Webminar: Structural Approach to characterize the Metabolic Activation Pathway Of a Nucleotide analogue : From Pro-Drug to Target

Summary: Bemnifosbuvir (AT-527) is a guanosine analog currently in clinical trial phase III against SARS-CoV-2, and phase II (with Ruzasvir) against HCV (1,2). This purine nucleotide analogue (NA) is modified at three positions relative to their natural nucleotide counterparts: the purine N⁶, the 2'-ribose, and the 5'-phosphate.

Here we show this NAs require a minimal set of 5 enzymes, namely CatA/CES1 esterase, HINT1, ADALP1, GUK1, and NDPK for activation to their common 5'-triphosphate active form AT-9010, with an obligate order of reactions prior hitting the main component of the Sars-CoV2 replication complex. AT-9010 inhibits essential viral activities enzymes, accounting for broad spectrum antiviral potency (3). Enzyme assays together with structural data at atomic resolution illuminate N⁶-purine deamination compatible with metabolic activation by the human ADALP1 enzyme. Crystal structures of human HINT1, ADALP1, GUK1, and NDPK at 2.09, 2.44, 1.76, and 1.9 Å resolution, respectively, with respective precursors of AT-9010 complete the five-steps activation pathway from the orally available drug Bemnifosbuvir to AT-9010, pointing to key drug-protein contacts along the activation pathway as well as chemical positions candidate for further modification on the purine nucleotide.

Our work provides a structural and functional framework to integrate antiviral nucleotide analogue design from drug bio-availability and metabolic activation to antiviral potency.

Ref : Chazot A, Zimberger C, Feracci M, Moussa A, Good S, Sommadossi JP, Alvarez K, Ferron F, Canard B. The activation cascade of the broad-spectrum antiviral bemnifosbuvir characterized at atomic resolution. PLoS Biol. 2024 Aug 27;22(8):e3002743. doi: 10.1371/journal.pbio.3002743