Antonio Lavecchia

Machine-learning approaches in drug discovery: methods and applications.

During the past decade, virtual screening (VS) has evolved from traditional similarity searching, which utilizes single reference compounds, into an advanced application domain for data mining and machine-learning approaches, which require large and representative training-set compounds to learn robust decision rules. The explosive growth in the amount of public domain-available chemical and biological data has generated huge effort to design, analyze, and apply novel learning methodologies. Here, I focus on machine-learning techniques within the context of ligand-based VS (LBVS). In addition, I analyze several relevant VS studies from recent publications, providing a detailed view of the current state-of-the-art in this field and highlighting not only the problematic issues, but also the successes and opportunities for further advances.

Insights into the mechanism of partial agonism: crystal structures of the peroxisome proliferator-activated receptor gamma ligand-binding domain in the complex with two enantiomeric ligands.

The peroxisome proliferator-activated receptors (PPARs) are transcriptional regulators of glucose and lipid metabolism. They are activated by natural ligands, such as fatty acids, and are also targets of synthetic antidiabetic and hypolipidemic drugs. By using cell-based reporter assays, we studied the transactivation activity of two enantiomeric ureidofibrate-like derivatives. In particular, we show that the R-enantiomer, (R)-1, is a full agonist of PPARγ, whereas the S-enantiomer, (S)-1, is a less potent partial agonist. Most importantly, we report the x-ray crystal structures of the PPARγ ligand binding domain complexed with the R- and the S-enantiomer, respectively. The analysis of the two crystal structures shows that the different degree of stabilization of the helix 12 induced by the ligand determines its behavior as full or partial agonist. Another crystal structure of the PPARγ·(S)-1 complex, only differing in the soaking time of the ligand, is also presented. The comparison of the two structures of the complexes with the partial agonist reveals significant differences and is suggestive of the possible coexistence in solution of transcriptionally active and inactive forms of helix 12 in the presence of a partial agonist. Mutation analysis confirms the importance of Leu465, Leu469, and Ile472 in the activation by (R)-1 and underscores the key role of Gln286 in the PPARγ activity.

Crystal Structure of the Peroxisome Proliferator-Activated Receptor γ (PPARγ) Ligand Binding Domain Complexed with a Novel Partial Agonist: A New Region of the Hydrophobic Pocket Could Be Exploited for Drug Design

The peroxisome proliferator-activated receptors (PPARs) are ligand-dependent transcription factors regulating glucose and lipid metabolism. The search for new PPAR ligands with reduced adverse effects with respect to the marketed antidiabetic agents thiazolidinediones (TZDs) and the dual-agonists glitazars is highly desired. We report the crystal structure and activity of the two enantiomeric forms of a clofibric acid analogue, respectively complexed with the ligand-binding domain (LBD) of PPARgamma, and provide an explanation on a molecular basis for their different potency and efficacy against PPARgamma. The more potent S-enantiomer is a dual PPARalpha/PPARgamma agonist which presents a partial agonism profile against PPARgamma. Docking of the S-enantiomer in the PPARalpha-LBD has been performed to explain its different subtype pharmacological profile. The hypothesis that partial agonists show differential stabilization of helix 3, when compared to full agonists, is also discussed. Moreover, the structure of the complex with the S-enantiomer reveals a new region of the PPARgamma-LBD never sampled before by other ligands.

STAT-3 Inhibitors: State of the Art and New Horizons for Cancer Treatment

The signal transducers and activators of transcription (STATs) include a class of cytoplasmic signaling proteins whose role in the regulation of cell growth and survival is mediated by phosphorylation of a critical tyrosine residue within the STAT protein. This occurs in response to cytokines and growth factors modulating the expression of specific target genes. In particular, phosphorylation induces STAT:STAT dimer formation between two monomers, via reciprocal phosphoTyr (pTyr)-SH2 domain interactions. To date, seven members of the STAT family, all with different roles, have been identified in mammals. After dimerization, phosphorylated STATs enter the nucleus and, working co-ordinately with other transcriptional co-activators and transcription factors, induce increased transcriptional initiation. In healthy human and animal cells, ligand-dependent activation of STATs is a transient process, lasting for several minutes to several hours. In contrast, in many cancerous cell lines and tumors, where growth factor dysregulation is frequently at the heart of cellular transformation, the STAT proteins (in particular STAT1, 3 and 5) are persistently tyrosine-phosphorylated or activated; abnormal levels of STAT3 activation have been observed in breast, ovarian, prostate, hematological and head and neck cancer cell lines. Thus, in this review, we examine the most important classes of agents designed to disrupt STAT3 signaling, with particular regard to STAT3 dimerization inhibitors, which could play a significant role in the future of cancer and adjuvant cancer therapies.

In silico methods to address polypharmacology: current status, applications and future perspectives

Polypharmacology, a new paradigm in drug discovery that focuses on multi-target drugs (MTDs), has potential application for drug repurposing, the process of finding new uses for existing approved drugs, prediction of off-target toxicities and rational design of MTDs. In this scenario, computational strategies have demonstrated great potential in predicting polypharmacology and in facilitating drug repurposing. Here, we provide a comprehensive overview of various computational approaches that enable the prediction and analysis of in vitro and in vivo drug-response phenotypes and outline their potential for drug discovery. We give an outlook on the latest advances in rational design of MTDs and discuss possible future directions of algorithm development in this field.

Synthesis, Pharmacological Characterization, and Docking Analysis of a Novel Family of Diarylisoxazoles as Highly Selective Cyclooxygenase-1 (COX-1) Inhibitors

3-(5-Chlorofuran-2-yl)-5-methyl-4-phenylisoxazole (P6), a known selective cyclooxygenase-1 (COX-1) inhibitor, was used to design a new series of 3,4-diarylisoxazoles in order to improve its biochemical COX-1 selectivity and antiplatelet efficacy. Structure-activity relationships were studied using human whole blood assays for COX-1 and COX-2 inhibition in vitro, and results showed that the simultaneous presence of 5-methyl (or -CF3), 4-phenyl, and 5-chloro(-bromo or -methyl)furan-2-yl groups on the isoxazole core was essential for their selectivity toward COX-1. 3g, 3s, 3d were potent and selective COX-1 inhibitors that affected platelet aggregation in vitro through the inhibition of COX-1-dependent thromboxane (TX) A2. Moreover, we characterized their kinetics of COX-1 inhibition. 3g, 3s, and 3d were more potent inhibitors of platelet COX-1 and aggregation than P6 (named 6) for their tighter binding to the enzyme. The pharmacological results were supported by docking simulations. The oral administration of 3d to mice translated into preferential inhibition of platelet-derived TXA2 over protective vascular-derived prostacyclin (PGI2).

Indolylarylsulfones Bearing Natural and Unnatural Amino Acids. Discovery of Potent Inhibitors of HIV-1 Non-Nucleoside Wild Type and Resistant Mutant Strains Reverse Transcriptase and Coxsackie B4 Virus

New potent indolylarylsulfone (IAS) HIV-1 NNRTIs were obtained by coupling natural and unnatural amino acids to the 2-carboxamide and introducing different electron-withdrawing substituents at position 4 and 5 of the indole nucleus. The new IASs inhibited the HIV-1 replication in human T-lymphocyte (CEM) cells at low/subnanomolar concentration and were weakly cytostatic. Against the mutant L100I, K103N, and Y181C RT HIV-1 strains in CEM cells, sulfones 3, 4, 19, 27, and 31 were comparable to EFV. The new IASs were inhibitors to Coxsackie B4 virus at low micromolar (2-9 microM) concentrations. Superimposition of PLANTS docked conformations of IASs 19 and 9 revealed different hydrophobic interactions of the 3,5-dimethylphenyl group, for which a staking interaction with Tyr181 aromatic side chain was observed. The binding mode of 19 was not affected by the L100I mutation and was consistent with the interactions reported for the WT strain.

Conformational analysis of furanoid ε-sugar amino acid containing cyclic peptides by NMR spectroscopy, molecular dynamics simulation, and X-ray crystallography: Evidence for a novel turn structure

Sugar amino acids (SAAs) are useful building blocks for the design of peptidomimetics and peptide scaffolds. The three-dimensional structures of cyclic hybrid molecules containing the furanoid epsilon-SAA III and several amino acids were elucidated to study the preferred conformation of such an epsilon-SAA and its conformational influence on the backbone of cyclic peptides. NMR-based molecular dynamics simulations and empirical calculations of the cyclic tetramer 1, consisting of two copies of the SAA residue and two amino acids, revealed that it is conformationally restrained. The two SAA residues adopt different conformations. One of them forms an unusual turn, stabilized by an intraresidue nine-member hydrogen bond. The methylene functionalities of the other SAA residue are positioned in such a way that an intraresidue H bond is not possible. The X-ray crystal structure of 1 strongly resembles the solution conformation. Molecular dynamics calculations in combination with NMR analysis were also performed for compounds 2 and 3, which contain the RGD (Arg-Gly-Asp) consensus sequence and were previously shown to inhibit alpha(IIb)beta(3)-receptor-mediated platelet aggregation. The biologically most active compound 2 adopts a preferred conformation with the single SAA residue folded into the nine-member H bond-containing turn. Compound 3, containing an additional valine residue, as compared with compound 2, is conformational flexible. Our studies demonstrate that the furanoid epsilon-SAA III is able to introduce an unusual intraresidue hydrogen bond-stabilized beta-turn-like conformation in two of the three cyclic structures.

CDC25 Phosphatase Inhibitors: An Update

The cell division cycle 25 (CDC25) family of proteins is a group of highly conserved dual-specificity phosphatases. They are key regulators of normal cell division and the cell response to DNA damage, and play a fundamental role in transitions between cell cycle phases during normal cell division, via the activation of CdK/cyclin complexes. Their abnormal expression, detected in a number of tumors, often correlated with a poor clinical prognosis, implies that their dysregulation is involved in malignant transformation. Thus, inhibition of these proteins represents an attractive therapeutic target in oncology, as evidenced from many patents and papers published on the subject in recent years. Hence, this review aims to provide an overview of recent developments in the field of CDC25 phosphatase inhibitor design since 2008.

Modeling of Cdc25B dual specifity protein phosphatase inhibitors: docking of ligands and enzymatic inhibition mechanism.

The Cdc25 dual specificity phosphatases have central roles in coordinating cellular signalling processes and cell proliferation. It has been reported that an improper amplification or activation of these enzymes is a distinctive feature of a number of human cancers, including breast cancers. Thus, the inhibition of Cdc25 phosphatases might provide a novel approach for the discovery of new and selective antitumor agents. By using the crystal structure of the catalytic domain of Cdc25B, structural models for the interaction of various Cdc25B inhibitors (1–13) with the enzyme were generated by computational docking. The parallel use of two efficient and predictive docking programs, AutoDock and GOLD, allowed mutual validation of the predicted binding poses. To evaluate their quality, the models were validated with known structure–activity relationships and site‐directed mutagenesis data. The results provide an improved basis for structure‐based ligand design and suggest a possible explanation for the inhibition mechanism of the examined Cdc25B ligands. We suggest that the recurring motif of a tight interaction between the inhibitor and the two arginine residues, 482 and 544, is of prime importance for reversible enzyme inhibition. In contrast, the irreversible inhibition mechanism of 1–4 seems to be associated with the close vicinity of the quinone ring and the Cys473 catalytic thiolate. We believe that this extensive study might provide useful hints to guide the development of new potent Cdc25B inhibitors as novel anticancer drugs.