Maria Rosa Buemi

Synthesis and biological characterization of 3-substituted-1H-indoles as ligands of GluN2B-containing N-methyl-D-aspartate receptors.

As an extension of our studies, novel indole derivatives were rationally designed and synthesized as ligands targeted to GluN2B/NMDA receptors. The 2-(4-benzylpiperidin-1-yl)-1-(6-hydroxy-1H-indol-3-yl)ethanone (4i) and 1-(4-benzylpiperidin-1-yl)-2-(6-hydroxy-1H-indol-3-yl)ethane-1,2-dione (6i) showed high binding affinity in [3H]ifenprodil displacement assay. By computational studies, we suggested the hypothetical interactions playing a significant role during the binding process. However, in functional and in vivo studies the most potent compound 4i did not show any activity whereas it displayed relevant affinity toward the σ2 receptor.

Indole derivatives as dual-effective agents for the treatment of neurodegenerative diseases: Synthesis, biological evaluation, and molecular modeling studies

Several indole derivatives, that were highly potent ligands of GluN2B-subunit-containing N-methyl-D-aspartate (NMDA) receptor, also demonstrated antioxidant properties in ABTS method. In particular, the 2-(4-benzylpiperidin-1-yl)-1-(5-hydroxy-1H-indol-3-yl)ethanone (1) proved to be a dual-effective neuroprotective agent. With the aim to increase the antioxidant properties we added a catechol moiety onto piperidine moiety. The designed hybrid derivative 3,4-dihydroxy-N-[1-[2-(5-hydroxy-1H-indol-3-yl)-2-oxoethyl]piperidin-4-yl]benzamide (10) was the most effective antioxidant agent (>94.1 ± 0.1% of inhibition at 17 μM) and showed GluN2B/NMDA receptor affinity at low micromolar concentration (IC₅₀ 0.66 μM). By means of computational studies we explored the effect of the presence of this antioxidant fragment during the recognition process to binding pocket.

Probing Molecular Interactions between Human Carbonic Anhydrases (hCAs) and a Novel Class of Benzenesulfonamides

On the basis of X-ray crystallographic studies of the complex of hCA II with 4-(3,4-dihydro-1H-isoquinoline-2-carbonyl)benzenesulfonamide (3) (PDB code 4Z1J ), a novel series of 4-(1-aryl-3,4-dihydro-1H-isoquinolin-2-carbonyl)benzenesulfonamides (23-33) was designed. Specifically, our idea was to improve the selectivity toward druggable isoforms through the introduction of additional hydrophobic/hydrophilic functionalities. Among the synthesized and tested compounds, the (R,S)-4-(6,7-dihydroxy-1-phenyl-3,4-tetrahydroisoquinoline-1H-2-carbonyl)benzenesulfonamide (30) exhibited a remarkable inhibition for the brain-expressed hCA VII (Ki = 0.20 nM) and selectivity over wider distributed hCA I and hCA II isoforms. By enantioselective HPLC, we solved the racemic mixture and ascertained that the two enantiomers (30a and 30b) are equiactive inhibitors for hCA VII. Crystallographic and docking studies revealed the main interactions of these inhibitors into the carbonic anhydrase (CA) catalytic site, thus highlighting the relevant role of nonpolar contacts for this class of hCA inhibitors.

Carbonic anhydrase inhibitors: Design, synthesis and structural characterization of new heteroaryl-N-carbonylbenzenesulfonamides targeting druggable human carbonic anhydrase isoforms.

A set of heteroaryl-N-carbonylbenzenesulfonamides has been designed, synthesized, and screened as inhibitors of human carbonic anhydrases (hCAs). The new sulfonamide derivatives were tested against hCA I, hCA II, hCA VII, hCA IX, and hCA XII isoforms using acetazolamide (AAZ, 1) and topiramate (TPM, 2) as reference compounds. Six compounds were low nanomolar inhibitors of tumor-associated hCA IX isoform (Ki values < 10 nM); among them we identified three arylsulfonamides showing unexpected inefficacy over brain distributed hCA VII isoform (hCA IX/hCA VII selectivity ratio > 1500 for compound 5c). Thus, these compounds can offer the opportunity to highlight the interactions preventing the inhibition of hCA VII mainly expressed in central nervous system. Thereby, we used structural and computational techniques to study in depth the interaction with hCAs. In an effort to confirm the inhibitory action we determined crystal structures of five selected heteroaryl-N-carbonylbenzenesulfonamides (4a, 4b, 4e, 5c, and 5e) in complex with hCA II. Moreover, to explore the lack of inhibitory effects of selected compounds (e.g.4b and 5c) we also performed docking studies into hCA VII catalytic site.

In Vivo Evaluation of Selective Carbonic Anhydrase Inhibitors as Potential Anticonvulsant Agents

Epilepsy is a common neurological disorder caused by an imbalance between inhibitory and excitatory neurotransmission. It is well known that neuronal excitability is related to γ‐aminobutyric acid (GABA)ergic depolarization. HCO3−‐dependent depolarization can be suppressed by membrane‐permeable inhibitors of carbonic anhydrase. We previously identified some isoquinoline sulfonamides as potent and selective inhibitors of the human carbonic anhydrase II and VII (hCA II and hCA VII) isoforms. Given that hCA II and hCA VII are specific isoforms involved in GABA‐mediated neuronal excitation, we hypothesized that they could represent the biological target for the development of new anticonvulsant agents. Therefore, for selected isoquinoline sulfonamides, we preliminarily tested their ability to prevent audiogenic seizures in DBA/2 mice. All compounds were evaluated after intraperitoneal administration, and some of them proved to protect the mice against convulsions. Among this series of compounds, several derivatives showed combined in vivo efficacy with inhibitory effects toward the targeted carbonic anhydrases (i.e., hCA II and hCA VII). Specifically, the most interesting molecule was 1‐(4‐aminophenyl)‐6,7‐dimethoxy‐3,4‐dihydroisoquinoline‐2(1H)‐sulfonamide (6), which proved to be a more active and selective hCA VII inhibitor than the reference compound topiramate. Further studies to explore the in vivo pharmacokinetic profile of the most active compounds may help to provide insight into the future design of selective hCA VII inhibitors.

From NMDA receptor antagonists to discovery of selective σ2 receptor ligands

Following previous studies focused on the search for new molecules targeting GluN2B-containing NMDA, a small series of 1-(1H-indol-3-yl)-2-(4-phenylpiperidin-1-yl)ethanone derivatives has been synthesized by using Microwave Assisted Organic Synthesis (MAOS). Given that GluN2B ligands frequently exert off-target effects we also tested their affinity towards sigma receptors. Binding assay revealed that only the 1-(5-hydroxy-1H-indol-3-yl)-2-(4-phenylpiperidin-1-yl)ethanone (7a) retained GluN2B affinity. Interestingly, the 5-methoxyindoles 5a and 6a were efficient and selective ligands toward σ₂ receptor (Ki values of 10nM and 20 nM, respectively). Thus, in this case the discovery of new σ₂ receptor selective ligands was an unexpected result emerging from the screening of cross-activity against other CNS receptors.

Searching for novel N1-substituted benzimidazol-2-ones as non-nucleoside HIV-1 RT inhibitors

Non-nucleoside reverse transcriptase inhibitors (NNRTIs) represent an integral part of the currently available combination antiretroviral therapy (cART) contributing to reduce the AIDS-mortality and turned the disease from lethal to chronic. In this context we recently reported a series of 6-chloro-1-(3-methylphenylsulfonyl)-1,3-dihydro-2H-benzimidazol-2-ones as potent non-nucleoside HIV-1 reverse transcriptase inhibitors. In this paper, we describe the design and the synthesis of two novel series of benzimidazolone analogues in which the linker moiety between the phenyl ring and the sulfonyl group was modified and new small lipophilic groups on the benzyl sulfonyl pendant were introduced. All the new obtained compounds were evaluated as RT inhibitors and were also tested against RTs containing single amino acid mutations. Finally, molecular docking studies were performed in order to rationalize the observed activity of the most promising compound.

Targeting GluN2B-containing N-Methyl-D-aspartate receptors: design, synthesis, and binding affinity evaluation of novel 3-substituted indoles.

In an effort to improve our knowledge about structure–affinity relationships (SARs) for a class of 3‐substituted‐indole derivatives as GluN2B‐containing N‐methyl‐D‐aspartate‐type receptor (NMDAR) ligands, we herein describe the design, synthesis, and preliminary screening of a new series of molecules. The in vitro determination of binding affinities suggested that 5‐hydroxy‐ and 6‐hydroxyindole derivatives 12 and 13 were active ligands. Generally, the novel compounds proved to be less potent than their homologs previously reported as promising neuroprotective agents. In fact, our lead compound 3‐(4‐benzylpiperidin‐1‐yl)‐1‐(5‐hydroxy‐1H‐indol‐3‐yl)ethan‐1‐one (2) was about 10‐fold more active than the new propan‐1‐one derivative (12). To rationalize the low potency of the new analog 12, docking studies were also performed and the in silico results were consistent with the in vitro data.

Structural optimization of N 1 -aryl-benzimidazoles for the discovery of new non-nucleoside reverse transcriptase inhibitors active against wild-type and mutant HIV-1 strains

Non-nucleoside reverse transcriptase inhibitors (NNRTIs) are recommended components of preferred combination antiretroviral therapies used for the treatment of human immunodeficiency virus (HIV) infection. These regimens are extremely effective in suppressing virus replication. Recently, our research group identified some N1-aryl-2-arylthioacetamido-benzimidazoles as a novel class of NNRTIs. In this research work we report the design, the synthesis and the structure-activity relationship studies of new compounds (20-34) in which some structural modifications have been introduced in order to investigate their effects on reverse transcriptase (RT) inhibition and to better define the features needed to increase the antiviral activity. Most of the new compounds proved to be highly effective in inhibiting both RT enzyme at nanomolar concentrations and HIV-1 replication in MT4 cells with minimal cytotoxicity. Among them, the most promising N1-aryl-2-arylthioacetamido-benzimidazoles and N1-aryl-2-aryloxyacetamido-benzimidazoles were also tested toward a panel of single- and double-mutants strain responsible for resistance to NNRTIs, showing in vitro antiviral activity toward single mutants L100I, K103N, Y181C, Y188L and E138K. The best results were observed for derivatives 29 and 33 active also against the double mutants F227L and V106A. Computational approaches were applied in order to rationalize the potency of the new synthesized inhibitors.

Rational Design, Synthesis and Evaluation of Coumarin Derivatives as Protein‐protein Interaction Inhibitors

Herein we describe the design and synthesis of a new series of coumarin derivatives searching for novel HIV‐1 integrase (IN) allosteric inhibitors. All new obtained compounds were tested in order to evaluate their ability to inhibit the interaction between the HIV‐1 IN enzyme and the nuclear protein lens epithelium growth factor LEDGF/p75. A combined approach of docking and molecular dynamic simulations has been applied to clarify the activity of the new compounds. Specifically, the binding free energies by using the method of molecular mechanics‐generalized Born surface area (MM‐GBSA) was calculated, whereas hydrogen bond occupancies were monitored throughout simulations methods.