Discovery of Covalent Inhibitors of Glyceraldehyde-3-phosphate Dehydrogenase, A Target for the Treatment of Malaria

We developed a new class of covalent inhibitors of Plasmodium falciparum glyceraldehyde-3-phosphate dehydrogenase, a validated target for the treatment of malaria, by screening a small library of 3-bromo-isoxazoline derivatives that inactivate the enzyme through a covalent, selective bond to the catalytic cysteine, as demonstrated by mass spectrometry. Substituents on the isoxazolinic ring modulated the potency up to 20-fold, predominantly due to an electrostatic effect, as assessed by computational analysis

p53 Functional Inhibitors Behaving Like Pifithrin‑β Counteract the Alzheimer Peptide Non-β-amyloid Component Effects in Human SH-SY5Y Cells

Alzheimer’s disease (AD) develops from a complex setting of genetic and biochemical alterations, including an increased level of p53 in the brain. Here, the robust and specific activation of p53 by the fibrillar non-β-amyloid component (NAC) of AD was demonstrated in human neuroblastoma SH-SY5Y cells. For the first time, the increase in the level of p53 target gene transcription, the cell cycle arrest, and the induction of apoptosis elicited by NAC were evidenced. These effects were counterbalanced by pifithrin-β, a small molecule interfering with the p53 functions. Using the structure of a pifithrin-β analogue as a reference, a pharmacophore-based virtual screening of the ZINC database was performed. Among the resulting hits, 20 druglike heterocyclic compounds were selected and evaluated for their neuroprotective activity against fibrillar NAC in the human SH-SY5Y cellular model. Three compounds exhibited neuroprotective effects. In particular, 2-(4-methoxyphenyl)-7-methyl-7<i>H</i>-pyrazolo­[4,3-<i>e</i>]­[1,2,4]­triazolo­[1,5-<i>c</i>]­pyrimidine resulted in a promising lead compound for further development of anti-AD agents in terms of neuroprotection, reducing the rate of NAC-induced cell death with an activity higher than that of pifithrin-β, as a result of a more effective functional inhibition of p53 target gene transcription

Protein Flexibility in Virtual Screening: The BACE‑1 Case Study

Simulating protein flexibility is a major issue in the docking-based drug-design process for which a single methodological solution does not exist. In our search of new anti-Alzheimer ligands, we were faced with the challenge of including receptor plasticity in a virtual screening campaign aimed at finding new β-secretase inhibitors. To this aim, we incorporated protein flexibility in our simulations by using an ensemble of static X-ray enzyme structures to screen the National Cancer Institute database. A unified description of the protein motion was also generated by computing and combining a set of grid maps using an energy weighting scheme. Such a description was used in an energy-weighted virtual screening experiment on the same molecular database. Assessment of the enrichment factors from these two virtual screening approaches demonstrated comparable predictive powers, with the energy-weighted method being faster than the ensemble method. The <i>in vitro</i> evaluation demonstrated that out of the 32 tested ligands, 17 featured the predicted enzyme inhibiting property. Such an impressive success rate (53.1%) demonstrates the enhanced power of the two methodologies and suggests that energy-weighted virtual screening is a more than valid alternative to ensemble virtual screening given its reduced computational demands and comparable performance

Selective Arylsulfonamide Inhibitors of ADAM-17: Hit Optimization and Activity in Ovarian Cancer Cell Models

Activated leukocyte cell adhesion molecule (ALCAM) is expressed at the surface of epithelial ovarian cancer (EOC) cells and is released in a soluble form (sALCAM) by ADAM-17-mediated shedding. This process is relevant to EOC cell motility and invasiveness, which is reduced by inhibitors of ADAM-17. In addition, ADAM-17 plays a key role in EGFR signaling and thus may represent a useful target in anticancer therapy. Herein we report our hit optimization effort to identify potent and selective ADAM-17 inhibitors, starting with previously identified inhibitor <b>1</b>. A new series of secondary sulfonamido-based hydroxamates was designed and synthesized. The biological activity of the newly synthesized compounds was tested in vitro on isolated enzymes and human EOC cell lines. The optimization process led to compound <b>21</b>, which showed an IC₅₀ of 1.9 nM on ADAM-17 with greatly increased selectivity. This compound maintained good inhibitory properties on sALCAM shedding in several in vitro assays

Structure-Based Lead Optimization and Biological Evaluation of BAX Direct Activators as Novel Potential Anticancer Agents

The first direct activator of BAX, a pro-apoptotic member of the BCL-2 family, has been recently identified. Herein, a structure-based lead optimization turned out into a small series of analogues, where <b>8</b> is the most potent compound published so far. <b>8</b> was used as pharmacological tool to ascertain, for the first time, the anticancer potential of BAX direct activators and the obtained results would suggest that BAX direct activators are potential future anticancer drugs rather than venoms

Structure-Based Optimization of Tyrosine Kinase Inhibitor <b>CLM3</b>. Design, Synthesis, Functional Evaluation, and Molecular Modeling Studies.

Recent advances in the knowledge of thyroid carcinomas development identified receptor tyrosine kinases, like VEGFR2 and RET, as viable and promising targets. Accordingly, their inhibition is emerging as the major therapeutic strategy to treat these pathologies. In this study we describe the synthesis and the functional evaluation of three different series of 4-substituted pyrazolo­[3,4-<i>d</i>]­pyrimidine derivatives, <b>8a</b>–<b>g</b>, <b>9a</b>–<b>g</b>, and <b>10a</b>–<b>g</b>, designed exploiting a structure-based optimization of the previously developed inhibitor <b>CLM3</b>. Compared to the lead, the novel compounds markedly improved both their inhibitory profile against the target proteins, VEGFR2 and RET, and their antiproliferative efficacy against the medullary thyroid cancer cell line TT. Significantly, compounds <b>8b</b>, <b>9c</b>, and <b>10c</b> proved to block the kinase activity of the mutant RET^V804L, which still lacks effective inhibitors

Identification of Glycogen Synthase Kinase-3 Inhibitors with a Selective Sting for Glycogen Synthase Kinase-3α

The glycogen synthase kinase-3 (GSK-3) has been linked to the pathogenesis of colorectal cancer, diabetes, cardiovascular disease, acute myeloid leukemia (AML), and Alzheimer’s disease (AD). The debate on the respective contributions of GSK-3α and GSK-3β to AD pathology and AML is ongoing. Thus, the identification of potent GSK-3α-selective inhibitors, endowed with favorable pharmacokinetic properties, may elucidate the effect of GSK-3α inhibition in AD and AML models. The analysis of all available crystallized GSK-3 structures provided a simplified scheme of the relevant hot spots responsible for ligand binding and potency. This resulted in the identification of novel scorpion shaped GSK-3 inhibitors. It is noteworthy, compounds <b>14d</b> and <b>15b</b> showed the highest GSK-3α selectivity reported so far. In addition, compound <b>14d</b> did not display significant inhibition of 48 out of 50 kinases in the test panel. The GSK-3 inhibitors were further profiled for efficacy and toxicity in the wild-type (wt) zebrafish embryo assay

Simultaneous Targeting of RGD-Integrins and Dual Murine Double Minute Proteins in Glioblastoma Multiforme

In the fight against Glioblastoma Multiforme, recent literature data have highlighted that integrin α5β1 and p53 are part of convergent pathways in the control of glioma apoptosis. This observation prompted us to seek a molecule able to simultaneously modulate both target families. Analyzing the results of a previous virtual screening against murine double minute 2 protein (MDM2), we envisaged that Arg-Gly-Asp (RGD)-mimetic molecules could be inhibitors of MDM2/4. Herein, we present the discovery of compound <b>7</b>, which inhibits both MDM2/4 and α5β1/αvβ3 integrins. A lead optimization campaign was carried out on <b>7</b> with the aim to preserve the activities on integrins while improving those on MDM proteins. Compound <b>9</b> turned out to be a potent MDM2/4 and α5β1/αvβ3 blocker. In p53-wild type glioma cells, <b>9</b> arrested cell cycle and proliferation and strongly reduced cell invasiveness, emerging as the first molecule of a novel class of integrin/MDM inhibitors, which might be especially useful in subpopulations of patients with glioblastoma expressing a functional p53 concomitantly with a high level of α5β1 integrin

Simultaneous Targeting of RGD-Integrins and Dual Murine Double Minute Proteins in Glioblastoma Multiforme

In the fight against Glioblastoma Multiforme, recent literature data have highlighted that integrin α5β1 and p53 are part of convergent pathways in the control of glioma apoptosis. This observation prompted us to seek a molecule able to simultaneously modulate both target families. Analyzing the results of a previous virtual screening against murine double minute 2 protein (MDM2), we envisaged that Arg-Gly-Asp (RGD)-mimetic molecules could be inhibitors of MDM2/4. Herein, we present the discovery of compound <b>7</b>, which inhibits both MDM2/4 and α5β1/αvβ3 integrins. A lead optimization campaign was carried out on <b>7</b> with the aim to preserve the activities on integrins while improving those on MDM proteins. Compound <b>9</b> turned out to be a potent MDM2/4 and α5β1/αvβ3 blocker. In p53-wild type glioma cells, <b>9</b> arrested cell cycle and proliferation and strongly reduced cell invasiveness, emerging as the first molecule of a novel class of integrin/MDM inhibitors, which might be especially useful in subpopulations of patients with glioblastoma expressing a functional p53 concomitantly with a high level of α5β1 integrin