Structural basis of nirmatrelvir and ensitrelvir activity against naturally occurring polymorphisms of the SARS-CoV-2 main protease.

SARS-CoV-2 is the causative agent of COVID-19. The main viral protease (Mpro) is an attractive target for antivirals. The clinically approved drug nirmatrelvir and the clinical candidate ensitrelvir have so far showed great potential for treatment of viral infection. However, the broad use of antivirals is often associated with resistance generation. Herein, we enzymatically characterized 14 naturally occurring Mpro polymorphisms that are close to the binding site of these antivirals. Nirmatrelvir retained its potency against most polymorphisms tested, while mutants G143S and Q189K were associated with diminished inhibition constants. For ensitrelvir, diminished inhibition constants were observed for polymorphisms M49I, G143S, and R188S, but not for Q189K, suggesting a distinct resistance profile between inhibitors. In addition, the crystal structures of selected polymorphisms revealed interactions that were critical for loss of potency. In conclusion, our data will assist the monitoring of potential resistant strains, support the design of combined therapy, as well as assist the development of the next generation of Mpro inhibitors.

Age-related tolerance to paraquat-induced parkinsonism in Drosophila melanogaster.

Paraquat (PQ) is a widely used herbicide that can cross the dopaminergic neuronal membrane, accumulate in mitochondria and damage complex I of the electron transport chain, leading to neuronal death. In Drosophila melanogaster, PQ exposure leads to the development of parkinsonism and is a classical model for studying Parkinson's Disease (PD). Muscle mitochondrial dysfunction, affecting survival and locomotion, is described in familial PD in D. melanogaster mutants. However, no study has shown the effects of PQ-induced parkinsonism in D. melanogaster regarding muscle ultrastructure and locomotor behavior at different ages. Thus, we evaluated survival, locomotion, and morphological parameters of mitochondria and myofibrils using transmission electron microscopy in 2 and 15-day-old D. melanogaster, treated with different PQ doses: control, 10, 50, 100, 150, and 200 mM. PQ100mM presented 100% lethality in 15-day-old D. melanogaster, while in 2-day-old animals PQ150mM produced 20% lethality. Bradykinesia was only observed in 15-day-old D. melanogaster treated with PQ10 mM and PQ50 mM. However, these results are unlikely to be associated with changes to morphology. Taken together, our data indicate pathophysiological differences between PQ-induced parkinsonism and familial parkinsonism in D. melanogaster (resultant from gene mutations), demonstrating for the first time a differential susceptibility to PQ in two developmental stages.