Gediminas Mikutis

SnAP reagents for the transformation of aldehydes into substituted thiomorpholines--an alternative to cross-coupling with saturated heterocycles.

Saturated N-heterocycles are increasingly common scaffolds for the discovery and development of biologically active small molecules. Unlike their aromatic counterparts, they cannot be easily appended to a substrate by cross-coupling reactions. The most reliable method for constructing 2-aryl piperidines and pyrrolidines is an effective but laborious lithiation, transmetallation, and Pd-catalyzed cross-coupling. Unfortunately, this method is not well-suited for cross-coupling morpholines, thiomorpholines, or piperazines. Researchers have therefore sought to identify new synthetic methods for the preparation of substituted, saturated N-heterocycles by C H functionalization. For example, the groups of Nakamura, Sames, MacMillan, Maulide, and others have reported a-arylation of N-heterocycles, but these reactions require N-aryl substituents that are difficult to remove or further elaborate. Herein, we disclose SnAP reagents for the facile conversion of aldehydes into N-unprotected, 3-thiomorpholines (Figure 1). This strategy has the potential to be a general approach to installing saturated heterocycles using aldehydes as a synthetic handle. Its successful execution relies on the use of radical chemistry to overcome a long-standing challenge in organic synthesis: C C bond-forming addition to unactivated primary imines. At the outset of our studies, we established two key design parameters for reaction development. First, we required that our approach deliver N-unprotected heterocycles directly from the reaction. Second, we wanted the N-heterocycle substituents to be derived from a stable, widely available functional group, ideally halide, aldehyde, or organoboronic acid. After several unsuccessful attempts, our studies led us to an unusual approach: intramolecular C C bond-forming radical cyclization of an imine bearing a pendant nucleophilic carbon. For our initial studies, we selected the preparation of 3-substituted thiomorpholines from aldehydes, as there are presently very few methods for their synthesis. Amino tributylstannane 1 (SnAP-TM) is readily obtained in one step by S-alkylation of 2-aminoethanethiol with commercially available tributyl(iodomethyl)-stannane. Condensation with an aldehyde gives the corresponding imine, which we endeavored to cyclize to give N-unprotected 3-thiomorpholines. The poor electrophilicity of this imine, which lacks either the N-sulfonyl or N-aryl group typically found in imine electrophiles, presented numerous challenges. Initial attempts with stannane–lithium exchange or Lewis acid induced cyclization proved unsuccessful (Table 1, entry 1). Inspired by reports from the group of Kagoshima on the intermolecular addition of organostannanes to N-phenyl or N-paramethoxyphenyl imines using stoichiometric Cu(OTf)2, [12] we Figure 1. Approaches to cross-coupling with saturated heterocycles. Boc= tert-butoxycarbonyl, Fg= functional group.

Systematic evaluation and optimization of modification reactions of oligonucleotides with amines and carboxylic acids for the synthesis of dna-encoded chemical libraries

DNA-encoded chemical libraries are collections of small molecules, attached to DNA fragments serving as identification barcodes, which can be screened against multiple protein targets, thus facilitating the drug discovery process. The preparation of large DNA-encoded chemical libraries crucially depends on the availability of robust synthetic methods, which enable the efficient conjugation to oligonucleotides of structurally diverse building blocks, sharing a common reactive group. Reactions of DNA derivatives with amines and/or carboxylic acids are particularly attractive for the synthesis of encoded libraries, in view of the very large number of building blocks that are commercially available. However, systematic studies on these reactions in the presence of DNA have not been reported so far. We first investigated conditions for the coupling of primary amines to oligonucleotides, using either a nucleophilic attack on chloroacetamide derivatives or a reductive amination on aldehyde-modified DNA. While both methods could be used for the production of secondary amines, the reductive amination approach was generally associated with higher yields and better purity. In a second endeavor, we optimized conditions for the coupling of a diverse set of 501 carboxylic acids to DNA derivatives, carrying primary and secondary amine functions. The coupling efficiency was generally higher for primary amines, compared to secondary amine substituents, but varied considerably depending on the structure of the acids and on the synthetic methods used. Optimal reaction conditions could be found for certain sets of compounds (with conversions >80%), but multiple reaction schemes are needed when assembling large libraries with highly diverse building blocks. The reactions and experimental conditions presented in this article should facilitate the synthesis of future DNA-encoded chemical libraries, while outlining the synthetic challenges that remain to be overcome.

Tankyrase 1 Inhibitors with Drug-like Properties Identified by Screening a DNA-Encoded Chemical Library

We describe the synthesis and screening of a DNA-encoded chemical library containing 76230 compounds. In this library, sets of amines and carboxylic acids are directly linked producing encoded compounds with compact structures and drug-like properties. Affinity screening of this library yielded inhibitors of the potential pharmaceutical target tankyrase 1, a poly(ADP-ribose) polymerase. These compounds have drug-like characteristics, and the most potent hit compound (X066/Y469) inhibited tankyrase 1 with an IC50 value of 250 nM.

Cap-and-Catch Purification for Enhancing the Quality of Libraries of DNA Conjugates.

The potential of DNA-encoded combinatorial libraries (DECLs) as tools for hit discovery crucially relies on the availability of methods for their synthesis at acceptable purity and quality. Incomplete reactions in the presence of DNA can noticeably affect the purity of DECLs and methods to selectively remove unreacted oligonucleotide-based starting products would likely enhance the quality of DECL screening results. We describe an approach to selectively remove unreacted oligonucleotide starting products from reaction mixtures and demonstrate its applicability in the context of acylation of amino-modified DNA. Following an amide bond forming reaction, we treat unreacted amino-modified DNAs with biotinylating reagents and isolate the corresponding biotinylated oligonucleotides from the reaction mixture by affinity capture on streptavidin-coated sepharose. This approach, which yields the desired DNA-conjugate at enhanced purity, can be applied both to reactions performed in solution and to procedures in which DNA is immobilized on an anion exchange solid support.

Submicrometer-Sized Thermometer Particles Exploiting Selective Nucleic Acid Stability

Encapsulated nucleic acid selective damage quantification by real-time polymerase chain reaction is used as sensing mechanism to build a novel class of submicrometer size thermometer. Thanks to the high thermal and chemical stability, and the capability of storing the read accumulated thermal history, the sensor overcomes some of current limitations in small scale thermometry.

Silica-Encapsulated DNA-Based Tracers for Aquifer Characterization

Environmental tracing is a direct way to characterize aquifers, evaluate the solute transfer parameter in underground reservoirs, and track contamination. By performing multitracer tests, and translating the tracer breakthrough times into tomographic maps, key parameters such as a reservoirs effective porosity and permeability field may be obtained. DNA, with its modular design, allows the generation of a virtually unlimited number of distinguishable tracers. To overcome the insufficient DNA stability due to microbial activity, heat, and chemical stress, we present a method to encapsulated DNA into silica with control over the particle size. The reliability of DNA quantification is improved by the sample preservation with NaN3 and particle redispersion strategies. In both sand column and unconsolidated aquifer experiments, DNA-based particle tracers exhibited slightly earlier and sharper breakthrough than the traditional solute tracer uranine. The reason behind this observation is the size exclusion effect, whereby larger tracer particles are excluded from small pores, and are therefore transported with higher average velocity, which is pore size-dependent. Identical surface properties, and thus flow behavior, makes the new material an attractive tracer to characterize sandy groundwater reservoirs or to track multiple sources of contaminants with high spatial resolution.