Production, structural and functional validation of the BDNF neurotrophin and its recombinant proisoforms with emphasis in depressive disorder

• Grant number: 23/05079-6
• Support Opportunities: Regular Research Grants
• Start date: October 01, 2023
• End date: December 31, 2025
• Field of knowledge: Biological Sciences - Pharmacology - Biochemical and Molecular Pharmacology
• Principal Investigator: Renê de Oliveira Beleboni
• Grantee: Renê de Oliveira Beleboni
• Host Institution: Universidade de Ribeirão Preto (UNAERP). Campus Ribeirão Preto. Ribeirão Preto , SP, Brazil
• Associated researchers: Ana Lucia Fachin Saltoratto ; Mozart de Azevedo Marins ; Rodrigo Guerino Stabeli ; Suzelei de Castro França
• Associated scholarship(s): 25/06653-3 - Neuroprotection Mediated by BDNF and its Recombinant Isoforms in Neuron-Like Cells Exposed to Glutamatergic Hyperexcitability, BP.IC; 24/14919-0 - PRODUCTION, PURIFICATION AND STRUCTURAL VALIDATION OF THE BDNF NEUROTROPHIN AND ITS RECOMBINANT PROISOFORMS WITH EMPHASIS IN DEPRESSIVE DISORDER, BP.IC

Abstract

Depression is a neuropsychiatric disorder that affects 10-20% of the world population, and is therefore a serious universal public health problem. Although the understanding, diagnosis/monitoring and treatment of this disorder have progressed in recent decades, much still needs to evolve in order for more significant advances impact the reality of medical practice and patients' health. Regarding the neurobiological explanations of depression, the modern complementation of the monoamine theory by the neurotrophin theory has pointed to the neurotrophic factor BDNF and its pro-isoforms as entities with great practical potential as biomarker and/or treating the disorder. The BDNF factor is found at reduced levels, especially in the hippocampus and prefrontal cortex of depressed patients, and increases in level after effective treatment with conventional antidepressants. Blood levels of BDNF accompany the reduction of its brain content and severity of the disorder, and also rise in the course of its effective treatment. The objective of this work will be to provide the recombinant and nationalized production of BDNF and its human and main proisoforms, as well as to provide the diverse structural/functional validation of these actives with an innovative emphasis on the potential treatment/biomarker of depression. Heterologous production will take place in E. coli using pET-26b(+) plasmid expression vectors with expected initial productivity in the mg/L range. Purification will be carried out using affinity chromatography in a nickel column, and the actives will then be quantified and analyzed for the degree of homogeneity/purity. The structural identification/validation will be carried out by mass spectrometry, while the functional validation will be carried out by various in vitro and in vivo tests. Functional validation will occur simultaneously with the identification of the best active(s) in terms of extending the study of alternative routes of administration, especially intranasal, and increasing general brain bioavailability and/or directed to specific areas of the brain. In the context of this last issue, it is necessary to consider the hypothesis that pro-BDNF isoforms seem to be more protected from proteolysis than mature BDNF, in addition to being able to target more specific areas of the brain, where they are more important, and easily activated. In this context, brain bioavailability and targeting can vary widely between BDNF and proisoforms and, above all, between different proisoforms. Still in the stock of actions of this project and based on the evaluation of performance in the functional validation, the best asset(s) will be elected for tests on a chronic scale of the disease and treatment in animal models of choice. Indeed, this will additionally make it possible to monitor the role of blood BDNF as a biomarker in the evolution of the treatment provided by itself and/or from the activation of proisoform(s) at the brain level. Finally, and still conditioned to the results obtained, it is part of the proposal to request the patenting of the use of the active(s) of greatest performance/interest for the treatment of depression, as well as to initiate studies for pilot/semi scaling. -industrial production of these same assets. In this way, the present project has marked biotechnological value, as well as great medical and industrial-pharmaceutical interest. It has an anticipatory market function and establishes an expanded biochemical-pharmacological production and study arrangement for innovative actives with a great future. (AU)

Scientific publications

(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)

RODRIGUES, ELLEN F.; FACHIN, ANA L.; MARINS, MOZART; STABELI, RODRIGO G.; BELEBONI, RENE O.. Challenges in recombinant brain-derived neurotrophic factor production. TRENDS IN BIOTECHNOLOGY, v. 42, n. 5, p. 4-pg., 2024-05-01. (23/05079-6)