Development of a polymer system for the delivery of daunorubicin to tumor cells to overcome drug resistance

Method for the synthesis of polymeric nanoparticles (NP) with encapsulated daunorubicin (DNR) was developed on the basis of double emulsion solvent evaporation technique using biodegradable poly(lactide-co-glycolide) (PLGA), which is aimed at customization of pharmacokinetic properties of the preparation, enhanced accumulation of DNR in tumor cells and prolongation of its action. The obtained polymer nanoparticles (DNR-PLGA) had average size ranging around 138±36 nm, with zeta-potential of –25.3 mV and the polydispersity index (PDI) of 0.072. The release kinetics of DNR from polymer nanoparticles at pH 7.4 and 5.0 has been studied. In vitro studies showed similar specific activity of DNR- PLGA in K562 and MCF-7 cancer cell lines together with an increase in activity in K562 Adr and MCF-7 Adr cell lines, which are anthracycline resistant, by 1.6 and 3.4 times. The study demonstrated the efficacy of the developed PLGA-based DNR delivery system in the improvement of antitumor effect of DNR, overcoming multidrug resistance in cancer cells, and also in the decrease in nonspecific toxicity of the preparation.     

Interaction of an anthracycline disaccharide with ctDNA: Investigation by spectroscopic technique and modeling studies

This study was designed to examine the interaction of an anthracycline disaccharide, 4′-O-(β-L-oleandrosyl) daunorubicin (DNR–D2), with calf thymus deoxyribonucleic acid (ctDNA) by UV–vis in combination with fluorescence spectroscopy and molecular modeling techniques under physiological conditions (Britton–Robinson buffer solutions, pH 7.4). By the analysis of UV–vis and fluorescence spectrum, it was observed that the binding mode between DNR–D2 and ctDNA might be intercalation, and fluorescence quenching mechanism of DNR–D2 by ctDNA was a static quenching type. Upon binding to ctDNA, the anthraquinone chromophore of DNR–D2 could slide into the C–G rich region of ctDNA. Hydrogen bonding forces may play an essential role in the binding of DNR–D2 to ctDNA. Furthermore, the results obtained from computational modeling corroborated the experimental results obtained from spectroscopic investigations. These studies are valuable for a better understanding the datailed mode of DNR–D2–DNA interaction, which should be important in deeper insight into the therapeutic efficiency of DNR–D2.     

One-Pot Green Synthesis of Hematite (α-Fe<Subscript>2</Subscript>O<Subscript>3</Subscript>) Nanoparticles by Ultrasonic Irradiation and Their In Vitro Cytotoxicity on Human Keratinocytes CRL-2310

Greener and ecofriendly approaches to the synthesis of hematite (α-Fe₂O₃) nanoparticles are important for various biomedical applications. The authors describe on a facile, one-pot method for synthesizing hematite nanoparticles (HNPs) using ultrasonic irradiation of iron(III) oxide solution containing the aqueous root extract of Arisaema amurense, which was used as both reducing and stabilizing agents. The synthesized HNPs were characterized by X-ray diffraction (XRD), Fourier transform infrared (FT-IR) spectroscopy, thermogravimetric analysis (TGA), scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDS), transmission electron microscopy (TEM), and vibrating sample magnetometry (VSM). FT-IR analysis indicated the presence of stabilizing groups such as hydroxyl (–OH), C–O, and amide (–NH–) on the surfaces of HNPs. TEM analysis revealed the formation of near spherical HNPs of average size 24.55 ± 6.9 nm. VSM confirmed the ferromagnetic nature of the HNPs with a saturation magnetization (Ms) at 1.25 emu/g and remanent magnetization (Mr) at 0.50 emu/g at 301 K. The electrochemical behavior of glassy carbon (GC)/HNPs electrode was studied using cyclic voltammetry (CV). MTT assays of the HNPs exhibited in vitro concentration-dependent cytotoxicity to human keratinocytes CRL-2310, which indicated the synthesized HNPs are compatible with requirements for in vivo biomedical applications at lower concentrations.     

A novel aptamer functionalized CuInS2 quantum dots probe for daunorubicin sensing and near infrared imaging of prostate cancer cells

In this paper, a novel daunorubicin (DNR)-loaded MUC1 aptamer-near infrared (NIR) CuInS₂ quantum dot (DNR–MUC1–QDs) conjugates were developed, which can be used as a targeted cancer imaging and sensing system. After the NIR CuInS₂ QDs conjugated with the MUC1 aptamer–(CGA)₇, DNR can intercalate into the double-stranded CG sequence of the MUC1–QDs. The incorporation of multiple CG sequences within the stem of the aptamers may further increase the loading efficiency of DNR on these conjugates. DNR–MUC1–QDs can be used to target prostate cancer cells. We evaluated the capacity of MUC1–CuInS₂ QDs for delivering DNR to cancer cells in vitro, and its binding affinity to MUC1-positive and MUC1-negative cells. This novel aptamer functionalized QDs bio-nano-system can not only deliver DNR to the targeted prostate cancer cells, but also can sense DNR by the change of photoluminescence intensity of CuInS₂ QDs, which concurrently images the cancer cells. The quenched fluorescence intensity of MUC1–QDs was proportional to the concentration of DNR in the concentration ranges of 33–88 nmol L⁻¹. The detection limit (LOD) for DNR was 19 nmol L⁻¹. We demonstrate the specificity and sensitivity of this DNR–MUC1–QDs probe as a cancer cell imaging, therapy and sensing system in vitro.     

Same-single-cell analysis using the microfluidic biochip to reveal drug accumulation enhancement by an amphiphilic diblock copolymer drug formulation

Multidrug resistance (MDR) is one of the major obstacles in drug delivery, and it is usually responsible for unsuccessful cancer treatment. MDR may be overcome by using MDR inhibitors. Among different classes of these inhibitors that block drug efflux mediated by permeability-glycoprotein (P-gp), less toxic amphiphilic diblock copolymers composed of methoxypolyethyleneglycol-block-polycaprolactone (MePEG-b-PCL) have been studied extensively. The purpose of this work is to evaluate how these copolymer molecules can reduce the efflux, thereby enhancing the accumulation of P-gp substrates (e.g., daunorubicin or DNR) in MDR cells. Using conventional methods, it was found that the low-molecular-weight diblock copolymer, MePEG₁₇-b-PCL₅ (PCL5), enhanced drug accumulation in MDCKII-MDR1 cells, but the high-molecular-weight version, MePEG₁₁₄-b-PCL₂₀₀ (PCL200), did not. However, when PCL200 was mixed with PCL5 (and DNR) in order to encapsulate them to facilitate drug delivery, there was no drug enhancement effect attributable to PCL5, and the reason for this negative result was unclear. Since drug accumulation measured on different cell batches originated from single cells, we employed the same-single-cell analysis in the accumulation mode (SASCA-A) to find out the reason. A microfluidic biochip was used to select single MDR cells, and the accumulation of DNR was fluorescently measured in real time on these cells in the absence and presence of PCL5. The SASCA-A method allowed us to obtain drug accumulation information faster in comparison to conventional assays. The SASCA-A results, and subsequent curve-fitting analysis of the data, have confirmed that when PCL5 was encapsulated in PCL200 nanoparticles as soon as they were synthesized, the ability of PCL5 to enhance DNR accumulation was retained, thus suggesting PCL200 as a promising delivery system for encapsulating P-gp inhibitors, such as PCL5. Graphical Abstract

?     

Layered microcapsules for daunorubicin loading and release as well as in vitro and in vivo studies

Anti‐cancer drug daunorubicin (DNR) was encapsulated in preformed multilayer microcapsules and was applied in tumor treatment by in vitro cell culture and in vivo animal experiments. The microcapsules were fabricated by an alternate deposition of oppositely charged polysaccharides, i.e. chitosan and alginate onto carboxymethyl cellulose (CMC) doped CaCO₃ colloidal particles in a sequential assembly procedure, followed by crosslinking of the capsule shells with glutaraldehyde (GA) and removal of the templates by disodium ethylenediaminetetraacetic acid (EDTA). The as‐prepared microcapsules showed strong ability to induce the positively charged DNR to deposit into the microcapsule interiors. Confocal microscopy and transmission electron microscopy observed homogeneous distribution of the drug within microcapsules. The loaded DNR could be released again, following a diffusion‐controlled model at the initial stage. In vitro experiments demonstrated that the encapsulated DNR can effectively induce the apoptosis of BEL‐7402 tumor cells, as evidenced by various microscopy techniques after acridine orange (AO), Hoechst 33342, and osmium tetraoxide staining. By seeding the BEL‐7402 hepatoma cells into BALB/c/nu mice, tumors were created for the animal experiments. The results showed that the encapsulated DNR had better efficacy than that of the free drug in terms of tumor inhibition in a 4 week in vivo culture period. Copyright © 2007 John Wiley & Sons, Ltd.     

Real-time detection of the interaction between anticancer drug daunorubicin and cancer cells by Au-MCNT nanocomposites modified electrodes

In this study,we have prepared the blending of gold nanoparticles-multiwalled nanotubes (Au-MCNTs) and then applied the new nanocomposites to modify the glassy carbon electrode (GCE) for highly sensitive detection of the interaction between anticancer drug daunorubicin (DNR) and cancer cells. Electrochemical analysis indicates that the Au-MCNT modified GCE shows high sensitivity and could track the real time response of cancer cells under DNR treatments. Therefore,this new nano-interface and Au-MCNT modifie...     

Calcium Phosphate Hybrid Nanoparticles: Self‐Assembly Formation,Characterization, and Application as an Anticancer Drug Nanocarrier

Calcium phosphate hybrid nanoparticles (CaP‐HNPs) have been synthesized in aqueous solution through self‐assembly by using two oppositely charged polyelectrolytes (poly(diallyldimethylammonium chloride) (PDADMAC) and poly(acrylate sodium) (PAS)) as dual templates. First, the PAS/Ca²⁺ and PDADMAC/PO₄^3? complexes form through electrostatic interactions and then two complexes self‐assemble into CaP‐HNPs after mixing them together. The as‐prepared CaP‐HNPs exhibit a spherical morphology with a narrow size distribution, good dispersibility, and high colloidal stability in water. The CaP‐HNPs are explored as a nanocarrier for the anticancer drug docetaxel (Dtxl). The CaP‐HNPs show excellent biocompatibility, high drug‐loading capacity, pH‐sensitive drug‐release behavior, and high anticancer effect after being loaded with Dtxl. Therefore, the as‐prepared CaP‐HNPs are promising drug nanocarriers for cancer therapy.     

Efficient intracellular delivery of camptothecin by silica/titania hollow nanoparticles

Silica/titania hollow nanoparticles (HNPs) with 50?nm were fabricated by using the dissolution and redeposition method and modified with anti-[human epidermal growth factor receptor 2] monoclonal antibody (herceptin), and their application as camptothecin (CPT) delivery agents to human breast cancer SK-BR-3 cells was investigated. Although the diameter of herceptin-modified HNPs (HER-HNP) is smaller than that of other reported mesoporous silica nanoparticles, the extensive hollow cavity of HNPs (ca. 30?nm) allowed the loading of a large amount of CPT. CPT-loaded HER-HNP (HER-HNP-CPT) did not release CPT in phosphate-buffered saline over a period of 24?h, however, HER-HNP-CPT in a hydrophobic solvent released its entire load of CPT. In addition, HER-HNPs were efficiently internalized owing to their herceptin conjugation and optimized size. To evaluate in vitro antitumor efficacy, apoptosis/necrosis and viability of HER-HNP-CPT-treated cells were investigated. When the cells were treated with HER-HNP-CPT for 30?min, a few apoptotic cells were observed. After 24?h, the viability of HER-HNP-CPT-treated SK-BR-3 decreased to 60?%, which revealed highly efficient chemotherapy. However, CPT loaded into HNP or HER-HNP had no significant effects on the viability of macrophages. Judging from these data, HER-HNPs are very suitable for application in anticancer therapy. A HER-HNP-CPT drug delivery system offers a new direction for a hydrophobic anticancer drug carrier and can be expanded to practical applications with further studies.     

Simultaneous determination of cytosine arabinoside, its nucleotides and metabolites by ion-pair high-performance liquid chromatography

Currently available high-performance liquid chromatographic assays for cytosine arabinoside (ara-C) and its metabolites suffer from two major shortcomings: inability to resolve both ara-C and its nucleotides in a single chromatographic step and/or inadequate sensitivity to allow quantitation of intracellular cytosine arabinofuranoside-5'-triphosphate (ara-CTP) without the use of radiolabelled drug. In this paper, we describe a new ion-pairing high-performance liquid chromatographic assay for ara-C in biological samples that can separate ara-C from its nucleotides, metabolites, and naturally occurring ribonucleotides in a single chromatographic step with a lower limit of quantitation of 5 pmol for ara-C and 10 pmol for ara-CTP. Examples of the utility of this assay are shown in studies of intracellular pharmacokinetics of ara-C in cultured human breast cancer cells and in analysis of plasma nucleoside levels in patients receiving high-dose thymidine chemotherapy. We conclude that this assay provides a rapid and versatile system that can be applied to the study of both cellular and plasma nucleoside pharmacokinetics.     

β-环糊精修饰电极对柔红霉素的电化学定量测定

柔红霉素(DNR)是强效的抗白血病制剂,但具有心脏毒性,基因毒性和生殖器官毒性。因此,研究柔红霉素的用量,建立多种微量检测方法,在临床医学方面具有积极的和现实的意义。本文研究了DNR在β-环糊精（β-CD）修饰金电极上的电化学行为,发现DNR能在β-CD俢饰电极上发生包络反应,其包络常数为6.1×104 L•mol-1。而且,只有包络的DNR能进行不可逆的电化学反应。其速率常数为28.03 s-1。在3-30 μmol• L-1的浓度范围内,DNR还原峰峰电流与DNR浓度成线性关系,因此,β-CD修饰电极可用于DNR的定量检测。在pH=7.0的中性电解液和温度接近40℃时,DNR在β-CD修饰电极上有最好的电化学活性。此时,β-CD修饰电极对DNR有较低的检出极限。     

纳米金-多壁碳纳米管复合物修饰电极对柔红霉素作用癌细胞的实时检测

本文通过制备纳米金-多壁碳纳米管复合物(Au-MCNT)修饰玻碳电极,建立了抗癌药物柔红霉素(DNR)作用癌细胞的高灵敏检测方法,研究并追踪了DNR与癌细胞相互作用过程中细胞对DNR的电化学实时响应.结果表明,Au-MCNT修饰电极能实现抗癌药物DNR作用癌细胞的高灵敏实时检测.基于该纳米复合材料的电化学药物传感分析方法可作为抗癌研究中一种方便、快捷、灵敏的实时检测手段,在生物医学等领域具有良好的应用前景.     

Novel synthetic route for covalent coupling of biomolecules on super‐paramagnetic hybrid nanoparticles

The immobilization of biomolecules on magnetic nanoparticles is an issue with high potential in different fields. We describe herein a new strategy to immobilize biomolecules on super‐paramagnetic nanoparticles based on the reactivity of vinyl sulfone groups with naturally occurring functional groups present in biomolecules (amine and thiol). A new monomer containing a polymerizable methacryloyl group and a secondary amine group was synthesized and used to prepare super‐paramagnetic hybrid nanoparticles (SP‐HNPs) by two‐step miniemulsion polymerization. The Michael addition reaction of divinyl sulfone (DVS) to the secondary amine groups localized on the nanoparticles surface allows the introduction of the vinyl sulfone function in the SP‐HNPs (SP‐HNPs‐VS). The morphology of the functionalized SP‐HNPs was characterized by transmission electron microscopy (TEM), high‐resolution transmission electron microscopy (HRTEM), dynamic light scattering, and magnetic susceptibility. The capacity of SP‐HNPs‐VS for the immobilization of biomolecules was evaluated with three model proteins: avidin, invertase, and horseradish peroxidase (HRP). The model proteins were successfully immobilized in mild aqueous conditions compatible with the biological nature of the enzymes. © 2012 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem, 2012     

A label-free method for identifying electroactive G-quadruplex-binding ligand

Ligands of G-quadruplex have received great attention because of their potential anticancer activity. Here, we designed a label-free electrochemical method to identify electrochemically active molecules that stabilize G-quadruplexes in homogeneous medium. Daunorubicin (DNR) was used as model drug. In the presence of GDNA, the voltammetry of DNR changed. The binding constant (β) and binding-site number (m) of GDNA – mDNR are simultaneously obtained from dependence of the current on the amount of added GDNA in voltammetry, concluding that β = (2.37 ± 2.25) × 10⁷ M^? 1and m was near 1, respectively. Control experiment was performed, indicating that this method can specific identify G-quadruplex-binding ligand. The interaction mechanism of DNR with GDNA was also investigated by circular dichroism (CD). The result demonstrated that it is a simple and selective method for identification electroactive ligand with potential anticancer activity.     

Possibility to Biotransform Anthracyclines by Peroxidases Produced by Bjerkandera adusta CCBAS 930 with Reduction of Geno- and Cytotoxicity and Pro-Oxidative Activity

The aim of this study was to evaluate the bioremoval mechanism of anthracycline antibiotics by the white-rot fungus B. adusta CCBAS 930. The activity of oxidoreductases and levels of phenolic compounds and free radicals were determined during the biotransformation of anthraquinone antibiotics: daunomycin (DNR) and doxorubicin (DOX) by B. adusta strain CCBAS 930. Moreover, phytotoxicity (Lepidium sativum L.), ecotoxicity (Vibrio fischeri), genotoxicity and cytotoxicity of anthraquinone dyes were evaluated before and after biological treatment. More than 80% and 90% of DNR and DOX were removed by biodegradation (decolorization). Initial solutions of DNR and DOX were characterized by eco-, phyto-, geno- and cytotoxicity. Despite efficient decolorization, secondary metabolites, toxic to bacteria, formed during biotransformation of anthracycline antibiotics in B. adusta CCBAS 930 cultures. DNR and DOX metabolites did not increase reactive oxygen species (ROS) production in human fibroblasts and resazurin reduction. DNR metabolites did not change caspase-3 activity.     

Silica-PMMA hairy nanoparticles prepared via phase transfer-assisted aqueous miniemulsion atom transfer radical polymerization

Hairy nanoparticles (HNPs) constitute a class of hybrid nanocomposites that are resistant to aggregation and agglomeration, although the green, large-scale synthesis of HNPs remains a challenge. In this work, 25 nm-diameter silica-core HNPs with a poly(methyl methacrylate) (PMMA) shell were synthesized using a graft-from approach in aqueous miniemulsion, employing atom transfer radical polymerization with activators regenerated by electron transfer (ARGET-ATRP). In particular, this work used tetrabutylammonium bromide (TBAB)-assisted phase transfer of monomer, markedly improving upon earlier methods by showing that phase transfer could take place in the absence of organic solvents. Furthermore, syntheses with selected monomer addition rates produced HNP graft densities ranging from 0.011 to 0.017 chains/nm² and shell thicknesses ranging from 2.5 to 11 nm. Finally, analysis of reaction kinetics revealed that shell growth reached completion in as little as 2 hr, confirmed by the synthesis of >1 g of PMMA-shell HNPs in a reduced timeframe.     

Unconventional Route to Uniform Hollow Semiconducting Nanoparticles with Tailorable Dimensions,Compositions, Surface Chemistry,and Near‐Infrared Absorption

Despite impressive recent advances in the synthesis of lead chalcogenide solid nanoparticles, there are no examples of lead chalcogenide hollow nanoparticles (HNPs) with controlled diameter and shell thickness as current synthetic approaches for HNPs have inherent limitations associated with their complexity, inability to precisely control the dimensions, and limited possibilities with regard to applicable materials. Herein, we report on an unconventional strategy for crafting uniform lead chalcogenide (PbS and PbTe) HNPs with tailorable size, surface chemistry, and near‐IR absorption. Amphiphilic star‐like triblock copolymers [polystyrene‐block‐poly(acrylic acid)‐block‐polystyrene and polystyrene‐block‐poly(acrylic acid)‐block‐poly(3,4‐ethylenedioxythiophene)] were rationally synthesized and exploited as nanoreactors for the formation of uniform PbS and PbTe HNPs. Compared to their solid counterparts, the near‐IR absorption of the HNPs is blue‐shifted owing to the hollow interior. This strategy can be readily extended to other types of intriguing low‐band‐gap HNPs for diverse applications.     

Unconventional Route to Uniform Hollow Semiconducting Nanoparticles with Tailorable Dimensions,Compositions, Surface Chemistry,and Near‐Infrared Absorption

Despite impressive recent advances in the synthesis of lead chalcogenide solid nanoparticles, there are no examples of lead chalcogenide hollow nanoparticles (HNPs) with controlled diameter and shell thickness as current synthetic approaches for HNPs have inherent limitations associated with their complexity, inability to precisely control the dimensions, and limited possibilities with regard to applicable materials. Herein, we report on an unconventional strategy for crafting uniform lead chalcogenide (PbS and PbTe) HNPs with tailorable size, surface chemistry, and near‐IR absorption. Amphiphilic star‐like triblock copolymers [polystyrene‐block‐poly(acrylic acid)‐block‐polystyrene and polystyrene‐block‐poly(acrylic acid)‐block‐poly(3,4‐ethylenedioxythiophene)] were rationally synthesized and exploited as nanoreactors for the formation of uniform PbS and PbTe HNPs. Compared to their solid counterparts, the near‐IR absorption of the HNPs is blue‐shifted owing to the hollow interior. This strategy can be readily extended to other types of intriguing low‐band‐gap HNPs for diverse applications.     

Size-tunable silicon/iron oxide hybrid nanoparticles with fluorescence, superparamagnetism, and biocompatibility

Magnetic/fluorescent composite materials have become one of the most important tools in the imaging modality in vivo using magnetic resonance imaging (MRI) monitoring and fluorescence optical imaging. We report herein on a simplified procedure to synthesize hybrid nanoparticles (HNPs) that combine silicon and magnetic iron oxides consisting of magnetite (Fe(3)O(4)) and maghemite (γ-Fe(2)O(3)). Intriguingly, our unique synthetic approach can control magnetic and optical behaviors by reducing the particle size, demonstrating that the HNPs with the mean diameter of 3.0 nm exhibit superparamagnetic behavior and green fluorescence in an aqueous solution, ambient air, and a cellular environment, whereas the HNPs with the mean diameter more than 5.0 nm indicate ferromagnetic behavior without fluorescence. Additionally, both HNPs with different diameters possess excellent magnetic responsivity for external applied magnetic field and good biocompatibility due to the low cytotoxicity. Our biocompatible HNPs with the superparamagnetism can provide an attractive approach for diagnostic imaging system in vivo.     

Study of human neutrophil peptides in saliva by matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry

Matrix‐assisted laser desorption/ionization time‐of‐flight (MALDI‐TOF) mass spectrometry is used to rapidly characterize the human neutrophil peptides – HNP 1, 2, and 3 – in saliva. The saliva excreted from the parotid and sublingual/submandibular glands of 70 individuals were collected and examined using MALDI‐TOF. The MALDI approach requires no sample pretreatment other than mixing the saliva‐absorbing material with the matrix and drying under ambient conditions. Tissue paper was the best material for collecting the saliva samples because of its strong texture and high absorbance, and sinapinic acid was the best MALDI matrix for the analysis of the HNPs. HNPs were detected in almost all the samples collected from the parotid glands, with no obvious differences among age or gender. In contrast, the distribution of the HNPs in the samples collected from the sublingual/submandibular glands was age‐dependent: no HNPs were detected for those collected from individuals younger than 30, but the HNPs were present in all of the samples collected from those older than 60 years. The increased probability of detecting saliva HNPs with age suggests that HNPs may function as a biomarker for aging. Copyright © 2009 John Wiley & Sons, Ltd.