Diethyldithiocarbamate encapsulation reduces toxicity and promotes leishmanicidal effect through apoptosis-like mechanism in promastigote and ROS production by macrophage.

The use of compounds from natural or synthetic sources and nanotechnology may represent an alternative to develop new drugs for the leishmaniasis treatment. DETC is an inhibitor of the SOD1 enzyme, which leads to increased ROS production, important for the elimination of Leishmania. Thus, our objective was to assess the leishmanicidal in vitro effect of free Diethydithiocarbamate (DETC) and DETC loaded in beeswax-copaiba oil nanoparticles (DETC-Beeswax-CO Nps) on L. amazonensis forms and elucidate the possible mechanisms involved in the parasite death. DETC-Beeswax-CO Nps presented size below 200 nm, spherical morphology, negative zeta potential, and high encapsulation efficiency. Free DETC reduced the viability of promastigotes and increase ROS production, lower the mitochondrial membrane potential, cause phosphatidylserine exposure, and enhance plasma membrane permeability, in addition to promoting morphological changes in the parasite. Free DETC proved toxic in the assessment of toxicity to murine macrophages, however, the encapsulation of this compound was able to reduce these toxic effects on macrophages. DETC-Beeswax-CO Nps exerted anti-amastigote effect by enhancing the production of ROS, superoxide anion, TNF-α, IL-6, and reduced IL-10 in macrophages. Therefore, free DETC induces antipromastigote effect by apoptosis-like; and DETC-Beeswax-CO Nps exerted anti-leishmanial effect due to pro-oxidant and pro-inflammatory response.

Doxorubicin intercalated copper diethyldithiocarbamate functionalized layered double hydroxide hybrid nanoparticles for targeted therapy of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the deadliest cancers due to its long incubation period and low cure rate. Layered double hydroxide (LDH) nanoparticles have attracted considerable research interest in the field of nanomedicine owing to their surface effects and good biocompatibility. In this research, we synthesized a hexagonal nanoparticle by the co-precipitation method, referred to as Cu-Al LDH. As an alternative to traditional drug-loading methods, sodium diethyldithiocarbamate (DDC) was introduced and combined with Cu2+ in LDHs to form a diethyldithiocarbamate-copper complex (Cu(DDC)2), which was not only the composition of carrier materials but also an effective component for cancer therapy. Doxorubicin (DOX) was also encapsulated into LDHs due to the clinical relevance of DOX treatment for HCC. Formulations of the Cu(DDC)2 and DOX co-loaded nanoparticles were optimized to precisely control the Cu(DDC)2/DOX ratio. The nanoparticles were coated with polyethylene glycol-graft-polyglutamic acid (PEG-PLG) through electrostatic adsorption to improve the stability of the nanoparticles. The outer layer was decorated with hyaluronic acid (HA) to achieve specific targeting of tumors. Compared with non-HA coated nanoparticles, HA coated nanoparticles showed greater cellular uptake in Hep G2 cells, which could cause higher cytotoxicity. In addition, targeted nanoparticles effectively inhibited tumor growth in mouse models of ectopic hepatocellular carcinoma. It can be concluded that there is a great potential for synergistic cancer therapy using the novel DOX intercalated Cu(DDC)2 functionalized layered double hydroxide hybrid nanoparticles.

Development and optimization of an injectable formulation of copper diethyldithiocarbamate, an active anticancer agent.

Copper diethyldithiocarbamate (Cu(DDC)2) is the active anticancer agent generated when disulfiram (DSF) is provided in the presence of copper. To date, research directed toward repurposing DSF as an anticancer drug has focused on administration of DSF and copper in combination, efforts that have proven unsuccessful in clinical trials. This is likely due to the inability to form Cu(DDC)2 at relevant concentrations in regions of tumor growth. Little effort has been directed toward the development of Cu(DDC)2 because of the inherent aqueous insolubility of the complex. Here, we describe an injectable Cu(DDC)2 formulation prepared through a method that involves synthesis of Cu(DDC)2 inside the aqueous core of liposomes. Convection-enhanced delivery of a Cu(DDC)2 formulation prepared using 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC)/cholesterol liposomes into a rat model of F98 glioma engendered a 25% increase in median survival time relative to vehicle-treated animals. In a murine subcutaneous MV-4-11 model, treatment resulted in a 45% reduction in tumor burden when compared to controls. Pharmacokinetic studies indicated that the Cu(DDC)2 was rapidly eliminated after intravenous administration while the liposomes remained in circulation. To test whether liposomal lipid composition could increase Cu(DDC)2 circulation lifetime, a number of different formulations were evaluated. Studies demonstrated that liposomes composed of DSPC and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-n-(carboxy[polyethylene glycol]-2000) (95:5) enhanced Cu(DDC)2 concentrations in the circulation as reflected by a 4.2-fold increase in plasma AUC(0-∞) relative to the DSPC/cholesterol formulation. The anticancer activity of this Cu(DDC)2 formulation was subsequently evaluated in the MV-4-11 model. At its maximum tolerated dose, this formulation exhibited comparable activity to the DSPC/cholesterol formulation. This is the first report demonstrating the therapeutic effects of an injectable Cu(DDC)2 formulation in vivo.

Ammonium salts of carbamodithioic acid as potent vaginal trichomonacides and fungicides.

Chemical attenuation of the reactive oxygen species (ROS)-sensitive anaerobes Trichomonas vaginalis, which is the most prevalent non-viral sexually transmitted infection, and two often coexisting vaginal infections, namely Candida albicans and Staphylococcus aureus, which are opportunistic reproductive tract infections, was attempted with novel ammonium salts of carbamodithioic acid through inhibition of free thiols. In vitro and in vivo efficacies of the designed compounds were evaluated as topical vaginal microbicides. Five compounds showed exceptional activity against drug-resistant and -susceptible strains with negligible toxicity to host (HeLa) cells in vitro in comparison with the standard vaginal microbicide nonoxynol-9 (N-9), without disturbing the normal vaginal flora (i.e. Lactobacillus). The compounds significantly inhibited the cytopathic effects of Trichomonas on HeLa cells in vitro with efficacies comparable with metronidazole (MTZ); however, their efficacy to rescue host cells from co-infection (protozoal and fungal) was greater than that of MTZ. The compounds inhibited ß-haemolysis of red blood cells caused by Trichomonas and were found to be active in vivo in the mouse subcutaneous abscess assay. Some compounds rapidly immobilized human sperm. A mechanism involving inhibition of free thiols and consequently the cysteine proteases of T. vaginalis by the new compounds has been proposed. Thus, a unique scaffold of antimicrobial agents has been discovered that warrants further investigation for development as contraceptive vaginal microbicides.

The importance of therapeutic drug monitoring (TDM) for parenteral busulfan dosing in conditioning regimen for hematopoietic stem cell transplantation (HSCT) in children.

BACKGROUND: Series of observations indicate PK/PD variability challenging the accuracy of the body-weight based busulfan (Bu) dosing schedule for (HSCT) conditioning therapy. The purpose of this communication is to describe the frequency of dose changes in initially body-weight-based fixed IV Bu dose and to emphasize the importance of TDM. MATERIAL AND METHODS: Sixty-two children (ages 2 months-18 years) were treated with IV busulfan doses based on body weight for myeloablation. TDM utilizing a limited sample strategy (trough concentration immediately before the 5th dose, followed by samples immediately after the end of the 2-h infusion peak, 4 h, and 6 h from initiation of the infusion) was performed in 46 of 62 subjects. Busulfan concentrations were determined by high-performance liquid chromatography (HPLC). AUC was calculated according to the trapezoidal rule. RESULTS: We observed trough levels of 25-1244 µg/L, peak levels of 849-4586 µg/L, and AUC of 2225-12818 µg/L·h following body weight-based high-dose busulfan. The doses were changed in 54% of cases. AUC in 5 of 9 patients with VOD were within target, in 3 patients AUS was higher, and in 1 patient AUC was lower. One of the 2 patients with neurotoxicity had higher AUC. Engraftment was 100%, but relapse occurred in 25% of cases. CONCLUSIONS: Our results demonstrate that even with IV busulfan, intra-individual PK/PD variability is challenging. Although AUC does not necessarily correspond with outcomes (due to the role of other factors the fact that doses were changed in 54% of cases underlines the importance of TDM.

Cytoprotective effect of eckol against oxidative stress-induced mitochondrial dysfunction: involvement of the FoxO3a/AMPK pathway.

This study investigated the cytoprotective effect of Ecklonia cava-derived eckol against H2O2-induced mitochondrial dysfunction in Chang liver cells. While H2O2 augmented levels of mitochondrial reactive oxygen species (ROS), eckol decreased it. Eckol also attenuated high intracellular Ca(2+) levels stimulated by H2O2 and recovered H2O2-diminished ATP levels and succinate dehydrogenase activity. Eckol time-dependently increased the expression of manganese superoxide dismutase (Mn SOD), a mitochondrial antioxidant enzyme with cytoprotective effect against oxidative stress. Eckol recovered Mn SOD expression and activity that were decreased by H2O2. Finally, eckol induced Mn SOD through phosphorylated AMP-activated protein kinase (AMPK) and forkhead box O3a (FoxO3a). Specific silencing RNAs (siRNAs) against FoxO3a and AMPK reduced eckol-stimulated Mn SOD expression, and diethyldithiocarbamate (Mn SOD inhibitor) and siRNA against Mn SOD reduced the cytoprotective effect of eckol against H2O2-provoked cell death. These results demonstrate that eckol protects cells from mitochondrial oxidative stress by activating AMPK/FoxO3a-mediated induction of Mn SOD.

Dithiocarb (N,N-diethyldithiocarbamate, DEDTC) decreases levels of biogenic monoamines in the adult mouse brain.

AIMS: Dithiocarb (diethyldithiocarbamate, DEDTC) belongs to the group of dithiocarbamates and is the main metabolite of disulphiram, a drug of choice for the treatment of alcohol dependence. Its therapeutic potential relays on its ability to create an unpleasant aversive reaction following the ingestion of alcohol, and this effect is usually accompanied by neurobehavioural symptoms. Most of these can be attributed to the impaired metabolism of brain biogenic amines. METHODS: To gain new insights into the dithiocarbamates and their effects on neurotransmitter systems, an in vivo experimental model based on daily injections of DEDTC in adult mice for 7 days was established. To this end, the concentrations of the three major brain monoamines, dopamine (DA), noradrenaline (NA) and serotonin (5-HT) were measured in whole brain extracts with high-performance liquid chromatography (HPLC). The levels of D2 dopamine receptor (D2R) were evaluated by Western blot and by immunohistochemical techniques the cell pattern of tyrosine hydroxylase (TH), dopa beta hydroxylase (DBH) and choline acetyltransferase ChAT) were analysed. RESULTS: A significant reduction in DA and 5-HT levels was observed, whereas NA was not affected. Moreover, decreases in D2R levels, as well as in enzymes such as TH, DBH and ChAT, were found. CONCLUSIONS: Our data suggest that DEDTC provokes alterations in biogenic amines and in different substrates of neurotransmitter systems, which could explain some of the neurobehavioural effects observed in patients treated with disulphiram.

Single-cell resolution mapping of neuronal damage in acute focal cerebral ischemia using thallium autometallography.

Neuronal damage shortly after onset or after brief episodes of cerebral ischemia has remained difficult to assess with clinical and preclinical imaging techniques as well as with microscopical methods. We here show, in rodent models of middle cerebral artery occlusion (MCAO), that neuronal damage in acute focal cerebral ischemia can be mapped with single-cell resolution using thallium autometallography (TlAMG), a histochemical technique for the detection of the K(+)-probe thallium (Tl(+)) in the brain. We intravenously injected rats and mice with thallium diethyldithiocarbamate (TlDDC), a lipophilic chelate complex that releases Tl(+) after crossing the blood-brain barrier. We found, within the territories of the affected arteries, areas of markedly reduced neuronal Tl(+) uptake in all animals at all time points studied ranging from 15 minutes to 24 hours after MCAO. In large lesions at early time points, areas with neuronal and astrocytic Tl(+) uptake below thresholds of detection were surrounded by putative penumbral zones with preserved but diminished Tl(+) uptake. At 24 hours, the areas of reduced Tl(+)uptake matched with areas delineated by established markers of neuronal damage. The results suggest the use of (201)TlDDC for preclinical and clinical single-photon emission computed tomography (SPECT) imaging of hyperacute alterations in brain K(+) metabolism and prediction of tissue viability in cerebral ischemia.

Electrophilic adduction of ubiquitin activating enzyme E1 by N,N-diethyldithiocarbamate inhibits ubiquitin activation and is accompanied by striatal injury in the rat.

Previous studies have shown ubiquitin activating enzyme E1 to be sensitive to adduction through both Michael addition and SN(2) chemistry in vitro. E1 presents a biologically important putative protein target for adduction due to its role in initiating ubiquitin based protein processing and the involvement of impaired ubiquitin protein processing in two types of familial Parkinson's disease. We tested whether E1 is susceptible to xenobiotic-mediated electrophilic adduction in vivo and explored the potential contribution of E1 adduction to neurodegenerative events in an animal model. N,N-Diethyldithiocarbamate (DEDC) was administered to rats using a protocol that produces covalent cysteine modifications in vivo, and brain E1 protein adducts were characterized and mapped using shotgun LC-MS/MS. E1 activity, global and specific protein expression, and protein carbonyls were used to characterize cellular responses and injury in whole brain and dorsal striatal samples. The data demonstrate that DEDC treatment produced S-(ethylaminocarbonyl) adducts on Cys234 and Cys179 residues of E1 and decreased the levels of activated E1 and total ubiquitinated proteins. Proteomic analysis of whole brain samples identified expression changes for proteins involved in myelin structure, antioxidant response, and catechol metabolism, systems often disrupted in neurodegenerative disease. Our studies also delineated localized injury within the striatum as indicated by decreased levels of tyrosine hydroxylase, elevated protein carbonyl content, increased antioxidant enzyme and α-synuclein expression, and enhanced phosphorylation of tau and tyrosine hydroxylase. These data are consistent with E1 having similar susceptibility to adduction in vivo as previously reported in vitro and support further investigation into environmental agent adduction of E1 as a potential contributing factor to neurodegenerative disease. Additionally, this study supports the predictive value of in vitro screens for identifying sensitive protein targets that can be used to guide subsequent in vivo experiments.

Defensive effect of natrium diethyldithiocarbamate trihydrate (NDDCT) and lisinopril in DOCA-salt hypertension-induced vascular dementia in rats.

RATIONALE: Vascular dementia and hypertension are increasing day by day, with a high degree of co-occurrence. Tremendous amount of research work is required so that new pharmacological agents may be identified for their appropriate therapeutic utility to combat different dementing disorders. OBJECTIVES: This study investigates the effect of natrium diethyldithiocarbamate trihydrate (NDDCT), a nuclear factor kappa-B (NF-κB) inhibitor, as well as lisinopril, an angiotensin converting enzyme (ACE) inhibitor, on deoxycorticosterone acetate (DOCA) hypertension-induced vascular dementia in rats. METHODS: DOCA was used to induce hypertension and associated vascular dementia. Morris water maze (MWM) was used for testing learning and memory. Endothelial function was assessed by acetylcholine-induced endothelium-dependent relaxation of aortic strips. Different biochemical estimations were used to assess oxidative stress (aortic superoxide anion, serum and brain thiobarbituric acid reactive species, and brain glutathione), nitric oxide levels (serum nitrite/nitrate), and cholinergic activity (brain acetyl cholinesterase activity). RESULTS: DOCA treatment significantly raised the mean arterial blood pressure of rats, and these hypertensive rats performed poorly on MWM, reflecting impairment of learning and memory. DOCA treatment also impaired vascular endothelial function and different biochemical parameters. Treatments of NDDCT as well as lisinopril significantly attenuated DOCA hypertension-induced impairment of learning and memory, endothelial dysfunction, and changes in various biochemical levels. CONCLUSIONS: DOCA-salt hypertension induces vascular dementia in rats. NF-κB as well as ACE inhibitors may be considered as potential pharmacological agents for the management of hypertension-induced vascular dementia.

Ameliorative potential of sodium cromoglycate and diethyldithiocarbamic acid in restraint stress-induced behavioral alterations in rats.

The present study was designed to investigate the ameliorative effects of sodium cromoglycate and diethyldithiocarbamic acid in acute stress-induced behavioral alterations in rats subjected to restraint stress. The rats were placed in the restrainer (5.5 cm in diameter and 18 cm in length) for 3.5 h. Restraint stress-induced behavioral alterations were assessed using the hole-board, social interactions and open field tests. Restraint stress resulted in a decrease in the frequency of head dips, rearing in the hole board, line crossings and rearings in the open field, and an increase in avoidance behaviors in the social interaction tests. Sodium cromoglycate (25 mg/kg and 50 mg/kg, ip), a mast cell stabilizer, and diethyldithiocarbamic acid (75 mg/kg and 150 mg/kg, ip), a selective NF-κB inhibitor, were employed to modulate restraint stress-induced behavioral changes. The administration of sodium cromoglycate and diethyldithiocarbamic acid significantly attenuated the restraint stress-induced behavioral changes. The noted beneficial effects of sodium cromoglycate and diethyldithiocarbamic acid may possibly be attributed to mast cell stabilization and inhibition of NF-κB activity, respectively.

Preparation of the core-shell structure adriamycin lipiodol microemulsions and their synergistic anti-tumor effects with diethyldithiocarbamate in vivo.

We prepared the core-shell structure adriamycin lipiodol microemulsions (ADM-CSLMs) and evaluated their in vivo antitumor effects in combination with Diethyldithiocarbamate (DDC). Two types of ADM-CSLMs, adriamycin liposome-lipiodol microemulsion(ADM-LLM) and adriamycin microsphere lipiodol microemulsion (ADM-MLM), were prepared through the emulsification method. The drug loading and encapsulation efficiency of ADM-CSLMs were measured by the high-performance liquid chromatograph (HPLC). The size and shape of the ADM-CSLMs were determined by an atom force microscopy (AFM), a transmission electron microscopy (TEM), and a particle size analyzer, respectively. The synergistic effects of DDC and ADM-CSLMs for cancer treatment of carcinoma drug-resistance cell was evaluated by the MTT method, the activation of superoxide dismutase (SOD) was detected by chemiluminescence, and the ADM accumulation in cells was measured by flow cytometry. Walker-256 carcinoma was transplanted to the livers of the male SD rats, ADM-CSLMs were administrated to the livers of the rats by intervention hepatic artery embolization through microsurgery. The tumor growth and animal survival were evaluated. The results show that the average diameter of ADM-LLM and ADM-MLM were 4.23 ± 1.2 µm and 4.67 ± 1.4 µm, respectively, and their ADM encapsulation efficiency were 83.7% and 87.2% with respect to loading efficiency of 82 µg/ml and 91 µg/ml. The tumor growth and animal survival in two of the ADM-CSLMs combined with DDC groups were significantly higher than that of ADM only treatment, ADM liposome combined with DDC (P < 0.01), as well as the ADM microsphere combined with DDC (P < 0.01). Therefore, ADM-CSLMs are useful carriers for the treatment of carcinoma and their anti-tumor effect can be enhanced by DDC in a suitable concentration.

Attenuation of renal excretory responses to ANG II during inhibition of superoxide dismutase in anesthetized rats.

To examine the functional interaction between superoxide dismutase (SOD) and NADPH oxidase activity, we assessed renal responses to acute intra-arterial infusion of ANG II (0.5 ng x kg(-1) x min(-1)) before and during administration of a SOD inhibitor, diethyldithiocarbamate (DETC, 0.5 mg x kg(-1) x min(-1)), in enalaprilat-pretreated (33 microg x kg(-1) x min(-1)) rats (n = 11). Total (RBF) and regional (cortical, CBF; medullary; MBF) renal blood flows were determined by Transonic and laser-Doppler flowmetry, respectively. Renal cortical and medullary tissue NADPH oxidase activity in vitro was determined using the lucigenin-chemiluminescence method. DETC treatment alone resulted in decreases in RBF, CBF, MBF, glomerular filtration rate (GFR), urine flow (V), and sodium excretion (U(Na)V) as reported previously. Before DETC, ANG II infusion decreased RBF (-18 +/- 3%), CBF (-16 +/- 3%), MBF [-5 +/- 6%; P = not significant (NS)], GFR (-31 +/- 4%), V (-34 +/- 2%), and U(Na)V (-53 +/- 3%). During DETC infusion, ANG II also caused similar reductions in RBF (-20 +/- 4%), CBF (-19 +/- 3%), MBF (-2 +/- 2; P = NS), and in GFR (-22 +/- 7%), whereas renal excretory responses (V; -12 +/- 2%; U(Na)V; -24 +/- 4%) were significantly attenuated compared with those before DETC. In in vitro experiments, ANG II (100 muM) enhanced NADPH oxidase activity both in cortical [13,194 +/- 1,651 vs. 20,914 +/- 2,769 relative light units (RLU)/mg protein] and in medullary (21,296 +/- 2,244 vs. 30,597 +/- 4,250 RLU/mg protein) tissue. Application of DETC (1 mM) reduced the basal levels and prevented ANG II-induced increases in NADPH oxidase activity in both tissues. These results demonstrate that renal excretory responses to acute ANG II administration are attenuated during SOD inhibition, which seems related to a downregulation of NADPH oxidase in the deficient condition of SOD activity.

High-resolution mapping of neuronal activity using the lipophilic thallium chelate complex TlDDC: protocol and validation of the method.

In neurons the rate of K(+)-uptake increases with increasing activity. K(+)-analogues like the heavy metal ion thallium (Tl(+)) can be used, therefore, as tracers for imaging neuronal activity. However, when water-soluble Tl(+)-salts are injected systemically only minute amounts of the tracer enter the brain and the Tl(+)-uptake patterns are influenced by regional differences in blood-brain barrier (BBB) K(+)-permeability. We here show that the BBB-related limitations in using Tl(+) for imaging neuronal activity are no longer present when the lipophilic Tl(+) chelate complex thallium diethyldithiocarbamate (TlDDC) is applied. We systemically injected rodents with TlDDC and mapped the Tl(+)-distribution in the brain using an autometallographic (AMG) technique, a histochemical method for detecting heavy metals. We find that Tl(+)-doses for optimum AMG staining could be substantially reduced, and regional differences attributable to differences in BBB K(+)-permeability were no longer detectable, indicating that TlDDC crosses the BBB. At the cellular level, however, the Tl(+)-distribution was essentially the same as after injection of water-soluble Tl(+)-salts, indicating Tl(+)-release from TlDDC prior to neuronal or glial uptake. Upon sensory stimulation or intracortical microstimulation neuronal Tl(+)-uptake increased after TlDDC injection, upon muscimol treatment neuronal Tl(+)-uptake decreased. We present a protocol for mapping neuronal activity with cellular resolution, which is based on intravenous TlDDC injections during ongoing activity in unrestrained behaving animals and short stimulation times of 5 min.

Persistent pain is dependent on spinal mitochondrial antioxidant levels.

Reactive oxygen species (ROS) scavengers have been shown to relieve persistent pain; however, the mechanism is not clearly understood. Superoxide produced from mitochondrial oxidative phosphorylation is considered the major source of ROS in neurons during excitation where mitochondrial superoxide levels are normally controlled by superoxide dismutase (SOD-2). The present study hypothesizes that capsaicin-induced secondary hyperalgesia is a consequence of superoxide build-up in spinal dorsal horn neurons and SOD-2 is a major determinant. To test this hypothesis, the spinal levels of SOD-2 activity, inactivated SOD-2 proteins, and mitochondrial superoxide were measured and correlated to the levels of capsaicin-induced secondary hyperalgesia in mice with and without SOD-2 manipulations. The data suggest that superoxide accumulation is a culprit in the abnormal sensory processing in the spinal cord in capsaicin-induced secondary hyperalgesia. Our studies also support the notion that SOD-2 nitration is a critical mechanism that maintains elevated superoxide levels in the spinal cord after capsaicin treatment. Finally, our findings suggest a therapeutic potential for the manipulation of spinal SOD-2 activity in pain conditions.

Nitrogen substituent polarity influences dithiocarbamate-mediated lipid oxidation, nerve copper accumulation, and myelin injury.

Dithiocarbamates have a wide spectrum of applications in industry, agriculture, and medicine, with new applications being investigated. Past studies have suggested that the neurotoxicity of some dithiocarbamates may result from copper accumulation, protein oxidative damage, and lipid oxidation. The polarity of a dithiocarbamate's nitrogen substituents influences the lipophilicity of the copper complexes that it generates and thus potentially determines its ability to promote copper accumulation within nerve and induce myelin injury. In the current study, a series of dithiocarbamate-copper complexes differing in their lipophilicity were evaluated for their relative abilities to promote lipid peroxidation determined by malondialdehyde levels generated in an ethyl arachidonate oil-in-water emulsion. In a second component of this study, rats were exposed to either N,N-diethyldithiocarbamate or sarcosine dithiocarbamate; both generated dithiocarbamate-copper complexes that were lipid- and water-soluble, respectively. Following the exposures, brain, tibial nerve, spinal cord, and liver tissue copper levels were measured by inductively coupled mass spectroscopy to assess the relative abilities of these two dithiocarbamates to promote copper accumulation. Peripheral nerve injury was evaluated using grip strengths, nerve conduction velocities, and morphologic changes at the light microscope level. Additionally, the protein expression levels of glutathione transferase alpha and heme-oxygenase-1 in nerve were determined, and the quantity of protein carbonyls was measured to assess levels of oxidative stress and injury. The data provided evidence that dithiocarbamate-copper complexes are redox active and that the ability of dithiocarbamate complexes to promote lipid peroxidation is correlated to the lipophilicity of the complex. Consistent with neurotoxicity requiring the formation of a lipid-soluble copper complex, significant increases in copper accumulation, oxidative stress, and myelin injury were produced by N,N-diethyldithiocarbamate but not by sarcosine dithiocarbamate.

Nuclear factor-kappa-B inhibitor modulates the development of opioid dependence in a mouse model of naloxone-induced opioid withdrawal syndrome.

The present study was designed to investigate the effect of diethyl dithiocarbamic acid sodium salt trihydrate (DDA), a selective inhibitor of nuclear factor-kappa-B, on the development of morphine dependence in a mouse model of naloxone-induced opioid withdrawal syndrome. Morphine (5 mg/kg, intraperitoneally) was administered twice daily for a period of 5 days, after which a single injection of naloxone (8 mg/kg, intraperitoneally) precipitated an opioid withdrawal syndrome in mice. Behavioral observations were made for a period of 30 min immediately after naloxone treatment. Withdrawal syndrome was quantitatively assessed in terms of withdrawal severity score and the frequency of jumping, rearing, forepaw licking, and circling. DDA markedly and dose-dependently (P<0.01) attenuated the morphine-naloxone-induced experimental opioid withdrawal syndrome. However, DDA administration did not alter locomotor activity, thus ruling out any sedative action of DDA per se. Further, DDA pretreatment did not alter the acute analgesic effect of morphine. The results suggest that nuclear factor-kappa-B is involved in the development of opioid dependence and the precipitation of its withdrawal syndrome, and thus, may serve as a viable pharmacological target to tackle the problem of opioid addiction.

NF-kappaB pathway inhibitors preferentially inhibit breast cancer stem-like cells.

Accumulating evidence indicates that breast cancer is caused by cancer stem cells and cure of breast cancer requires eradication of breast cancer stem cells. Previous studies with leukemia stem cells have shown that NF-kappaB pathway is important for leukemia stem cell survival. In this study, by using MCF7 sphere cells as model of breast cancer stem-like cells, we evaluated the effect of NF-kappaB pathway specific inhibitors on human breast cancer MCF7 sphere cells. Three inhibitors including parthenolide (PTL), pyrrolidinedithiocarbamate (PDTC) and its analog diethyldithiocarbamate (DETC) were found to preferentially inhibit MCF7 sphere cell proliferation. These compounds also showed preferential inhibition in term of proliferation and colony formation on MCF7 side population (SP) cells, a small fraction of MCF7 cells known to enrich in breast cancer stem-like cells. The preferential inhibition effect of these compounds was due to inhibition of the NF-kappaB activity in both MCF7 sphere and MCF7 cells, with higher inhibition effect on MCF7 sphere cells than on MCF7 cells. PDTC was further evaluated in vivo and showed significant tumor growth inhibition alone but had better tumor growth inhibition in combination with paclitaxel in the mouse xenograft model than either PDTC or paclitaxel alone. This study suggests that breast cancer stem-like cells could be selectively inhibited by targeting signaling pathways important for breast cancer stem-like cells.

Camostat, an oral trypsin inhibitor, reduces pancreatic fibrosis induced by repeated administration of a superoxide dismutase inhibitor in rats.

BACKGROUND AND AIM: An oral trypsin inhibitor, camostat (CM), has a beneficial effect on chronic pancreatitis, but its mechanism is not yet fully understood. Recently, pancreatic stellate cells (PSC) have been reported to play an essential role in pancreatic fibrosis. An experimental model of pancreatic fibrosis induced by a superoxide dismutase (SOD) inhibitor (diethyldithiocarbamate [DDC]) was developed in rats. Thus, the effect of an oral trypsin inhibitor on pancreatic fibrosis and PSC was investigated. METHODS: Pancreatic fibrosis was induced in rats using DDC (DDC rats). DDC + CM rats were administered DDC, and subsequently were fed a diet containing CM. Immunohistochemistry of the pancreas was performed with monoclonal anti-alpha-smooth muscle actin (alpha-SMA) antibody and anti-desmin antibody. RESULTS: The DDC rats showed a significant increase in alpha-SMA-positive cells or desmin-positive cells compared with control rats. These significant increases in the fibrotic area improved after treatment with CM. The level of prolyl hydroxylase in the pancreas, which significantly increased as a result of DDC, decreased after treatment with CM. CONCLUSION: Camostat has a beneficial effect on pancreatic fibrosis induced by the administration of a SOD inhibitor, which inhibits the proliferation and activation of PSC.

An ocular drug delivery system containing zinc diethyldithiocarbamate and HPbetaCD inclusion complex--corneal permeability, anti-cataract effects and mechanism studies.

Our purpose was to study the formulation and anti-cataract effects of aqueous eye drops containing a high concentration of zinc diethyldithiocarbamate (Zn-DDC). A possible mechanism of the anti-cataract effect of Zn-DDC was also studied. Zn-DDC and hydroxypropyl-beta-cyclodextrin (HPbetaCD) inclusion complex (Zn-DDC/HPbetaCD) was studied using the saturation solution method and characterized by differential scanning calorimetry (DSC) and Fourier transform infrared spectroscopy (IR). Suitable formulations for Zn-DDC eye drops were established by means of in-vitro trans-corneal penetration experiments. The anti-cataract effect of the selected formulation was demonstrated by the delay in lens opacity development in hereditary shumuya cataract rats (SCRs). Semiquantitative reverse transcription polymerase chain reaction (RT-PCR) was performed to study the effect of diethyldithiocarbamate (DDC), a metabolite of Zn-DDC, on the transcription inducible nitric oxide synthase (iNOS) mRNA in human lens epithelial cells (HLEC). In the presence of 22% (w/v) HPbetaCD, the solubility of Zn-DDC in water (0.2 mM) was increased almost 850 fold (to 17 mM), by the formation of Zn-DDC/HPbetaCD. The stoichiometry of Zn-DDC inclusion was 1:1. The Zn-DDC/HPbetaCD stability constant, Ks (1:1) was estimated to be 3453 M(-1). The ophthalmic preparation containing 0.1% HPMC and 0.1% poloxamer 188 (P188) exhibited better permeability than the others in-vitro, and significantly delayed cataract formation in SCRs compared with non-treated SCRs. DDC inhibits the transcription of iNOS mRNA in HLEC. We concluded that this drug delivery system increases both the drug solubility in aqueous eye drops and the permeability of drug through the rabbit cornea, by the formation of a drug-cyclodextrin inclusion complex and the addition of polymers and penetration enhancers. The preparation effectively prevented the development of cataracts in SCRs. DDC, the metabolite of Zn-DDC, may be one of the factors in the prevention of cataract formation because it inhibits the transcription of iNOS mRNA.