Fluorovinylsulfones and -Sulfonates as Potent Covalent Reversible Inhibitors of the Trypanosomal Cysteine Protease Rhodesain: Structure–Activity Relationship, Inhibition Mechanism, Metabolism, and In Vivo Studies

Rhodesain is a major
cysteine protease of Trypanosoma
brucei rhodesiense, a pathogen causing Human African
Trypanosomiasis, and a validated drug target. Recently, we reported
the development of α-halovinylsulfones as a new class of covalent
reversible cysteine protease inhibitors. Here, α-fluorovinylsulfones/-sulfonates
were optimized for rhodesain based on molecular modeling approaches. 2d, the most potent and selective inhibitor in the series,
shows a single-digit nanomolar affinity and high selectivity toward
mammalian cathepsins B and L. Enzymatic dilution assays and MS experiments
indicate that 2d is a slow-tight binder (Ki = 3 nM). Furthermore, the nonfluorinated 2d-(H) shows favorable metabolism and biodistribution by accumulation in
mice brain tissue after intraperitoneal and oral administration. The
highest antitrypanosomal activity was observed for inhibitors with
an N-terminal 2,3-dihydrobenzo­[b]­[1,4]­dioxine group
and a 4-Me-Phe residue in P2 (2e/4e) with
nanomolar EC50 values (0.14/0.80 μM). The different
mechanisms of reversible and irreversible inhibitors were explained
using QM/MM calculations and MD simulations.