In Vitro, In Silico, and In Vivo Analyses of Novel Aromatic Amidines against Trypanosoma cruzi

Abstract

Five bis-arylimidamides were assayed as anti-Trypanosoma cruzi agents by in vitro, in silico, and in vivo approaches. None were considered to be pan-assay interference compounds. They had a favorable pharmacokinetic landscape and were active against trypomastigotes and intracellular forms, and in combination with benznidazole, they gave no interaction. The most selective agent (28SMB032) tested in vivo led to a 40% reduction in parasitemia (0.1 mg/kg of body weight/5 days intraperitoneally) but without mortality protection. In silico target fishing suggested DNA as the main target, but ultrastructural data did not match.

Keywords: Chagas disease; arylimidamides; experimental chemotherapy; in vivo.

FIG 1

(A) In vitro activities (EC₅₀s) and selectivity indices (SI) of 28SMB032 (A and C) and benznidazole (A and D) against intracellular forms of T. cruzi and the respective LC₅₀ upon primary cardiac cell (CC) cultures. (B to D) Light microscopy images of CC cultures infected with T. cruzi (Y strain) untreated (B) or exposed to 1.25 µM of 28SMB032 (C) and benznidazole (D). Original magnification, ×215. Values in parentheses are standard errors of the means for 95% confidence intervals.

FIG 2

Scanning electron microscopy examination of T. cruzi bloodstream trypomastigotes (Y strain). (A) Control with no compound exposure. Treatment with 0.88 μM 31DAP069 (B), 0.35 μM 36DAP015 (C), and 10.48 μM 28SMB032 (D) resulted in no morphological alteration for the parasites shown in panel B or C, but the parasites in panels D1 to D4 exhibited body retraction, and those in panels D2 and D3 exhibited a twist of the parasite body. Bars = 1 μm.

FIG 3

Ultrastructural effects of AIAs in T. cruzi bloodstream trypomastigotes (Y strain). (A1 and A2) Controls with no compound exposure display the characteristic morphology. Treatment with 0.88 μM 31DAP069 (panels B), 0.35 μM 36DAP015 (panels C), and 10.48 μM 28SMB032 (panels D) resulted in several insults, including dilatation of the flagellar pocket (black star in panels B1, C4, and D6), concentric membranar structures and myelin figures in the cytosol (black arrows in panels B2, B3, C3, and D4), disruption of the Golgi apparatus (asterisk in panels C1 and D7), an endoplasmic reticulum surrounding cytosolic structures (arrowheads in panels C2 and D3), and detachment of the nuclear (short arrow in panel D8) and plasma (short arrow in panel D6) membranes. No alterations were detected in subpellicular microtubules (thin arrows in panel D5) and on the parasite kinetoplast DNA (kDNA) (double arrows in panels D1 and D2). G, Golgi cisternae; M, mitochondria; N, nuclei; F, flagellum. Bars = 500 nm.