Genetic Analysis in UK Biobank Links Insulin Resistance and Transendothelial Migration Pathways to Coronary Artery Disease

UK Biobank is among the world’s largest repositories for phenotypic and genotypic information in individuals of European ancestry1. We performed a genome-wide association study in UK Biobank testing ~9 million DNA sequence variants for association with coronary artery disease (4,831 cases; 115,455 controls) and carried out meta-analysis with previously published results. We identified fifteen novel loci, bringing the total number of coronary artery disease-associated loci to 95. Phenome-wide association scanning revealed that CCDC92 likely affects coronary artery disease through insulin resistance pathways whereas experimental analysis suggests that ARHGEF26 impacts the transendothelial migration of leukocytes

Characterization of the Prion Protein Binding Properties of Antisense Oligonucleotides

Antisense oligonucleotides (ASOs) designed to lower prion protein (PrP) expression in the brain through RNase H1-mediated degradation of PrP RNA are in development as prion disease therapeutics. ASOs were previously reported to sequence-independently interact with PrP and inhibit prion accumulation in cell culture, yet in vivo studies using a new generation of ASOs found that only PrP-lowering sequences were effective at extending survival. Cerebrospinal fluid (CSF) PrP has been proposed as a pharmacodynamic biomarker for trials of such ASOs, but is only interpretable if PrP lowering is indeed the relevant mechanism of action in vivo and if measurement of PrP is unconfounded by any PrP–ASO interaction. Here, we examine the PrP-binding and antiprion properties of ASOs in vitro and in cell culture. Binding parameters determined by isothermal titration calorimetry were similar across all ASOs tested, indicating that ASOs of various chemistries bind full-length recombinant PrP with low- to mid-nanomolar affinity in a sequence-independent manner. Nuclear magnetic resonance, dynamic light scattering, and visual inspection of ASO–PrP mixtures suggested, however, that this interaction is characterized by the formation of large aggregates, a conclusion further supported by the salt dependence of the affinity measured by isothermal titration calorimetry. Sequence-independent inhibition of prion accumulation in cell culture was observed. The inefficacy of non-PrP-lowering ASOs against prion disease in vivo may be because their apparent activity in vitro is an artifact of aggregation, or because the concentration of ASOs in relevant compartments within the central nervous system (CNS) quickly drops below the effective concentration for sequence-independent antiprion activity after bolus dosing into CSF. Measurements of PrP concentration in human CSF were not impacted by the addition of ASO. These findings support the further development of PrP-lowering ASOs and of CSF PrP as a pharmacodynamic biomarker

Single Diastereomer of a Macrolactam Core Binds Specifically to Myeloid Cell Leukemia 1 (MCL1)

A direct binding screen of 100 000 sp³-rich molecules identified a single diastereomer of a macrolactam core that binds specifically to myeloid cell leukemia 1 (MCL1). A comprehensive toolbox of biophysical methods was applied to validate the original hit and subsequent analogues and also established a binding mode competitive with NOXA BH3 peptide. X-ray crystallography of ligand bound to MCL1 reveals a remarkable ligand/protein shape complementarity that diverges from previously disclosed MCL1 inhibitor costructures

A Maltose-Binding Protein Fusion Construct Yields a Robust Crystallography Platform for MCL1

<div><p>Crystallization of a maltose-binding protein MCL1 fusion has yielded a robust crystallography platform that generated the first apo MCL1 crystal structure, as well as five ligand-bound structures. The ability to obtain fragment-bound structures advances structure-based drug design efforts that, despite considerable effort, had previously been intractable by crystallography. In the ligand-independent crystal form we identify inhibitor binding modes not observed in earlier crystallographic systems. This MBP-MCL1 construct dramatically improves the structural understanding of well-validated MCL1 ligands, and will likely catalyze the structure-based optimization of high affinity MCL1 inhibitors.</p></div

The structure of Apo MBP-MCL1 determined at 1.90 Å.

<p>(A) The MBP domain (red) is connected by a short GS linker (orange) to MCL1 173–321 (blue). A portion of alpha helix four is not ordered in the structure (red dashed-line). Maltose ligand is shown in yellow. (B) The MCL1 domain is structurally very similar to the NMR structure of Apo-MCL1 (gray).</p

The structure of MBP-MCL1 bound to fragment 5 determined at 1.9 Å.

<p>Fragment <b>5</b> (yellow) binds similarly in comparison to the elaborated ligand from PDB ID 4OQ6 (gray).</p

The conformational flexibility of the binding pocket of MCL1.

<p>Surface representations are shown as side views and ligands are shown as yellow sticks. (A and B) Fragment 4 maps onto L78 of NoxaB from PDB ID 2NLA, with only minor structural perturbation of the BH3-binding groove of MCL1. In contrast, binding of fragment 6 creates a significant pocket (C) which is further expanded upon binding of ligand 1 (D).</p

Binding affinity (K_D) of ligands to MCL1 and MBP-MCL1.

<p>All experiments are n ≥ 3, and averaged values for K_D are reported.</p><p>Binding affinity (K_D) of ligands to MCL1 and MBP-MCL1.</p

Comparison of PDB 4HW3 and MBP-MCL1 with fragment 4.

<p>The structure of MBP-MCL1 with fragment <b>4</b> (yellow) determined to 2.4 Å (blue) overlaid with the structure of MCL1 171–323 determined at 2.4 Å (PDB ID 4HW3, gray). The carboxylic acid of 4HW3 adopts multiple conformations depending on the chain; only chain A is shown for clarity.</p