Discovery of New Uncompetitive Inhibitors of Glucose-6-Phosphate Dehydrogenase

Abstract

The enzyme glucose-6-phosphate dehydrogenase (G6PDH) catalyzes the first step of the oxidative branch of the pentose phosphate pathway, which provides cells with NADPH, an essential cofactor for many biosynthetic pathways and antioxidizing enzymes. In Trypanosoma cruzi, the G6PDH has being pursued as a relevant target for the development of new drugs against Chagas disease. At present, the best characterized inhibitors of T. cruzi G6PDH are steroidal halogenated compounds derivatives from the mammalian hormone precursor dehydroepiandrosterone, which indeed are also good inhibitors of the human homologue enzyme. The lack of target selectivity might result in hemolytic side effects due to partial inhibition of human G6PDH in red blood cells. Moreover, the treatment of Chagas patients with steroidal drugs might also cause undesired androgenic side effects. Aiming to identify of new chemical classes of T. cruzi G6PDH inhibitors, we performed a target-based high-throughput screen campaign against a commercial library of diverse compounds. Novel TcG6PDH inhibitors were identified among thienopyrimidine and quinazolinone derivatives. Preliminary structure activity relationships for the identified hits are presented, including structural features that contribute for selectivity toward the parasite enzyme. Our results indicate that quinazolinones are promising hits that should be considered for further optimization.

A small molecule G6PD inhibitor reveals immune dependence on pentose phosphate pathway

“…We combed the literature for a compound series that could serve as a suitable chemical starting point for inhibitor discovery. Our search identified a non-steroidal aminoquinazolinone series that was recently discovered and optimized against Trypanosoma cruzi G6PD 26 . Synthesis of representative compounds identified G6PDi-precursor ( 5 ) with low micromolar in vitro activity against human G6PD ( Figure 2a – b ).…”

Mutations in the tetramer interface of human glucose‐6‐phosphate dehydrogenase reveals kinetic differences between oligomeric states

“…The cloning of the expression vector pET_SU-MO_HsDG6PDH encoding for a short construction of HsG6PDH (HsDG6PDH, residues 29-511) with a N-terminal His-SUMO tag was described elsewhere [10]. The single HsDG6PDH mutants A277C and E347A (HsDG6PDH_ A277C and HsDG6PDH_E347A, respectively) were generated using the QuickChange II XL Site-Directed Mutagenesis Kit (Stratagene, La Jolla, CA, USA), following the manufacturer's instructions and with the oligonucleotides: 5 0 -GGCCATGGAGAAGCCCTGCTCCACC AACTCAG-3 0 and 5 0 -CTGAGTTGGTGGAGCAGGG CTTCTCCATGGCC-3 0 for A277C and 5 0 -CTATGTG GAGAATGCGAGGTGGGATGGGG-3 0 and 5 0 -CCCCA TCCCACCTCGCATTCTCCACATAG-3 0 for E347A (nucleotides underlined indicate the mutation positions).…”

“…The expression and purification of both HsDG6PDH mutants followed the protocol previously established for HsDG6PDH [10]. Briefly, Escherichia coli BL21 (DE3) cells harboring either HsDG6PDH_A277C or HsDG6PDH _E347A vectors were grown in ZYM5052 autoinduction medium [11] for 48 h at room temperature and 250 rpm.…”

The structure of a Trypanosoma cruzi glucose‐6‐phosphate dehydrogenase reveals differences from the mammalian enzyme

“…Additionally, steroids and quinazolinones were shown to inhibit T. cruzi G6PDH and to kill epimastigote forms of the parasite in vitro , suggesting that G6PDH is an attractive target for development of new trypanocidal drugs. Some efforts have already been made to develop G6PDH inhibitors with therapeutic utility against trypanosomiasis and also cancer . Steroids and quinazolinones represent the most potent G6PDH inhibitors known to date.…”