Development of setups for real-time analysis of the effluent of a microreactor by mass spectrometry.

Monitoring a synthesis reaction in real time could allow not only the detection of the intermediates involved in the synthesis, to better understand its mechanisms, but also the impurities. Spectroscopic methods could be performed but are not so performant when analyzing complex mixtures and could require specific properties for the detection of the molecules of interest, the presence of a chromophore moiety for example. Mass spectrometry (MS) may overcome these limitations and is able to reach the accuracy and sensitivity required to efficiently detect, quantify, identify, and characterize the reagents and species produced during the synthesis. This is why the hyphenation of a microreactor with MS has already allowed synthesis processes to be monitored, but most of the time it targets a specific reaction or compounds and involves solvents compatible with MS. In this study, a universal setup for the hyphenation of a microreactor with MS and based on two valves has been developed. This two-valve setup has proven itself for the analysis of molecules of different nature and hydrophilicity, soluble in a large number of solvents even in non-MS-compatible ones. The developed setup evidenced a good repeatability and a linear response for the detection of the studied compounds. In addition, the dilution step included in the two-valve setup allows the MS monitoring of compounds initially synthesized at different concentrations. Finally, it was successfully used to study an amination reaction allowing the detection of the reaction products in 4 min with good repeatability as RSD values of MS signals were lower than 17%.

Synthesis and multiparametric evaluation of thiadiazoles and oxadiazoles as diacylglycerol acyltransferase type 1 inhibitors.

Chemical modulation of a formerly disclosed DGAT-1 inhibitor resulted in the identification of a compound with a suitable profile for preclinical development. Optimisation of solubility is discussed and a PK/PD study is presented.

Discovery of SAR184841, a potent and long-lasting inhibitor of 11ß-hydroxysteroid dehydrogenase type 1, active in a physiopathological animal model of T2D.

Starting from 11ß-HSD1 inhibitors that were active ex vivo but with Cyp 3A4 liability, we obtained a new series of adamantane ureas displaying potent inhibition of both human and rodent 11ß-HSD1 enzymes, devoid of Cyp 3A4 interactions, and rationally designed to provide long-lasting inhibition in target tissues. Final optimizations lead to SAR184841 with good oral pharmacokinetic properties showing in vivo activity and improvement of metabolic parameters in a physiopathological model of type 2 diabetes.

Synthesis of trifluoromethyl N,N-aminals from nitrogen containing heterocycles by using a plasma flow microreactor.

Trifluoromethyl N,N-aminals are important precursors allowing access to fluorinated building blocks. In this work, the direct synthesis of trifluoromethyl N,N-aminals from nitrogen containing heterocycles is reported using argon plasma in a continuous flow microreactor without any additives or metal catalysts. Their transformation to N-trifluoroethyl amines is also reported.

TEAD Inhibitors Sensitize KRASG12C Inhibitors via Dual Cell Cycle Arrest in KRASG12C-Mutant NSCLC.

KRASG12C is one of the most common mutations detected in non-small cell lung cancer (NSCLC) patients, and it is a marker of poor prognosis. The first FDA-approved KRASG12C inhibitors, sotorasib and adagrasib, have been an enormous breakthrough for patients with KRASG12C mutant NSCLC; however, resistance to therapy is emerging. The transcriptional coactivators YAP1/TAZ and the family of transcription factors TEAD1-4 are the downstream effectors of the Hippo pathway and regulate essential cellular processes such as cell proliferation and cell survival. YAP1/TAZ-TEAD activity has further been implicated as a mechanism of resistance to targeted therapies. Here, we investigate the effect of combining TEAD inhibitors with KRASG12C inhibitors in KRASG12C mutant NSCLC tumor models. We show that TEAD inhibitors, while being inactive as single agents in KRASG12C-driven NSCLC cells, enhance KRASG12C inhibitor-mediated anti-tumor efficacy in vitro and in vivo. Mechanistically, the dual inhibition of KRASG12C and TEAD results in the downregulation of MYC and E2F signatures and in the alteration of the G2/M checkpoint, converging in an increase in G1 and a decrease in G2/M cell cycle phases. Our data suggest that the co-inhibition of KRASG12C and TEAD leads to a specific dual cell cycle arrest in KRASG12C NSCLC cells.

Thiadiazoles as new inhibitors of diacylglycerol acyltransferase type 1.

A novel class of DGAT1 inhibitors containing a thiadiazole core has been discovered. Chemical optimization lead to inhibitors of human DGAT1 with an appropriate ADME profile and that show in vivo activity in target tissues.

Plasma flow chemistry for direct N-acylation of amines by esters.

The direct N-acylation of amines by esters in a microreactor is described using argon plasma, thereby producing amides in good to excellent yields.

Pyrrolidine-pyrazole ureas as potent and selective inhibitors of 11ß-hydroxysteroid-dehydrogenase type 1.

A High Throughput Screening campaign allowed the identification of a novel class of ureas as 11ß-HSD1 inhibitors. Rational chemical optimization provided potent and selective inhibitors of both human and murine 11ß-HSD1 with an appropriate ADME profile and ex vivo activity in target tissues.

Recent Therapeutic Approaches to Modulate the Hippo Pathway in Oncology and Regenerative Medicine.

The Hippo pathway is an evolutionary conserved signaling network that regulates essential processes such as organ size, cell proliferation, migration, stemness and apoptosis. Alterations in this pathway are commonly found in solid tumors and can lead to hyperproliferation, resistance to chemotherapy, compensation for mKRAS and tumor immune evasion. As the terminal effectors of the Hippo pathway, the transcriptional coactivators YAP1/TAZ and the transcription factors TEAD1-4 present exciting opportunities to pharmacologically modulate the Hippo biology in cancer settings, inflammation and regenerative medicine. This review will provide an overview of the progress and current strategies to directly and indirectly target the YAP1/TAZ protein-protein interaction (PPI) with TEAD1-4 across multiple modalities, with focus on recent small molecules able to selectively bind to TEAD, block its autopalmitoylation and inhibit YAP1/TAZ-TEAD-dependent transcription in cancer.

High-Throughput Cellular RNA Sequencing (HiCAR-Seq): Cost-Effective, High-Throughput 3' mRNA-Seq Method Enabling Individual Sample Quality Control.

High-throughput screening is one of the pillars of drug development. Unbiased transcriptome profiling is now widely used for a deeper understanding of a drug's mechanisms of action, off target effects, and cytotoxicity. Although currently available high-throughput RNA-Seq (HT RNA-Seq) methods such as PLATE-Seq, DRUG-Seq, and BRB-Seq serve these purposes, the inherent nature of these methods does not allow sample-wise sequencing library quality control. Here, we describe an HTR method called High-throughput CellulAr RNA Sequencing (HiCAR-Seq). HiCAR-Seq was optimized to work directly on cultured cells (as little as 1,000 cells) or 10 ng of total RNA. HiCAR-Seq involves reverse transcription from cultured cells or total RNA using oligo-dT primers followed by the PCR amplification of full-length cDNAs using sample-specific barcode primers in individual plate wells. Amplification of cDNA from every sample can be verified using Bioanalyzer. This step not only reveals cDNA amplification but also provides greater precision for pooling equal concentrations of cDNA from different samples. A single pooled cDNA library is made suitable for sequencing on Illumina sequencers using a tagmentation kit. Because HiCAR-Seq targets a small region at the 3' of the mRNAs, as little as 3 to 4 million reads/sample are enough to infer changes in gene expression in human or mouse cells. We believe that HiCAR-Seq represents a robust and competitive addition to the existing set of transcriptome-based high-throughput screening methods. © 2020 Wiley Periodicals LLC. Basic Protocol 1: cDNA synthesis and barcoding/enrichment PCR Basic Protocol 2: Nextera tagmentation/amplification, quantification, and sequencing.

Regio- and Stereocontrolled Formation of Chiral Epoxy Oxazolidines via Bromocarbamation of N-Boc Alkenyl Oxazolidines. Application to Asymmetric Synthesis.

Treatment of alpha-alkenyl N-Boc oxazolidines with N-bromosuccinimide leads to epoxy oxazolidines via a bromocyclocarbamation reaction which is completely stereoselective. Action of sodium azide on these epoxides, followed by a few functional group manipulations, eventually affords chiral beta-amino alcohols which are intermediates for the enantioselective synthesis of bioactive products: the anti side chain of taxol and a hydroxyethylamine isostere. Both the bromocarbamation cyclization and the nucleophilic cleavage of epoxides are totally regioselective. AM1 calculations suggest that this selectivity is controlled by the positive charge distribution at the electrophilic centers.