One Scaffold, Different Organelle Sensors: pH-Activable Fluorescent Probes for Targeting Live Microglial Cell Organelles.

Targeting live cell organelles is essential for imaging, understanding, and controlling specific biochemical processes. Typically, fluorescent probes with distinct structural scaffolds are used to target specific cell organelles. Here, we have designed a modular one-step synthetic strategy using a common reaction intermediate to develop new lysosomal, mitochondrial, and nucleus-targeting pH-activable fluorescent probes that are all based on a single boron dipyrromethane scaffold. The divergent cell organelle targeting was achieved by synthesizing probes with specific functional group changes to the central scaffold resulting in differential fluorescence and pKa . Specifically, we show that the functional group transformation of the same scaffold influences cellular localization and specificity of pH-activable fluorescent probes in live primary microglial cells with pKa values ranging from ∼3.2-6.0. We introduce a structure-organelle-relationship (SOR) framework to target nuclei (NucShine), lysosomes (LysoShine), and mitochondria (MitoShine) in live microglia. This work will result in future applications of SOR beyond imaging to target and control organelle-specific biochemical processes in disease-specific models.

Access to Imidazolidine-Fused Sulfamidates and Sulfamides Bearing a Quaternary Center via 1,3-Dipolar Cycloaddition of Nonstabilized Azomethine Ylides.

A 1,3-dipolar cycloaddition reaction of nonstabilized azomethine ylides and cyclic N-sulfonyl imines has been developed providing a workable access to imidazolidine-fused sulfamidates, sulfamides, and benzosultams bearing a quaternary center. Distinct from the available literature, this current work enables to make entry, for the first time, into the novel imidazolidine-fused sulfamidates and sulfamides. Furthermore, the selective imidazolidine ring opening accompanied by CH2 extrusion yielded tetra-substituted sulfamidates with an aminomethyl group. In addition, imidazolidine ring opening coupled with SO2 extrusion provided synthetically useful 1,2-diamines.

Intramolecular Acylation of Unactivated Pyridines or Arenes via Multiple C-H Functionalizations: Synthesis of All Four Azafluorenones and Fluorenones.

An unprecedented intramolecular acylation of unactivated pyridines via multiple C(sp3/sp2)-H functionalizations of a methyl, hydroxymethyl, or aldehyde group has been developed providing a general access to all four azafluorenones. The application of this protocol is further demonstrated to the synthesis of azafluorenone related fused nitrogen heterocycles and fluorenones. In addition, design and synthesis of a novel fluorene based organic emitter for potential use in organic light emitting devices (OLEDs) is also reported.

Intramolecular Minisci acylation under silver-free neutral conditions for the synthesis of azafluorenones and fluorenones.

Despite its synthetic potential, intramolecular acylation by the Minisci reaction remains unexplored. The development of a new intramolecular Minisci acylation under silver-free neutral conditions providing access to azafluorenones and fluorenones is described. Distinct from the current literature known approaches for Minisci acylation, the report described herein features a method that: (a) avoids the use of silver that is invariably used in Minisci acylation, (b) does not require any acidic conditions for the activation of pyridines, and

Implications of dynamic imine chemistry for the sustainable synthesis of nitrogen heterocycles via transimination followed by intramolecular cyclisation.

An exploration of a tandem approach to the sustainable synthesis of N-heterocycles from readily available N-aryl benzylamines or imines and ortho-substituted anilines is described, which demonstrates, for the first time, an important synthetic application of dynamic imine chemistry. The key features to the successful development of this protocol include the utilisation of N-aryl benzylamines as imine precursors in transimination, the occurrence of transimination in acetonitrile in the absence of any catalysts, an intramolecular nucleophilic addition occurring in the newly formed imine causing irreversible transimination, and the tandem event occurring under green conditions.

Palladium-catalyzed intramolecular oxidative coupling involving double C(sp(2))-H bonds for the synthesis of annulated biaryl sultams.

The palladium-catalyzed intramolecular oxidative coupling described herein involves a double C(sp(2))-H bond functionalization in sulfonanilides, providing a workable access to biaryl sultams annulated into a six-membered ring that are otherwise difficult to obtain by literature methods. The other synthetic applications of this protocol including the synthesis of biaryl sultams containing a seven-membered ring and analogous sultones are also presented.

Monitoring phagocytic uptake of amyloid ß into glial cell lysosomes in real time.

Phagocytosis by glial cells is essential to regulate brain function during health and disease. Therapies for Alzheimer's disease (AD) have primarily focused on targeting antibodies to amyloid ß (Aß) or inhibitng enzymes that make it, and while removal of Aß by phagocytosis is protective early in AD it remains poorly understood. Impaired phagocytic function of glial cells during later stages of AD likely contributes to worsened disease outcome, but the underlying mechanisms of how this occurs remain unknown. We have developed a human Aß1-42 analogue (AßpH) that exhibits green fluorescence upon internalization into the acidic organelles of cells but is non-fluorescent at physiological pH. This allowed us to image, for the first time, glial uptake of AßpH in real time in live animals. We find that microglia phagocytose more AßpH than astrocytes in culture, in brain slices and in vivo. AßpH can be used to investigate the phagocytic mechanisms responsible for removing Aß from the extracellular space, and thus could become a useful tool to study Aß clearance at different stages of AD.

Accelerated Reactivity Mechanism and Interpretable Machine Learning Model of N-Sulfonylimines toward Fast Multicomponent Reactions.

We introduce chemical reactivity flowcharts to help chemists interpret reaction outcomes using statistically robust machine learning models trained on a small number of reactions. We developed fast N-sulfonylimine multicomponent reactions for understanding reactivity and to generate training data. Accelerated reactivity mechanisms were investigated using density functional theory. Intuitive chemical features learned by the model accurately predicted heterogeneous reactivity of N-sulfonylimine with different carboxylic acids. Validation of the predictions shows that reaction outcome interpretation is useful for human chemists.

Spectral deep learning for prediction and prospective validation of functional groups.

State-of-the-art identification of the functional groups present in an unknown chemical entity requires the expertise of a skilled spectroscopist to analyse and interpret Fourier transform infra-red (FTIR), mass spectroscopy (MS) and/or nuclear magnetic resonance (NMR) data. This process can be time-consuming and error-prone, especially for complex chemical entities that are poorly characterised in the literature, or inefficient to use with synthetic robots producing molecules at an accelerated rate. Herein, we introduce a fast, multi-label deep neural network for accurately identifying all the functional groups of unknown compounds using a combination of FTIR and MS spectra. We do not use any database, pre-established rules, procedures, or peak-matching methods. Our trained neural network reveals patterns typically used by human chemists to identify standard groups. Finally, we experimentally validated our neural network, trained on single compounds, to predict functional groups in compound mixtures. Our methodology showcases practical utility for future use in autonomous analytical detection.

Rapid, Refined, and Robust Method for Expression, Purification, and Characterization of Recombinant Human Amyloid beta 1-42.

Amyloid plaques found in the brains of Alzheimer's disease patients primarily consists of amyloid beta 1-42 (Aß42). Commercially, Aß42 is synthesized using high-throughput peptide synthesizers resulting in the presence of impurities and the racemization of amino acids that affects its aggregation properties. Furthermore, the repeated purchase of even a small quantity (~1 mg) of commercial Aß42 can be expensive for academic researchers. Here, we describe a detailed methodology for robust expression of recombinant human Aß(M1-42) in Rosetta(DE3)pLysS and BL21(DE3)pLysS competent E. coli using standard molecular biology techniques with refined and rapid one-step analytical purification techniques. The peptide is isolated and purified from transformed cells using an optimized reverse-phase high-performance liquid chromatography (HPLC) protocol with commonly available C18 columns, yielding high amounts of peptide (~15-20 mg per 1 L culture) within a short period of time. The recombinant human Aß(M1-42) forms characteristic aggregates similar to synthetic Aß42 aggregates as verified by western blotting and atomic force microscopy to warrant future biological use. Our rapid, refined, and robust technique produces pure recombinant human Aß(M1-42) that may be used to synthesize chemical probes and in several downstream in vitro and in vivo assays to facilitate Alzheimer's disease research.

Scope of Successive C-H Functionalizations of the Methyl Group in 3-Picolines: Intramolecular Carbonylation of Arenes to the Metal-Free Synthesis of 4-Azafluorenones.

A transition-metal-free, t-BuOOH mediated intramolecular carbonylation of arenes in 2-aryl-3-picolines via oxidative C-H functionalizations of the methyl group has been developed, providing an expedient synthesis of 4-azafluorenones. Distinct from the current literature wherein methylarenes have been used as acylating agents, 2-aryl-3-picolines in this study are transformed into aldehydes, which give 4-azafluorenones upon rapid intramolecular acylation. The study demonstrates the first example of intramolecular carbonylation of arenes utilizing a methyl group as latent carbonyl functionality.

Access to biaryl sulfonamides by palladium-catalyzed intramolecular oxidative coupling and subsequent nucleophilic ring opening of heterobiaryl sultams with amines.

The installation of sulfonamide pharmacophores on heterobiaryls has successfully been executed by a previously unknown palladium-catalyzed intramolecular oxidative coupling in N-arylsulfonyl heterocycles followed by novel ring opening of heterobiaryl sultams with amine nucleophiles. The protocol has a wide scope of substrates warranting broad applications in the synthesis of heterobiaryls containing an o-sulfonyl or carboxyl functional group.

Palladium-catalyzed regio- and chemoselective ortho-benzylation of C-H bond using a functionalizable primary amide directing group: a concise synthesis of dibenzo[b,e]azepin-6-ones.

A palladium-catalyzed regio- and chemoselective direct benzylation of primary benzamides with 2-bromobenzyl bromides under a mild basic condition has been developed affording various substituted diarylmethanes in good yields. Furthermore, the directing amide group (-CONH2) was subjected to intramolecular N-arylation with the aryl bromide moiety present in diarylmethanes leading to a concise synthesis of dibenzoazepinones.