Pyridine-based strategies towards nitrogen isotope exchange and multiple isotope incorporation.

Isotopic labeling is at the core of health and life science applications such as nuclear imaging, metabolomics and plays a central role in drug development. The rapid access to isotopically labeled organic molecules is a sine qua non condition to support these societally vital areas of research. Based on a rationally driven approach, this study presents an innovative solution to access labeled pyridines by a nitrogen isotope exchange reaction based on a Zincke activation strategy. The technology conceptualizes an opportunity in the field of isotope labeling. 15N-labeling of pyridines and other relevant heterocycles such as pyrimidines and isoquinolines showcases on a large set of derivatives, including pharmaceuticals. Finally, we explore a nitrogen-to-carbon exchange strategy in order to access 13C-labeled phenyl derivatives and deuterium labeling of mono-substituted benzene from pyridine-2H5. These results open alternative avenues for multiple isotope labeling on aromatic cores.

Easy-to-Implement Hydrogen Isotope Exchange for the Labeling of N-Heterocycles, Alkylkamines, Benzylic Scaffolds, and Pharmaceuticals.

Facilitating access to deuterated and tritiated complex molecules is of paramount importance due to the fundamental role of isotopically labeled compounds in drug discovery and development. Deuterated analogues of drugs are extensively used as internal standards for quantification purposes or as active pharmaceutical ingredients, whereas tritiated drugs are essential for preclinical ADME studies. In this report, we describe the labeling of prevalent substructures in FDA-approved drugs such as azines, indoles, alkylamine moieties, or benzylic carbons by the in situ generation of Rh nanoparticles able to catalyze both C(sp2)-H and C(sp3)-H activation processes. In this easy-to-implement labeling process, Rh nanocatalysts are formed by decomposition of a commercially available rhodium dimer under a deuterium or tritium gas atmosphere (1 bar or less), using the substrate itself as a surface ligand to control the aggregation state of the resulting metallic clusters. It is noteworthy that the size of the nanoparticles observed is surprisingly independent of the substrate used and is homogeneous, as evidenced by transmission electron microscopy experiments. This method has been successfully applied to the one-step synthesis of (1) deuterated pharmaceuticals usable as internal standards for MS quantification and (2) tritiated drug analogues with very high molar activities (up to 113 Ci/mmol).

Monitoring In Vivo Performances of Protein-Drug Conjugates Using Site-Selective Dual Radiolabeling and Ex Vivo Digital Imaging.

In preclinical models, the development and optimization of protein-drug conjugates require accurate determination of the plasma and tissue profiles of both the protein and its conjugated drug. To this aim, we developed a bioanalytical strategy based on dual radiolabeling and ex vivo digital imaging. By combining enzymatic and chemical reactions, we obtained homogeneous dual-labeled anti-MMP-14 Fabs (antigen-binding fragments) conjugated to monomethyl auristatin E where the protein scaffold was labeled with carbon-14 (14C) and the conjugated drug with tritium (3H). These antibody-drug conjugates with either a noncleavable or a cleavable linker were then evaluated in vivo. By combining liquid scintillation counting and ex vivo dual-isotope radio-imaging, it was possible not only to monitor both components simultaneously during their circulation phase but also to quantify accurately their amount accumulated within the different organs.

Gemcitabine lipid prodrug nanoparticles: Switching the lipid moiety and changing the fate in the bloodstream.

A simple approach to achieve a lipoprotein (LP)-mediated drug delivery is to trigger the spontaneous drug insertion into endogenous lipoproteins in the bloodstream, by means of its chemical modification. Nanoparticles (NPs) made of the squalene-gemcitabine (SQGem) conjugate were found to have a high affinity for plasma lipoproteins while free gemcitabine did not, suggesting a key role of the lipid moiety in this event. Whether the drug conjugation to cholesterol, one of the major lipoprotein-transported lipids, could also promote an analogous interaction was a matter of question. NPs made of the cholesterol-gemcitabine conjugate (CholGem) have been herein thoroughly investigated for their blood distribution profile both in vitro and in vivo. Unexpectedly, contrarily to SQGem, no trace of the CholGem prodrug could be found in the lipoprotein fractions, nor was it interacting with albumin. The investigation of isolated NPs and NPs/LPs physical mixtures provided a further insight into the lack of interaction of CholGem NPs with LPs. Although essential for allowing the self-assembly of the prodrug into nanoparticles, the lipid moiety may not be sufficient to elicit interaction of the conjugated drug with plasma lipoproteins but the whole NP physicochemical features must be carefully considered.

Nanocatalyzed Hydrogen Isotope Exchange.

Recently, hydrogen isotope exchange (HIE) reactions have experienced impressive development due to the growing importance of isotope containing compounds in various fields including materials and life sciences, in addition to their classical use for mechanistic studies in chemistry and biology. Tritium-labeled compounds are also of crucial interest to study the in vivo fate of a bioactive substance or in radioligand binding assays. Over the past few years, deuterium-labeled drugs have been extensively studied for the improvement of ADME (absorption, distribution, metabolism, excretion) properties of existing bioactive molecules as a consequence of the primary kinetic isotope effect. Furthermore, in the emergent "omic" fields, the need for new stable isotopically labeled internal standards (SILS) for quantitative GC- or LC-MS analyses is increasing. Because of their numerous applications, the development of powerful synthetic methods to access deuterated and tritiated molecules with either high isotope incorporation and/or selectivities is of paramount importance.HIE reactions allow a late-stage incorporation of hydrogen isotopes in a single synthetic step, thus representing an advantageous alternative to conventional multistep synthesis approaches which are time- and resource-consuming. Moreover, HIE reactions can be considered as the most fundamental C-H functionalization processes and are therefore of great interest for the chemists' community. Depending on the purpose, HIE reactions must either be highly regioselective or allow a maximal incorporation of hydrogen isotopes, sometimes both. In this context, metal-catalyzed HIE reactions are generally performed using either homogeneous or heterogeneous catalysis which may have considerable drawbacks including an insufficient isotope incorporation and a lack of chemo- and/or regioselectivity, respectively.Over the past 6 years, we have shown that nanocatalysis can be considered as a powerful tool to access complex labeled molecules (e.g., pharmaceuticals, peptides and oligonucleotides) via regio- and chemoselective or even enantiospecific labeling processes occurring at the surface of metallic nanoclusters (Ru or Ir). Numerous heterocyclic (both saturated and unsaturated) and acyclic scaffolds have been labeled with an impressive functional group tolerance, and highly deuterated compounds or high molar activity tritiated drugs have been obtained. An insight into mechanisms has also been provided by theoretical calculations to explain the regioselectivities of the isotope incorporation. Our studies have suggested that undisclosed key intermediates, including 4- and 5-membered dimetallacycles, account for the particular regioselectivities observed during the process, in contrast to the 5- or 6-membered metallacycle key intermediates usually encountered in homogeneous catalysis. These findings together with the important number of available coordination sites explain the compelling reactivity of metal nanoparticles, in between homogeneous and heterogeneous catalysis. They represent innovative tools combining the advantages of both methods for the isotopic labeling and activation of C-H bonds of complex molecules.

Chemical Synthesis of [2H]-Ethyl Tosylate and Exploration of Its Crypto-optically Active Character Combining Complementary Spectroscopic Tools.

Small chiral molecules are excellent candidates to push the boundaries of enantiodiscrimination analytical techniques. Here is reported the synthesis of two new deuterated chiral probes, (R)- and (S)-[2H]-ethyl tosylate, obtained with high enantiomeric excesses. Due to their crypto-optically active properties, the discrimination of each enantiomer is challenging. Whereas their enantiopurity is determined by 2H NMR in chiral anisotropic media, their identification was performed by combining quantum chemical calculations and vibrational circular dichroism analysis.

Tuning the Reactivity of a Heterogeneous Catalyst using N-Heterocyclic Carbene Ligands for C-H Activation Reactions.

We report the dramatic impact of the addition of N-heterocyclic carbenes (NHCs) on the reactivity and selectivity of heterogeneous Ru catalysts in the context of C-H activation reactions. Using a simple and robust method, we prepared a series of new air-stable catalysts starting from commercially available Ru on carbon (Ru/C) and differently substituted NHCs. Associated with C-H deuteration processes, depending on Ru/C-NHC ratios, the chemical outcome can be controlled to a large extent. Indeed, tuning the reactivity of the Ru catalyst with NHC enabled: 1) increased chemoselectivity and the regioselectivity for the deuteration of alcohols in organic media; 2) the synthesis of fragile pharmaceutically relevant deuterated heterocycles (azine, purine) that are otherwise completely reduced using unmodified commercial catalysts; 3) the discovery of a novel reactivity for such heterogeneous Ru catalysts, namely the selective C-1 deuteration of aldehydes.

Chemoselective H/D exchange catalyzed by nickel nanoparticles stabilized by N-heterocyclic carbene ligands.

With this work, we report the synthesis and full characterization of nickel nanoparticles (NPs) stabilized by N-heterocyclic carbene (NHC) ligands, namely 1,3-bis(cyclohexyl)-1,3-dihydro-2H-imidazol-2-ylidene (ICy) and 1,3-bis(2,4,6-trimethylphenyl)-1,3-dihydro-2H-imidazol-2-ylidene (IMes). Although the resulting NPs have the same size, they display different magnetic properties and different reactivities, which result from ligand effects. In the context of H/D exchange on pharmaceutically relevant heterocycles, Ni@NHC shows a high chemoselectivity, avoiding the formation of undesired reduced side-products and enabling a variety of H/D exchange on nitrogen-containing aromatic compounds. Using 2-phenylpyridine as a model substrate, it was observed that deuteration occurred preferably at the α position of the nitrogen atom, which is the most accessible position for the C-H activation. In addition, Ni@IMes NPs are also able to fully deuterate the ortho positions of the phenyl substituents.

Delivery of siRNA to Ewing Sarcoma Tumor Xenografted on Mice, Using Hydrogenated Detonation Nanodiamonds: Treatment Efficacy and Tissue Distribution.

Nanodiamonds of detonation origin are promising delivery agents of anti-cancer therapeutic compounds in a whole organism like mouse, owing to their versatile surface chemistry and ultra-small 5 nm average primary size compatible with natural elimination routes. However, to date, little is known about tissue distribution, elimination pathways and efficacy of nanodiamonds-based therapy in mice. In this report, we studied the capacity of cationic hydrogenated detonation nanodiamonds to carry active small interfering RNA (siRNA) in a mice model of Ewing sarcoma, a bone cancer of young adults due in the vast majority to the EWS-FLI1 junction oncogene. Replacing hydrogen gas by its radioactive analog tritium gas led to the formation of labeled nanodiamonds and allowed us to investigate their distribution throughout mouse organs and their excretion in urine and feces. We also demonstrated that siRNA directed against EWS-FLI1 inhibited this oncogene expression in tumor xenografted on mice. This work is a significant step to establish cationic hydrogenated detonation nanodiamond as an effective agent for in vivo delivery of active siRNA.

Formal enantioselective synthesis of nhatrangin A.

A new and straightforward synthesis of the C1-C7 core fragment of nhatrangin A was achieved in 14 steps from achiral 3-hydroxybenzaldehyde, without the need of chiral reagents or enzymatic resolution to introduce the chiral centers. The key asymmetric steps include in particular a highly enantioselective organocatalyzed Michael addition on an aryl vinyl ketone, a Sharpless asymmetric epoxidation and a subsequent regioselective ring opening of the resulting chiral epoxide. This work represents the first formal enantioselective synthesis of nhatrangin A.

Multiple Site Hydrogen Isotope Labelling of Pharmaceuticals.

Radiolabelling is fundamental in drug discovery and development as it is mandatory for preclinical ADME studies and late-stage human clinical trials. Herein, a general, effective, and easy to implement method for the multiple site incorporation of deuterium and tritium atoms using the commercially available and air-stable iridium precatalyst [Ir(COD)(OMe)]2 is described. A large scope of pharmaceutically relevant substructures can be labelled using this method including pyridine, pyrazine, indole, carbazole, aniline, oxa-/thia-zoles, thiophene, but also electron-rich phenyl groups. The high functional group tolerance of the reaction is highlighted by the labelling of a wide range of complex pharmaceuticals, containing notably halogen or sulfur atoms and nitrile groups. The multiple site hydrogen isotope incorporation has been explained by the in situ formation of complementary catalytically active species: monometallic iridium complexes and iridium nanoparticles.

NHC-Stabilized Iridium Nanoparticles as Catalysts in Hydrogen Isotope Exchange Reactions of Anilines.

The preparation of N-heterocyclic carbene-stabilized iridium nanoparticles and their application in hydrogen isotope exchange reactions is reported. These air-stable and easy-to-handle iridium nanoparticles showed a unique catalytic activity, allowing selective and efficient hydrogen isotope incorporation on anilines using D2 or T2 as isotopic source. The usefulness of this transformation has been demonstrated by the deuterium and tritium labeling of diverse complex pharmaceuticals.

Hydrogen Isotope Exchange Catalyzed by Ru Nanocatalysts: Labelling of Complex Molecules Containing N-Heterocycles and Reaction Mechanism Insights.

Ruthenium nanocatalysis can provide effective deuteration and tritiation of oxazole, imidazole, triazole and carbazole substructures in complex molecules using D2 or T2 gas as isotopic sources. Depending on the substructure considered, this approach does not only represent a significant step forward in practice, with notably higher isotope uptakes, a broader substrate scope and a higher solvent applicability compared to existing procedures, but also the unique way to label important heterocycles using hydrogen isotope exchange. In terms of applications, the high incorporation of deuterium atoms, allows the synthesis of internal standards for LC-MS quantification. Moreover, the efficacy of the catalyst permits, even under subatmospheric pressure of T2 gas, the preparation of complex radiolabeled drugs owning high molar activities. From a fundamental point of view, a detailed DFT-based mechanistic study identifying undisclosed key intermediates, allowed a deeper understanding of C-H (and N-H) activation processes occurring at the surface of metallic nanoclusters.

Using hydrogen isotope incorporation as a tool to unravel the surfaces of hydrogen-treated nanodiamonds.

We report here on a robust and easy-to-implement method for the labelling of detonation nanodiamonds (DND) with hydrogen isotopes (deuterium and tritium), using thermal annealing performed in a closed system. With this method, we have synthesized and fully characterized (FTIR, Raman, DLS, 3H/2H/1H and 13C MAS NMR) deuterium-treated and tritium-treated DND and demonstrated the usefulness of isotope incorporation in investigating the surface chemistry of such nanomaterials. For instance, surface treatment with deuterium coupled to FTIR spectroscopy allowed us to discriminate the origin of C-H terminations at the DND surface after the hydrogenation process. As a complementary, tritium appeared very useful for quantification purposes, while 1,2,3H NMR confirmed the nature of the C-1,2,3H bonds created. This isotopic study provides new insights into the characteristics of hydrogen-treated DND.

Efficient Access to Deuterated and Tritiated Nucleobase Pharmaceuticals and Oligonucleotides using Hydrogen-Isotope Exchange.

A general approach for the efficient hydrogen-isotope exchange of nucleobase derivatives is described. Catalyzed by ruthenium nanoparticles, using mild reaction conditions, and involving either D2 or T2 as isotopic sources, this reaction possesses a wide substrate scope and a high solvent tolerability. This novel method facilitates the access to essential diagnostic tools in drug discovery and development: tritiated pharmaceuticals with high specific activities and deuterated oligonucleotides suitable for use as internal standards during LC-MS quantification.

Ruthenium-catalyzed hydrogen isotope exchange of C(sp3)-H bonds directed by a sulfur atom.

We present here the first example of C(sp3)-H activation directed by a sulfur atom. Based on this transformation catalyzed by Ru/C, we have developed a hydrogen isotope exchange reaction for the deuterium and tritium labelling of thioether substructures in complex molecules.

Design and Synthesis of New Circularly Polarized Thermally Activated Delayed Fluorescence Emitters.

This work describes the first thermally activated delayed fluorescence material enabling circularly polarized light emission through chiral perturbation. These new molecular architectures obtained through a scalable one-pot sequential synthetic procedure at room temperature (83% yield) display high quantum yield (up to 74%) and circularly polarized luminescence with an absolute luminescence dissymmetry factor, |glum|, of 1.3 × 10(-3). These chiral molecules have been used as an emissive dopant in an organic light emitting diode exhibiting external quantum efficiency as high as 9.1%.

C-Glucosylated malonitrile as a key intermediate towards carbohydrate-based glycogen phosphorylase inhibitors.

Glycogen utilization involves glycogen phosphorylase, an enzyme which appears to be a potential target for the regulation of glycaemia, as the liver isoform is a major player for hepatic glucose output. A single C-glucosylated malonitrile allowed for the synthesis of three glucose-based derivatives namely bis-oxadiazoles, bis-amides and a C-glucosylated tetrahydropyrimidin-2-one. When evaluated as glycogen phosphorylase inhibitors, two of the synthesized compounds displayed inhibition in the sub-millimolar range. In silico studies revealed that only one out of the bis-amides obtained and the C-glucosylated tetrahydropyrimidin-2-one may bind at the catalytic site.