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Use of phage display as a tool in the diagnosis and control of diseases transmitted by hematophagous vectors

Grant number: 19/03779-5
Support type:Research Projects - Thematic Grants
Duration: November 01, 2019 - October 31, 2024
Field of knowledge:Biological Sciences - Biochemistry - Chemistry of Macromolecules
Principal Investigator:Aparecida Sadae Tanaka
Grantee:Aparecida Sadae Tanaka
Home Institution: Escola Paulista de Medicina (EPM). Universidade Federal de São Paulo (UNIFESP). Campus São Paulo. São Paulo , SP, Brazil
Assoc. researchers:Alexandre Keiji Tashima ; Francisco Jose Alves Lemos ; Isaias Glezer ; Itabajara da Silva Vaz Junior ; Jayme Augusto de Souza-Neto ; Luciano Puzer ; Vitor Marcelo Silveira Bueno Brandão de Oliveira
Associated grant(s):20/05451-4 - COVID-19: selection of specific inhibitors for main protease Mpro of SARS-CoV-2 by phage display, AP.R
Associated scholarship(s):20/02433-5 - Library construction of cDNA fragments of R. microplus proteolytic enzymes (peptidases) and serpins on the surface of filamentous phages, BP.PD
19/26101-4 - Biochemical characterization of the putative Kazal serinoprotease inhibitor (AaKz211) modulated in dengue virus 2- infected Aedes aegypti mosquitoes, BP.IC

Abstract

Neglected diseases are responsible for epidemics with countless deaths each year in many countries, especially in developing countries. Most neglected diseases are transmitted by hematophagous vectors. Over the last decades, our group has been working on the prospection of active molecules produced by hematophagous arthropods, vectors of diseases. In the last project, our group evaluated the role of previously prospective molecules in the interaction of vectors with their etiological agents. In the present project, we are going to use phage display, vector proteins and flaviviruses (eg DENV-2) to identify cyclic peptides (using template peptide SFTI - sunflower trypsin inhibitor) and antibodies as targets for diagnosis, antiviral and vector control. The vectors targeted by this project will be the mosquito Aedes aegypti (vector of dengue, yellow fever, zika) and the tick of the species Rhipicephalus (Boophilus) microplus, ectoparasite of cattle. Our group already has extensive experience in the phage display technique using filamentous phage, and to extend this experience, T7 phage libraries and libraries of antibody fragments displayed on phage will be constructed in collaboration with international researchers (Dr. A. Mulenga, USA and Dr. M Hust, Germany). The cyclic peptide libraries will be used in the selection of inhibitors for dengue 2 proteases and digestive proteases of Ae. aegypti larvae. The libraries of tick intestine cDNA fragments will be screened for immunoglobulins from resistant or immunized bovines and the libraries of recombinant human antibody fragments will be screened for flavivirus prM proteins. The selected phages will be sequenced, the sequences of peptides obtained with high frequency will synthesized by company. Selected cDNA fragments and antibody fragments will be produced in bacteria or yeast. Recombinant proteins or peptides will be tested as antivirals, larvicide, antigens for tick control and differential diagnosis of flavivirus (dengue, yellow fever and zika). The molecules identified with biotechnological potential should be used for vector control, viral replication control (DENV-2) and flavivirus diagnosis. For the success of this project, in addition to international collaborations, we will have the collaboration of Brazilian researchers from UNIFESP, UFABC, UNESP, UENF, UFRGS and Fiocruz de Campo Grande, as well as technicians, postgraduate students and postdoctoral fellows. The project also aims to train human resources. (AU)