A Novel Protocol for the Synthesis of 1,2,4-Oxadiazoles Active against Trypanosomatids and Drug-Resistant Leukemia Cell Lines

Abstract

Cancer and parasitic diseases, such as leishmaniasis and Chagas disease, share similarities that allow the co-development of new antiproliferative agents as a strategy to quickly track the discovery of new drugs. This strategy is especially interesting regarding tropical neglected diseases, for which chemotherapeutic alternatives are extremely outdated. We designed a series of (E)-3-aryl-5-(2-aryl-vinyl)-1,2,4-oxadiazoles based on the reported antiparasitic and anticancer activities of structurally related compounds. The synthesis of such compounds led to the development of a new, fast, and efficient strategy for the construction of a 1,2,4-oxadiazole ring on a silica-supported system under microwave irradiation. One hit compound (23) was identified during the in vitro evaluation against drug-sensitive and drug-resistant chronic myeloid leukemia cell lines (EC₅₀ values ranging from 5.5 to 13.2 µM), Trypanosoma cruzi amastigotes (EC₅₀ = 2.9 µM) and Leishmania amazonensis promastigotes (EC₅₀ = 12.2 µM) and amastigotes (EC₅₀ = 13.5 µM). In silico studies indicate a correlation between the in vitro activity and the interaction with tubulin at the colchicine binding site. Furthermore, ADMET in silico predictions indicate that the compounds possess a high druggability potential due to their physicochemical, pharmacokinetic, and toxicity profiles, and for hit 23, it was identified by multiple spectroscopic approaches that this compound binds with human serum albumin (HSA) via a spontaneous ground-state association with a moderate affinity driven by entropically and enthalpically energies into subdomain IIA (site I) without significantly perturbing the secondary content of the protein.

Keywords: Leishmania amazonensis; Trypanosoma cruzi; anticancer; chagas disease; chronic myeloid leukemia; molecular docking.

Figure 1

Chemical structures of compounds with described antitrypanosomal and antimitotic activity bearing the (E)-2-aryl-vinyl (in blue) or the 3-aryl-1,2,4-oxadiazole (in red) moieties.

Figure 2

Chemical structures of (E)-3-aryl-5-(2-aryl-vinyl)-1,2,4-oxadiazoles (9–25) planned for this work.

Scheme 1

Synthesis of 1,2,4-oxadiazoles 9–25. Reaction conditions: (i) NH₂OH∙HCl (2.0 eqv), K₂CO₃ (1.5 eqv), 8-hydroxyquinoline (0.35 mol %), EtOH/H₂O 1:1, reflux, 60 min. (ii) (COCl)₂ (solvent), r.t., 30 min. (iii) benzamidoxime (27), acid chloride 29 (1.2 eqv), K₂CO₃ (1.5 eqv), DCM, r.t., 30 min., then silica gel (solvent free), microwave irradiation, 75 W, 105 °C, 5–45 min.

Scheme 2

Proposed mechanism for the cyclization reaction in the presence of silica gel to yield 1,2,4-oxadiazoles.

Figure 3

Comparison between the effects of treatment with 1,2,4-oxadiazoles against intracellular amastigote and host cell growth. (A) Compounds 14, 17, 25, and 26 against T. cruzi and LLC-MK2 cells. (B) Compounds 14, 25, and 26 against L. amazonensis and RAW 264.7 cells. * = p < 0.05; ** = p < 0.01; **** = p < 0.0001.

Figure 4

Toxicity of compounds 9–25 to murine splenocytes (BALB/c mice). Statistical significance was determined in comparison to the untreated control. * = p < 0.05; *** = p < 0.001; **** = p < 0.0001.

Figure 5

Best docking poses at CBS (α and β monomers of bovine tubulin in purple and orange, respectively) obtained for compounds 12 (A, yellow carbon atoms), 15 (B, purple carbon atoms), 23 (C, cyan carbon atoms), and 24 (D, gray carbon atoms). Hydrogen bonds are in dashed yellow lines. Hydrogen atoms have been omitted for better visualization.

Figure 6

Best docking poses at CBS (α and β monomers of bovine tubulin in purple and orange, respectively). Colchicine (magenta), 12 (yellow), 15 (purple), 23 (cyan), and 24 (gray).

Figure 7

(A) Steady-state fluorescence emission spectra (λ_exc = 280 nm) for HSA without and upon successive additions of 23 at pH = 7.4 and 310 K. (B) Stern-Volmer plots for the HSA:23 interaction in PBS. (C) Time-resolved fluorescence decays for HSA (1.0 × 10⁻⁵ M) without and with 23 (1.32 × 10⁻⁵ M) at pH = 7.4 and 296 K. The residuals were obtained by biexponential treatment with χ² values of 1.027 and 1.142 for HSA and HSA:23, respectively. (D) Double-logarithmic plots for HSA:23 in PBS. (E) Stern-Volmer plots for HSA:23 without and with the site markers warfarin, ibuprofen, or digitoxin at 310 K. (F) Van’t Hoff plot based on K_SV values for HSA:23. (G) Secondary structure content of HSA (1.0 × 10⁻⁶ M) without and with 23 (proportion 1:10). SF spectra of HSA without and upon successive additions of 23 at (H) Δλ = 15 nm for Tyr residues and (I) Δλ = 60 nm for Trp residues in PBS at room temperature. Concentrations: HSA = 1.0 × 10⁻⁵ M; 23 = 0.17, 0.33, 0.50, 0.66, 0.83, 0.99, 1.15, and 1.32 × 10⁻⁵ M.