Rosaria Gitto

Pharmacophore-Based Discovery of Small-Molecule Inhibitors of Protein–Protein Interactions between HIV-1 Integrase and Cellular Cofactor LEDGF/p75

The cellular protein lens epithelium‐derived growth factor, or transcriptional coactivator p75 (LEDGF/p75), plays a crucial role in HIV integration. The protein–protein interactions (PPIs) between HIV‐1 integrase (IN) and its cellular cofactor LEDGF/p75 may therefore serve as targets for the development of new anti‐HIV drugs. In this work, a structure‐based pharmacophore model for potential small‐molecule inhibitors of HIV‐1 IN–LEDGF/p75 interaction was developed using the LigandScout software. The 3D model obtained was used for virtual screening of our in‐house chemical database, CHIME, leading to the identification of compound CHIBA‐3002 as an interesting hit for further optimization. The rational design, synthesis and biological evaluation of four derivatives were then carried out. Our studies resulted in the discovery of a new and more potent small molecule (7, CHIBA‐3003) that is able to interfere with the HIV‐1 IN–LEDGF/p75 interaction at micromolar concentration, representing one of the first compounds to show activity against these specific PPIs. Docking simulations were subsequently performed in order to investigate the possible binding mode of our new lead compound to HIV‐1 IN. This study is a valid starting point for the identification of anti‐HIV agents with a different mechanism of action from currently available antiviral drugs.

5H-[1,2,4]Oxadiazolo[5,4-d][1,5]benzothiazepines as anticonvulsant agents in DBA/2 mice

Summary A series of 3 a ,4-dihydro-5 H -[1,2,4]oxadiazolo[5,4- d ][1,5]benzothiazepines have been synthesized by 1,3-dipolar cycloaddition reaction of benzonitriloxide to the C N double bond of 1,5-benzothiazepine derivatives, and the stereochemical features of compounds obtained have been determined by NMR spectroscopy. The results of evaluation of their activity in preventing seizures induced by audiogenic stimulation in DBA/2 mice are also reported and discussed. The 5-(4-bromophenyl)-1,3-diphenyl derivative 3b , the most active compound of the series, is over 20 times more active than the parent benzothiazepine 1b and shows an activity comparable to clobazam and better than desmethylclobazam.

GYKI 52466 and related 2,3-benzodiazepines as anticonvulsant agents in DBA/2 mice.

The behavioural and anticonvulsant effects of several 1-aryl-3,5-dihydro-4H-2,3-benzodiazepin-4-ones (2,3-BZs) and of 11b-aryl-7,11-dihydro-3-phenyl[1,2,4]oxadiazolo[5,4-a][2,3]benz odiazepin-6-ones (2,3-OBZs) were studied after intraperitoneal (i.p.) administration in DBA/2 mice, a strain genetically susceptible to sound-induced seizures. The seizures were evoked by means of auditory stimulation (109 dB, 12-16 kHz) in animals placed singly under a hemispheric Perspex dome. The 2,3-benzodiazepines studied after 30 min pretreatment were generally less potent than the related derivative 1-(4-aminophenyl)-4-methyl-7,8-methylenedioxy-5H-2,3-benzodiazepine hydrochloride (GYKI 52466) except 3,5-dihydro-7,8-dimethoxy-1-phenyl-4H-2,3-benzodiazepin-4-one (2,3-BZ-2) and 2,3-BZ-2M (3-methyl derivative of 2,3-BZ-2) which showed comparable activity. Thirty minutes after i.p. administration of 2,3-benzodiazepines, the rank order of potency for anticonvulsant activity against clonus was 2,3-BZ-2 > GYKI 52466 > 2,3-BZ-2M > 2,3-BZ-1 > 2,3-BZ-3, > 2,3-OBZ-1, > 2,3-OBZ-2 2,3-OBZ-3. The intracerebroventricular (i.c.v.) injection of aniracetam on it own (12.5 - 100 nmol/mouse) had no convulsant activity, but it reversed the anticonvulsant effects of some 2,3-benzodiazepines. In particular, the pharmacological actions of GYKI 52466, 2,3-BZ-2 and 2,3-BZ-2M, which proved to be the most potent 2,3-benzodiazepine derivatives as anticonvulsants, were significantly reduced by an i.c.v. pretreatment with aniracetam (50 nmol/mouse). Concomitant treatment with aniracetam (50 nmol/mouse) shifted to the right the dose-response curves and significantly increased the ED50 values for GYKI 52466, 2,3-BZ-2 and 2,3-BZ-2M. After 30 min pretreatment 2,3-BZ-2 showed a similar potency to GYKI 52466 in antagonizing seizures induced by i.c.v. administration of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), thus suggesting a clear involvement of AMPA receptors in the anticonvulsant activity of these compounds. In addition, 2,3-BZ-2 and 2,3-BZ-2M showed anticonvulsant properties longer lasting than GYKI 52466.

AMPA receptor antagonists as potential anticonvulsant drugs.

Over the last years alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid glutamate receptors (AMPARs) have been intensively studied owing to their crucial role in physiological and pathological processes. Efforts targeting AMPAR have been focused on identification of ligands as potential therapeutic agents useful in the prevention and treatment of a variety of neurological and non-neurological diseases. In particular, extensive work was addressed to the discovery of selective antagonists some of which proved to be potent anticonvulsant agents.

Comparative anticonvulsant activity of some 2,3-benzodiazepine derivatives in rodents

The anticonvulsant activities of some 2-amino-3-(3-hydroxy-5-methylisoxazol-4-yl)propionic acid (AMPA)/kainate receptor antagonists, noncompetitive (2,3-benzodiazepines) and a competitive 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)-quinoxaline (NBQX), were compared in different experimental seizure models. In particular, compounds were evaluated against audiogenic seizure in DBA/2 mice, maximal electroshock seizure (MES) test and various chemoconvulsant models; both groups showed a protective action against audiogenic seizure, MES- and pentylenetetrazole (PTZ)-induced seizures. All 2,3-benzodiazepines were also protective against clonic and tonic seizures and lethality induced by 4-aminopyridine, kainate, AMPA and 3-mercaptopropionic acid but were ineffective against NMDA-induced seizures. NBQX was unable to affect 4-aminopyridine-, mercaptopropionic acid- and NMDA-induced seizures. The duration of anticonvulsant action of 33 micromol/kg of some 2,3-benzodiazepine in DBA/2 mice, genetically susceptible to audiogenic seizures, was also investigated. The derivatives possessing a thiocarbonyl group at the C-4 position of heptatomic ring showed higher anticonvulsant activities and longer lasting protective effects. We conclude that all 2,3-benzodiazepines studied are effective against various models of experimental epilepsy and the presence of thiocarbonyl groups at the C-4 position of heptatomic ring is able to increase the anticonvulsant effect of these compounds.

AMPA receptor antagonists

It is now generally recognised that glutamate is the major excitatory neurotransmitter in vertebrate central nervous system (CNS). It acts at ionotropic and metabotropic receptors which appear to play important roles in all aspects of CNS functions. The ionotropic receptors, which are involved in fast synaptic transmission, belong to three subtypes named after three relatively selective agonists: NMDA (N-methyl-D-aspartate, AMPA [2- amino-3-(3-hydroxy-5-methyl-4-isoxazolyl)-propionate] and KA (kainate) receptors. The compounds acting as AMPA antagonists are potentially useful for the prevention and treatment of a broad range of acute and chronic neurological disorders. This article describes the development of recent AMPA receptor antagonists, reviewing both the primary and patent literature.

HIV-1 integrase strand-transfer inhibitors: design, synthesis and molecular modeling investigation.

This study is focused on a new series of benzylindole derivatives with various substituents at the benzene-fused ring, suggested by our 3D pharmacophore model developed for HIV-1 integrase inhibitors (INIs). All synthesized compounds proved to be active in the nanomolar range (6-35 nM) on the strand-transfer step (ST). In particular, derivative 4-[1-(4-fluorobenzyl)-5,7-dimethoxy-1H-indol-3-yl]-2-hydroxy-4-oxobut-2-enoic acid (8e), presenting the highest best-fit value on pharmacophore model, showed a potency comparable to that of clinical INSTIs GS 9137 (1) and MK-0518 (2). The binding mode of our molecules has been investigated using the recently published crystal structure of the complex of full-length integrase from the prototype foamy virus in complex with its cognate DNA (PFV-IN/DNA). The results highlighted the ability of derivative 8e to assume the same binding mode of MK-0518 and GS 9137.

Comparative anticonvulsant activity of N-acetyl-1-aryl-6,7-dimethoxy-1,2,3,4-tetrahydroisoquinoline derivatives in rodents

The anticonvulsant activity of competitive 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo (F)-quinoxaline (NBQX) and noncompetitive 2,3-benzodiazepines and tetrahydroisoquinolines (THIQs) AMPA/kainate receptor antagonists, was tested in different experimental seizure models and compared with diazepam, a conventional antiepileptic drug acting on GABAergic neurotransmission. In particular, the compounds were evaluated against audiogenic and maximal electroshock seizures (MES) test and pentetrazol (PTZ) seizures model, and all of them showed protective action. In addition, NBQX, 2,3-benzodiazepines and THIQs, but not diazepam, were also protective against clonic and tonic seizures and lethality induced by kainate, AMPA and ATPA, but were ineffective against NMDA-induced seizures. Only 2,3-benzodiazepines and some THIQs were able to affect 4-aminopyridine- and mercaptopropionic-acid-induced seizures. The duration of anticonvulsant action of 33 micromol/kg of some 2,3-benzodiazepines and THIQs was also investigated in DBA/2 mice, a strain genetically susceptible to audiogenic seizures, and it was observed that the derivative THIQ-10c, possessing an acetyl group at the N-2 and a chlorine atom on the C-1 phenyl ring, showed higher anticonvulsant activity and longer-lasting protective effects.

Annelated 1,5-Benzodiazepines. Part 1. Three, Four, and Five membered Rings

This review describes the synthetic approaches to mono and diannelated 1,5-benzodiazepines with three, four, and five-membered rings fused to different positions of the 1,5-benzodiazepine skeleton

Structure-based screening for the discovery of new carbonic anhydrase VII inhibitors

Among the different mammalian isoforms of Carbonic Anhydrase, the hCA VII is mainly expressed in the brain where it is involved in several neurological diseases. Thereby hCA VII has been validated as an attractive target for the discovery of selective inhibitors for the treatment of epilepsy and neurological pain. To identify new chemical entities as carbonic anhydrase inhibitors (CAIs) targeting hCA VII, we used a structure-based approach. By means of LigandScout software we built pharmacophore models from crystal structures of two well-known CAIs in complex with hCA VII. A merged pharmacophore hypothesis has been obtained. Subsequently, a focused library of compounds was screened against pharmacophore model and the most interesting hits were docked into the crystal structure of hCA VII. As a result, we identified new compounds displaying significant CA inhibitory effects in the nanomolar range.