efficacy and availability, so research to discover new drugs for the treatment of Chagas disease is imperative. Many drugs available on the market are natural products as found in nature or compounds designed based on the structure and activity of these natural products. METHODS: This study evaluated the in vitro antiparasitic activity of a series of previously synthesized stilbene and terphenyl compounds in T. cruzi epimastigotes and intracellular amastigotes. The action of the most selective compounds was investigated by flow cytometric analysis to evaluate the mechanism of cell death. The ability to induce apoptosis or caspase-1 inflammasomes was assayed in macrophages infected with T. cruzi after treatment, comparing it with that of Nifurtimox. RESULTS: The stilbene ST18 was the most potent compound of the series. It was slightly less active than Nifurtimox in epimastigotes but most active in intracellular amastigotes. Compared to Nifurtimox, it was markedly less cytotoxic when tested in vitro on normal cells. ST18 was able to induce a marked increase in parasites positive for Annexin V and monodansylcadaverine. Moreover, ST18 induced the activation, in infected macrophages, of caspase-1, a conserved enzyme that plays a major role in controlling parasitemia, host survival and the onset of the adaptive immune response in Trypanosoma infection. CONCLUSIONS: The antiparasitic activity of ST18 together with its ability to activate caspase-1 in infected macrophages and its low toxicity toward normal cells makes this compound interesting for further clinical investigation.

Molecular Diagnosis of Leishmaniasis: Quantification of Parasite Load by a Real-Time PCR Assay with High Sensitivity.

Real-time PCR was developed to quantify Leishmania infantum kinetoplast DNA and optimized to achieve a sensitivity of 1 parasite/mL. For this purpose, we cloned the conserved kDNA fragment of 120 bp into competent cells and correlated them with serial dilutions of DNA extracted from reference parasite cultures calculating that a parasite cell contains approximately 36 molecules of kDNA. This assay was applied to estimate parasite load in clinical samples from visceral, cutaneous leishmaniasis patients and infected dogs and cats comparing with conventional diagnosis. The study aimed to propose a real-time PCR for the detection of Leishmania DNA from clinical samples trying to solve the diagnostic problems due to the low sensitivity of microscopic examination or the low predictive values of serology and resolve problems related to in vitro culture. The quantitative PCR assay in this study allowed detection of Leishmania DNA and quantification of considerably low parasite loads in samples that had been diagnosed negative by conventional techniques. In conclusion, this quantitative PCR can be used for the diagnosis of both human, canine and feline Leishmaniasis with high sensitivity and specificity, but also for evaluating treatment and the endpoint determination of leishmaniasis.