Defect-free graphene enhances enzyme delivery to fibroblasts derived from patients with lysosomal storage disorders.

Enzyme replacement therapy shows remarkable clinical improvement in treating lysosomal storage disorders. However, this therapeutic approach is hampered by limitations in the delivery of the enzyme to cells and tissues. Therefore, there is an urgent, unmet clinical need to develop new strategies to enhance the enzyme delivery to diseased cells. Graphene-based materials, due to their dimensionality and favourable pattern of interaction with cells, represent a promising platform for the loading and delivery of therapeutic cargo. Herein, the potential use of graphene-based materials, including defect-free graphene with positive or negative surface charge and graphene oxide with different lateral dimensions, was investigated for the delivery of lysosomal enzymes in fibroblasts derived from patients with Mucopolysaccharidosis VI and Pompe disease. We report excellent biocompatibility of all graphene-based materials up to a concentration of 100 µg mL-1 in the cell lines studied. In addition, a noticeable difference in the uptake profile of the materials was observed. Neither type of graphene oxide was taken up by the cells to a significant extent. In contrast, the two types of graphene were efficiently taken up, localizing in the lysosomes. Furthermore, we demonstrate that cationic graphene flakes can be used as carriers for arylsulfatase B enzyme, for the delivery of the lacking enzyme to the lysosomes of Mucopolysaccharidosis VI fibroblasts. Arylsulfatase B complexed with cationic graphene flakes not only retained the enzymatic activity, but also exerted biological effects almost twice as high as arylsulfatase B alone in the clearance of the substrate in Mucopolysaccharidosis VI fibroblasts. This study lays the groundwork for the potential use of graphene-based materials as carriers for enzyme replacement therapy in lysosomal storage disorders.

Effects of Lateral Size, Thickness, and Stabilizer Concentration on the Cytotoxicity of Defect-Free Graphene Nanosheets: Implications for Biological Applications.

In this work, we apply liquid cascade centrifugation to highly concentrated graphene dispersions produced by liquid-phase exfoliation in water with an insoluble bis-pyrene stabilizer to obtain fractions containing nanosheets with different lateral size distributions. The concentration, stability, size, thickness, and the cytotoxicity profile are studied as a function of the initial stabilizer concentration for each fraction. Our results show that there is a critical initial amount of stabilizer (0.4 mg/mL) above which the dispersions show reduced concentration, stability, and biocompatibility, no matter the lateral size of the flakes.

Triflic acid-mediated N-heteroannulation of ß-anilino-ß-(methylthio)acrylonitriles: a facile synthesis of 4-amino-2-(methylthio)quinolines.

Various functionalised 4-amino-2-(methylthio)quinolines are synthesised through triflic acid-mediated N-heteroannulation of α-functionalized-ß-anilino-ß-(methylthio)acrylonitriles for the first time. The N-heteroannulation process is highly chemoselective and has mild reaction conditions. However, this process fails in the absence of the ß-methylthio group in the acrylonitriles. In addition, a new double N-heteroannulation process is demonstrated to synthesise indolo[3,2-c]quinolines from non-heterocyclic precursors. Natural product isocryptolepine is synthesised in four steps from an acyclic precursor.

Enhanced liquid phase exfoliation of graphene in water using an insoluble bis-pyrene stabiliser.

Stabilisers, such as surfactants, polymers and polyaromatic molecules, offer an effective way to produce graphene dispersions in water by Liquid Phase Exfoliation (LPE) without degrading the properties of graphene. In particular, pyrene derivatives provide better exfoliation efficiency than traditional surfactants and polymers. A stabiliser is expected to be relatively soluble in order to disperse hydrophobic graphene in water. Here, we show that exfoliation can also be achieved with insoluble pyrene stabilisers if appropriately designed. In particular, bis-pyrene stabilisers (BPSs) functionalised with pyrrolidine provide a higher exfoliation efficiency and percentage of single layers compared to traditional pyrene derivatives under the same experimental conditions. This is attributed to the enhanced interactions between BPS and graphene, provided by the presence of two pyrene binding groups. This approach is therefore attractive not only to produce highly concentrated graphene, but also to use graphene to disperse insoluble molecules in water. The enhanced adsorption of BPS on graphene, however, is reflected in higher toxicity towards human epithelial bronchial immortalized cells, limiting the use of this material for biomedical applications.

Transition metal-free cross-dehydrogenative arylation of unactivated benzylic C-H bonds.

The cross-dehydrogenative arylation of benzylic C-H bonds with arenes provides straightforward access to synthetically useful 1,1-diarylmethanes, from readily available starting materials. Current approaches suffer from limited substrate scope, requirement for large excesses of alkyl arene and/or non-trivial reaction set up. We report a transition metal-free cross-dehydrogenative arylation of benzylic C-H bonds using alkyl benzene derivatives and electron-rich arenes as coupling partners. The method proceeds through the in situ generation of a reactive benzyl fluoride intermediate which then reacts with the nucleophilic arene. The reaction tolerates a wide variety of functional groups including unprotected polar functionality and has been applied to the late-stage benzylation of several biologically relevant molecules.

Ag/Pd Cocatalyzed Direct Arylation of Fluoroarene Derivatives with Aryl Bromides.

Diverse C-H functionalizations catalyzed by Pd employ Ag(I) salts added as halide abstractors or oxidants. Recent reports have shown that Ag can also perform the crucial C-H activation step in several of these functionalizations. However, all of these processes are limited by the wasteful requirement for (super)stoichiometric Ag(I) salts. Herein, we report the development of a Ag/Pd cocatalyzed direct arylation of (fluoroarene) chromium tricarbonyl complexes with bromoarenes. The small organic salt, NMe4OC(CF3)3, added as a halide abstractor, enables the use of a catalytic amount of Ag, reversing the rapid precipitation of AgBr. We have shown through H/D scrambling and kinetic studies that a (PR3)Ag-alkoxide is responsible for C-H activation, a departure from previous studies with Ag carboxylates. Furthermore, the construction of biaryls directly from the simple arene is achieved via a one-pot chromium tricarbonyl complexation/C-H arylation/decomplexation sequence using (pyrene)Cr(CO)3 as a Cr(CO)3 donor.

Palladium catalysed C-H arylation of pyrenes: access to a new class of exfoliating agents for water-based graphene dispersions.

A new and diverse family of pyrene derivatives was synthesised via palladium-catalysed C-H ortho-arylation of pyrene-1-carboxylic acid. The strategy affords easy access to a broad scope of 2-substituted and 1,2-disubstituted pyrenes. The C1-substituent can be easily transformed into carboxylic acid, iodide, alkynyl, aryl or alkyl functionalities. This approach gives access to arylated pyrene ammonium salts, which outperformed their non-arylated parent compound during aqueous Liquid Phase Exfoliation (LPE) of graphite and compare favourably to state-of-the-art sodium pyrene-1-sulfonate PS1. This allowed the production of concentrated and stable suspensions of graphene flakes in water.

Ag(I)-C-H Activation Enables Near-Room-Temperature Direct α-Arylation of Benzo[ b]thiophenes.

The first example of near-room-temperature α-arylation of benzo[ b]thiophenes is reported. The discovery rests on the observation of a switch in α-/ß-regioselectivity at different loadings of Pd2(dba)3·CHCl3 in the coupling between benzo[ b]thiophene and 4-iodotoluene. We show that this unprecedented regioselectivity switch is driven by a Ag(I)-mediated C-H activation at the α-C-H position, which becomes the dominant mode of reactivity at low concentrations of Pd. Competition experiments, kinetic studies, KIE, and D/H scrambling experiments have been carried out supporting this mechanism.

Transition-Metal-Free Decarboxylative Iodination: New Routes for Decarboxylative Oxidative Cross-Couplings.

Constructing products of high synthetic value from inexpensive and abundant starting materials is of great importance. Aryl iodides are essential building blocks for the synthesis of functional molecules, and efficient methods for their synthesis from chemical feedstocks are highly sought after. Here we report a low-cost decarboxylative iodination that occurs simply from readily available benzoic acids and I2. The reaction is scalable and the scope and robustness of the reaction is thoroughly examined. Mechanistic studies suggest that this reaction does not proceed via a radical mechanism, which is in contrast to classical Hunsdiecker-type decarboxylative halogenations. In addition, DFT studies allow comparisons to be made between our procedure and current transition-metal-catalyzed decarboxylations. The utility of this procedure is demonstrated in its application to oxidative cross-couplings of aromatics via decarboxylative/C-H or double decarboxylative activations that use I2 as the terminal oxidant. This strategy allows the preparation of biaryls previously inaccessible via decarboxylative methods and holds other advantages over existing decarboxylative oxidative couplings, as stoichiometric transition metals are avoided.

Ruthenium-Catalyzed C-H Arylation of Benzoic Acids and Indole Carboxylic Acids with Aryl Halides.

Herein we report the first Ru-catalyzed C-H arylation of benzoic acids with readily available aryl (pseudo)halides. The reaction, which does not require the use of silver salt additives, allows the arylation of previously challenging hindered benzoic acids and the use of generally unreactive ortho-substituted halorarenes. Furthermore, our new protocol can efficiently be applied to indole carboxylic acids, thus allowing access to C7-, C6-, C5- and C4-arylated indole compounds, a departure from the classical enhanced reactivity of the C2 and C3 positions of indole.