Iloprost Attenuates Oxidative Stress-Dependent Activation of Collagen Synthesis Induced by Sera from Scleroderma Patients in Human Pulmonary Microvascular Endothelial Cells.

Endothelial cell injury is an early event in systemic sclerosis (SSc) pathogenesis and several studies indicate oxidative stress as the trigger of SSc-associated vasculopathy. Here, we show that circulating factors present in sera of SSc patients increased reactive oxygen species (ROS) production and collagen synthesis in human pulmonary microvascular endothelial cells (HPMECs). In addition, the possibility that iloprost, a drug commonly used in SSc therapy, might modulate the above-mentioned biological phenomena has been also investigated. In this regard, as compared to sera of SSc patients, sera of iloprost-treated SSc patients failed to increased ROS levels and collagen synthesis in HPMEC, suggesting a potential antioxidant mechanism of this drug.

Protective Effect of Cyclically Pressurized Solid⁻Liquid Extraction Polyphenols from Cagnulari Grape Pomace on Oxidative Endothelial Cell Death.

The aim of this work is the evaluation of a green extraction technology to exploit winery waste byproducts. Specifically, a solid⁻liquid extraction technology (Naviglio Extractor®) was used to obtain polyphenolic antioxidants from the Cagnulari grape marc. The extract was then chemically characterized by spectrophotometric analysis, high-performance liquid chromatography, and mass spectrometry, revealing a total polyphenol content of 4.00 g/L ± 0.05, and the presence of anthocyanins, one of the most representative groups among the total polyphenols in grapes. To investigate potential biological activities of the extract, its ability to counteract hydrogen peroxide-induced oxidative stress and cell death was assessed in primary human endothelial cells. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test, used to assess potential extract cytotoxicity, failed to show any deleterious effect on cultured cells. Fluorescence measurements, attained with the intracellular reactive oxygen species (ROS) probe 2',7'-dichlorodihydrofluorescein diacetate (H2DCF-DA), revealed a strong antioxidant potential of the marc extract on the used cells, as indicated by the inhibition of the hydrogen peroxide-induced ROS generation and the counteraction of the oxidative-induced cell death. Our results indicate the Naviglio extraction, as a green technology process, can be used to exploit wine waste to obtain antioxidants which can be used to produce enriched foods and nutraceuticals high in antioxidants.

N- and S-homocysteinylation reduce the binding of human serum albumin to catechins.

PURPOSE: The dietary flavonoids epicatechin (EC), epigallocatechin (EGC), epicatechin gallate (ECG) and epigallocatechin gallate (EGCG) have been shown to interact with circulating albumin for transport in blood to different body tissues. This interaction may modulate their bioavailability and effectiveness. METHODS: Using affinity capillary electrophoresis to assess binding constants (K b), we investigated whether posttranslational modification of human serum albumin (HSA) through N- and S-homocysteinylation, commonly observed in hyperhomocysteinemia, may modify its interaction with catechins. RESULTS: S-Hcy HSA had lower Kb values toward EC (14 %), EGC (18 %), ECG (24 %) and EGCG (30 %). Similarly, N-Hcy HSA had lower Kb values toward EC (17 %), EGC (22 %), ECG (23 %) and EGCG (32 %). No differences were observed in the affinity between catechins, albumin and mercaptalbumin. CONCLUSION: Therefore, HSA posttranslational modifications typical of hyperhomocysteinemia reduce its affinity to catechins, potentially affecting their pharmacokinetics and availability at the active sites.

Protective Effect of Knee Postoperative Fluid on Oxidative-Induced Damage in Human Knee Articular Chondrocytes.

The oxidative-stress-elicited deterioration of chondrocyte function is the initial stage of changes leading to the disruption of cartilage homeostasis. These changes entail a series of catabolic damages mediated by proinflammatory cytokines, MMPs, and aggrecanases, which increase ROS generation. Such uncontrolled ROS production, inadequately balanced by the cellular antioxidant capacity, eventually contributes to the development and progression of chondropathies. Several pieces of evidence show that different growth factors, single or combined, as well as anti-inflammatory cytokines and chemokines, can stimulate chondrogenesis and improve cartilage repair and regeneration. In this view, hypothesizing a potential growth-factor-associated action, we investigate the possible protective effect of post-operation knee fluid from patients undergoing prosthesis replacement surgery against ROS-induced damage on normal human knee articular chondrocytes (HKACs). To this end, HKACs were pre-treated with post-operation knee fluid and then exposed to H2O2 to mimic oxidative stress. Intracellular ROS levels were measured by using the molecular probe H2DCFDA; cytosolic and mitochondrial oxidative status were assessed by using HKACs infected with lentiviral particles harboring the redox-sensing green fluorescent protein (roGFP); and cell proliferation was determined by measuring the rate of DNA synthesis with BrdU incorporation. Moreover, superoxide dismutase (SOD), catalase, and glutathione levels from the cell lysates of treated cells were also measured. Postoperative peripheral blood sera from the same patients were used as controls. Our study shows that post-operation knee fluid can counteract H2O2-elicited oxidative stress by decreasing the intracellular ROS levels, preserving the cytosolic and mitochondrial redox status, maintaining the proliferation of oxidatively stressed HKACs, and upregulating chondrocyte antioxidant defense. Overall, our results support and propose an important effect of post-operation knee fluid substances in maintaining HKAC function by mediating cell antioxidative system upregulation and protecting cells from oxidative stress.

Cytoprotective, antioxidant, and anti-migratory activity of Pistacia lentiscus L. supercritical carbon dioxide extract on primary human endothelial cells.

Green chemistry is a useful tool for producing valuable chemicals from biomass. However, extracted compounds need to be tested for safety and efficacy before their use in humans. Here we investigate the chemical composition and biological effects of a leaves Pistacia lentiscus L. supercritical carbon dioxide (SCCO2) extract. Terpenes represented the main extract fraction, with Germacrene D (11.18%), delta-cadinene (10.54%), and alpha-pinene (8.7%) the most abundant molecules. Challenged with endothelial cells (ECs), increasing extract concentrations failed to affect cell proliferation or promote cell toxicity. ROS assessment in unstressed and H2O2-treated ECs revealed an extract dose-dependent antioxidant activity. Exposition of H2O2-treated ECs to increasing extract concentrations dose-dependently counteracted H2O2-induced cell impairments. The extract significantly counteracted fetal calf serum-induced ECs migration. For the first time, we report that a SCCO2 extract obtained from PL leaves is safe on ECs and may be a useful source of valuable compounds with vasculoprotective properties.

NADPH-derived ROS generation drives fibrosis and endothelial-to-mesenchymal transition in systemic sclerosis: Potential cross talk with circulating miRNAs.

Systemic sclerosis (SSc) is an immune disorder characterized by diffuse fibrosis and vascular abnormalities of the affected organs. Although the etiopathology of this disease is largely unknown, endothelial damage and oxidative stress appear implicated in its initiation and maintenance. Here, we show for the first time that circulating factors present in SSc sera increased reactive oxygen species (ROS) production, collagen synthesis, and proliferation of human pulmonary microvascular endothelial cells (HPMECs). The observed phenomena were also associated with endothelial to mesenchymal transition (EndMT) as indicated by decreased von Willebrand factor (vWF) expression and increased alpha-smooth muscle actin, respectively, an endothelial and mesenchymal marker. SSc-induced fibroproliferative effects were prevented by HPMECs exposition to the NADPH oxidase inhibitor diphenyleneiodonium, demonstrating ROS's causative role and suggesting their cellular origin. Sera from SSc patients showed significant changes in the expression of a set of fibrosis/EndMT-associated microRNAs (miRNA), including miR-21, miR-92a, miR-24, miR-27b, miR-125b, miR-29c, and miR-181b, which resulted significantly upregulated as compared to healthy donors sera. However, miR29b resulted downregulated in SSc sera, whereas no significant differences were found in the expression of miR-29a in the two experimental groups of samples. Taking together our data indicate NADPH oxidase-induced EndMT as a potential mechanism of SSc-associated fibrosis, suggesting fibrosis-associated miRNAs as potentially responsible for initiating and sustaining the vascular alterations observed in this pathological condition.

Antioxidant Properties of Olive Mill Wastewater Polyphenolic Extracts on Human Endothelial and Vascular Smooth Muscle Cells.

This work aims to analyze the chemical and biological evaluation of two extracts obtained by olive mill wastewater (OMW), an olive oil processing byproduct. The exploitation of OMW is becoming an important aspect of development of the sustainable olive oil industry. Here we chemically and biologically evaluated one liquid (L) and one solid (S) extract obtained by liquid-liquid extraction followed by acidic hydrolysis (LLAC). Chemical characterization of the two extracts indicated that S has higher phenol content than L. Hydroxytyrosol and tyrosol were the more abundant phenols in both OMW extracts, with hydroxytyrosol significantly higher in S as compared to L. Both extracts failed to induce cell death when challenged with endothelial cells and vascular smooth muscle cells in cell viability experiments. On the contrary, the higher extract dosages employed significantly affected cell metabolic activity, as indicated by the MTT tests. Their ability to counteract H2O2-induced oxidative stress and cell death was assessed to investigate potential antioxidant activities of the extracts. Fluorescence measurements obtained with the reactive oxygen species (ROS) probe H2DCF-DA indicated strong antioxidant activity of the two OMW extracts in both cell models, as indicated by the inhibition of H2O2-induced ROS generation and the counteraction of the oxidative-induced cell death. Our results indicate LLAC-obtained OMW extracts as a safe and useful source of valuable compounds harboring antioxidant activity.

Flavin Oxidase-Induced ROS Generation Modulates PKC Biphasic Effect of Resveratrol on Endothelial Cell Survival.

BACKGROUND: Dietary intake of natural antioxidants is thought to impart protection against oxidative-associated cardiovascular diseases. Despite many in vivo studies and clinical trials, this issue has not been conclusively resolved. Resveratrol (RES) is one of the most extensively studied dietary polyphenolic antioxidants. Paradoxically, we have previously demonstrated that high RES concentrations exert a pro-oxidant effect eventually elevating ROS levels leading to cell death. Here, we further elucidate the molecular determinants underpinning RES-induced oxidative cell death. METHODS: Using human umbilical vein endothelial cells (HUVECs), the effect of increasing concentrations of RES on DNA synthesis and apoptosis was studied. In addition, mRNA and protein levels of cell survival or apoptosis genes, as well as protein kinase C (PKC) activity were determined. RESULTS: While high concentrations of RES reduce PKC activity, inhibit DNA synthesis and induce apoptosis, low RES concentrations elicit an opposite effect. This biphasic concentration-dependent effect (BCDE) of RES on PKC activity is mirrored at the molecular level. Indeed, high RES concentrations upregulate the proapoptotic Bax, while downregulating the antiapoptotic Bcl-2, at both mRNA and protein levels. Similarly, high RES concentrations downregulate the cell cycle progression genes, c-myc, ornithine decarboxylase (ODC) and cyclin D1 protein levels, while low RES concentrations display an increasing trend. The BCDE of RES on PKC activity is abrogated by the ROS scavenger Tempol, indicating that this enzyme acts downstream of the RES-elicited ROS signaling. The RES-induced BCDE on HUVEC cell cycle machinery was also blunted by the flavin inhibitor diphenyleneiodonium (DPI), implicating flavin oxidase-generated ROS as the mechanistic link in the cellular response to different RES concentrations. Finally, PKC inhibition abrogates the BCDE elicited by RES on both cell cycle progression and pro-apoptotic gene expression in HUVECs, mechanistically implicating PKC in the cellular response to different RES concentrations. CONCLUSIONS: Our results provide new molecular insight into the impact of RES on endothelial function/dysfunction, further confirming that obtaining an optimal benefit of RES is concentration-dependent. Importantly, the BCDE of RES could explain why other studies failed to establish the cardio-protective effects mediated by natural antioxidants, thus providing a guide for future investigation looking at cardio-protection by natural antioxidants.

Antioxidant Activity Mediates Pirfenidone Antifibrotic Effects in Human Pulmonary Vascular Smooth Muscle Cells Exposed to Sera of Idiopathic Pulmonary Fibrosis Patients.

Idiopathic pulmonary fibrosis (IPF) is a chronic lung disease characterized by an exacerbated fibrotic response. Although molecular and cellular determinants involved in the onset and progression of this devastating disease are largely unknown, an aberrant remodeling of the pulmonary vasculature appears to have implications in IPF pathogenesis. Here, we demonstrated for the first time that an increase of reactive oxygen species (ROS) generation induced by sera from IPF patients drives both collagen type I deposition and proliferation of primary human pulmonary artery smooth muscle cells (HPASMCs). IPF sera-induced cellular effects were significantly blunted in cells exposed to the NADPH oxidase inhibitor diphenyleneiodonium (DPI) proving the causative role of ROS and suggesting their potential cellular source. Contrary to IPF naive patients, sera from Pirfenidone-treated IPF patients failed to significantly induce both ROS generation and collagen synthesis in HPASMCs, mechanistically implicating antioxidant properties as the basis for the in vivo effect of this drug.

Oxidative stress-induced Akt downregulation mediates green tea toxicity towards prostate cancer cells.

Green tea consumption has been shown to possess cancer chemopreventive activity. Polyphenol E (PE) is a widely used standardized green tea extract formulation. This study was designed to investigate the impact of PE on prostate cancer cells (PC3), analyze the potential signals involved and elucidate whether anti- or pro-oxidant effects may be implicated. Treatment of PC3 cells with 30 and 100µg/ml PE significantly decreased cell viability and proliferation. At the tested concentrations, PE did not exert any antioxidant activity, eliciting instead a pro-oxidant effect at concentrations 30 and 100µg/ml, which was consistent with the observed PE cytotoxicity. PE-induced cell death was associated with mitochondrial dysfunction and downregulation of Akt activation, thus suggesting their implication in the PE-elicited cell dysfunction. Cell exposure to the ROS scavenger N-Acetyl Cysteine prevented PE-induced ROS increase, pAkt impairment, and cell death, clearly indicating the causative role of ROS in the observed phenomena. Failure of PE to induce PC3 damage in cells overexpressing Akt further confirms its implication in the PE-elicited cell death. Our findings showed an association between the antiproliferative and the pro-oxidant effect elicited by PE on PC3 cells and delineates a molecular signaling pattern potentially implicated in the toxicity of PE towards prostate cancer cells.

Human Serum Albumin Increases the Stability of Green Tea Catechins in Aqueous Physiological Conditions.

Epicatechin (EC), epigallocatechin (EGC), epicatechingallate (ECG) and epigallocatechingallate (EGCG) are antioxidants present in the green tea, a widely used beverage whose health benefits are largely recognized. Nevertheless, major physicochemical limitations, such as the high instability of catechins, pose important questions concerning their potential pharmacological use. Recent studies indicate that binding of catechins with plasmatic proteins may modulate their plasma concentration, tissue delivery and biological activity. After 5 minutes of incubation with HSA both ECG and EGCG were fully bound to HSA, while after 48h incubation only 41% of EC and 70% of EGC resulted linked. HSA had a strong stabilizing effect on all catechins, which could be found in solution between 29 and 85% even after 48h of incubation. In the absence of HSA, EGC and EGCG disappeared in less than 24h, while ECG and EC were found after 48h at 5 and 50%, respectively. The stabilizing effect of HSA toward EGCG, obtained in aqueous physiological conditions, resulted stronger in comparison to cysteine and HCl, previously reported to stabilize this polyphenol. Because of the multitude of contradictory data concerning in vivo and in vitro antioxidant-based experimentations, we believe our work may shed some light on this debated field of research.

Resveratrol alters human endothelial cells redox state and causes mitochondrial-dependent cell death.

Studies analyzing the impact of natural antioxidants (NA) on Endothelial Cells (ECs) have dramatically increased during the last years, since a deregulated ECs redox state is at the base of the onset and progression of several cardiovascular diseases. However, whether NA can provide cardiovascular benefits is still a controversial area of debate. Resveratrol (RES), a natural polyphenol found in grapes, is believed to provide cardiovascular benefits by virtue of its antioxidant effect on the endothelium. Here, we report that tissue-attainable doses of resveratrol increased the intracellular oxidative state, thus affecting mitochondrial membrane depolarization and inducing EC death. Cyclosporine A, a mitochondrial permeability transition pore inhibitor, prevented oxidative-mediated cell death, thus implicating mitochondria in resveratrol-induced EC impairment. The specific cytochrome P450 (CYP) 2C9 inhibitor, sulfaphenazole, counteracted both oxidative stress and mitochondrial membrane depolarization, providing EC protection against resveratrol-elicited pro-oxidant effects. Our findings strongly suggest that CYP2C9 mediates resveratrol-induced oxidative stress leading to mitochondria impairment and EC death.

NAMI-A inhibits the PMA-induced ODC gene expression in ECV304 cells: involvement of PKC/Raf/Mek/ERK signalling pathway.

Imidazolium trans-imidazole dimethyl sulfoxide tetrachlororuthenate (NAMI-A) is a new compound active against lung metastasis of solid metastasizing tumours. While its in vivo effect has been studied, the molecular insights that underlie its action are largely unknown. Among the possible pathways responsible for malignant transformation, PKC arose as one of the most promising targets for new antineoplastic drugs. We demonstrated the capability of NAMI-A of inhibiting PMA induced-PKC activity in ECV304 in a dose-dependent fashion. Furthermore, NAMI-A through modulation of PKC activity has been proved capable of reducing the phorbol ester induced expression of ornithine decarboxilase (ODC) gene and to abrogate the activation of the Raf/MEK/ERK pathway. Taken together these results suggest that many of the in vivo outcomes of NAMI-A treatment may be the result of a direct action on PKC.

Evaluation of non-covalent interactions between serum albumin and green tea catechins by affinity capillary electrophoresis.

The natural antioxidant-associated biological responses appear contradictory since biologically active dosages registered in vitro experiments are considerably higher if compared to concentrations found in vivo. The recent research indicates that natural antioxidants, including the major catechins of green tea epicatechin (EC), epigallocatechin (EGC), epicatechingallate (ECG) and epigallocatechingallate (EGCG) form non-covalent complexes with albumin, a crucial aspect that may modulate their plasma concentration, tissue delivery and biological activity. Affinity capillary electrophoresis (ACE) was used to characterize the binding of the four catechins to human serum albumin (HSA) and bovine serum albumin (BSA) at near-physiological conditions: 10 mmol/L phosphate buffer, HEPES 50 mmol/L (pH 7.5), temperature 37°C. The studied flavonoids displayed affinities toward the albumin with binding constants in the range 10(3)-10(5)M(-1), with a greater affinity of catechins toward HSA than BSA (between 3 and 3.5 fold higher). We also confirmed that catechins having a galloyl moiety (ECG and EGCG) have a higher binding affinity toward albumin than the catechins lacking the galloyl moiety (EC and EGC), and that for both albumins the order of affinity is EC

Oxidative stress-dependent activation of collagen synthesis is induced in human pulmonary smooth muscle cells by sera from patients with scleroderma-associated pulmonary hypertension.

Pulmonary arterial hypertension is a major complication of systemic sclerosis. Although oxidative stress, intima hyperplasia and a progressive vessel occlusion appear to be clearly involved, the fine molecular mechanisms underpinning the onset and progression of systemic sclerosis-associated pulmonary arterial hypertension remain largely unknown. Here we shows for the first time that an increase of NADPH-derived reactive oxygen species production induced by sera from systemic sclerosis patients with pulmonary arterial hypertension drives collagen type I promoter activity in primary human pulmonary artery smooth muscle cells, suggesting that antioxidant-based therapies should be considered in the treatment of systemic sclerosis-associated vascular diseases.

Different redox response elicited by naturally occurring antioxidants in human endothelial cells.

Evidences that higher natural antioxidant (NA) intake provides protection against cardiovascular disease (CVD) are contradictory. Oxidative-induced endothelial cells (ECs) injury is the key step in the onset and progression of CVD and for this reason the cellular responses resulting from NA interaction with ECs are actively investigated. This study was designed to investigate the direct impact of different naturally occurring antioxidants on the intracellular ROS levels in cultured human ECs. NA-induced redox changes, in terms of modulation of the intracellular ROS levels, were assessed by using the ROS fluorescent probe 2',7'-dichlorodihydrofluorescein diacetate (H2DCF-DA). While caffeic and caftaric acid exerted an anti-oxidant effect, both coumaric acid and resveratrol were pro-oxidant. Anti- and pro-oxidant effects of the tested compounds were concentration dependent, showing the induction or the tendency to promote a pro-oxidant outcome with increasing concentrations. Interestingly, the anti- and pro-oxidant behavior of chlorogenic and ferulic acid was dependent on the basal intracellular redox state. Our data indicate that naturally occurring antioxidants are able to induce a rapid modification of the intracellular ROS levels in human ECs, which is dependent on both the applied concentration and the intracellular redox state.

Coumaric acid induces mitochondrial damage and oxidative-mediated cell death of human endothelial cells.

Evidence that higher natural antioxidants (NA) intake provides cardiovascular protection is contradictory. The endothelium plays a pivotal role in cardiovascular homeostasis, and for this reason, the molecular events resulting from the interaction of NA with endothelial cells (ECs) are actively investigated. Here, we show that moderately high doses of coumaric acid (CA) induced intracellular reactive oxygen species (ROS) production, mitochondrial membrane depolarization and ECs death. Treatment of ECs with cyclosporine A, a mitochondrial permeability transition pore inhibitor, prevented the oxidative-mediated cell damage indicating mitochondrial involvement in CA-induced ECs impairment. CA-induced intracellular ROS generation was counteracted by the specific cytochrome P450 (CYP) 2C9 inhibitor sulfaphenazole (SPZ). SPZ also prevented CA-induced mitochondrial membrane depolarization and ECs death, implicating CYP2C9 in mediating the cellular response upon CA treatment. Our results indicate that moderately high doses of CA can promote CYP2C9-mediated oxidative stress eliciting mitochondrial-dependent ECs death and may pave the way toward mechanistic insight into NA effects on cardiovascular cells.

Apricot melanoidins prevent oxidative endothelial cell death by counteracting mitochondrial oxidation and membrane depolarization.

The cardiovascular benefits associated with diets rich in fruit and vegetables are thought to be due to phytochemicals contained in fresh plant material. However, whether processed plant foods provide the same benefits as unprocessed ones is an open question. Melanoidins from heat-processed apricots were isolated and their presence confirmed by colorimetric analysis and browning index. Oxidative injury of endothelial cells (ECs) is the key step for the onset and progression of cardiovascular diseases (CVD), therefore the potential protective effect of apricot melanoidins on hydrogen peroxide-induced oxidative mitochondrial damage and cell death was explored in human ECs. The redox state of cytoplasmic and mitochondrial compartments was detected by using the redox-sensitive, fluorescent protein (roGFP), while the mitochondrial membrane potential (MMP) was assessed with the fluorescent dye, JC-1. ECs exposure to hydrogen peroxide, dose-dependently induced mitochondrial and cytoplasmic oxidation. Additionally detected hydrogen peroxide-induced phenomena were MMP dissipation and ECs death. Pretreatment of ECs with apricot melanoidins, significantly counteracted and ultimately abolished hydrogen peroxide-induced intracellular oxidation, mitochondrial depolarization and cell death. In this regard, our current results clearly indicate that melanoidins derived from heat-processed apricots, protect human ECs against oxidative stress.

Prune melanoidins protect against oxidative stress and endothelial cell death.

The health-promoting effects of fruit and vegetable consumption are thought to be due to phytochemicals contained in fresh plant material. Whether processed plant foods provide the same benefits as unprocessed ones is an open question. Melanoidins from heat-processed plums (prunes) were isolated and their presence confirmed by hydroxymethylfurfural content and browning index. Oxidative-induced endothelial cell (EC) damage is the trigger for the development of cardiovascular diseases (CVD); therefore the potential protective effect of prune melanoidins on hydrogen peroxide-induced oxidative cell damage was investigated on human endothelial ECV304 cells. Cytoplasmic and mitochondrial redox status was assessed by using the novel, redox-sensitive, ratiometric fluorescent protein sensor (roGFP), while mitochondrial membrane potential (MMP) was investigated with the fluorescent dye, JC-1. Treatment of ECV304 cells with hydrogen peroxide dose-dependently induced both mitochondrial and cytoplasmic oxidation, in addition to MMP dissipation, with ensuing cell death. Pretreatment of ECV304 with prune melanoidins, significantly counteracted and ultimately abolished hydrogen peroxide elicited phenomena, clearly indicating that these polymers protect human EC against oxidative stress.

Novel docetaxel-loaded nanoparticles based on poly(lactide-co-caprolactone) and poly(lactide-co-glycolide-co-caprolactone) for prostate cancer treatment: formulation, characterization, and cytotoxicity studies.

Docetaxel (Dtx) chemotherapy is the optional treatment in patients with hormone-refractory metastatic prostate cancer, and Dtx-loaded polymeric nanoparticles (NPs) have the potential to induce durable clinical responses. However, alternative formulations are needed to overcome the serious side effects, also due to the adjuvant used, and to improve the clinical efficacy of the drug.In the present study, two novel biodegradable block-copolymers, poly(lactide-co-caprolactone) (PLA-PCL) and poly(lactide-co-caprolactone-co-glycolide) (PLGA-PCL), were explored for the formulation of Dtx-loaded NPs and compared with PLA- and PLGA-NPs. The nanosystems were prepared by an original nanoprecipitation method, using Pluronic F-127 as surfactant agent, and were characterized in terms of morphology, size distribution, encapsulation efficiency, crystalline structure, and in vitro release. To evaluate the potential anticancer efficacy of a nanoparticulate system, in vitro cytotoxicity studies on human prostate cancer cell line (PC3) were carried out. NPs were found to be of spherical shape with an average diameter in the range of 100 to 200 nm and a unimodal particle size distribution. Dtx was incorporated into the PLGA-PCL NPs with higher (p < 0.05) encapsulation efficiency than that of other polymers. Differential scanning calorimetry suggested that Dtx was molecularly dispersed in the polymeric matrices. In vitro drug release study showed that release profiles of Dtx varied on the bases of characteristics of polymers used for formulation. PLA-PCL and PLGA-PCL drug loaded NPs shared an overlapping release profiles, and are able to release about 90% of drug within 6 h, when compared with PLA- and PLGA-NPs. Moreover, cytotoxicity studies demonstrated advantages of the Dtx-loaded PLGA-PCL NPs over pure Dtx in both time- and concentration-dependent manner. In particular, an increase of 20% of PC3 growth inhibition was determined by PLGA-PCL NPs with respect to free drug after 72 h incubation and at all tested Dtx concentration. In summary, PLGA-PCL copolymer may be considered as an attractive and promising polymeric material for the formulation of Dtx NPs as delivery system for prostate cancer treatment, and can also be pursued as a validated system in a more large context.