Discovery of a novel ligand that modulates the protein–protein interactions of the AAA+ superfamily oncoprotein reptin

Developing approaches to discover protein–protein interactions (PPIs) remains a fundamental challenge. A chemical biology platform is applied here to identify novel PPIs for the AAA+ superfamily oncoprotein reptin. An in silico screen coupled with chemical optimization provided Liddean, a nucleotide-mimetic which modulates reptin''s oligomerization status, protein-binding activity and global conformation. Combinatorial peptide phage library screening of Liddean-bound reptin with next generation sequencing identified interaction motifs including a novel reptin docking site on the p53 tumor suppressor protein. Proximity ligation assays demonstrated that endogenous reptin forms a predominantly cytoplasmic complex with its paralog pontin in cancer cells and Liddean promotes a shift of this complex to the nucleus. An emerging view of PPIs in higher eukaryotes is that they occur through a striking diversity of linear peptide motifs. The discovery of a compound that alters reptin''s protein interaction landscape potentially leads to novel avenues for therapeutic development.     

Second-generation DNA-encoded multiple display on a constant macrocyclic scaffold enabled by an orthogonal protecting group strategy

DNA-encoded chemical library(DEL) represents an emerging drug discovery technology to construct compound libraries with abundant chemical combinations. While drug-like small molecule DELs facilitate the discovery of binders against targets with defined pockets, macrocyclic DELs harboring extended scaffolds enable targeting of the protein–protein interaction(PPI) interface. We previously demonstrated the design of the first-generation DNA-encoded multiple display based on a constant macrocyclic s...     

Engineered Fully Human Single-Chain Monoclonal Antibodies to PIM2 Kinase

Proviral integration site of Moloney virus-2 (PIM2) is overexpressed in multiple human cancer cells and high level is related to poor prognosis; thus, PIM2 kinase is a rational target of anti-cancer therapeutics. Several chemical inhibitors targeting PIMs/PIM2 or their downstream signaling molecules have been developed for treatment of different cancers. However, their off-target toxicity is common in clinical trials, so they could not be advanced to official approval for clinical application. Here, we produced human single-chain antibody fragments (HuscFvs) to PIM2 by using phage display library, which was constructed in a way that a portion of phages in the library carried HuscFvs against human own proteins on their surface with the respective antibody genes in the phage genome. Bacterial derived-recombinant PIM2 (rPIM2) was used as an antigenic bait to fish out the rPIM2-bound phages from the library. Three E. coli clones transfected with the HuscFv genes derived from the rPIM2-bound phages expressed HuscFvs that bound also to native PIM2 from cancer cells. The HuscFvs presumptively interact with the PIM2 at the ATP binding pocket and kinase active loop. They were as effective as small chemical drug inhibitor (AZD1208, which is an ATP competitive inhibitor of all PIM isoforms for ex vivo use) in inhibiting PIM kinase activity. The HuscFvs should be engineered into a cell-penetrating format and tested further towards clinical application as a novel and safe pan-anti-cancer therapeutics.     

Isolation and synthesis of pygmanilines,phenylurea derivatives from the Northeastern Atlantic lichen Lichina pygmaea

A library of 864 extracts prepared from Irish marine organisms was screened in the search for novel antioxidants. Chemical investigation of the most promising extract obtained from the common lichen Lichina pygmaea led to the isolation of five resorcinol derivatives. An unusual urea moiety was found embedded in two of these compounds, pygmanilines A (1) and B (2) and their structures were confirmed by one-step chemical syntheses.     

Design and synthesis of 2(1H)-pyrazinones as inhibitors of protein kinases

Kinase enzymes play a key role in the development and progression of cancer. Inhibitors of deregulated kinases are effective small molecule anticancer drugs. The 2(1H)-pyrazinone heterocycle is a previously unexploited motif that can fulfil the structural requirements for ATP-competitive inhibition of kinases. Rapid solution-phase syntheses of novel 3,5- and 3,6-disubstituted-2(1H)-pyrazinones were developed through selective, sequential substitution of 2,5-dihalo-3-benzyloxypyrazine and 3,5-dihalo-2(1H)-pyrazinone intermediates. Palladium-catalysed cross-couplings and S_NAr reactions were used to introduce substituents chosen on the basis of the calculated physicochemical properties of the target pyrazinones. Representative compounds demonstrated good solubility, kinase inhibitory activity and antiproliferative activity in human tumour cells, confirming the suitability of this chemical class as a kinase-focused library.     

Diversity-oriented decoration of pyrrolo[1,2-a]pyrazines

Diversity-oriented synthesis of a chemical library based on a pyrrolo[1,2-a]pyrazine core is described by using palladium-catalyzed direct C6 arylation of pyrrolo[1,2-a]pyrazines with various aryl bromides. The starting materials, pyrrolo[1,2-a]pyrazines, were easily synthesized by the base-mediated N-alkylation of pyrrole-2-carboxaldehyde with several 2-bromoacetophenones followed by dehydrative cyclization with incorporation of nitrogen by the action of ammonium acetate. Introduction of other functional groups on this chemical scaffold is also discussed herein.     

Development of P1-monodentate diamidophosphites with a C1-symmetric 1,2-diamine backbone: the effects of substituents in the 1,3,2-diazaphospholidine cycle on Pd-catalyzed asymmetric allylations

We have designed and synthesized a small library of modular monodentate diamidophosphite ligands with stereogenic phosphorus atoms. The library was prepared efficiently from the commercially available and inexpensive (S)-N-Boc-amino acids. These novel ligands were screened in the Pd-catalyzed asymmetric allylations of (E)-1,3-diphenylallyl acetate with dimethyl malonate as the C-nucleophile with up to 93% ee being obtained. The results showed that the different substituents in the 1,3,2-diazaphospholidine cycle had remarkable effects on the enantioselectivity.     

Optimisation d’une sélection in vitro d’enzymes

Optimisation of an in vitro enzyme selection. Isolation of a catalyst for a given chemical reaction may be achieved by in vitro selection of enzymes from a protein library. Here, we investigate the polymerisation reaction on filamentous phage and the cross-linking of substrate on phage to optimise an in vitro selection for DNA polymerase activity. The efficiency of the optimised selection is measured by enrichment factors up to 3.8 × 10³, the highest described so far for an in vitro selection of proteins for catalysis. It should be useful for directed polymerase evolution towards novel catalytic activities. To cite this article: E. Orsi, J.-L. Jestin, C. R. Chimie 6 (2003).     

Synthesis of an octahydroindolinone scaffold for a diversity-based chemical compound library

The synthesis of a chemical compound library using diversity-oriented synthesis (DOS) is discussed. The library is structurally inspired by the Amaryllidaceae alkaloids, a family of natural products which has been known to demonstrate potent antiviral and antineoplastic activity. Highlights of this work include the rapid, high-yielding construction of the octahydroindolinone core and the solid-phase diversification of the lactam using a neutral phosphazene base.     

Selection of phage antibodies with GPX activity by combination of phage displayed antibody library with chemical modification and their characterization using a surface plasmon resonance biosensor

Glutathione peroxidase (GPX) is an important antioxidant enzyme, which plays an important role in scavenging reactive oxygen species. To obtain humanized GPX catalytic antibodies, the phage displayed human antibody library on the surface of the filamentous bacteriophage was used to select novel antibodies by repetitive screening. Phage antibodies B8, H6 and C1 with the GSH-binding site were obtained from the library by enzyme-linked immunosorbent assay (ELISA) analysis with four rounds of selection against three haptens, S-2,4-dinitrophenyl t-butyl ester [GSH-S-DNP-Bu (B)], S-2,4-dinitrophenyl t-hexyl ester [GSH-S-DNP-He (H)] and S-2,4-dinitrophenyl cycle-hexyl ester [GSH-S-DNP-cHe (C)], and characterized using surface plasmon resonance (SPR) biosensor. The gold layer was modified by dithiodiglycolic acid (DDA) and three haptens were easily attached to DDA by self-assembling to form a biosensor membrane. The membrane bounds specifically corresponding antibodies. The kinetic process of the reaction between phage antibodies and their haptens was studied by SPR biosensor. In order to improve selectivity, chemical modification was used to incorporate directly catalytic group selenocysteine (Sec) into selected phage clone B8, H6 and C1 to form Se-B8, Se-H6 and Se-C1, respectively. The GPX activities of Se-B8, Se-H6 and Se-C1 were found to be 3000, 2000 and 700 units/μmol, respectively. Compared with conventional ELISA analysis, the proposed method based on SPR biosensor is much more rapid and simpler.     

Synthesis and screening of peptoid arrays on cellulose membranes

Rapid synthesis and screening of compound libraries enables the accelerated identification of novel protein ligands in order to support processes like analysis of protein interactions, drug target discovery or lead structure discovery. SPOT synthesis—a well established method for the rapid preparation of peptide arrays—has recently been extended to the field of nonpeptides. In this contribution we report on the systematic evaluation of the SPOT technique for the assembly of N-alkylglycine (peptoid) library arrays. In the course of this investigation bromoacetic acid 2,4-dinitrophenylester (1a) was identified to be the most suited agent for bromoacetylation in terms of yield and N-selectivity enabling straightforward submonomer synthesis on hydroxy-group rich cellulose membranes. The potential of this method for the rapid identification of novel nonpeptidic protein ligands was demonstrated by synthesis and screening of a library consisting of 8000 peptoids and peptomers (i.e. their hybrids with α-substituted amino acids) allowing the identification of micromolar ligands for the monoclonal antibody Tab-2.     

Chemical Probes for the Functionalization of Polyketide Intermediates

A library of functionalized chemical probes capable of reacting with ketosynthase‐bound biosynthetic intermediates was prepared and utilized to explore in vivo polyketide diversification. Fermentation of ACP mutants of S. lasaliensis in the presence of the probes generated a range of unnatural polyketide derivatives, including novel putative lasalocid A derivatives characterized by variable aryl ketone moieties and linear polyketide chains (bearing alkyne/azide handles and fluorine) flanking the polyether scaffold. By providing direct information on microorganism tolerance and enzyme processing of unnatural malonyl‐ACP analogues, as well as on the amenability of unnatural polyketides to further structural modifications, the chemical probes constitute invaluable tools for the development of novel mutasynthesis and synthetic biology.     

Participation of compact planar 1,3,5-tri(buta-2,3-dien-1-yl)-1,3,5-triazinane-2,4,6-trione in Pd(0) catalysed seven component cascade reactions delivers novel tunable molecular architecture

We report the spatially controlled, protecting group free, catalytic assembly of a library of nineteen 7-component cascade products generated from a novel planar trisallenyl 1,3,5-triazinane-2,4,6-trione core in combination with aryl iodides and amines with excellent regio and good stereoaselectivity for Z,Z,Z-isomers.     

Synthesis of novel spiro imidazolium salts as chiral ionic liquids

A library of 13 novel chiral spiro imidazolium salts has been synthesized. The effects of N-substituents and counteranions on the melting point of spiro bis(imidazolium) salts are studied in efforts toward the development of room temperature chiral ionic liquids.     

A green,isocyanide-based three-component reaction approach for the synthesis of multisubstituted ureas and thioureas

A one-pot, isocyanide based multicomponent protocol was presented starting from secondary amines towards (thio)urea derivatives and utilized for the construction of a diverse 27-membered chemical library. Following a green compatible microwave assisted condition, the formed N,N′-multisubstituted (thio)ureas were obtained in up to 85% yield.     

MEMES: Machine learning framework for Enhanced MolEcular Screening

In drug discovery applications, high throughput virtual screening exercises are routinely performed to determine an initial set of candidate molecules referred to as “hits”. In such an experiment, each molecule from a large small-molecule drug library is evaluated in terms of physical properties such as the docking score against a target receptor. In real-life drug discovery experiments, drug libraries are extremely large but still there is only a minor representation of the essentially infinite chemical space, and evaluation of physical properties for each molecule in the library is not computationally feasible. In the current study, a novel Machine learning framework for Enhanced MolEcular Screening (MEMES) based on Bayesian optimization is proposed for efficient sampling of the chemical space. The proposed framework is demonstrated to identify 90% of the top-1000 molecules from a molecular library of size about 100 million, while calculating the docking score only for about 6% of the complete library. We believe that such a framework would tremendously help to reduce the computational effort in not only drug-discovery but also areas that require such high-throughput experiments.

A novel machine learning framework based on Bayesian optimization for efficient sampling of chemical space. The framework is able to identify 90% of top-1000 hits by only sampling 6% of the complete dataset containing ∼100 million compounds.     

Prediction of cathodic E1/21 and E1/22 values for viologen-containing conjugated unimers and dimers from calculated pKb values of the aromatic substituents

A library of 18 conjugated, rod-like compounds with either one or two viologen residues were synthesized and characterized electrochemically. Containing up to 8 aromatic/heterocyclic rings in conjugation, the members of the library differ in the substitution pattern of electron-withdrawing or -donating groups on the aromatic substituents of the viologen units. The first and second half-wave potentials of each member were found to be linearly correlated with the calculated pK_b values of the aromatic end-groups. This relationship will enable the half-wave potentials of related, novel, substituted viologen species to be predicted using a simple, empirical formula.     

Palladium-catalyzed C3-selective mono-arylation of 4-hydroxycoumarin

4-Hydroxycoumarin (1) is a useful synthon in synthetic organic chemistry and a basic skeleton of several bioactive compounds. The direct functionalization of 1 with novel catalytic system would provide an efficient meaning for construction of a chemical library. Here, we developed a new catalytic system using palladium(II) acetate for the C3 regioselective mono-arylation of 1. The 3-mono-arylated and the 3,4-disubstituted coumarins were synthesized to enable derivatization. This palladium-catalyzed direct arylation of 1 is effective for rapid diversification and chemical library construction.     

Bioresorbable tyrosol-derived poly(ester-arylate)s with tunable properties

We designed and developed a novel library of tyrosol-derived poly(ester-arylate)s that exhibit tunable chemical, thermal, mechanical, and degradative properties. To build the library, the diphenols 4-hydroxyphenethyl 2-(4-hydroxyphenyl)acetate (HTy) and 4-hydroxyphenethyl 3-(4-hydroxyphenyl)propanoate (DTy) are synthesized and subsequently polymerized with various diacids. Characterization of library members is performed in order to assess the impact of chemical structure on polymer properties. Specifically, the relative influence of diphenol pseudosymmetry versus asymmetry, diacid carbon chain length, and diacid bond rigidity on resulting properties is investigated. Diphenol choice greatly impacts resulting polymer thermal properties and processability. HTy-containing polymers generally have lower melting temperatures compared to their DTy-derived counterparts and are easier to quench in the amorphous phase. As a result, processing results in greater tunability for HTy-derived polymers. One specific example was pHTy3, which increased its tensile modulus from 1 GPa to 3 GPa upon drawing. Diacid lengths and bond rigidity also significantly influence thermal, mechanical, and degradative properties. In all, members of this library can be synthesized efficiently, with high molecular weight and exhibit a wide range of properties, motivating future commercial translation.