A tau homeostasis signature is linked with the cellular and regional vulnerability of excitatory neurons to tau pathology.

Excitatory neurons are preferentially impaired in early Alzheimer's disease but the pathways contributing to their relative vulnerability remain largely unknown. Here we report that pathological tau accumulation takes place predominantly in excitatory neurons compared to inhibitory neurons, not only in the entorhinal cortex, a brain region affected in early Alzheimer's disease, but also in areas affected later by the disease. By analyzing RNA transcripts from single-nucleus RNA datasets, we identified a specific tau homeostasis signature of genes differentially expressed in excitatory compared to inhibitory neurons. One of the genes, BCL2-associated athanogene 3 (BAG3), a facilitator of autophagy, was identified as a hub, or master regulator, gene. We verified that reducing BAG3 levels in primary neurons exacerbated pathological tau accumulation, whereas BAG3 overexpression attenuated it. These results define a tau homeostasis signature that underlies the cellular and regional vulnerability of excitatory neurons to tau pathology

Streamlining bioactive molecular discovery through integration and automation

The discovery of bioactive small molecules is generally driven via iterative design–make–purify–test cycles. Automation is routinely harnessed at individual stages of these cycles to increase the productivity of drug discovery. Here, we describe recent progress to automate and integrate two or more adjacent stages within discovery workflows. Examples of such technologies include microfluidics, liquid-handling robotics and affinity-selection mass spectrometry. The value of integrated technologies is illustrated in the context of specific case studies in which modulators of targets, such as protein kinases, nuclear hormone receptors and protein–protein interactions, were discovered. We note that to maximize impact on the productivity of discovery, each of the integrated stages would need to have both high and matched throughput. We also consider the longer-term goal of realizing the fully autonomous discovery of bioactive small molecules through the integration and automation of all stages of discovery

Synthetic Studies Directed toward Dideoxy Lomaiviticinone Lead to Unexpected 1,2-Oxazepine and Isoxazole Formation

In the course of studies directed toward the synthesis of dideoxy lomaiviticinone, 3-(nitromethyl)cyclohexenones <b>2a</b> (X = H) and <b>2b</b> (X = I) were prepared. The corresponding enolates were reacted with naphthazarin (<b>1</b>) and unexpectedly afforded 1,2-oxazepine <b>3</b> and isoxazole <b>4</b>, respectively. Rationale for their formation is proposed

Integrated Platform for Expedited Synthesis–Purification–Testing of Small Molecule Libraries

The productivity of medicinal chemistry programs can be significantly increased through the introduction of automation, leading to shortened discovery cycle times. Herein, we describe a platform that consolidates synthesis, purification, quantitation, dissolution, and testing of small molecule libraries. The system was validated through the synthesis and testing of two libraries of binders of polycomb protein EED, and excellent correlation of obtained data with results generated through conventional approaches was observed. The fully automated and integrated platform enables batch-supported compound synthesis based on a broad array of chemical transformations with testing in a variety of biochemical assay formats. A library turnaround time of between 24 and 36 h was achieved, and notably, each library synthesis produces sufficient amounts of compounds for further evaluation in secondary assays thereby contributing significantly to the shortening of medicinal chemistry discovery cycles

A Fluorogenic Aryl Fluorosulfate for Intraorganellar Transthyretin Imaging in Living Cells and in <i>Caenorhabditis elegans</i>

Fluorogenic probes, due to their often greater spatial and temporal sensitivity in comparison to permanently fluorescent small molecules, represent powerful tools to study protein localization and function in the context of living systems. Herein, we report fluorogenic probe <b>4</b>, a 1,3,4-oxadiazole designed to bind selectively to transthyretin (TTR). Probe <b>4</b> comprises a fluorosulfate group not previously used in an environment-sensitive fluorophore. The fluorosulfate functional group does not react covalently with TTR on the time scale required for cellular imaging, but does red shift the emission maximum of probe <b>4</b> in comparison to its nonfluorosulfated analogue. We demonstrate that probe <b>4</b> is dark in aqueous buffers, whereas the TTR·<b>4</b> complex exhibits a fluorescence emission maximum at 481 nm. The addition of probe <b>4</b> to living HEK293T cells allows efficient binding to and imaging of exogenous TTR within intracellular organelles, including the mitochondria and the endoplasmic reticulum. Furthermore, live <i>Caenorhabditis elegans</i> expressing human TTR transgenically and treated with probe <b>4</b> display TTR·<b>4</b> fluorescence in macrophage-like coelomocytes. An analogue of fluorosulfate probe <b>4</b> does react selectively with TTR without labeling the remainder of the cellular proteome. Studies on this analogue suggest that certain aryl fluorosulfates, due to their cell and organelle permeability and activatable reactivity, could be considered for the development of protein-selective covalent probes