Polymer-drug conjugates for combination anticancer therapy: investigating the mechanism of action.

We developed a family of polymer-drug conjugates carrying the combination of the anticancer agent epirubicin (EPI) and nitric oxide (NO). EPI-PEG-(NO)8, carrying the highest content of NO, displayed greater activity in Caco-2 cells while it decreased toxicity against endothelium cells and cardiomyocytes with respect to free EPI. FACS and confocal microscopy confirmed conjugates internalization. Light scattering showed formation of micelle whose size correlated with internalization rate. EPI-PEG-(NO)8 showed increased bioavailability in mice compared to free EPI.

Chronic toxicity of ibuprofen to Daphnia magna: Effects on life history traits and population dynamics

The non-steroidal anti-inflammatory drug (NSAID) ibuprofen (IB) is a widely used pharmaceutical that can be found in several freshwater ecosystems. Acute toxicity studies with Daphnia magna suggest that the 48 h EC50 (immobilisation) is 10-100 mg IB l(-1). However, there are currently no chronic IB toxicity dataon arthropod populations, and the aquatic life impacts of such analgesic drugs are still undefined. We performed a 14-day exposure of D. magna to IB as a model compound (concentration range: 0, 20, 40 and 80 mg IB l(-1)) measuring chronic effects on life history traits and population performance. Population growth rate was significantly reduced at all IB concentrations, although survival was only affected at 80 mg IB l(-1). Reproduction, however, was affected at lower concentrations of IB (14-day EC50 of 13.4 mg IB l(-1)), and was completely inhibited at the highest test concentration. The results from this study indicate that the long-term crustacean population consequences of a chronic IB exposure at environmentally realistic concentrations (ng l(-1) to mu g l(-1)) would most likely be of minor importance. We discuss our results in relation to recent genomic studies, which suggest that the potential mechanism of toxicity in Daphnia is similar to the mode of action in mammals, where IB inhibits eicosanoid biosynthesis. (C) 2007 Elsevier Ireland Ltd. All rights reserved.

Polymer-drug conjugates: towards a novel approach for the treatment of endrocine-related cancer

The last decade has seen successful clinical application of polymer–protein conjugates (e.g. Oncaspar, Neulasta) and promising results in clinical trials with polymer–anticancer drug conjugates. This, together with the realisation that nanomedicines may play an important future role in cancer diagnosis and treatment, has increased interest in this emerging field. More than 10 anticancer conjugates have now entered clinical development. Phase I/II clinical trials involving N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer-doxorubicin (PK1; FCE28068) showed a four- to fivefold reduction in anthracycline-related toxicity, and, despite cumulative doses up to 1680 mg/m2 (doxorubicin equivalent), no cardiotoxicity was observed. Antitumour activity in chemotherapy-resistant/refractory patients (including breast cancer) was also seen at doxorubicin doses of 80–320 mg/m2, consistent with tumour targeting by the enhanced permeability (EPR) effect. Hints, preclinical and clinical, that polymer anthracycline conjugation can bypass multidrug resistance (MDR) reinforce our hope that polymer drugs will prove useful in improving treatment of endocrine-related cancers. These promising early clinical results open the possibility of using the water-soluble polymers as platforms for delivery of a cocktail of pendant drugs. In particular, we have recently described the first conjugates to combine endocrine therapy and chemotherapy. Their markedly enhanced in vitro activity encourages further development of such novel, polymer-based combination therapies. This review briefly describes the current status of polymer therapeutics as anticancer agents, and discusses the opportunities for design of second-generation, polymer-based combination therapy, including the cocktail of agents that will be needed to treat resistant metastatic cancer.

In vitro basal cytotoxicity assay applied to estimate acute oral systemic toxicity of grandisin and its major metabolite

Preclinical investigations can start with preliminary in vitro studies before using animal models. Following this approach, the number of animals used in preclinical acute toxicity testing can be reduced. In this study, we employed an in-house validated in vitro cytotoxicity test based on the Spielmann approach for toxicity evaluation of the lignan grandisin, a candidate anticancer agent, and its major metabolite. the 4-O-demethylgrandisin, by neutral red uptake (NRU) assay, on mouse fibroblasts Balb/c 3T3 cell line. Using different concentrations of grandisin and its major metabolite (2.31; 1.16; 0.58; 0.29; 0.14; 0.07; 0.04; 0.002 mu M) in Balb/c 3T3-A31 NRU cytotoxicity assay, after incubation for 48 h, we obtained IC(50) values for grandisin and its metabolite of 0.078 and 0.043 mu M, respectively. The computed LD(50) of grandisin and 4-O-demethylgrandisin were 617.72 and 429.95 mg/kg, respectively. Both were classified under the Globally Harmonized System as category 4. Since pharmacological and toxicological data are crucial in the developmental stages of drug discovery, using an in vitro assay we demonstrated that grandisin and its metabolite exhibit distinct toxicity profiles. Furthermore, results presented in this work can contribute to reduce the number of animals required in subsequent pharmacological/toxicological studies. (C) 2010 Elsevier GmbH. All rights reserved.

Complexation of the anti-Trypanosoma cruzi drug benznidazole improves solubility and efficacy

The ruthenium complex,trans-[Ru(Bz)(NH3)(4)SO2](CF3SO3)(2) 1, Bz = benznidazole (N-benzyl-2-(2-nitro-1H-imidazol-1-yl)acetamide), is more hydrosoluble and more active (IC50try/1 h = 79 +/- 3 mu M) than free benznidazole 2 (IC50try/1 h > 1 mM). 1 also exhibits low acute toxicity in vitro (IC50macrophages > 1 mM) and in vivo (400 mu mol/kg < LD50 < 600 mu mol/kg) and the formation of hydroxylamine is more favorable in 1 than in 2 by 9.6 kcal/mol. In murine acute models of Chagas` disease, 1 was more active than 2 even when only one dose was administrated. Moreover, 1 at a thousand-fold smaller concentration than the considered optimal dose for 2 (385 mu mol/kg/day = 100 mg/kg/day), proved to be sufficient to protect all infected mice, eliminating the amastigotes in their hearts and skeletal muscles as observed in H&E micrographics.

Similarity between the in vitro activity and toxicity of two different fungizone™ / lipofundin™ admixtures

Amphotericin B (AmB), an antifungal agent that presents a broad spectrum of activity, remains the gold standard in the antifungal therapy. However, sometimes the high level of toxicity forbids its clinical use. The aim of this work was to evaluate and compare the efficacy and toxicity in vitro of Fungizon™ (AmB-D) and two new different AmB formulations. Methods: three products were studied: Fungizon™, and two Fungizon™ /Lipofundin™ admixtures, which were diluted through two methods: in the first one, Fungizon™ was previously diluted with water for injection and then, in Lipofundin™ (AmB-DAL); the second method consisted of a primary dilution of AmB-D as a powder in the referred emulsion (AmB-DL). For the in vitro assay, two cell models were used: Red Blood Cells (RBC) from human donors and Candida tropicallis (Ct). The in vitro evaluation (K+ leakage, hemoglobin leakage and cell survival rate-CSR) was performed at four AmB concentrations (from 50 to 0.05mg.L-1). Results: The results showed that the action of AmB was not only concentration dependent, but also cellular type and vehicle kind dependent. At AmB concentrations of 50 mg.L-1, although the hemoglobin leakage for AmB-D was almost complete (99.51), for AmB-DAL and AmB-DL this value tended to zero. The p = 0.000 showed that AmB-D was significantly more hemolytic. Conclusion: The Fungizon™- Lipofundin™ admixtures seem to be the more valuable AmB carrier systems due to their best therapeutic index presented

Stability and Local Toxicity Evaluation of a Liposomal Prilocaine Formulation

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Nanotechnology-based Drug Delivery Systems for Dermatomycosis Treatment

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Amphotericin B Microemulsion Reduces Toxicity and Maintains the Efficacy as an Antifungal Product

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

AC Biosusceptometry as a Tool for Monitoring the Gastrointestinal Transit of Multiparticulate Drug Delivery System

Drug delivery systems based on natural polysaccharides, such as chitosan (CS) and pectin (PC), rather than on synthetic polymers, have been widely studied. Some reasons for that are low toxicity and costs and high biodegradability of the formers. A multiparticulate system based on CS and PC was developed in our laboratories, including the addition of an enteric polymer, cellulose acetate phtalate (CAP). Such improvement promoted stronger gastric and enteric resistances, as assessed in vitro, making the systems more selective to enzymatic degradation in the colon. Although in vitro dissolution tests can simulate some properties concerning the gastrointestinal transit (GT), collaborating to characterize the systems behavior in the biological fluids, frequently they do not result in satisfactory in vitro/in vivo correlations. The objective of this work was to follow in vivo the GT of the particles developed by means of AC biosusceptometry (ACB), a non-invasive and of low cost methodology. The particles containing ferrite in powder form were prepared by complex coacervation using an ideal 3:1:1 mass ratio for PC:CS:CAP. The magnetic particles were administered to healthy volunteers by oral route. The GT was monitored by using multi-sensor ACB system and the signal acquisition was performed every IS min until the colonic region was reached. By means of ACB technique, it was possible to acquiring images generated by the magnetic particles within the whole gastrointestinal tract including the colonic region. Variable particles transit times were observed among the volunteers, but without interference on the mapping of the particles until the colonic region. The particles were able to produce magnetic field strong enough to generate signals adequate for mapping the particles. The results suggest that integral particles reached the colon, after they resisted against gastric and enteric media. Studies associating transit time and in vivo drug release are in development in order to confirm the efficiency of the systems.

Electrochemical Reduction Using Glassy Carbon Electrode in Aqueous Medium of a Potential Anti-Chagas Drug: NFOH

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Protection of insulin-producing cells against toxicity of dexamethasone by catalase overexpression

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

REACTIVE OXYGEN GENERATION BY AZOMETHINE-H - A NEW ANTIMALARIAL DRUG

Superoxide radical (O-2(-)) is a free radical that may be involved in various toxic processes. Cu-Zn superoxide dismutase catalyzes the dismutation of the superoxide free radical and protects cells from oxidative damage. A rat bioassay validated for the identification of the toxic effects of azomethine H revealed increased serum activities of amylase, alanine transaminase, and alkaline phosphatase. The lipoperoxide and bilirubin concentrations were also increased in animals that received azomethine H (1 g/kg) from ascorbic or hydrochloric acid solutions. Azomethine H increased Cu-Zn superoxide dismutase activity. This elevation of Cu-Zn superoxide dismutase activity was highest on the 7th day and was at levels comparable with those of control rats from day 60 onwards. Superoxide is an important intermediate in the action and toxicity of azomethine H.

Aromatic antiepileptic drugs and mitochondrial toxicity: Effects on mitochondria isolated from rat liver

Idiosyncratic hepatotoxicity is a well-known complication associated with aromatic antiepileptic drugs (AAED), and it has been suggested to occur due to the accumulation of toxic arene oxide metabolites. Although there is clear evidence of the participation of an immune process, a direct toxic effect involving mitochondria dysfunction is also possible. The effects of AAED on mitochondrial function have not been studied yet. Therefore, we investigated, in vitro, the cytotoxic mechanism of carbamazepine (CB), phenytoin (PT) and phenobarbital (PB), unaltered and bioactivated, in the hepatic mitochondrial function. The murine hepatic microsomal system was used to produce the anticonvulsant metabolites. All the bioactivated drugs (CB-B, PB-B, PT-B) affected mitochondrial function causing decrease in state three respiration, RCR, ATP synthesis and membrane potential, increase in state four respiration as well as impairment of Ca(2+) uptake/release and inhibition of calcium-induced swelling. As an unaltered drug, only PB, was able to affect mitochondrial respiration (except state four respiration) ATP synthesis and membrane potential; however, Ca(2+) uptake/release as well as swelling induction were not affected. The potential to induce mitochondrial dysfunction was PT-B > PB-B > CB-B > PB. Results suggest the involvement of mitochondrial toxicity in the pathogenesis of AAED-induced hepatotoxicity. (C) 2008 Elsevier Ltd. All rights reserved.

Long-term toxicity following acute administration of nickel

Nickel compounds have high potential risk for the health of populations and for this reason their toxic effects should be urgently established. To determine the effect of nickel monosulfide in the muscle at the injection site on pancreatic, hepatic, and osteogenic lesions and the potential therapeutic effect of Cu-Zn superoxide dismutase (SOD), male Wistar rats received single intramuscular injections of nickel monosulfide (NiS - 7 mg Ni2+/Kg). A group of these experimental rats were injected intraperitoneally, with a single weekly dose of SOD covalently linked to polyethylene glycol (SOD-PEG). Rats were sacrificed at 2, 4, 6, and 8 months after Ni2+ injection. Nickel monosulfide produced tumors at the injection site. The increased phospholipid, alanine transaminase (ALT), alkaline phosphatase (ALP), and amylase levels in serum, in absence of SOD-PEG, reflected the toxic effects on pancreatic, hepatic, and osteogenic tissues of rats. SOD activity was increased in serum of rats receiving SOD-PEG throughout the experiment, and no significant difference was observed in biochemical parameters of control and experimental rats in presence of SOD- PEG. Superoxide radical generated by Ni2+ is of primary importance in the development of tumors at the injection site. Superoxide anion (O2 -) is also an important toxic intermediate with respect to hepatic, pancreatic, and osteogenic injury, since SOD-PEG has a potential therapeutic effect.