Biocatalytic stereoinversion of D-para-bromophenylalanine in a one-pot three-enzyme reaction

Halogenated derivatives of phenylalanine can be used as cross-coupling reagents for making drug-like molecules, and pure enantiomers of these precursors are therefore highly desirable. In our exploration of enzymatic routes to simplify the deracemisation process, the application of two enzymes, D-amino acid transaminase and phenylalanine dehydrogenase, both from Lysinibacillus sphaericus, has given promising results for the stereo-inversion of D-enantiomers of para-bromophenylalanine as the model substrate and also p-chloro/fluorophenylalanine and tyrosine. The addition of a coenzyme recycling system using ethanol and alcohol dehydrogenase reduced the amount of coenzyme needed for the reaction catalysed by phenylalanine dehydrogenase, reducing cost and permitting efficient and complete conversion of the racemic amino acids to the L-enantiomer. Relative proportions of the enzymes were optimized. The high purity of the L-enantiomer, with an ee over 99%, and the ease of the process make it an ideal alternative for deracemisation of the studied compounds.Irish Research Counci

Dissecting RAF Inhibitor Resistance by Structure-based Modeling Reveals Ways to Overcome Oncogenic RAS Signaling

Clinically used RAF inhibitors are ineffective in RAS-mutant tumors because they enhance homo- and heterodimerization of RAF kinases, leading to paradoxical activation of ERK signaling. Overcoming enhanced RAF dimerization and the resulting resistance is a challenge for drug design. Combining multiple inhibitors could be more effective, but it is unclear how the best combinations can be chosen. We built a next-generation mechanistic dynamic model to analyze combinations of structurally different RAF inhibitors, which can efficiently suppress MEK/ERK signaling. This rule-based model of the RAS/ERK pathway integrates thermodynamics and kinetics of drug-protein interactions, structural elements, post-translational modifications and cell mutational status as model rules to predict RAF inhibitor combinations for inhibiting ERK activity in oncogenic RAS and/or BRAFV600E backgrounds. Predicted synergistic inhibition of ERK signaling was corroborated by experiments in mutant NRAS, HRAS and BRAFV600E cells, and inhibition of oncogenic RAS signaling was associated with reduced cell proliferation and colony formation.European Commission Horizon 2020Science Foundation Irelan

Investigating Polypharmacology through Targeting Known Human Neutrophil Elastase Inhibitors to Proteinase 3

Using a combination of multisite λ−dynamics (MSλD) together with in vitro IC50 assays, we evaluated the polypharmacological potential of a scaffold currently in clinical trials for inhibition of human neutrophil elastase (HNE), targeting cardiopulmonary disease, for efficacious inhibition of Proteinase 3 (PR3), a related neutrophil serine proteinase. The affinities we observe suggest that the dihydropyrimidinone scaffold can serve as a suitable starting point for the establishment of polypharmacologically targeting both enzymes and enhancing the potential for treatments addressing diseases like chronic obstructive pulmonary disease

Investigating Polypharmacology through Targeting Known Human Neutrophil Elastase Inhibitors to Proteinase 3

Using a combination of multisite λ−dynamics (MSλD) together with in vitro IC50 assays, we evaluated the polypharmacological potential of a scaffold currently in clinical trials for inhibition of human neutrophil elastase (HNE), targeting cardiopulmonary disease, for efficacious inhibition of Proteinase 3 (PR3), a related neutrophil serine proteinase. The affinities we observe suggest that the dihydropyrimidinone scaffold can serve as a suitable starting point for the establishment of polypharmacologically targeting both enzymes and enhancing the potential for treatments addressing diseases like chronic obstructive pulmonary disease