Next generation of antiretroviral agents targeting the RNA binding site of the HIV-1 cellular cofactor DDX3: an innovative therapeutic approach

Introduction: Efficacy of currently approved anti-HIV drugs is hampered by mutations of the viral enzymes, leading invariably to drug resistance and chemotherapy failure. Recent data suggest that cellular co-factors also represent useful targets for anti-HIV therapy. We have recently provided evidence for the possibility to block HIV-1 replication by targeting its cellular cofactor DDX3. Material and methods: Molecular modeling and in silico technologies were applied to rationally design small molecules specifically targeting the RNA binding site of human DDX3. Biochemical studies of mutated DDX3 enzymes were also used to identify additional potential drug binding sites. Results Optimization of compounds identified by application of a high-throughput docking approach afforded a promising lead compound which proved to inhibit both the helicase and ATPase activity of DDX3 and to reduce the viral load of peripheral blood mononuclear cells (PBMC) infected with HIV-1. A novel interaction site has been also identified in DDX3, which, when blocked, can reduce viral replication, representing an additional target for small molecules inhibitors. Conclusions: We have identified the first inhibitors of HIV-1 replication targeting the RNA binding site of the cellular cofactor human DDX3. These compounds may offer superior selectivity over the ATP-competitive inhibitors previously developed. In addition, a novel RNA interacting motif specific to DDX3 has been identified, opening new venues for HIV-1 drug development

Radial access for percutaneous coronary procedure: relationship between operator expertise and complications

Objective The aim of this study was to investigate (1) whether the learning curve of new catheterization laboratory operators increases the incidence of complications of transradial access during percutaneous coronary interventions and (2) whether manual compression with a two-step approach is safe and efficient for radial access hemostasis. Methods We performed a prospective study with all consecutive patients who underwent a coronary diagnostic or intervention procedure with radial access. The primary end point was a composite of pulseless radial artery of the wrist and hematoma evaluated after 24 hours. The secondary end point of efficacy was defined as the presence of bleeding or hematoma after 30 seconds. Results From March 2016 to June 2016, 150 consecutive patients, of whom 147 underwent coronary angiography and/or percutaneous coronary intervention through radial access, were included in the present study. The primary end point was present in 33%, but pulseless radial artery of the wrist was present only in 5.3%. We found that the incidence of primary end point was statistically different according to the number of puncture attempts, with a cutoff of two punctures with blood. The secondary end point of safety was present only in 4.7% of the cases. Conclusion Radial access is feasible and safe even if performed by training physicians. Manual compression with early evaluation after 30 seconds is a safe technique for managing the radial access after sheath removal

Protein Dynamics of the HIF-2\u3b1 PAS-B Domain upon Heterodimerization and Ligand Binding

Hypoxia-Inducible Factor (HIF) transcription factors are heterodimeric proteins involved in the regulation of oxygen homeostatis. Their upregulation has been related to several tumors with a remarkably poor clinical outcome. The recent discovery of a druggable cavity in the HIF-2\u3b1 PAS-B domain has opened an unprecedented opportunity for targeting the HIF-2\u3b1 transcription factor in view of pharmaceutical strategies. Coincidentally, a novel compound able to selectively disrupt the HIF heterodimerization with a submicromolar activity has been reported. In this work, we investigated the molecular mechanisms responsible for the inhibition by comparing the dynamical features of the HIF-2\u3b1 PAS-B monomer and the HIF-2\u3b1 PAS-B/HIF-1\u3b2 PAS-B complex, in the ligand-bound and -unbound states. Plain and biased Molecular Dynamics were used to characterize the differential conformational changes both structurally and energetically

COVID-19 Repurposed Therapeutics Targeting the Viral Protease and Spike-protein:ACE2 Interface using MD-based Pharmacophore and Consensus Virtual Screening

Molecular dynamics (MD) and enhanced sampling MD was performed for 100 ns on the biological assembly of the COVID-19 protease (6LU7), and a template of the COVID-19 S-protein:ACE2 receptor interface (99.88% coverage of 6M0J; model03, swissmodel). Apo-site pharmacophores of the resulting structural clusters were used to mine the FDA database (8700 compounds), and a multi-target library was developed from MD-based hits in high affinity sites across 100 ns. Consensus hits from high throughput docking in crystal structures 5R82, 6LU7 and 6Y2F (protease), and 6VW1 (S-protein:ACE2) were also added, and the resulting libraries were re-docked into MD sites to collect potential COVID-19 re-purposed therapeutics by estimated binding energies. </p

Structure-based design of small-molecule protein\u2013protein interaction modulators: the story so far

As the pivotal role of protein\u2013protein interactions in cell growth, transcriptional activity, intracellular trafficking, signal transduction and pathological conditions has been assessed, experimental and in silico strategies have been developed to design protein\u2013protein interaction modulators. State-of-the-art structure-based design methods, mainly pharmacophore modeling and docking, which have succeeded in the identification of enzyme inhibitors, receptor agonists and antagonists, and new tools specifically conceived to target protein\u2013protein interfaces (e.g., hot-spot and druggable pocket prediction methods) have been applied in the search for small-molecule protein\u2013protein interaction modulators. Many successful applications of structure-based design approaches that, despite the challenge of targeting protein\u2013protein interfaces with small molecules, have led to the identification of micromolar and submicromolar hits are reviewed here

The HIF-2α PAS-B/HIF-1β PAS-B complex.

<p>A. The HIF-1β PAS-B domain is shown as blue ribbons, whereas the HIF-2α PAS-B is colored in white except for the three central β-strands of the β-sheet surface (Aβ, Iβ, and Hβ strands, in yellow). The eight crystallographic water molecules are also shown as van der Waals spheres. B. Details on important aminoacids at the interface between domains. In particular, aminoacids involved in heterodimerization (Gln322, Met338, and Tyr342) and retro-mutated aminoacids (Arg247 and Glu362) are shown as sticks. The HIF-2α PAS-B Connolly surface is shown in transparent.</p