Discovery of orexant and anorexant agents with indazole scaffold endowed with peripheral antiedema activity

CB1 receptors and endocannabinoids are integrated components of neuronal networks controlling different organism’s functions, such as appetite and food intake in the hypothalamus. A series of Rimonabant/Fubinaca hybrids have been synthesized in solution as C-terminal amides, acids, methyl esters and N-methyl amides. These compounds have been studied in cannabinoid receptor binding assay and functional receptor assay in vitro, the most active among them as agonist (LONI 11) and antagonist (LONI 4) were tested in vivo to evaluate their ability to stimulate or suppress the feeding behavior after i.p. administration. For LONI 11 formalin test and tail flick tests after s.c. and i.c.v. routes respectively, were also performed in vivo with the aim to investigate the antinociceptive effect at the central or peripheral level. In the Zymosan-induced edema and hyperalgesia, LONI 11 reduced the % paw volume increase and % paw latency after s.c. administration, also suggesting a potential anti-inflammatory activity at the periphery. Keywords. Cannabinoid receptor, Rimonabant, food intake, anorexant agent, edema

Food-inspired peptides from spinach Rubisco endowed with antioxidant, antinociceptive and anti-inflammatory properties

Rubiscolin-6 (amino acid sequence: YPLDLF) is a selective & delta;-opioid receptor peptide isolated from spinach Rubisco. Its synthetic analogue, peptide YPMDIV is the most potent described so far for its increased opioid activity, thus in this work it was considered as lead compound for the design of twelve new analogues e.g. LMAS112. Firstly all the novel compounds have been tested for their antinociceptive and anti-inflammatory capacity in vitro and in vivo in order to evaluate their ability to maintain or loss the original activity. Among them peptides LMAS5-8 gave the best results, thus their antioxidant properties have been investigated along with their enzymatic inhibitory ability. Peptide LMAS6 shows a strong antioxidant (154.25 mg TE/g CUPRAC) and inhibitor activity on tyrosinase (84.49 mg KAE/g), indicating a potential role in food industry as anti-browning agent, while peptides LMAS5 and LMAS7 possess a modest cholinesterase inhibitory activity suggesting a conceivable use for nutraceuticals production

Acupuncture decreases pain and MIF salivary levels in men and women

Acupuncture is used worldwide to treat many diseases, including painful conditions. Acupuncture-induced effects on pain were studied in men and women to understand the various factors involved in this positive procedure able to cure patients without side effects. Vital parameters (heart rate, blood pressure, temperature) were measured and questionnaires about quality of life (SF-36), mood state (POMS) and pain condition (QUID) were administered to men and women requesting acupuncture treatment for pain. Macrophage inhibitory factor (MIF) was determined in the saliva. All parameters were determined before the first and tenth acupuncture sessions. Pain scores (questionnaire and scales) significantly decreased from the first to the last acupuncture session in both sexes. Vital parameters were not affected, while the other questionnaires indicated a general improvement in the quality of life. MIF salivary levels were higher in males than in females and were significantly decreased by acupuncture in both sexes

Gender differences in pain and its relief

There is much evidence to suggest that gender is an important factor in the modulation of pain. Literature data strongly suggest that men and women differ in their responses to pain: they are more variable in women than men, with increased pain sensitivity and many more painful diseases commonly reported among women. Gender differences in pharmacological therapy and non-pharmacological pain interventions have also been reported, but these effects appear to depend on the treatment type and characteristics. It is becoming very evident that gender differences in pain and its relief arise from an interaction of genetic, anatomical, physiological, neuronal, hormonal, psychological and social factors which modulate pain differently in the sexes. Experimental data indicate that both a different modulation of the endogenous opioid system and sex hormones are factors influencing pain sensitivity in males and females. This brief review will examine the literature on sex differences in experimental and clinical pain, focusing on several biological mechanisms implicated in the observed gender-related differences. 

Opioid Receptor Activity and Analgesic Potency of DPDPE Peptide Analogues Containing a Xylene Bridge

d-Pen2,d-Pen5 enkephalin (DPDPE) is one of the most selective synthetic peptide agonists targeting the delta-opioid receptor. Three cyclic analogues of DPDPE containing a xylene bridge in place of disulfide bond have been synthesized and fully characterized as opioid receptors agonists. The in vitro activity was investigated showing a good affinity of 7a-c for mu- and delta-receptors. In vivo biological assays revealed that 7b is the most potent analogue with the ability to maintain high level of analgesia from 15 to 60 min following intracerebroventricular (i.c.v.) administration, whereas DPDPE was slightly active until 45 min. Compound 7b induced long lasting analgesia also after subcutaneous administration, whereas DPDPE was inactive

IL-17A neutralizing antibody regulates monosodium urate crystal-induced gouty inflammation

Gout is a paradigm of acute, self-limiting inflammation caused by the deposition of monosodium urate (MSU) crystals within intra-and/or peri-articular areas, leading to excruciating pain, joint swelling and stiffness. The infiltration of leukocytes drives the inflammatory response and remains an attractive target for therapeutic intervention. In this context, emerging evidence supports the view that systemic differentiation of Th17 cells and their in situ infiltration as one of the potential mechanisms by which these cells, and their main product IL-17, causes damage to target tissues. To test if IL-17 was having a detrimental role in gouty onset and progression we targeted this cytokine, using a neutralizing antibody strategy, in an experimental model of gout. Joint inflammation was induced in CD-1 mice by the intra-articular (i.a.) administration of MSU crystals (200 μg/20 μl). Animals from IL-17Ab-treated groups received 1, 3 and 10 μg (i.a.) in 20 μl of neutralizing antibody after MSU crystals administration. Thereafter, joints were scored macroscopically, and knee joint oedema determined with a caliper. Histological analysis, myeloperoxidase assay and western blots analysis for COX-2/mPGEs-1/IL-17R pathway were conducted at 18 h (peak of inflammation) to evaluate leukocytes infiltration and activation, followed by the analysis, in situ, of pro/anti-inflammatory cytokines and chemokines. Flow cytometry was also used to evaluate the modulation of infiltrated inflammatory monocytes and systemic Th17 and Treg profile. Treatment with IL-17Ab revealed a dose-dependent reduction of joint inflammation scores with maximal inhibition at 10 μg. The neutralizing antibody was also able to significantly reduce leukocytes infiltration and MPO activity as well the expression of JE, IL-1α, IL-1β, IL-16, IL-17, C5a, BLC and, with a less extent IP-10, Rantes, KC, TIMP-1, SDF-1 and metalloproteinases in inflamed tissues. Biochemical analysis also revealed that IL-17Ab treatment modulated COX-2/mPGEs-1 pathway (and related PGE2 production) without interfering with IL-17R expression. Furthermore, flow cytometry analysis highlighted a selective modulation of infiltrating inflammatory monocytes (B220-/GR1hi-F480hi/CD115+) and circulating Th17, but not Treg, cells after IL-17Ab treatment. Collectively the results of this study report for the first time, that i.a. injection of MSU crystals stimulates in vivo production of Th17 cells and Th17-related inflammatory cyto-chemokines. In addition, we have demonstrated that the administration of a neutralizing antibody against IL-17 attenuates joint symptoms, swelling and leukocytes infiltration to the inflamed tissue, possibly providing a new strategy for the treatment of gouty inflammation and/or arthritis

Design, synthesis and biological profile of mixed opioid agonist/N-VGCC blocker peptides

In this paper we reported the synthesis and the in vitro and the in vivo biological evaluation of linear pseudo-peptides incorporating the N-VGCC blocker tripeptide Phe-NMe-Leu-Tyr(OBz)-NtBu and the biphalin pharmacophore Tyr-D-Ala-Gly-Phe. The novel sequences have been designed by using amino acids of different length to join the two pharmacophores and explore the structure-activity relationships of the novel compounds

Pharmacological characterization of new compounds and herbal derivatives for pain and inflammation treatment

Introduction: Despite the availability of several analgesic and anti-inflammatory options, the effective treatment of pain and inflammation is still a challenge for clinicians, and the balance between efficacy and safety aspects remains both crucial and difficult. Thus, the identification of new potential targets which may affect pain and inflammatory processes is becoming an urgent clinical and therapeutic need. This scientific project derives from a close collaboration with various national and foreign research groups, in order to establish an efficient and highly multidisciplinary approach to investigate the pharmacological profile of new compounds and herbal derivatives with significant antinociceptive and antinflammatory effects to be used for the treatment of pain and inflammation. Materials and Methods: The research approach involves the in vivo studies of natural (ammonium glycyrrhizinate; rubiscolin-6 and soymorphin-6) and non-natural compounds (C-terminal amides derivatives of rubiscolin-6 and soymorphin-6; six analogues of endomorphin-2 and DAPEA; two compounds obtained in silico studies; a series of lonidamine with high CB1 receptor affinity and new pH-sensitive formulation of niosomes containing Polysorbate 20 derivatized by Glycine and loaded with ibuprofen) using murine models of pain and inflammation. All experiments were carried out using male CD-1 mice weighing 25 g. Results: The results suggest the potential use of ammonium glycyrrhizinate for clinical treatment of pain and/or inflammatory-related diseases, included diabetic peripheral neuropathy. The results obtained by in vivo studies testing two natural compounds, rubiscolin-6 and soymorphin-6, and their C-terminal amides derivatives, show that rubiscolin-6 was able to increase the nociceptive threshold to thermal stimuli after supraspinal administration, but was ineffective after subcutaneous administration, in an experimental paradigm of chemical-induced nociception as the formalin test. On the other hand, instead, its derivatives (rubiscolin-6 C-amide) centrally administered demonstrates a strong antinociceptive effect higher than rubiscolin-6, and it is effective after subcutaneous administration. Both soymorphin-6 and its derivative soymorphin-6 C-amide induced a robust antinociceptive effect after central administration in tail flick test but both peptides have not been able to change the behavioral response to chemical-induced nociception in the formalin test. The in vivo antinociceptive profile of new six tetrapeptide models containing α-amino-γ-lactam of Freidinger in position 2 and 3, as analogues of endomorphin-2 (EM-2) and DAPEA, revealed that one of them (peptide A2D) exhibited a strong antinociceptive effect in vivo after intracerebroventricular administration, performing better than the parent compounds EM-2 and DAPEA. Peptide A3D is also able to produce antinociceptive effect both in the acute and in the inflammatory phase of the formalin test. The in vivo results obtained from two compounds (tripeptides 6 and 11) derived from previous in silico studies revealed their ability to induce an antinociceptive effect after intracerebroventricular and subcutaneous administrations in the tail flick and formalin tests, respectively. Subsequently, it was analyzed the in vivo orexant/anorexant and antinociceptive profile of a series of lonidamine joined Leu-, tert/Leu- and Val-amino acids with different C-terminal functional groups (LONI 1-4,11) as novel compounds with high CB1 receptor affinity and selectivity with different biological activity depending on the C-terminal substitution and amino acid residues and endowed with activities. The results demonstrated that LONI2 and LONI4 were able to inhibit food intake (anorexant effect) consistent with an inverse agonism at CB1 receptors. On the other hand, LONI11, an agonist towards cannabinoid receptors CB1 and CB2, proved to carry out a significant orexant an antinociceptive effect at the central and periphery levels. Finally, a new pH-sensitive formulation of niosomes containing Polysorbate 20 derivatized by Glycine and loaded with ibuprofen (NioIbu) was in vivo tested, using models of pain and inflammation. The results demonstrated that NioIbu, administered 2h before testing, reduced nociception, whereas the free form of ibuprofen was ineffective. Conclusion: In conclusion, these findings create prospects for conducting clinical studies with the compounds presented, which show in fact their effectiveness for chronic pain and inflammation control

Flavonoid-statins interactions

Background: Statins and dietary modifications are the cornerstone of hypercholesterolemia management. Even if flavonoids could play a role in the prevention of hypercholesterolemia, the question whether there is a helpful or dangerous association between flavonoid-rich foods and statins has remained unanswered for a long time1. Therefore, although it is known that 58% of the more serious side effects of statins is related to interactions with other drugs and occurs especially when using statins metabolized at the level of cytochrome P450 (CYP)2, the aim of our work is to analyse the potential interaction between flavonoid-rich foods and statins in terms of musculoskeletal symptoms, on the basis of the type of statin metabolism. Methods: Subjects that use statins metabolised by CYP3A4 (N=20), CYP2C9 (N=20), statins minimally or not metabolised by CYP (N=20), and healthy control subjects (N=20) have been recruited. Data have been collected through questionnaires and a 7-day food diary, in order to evaluate the characteristics (pathologies, body mass index, etc.), the eating habits of the subjects, the adherence to the Mediterranean diet and the presence of musculoskeletal disorders. Results: The Mediterranean diet-statins interaction recently documented3 could be particularly accentuated for statins metabolised by CYP. Conclusions: eating habits should be considered not only during drug therapy but also during the use of nutraceuticals. [1] ISSN: 1875-5453. 2015. 16(9):833-46. [2] ISSN: 1972-6481. 2016. 17(6):447-55. [3] ISSN: 1874-1754. 2019. 276:248-25

Ammonium Glycyrrhizinate Prevents Apoptosis and Mitochondrial Dysfunction Induced by High Glucose in SH-SY5Y Cell Line and Counteracts Neuropathic Pain in Streptozotocin-Induced Diabetic Mice

Glycyrrhiza glabra, commonly known as liquorice, contains several bioactive compounds such as flavonoids, sterols, triterpene, and saponins; among which, glycyrrhizic acid, an oleanane-type saponin, is the most abundant component in liquorice root. Diabetic peripheral neuropathy is one of the major complications of diabetes mellitus, leading to painful condition as neuropathic pain. The pathogenetic mechanism of diabetic peripheral neuropathy is very complex, and its understanding could lead to a more suitable therapeutic strategy. In this work, we analyzed the effects of ammonium glycyrrhizinate, a derivate salt of glycyrrhizic acid, on an in vitro system, neuroblastoma cells line SH-SY5Y, and we observed that ammonium glycyrrhizinate was able to prevent cytotoxic effect and mitochondrial fragmentation after high-glucose administration. In an in vivo experiment, we found that a short-repeated treatment with ammonium glycyrrhizinate was able to attenuate neuropathic hyperalgesia in streptozotocin-induced diabetic mice. In conclusion, our results showed that ammonium glycyrrhizinate could ameliorate diabetic peripheral neuropathy, counteracting both in vitro and in vivo effects induced by high glucose, and might represent a complementary medicine for the clinical management of diabetic peripheral neuropathy