Annalina Lapucci

Novel transthyretin amyloid fibril formation inhibitors: synthesis, biological evaluation, and X-ray structural analysis.

Transthyretin (TTR) is one of thirty non-homologous proteins whose misfolding, dissociation, aggregation, and deposition is linked to human amyloid diseases. Previous studies have identified that TTR amyloidogenesis can be inhibited through stabilization of the native tetramer state by small molecule binding to the thyroid hormone sites of TTR. We have evaluated a new series of β-aminoxypropionic acids (compounds 5–21), with a single aromatic moiety (aryl or fluorenyl) linked through a flexible oxime tether to a carboxylic acid. These compounds are structurally distinct from the native ligand thyroxine and typical halogenated biaryl NSAID-like inhibitors to avoid off-target hormonal or anti-inflammatory activity. Based on an in vitro fibril formation assay, five of these compounds showed significant inhibition of TTR amyloidogenesis, with two fluorenyl compounds displaying inhibitor efficacy comparable to the well-known TTR inhibitor diflunisal. Fluorenyl 15 is the most potent compound in this series and importantly does not show off-target anti-inflammatory activity. Crystal structures of the TTR∶inhibitor complexes, in agreement with molecular docking studies, revealed that the aromatic moiety, linked to the sp2-hybridized oxime carbon, specifically directed the ligand in either a forward or reverse binding mode. Compared to the aryl family members, the bulkier fluorenyl analogs achieved more extensive interactions with the binding pockets of TTR and demonstrated better inhibitory activity in the fibril formation assay. Preliminary optimization efforts are described that focused on replacement of the C-terminal acid in both the aryl and fluorenyl series (compounds 22–32). The compounds presented here constitute a new class of TTR inhibitors that may hold promise in treating amyloid diseases associated with TTR misfolding.

Synthesis and pharmacological evaluation of multifunctional tacrine derivatives against several disease pathways of AD.

A novel series of tacrine derivatives were designed and synthesized by combining caffeic acid (CA), ferulic acid (FA) and lipoic acid (LA) with tacrine. The antioxidant study revealed that all the hybrids have much more antioxidant capacities compared to CA. Among these compounds, 1b possessed a good ability to inhibit the β-amyloid protein (Aβ) self-aggregation, sub-micromole acetylcholinesterase (AChE)/butyrylcholinesterase (BuChE) inhibitory, modest BACE1 inhibitory. Moreover, compound 1b also was a DPPH radical scavenger and copper chelatory as well as had potent neuroprotective effects against glutamate-induced cell death with low toxicity in HT22 cells. Our findings suggest that the compound 1b might be a promising lead multi-targeted ligand and worthy of further developing for the therapy of Alzheimers disease.

Design and synthesis of 2-oxindole based multi-targeted inhibitors of PDK1/Akt signaling pathway for the treatment of glioblastoma multiforme

Aggressive behavior and diffuse infiltrative growth are the main features of Glioblastoma multiforme (GBM), together with the high degree of resistance and recurrence. Evidence indicate that GBM-derived stem cells (GSCs), endowed with unlimited proliferative potential, play a critical role in tumor development and maintenance. Among the many signaling pathways involved in maintaining GSC stemness, tumorigenic potential, and anti-apoptotic properties, the PDK1/Akt pathway is a challenging target to develop new potential agents able to affect GBM resistance to chemotherapy. In an effort to find new PDK1/Akt inhibitors, we rationally designed and synthesized a small family of 2-oxindole derivatives. Among them, compound 3 inhibited PDK1 kinase and downstream effectors such as CHK1, GS3Kα and GS3Kβ, which contribute to GCS survival. Compound 3 appeared to be a good tool for studying the role of the PDK1/Akt pathway in GCS self-renewal and tumorigenicity, and might represent the starting point for the development of more potent and focused multi-target therapies for GBM.

Synthesis of novel 3,5-disubstituted-2-oxindole derivatives as antitumor agents against human nonsmall cell lung cancer

This study was aimed at investigating the antitumor activity of novel 2-oxindole derivatives against a well-characterized human nonsmall cell lung cancer (NSCLC) cell line. Test compounds produced an antiproliferative activity in the low micromolar/submicromolar range of concentrations and significantly induced typical apoptotic morphology with cell shrinkage, nuclear condensation and fragmentation, and rupture of cells into debris in a relatively low percentage of A549 cells. Cell cycle arrest occurred at the G1/S phase (1a and 2), and Akt phosphorylation was significantly inhibited at Thr308 and Ser473. The most active compound (1a) has an IC50 6-fold lower than the Akt inhibitor, perifosine. These data suggest that the new compounds may be cytostatic and may have maximum clinical effects in NSCLC patients who do not respond to EGFR inhibitors. These findings prompt us to further explore the oxindole structure as leading scaffold to design new molecules with potent antitumor activity against NSCLC.

Therapeutic potential of sulindac hydroxamic acid against human pancreatic and colonic cancer cells

The non-steroidal anti-inflammatory drug (NSAID) sulindac exhibits cyclooxygenase (COX)-dependent and COX-independent chemopreventive properties in human cancer. The present study was aimed at investigating whether the hydroxamic acid substitution for the carboxylic acid group could enhance the in vitro antitumor and antiangiogenic activities of sulindac. Characterization tools used on this study included analyses of cell viability, caspase 3/7 induction, DNA fragmentation, and gene expression. Our findings demonstrate that the newly synthesized hydroxamic acid derivative of sulindac and its sulfone and sulfide metabolites were characterized by a good anticancer activity on human pancreatic and colon cancer cells, both in terms of potency (IC(50) mean values from 6 ± 1.1 μM to 64 ± 1.1 μM) and efficacy (E(max) of ∼100%). Hydroxamic acid derivatives trigger a higher degree of apoptosis than carboxylic acid counterparts, increase bax/bcl-2 expression ratio and induce caspase 3/7 activation. Most notably, these compounds significantly inhibit proangiogenic growth factor-stimulated proliferation of vascular endothelial cell (HUVEC) at sub-micromolar concentrations. Our data also provide evidence that the COX-active metabolite of sulindac hydroxamic acid were the most active of the series and selective inhibition of COX-1 but not COX-2 can mimic its effects, suggesting that COX inhibition could only play a partial role in the mechanism of compound action. In conclusion, these data demonstrate that substitution of the carboxylic acid group with the hydroxamic acid moiety enhances in vitro antiproliferative, proapoptotic and antiangiogenic properties of sulindac, therefore increasing the therapeutic potential of this drug.

Synthesis and Biological Evaluation of 2′‐Oxo‐2,3‐dihydro‐3′H‐ spiro[chromene‐4,5′‐[1,3]oxazolidin]‐3′yl]acetic Acid Derivatives as Aldose Reductase Inhibitors

Aldose reductase (ARL2) is the first enzyme in the polyol pathway which catalyzes the NADPH‐dependent reduction of glucose to sorbitol. Its involvement on diabetic complications makes this enzyme a challenge therapeutic target widely investigated to limit and/or prevent them. On this basis, a limited series of 4‐spiro‐oxazolidinone‐benzopyran derivatives (1–7) were synthesized to evaluate them as potential ARL2 inhibitors. The activity was determined spectrophotometrically by monitoring the oxidation of NADPH catalyzed by ALR2. Within the series of compounds, the 4‐methoxy derivative 1b showed to be the most active compound, exhibiting inhibitory levels in the submicromolar range. In addition, the activity against the aldehyde reductase isoform (ARL1) was also evaluated. Unlike sorbinil (reference drug) that lack of selectivity towards the two enzyme all the tested compounds resulted to be devoid of ARL1 inhibitory activity (IC50 > 10 µM), thus proving to be selective.

SYNTHESIS, ANTIINFLAMMATORY ACTIVITY AND MOLECULAR-ORBITAL STUDIES OF A SERIES OF BENZYLIDENEAMINOXYPROPIONIC ACIDS SUBSTITUTED ON THE PHENYL RING

Abstract An examination of the antiinflammatory properties of certain β-aminoxypropionic acids A (AOPAs) previously synthesized as analogs of antiinflammatory drugs with an arylacetic structure B (ArAAs), indicated that the most active acid is the ( E )-3-(benzylideneaminoxy)-propionic acid ( 1 ), which was found to possess an activity comparable to that of Diclofenac ( 8 ). In an attempt to verify whether appropriate substitutions on the aromatic ring of 1 can modulate the antiinflammatory properties of this class of drugs, a series of benzylideneaminoxypropionic acids (BAOPAs) were synthesized, in which the phenyl group is substituted, in its 3 possible positions, by groups possessing different electronic, steric, and lipophilic characteristics. The antiinflammatory activity of the new compounds was determined by carrageenan-induced rat paw edema, using Diclofenac ( 8 ) as the reference drug. The pharmacological results revealed that among the BAOPAs examined, the most active are the m -chloro ( 7b ) and p -ethoxy ( 7p ) substituted compounds, which exhibit an antiinflammatory activity comparable to that of 1 . Quantum mechanical calculations were also carried out in order to gain insight into the possible correlations between the pharmacological activity observed and the electronic and conformational effects induced by the presence of the various substituents.

3-(Methyleneaminoxy)methylpiperidine derivatives as uptake inhibitors of biogenic amines in the brain synaptosomal fraction

Abstract A series of 3-(methyleneaminoxy)methylpiperidines (5a–h) and their corresponding N-methyl derivatives (6a–h) with a variety of substituents on the imino carbon were synthesized and tested for their potential antidepressant properties; their capacity to inhibit the re-uptake of biogenic amines (NA, 5-HT and DA) in rabbit brain synaptosomal fractions was also evaluated. The biological results obtained for the piperidine derivatives 5a–h and 6a–h and viloxazine 1, the reference drug, on the 3 re-uptake systems revealed that compounds 5 and 6 are generally able to inhibit biogenic amine uptake. The IC50 values for 5 and 6 were often lower than that of viloxazine 1, in particular for the serotonin- and/or dopamine-uptake systems. A higher activity was found for compounds substituted with at least one phenyl ring on the imino carbon with respect to completely aliphatic systems, and for N-unsubstituted compounds with respect to N-methyl-substituted compounds.

Synthesis, molecular docking and binding studies of selective serotonin transporter inhibitors.

With the aim of obtaining compounds possessing high SERT selectivity, in the present work we synthesized and studied the inhibition of serotonin (SERT), dopamine (DAT) and norepinephrine (NET) transporters by docking studies and experimental binding measurements of a series of 4-(aryl)piperidin-3-one O-4-benzyl oxime hydrochlorides (1-10) of both E and Z configuration. E configuration compounds showed high SERT binding affinities (K(i) = 10-98 nM) and high SERT selectivities over both NET and DAT. The molecular docking studies allowed a rationalization of the molecular basis of drug-SERT interactions both of the synthesized compounds and paroxetine and fluoxetine used as reference antidepressant drugs.

Synthesis and in-vitro antitumour activity of new naphthyridine derivatives on human pancreatic cancer cells.

Objectives The aim of the study was to evaluate the antitumour effect in vitro of newly synthesized 7‐substituted 2,3‐dihydro‐1,8‐naphthyridines.