Thiol Reactivity of N-Aryl α-Methylene-γ-lactams: A Reactive Group for Targeted Covalent Inhibitor Design

Abstract

Kinase activity can be modulated reversibly or irreversibly by the reaction of targeted covalent inhibitors with nucleophilic residues in protein active sites. Herein, we present thiol reactivity studies that support α-methylene-γ-lactams as tunable surrogates for the highly reactive α-methylene-γ-lactones. The reactivity of the α-methylene is modulated via the N substituent, and the reaction rates toward glutathione were determined via mass spectrometry. Density functional theory calculations of transition states of thiol additions to α-methylene-γ-lactams revealed that the use of the M06-2X functional with the SMD solvation model and methyl thiolate as a model nucleophile reliably predicts the relative reactivities of the α-methylene-γ-lactams, and quasiharmonic approximations improve the agreement between experiment and computation.

Thiol Reactivity of N-Aryl α-Methylene-γ-lactams: Influence of the Guaianolide Structure

“…Conditions were selected to ensure pseudo-first-order kinetics and rate constants and to the benchmark thiol reactivity relative to those previously reported for N -aryl 3-methylene-2-pyrrolidinones and N -aryl acrylamides (see the Supporting Information). ,,, Thiol addition occurs at the exocyclic methylene of the lactam, as evidenced by the disappearance of the methylene protons in the 1 H NMR when 20b was reacted with cysteamine (see Figure S2). Moreover, Hammett linear free-energy relationship studies for the reaction of GSH with the lactams support this chemoselectivity conclusion ( vide infra ).…”

“…To understand the reactivity differences between the corresponding cis- and trans -lactams, the measured rate constants were benchmarked against the reactivity data for the N -aryl 3-methylene-2-pyrrolidinones that our lab recently reported (see the Supporting Information). Rate constants for the cis -lactams 18a – 24a were nearly identical to those for the N -aryl 3-methylene-2-pyrrolidinone series S1 – S4 (compare blue and gray plots, respectively, in Figure ). On the contrary, the trans -lactams 18b – 24b showed a 10-fold rate increase when compared to the pyrrolidinone series S1 – S4 (compare orange and gray plots, respectively, in Figure ).…”

“…Additionally, it expands the chemical space of natural product-derived lead compounds for TCI development to include natural products previously considered over-reactive and therapeutically unstable. This strategy is based on the knowledge that the reactivity of the lactam is within the realm of warhead fragments currently seeing success in TCI development and builds upon our previous studies, where reaction kinetics revealed a linear free-energy relationship with the Hammett parameters of the N -aryl group of the α-methylene-γ-lactam and density functional theory (DFT) calculations accurately predicted relative reactivities (Figure B) …”

“…Notably, Pfizer recently unveiled a coronavirus disease 2019 (COVID-19) antiviral clinical candidate equipped with a γ-lactam adjacent to a CRG that reacts with Cys145 of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) main protease. , The 5,7,5-ring system is accessed using a highly convergent strategy involving the early stage installation of the allenyl moiety and an allenic Pauson–Khand reaction (APKR) developed in our group . Thiol reactivities toward glutathione (GSH) were determined for these analogues under biomimetic conditions using previously reported reaction kinetics protocols. , We expect findings from this work to inform the design of other molecularly complex compounds possessing CRGs. Additionally, these lactam analogs will enhance structure–activity relationship studies of 6,12-guaianolides through their moderated thiol reactivity.…”

One-Pot Diastereoselective Synthesis of Pyrrolopiperazine-2,6-diones by a Ugi/Nucleophilic Substitution/N-Acylation Sequence

“…This route allows high chemical yields and an almost quantitative diastereoselectivity of diketopiperazine–pyrrolidinone derivatives bearing an unsubstituted C8a position and different substituents in the C4 position ( Scheme 1 d). Given the interest in the α-alkylidene-γ-lactam scaffold in the development of compounds with biological properties as Michael acceptors toward bionucleophiles, 16 α-methylidene-γ-lactams fused with 2,6-diketopiperazines, systems which have not been described so far, were also synthesized following a similar strategy.…”