Planejamento e síntese de inibidores de di-hidrofolato redutase para micobactérias não-tuberculosas

Author(s)

Matheus Andrade Meirelles

• Document type: Doctoral Thesis
• Press: Campinas, SP.
• Institution: Universidade Estadual de Campinas (UNICAMP). Instituto de Química
• Defense date: 2023-02-13

Examining board members

Ronaldo Aloise Pilli; Carlos Roque Duarte Correia; Daniel Fábio Kawano; Roberto Parise Filho

• Advisor: Rafael Lemos Miguez Couñago; Ronaldo Aloise Pilli

Abstract

Nontuberculous mycobacteria (NTM) are opportunistic pathogens responsible for lung diseases in immunocompromised patients or patients with pre-existing lung diseases such as cystic fibrosis and chronic obstructive pulmonary disease. The incidence and prevalence of these diseases are increasing worldwide due to the lack of effective drugs. In this work, we aim to modify the chemical structure of compound P218, a dihydrofolate reductase (DHFR) inhibitor developed for malaria, to access new DHFR inhibitors for NTM. Through a visual inspection of the crystal structures of the M. abscessus and human enzymes complexed with P218, we decided to modify the position containing the ethyl group in the pyrimidine ring to obtain more selective analogues. Then, we designed and synthesized five generations of DHFR inhibitors for Mycobacterium avium and Mycobacterium abscessus. A target product profile (TPP) was defined to monitor the development of new molecules and the progress of the project. The base (TMP)2Zn·2MgCl2·2LiCl and palladium catalyzed reactions were instrumental to access the desired modification at the C-6 position in good yields. Moreover, the new synthetic strategy was used as a reliable method to synthesize P218, and this route was also used to incorporate nitrogen-containing groups at the C-6 position (fourth-generation analogues). Finally, we developed a synthetic route that allowed the late-stage modification of the P218 tail, which was employed to prepare the fifth-generation analogues. The analogue containing the cyclohexyl ring (29.5) maintained the high activity for both bacterial enzymes being 8-fold less active against the human DHFR compared to P218. More impressive enzymatic profiles were achieved with four analogues (34.1, 34.5, 36.2, 36.3), which were potent against the M. avium and M. abscessus DHFR (Ki < 1 nM) and inactive against the human (KiHs > 5000 nM). A collaboration with the Seattle Children’s Hospital and the University of Kansas enabled the cocrystalization of six P218 analogues with M. ulcerans DHFR, which helped to understand their biding mode and possible future directions in developing DHFR inhibitors. Five analogues (F14.4, 29.5, 33.3, 34.6, 44.6) exhibited higher potency than the antibiotic amikacin against M. avium subp. hominissuis, with MIC90MAH < 1 ?M in whole-cell inhibition assay, which was performed in collaboration with the McGill University, Canada. Compounds 29.5 and 44.6 also showed similar potency to amikacin against M. asbcessus. DMPK studies in collaboration with the University of Dundee, Scotland, showed that the P218 analogues F14.4, 29.5, 33.3, and 34.6 presented excellent metabolic stability in the presence of both mouse and human microsomes and hepatocytes. Although solubility may have to be improved, the compounds also demonstrated good permeability in the MDCK assay. The modification of the C-6 position was essential in developing potent and selective P218 analogues against M. avium and M. abscessus. The excellent drug profile exhibited by some of the synthesized molecules (F14.4, 29.5, 33.3, and 34.6) indicates that these compounds could represent promising lead compounds (AU)

• FAPESP's process: 19/20735-1 - Design and synthesis of dihydrofolate reductase inhibitors for nontuberculous mycobacteria
• Grantee: Matheus Andrade Meirelles
• Support Opportunities: Scholarships in Brazil - Doctorate