Non-peptidic Cruzain Inhibitors with Trypanocidal Activity Discovered by Virtual Screening and In Vitro Assay

Author(s):Show less -

Wiggers, Helton J. ^{[1, 2]} ; Rocha, Josmar R. ^[2] ; Fernandes, William B. ^{[2, 3]} ; Sesti-Costa, Renata ^[4] ; Carneiro, Zumira A. ^[4] ; Cheleski, Juliana ^[2] ; da Silva, Alberico B. F. ^[2] ; Juliano, Luiz ^[5] ; Cezari, Maria H. S. ^[5] ; Silva, Joao S. ^[4] ; McKerrow, James H. ^[3] ; Montanari, Carlos A. ^{[1, 2]}

Affiliation

^[1] Univ Fed Sao Carlos, Dept Quim, BR-13560 Sao Carlos, SP - Brazil

^[2] Univ Sao Paulo, Inst Quim Sao Carlos, Grp Quim Med IQSC USP, Sao Carlos, SP - Brazil

^[3] Univ Calif San Francisco, Dept Pathol, Ctr Discovery & Innovat Parasit Dis, San Francisco, CA 94140 - USA

^[4] Univ Sao Paulo, Fac Med Ribeirao Preto, Dept Bioquim & Imunol, BR-14049 Ribeirao Preto, SP - Brazil

^[5] Univ Fed Sao Paulo, Escola Paulista Med, Dept Biofis, Sao Paulo - Brazil

• Document type: Journal article
• Source: PLoS Neglected Tropical Diseases; v. 7, n. 8 AUG 2013.
• Web of Science Citations: 41

Abstract

A multi-step cascade strategy using integrated ligand-and target-based virtual screening methods was developed to select a small number of compounds from the ZINC database to be evaluated for trypanocidal activity. Winnowing the database to 23 selected compounds, 12 non-covalent binding cruzain inhibitors with affinity values (K-i) in the low micromolar range (3-60 mu M) acting through a competitive inhibition mechanism were identified. This mechanism has been confirmed by determining the binding mode of the cruzain inhibitor Nequimed176 through X-ray crystallographic studies. Cruzain, a validated therapeutic target for new chemotherapy for Chagas disease, also shares high similarity with the mammalian homolog cathepsin L. Because increased activity of cathepsin L is related to invasive properties and has been linked to metastatic cancer cells, cruzain inhibitors from the same library were assayed against it. Affinity values were in a similar range (4-80 mu M), yielding poor selectivity towards cruzain but raising the possibility of investigating such inhibitors for their effect on cell proliferation. In order to select the most promising enzyme inhibitors retaining trypanocidal activity for structure-activity relationship (SAR) studies, the most potent cruzain inhibitors were assayed against T. cruzi-infected cells. Two compounds were found to have trypanocidal activity. Using compound Nequimed42 as precursor, an SAR was established in which the 2-acetamidothiophene-3-carboxamide group was identified as essential for enzyme and parasite inhibition activities. The IC50 value for compound Nequimed42 acting against the trypomastigote form of the Tulahuen lacZ strain was found to be 10.6 +/- 0.1 mu M, tenfold lower than that obtained for benznidazole, which was taken as positive control. In addition, by employing the strategy of molecular simplification, a smaller compound derived from Nequimed42 with a ligand efficiency (LE) of 0.33 kcal mol(-1) atom(-1) (compound Nequimed176) is highlighted as a novel non-peptidic, non-covalent cruzain inhibitor as a trypanocidal agent candidate for optimization. (AU)

• FAPESP's process: 11/01893-3 - Optimizing trypanosomatid agents by integration of in silico, calorimetry and cell-based assays
• Grantee: Carlos Alberto Montanari
• Support type: Regular Research Grants