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Neuropeptide S (NPS) regulates different biological functions by selectively activating a G protein coupled receptor named NPSR. Previous studies identified [ d-Pen]NPS as a NPSR antagonist. This study investigated the structure activity relationship of [ d-Pen]NPS by alkylation of the thiol group of d-Pen with different aliphatic and aromatic moieties. Nine compounds were synthesised and assayed in calcium mobilization studies, performed in HEK293 cells expressing the recombinant mouse NPSR. All compounds behaved as pure NPSR antagonists of moderate to high potency, in a similar manner to the reference peptide. The most potent compounds were obtained using alkylating groups as cyclic aromatic moieties. [ d-Pen( p-BuBzl)]NPS, the best compound of this series, was further investigated in vivo, where it fully prevented the stimulatory effects of supraspinal NPS on mouse locomotor activity. [ d-Pen( p-BuBzl)]NPS is proposed as a novel pharmacological tool which could be useful to investigate in vitro and in vivo biological functions regulated by the NPS/NPSR receptor system. [ABSTRACT FROM AUTHOR]

Background and purpose:Nocistatin (NST) is a neuropeptide generated from cleavage of the nociceptin/orphanin FQ (N/OFQ) precursor. Evidence has been presented that NST acts as a functional antagonist of N/OFQ, although NST receptor and transduction pathways have not yet been identified. We previously showed that N/OFQ inhibited [3H]5-hydroxytryptamine ([3H]5-HT) release from mouse cortical synaptosomes via activation of NOP receptors. We now investigate whether NST regulates [3H]5-HT release in the same preparation.Experimental approach:Mouse and rat cerebrocortical synaptosomes in superfusion, preloaded with [3H]5-HT and stimulated with 1 min pulses of 10 mM KCl, were used.Key results:Bovine NST (b-NST) inhibited the K+-induced [3H]5-HT release, displaying similar efficacy but lower potency than N/OFQ. b-NST action underwent concentration-dependent and time-dependent desensitization, and was not prevented either by the NOP receptor antagonist [Nphe1 Arg14,Lys15]N/OFQ(1-13)-NH2 (UFP-101) or by the non-selective opioid receptor antagonist, naloxone. Contrary to N/OFQ, b-NST reduced [3H]5-HT release from synaptosomes obtained from NOP receptor knockout mice. However, both N/OFQ and NST were ineffective in synaptosomes pre-treated with the Gi/o protein inhibitor, Pertussis toxin. NST-N/OFQ interactions were also investigated. Co-application of maximal concentrations of both peptides did not result in additive effects, whereas pre-application of maximal b-NST concentrations partially attenuated N/OFQ inhibition.Conclusions and implications:We conclude that b-NST inhibits [3H]5-HT release via activation of Gi/o protein linked pathways, not involving classical opioid receptors and the NOP receptor. The present data strengthen the view that b-NST is, per se, a biologically active peptide endowed with agonist activity.British Journal of Pharmacology (2007) 152, 549–555; doi:10.1038/sj.bjp.0707377; published online 9 July 2007 [ABSTRACT FROM AUTHOR]

In the present study we describe the in vitro pharmacological characterization of the nociceptin/orphanin FQ (N/OFQ) receptor (NOP) ligand Ac-RYYRWK-NH2 and the synthesis and biological evaluation of 13 Trp5 substituted Ac-RYYRWK-NH2 analogs. Results indicate that Ac-RYYRWK-NH2 behaves as a highly potent and selective partial agonist at the NOP receptors and that the whole indole moiety of the Trp5 side chain is not required, being a phenyl-ethyl side chain already sufficient for maintaining high potency. [ABSTRACT FROM AUTHOR]

In the present study we describe the in vitro pharmacological characterization of the nociceptin/orphanin FQ (N/OFQ) receptor (NOP) ligand Ac-RYYRWK-NH2 and the synthesis and biological evaluation of 13 Trp5 substituted Ac-RYYRWK-NH2 analogs. Results indicate that Ac-RYYRWK-NH2 behaves as a highly potent and selective partial agonist at the NOP receptors and that the whole indole moiety of the Trp5 side chain is not required, being a phenyl-ethyl side chain already sufficient for maintaining high potency. [ABSTRACT FROM AUTHOR]

Background and Purpose: Fentanyl analogues have been implicated in many cases of intoxication and death with overdose worldwide. The aim of this study is to investigate the pharmaco-toxicology of two fentanyl analogues: butyrylfentanyl (BUF) and 4-fluorobutyrylfentanyl (4F-BUF)., Experimental Approach: In vitro, we measured agonist opioid receptor efficacy, potency, and selectivity and ability to promote interaction of the μ receptor with G protein and β-arrestin 2. In vivo, we evaluated thermal antinociception, stimulated motor activity and cardiorespiratory changes in female and male CD-1 mice injected with BUF or 4F-BUF (0.1-6 mg·kg -1 ). Opioid receptor specificity was investigated using naloxone (6 mg·kg -1 ). We investigated the possible role of stress in increasing cardiorespiratory toxicity using the corticotropin-releasing factor 1 (CRF 1 ) antagonist antalarmin (10 mg·kg -1 )., Key Results: Agonists displayed the following rank of potency at μ receptors: fentanyl > 4F-BUF > BUF. Fentanyl and BUF behaved as partial agonists for the β-arrestin 2 pathway, whereas 4F-BUF did not promote β-arrestin 2 recruitment. In vivo, we revealed sex differences in motor and cardiorespiratory impairments but not antinociception induced by BUF and 4F-BUF. Antalarmin alone was effective in blocking respiratory impairment induced by BUF in both sexes but not 4F-BUF. The combination of naloxone and antalarmin significantly enhanced naloxone reversal of the cardiorespiratory impairments induced by BUF and 4F-BUF in mice., Conclusion and Implications: In this study, we have uncovered a novel mechanism by which synthetic opioids induce respiratory depression, shedding new light on the role of CRF 1 receptors in cardiorespiratory impairments by μ agonists., (© 2024 The Author(s). British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.)

Background: A neuroinflammatory disease such as Alzheimer's disease, presents a significant challenge in neurotherapeutics, particularly due to the complex etiology and allostatic factors, referred to as CNS stressors, that accelerate the development and progression of the disease. These CNS stressors include cerebral hypo-glucose metabolism, hyperinsulinemia, mitochondrial dysfunction, oxidative stress, impairment of neuronal autophagy, hypoxic insults and neuroinflammation. This study aims to explore the efficacy and safety of DAG-MAG-ΒHB, a novel ketone diester, in mitigating these risk factors by sustaining therapeutic ketosis, independent of conventional metabolic pathways., Methods: We evaluated the intestinal absorption of DAG-MAG-ΒHB and the metabolic impact in human microglial cells. Utilizing the HMC3 human microglia cell line, we examined the compound's effect on cellular viability, Acetyl-CoA and ATP levels, and key metabolic enzymes under hypoglycemia. Additionally, we assessed the impact of DAG-AG-ΒHB on inflammasome activation, mitochondrial activity, ROS levels, inflammation and phagocytic rates., Results: DAG-MAG-ΒHB showed a high rate of intestinal absorption and no cytotoxic effect. In vitro, DAG-MAG-ΒHB enhanced cell viability, preserved morphological integrity, and maintained elevated Acetyl-CoA and ATP levels under hypoglycemic conditions. DAG-MAG-ΒHB increased the activity of BDH1 and SCOT, indicating ATP production via a ketolytic pathway. DAG-MAG-ΒHB showed remarkable resilience against low glucose condition by inhibiting NLRP3 inflammasome activation., Conclusions: In summary, DAG-MAG-ΒHB emerges as a promising treatment for neuroinflammatory conditions. It enhances cellular health under varying metabolic states and exhibits neuroprotective properties against low glucose conditions. These attributes indicate its potential as an effective component in managing neuroinflammatory diseases, addressing their complex progression.

Herein we present the biocatalysed preparation of a mono- N -carbamate-protected precursor of antitumoral Nutlin-3a through enantioselective alkoxycarbonylation of meso -1,2-disubstituted-1,2-diaminoethane using enzyme lipases and dialkyl carbonates as acylating agents. A series of supported or free lipase enzymes were screened in combination with commercially available diallyl, diethyl and dimethyl carbonates. The reactions were conducted at different temperatures, for different reaction times and with variable co-solvent systems to evaluate the effects on the enzyme catalytic activity. The best results in terms of conversion, enantiomeric excess and yield were obtained when lipase from Candida antarctica B (CAL-B) was used with diallyl carbonate (DAC) when conducting the reaction solventless at 75 °C., Competing Interests: The authors declare no conflicts of interest.

Some new hemorphin-4 analogs with structures of Xxx-Pro-Trp-Thr-NH 2 and Tyr-Yyy-Trp-Thr-NH 2 , where Xxx is 2-amino-3-(4-hydroxy-2,6-dimethylphenyl)propanoic acid or 2-amino-3-(4-dibenzylamino-2,6-dimethylphenyl)propanoic acid, and Yyy is (2S,4S)-4-amino-pyrrolidine-2-carboxylic acid, were synthesized and characterized by electrochemical and spectral analyses. In vivo anticonvulsant and antinociceptive activities of peptide derivatives were studied after intracerebroventricular injection in mice. The therapeutic effects of the modified peptides on seizures and pain in mice were evaluated to provide valuable insights into the potential applications of the novel compounds. Electrochemical characterization showed that the compounds behave as weak protolytes and that they are in a soluble, stable molecular form at physiological pH values. The antioxidant activity of the peptides was evaluated with voltammetric analyses, which were confirmed by applying the 2,2-Diphenyl-1-picrylhydrazyl method. The compounds showed satisfactory results regarding their structural stability, reaching the desired centers for the manifestation of biological activity without hydrolysis processes at 37°C and physiological pH. Dm-H4 and H4-P1 exhibited 100% and 83% potency to suppress the psychomotor seizures in the 6-Hz test compared to 67% activity of H4. Notably, only the H4-P1 had efficacy in blocking the tonic component in the maximal electroshock test with a potency comparable to H4. All investigated peptides containing unnatural conformationally restricted amino acids showed antinociceptive effects. The analogs Db-H4 and H4-P1 showed the most pronounced and long-lasting effect in both experimental models of pain induced by thermal and chemical stimuli. Dm-H4 produced a dose-dependent thermal antinociception and H4-P2 inhibited only formalin-induced pain behavior., (© 2024 Deutsche Pharmazeutische Gesellschaft.)

Various geraniol esters act as insect pheromones and display pharmacological activities, especially as neuroprotective agents. Therefore, the search for synthetic strategies alternative to traditional chemical synthesis could help designing ecofriendly routes for the preparation of such bioactive compounds. Hence, this work aims at the microwave-assisted enzymatic synthesis of geranyl esters in solvent-free systems. The process variables were optimized for the synthesis of geranyl acetoacetate, achieving 85% conversion after 60 min using a 1:5 substrates molar ratio (ester to geraniol), 80 °C and 8.4% of Lipozyme 435 lipase without removal of the co-produced methanol. On the other hand, a 95% conversion was reached after 30 min using 1:6 substrates molar ratio, 70 °C and 7% lipase in the presence of 5Å molecular sieves for the methanol capture. In addition, the lipase showed good reusability, maintaining the same activity for five reaction cycles. Finally, under the above optimized conditions, other geraniol esters were successfully synthetized such as the geranyl butyrate (98%), geranyl hexanoate (99%), geranyl octanoate (98%), and geranyl (R)-3-hydroxybutyrate (56%). These results demonstrate the microwave-assisted lipase-catalyzed transesterification in a solvent-free system as an excellent and sustainable catalytic methodology to produce geraniol esters., (© 2023. The Author(s).)

Finding a therapy for ischemia-reperfusion injury, which consists of cell death following restoration of blood flowing into the artery affected by ischemia, is a strong medical need. Nowadays, only the use of broad-spectrum molecular therapies has demonstrated a partial efficacy in protecting the organs following reperfusion, while randomized clinical trials focused on more specific drug targets have failed. In order to overcome this problem, we applied a combination of molecular modeling and chemical synthesis to identify novel spiropiperidine-based structures active in mitochondrial permeability transition pore opening inhibition as a key process to enhance cell survival after blood flow restoration. Our results were confirmed by biological assay on an in vitro cell model on HeLa and human renal proximal tubular epithelial cells and pave the way to further investigation on an in vivo model system.

Here, we propose the molecular hybridization of dihydroartemisinin (DHA) and ursodeoxycholic bile acid (UDCA), approved drugs, for the preparation of antiviral agents against SARS-CoV-2. DHA and UDCA were selected on the basis of their recently demonstrated in vitro activity against SARS-CoV-2. A selection of DHA-UDCA-based hybrids obtained by varying the nature of the linkage and the bile acid conjugation point as well as unconjugated DHA and UDCA were tested in vitro for cytotoxicity and anti-SARS-CoV-2 activity on Vero E6 and Calu-3 human lung cells. The hybrid DHA-t-UDCMe, obtained by conjugation via click chemistry on a gram scale, was identified as a potential candidate for SARS-CoV-2 infection treatment due to significant reduction of viral replication, possibly involving ACE2 downregulation, no cytotoxicity, and chemical stability., Competing Interests: The authors declare no competing financial interest., (© 2023 The Authors. Published by American Chemical Society.)

The enormous influence in terms of bioactivity, affinity, and selectivity represented by the replacement of (L)-2,6-dimethyl tyrosine (Dmt) instead of Phenylalanine (Phe) into Nociceptin/orphanin (N/OFQ) neuropeptide analogues has been well documented in the literature. More recently, the non-natural amino acid (L)-2-methyl tyrosine (Mmt), with steric hindrance included between Tyr and Dmt, has been studied because of the modulation of steric effects in opioid peptide chains. Here, we report a new synthetic strategy to obtain Mmt based on the well-known Pd-catalyzed ortho -C(sp 2 )-H activation approach, because there is a paucity of other synthetic routes in the literature to achieve it. The aim of this work was to force only the mono- ortho -methylation process over the double ortho -methylation one. In this regard, we are pleased to report that the introduction of the dibenzylamine moiety on a Tyr aromatic nucleus is a convenient and traceless solution to achieve such a goal. Interestingly, our method provided the aimed Mmt either as N -Boc or N -Fmoc derivatives ready to be inserted into peptide chains through solid-phase peptide synthesis (SPPS). Importantly, the introduction of Mmt in place of Phe 1 in the sequence of N/OFQ(1-13)-NH 2 was very well tolerated in terms of pharmacological profile and bioactivity.

Permeability transition pore (PTP) molecular composition and activity modulation have been a matter of research for several years, especially due to their importance in ischemia reperfusion injury (IRI). Notably, c subunit of ATP synthase (Csub) has been identified as one of the PTP-forming proteins and as a target for cardioprotection. Oligomycin A is a well-known Csub interactor that has been chemically modified in-depth for proposed new pharmacological approaches against cardiac reperfusion injury. Indeed, by taking advantage of its scaffold and through focused chemical improvements, innovative Csub-dependent PTP inhibitors (1,3,8-Triazaspiro[4.5]decane) have been synthetized in the past. Interestingly, four critical amino acids have been found to be involved in Oligomycin A-Csub binding in yeast. However, their position on the human sequence is unknown, as is their function in PTP inhibition. The aims of this study are to (i) identify for the first time the topologically equivalent residues in the human Csub sequence; (ii) provide their in vitro validation in Oligomycin A-mediated PTP inhibition and (iii) understand their relevance in the binding of 1,3,8-Triazaspiro[4.5]decane small molecules, as Oligomycin A derivatives, in order to provide insights into Csub interactions. Notably, in this study we demonstrated that 1,3,8-Triazaspiro[4.5]decane derivatives inhibit permeability transition pores through a F O -ATP synthase c subunit Glu 119 -independent mechanism that prevents Oligomycin A-related side effects.