The Serine hydrolase enzyme family is one of the largest and most diverse protein classes, including proteases, lipases, esterases, thioesterases, amidases and peptidases. All these enzymes utilize a base-activated Serine residue cleavage of amide bonds in substrates via a covalent acyl-enzyme intermediate.

Different specific covalent-acting chemical probes have increasingly been used in proteome-wide target identification and imaging and for finding inhibitors with high specificity among related enzymes and enzyme isoforms. Significant number of known covalent drugs and natural products have approved efficacy of such approach in drug discovery.

Our library of Serine focused covalent fragments is available in versatile pre-plated formats for most convenient and fast delivery. All compounds passed QC check (90%+ purity) before formatting to ensure quality of the library.

Examples of compounds in the library

Library Design

Recently elaborated approaches to parallel synthesis, allowed Enamine to synthesize the largest commercial source of covalent binders. We focused our efforts on synthesis of compounds with most reliable covalent warheads, with well determined reactivity. Our Serine focused library was designed based on a combination of specific moieties, reported to form covalent bonds particularly with Serine residue and presence of a drug-like, MedChem friendly core in a molecule. The library has also been shaped with Ro3 criteria to meet all requirements of FBDD.

The following functional groups, reported to form covalent bonds in proteins with Ser residue, have been used for construction of the library:

• Sulfonyl fluorides, fluorosulfonates and sulfamoyl fluorides
• Epoxides
• β-lactams and β-Propiolactone
• Boronic acids and pinacolates
• Aldehydes

Hits derived from this library can be easily followed with analogues from over 4.4M stock compounds or either synthesis of new compounds through REAL Database technology within 3 weeks only.

We provide Hit Confirmation support for all our libraries with the samples resupply from dry powders, QC check and HPLC repurification. Hits can be resynthesized in 2 weeks only.

Covalent chemical probes become an important tool in drug discovery within last few years. The impressive number of successful applications in protein drugability assessment, especially focusing on Cys residue, brought aspiration to discovery and synthesis of new covalent modifiers.

We used deep knowledge-based approach to design and synthesize our cysteine focused fragment library. Careful choice of covalent warheads based on their reactivity was performed by experienced chemists and reflected a small set of specific structural filters. Electrophilic fragments selected by structural filters were then analyzed to remove any molecules with trivial or unwanted structural features. The resulting set was refined with Ro3 criteria applied to “core structures” to yield 3 200 fragments able to form covalent bonds with cysteine residues.

Hits derived from this library can be easily followed with analogues from stock or either synthesis of new compounds through REAL Database technology.

Key features

• Only well-validated covalent warheads that form easy-to-interpret adducts.
• No overreactive, promiscuous binders.
• Attractive scaffolds, derived from the latest core building blocks.
• Plated at 100 mM concentration in DMSO.

The following warheads were used for the library construction:

• Acrylamides – 1,040
• Dimethylamine substituted acrylamides – 480
• Chloroacetamides – 800
• 2-Chloropropionamides – 320
• Chlorofluoroacetamides – 320
• Butynamides - 240

Examples of the molecules in Cysteine focused Library

Covalent chemical probes have become essential for drug discovery in recent years. The impressive number of successful applications in protein drugability assessment, proteomics, and investigation of protein functions brought aspiration to the discovery of new covalent modifiers.

We are continuously working on elaborating parallel chemistry approaches and the synthesis of new covalent compounds of various classes. Enamine Covalent Collection comprises over 133k covalent binders and is frequently updated with new warhead types. The acquired experience and knowledge on stability and selectivity were applied to the design of our new Covalent Fragment Library.

The renewed library has been designed to represent Enamine’s most recent scaffolds in combination with the most interesting covalent anchors. Careful choice of the latest attractive and highly tractable covalent functionalities makes this library the most reliable source of covalent fragments.

All compounds passed rigorous QC before formatting to ensure the high quality of our library.

Covalent Fragment Library is available for fast supply in multiple formats, including the following most popular options.

Key features

• Careful choice of warheads: only experimentally confirmed reactivity, no over-reactive warheads, no alternative reactive functionalities.
• Stable in DMSO and suitable for multiple freeze-thaw cycles.
• Advanced covalent binders: balanced reactivity covalent binders, no promiscuous binders
• Fragments are pre-plated by classes and can be acquired separately.
• The latest compounds featuring renowned Enamine Building Blocks.

Composition of the Library by the type of covalent binders

Examples of compounds in the library

Enamine’s Lipid GPCR Library has been designed in collaboration with NQuiX. Over 6 400 compounds have been selected from off-the-shelf collection or specifically synthesized for this project from Enamine REAL screening Collection. They were designed to target the family of eight endothelial differentiation gene (EDG) receptors (S1P1–5 and LPA1–3). The compounds may also be appropriate for screening at GPR3, GPR6 and GPR12 orphan receptors given some TM bundle binding site similarity and/or GPR23, GPR92 and P2Y5 given their more recent classification as additional, albeit distinct, LPA receptors.

Library design

The compounds have been selected using a combination of ligand-based methods using chemical fingerprints, 2D pharmacophores and 3D shape/feature matches (Figure 1). The training active compound set included 1 393 compounds (870 acidic and 523 non-acidic), collected across 73 papers (Figure 2). They represent a combination of compounds for expanding SAR around known chemotypes and scaffold hops seeking novelty. The current set of compounds covers the non-acidic classes of ligand, with acidic compounds being designed via a different procedure and added to the library separately.

Fig. 1.Dedicated design of the compounds included in the Library (general scheme).

Fig. 2. Distribution of the compounds in the training set.

Examples of the molecules

Z1576067638

Z740728124

Z641956124

Z1303449800

Z1523110947

Z1303440017

Z1576459756