Development of leishmanicidal drugs based on the selective inhibition of the enzyme dihydroorotate dehydrogenase

• Grant number: 12/25075-0
• Support Opportunities: Regular Research Grants
• Duration: April 01, 2013 - September 30, 2015
• Field of knowledge: Biological Sciences - Biophysics - Molecular Biophysics
• Principal Investigator: Maria Cristina Nonato
• Grantee: Maria Cristina Nonato
• Host Institution: Faculdade de Ciências Farmacêuticas de Ribeirão Preto (FCFRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil
• Associated researchers: Flavio da Silva Emery ; Renata Fonseca Vianna Lopez

Abstract

Leishmaniases are considered neglected tropical diseases due to the lack of investment from public and private sectors in the development of new methods of diagnosis and adequate therapies. Facing this tough reality, the Academy has had an important role in the identification and validation of macromolecular targets, as well as, in the search of bioactive molecules as prototypes for the development of new leishmanicidal therapies. Within this context, our laboratory has been working on the characterization and evaluation of the enzyme dihydroorotate dehydrogenase as a potential therapeutic target for tripanossomatid-related diseases, in special, the leishmaniases. During the last ten years, the enzyme DHODH from Leishmania major (LmDHODH) was deeply characterized through the application of molecular biology techniques, biochemistry, spectroscopy and X-ray crystallography. The enzyme dihydroorotate dehydrogenase (DHODH; E.C, 1.3.3.1) is a flavoenzyme that catalyzes the oxidation of (S)-dihydroorotate to orotate, the only redox reaction in the de novo pyrimidine biosynthesis pathway and has been deeply exploited in the development of new therapeutic strategies against antiproliferative and antiparasitic diseases. Our structural and functional evaluation of LmDHODH has allowed us to explore a more rational in vitro and in silico techniques in the search for potent and selective ligands against the parasites enzyme. Two molecules, named LCPSP3027 e LCPVL898, have been identified as LmDHODH inhibitors with IC50 of 1,66 ± 0,03 µM e 2,82 ± 0,02 µM, respectively. A careful search in the available literature, as well as against the Patent Depository performed by USP Patent Office revealed to be a new class of molecules not yet described as active against parasites. Those compounds have also displayed in vitro leishmanicidal activity against the promastigote form of Leishmania (V.) braziliensis (DL50 de 2.1 µM for LCPSP3027 e DL50 de 2.8 µM para LCPVL898), and the assays performed on macrophages demonstrated to be noncytotoxic molecules. Reduced levels of infectivity have also been observed when evaluating the amastigote form of Leishmania in presence of both compounds. At this point, we intend to move on, initially by developing a protocol for the synthesis of LCPSP3027 e LCPVL898 and their analogues, followed by the characterization of their mechanism of inhibition and the development of a pharmaceutical formulation that will further be used for our pre-clinical assays. The development of the present study, by using a multidisciplinary approach, will allow us to learn about the drug discovery pipeline, as well as, can be considered an important step in the search for an efficient therapy against different types of Leishmania. (AU)

Scientific publications

(9)

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

MAETANI, MICAH; KATO, NOBUTAKA; JABOR, VALQUIRIA A. P.; CALIL, FELIPE A.; NONATO, MARIA CRISTINA; SCHERER, CHRISTINA A.; SCHREIBER, STUART L.. Discovery of Antimalarial Azetidine-2-carbonitriles That Inhibit P-falciparum Dihydroorotate Dehydrogenase. ACS Medicinal Chemistry Letters, v. 8, n. 4, p. 438-442, APR 2017. (12/25075-0)

GARCIA REIS, RENATA ALMEIDA; LORENZATO, JR., EDER; SILVA, VALERIA CRISTINA; NONATO, MARIA CRISTINA. Recombinant production, crystallization and crystal structure determination of dihydroorotate dehydrogenase from Leishmania (Viannia) braziliensis. ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY COMMUNICATIONS, v. 71, n. 5, SI, p. 547-552, MAY 2015. (12/25075-0, 14/01257-8, 11/23504-9)

LEWIS, TIMOTHY A.; SYKES, DAVID B.; LAW, JASON M.; MUNOZ, BENITO; RUSTIGUEL, JOANE K.; NONATO, MARIA CRISTINA; SCADDEN, DAVID T.; SCHREIBER, STUART L.. Development of ML390: A Human DHODH Inhibitor That Induces Differentiation in Acute Myeloid Leukemia. ACS Medicinal Chemistry Letters, v. 7, n. 12, p. 1112-1117, DEC 2016. (12/25075-0)

REIS, RENATA A. G.; FERREIRA, PATRICIA; MEDINA, MILAGROS; NONATO, M. CRISTINA. The mechanistic study of Leishmania major dihydro-orotate dehydrogenase based on steady- and pre-steady-state kinetic analysis. Biochemical Journal, v. 473, n. 5, p. 651-660, MAR 1 2016. (12/25075-0, 11/23504-9)

AZEREDO, LUIS FELIPE S. P.; COUTINHO, JULIA P.; JABOR, VALQUIRIA A. P.; FELICIANO, PATRICIA R.; NONATO, MARIA CRISTINA; KAISER, CARLOS R.; MENEZES, CARLA MARIA S.; HAMMES, AMANDA S. O.; CAFFARENA, ERNESTO RAUL; HOELZ, LUCAS V. B.; et al. Evaluation of 7-arylaminopyrazolo[1,5-a]pyrimidines as anti-Plasmodium falciparum, antimalarial, and Pf-dihydroorotate dehydrogenase inhibitors. EUROPEAN JOURNAL OF MEDICINAL CHEMISTRY, v. 126, p. 72-83, JAN 27 2017. (12/25075-0)

PERES, RAPHAEL S.; SANTOS, GABRIELA B.; CECILIO, NERRY T.; JABOR, VALQURIA A. P.; NIEHUES, MICHAEL; TORRES, BRUNA G. S.; BUQUI, GABRIELA; SILVA, CARLOS H. T. P.; DALLA COSTA, TERESA; LOPES, NORBERTO P.; et al. Lapachol, a compound targeting pyrimidine metabolism, ameliorates experimental autoimmune arthritis. ARTHRITIS RESEARCH & THERAPY, v. 19, MAR 7 2017. (13/08216-2, 12/20990-2, 09/54014-7, 12/25075-0, 14/50265-3)

REIS, RENATA A. G.; CALIL, FELIPE ANTUNES; FELICIANO, PATRICIA ROSA; PINHEIRO, MATHEUS PINTO; NONATO, M. CRISTINA. The dihydroorotate dehydrogenases: Past and present. Archives of Biochemistry and Biophysics, v. 632, n. SI, p. 175-191, OCT 15 2017. (11/23504-9, 07/08703-0, 12/25075-0)

PADUA, RICARDO A. P.; TOMALERI, GIOVANI P.; REIS, RENATA A. G.; DAVID, JULIANA S.; SILVA, VALERIA C.; PINHEIRO, MATHEUS P.; NONATO, MARIA CRISTINA. ThermoFMN - a thermofluor assay developed for ligand-screening as an alternative strategy for drug discovery. Journal of the Brazilian Chemical Society, v. 25, n. 10, p. 1864+, OCT 2014. (11/23504-9, 07/08703-0, 08/11644-8, 11/14269-6, 12/25075-0)

AJALLA ALEIXO, MARIANA A.; RANGEL, VICTOR L.; RUSTIGUEL, JOANE K.; DE PADUA, RICARDO A. P.; NONATO, MARIA CRISTINA. Structural, biochemical and biophysical characterization of recombinant human fumarate hydratase. FEBS Journal, v. 286, n. 10, p. 1925-1940, MAY 2019. (12/25075-0)