Emanuela Arena

Anti‐amnesic properties of (±)‐PPCC, a novel sigma receptor ligand, on cognitive dysfunction induced by selective cholinergic lesion in rats

J. Neurochem. (2009) 109, 744–754.

Synthesis and resolution of cis-(+/-)-methyl (1R,2S/1S,2R)-2-[(4-hydroxy-4-phenylpiperidin-1-yl)methyl]-1-(4-methylphenyl)cyclopropanecarboxylate [(+/-)-PPCC)]: new sigma receptor ligands with neuroprotective effect.

The enantiomers of cis-(+/-)-methyl (1R,2S/1S,2R)-2-[(4-hydroxy-4-phenylpiperidin-1-yl)methyl]-1-(4-methylphenyl)cyclopropanecarboxylate [1, (+/-)-PPCC], a selective sigma ligand, were synthesized. The (+)- and (-)-enantiomers bind predominantly to sigma(1) receptors and have a reduced sigma(2) affinity. Both individually restore the astroglial oxidative status modified by glutamate, counteracting also transglutaminase-2 overexpression. They exhibited in vivo anti-opioid effects on kappa opioid (KOP) receptor-mediated analgesia. Our findings demonstrate that the enantiomers display mainly sigma(1) agonist activity and that they have neuroprotective effects.

Design and synthesis of new bifunctional sigma-1 selective ligands with antioxidant activity.

Herein we report the synthesis of new bifunctional sigma-1 (σ1)-selective ligands with antioxidant activity. To achieve this goal, we combined the structure of lipoic acid, a universal antioxidant, with an appropriate sigma aminic moiety. Ligands 14 and 26 displayed high affinity and selectivity for σ1 receptors (Kiσ1 = 1.8 and 5.5 nM; Kiσ2/σ1 = 354 and 414, respectively). Compound 26 exhibited in vivo antiopioid effects on kappa opioid (KOP) receptor-mediated analgesia. In rat liver and brain mitochondria (RLM, RBM), this compound significantly reduced the swelling and the oxidation of thiol groups induced by calcium ions. Our results demonstrate that the tested compound has protective effects against oxidative stress.

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.

Development of novel LP1-based analogues with enhanced delta opioid receptor profile

Pain relief achieved by co-administration of drugs acting at different targets is more effective than that obtained with conventional MOR selective agonists usually associated to relevant side effects. It has been demonstrated that simultaneously targeting different opioid receptors is a more effective therapeutic strategy. Giving the promising role for DOR in pain management, novel LP1-based analogues with different N-substituents were designed and synthesized with the aim to improve DOR profile. For this purpose, we maintained the phenyl ring in the N-substituent of 6,7-benzomorphan scaffold linked to an ethyl spacer bearing a hydroxyl/methyl or methoxyl group at carbon 2 or including it in a 1,4-benzodioxane ring. LP1 analogues were tested by competition binding assays. Compounds 6 (KiMOR=2.47nM, KiDOR=9.6nM), 7 (KiMOR=0.5nM and KiDOR=0.8nM) and 9 (KiMOR=1.08nM, KiDOR=6.6nM) retained MOR affinity but displayed an improved DOR binding capacity as compared to LP1 (KiMOR=0.83nM, KiDOR=29.1nM). Moreover, GPI and MVD functional assays indicated that compounds 6 (IC50=49.2 and IC50=10.8nM), 7 (IC50=9.9 and IC50=11.8nM) and 9 (IC50=21.5 and IC50=4.4nM) showed a MOR/DOR agonist profile, unlike LP1 that was a MOR agonist/DOR antagonist (IC50=1.9 and IC50=1240nM). Measurements of their antinociceptive effect was evaluated by mice radiant tail flick test displaying for compounds 6, 7 and 9 ED50 values of 1.3, 1.0 and 0.9mg/kg, i.p., respectively. Moreover, the antinociceptive effect of compound 9 was longer lasting with respect to LP1. In conclusion the N-substituent nature of compounds 6, 7 and 9 shifts the DOR profile of LP1 from antagonism to agonism.

(+)-Pentazocine reduces oxidative stress and apoptosis in microglia following hypoxia/reoxygenation injury

BACKGROUND Sigma-1 receptors (σ1R) are highly expressed in neurons as well as microglia and have been shown to modulate the inflammatory response in the central nervous system and thus may serve as possible target for neuroprotective strategies. The aim of the present study was to test the effect of (+)-pentazocine, a putative σ 1R agonist, in an in vitro model of microglia activation. METHODS Microglia (BV2 cells) was exposed (3h) to 1% oxygen and reoxygenation was allowed for 24h. Cells were treated with different concentrations (1, 10, 25 and 50μM) of (+)-pentazocine in the presence or absence of NE-100 (1μM), a well established σ1R antagonist. Cell viability and apoptosis were measured by cytofluorimetric analysis, whereas oxidative stress was evaluated by reduced glutathione (GSH) content and mitochondrial potential analysis. RESULTS Our results showed that (+)-pentazocine was able to increase cell viability and restore mitochondrial potential at all concentrations whereas only 1 and 10μM were able to reduce significantly apoptotic cell death, to restore reduced glutathione intracellular content and prevent ERK1/2 phosphorylation. All these effects were abolished by concomitant treatment with NE-100. CONCLUSIONS (+)-pentazocine exhibits significant dose dependent protective effects in our in vitro model of microglial activation thus suggesting that σ1R may represent a possible target for neuroprotection.

Nanoencapsulation strategies for the delivery of novel bifunctional antioxidant/σ1 selective ligands

Nowadays sigma-1 receptors are considered as new therapeutic objectives for central nervous system neurodegenerative diseases. Among different molecules, alpha lipoic acid has been identified as a natural potent antioxidant drug, whose therapeutic efficacy is limited by its many drawbacks, such as fast metabolism, poor bioavailability and high physico-chemical instability. Alfa-lipoic acid derivatives have been recently developed demonstrating their neuroprotective activity and effectiveness in different types of oxidative stress. In this work, two derivatives containing an amide or an ester functional group with different lipophilicity, were selected for their important affinity for sigma-1 receptors. Herein, in order to improve the in vitro stability and antioxidant effectiveness of alpha-lipoic acid derivatives, we focused our efforts in the nanoencapsulation strategies. Aqueous-core nanocapsules for the delivery of the hydrophilic compound and nanostructured lipid carrier for the lipophilic derivative, were properly designed and prepared using a direct or inverse eco-friendly organic solvent-free procedure. All nanosystems were characterized in terms of mean size, polydispersity, stability, morphology, encapsulation efficiency and in vitro release profiles. In order to evaluate the nanocarriers biocompatibility and antioxidant effectiveness, in vitro biological studies (cell viability, total antioxidant capacity and total oxidative status) were developed on primary human whole blood cell cultures, on both unloaded and derivatives-loaded nanodevices.

New bifunctional antioxidant/σ1 agonist ligands: Preliminary chemico-physical and biological evaluation

We previously reported bifunctional sigma-1 (σ1) ligands endowed with antioxidant activity (1 and 2). In the present paper, pure enantiomers (R)-1 and (R)-2 along with the corresponding p-methoxy (6, 11), p-fluoro derivatives (7, 12) were synthesized. σ1 and σ2 affinities, antioxidant properties, and chemico-physical profiles were evaluated. Para derivatives, while maintaining strong σ1 affinity, displayed improved σ1 selectivity compared to the parent compounds 1 and 2. In vivo evaluation of compounds 1, 2, (R)-1, 7, and 12 showed σ1 agonist pharmacological profile. Chemico-physical studies revealed that amides 2, 11 and 12 were more stable than corresponding esters 1, 6 and 7 under our experimental conditions. Antioxidant properties were exhibited by fluoro derivatives 7 and 12 being able to increase total antioxidant capacity (TAC). Our results underline that p-substituents have an important role on σ1 selectivity, TAC, chemical and enzymatic stabilities. In particular, our data suggest that new very selective compounds 7 and 12 could be promising tools to investigate the disorders in which σ1 receptor dysfunction and oxidative stress are contemporarily involved.

(+)-Methyl (1R,2S)-2-{[4-(4-Chlorophenyl)-4-hydroxypiperidin-1-yl]methyl}-1-phenylcyclopropanecarboxylate [(+)-MR200] Derivatives as Potent and Selective Sigma Receptor Ligands: Stereochemistry and Pharmacological Properties

Methoxycarbonyl-1-phenyl-2-cyclopropylmethyl based derivatives cis-(+)-1a [cis-(+)-MR200], cis-(-)-1a [cis-(-)-MR201], and trans-(±)-1a [trans-(±)-MR204], have been identified as new potent sigma (σ) receptor ligands. In the present paper, novel enantiomerically pure analogues were synthesized and optimized for their σ receptor affinity and selectivity. Docking studies rationalized the results obtained in the radioligand binding assay. Absolute stereochemistry was unequivocally established by X-ray analysis of precursor trans-(+)-5a as camphorsulfonyl derivative 9. The most promising compound, trans-(+)-1d, showed remarkable selectivity over a panel of more than 15 receptors as well as good chemical and enzymatic stability in human plasma. An in vivo evaluation evidenced that trans-(+)-1d, in contrast to trans-(-)-1d, cis-(+)-1d, or cis-(-)-1d, which behave as σ1 antagonists, exhibited a σ1 agonist profile. These data clearly demonstrated that compound trans-(+)-1d, due to its σ1 agonist activity and favorable receptor selectivity and stability, provided an useful tool for the study of σ1 receptors.

Novel Sigma Receptors Ligands-Nitric Oxide Photodonor: Molecular Hybrids for Double-Targeted Antiproliferative Effect

This contribution reports the synthesis and evaluation of novel hybrid compounds that conjugate a sigma (σ) receptor pharmacophore and a nitric oxide (NO) photodonor. All compounds preserve their capability to generate NO under visible light and possess overall σ receptor nanomolar affinity, with one of them (8b) exhibiting remarkable σ2 receptor selectivity. Compounds 8b, 11a, and 11b were tested on tumorigenic MCF-7 and A2058 cells expressing high levels of σ2 and σ1 receptor, respectively. Considerable loss of cell viability was detected under light excitation, while negligible effects in the dark were detected. Moreover, they did not show any significant cytotoxicity in the dark or under irradiation on nontumorigenic NCTC-2544 keratinocytes. NO-induced reduction of cellular viability was demonstrated by in-cell NO detection and total nitrite estimation. For the first time, a combination of σ receptor moieties and a NO photodonor is reported, providing distinctive ligands potentially useful for cancer management.