Pharmacokinetics and Safety of Rifamycin SV after Single and Multiple Doses of MMX® Modified Release Tablets in Healthy Male and Female Volunteers.

The primary objective of this single- and multiple-dose pharmacokinetic study was the investigation of rifamycin SV's pharmacokinetic profile in plasma and urine. All the 18 enrolled healthy men and post-menopausal women received modified release tablets containing 600 mg of the oral non-absorbable antibiotic, rifamycin SV, according to a multiple dose regimen: one tablet three times a day (daily intake: 1800 mg) for 14 consecutive days. Blood sampling and urine collection were performed up to 24 h post-dose after the first dose on Days 1 and 7. On average, on Day 1, Cmax,0-24 was 5.79 ± 4.24 ng/mL and was attained in a median time of 9 h. On Day 7, all the subjects had quantifiable levels of rifamycin SV in plasma at each sampling time. After a peak concentration attained 2 h post-dose (mean ± SD concentration: 10.94 ± 16.41 ng/mL), rifamycin SV decreased in plasma to levels similar to those of Day 1. The amounts of rifamycin SV excreted in urine paralleled the plasma concentration at the corresponding times. On Day 1, the total amount excreted in urine was 0.0013%, and was 0.0029% on Day 7. The study results confirmed those of the previous Phase I study: the systemic absorption of rifamycin SV was also proved negligible after 7 days of the 600 mg t.i.d. dose regimen of the newly formulated tablets, currently under development for the treatment of several small and large intestinal pathologies, including diarrhea-predominant irritable bowel syndrome, hepatic encephalopathy, and others. Registered at ClinicalTrials.gov with the identifier NCT02969252, last updated on 26JAN18.

An exploratory microdialysis study investigating the effect of repeated application of a diclofenac epolamine medicated plaster on prostaglandin concentrations in skeletal muscle after standardized physical exercise.

AIM: Muscle injuries and extensive exercise are associated with cyclo-oxygenase dependent formation of inflammatory prostaglandins. Since the effect of topical administration of non-steroidal anti-inflammatory drugs (NSAIDs) on local cyclo-oxygenase is unknown, the present exploratory, open label, non-randomized study set out to measure exercise induced release of prostaglandins before and after epicutaneous administration of diclofenac. METHODS: Microdialysis was used to determine the local interstitial concentration of PGE2 and 8-iso-PGF2α as well as diclofenac concentrations in the vastus lateralis under rest, dynamic exercise and during recovery in 12 healthy subjects at baseline and after a treatment phase applying a total of seven plasters medicated with 180 mg of diclofenac epolamine over 4 days. RESULTS: At baseline PGE2 concentrations were 1169 ± 780 pg ml(-1) at rest and 1287 ± 459 pg ml(-1) during dynamic exercise and increased to 2005 ± 1126 pg ml(-1) during recovery. After treatment average PGE2 concentrations were 997 ± 588 pg ml(-1) at rest and 1339 ± 892 pg ml(-1) during exercise. In contrast with the baseline phase no increase in PGE2 concentrations was recorded during the recovery period after treatment (PGE2 1134 ± 874 pg ml(-1)). 8-iso-PGF2α was neither affected by exercise nor by treatment with diclofenac. Local and systemic concentrations of diclofenac were highly variable but comparable with previous clinical pharmacokinetic studies. CONCLUSIONS: We can hypothesize an effect of topical diclofenac epolamine plaster on limiting the increase of local concentrations of the pro-inflammatory prostaglandin PGE2 induced in the muscle of healthy human subjects following standardized physical exercise. No effect of diclofenac treatment on 8-iso-PGF2α concentrations was observed, mainly since isoprostane is produced by a free radical-catalyzed lipid peroxidation mechanism independent of cyclo-oxygenases.

Dendritic localization of the translational repressor Pumilio 2 and its contribution to dendritic stress granules.

Pumilio (Pum) protein acts as a translational inhibitor in several organisms including yeast, Drosophila, Xenopus, and mammals. Two Pumilio genes, Pum1 and Pum2, have been identified in mammals, but their function in neurons has not been identified. In this study, we found that Pum2 mRNA is expressed during neuronal development and that the protein is found in discrete particles in both the cell body and the dendritic compartment of fully polarized neurons. This finding indicates that Pum2 is a novel candidate of dendritically localized ribonucleoparticles (RNPs). During metabolic stress, Pum2 is present in stress granules (SGs), which are subsequently detected in the somatodendritic domain. It remains excluded from processing bodies under all conditions. When overexpressed in neurons and fibroblasts, Pum2 induces the formation of SGs that also contain T-cell intracellular antigen 1 (TIA-1)-related protein, eukaryotic initiation factor 4E, poly(A)-binding protein, TIA-1, and other RNA-binding proteins including Staufen1 and Barentsz. This induction of SGs is dependent on the RNA-binding domain and a glutamine-rich region in the N terminus of Pum2. This glutamine-rich region behaves in a similar manner as TIA-1 and prion protein, two molecules with known roles in protein aggregation. Pum2 downregulation in neurons via RNA interference (RNAi) interferes with the formation of SGs during metabolic stress. Cotransfection with an RNAi-resistant portion of the Pum2 mRNA restores SG formation. These results suggest a role for Pum2 in dendritic RNPs and SG formation in mammalian neurons.

Molecular mechanisms involved in the asymmetric interaction between cannabinoid and opioid systems.

The aim of this work was to study the mechanism of cross-modulation between cannabinoid and opioid systems for analgesia during acute and chronic exposure. Acute coadministration of ineffectual subanalgesic doses of the synthetic cannabinoid CP-55,940 (0.2 mg/kg i.p.) and morphine (2.5 mg/kg i.p.) resulted in significant antinociception. In chronic studies, a low dose of CP-55,940 (0.2 mg/kg, i.p.) that per se did not induce analgesia in naive animals produced a significant degree of antinociception in rats made tolerant to morphine, whereas in rats made tolerant to CP-55,940, morphine challenge did not produce any analgesic response. To identify the mechanism of these asymmetric interactions during chronic treatment, we investigated the functional activity of cannabinoid and mu opioid receptors and their effects on the cyclic AMP (cAMP) cascade. Autoradiographic-binding studies indicated a slight but significant reduction in cannabinoid receptor levels in the hippocampus and cerebellum of morphine-tolerant rats, whereas CP-55,940-stimulated [35S]GTPgammaS binding showed a significant decrease in receptor/G protein coupling in the limbic area. In CP-55,940 exposed rats, mu opioid receptor binding was significantly raised in the lateral thalamus and periaqueductal gray (PAG), with an increase in DAMGO-stimulated [35S]GTPgammaS binding in the nucleus accumbens. Finally, we tested the cAMP system's responsiveness to the cannabinoid and opioid in the striatum and dorsal mesencephalon. In vivo chronic morphine did not affect CP-55,940's ability to inhibit forskolin-stimulated cAMP production in vitro and actually induced sensitization in striatal membranes. In contrast, in vivo chronic CP-55,940 desensitized DAMGO's efficacy in inhibiting forskolin-stimulated cAMP production in vitro. The alterations to the cAMP system seem to mirror the behavioral responses, indicating that the two systems may interact at the postreceptor level. This might open up new therapeutic opportunities for relief of chronic pain through cannabinoid-opioid coadministration.

Cannabidiol inhibits human glioma cell migration through a cannabinoid receptor-independent mechanism.

We evaluated the ability of cannabidiol (CBD) to impair the migration of tumor cells stimulated by conditioned medium. CBD caused concentration-dependent inhibition of the migration of U87 glioma cells, quantified in a Boyden chamber. Since these cells express both cannabinoid CB1 and CB2 receptors in the membrane, we also evaluated their engagement in the antimigratory effect of CBD. The inhibition of cell was not antagonized either by the selective cannabinoid receptor antagonists SR141716 (CB1) and SR144528 (CB2) or by pretreatment with pertussis toxin, indicating no involvement of classical cannabinoid receptors and/or receptors coupled to Gi/o proteins. These results reinforce the evidence of antitumoral properties of CBD, demonstrating its ability to limit tumor invasion, although the mechanism of its pharmacological effects remains to be clarified.

Antitumor effects of cannabidiol, a nonpsychoactive cannabinoid, on human glioma cell lines.

Recently, cannabinoids (CBs) have been shown to possess antitumor properties. Because the psychoactivity of cannabinoid compounds limits their medicinal usage, we undertook the present study to evaluate the in vitro antiproliferative ability of cannabidiol (CBD), a nonpsychoactive cannabinoid compound, on U87 and U373 human glioma cell lines. The addition of CBD to the culture medium led to a dramatic drop of mitochondrial oxidative metabolism [3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H tetrazolium bromide test] and viability in glioma cells, in a concentration-dependent manner that was already evident 24 h after CBD exposure, with an apparent IC(50) of 25 microM. The antiproliferative effect of CBD was partially prevented by the CB2 receptor antagonist N-[(1S)-endo-1,3,3-trimethylbicyclo[2,2,1]heptan-2-yl]-5-(4-chloro-3-methylphenyl)-1-(4-methylbenzyl)-pyrazole-3-carboxamide (SR144528; SR2) and alpha-tocopherol. By contrast, the CB1 cannabinoid receptor antagonist N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboximide hydrochloride (SR141716; SR1), capsazepine (vanilloid receptor antagonist), the inhibitors of ceramide generation, or pertussis toxin did not counteract CBD effects. We also show, for the first time, that the antiproliferative effect of CBD was correlated to induction of apoptosis, as determined by cytofluorimetric analysis and single-strand DNA staining, which was not reverted by cannabinoid antagonists. Finally, CBD, administered s.c. to nude mice at the dose of 0.5 mg/mouse, significantly inhibited the growth of subcutaneously implanted U87 human glioma cells. In conclusion, the nonpsychoactive CBD was able to produce a significant antitumor activity both in vitro and in vivo, thus suggesting a possible application of CBD as an antineoplastic agent.

Cannabinoids and opioids share cAMP pathway in rat splenocytes.

In the present work we investigated on rat splenocytes long-term interactions between opioid and cannabinoid drugs in terms of a common regulation of cAMP intracellular pathway. Both morphine and the synthetic cannabinoid compound CP-55,940 inhibited in a concentration-dependent manner the intracellular cAMP level in splenocytes stimulated by forskolin. The in vitro combination of submaximal concentrations of the two drugs did not yield any additive effect on the inhibition induced by the two drugs. In splenocytes taken from rats chronically treated with CP-55,940 (0.2 mg/kg i.p., twice a day for 4.5 days) or morphine (5 mg/kg s.c., twice a day for 6.5 days) and in vitro exposed to either CP-55,940 or morphine, it was found a desensitisation and cross-desensitisation to the inhibitory effects on cAMP production induced by the two drugs. Binding experiments on the cannabinoid receptors level in spleen coronal sections after in vivo chronic administration of morphine, revealed that there was no changes in the binding of [H3]-CP-55,940. Thus, these results strengthen the hypothesis of cAMP as part of the common intracellular pathway shared by opiates and cannabinoids at immune cell level.

Chronic morphine modulates the contents of the endocannabinoid, 2-arachidonoyl glycerol, in rat brain.

Opioids and cannabinoids are among the most widely consumed drugs of abuse in humans and the phenomena of cross-tolerance or mutual potentiation have been demonstrated between the two drugs. Several authors have suggested that both drugs share common links in their molecular mechanisms of action, although this has been a matter of controversy. Furthermore, no data exist on the possible adaptive changes in the contents of arachidonoylethanolamide (anandamide, AEA) and 2-arachidonoylglycerol (2-AG), the two major endogenous ligands for cannabinoid receptors, in morphine-tolerant rats. In the present work, we investigated the alterations in cannabinoid receptor functionality and endocannabinoid levels in rats chronically treated with morphine (5 mg/kg, s.c., twice a day for 5 days). Autoradiographic-binding studies using [(3)H]CP-55 940 revealed a slight but significant reduction in cannabinoid receptor level in the cerebellum and hippocampus of morphine-tolerant rats, while CP-55 940-stimulated [(35)S]GTPgammaS binding showed a strong decrease (40%) in receptor/G protein coupling in the limbic area of these animals. Moreover, in the same brain regions we measured, by isotope-dilution gas chromatography/mass spectrometry, the contents of AEA and 2-AG. Chronic morphine exposure produced a strong reduction in 2-AG contents without changes in AEA levels in several brain regions (ie striatum, cortex, hippocampus, limbic area, and hypothalamus). These findings clearly demonstrate that prolonged activation of opioid receptors could alter the cannabinoid system, in terms of both receptor functionality and endocannabinoid levels, and suggest the involvement of this system, alone or in combination with other mediators, in the phenomenon of morphine tolerance.