Andrea Beccari

Receptor binding mode and pharmacological characterization of a potent and selective dual CXCR1/CXCR2 non-competitive allosteric inhibitor

BACKGROUND AND PURPOSE DF 2156A is a new dual inhibitor of IL‐8 receptors CXCR1 and CXCR2 with an optimal pharmacokinetic profile. We characterized its binding mode, molecular mechanism of action and selectivity, and evaluated its therapeutic potential.

Allosteric inhibitors of chemoattractant receptors: opportunities and pitfalls

Given the central role of chemokines in infection, inflammation and immunity, chemokine receptors are a prime target for pharmacological intervention, and more so after the recent approval of chemokine receptor inhibitors for HIV. Allosteric inhibitors offer a largely unexploited opportunity to interfere with and modulate chemokine receptor activation and signaling. In addition to characterizing binding mode as a first step to understanding the specific mechanism underlying drug action, allosteric inhibitors pose new questions concerning different phases in drug discovery and pharmacological characterization, including the identification of appropriate screening tests, the evaluation of inhibitory effects on different signaling pathways and the implications of agonist- and signaling pathway-dependent inhibition for overall in vivo efficacy.

Targeting the minor pocket of C5aR for the rational design of an oral allosteric inhibitor for inflammatory and neuropathic pain relief

Significance Persistent pain in inflammatory and neuropathic conditions is often refractory to conventional analgesic therapy, with most patients suffering with unrelieved pain and serious treatment-related side effects. There is still a tremendous need to identify novel therapeutics for pain control with innovative biological mechanisms and minimal side effects. In this paper we challenge the hypothesis that a conserved structural motif across the G protein-coupled receptor family plays a regulatory role in the negative modulation of receptor activation and use a multidisciplinary approach to the rational drug design and characterization of a novel potent allosteric inhibitor of the C5a anaphylatoxin receptor (C5aR), thus providing a new promising avenue for the improvement of pharmacotherapy of chronic pain. Chronic pain resulting from inflammatory and neuropathic disorders causes considerable economic and social burden. Pharmacological therapies currently available for certain types of pain are only partially effective and may cause severe adverse side effects. The C5a anaphylatoxin acting on its cognate G protein-coupled receptor (GPCR), C5aR, is a potent pronociceptive mediator in several models of inflammatory and neuropathic pain. Although there has long been interest in the identification of C5aR inhibitors, their development has been complicated, as for many peptidomimetic drugs, mostly by poor drug-like properties. Herein, we report the de novo design of a potent and selective C5aR noncompetitive allosteric inhibitor, DF2593A, guided by the hypothesis that an allosteric site, the “minor pocket,” previously characterized in CXC chemokine receptors-1 and -2, is functionally conserved in the GPCR class. In vitro, DF2593A potently inhibited C5a-induced migration of human and rodent neutrophils. In vivo, oral administration of DF2593A effectively reduced mechanical hyperalgesia in several models of acute and chronic inflammatory and neuropathic pain, without any apparent side effects. Mechanical hyperalgesia after spared nerve injury was also reduced in C5aR−/− mice compared with WT mice. Furthermore, treatment of C5aR−/− mice with DF2593A did not produce any further antinociceptive effect compared with C5aR−/− mice treated with vehicle. The successful medicinal chemistry strategy confirms that a conserved minor pocket is amenable for the rational design of selective inhibitors and the pharmacological results support that the allosteric blockade of the C5aR represents a highly promising therapeutic approach to control chronic inflammatory and neuropathic pain.

LiGen: a high performance workflow for chemistry driven de novo design.

Tools for molecular de novo design are actively sought incorporating sets of chemical rules for fast and efficient identification of structurally new chemotypes endowed with a desired set of biological properties. In this paper, we present LiGen, a suite of programs which can be used sequentially or as stand-alone tools for specific purposes. In its standard application, LiGen modules are used to define input constraints, either structure-based, through active site identification, or ligand-based, through pharmacophore definition, to docking and to de novo generation. Alternatively, individual modules can be combined in a user-defined manner to generate project-centric workflows. Specific features of LiGen are the use of a pharmacophore-based docking procedure which allows flexible docking without conformer enumeration and accurate and flexible reactant mapping coupled with reactant tagging through substructure searching. The full description of LiGen functionalities is presented.

Molecular mechanism and functional role of brefeldin A-mediated ADP-ribosylation of CtBP1/BARS

ADP-ribosylation is a posttranslational modification that modulates the functions of many target proteins. We previously showed that the fungal toxin brefeldin A (BFA) induces the ADP-ribosylation of C-terminal–binding protein-1 short-form/BFA–ADP-ribosylation substrate (CtBP1-S/BARS), a bifunctional protein with roles in the nucleus as a transcription factor and in the cytosol as a regulator of membrane fission during intracellular trafficking and mitotic partitioning of the Golgi complex. Here, we report that ADP-ribosylation of CtBP1-S/BARS by BFA occurs via a nonconventional mechanism that comprises two steps: (i) synthesis of a BFA–ADP-ribose conjugate by the ADP-ribosyl cyclase CD38 and (ii) covalent binding of the BFA–ADP-ribose conjugate into the CtBP1-S/BARS NAD+-binding pocket. This results in the locking of CtBP1-S/BARS in a dimeric conformation, which prevents its binding to interactors known to be involved in membrane fission and, hence, in the inhibition of the fission machinery involved in mitotic Golgi partitioning. As this inhibition may lead to arrest of the cell cycle in G2, these findings provide a strategy for the design of pharmacological blockers of cell cycle in tumor cells that express high levels of CD38.

Use of experimental design to optimize docking performance: the case of LiGenDock, the docking module of LiGen, a new de novo design program.

On route toward a novel de novo design program, called LiGen, we developed a docking program, LiGenDock, based on pharmacophore models of binding sites, including a non-enumerative docking algorithm. In this paper, we present the functionalities of LiGenDock and its accompanying module LiGenPocket, aimed at the binding site analysis and structure-based pharmacophore definition. We also report the optimization procedure we have carried out to improve the cognate docking and virtual screening performance of LiGenDock. In particular, we applied the design of experiments (DoE) methodology to screen the set of user-adjustable parameters to identify those having the largest influence on the accuracy of the results (which ensure the best performance in pose prediction and in virtual screening approaches) and then to choose their optimal values. The results are also compared with those obtained by two popular docking programs, namely, Glide and AutoDock for pose prediction, and Glide and DOCK6 for Virtual Screening.

Accelerating a Geometric Approach to Molecular Docking with OpenACC

In a drug discovery process, the Molecular Docking task aims at estimating the three-dimensional pose of a molecule when it interacts with the target protein. This task is usually used to perform a screening on a large library of molecules to find the most promising candidates. The output of this task is used to estimate the actual strength of atomic interactions. In this document we focus on an application that performs molecular docking using geometrical features of the molecule and of the protein, to quickly screen the target chemical library. Due to the size of the chemical library and to the complexity of the task, the application is a typical batch job that runs in an HPC platform, optimized for CPU processing. Given the amount of parallelism of this application, we evaluate the possibility to run such application on a GPU node, leveraging the OpenACC directive language. Preliminary results show that we are able to achieve a significant speedup on the kernel that was the bottleneck on the CPU (up to 16x), while we achieve a speedup of 5x on the overall execution.

Receptor binding mode and pharmacological characterization of a potent and selective dual CXCR1/CXCR2 non-competitive allosteric inhibitor: Therapeutic potential of a CXCR1/CXCR2 inhibitor

BACKGROUND AND PURPOSE DF 2156A is a new dual inhibitor of IL‐8 receptors CXCR1 and CXCR2 with an optimal pharmacokinetic profile. We characterized its binding mode, molecular mechanism of action and selectivity, and evaluated its therapeutic potential.