The Impact of UFP-512 in Mice with Osteoarthritis Pain: The Role of Hydrogen Sulfide.

The pain-relieving properties of opioids in inflammatory and neuropathic pain are heightened by hydrogen sulfide (H2S). However, whether allodynia and functional and/or emotional impairments related to osteoarthritis (OA) could be reduced by activating δ-opioid receptors (DOR) and the plausible influence of H2S on these actions has not been completely established. In female C57BL/6J mice with OA pain generated via monosodium acetate (MIA), we analyze: (i) the effects of UFP-512 (a DOR agonist), given alone and co-administered with two H2S donors, on the symptoms of allodynia, loss of grip strength (GS), and anxiodepressive-like comportment; (ii) the reversion of UFP-512 actions with naltrindole (a DOR antagonist), and (iii) the impact of UFP-512 on the expression of phosphorylated NF-kB inhibitor alpha (p-IKBα) and the antioxidant enzymes superoxide dismutase 1 (SOD-1) and glutathione sulfur transferase M1 (GSTM1); and the effects of H2S on DOR levels in the dorsal root ganglia (DRG), amygdala (AMG), and hippocampus (HIP) of MIA-injected animals. Results showed that systemic and local administration of UFP-512 dose-dependently diminished the allodynia and loss of GS caused by MIA, whose effects were potentiated by H2S and reversed by naltrindole. UFP-512 also inhibited anxiodepressive-like behaviors, normalized the overexpression of p-IKBα in DRG and HIP, and enhanced the expression of SOD-1 and GSTM1 in DRG, HIP, and/or AMG. Moreover, the increased expression of DOR triggered by H2S might support the improved analgesic actions of UFP-512 co-administered with H2S donors. This study proposes the use of DOR agonists, alone or combined with H2S donors, as a new treatment for OA pain.

Prokineticin System Is a Pharmacological Target to Counteract Pain and Its Comorbid Mood Alterations in an Osteoarthritis Murine Model.

Osteoarthritis (OA) is the most prevalent joint disease associated with chronic pain. OA pain is often accompanied by mood disorders. We addressed the role of the Prokineticin (PK) system in pain and mood alterations in a mice OA model induced with monosodium iodoacetate (MIA). The effect of a PK antagonist (PC1) was compared to that of diclofenac. C57BL/6J male mice injected with MIA in the knee joint were characterized by allodynia, motor deficits, and fatigue. Twenty-eight days after MIA, in the knee joint, we measured high mRNA of PK2 and its receptor PKR1, pro-inflammatory cytokines, and MMP13. At the same time, in the sciatic nerve and spinal cord, we found increased levels of PK2, PKR1, IL-1ß, and IL-6. These changes were in the presence of high GFAP and CD11b mRNA in the sciatic nerve and GFAP in the spinal cord. OA mice were also characterized by anxiety, depression, and neuroinflammation in the prefrontal cortex and hippocampus. In both stations, we found increased pro-inflammatory cytokines. In addition, PK upregulation and reactive astrogliosis in the hippocampus and microglia reactivity in the prefrontal cortex were detected. PC1 reduced joint inflammation and neuroinflammation in PNS and CNS and counteracted OA pain and emotional disturbances.

Bv8 mediates myeloid cell migration and enhances malignancy of colorectal cancer.

Colorectal cancer (CRC) is the third most predominant malignancy in the world. Although the importance of immune system in cancer development has been well established, the underlying mechanisms remain to be investigated further. Here we studied a novel protein prokineticin 2 (Prok2, also known as Bv8) as a key pro-tumoral factor in CRC progression in in vitro and ex vivo settings. Human colorectal tumor tissues, myeloid cell lines (U937 cells and HL60 cells) and colorectal cancer cell line (Caco-2 cells) were used for various studies. Myeloid cell infiltration (especially neutrophils) and Bv8 accumulation were detected in human colorectal tumor tissue with immunostaining. The chemotactic effects of Bv8 on myeloid cells were presented in the transwell assay and chemotaxis assy. Cultured CRC cells treated with myeloid cells or Bv8 produced reactive oxygen species (ROS) and vascular endothelial growth factor (VEGF). Furthermore, ROS and VEGF acted as pro-angiogenesis buffer in myeloid cell-infiltrated CRC microenvironment. Moreover, myeloid cells or Bv8 enhanced energy consumption of glycolysis ATP and mitochondria ATP of CRC cells. Interestingly, myeloid cells increased CRC cell viability, but CRC cells decreased the viability of myeloid cells. ERK signalling pathway in CRC cells was activated in the presence of Bv8 or co-cultured myeloid cells. In conclusion, our data indicated the vital roles of Bv8 in myeloid cell infiltration and CRC development, suggesting that Bv8 may be a potential therapeutic target for colorectal cancer-related immunotherapy.

Prokineticin 1 is a novel factor regulating porcine corpus luteum function.

Prokineticin 1 (PROK1) is a pleiotropic factor secreted by endocrine glands; however, its role has not been studied in the corpus luteum (CL) during pregnancy in any species. The present study aimed to investigate the contribution of PROK1 in regulating processes related to porcine CL function and regression: steroidogenesis, luteal cell apoptosis and viability, and angiogenesis. The luteal expression of PROK1 was greater on Days 12 and 14 of pregnancy compared to Day 9. PROK1 protein expression during pregnancy increased gradually and peaked on Day 14, when it was also significantly higher than that on Day 14 of the estrous cycle. Prokineticin receptor 1 (PROKR1) mRNA abundance increased on Days 12 and 14 of pregnancy, whereas PROKR2 elevated on Day 14 of the estrous cycle. PROK1, acting via PROKR1, stimulated the expression of genes involved in progesterone synthesis, as well as progesterone secretion by luteal tissue. PROK1-PROKR1 signaling reduced apoptosis and increased the viability of luteal cells. PROK1 acting through PROKR1 stimulated angiogenesis by increasing capillary-like structure formation by luteal endothelial cells and elevating angiogenin gene expression and VEGFA secretion by luteal tissue. Our results indicate that PROK1 regulates processes vital for maintaining luteal function during early pregnancy and the mid-luteal phase.

Blocking prokineticin receptors attenuates synovitis and joint destruction in collagen-induced arthritis.

Rheumatoid arthritis (RA) is a chronic inflammatory disease mediated by an interdependent network of proinflammatory molecules such as chemokines. Prokineticin 2 (PK2) is a chemokine-like peptide that modulates nociceptive threshold and immuno-inflammatory processes via two G-protein-linked receptors, prokineticin receptor 1 and 2 (PKR1 and PKR2). In the present study, we investigated the effects of the prokineticin receptor antagonist PC1 on arthritic pain and the inflammatory response in type II collagen-induced arthritis (CIA) in mice. We demonstrated that PC1, administered subcutaneously from day 25 to day 35 after CIA, improved clinical signs of arthritis such as paw edema, pain, and impaired locomotor activity. In CIA mice, PC1 was also able to lower plasma malondialdehyde (MDA) levels, suggesting a role in reducing oxidative damage, as well as joint expression levels of PK2, PKRs, TNFα, IL-1ß, CD4, CD8, and NF-kB. These results suggest that blocking PKRs may be a successful strategy to control arthritic pain and pathology development. KEY MESSAGES: PK2/PKRs expression levels strongly increase in the synovium of RA mice. PC1 treatment shows anti-arthritic activity and reduces arthritis-induced pain. PC1 treatment significantly lowers synovial PK2/PKRs levels. PC1 treatment lowers plasma MDA levels and synovial levels of TNFα and IL -1ß PC1 treatment is a viable therapeutic option for RA.

Synthesis of Sulfonamides Incorporating Piperidinyl-Hydrazidoureido and Piperidinyl-Hydrazidothioureido Moieties and Their Carbonic Anhydrase I, II, IX and XII Inhibitory Activity.

Here we report a small library of hydrazinocarbonyl-ureido and thioureido benzenesulfonamide derivatives, designed and synthesized as potent and selective human carbonic anhydrase inhibitors (hCAIs). The synthesized compounds were evaluated against isoforms hCA I, II, IX and XII using acetazolamide (AAZ) as standard inhibitor. Several urea and thiourea derivatives showed inhibitory activity at low nanomolar levels with selectivity against the cytosolic hCA II isoform, as well as the transmembrane, tumor-associated enzymes hCA IX and XII. The thiourea derivatives showed enhanced potency as compared to urea analogues. Additionally, eight compounds 5g, 5m, 5o, 5q, 6l, 6j, 6o and 6u were selected for docking analysis on isoform I, II, IX, XII to illustrate the potential interaction with the enzyme to better understand the activity against the different isoforms.

Hydrogen Sulfide Increases the Analgesic Effects of µ- and δ-Opioid Receptors during Neuropathic Pain: Pathways Implicated.

Recent studies have revealed that hydrogen sulfide (H2S) increases the analgesic actions of the δ-opioid receptor (DOR) in inflammatory pain. However, the possible improvement of the analgesia of µ-opioid receptor (MOR) and DOR agonists during neuropathic pain, through pretreatment with two slow-releasing H2S donors-DADS (diallyl disulfide) and GYY4137 (morpholin-4-ium 4-methoxyphenyl(morpholino) phosphinodithioate dichloromethane complex)-is still unknown. In male C57BL/6J mice with neuropathic pain incited by chronic constriction of the sciatic nerve (CCI), we evaluated: (1) the influence of DADS (3.5 mg/kg) and GYY4137 (0.7 mg/kg) on the inhibition of the allodynia and hyperalgesia produced by the systemic or local administration of morphine (3 mg/kg or 65 µg) and UFP-512 (1 mg/kg or 12.5 µg); (2) the reversion of the antinociceptive actions of high doses of DADS (30 mg/kg) and GYY4137 (24 mg/kg) with MOR and DOR antagonists; and (3) the effects of H2S donors on oxidative stress, apoptotic responses, and MOR and DOR expression in the medial septum (MS) and dorsal root ganglia (DRG). The results revealed that both DADS and GYY4137 improved the antiallodynic effects of morphine and UFP-512, possibly by up-regulating MOR and DOR expression in DRG. The administration of MOR and DOR antagonists blocked the analgesic properties of DADS and GYY4137, revealing the feasible participation of the endogenous opioid system in H2S analgesic effects. Moreover, both H2S donors inhibited oxidative stress and apoptosis generated by CCI in the MS and/or DRG. This study suggests the co-treatment of H2S donors with MOR or DOR agonists as a potential therapy for neuropathic pain.

A Critical Role of δ-Opioid Receptor in Anti-microglial Activation Under Stress.

Microglia are involved in the regulation of cerebral homeostasis and pathogen confrontation. There is, however, evidence showing that excessive microglia activation is implicated in various age-related cerebral diseases. On the other hand, microglia may experience complex changes of polarization in pathological insults, i.e., from a proinflammatory M1 to an anti-inflammatory M2 phenotype, which differentially contribute to the exacerbation or alleviation of cellular injury. Remolding the phenotype of microglia or inhibiting the excessive activation of microglia seems to be a promising approach against neurodegenerative pathologies. Since δ-opioid receptor (DOR) activation exhibits a strong protective capacity against various neuronal injuries, especially the hypoxic/ischemic injury, we asked if the DOR-induced neuroprotection is associated with its effect on microglia. We explored this fundamental issue by using pharmacological and genetic approaches in the BV2 cell line, a general type of microglial cells. The results showed that DOR expression significantly increased in the activated microglial M2 phenotype, but slightly decreased in the microglial M1 phenotype. Hypoxia induced dual polarizations of BV2 cells with an increase in DOR expression. Administration of a specific DOR agonist, UFP-512, largely inhibited lipopolysaccharide (LPS) or hypoxia-induced microglial M1 activation and inflammatory activity with high concentrations of UFP-512 being effective to reverse the interleukin-4 (IL4)-induced microglial activation. Consistent with these observations, inhibiting DOR or knocking-down DOR promoted the excessive activation of BV2 cells in both M1 and M2 directions, while DOR overexpression did the opposite. Furthermore, the PC12 cells exposed to the conditioned medium of BV2 cells treated by UFP-512 grew better than those treated directly with UFP-512 under LPS or hypoxic insults. DOR inhibitor naltrindole could block all the effects of DOR activation. The medium from the BV2 cells with DOR knock-down decreased the viability of PC12 cell, while the medium from the BV2 cells with DOR overexpression largely attenuated LPS or hypoxic injury in the PC12 cells. These first data suggest a close linkage between DOR expression/function and microglial polarization and a critical role of DOR in negative controlling microglial activation. Our work provides a novel clue for new protective strategies against neurodegenerative pathophysiology through DOR-mediated regulation of microglia.

Investigation on Hydrazonobenzenesulfonamides as Human Carbonic Anhydrase I, II, IX and XII Inhibitors.

A small series of hydrazonobenzenesulfonamides was designed, synthesized and studied for their human carbonic anhydrase (hCA) inhibitory activity. The synthesized compounds were evaluated against hCA I, II, IX and XII isoforms using acetazolamide (AAZ) as the standard inhibitor. Various hydrazonosulfonamide derivatives showed inhibitory activity at low nanomolar levels with selectivity against the cytosolic hCA II isoform, as well as the transmembrane, tumor-associated enzymes hCA IX and XII. The most potent and selective hydrazones 8, 9, 10, 11, 19 and 24 were docked into isoforms I, II, IX and XII to better understand their activity and selectivity for the different CA isoforms.

Hydrogen Sulfide Inhibits Inflammatory Pain and Enhances the Analgesic Properties of Delta Opioid Receptors.

Chronic inflammatory pain is present in many pathologies and diminishes the patient's quality of life. Moreover, most current treatments have a low efficacy and significant side effects. Recent studies demonstrate the analgesic properties of slow-releasing hydrogen sulfide (H2S) donors in animals with osteoarthritis or neuropathic pain, but their effects in inflammatory pain and related pathways are not completely understood. Several treatments potentiate the analgesic actions of δ-opioid receptor (DOR) agonists, but the role of H2S in modulating their effects and expression during inflammatory pain remains untested. In C57BL/6J male mice with inflammatory pain provoked by subplantar injection of complete Freund's adjuvant, we evaluated: (1) the antiallodynic and antihyperalgesic effects of different doses of two slow-releasing H2S donors, i.e., diallyl disulfide (DADS) and phenyl isothiocyanate (P-ITC) and their mechanism of action; (2) the pain-relieving effects of DOR agonists co-administered with H2S donors; (3) the effects of DADS and P-ITC on the oxidative stress and molecular changes caused by peripheral inflammation. Results demonstrate that both H2S donors inhibited allodynia and hyperalgesia in a dose-dependent manner, potentiated the analgesic effects and expression of DOR, activated the antioxidant system, and reduced the nociceptive and apoptotic pathways. The data further demonstrate the possible participation of potassium channels and the Nrf2 transcription factor signaling pathway in the pain-relieving activities of DADS and P-ITC. This study suggests that the systemic administration of DADS and P-ITC and local application of DOR agonists in combination with slow-releasing H2S donors are two new strategies for the treatment of inflammatory pain.

The Antagonism of the Prokineticin System Counteracts Bortezomib Induced Side Effects: Focus on Mood Alterations.

The development of neuropathy and of mood alterations is frequent after chemotherapy. These complications, independent from the antitumoral mechanism, are interconnected due to an overlapping in their processing pathways and a common neuroinflammatory condition. This study aims to verify whether in mice the treatment with the proteasome inhibitor bortezomib (BTZ), at a protocol capable of inducing painful neuropathy, is associated with anxiety, depression and supraspinal neuroinflammation. We also verify if the therapeutic treatment with the antagonist of the prokineticin (PK) system PC1, which is known to contrast pain and neuroinflammation, can prevent mood alterations. Mice were treated with BTZ (0.4 mg/kg three times/week for 4 weeks); mechanical allodynia and locomotor activity were evaluated over time while anxiety (dark light and marble burying test), depression (sucrose preference and swimming test) and supraspinal neuroinflammation were checked at the end of the protocol. BTZ treated neuropathic mice develop anxiety and depression. The presence of mood alterations is related to the presence of neuroinflammation and PK system activation in prefrontal cortex, hippocampus and hypothalamus with high levels of PK2 and PKR2 receptor, IL-6 and TNF-α, TLR4 and an upregulation of glial markers. PC1 treatment, counteracting pain, prevented the development of supraspinal inflammation and depression-like behavior in BTZ mice.

Differential inflammation-mediated function of prokineticin 2 in the synovial fibroblasts of patients with rheumatoid arthritis compared with osteoarthritis.

Prokineticin 2 (PK2) is a secreted protein involved in several pathological and physiological processes, including the regulation of inflammation, sickness behaviors, and circadian rhythms. Recently, it was reported that PK2 is associated with the pathogenesis of collagen-induced arthritis in mice. However, the role of PK2 in the pathogenesis of rheumatoid arthritis (RA) or osteoarthritis (OA) remains unknown. In this study, we collected synovial tissue, plasma, synovial fluid, and synovial fibroblasts (SF) from RA and OA patients to analyze the function of PK2 using immunohistochemistry, enzyme-linked immunosorbent assays, and tissue superfusion studies. PK2 and its receptors prokineticin receptor (PKR) 1 and 2 were expressed in RA and OA synovial tissues. PKR1 expression was downregulated in RA synovial tissue compared with OA synovial tissue. The PK2 concentration was higher in RA synovial fluid than in OA synovial fluid but similar between RA and OA plasma. PK2 suppressed the production of IL-6 from TNFα-prestimulated OA-SF, and this effect was attenuated in TNFα-prestimulated RA-SF. This phenomenon was accompanied by the upregulation of PKR1 in OA-SF. This study provides a new model to explain some aspects underlying the chronicity of inflammation in RA.

Hair Growth Promotion by δ-Opioid Receptor Activation.

Literature has revealed that the delta opioid receptor (DOR) exhibited diverse pharmacological effects on neuron and skin. In the present study, we have investigated whether the activation of DOR has hair-growth promotion effects. Compared with other opioid receptor, DOR was highly expressed in epidermal component of hair follicle in human and rodents. The expression of DOR was high in the anagen phase, but it was low in the catagen and telogen phases during mouse hair cycle. Topical application of UFP-512, a specific DOR agonist, significantly accelerated the induction of the anagen in C3H mice. Topical application of UFP-512 also increased the hair length in hair organ cultures and promoted the proliferation and the migration of outer root sheath (ORS) cells. Similarly, pharmacological inhibition of DOR by naltrindole significantly inhibited the anagen transition process and decreased hair length in hair organ cultures. Thus, we further examined whether Wnt/ß-catenin pathway was related to the effects of DOR on hair growth. We found that Wnt/ß-catenin pathway was activated by UFP-512 and siRNA for ß-catenin attenuated the UFP-512 induced proliferation and migration of ORS cells. Collectively, result established that DOR was involved in hair cycle regulation, and that DOR agonists such as UFP-512 should be developed for novel hair-loss treatment.

Evidence-Based View of Safety and Effectiveness of Prokineticin Receptors Antagonists during Pregnancy.

Endocrine gland derived vascular endothelial growth factor (EG-VEGF) is a canonical member of the prokineticin (PROKs) family. It acts via the two G-protein coupled receptors, namely PROKR1 and PROKR2. We have recently demonstrated that EG-VEGF is highly expressed in the human placenta; contributes to placental vascularization and growth and that its aberrant expression is associated with pregnancy pathologies including preeclampsia and fetal growth restriction. These findings strongly suggested that antagonization of its receptors may constitute a potential therapy for the pregnancy pathologies. Two specific antagonists of PROKR1 (PC7) and for PROKR2 (PKRA) were reported to reverse PROKs adverse effects in other systems. In the view of using these antagonists to treat pregnancy pathologies, a proof of concept study was designed to determine the biological significances of PC7 and PKRA in normal pregnancy outcome. PC7 and PKRA were tested independently or in combination in trophoblast cells and during early gestation in the gravid mouse. Both independent and combined treatments uncovered endogenous functions of EG-VEGF. The independent use of antagonists distinctively identified PROKR1 and PROKR2-mediated EG-VEGF signaling on trophoblast differentiation and invasion; thereby enhancing feto-placental growth and pregnancy outcome. Thus, our study provides evidence for the potential safe use of PC7 or PKRA to improve pregnancy outcome.

δ-opioid receptor activation protects against Parkinson's disease-related mitochondrial dysfunction by enhancing PINK1/Parkin-dependent mitophagy.

Our previous studies have shown that the δ-opioid receptor (DOR) is an important neuroprotector via the regulation of PTEN-induced kinase 1 (PINK1), a mitochondria-related molecule, under hypoxic and MPP+ insults. Since mitochondrial dysfunctions are observed in both hypoxia and MPP+ insults, this study further investigated whether DOR is cytoprotective against these insults by targeting mitochondria. Through comparing DOR-induced responses to hypoxia versus MPP+-induced parkinsonian insult in PC12 cells, we found that both hypoxia and MPP+ caused a collapse of mitochondrial membrane potential and severe mitochondrial dysfunction. In sharp contrast to its inappreciable effect on mitochondria in hypoxic conditions, DOR activation with UFP-512, a specific agonist, significantly attenuated the MPP+-induced mitochondrial injury. Mechanistically, DOR activation effectively upregulated PINK1 expression and promoted Parkin's mitochondrial translocation and modification, thus enhancing the PINK1-Parkin mediated mitophagy. Either PINK1 knockdown or DOR knockdown largely interfered with the DOR-mediated mitoprotection in MPP+ conditions. Moreover, there was a major difference between hypoxia versus MPP+ in terms of the regulation of mitophagy with hypoxia-induced mitophagy being independent from DOR-PINK1 signaling. Taken together, our novel data suggest that DOR activation is neuroprotective against parkinsonian injury by specifically promoting mitophagy in a PINK1-dependent pathway and thus attenuating mitochondrial damage.

Prokineticin 1-prokineticin receptor 1 signaling promotes angiogenesis in the porcine endometrium during pregnancy†.

Pregnancy establishment in mammals, including pigs, requires proper communication between embryos and the maternal reproductive tract. Prokineticin 1 (PROK1) has been described as a secretory protein with pleiotropic functions and as a novel tissue-specific angiogenic factor. However, despite the studies performed mainly on human cell lines and in mice, the function of PROK1 in the endometrium during early pregnancy is still not fully elucidated. We hypothesized that PROK1 contributes to pregnancy establishment in pigs. The present study is the first to report that the expression of PROK1 and its receptor (PROKR1) is elevated in the porcine endometrium during the implantation and early placentation period. PROK1 protein was detected mainly in luminal epithelial cells, glandular epithelial cells, and blood vessels in the endometrium. Using the porcine in vivo model of unilateral pregnancy, we revealed that conceptuses induced the endometrial expression of PROK1 and PROKR1. Moreover, the embryonic signal, estradiol-17ß, as well as progesterone, stimulated the endometrial expression of PROK1 and PROKR1. We also evidenced that PROK1-PROKR1 signaling supports endometrial angiogenesis in pigs. The PROK1-stimulated proliferation of primary porcine endometrial endothelial (PEE) cells involved PI3K/AKT/mTOR, MAPK, cAMP, and NFKB signaling pathways. Furthermore, PROK1 via PROKR1 promoted the formation of capillary-like structures by PEE cells. PROK1 also stimulated VEGFA and PGF2α secretion, which in turn may indirectly support angiogenic changes within endometrial tissue. In summary, our study suggests that PROK1 acts as an embryonic signal mediator that regulates endometrial angiogenesis and secretory function during the implantation and early placentation period in pigs.

Opposite Roles of δ- and µ-Opioid Receptors in BACE1 Regulation and Alzheimer's Injury.

Alzheimer's disease (AD) is characterized by amyloid plaques and neurofibrillary tangles. Substantial evidence for AD pathogenesis suggests that ß-site APP cleaving enzyme 1 (BACE1) and γ-secretase enzyme initiate the amyloidogenic pathway and produces toxic Aß peptides that prone to aggregate in the brain. Therefore, the inhibition of BACE1 expression and function is an attractive strategy for AD therapy. In the present work, we made the first finding that activating δ-opioid receptors (DOR) with a specific DOR agonist significantly attenuated BACE1 expression and activity in the highly differentiated PC12 cells with mimicked AD injury, while the application of DOR inhibitor naltrindole reversed the UFP-512 effects, and even caused a major increase in BACE1 expression and activity as well as Aß42 production in physiological conditions. Knocking-down DOR also enhanced BACE1 protein expression and its activity for APP processing, associating with a significant increase in Aß42 production. In sharp contrast, activation of µ-opioid receptor (MOR) with DAMGO greatly promoted BACE1 expression and activity with an acceleration of APP cleavage, thus contributing to increased Aß42 production. DADLE, a less selective DOR agonist that may bind to MOR, had no stable inhibitory effect on BACE1. Similar results were also found in APP mutant (APPswe) SH-SY5Y cell line, providing further validation of the DOR action on BACE1 regulation. Our novel data demonstrated entirely different roles of DOR and MOR in the regulation of BACE1 expression and activity with DOR being neuroprotective against AD injury. These findings provided a novel clue for new strategies of AD therapy via targeting endogenous opioid receptors.

In-Vitro Evaluation of Antioxidant, Antiproliferative and Photo-Protective Activities of Benzimidazolehydrazone Derivatives.

In the search of multifunctional compounds we designed benzimidazole derivatives endowed with phenolic hydroxy groups and a hydrazone moiety as potential radical-scavenger and the antioxidant agents. The target molecules have been prepared by a simple synthetic procedure and tested for their antioxidant activity by DPPH, FRAP, and ORAC test, for photoprotective activity against UV rays and for antiproliferative activity against Colo-38 melanoma cells. Furthermore, two different dermocosmetic formulations were prepared with the compounds endowed with the best antioxidant and photoprotective profile and their release from formulation evaluated using Franz Cells system. High antioxidant activity is related to the presence of at least two hydroxy groups on arylidene moiety of benzimidazoles. Structure activity analysis revealed that the position of hydroxy groups is crucial for antioxidant activity as well as the presence of a 2-hydroxy-4-(diethylamino)arylidene group. The same correlation pattern was found to be related to photoprotective activity resulting in an UVA Protection Factor better than the commercial solar filter PBSA and antiproliferative activity against melanoma cells without producing cytotoxicity on normal keratinocytes. The release analysis indicated that high antioxidant activities are achieved with limited release at concentration compatible with the use as UV sunscreen filter.

Appliance of the piperidinyl-hydrazidoureido linker to benzenesulfonamide compounds: Synthesis, in vitro and in silico evaluation of potent carbonic anhydrase II, IX and XII inhibitors.

Herein we report on a new series of hydrazidoureidobenzensulfonamides investigated as inhibitors of the cytosolic human (h) hCA I and II isoforms, as well as the transmembrane, tumor-associated enzymes hCA IX and XII. The reported derivatives contain a 4-substituted piperidine fragment in which the hydrazidoureido linker has been involved as spacer between the benzenesulfonamide fragment which binds the zinc ion from the active site, and the tail of the inhibitor. Depending on the substitution pattern at the piperidine ring, low nanomolar inhibitors were detected against hCA II, hCA IX and hCA XII, making the new class of sulfonamides of interest for various pharmacologic applications.

Synthesis and evaluation of antioxidant and antiproliferative activity of 2-arylbenzimidazoles.

Three series of arylbenzimidazole derivatives 3-40, 45 have been simply synthesized and tested for their antioxidant capacity. The 2-arylbenzimidazoles were tested against various radicals by the DPPH, FRAP and ORAC tests and showed different activity profiles. It has been observed that the number and position of the hydroxy groups on the 2-aryl portion and the presence of a diethylamino group or a 2-styryl group are related to a good antioxidant capacity. Furthermore, benzimidazoles showed satisfactory SPF values ​​in vitro compared to the commercial PBSA filter, proving to have a good photoprotective profile. In particular, 2-arylbenzimidazole-5-sulphonic acids 15 and 38, the 2-styryl-benzimidazole 45 showed broad spectrum solar protection against UVA and UVB rays. The antiproliferative effect of the benzimidazoles was tested on human skin melanoma Colo-38 cells. The styrylbenzimidazole 45 exhibited antiproliferative effect at low micromolar concentration against Colo-38 cells and very low antiproliferative activity on normal HaCat keratinocyte cells.