Preparation of 4 '-spirocyclobutyl nucleoside analogues as novel and versatile adenosine scaffolds

Despite the large variety of modified nucleosides that have been reported, the preparation of constrained 4 '-spirocyclic adenosine analogues has received very little attention. We discovered that the [2+2]-cycloaddition of dichloroketene on readily available 4 '-exo-methylene furanose sugars efficiently results in the diastereoselective formation of novel 4 '-spirocyclobutanones. The reaction mechanism was investigated via density functional theory (DFT) and found to proceed either via a non-synchronous or stepwise reaction sequence, controlled by the stereochemistry at the 3 '-position of the sugar substrate. The obtained dichlorocyclobutanones were converted into nucleoside analogues, providing access to a novel class of chiral 4 '-spirocyclobutyl adenosine mimetics in eight steps from commercially available sugars. Assessment of the biological activity of designed 4 '-spirocyclic adenosine analogues identified potent inhibitors for protein methyltransferase target PRMT5

Comparison of Small Gut and Whole Gut Microbiota of First-Degree Relatives With Adult Celiac Disease Patients and Controls

Recent studies on celiac disease (CeD) have reported alterations in the gut microbiome. Whether this alteration in the microbial community is the cause or effect of the disease is not well understood, especially in adult onset of disease. The first-degree relatives (FDRs) of CeD patients may provide an opportunity to study gut microbiome in pre-disease state as FDRs are genetically susceptible to CeD. By using 16S rRNA gene sequencing, we observed that ecosystem level diversity measures were not significantly different between the disease condition (CeD), pre-disease (FDR) and control subjects. However, differences were observed at the level of amplicon sequence variant (ASV), suggesting alterations in specific ASVs between pre-disease and diseased condition. Duodenal biopsies showed higher differences in ASVs compared to fecal samples indicating larger disruption of the microbiota at the disease site. The duodenal microbiota of FDR was characterized by significant abundance of ASVs belonging to Parvimonas, Granulicatella, Gemella, Bifidobacterium, Anaerostipes, and Actinomyces genera. The duodenal microbiota of CeD was characterized by higher abundance of ASVs from genera Megasphaera and Helicobacter compared to the FDR microbiota. The CeD and FDR fecal microbiota had reduced abundance of ASVs classified as Akkermansia and Dorea when compared to control group microbiota. In addition, predicted functional metagenome showed reduced ability of gluten degradation by CeD fecal microbiota in comparison to FDRs and controls. The findings of the present study demonstrate differences in ASVs and predicts reduced ability of CeD fecal microbiota to degrade gluten compared to the FDR fecal microbiota. Further research is required to investigate the strain level and active functional profiles of FDR and CeD microbiota to better understand the role of gut microbiome in pathophysiology of CeD

Preparation of novel 4 '-spirocyclopropyl nucleoside analogues

The stereoselective preparation of a novel 4 '-spirocyclopropyl nucleoside analogue has been developed by using a semibenzilic Favorskii rearrangement of a 4 '-(2-chloro-3-oxocyclobutyl)spirofuranose as a key step. These chiral spirocyclic intermediates, readily obtained on a multigram scale from chiral-pool starting materials, were shown to be highly suitable precursors for achieving full stereoselectivity in the reduction-ring contraction sequence. The downstream introduction of a nucleobase through Vorbruggen glycosylation successfully resulted in the formation of the corresponding novel 4 '-spirocyclic nucleoside analogue in a stereospecific manner

Alchemical Free Energy Calculations on Membrane-Associated Proteins

<p>Membrane proteins have diverse functions within cells and are well-established drug targets. The advances in membrane protein structural biology have revealed drug and lipid binding sites on membrane proteins, while computational methods such as molecular simulations can resolve the thermodynamic basis of these interactions. Particularly, alchemical free energy calculations have shown promise in the calculation of reliable and reproducible binding free energies of protein–ligand and protein–lipid complexes in membrane-associated systems. In this review, we present an overview of representative alchemical free energy studies on G-protein-coupled receptors, ion channels, transporters as well as protein–lipid interactions, with emphasis on best practices and critical aspects of running these simulations. Additionally, we analyze challenges and successes when running alchemical free energy calculations on membrane-associated proteins. Finally, we highlight the value of alchemical free energy calculations calculations in drug discovery and their applicability in the pharmaceutical industry.</p&gt

Structure-function defects of the twinkle amino-terminal region in progressive external ophthalmoplegia

AbstractTWINKLE is a DNA helicase needed for mitochondrial DNA replication. In lower eukaryotes the protein also harbors a primase activity, which is lost from TWINKLE encoded by mammalian cells. Mutations in TWINKLE underlie autosomal dominant progressive external ophthalmoplegia (adPEO), a disorder associated with multiple deletions in the mtDNA. Four different adPEO-causing mutations (W315L, K319T, R334Q, and P335L) are located in the N-terminal domain of TWINKLE. The mutations cause a dramatic decrease in ATPase activity, which is partially overcome in the presence of single-stranded DNA. The mutated proteins have defects in DNA helicase activity and cannot support normal levels of DNA replication. To explain the phenotypes, we use a molecular model of TWINKLE based on sequence similarities with the phage T7 gene 4 protein. The four adPEO-causing mutations are located in a region required to bind single-stranded DNA. These mutations may therefore impair an essential element of the catalytic cycle in hexameric helicases, i.e. the interplay between single-stranded DNA binding and ATP hydrolysis

2,4-Diaminoquinazolines as Dual Toll-like Receptor (TLR) 7/8 Modulators for the Treatment of Hepatitis B Virus

A novel series of 2,4-diaminoquinazolines was identified as potent dual Toll-like receptor (TLR) 7 and 8 agonists with reduced off-target activity. The stereochemistry of the amino alcohol was found to influence the TLR7/8 selectivity with the (<i>R</i>) isomer resulting in selective TLR8 agonism. Lead optimization toward a dual agonist afforded (<i>S</i>)-3-((2-amino-8-fluoroquinazolin-4-yl)­amino)­hexanol <b>31</b> as a potent analog, being structurally different from previously described dual agonists (McGowan J. Med. Chem. 2016, 59, 7936). Pharmacokinetic and pharmacodynamic (PK/PD) studies revealed the desired high first pass profile aimed at limiting systemic cytokine activation. In vivo pharmacodynamic studies with lead compound <b>31</b> demonstrated production of cytokines consistent with TLR7/8 activation in mice and cynomolgus monkeys and ex vivo inhibition of hepatitis B virus (HBV)

2,4-Diaminoquinazolines as Dual Toll-like Receptor (TLR) 7/8 Modulators for the Treatment of Hepatitis B Virus

A novel series of 2,4-diaminoquinazolines was identified as potent dual Toll-like receptor (TLR) 7 and 8 agonists with reduced off-target activity. The stereochemistry of the amino alcohol was found to influence the TLR7/8 selectivity with the (<i>R</i>) isomer resulting in selective TLR8 agonism. Lead optimization toward a dual agonist afforded (<i>S</i>)-3-((2-amino-8-fluoroquinazolin-4-yl)­amino)­hexanol <b>31</b> as a potent analog, being structurally different from previously described dual agonists (McGowan J. Med. Chem. 2016, 59, 7936). Pharmacokinetic and pharmacodynamic (PK/PD) studies revealed the desired high first pass profile aimed at limiting systemic cytokine activation. In vivo pharmacodynamic studies with lead compound <b>31</b> demonstrated production of cytokines consistent with TLR7/8 activation in mice and cynomolgus monkeys and ex vivo inhibition of hepatitis B virus (HBV)