Conjugated metallothionein-carbon-doxorubicin nanotransporter for targeted breast cancer therapy.

Metalothionein (MT) is a polypeptide of molecular weight in the range of 6-10 kDa. MT typically contains 60 to 68 amino acid residues. MT is characterized by its unique content of metal ions as well as its sulfur content. Higher MT levels were observed in proliferating cells. This fact demonstrates the importance of MT in the process of cellular regulation (relationship to cancer). The most widely used drug for patients with breast cancer metastases is an anthracycline antitumor antibiotic doxorubicin (DOX). However, the clinical use of DOX is limited by dose-related heart muscle damage (cardiomyopathy), more prevalent with increasing cumulative doses. For this reason, creation of novel pharmaceutical formulations based on using alternative methods as nanocarriers for targeted drug delivery to tumour cells is a crucial task in modern pharmacology. The aim of this work was to design a nanotechnological construct. The construct is designed as two separate nanotransporters. The nanotransporter (A) is formed by an antibody-modified AgNPs particle and a carbon nanotube with encapsulated DOX (AgNPs/Ab1/MWCNT/DOX/ODN1). The nanotransporter (B) is engineered with SPION particle modified with antibody and with bound MT (SPION/Ab2/MT/ODN2). Construct AgNPs/Ab1/MWCNT/DOX/ODN1-SPION/Ab2/MT/ODN2 is formed using an oligonucleotide anchor. Individual parts of the nanotransporter were studied using appropriate methods. The presence of MT was monitored electrochemically by Brdicka method in connection with the transfer technique (AdTSV). Characteristic MT signals RS2CO (-1.15 V), Cat1 (-1.25 V), Cat2 (-1.45 V), Cat3 (-1.75 V) were observed at accumulation time of 120s. SDS PAGE confirmed the presence of MT on SPION nanoparticles at sizes 7 to 15 kDa. The DOX signal was fluorometrically monitored (Em 590 nm, Ex 490 nm). AgNPs sizes ranged from 15-20 nm, and the SPION nanoparticles ranged from 20-50 nm. Additionally, used AgNPs nanoparticles exhibited significant antiproliferative activity (growth inhibition by 20-40%) on a model culture S. Cerevisiae. Created nanoconstruct A showed growth inhibition for S. Cerevisiae by more than 50%. The nanoconstruct after these various analysis shows a high potential as an anticancer drug and may be an innovative way how to deal with the breast cancer in a targeted therapy

An insight into the complex roles of metallothioneins in malignant diseases with emphasis on (sub)isoforms/isoforms and epigenetics phenomena

L

Microarray analysis of metallothioneins in human diseases: a review

K

Zinc-modified nanotransporter of anticancer drugs for targeted therapy: biophysical analysis.

Modern anticancer therapy aims to increase the effectiveness of tumor treatment. The aim of this work was to propose a new nanotransporter for targeted delivery of anthracycline antibiotics, which is characterized by its bioavailability, increased uptake of the drug from the bloodstream at the site of tumor tissue and as well as low toxicity to non-target tissue. Chitosan nanoparticles have attracted great attention in the field of drug delivery due to their stability, low toxicity and easy preparation. Deacetylated chitosan skeleton is composed of glucosamine units and has a high density of charged amino groups which allow strong electrostatic interactions with biomolecules, transition metals (Zn, Se) and peptides. We obtained the encapsulation effectiveness of chitosan 20%. Electrochemical detection of the bounded Zn2+ ions into the chitosan structure showed shift from -0.99 to -0.93 V. This result proved the formation of a chitosan-zinc complex. The ability of metallothione in to quench the 2,2-diphenyl-1-picrylhydrazylradicalin the presence of 50 {aelig}M doxorubicin was confirmed by the change of relative absorbance in the range of 50 to 60%

Study of the effect of paracetamol binded in polymeric nanoparticles on dafnia magna.

Drugs are important xenobiotics in the environment. Their use increases with the growth of the human population, but also in agricultural primary production. Paracetamol (PAR) is a widely used analgesic and antipyretic and its production is still growing. Commonly available drug production technologies are being developed very intensively with nanotechnological modifications for their gradual and targeted release. Nanoparticles (ST/PAR) from starch were prepared: PAR (0, 1, 2, 3, 5 and 10 mg/L) was mixed with citric acid ester in a 1:8 v/v ratio for 30 min at 25 °C. By the centrifugation (16.000 g, 30 min) ST/PAR were obtained in the pellet. The effect of PAR was studied on Daphnia magna Straus (Cladocera, Crustacea). Adult females (70-400 mg) were used for self-evaluation. The EC50 was 3.749 mg/L after 48 h of PAR treatment. Total protein values determined by Lowry method were between 0.5-2.2 mg/mL and by Bradford method between 190-676 mg/L. Antioxidant activity values determined by CUPRAC method were between 4-15 μg/mL GAE and by ABTS method ranged between 40-103 μg/mL GAE. PAR values were between 9-40 μM. Subsequently, the biological activity of the prepared nanoparticles was tested

Zinc-modified nanotransporter of doxorubicine for multi-targeted therapy of prostate cancer cells.

Target therapy for oncologic diseases presents a big challenge for advance nanomedicine. In our work, we focused on multi-target approach development. Designed nanotransporter is based on polysaccharide chitosan which allows formation of nanoparticles. These nanoparticles can bind metal ions, mainly zinc (moreover, zinc stabilizes chitosan structure). The estimated zinc concentration was approximately 1 nmol/g of chitosan. In addition, chitosan nanoparticle (cage) irreversibly binds therapeutics which could be applied for targeted therapy of malignant tumours. Designed chitosan structure (LMQ, 10 g) encapsulation efficiency for doxorubicin was 50%. The pH change (tested interval 5 - 8) caused 20% release of doxorubicin from the nanocage. The nanotransporter is orientated to cancer tissue due the fact that the malignant cells highly express metallothionein (MT). The increased affinity of MT to zinc ions causes that the nanotransporter is preferentially bound to tumour regions with a high MT concentration. Our latest experimental results showed the changes in amino acid metabolism of prostate cancer signalized by increase in the amount of amino acid sarcosine. Therefore, the chitosan-based nanotransporter was modified by anti-sarcosine antibody. The functionality of designed nanotransporter was proved by ELISA with double detection of doxorubicin using fluorescence and by peroxidase activity of ABTS substrate. In another system, magnetic separation and identification of individual components of the nanotransporter were used. The sarcosine binding activity was estimated around 50%

A rapid method for the detection of sarcosine using SPIONs/Au/CS/SOX/NPs for prostate cancer sensing.

Background: Sarcosine is an amino acid that is formed by methylation of glycine and is present in trace amounts in the body. Increased sarcosine concentrations in blood plasma and urine are manifested in sarcosinemia and in some other diseases such as prostate cancer. For this purpose, sarcosine detection using the nanomedicine approach was proposed. In this study, we have prepared superparamagnetic iron oxide nanoparticles (SPIONs) with different modified surface area. Nanoparticles (NPs) were modified by chitosan (CS), and sarcosine oxidase (SOX). SPIONs without any modification were taken as controls. Methods and Results: The obtained NPs were characterized by physicochemical methods. The size of the NPs determined by the dynamic light scattering method was as follows: SPIONs/Au/NPs (100–300 nm), SPIONs/Au/CS/NPs (300–700 nm), and SPIONs/Au/CS/SOX/NPs (600–1500 nm). The amount of CS deposited on the NP surface was found to be 48 mg/mL for SPIONs/Au/CS/NPs and 39 mg/mL for SPIONs/Au/CS/SOX/NPs, and repeatability varied around 10%. Pseudo-peroxidase activity of NPs was verified using sarcosine, horseradish peroxidase (HRP) and 3,3′,5,5′-tetramethylbenzidine (TMB) as a substrate. For TMB, all NPs tested evinced substantial pseudo-peroxidase activity at 650 nm. The concentration of SPIONs/Au/CS/SOX/NPs in the reaction mixture was optimized to 0–40 mg/mL. Trinder reaction for sarcosine detection was set up at 510 nm at an optimal reaction temperature of 37 °C and pH 8.0. The course of the reaction was linear for 150 min. The smallest amount of NPs that was able to detect sarcosine was 0.2 mg/well (200 μL of total volume) with the linear dependence y = 0.0011x − 0.0001 and the correlation coefficient r = 0.9992, relative standard deviation (RSD) 6.35%, limit of detection (LOD) 5 μM. The suggested method was further validated for artificial urine analysis (r = 0.99, RSD 21.35%, LOD 18 μM). The calculation between the detected and applied concentrations showed a high correlation coefficient (r = 0.99). NPs were tested for toxicity and no significant growth inhibition was observed in any model system (S. cerevisiae, S. aureus, E. coli). The hemolytic activity of the prepared NPs was similar to that of the phosphate buffered saline (PBS) control. The reaction system was further tested on real urine specimens. Conclusion: The proposed detection system allows the analysis of sarcosine at micromolar concentrations and to monitor changes in its levels as a potential prostate cancer marker. The whole system is suitable for low-cost miniaturization and point-of-care testing technology and diagnostic systems. This system is simple, inexpensive, and convenient for screening tests and telemedicine applications

Phytotoxicity of silver nanoparticles (AgNPs) prepared by green synthesis using sage leaves (Salvia officinalis).

Silver nanoparticles (AgNPs) are widely investigated with regard to their physical, chemical, but also biological properties. Antibacterial and antitumor properties of AgNPs have been intensively studied. In addition, the synthesis using a green approach brings further significant biological properties. However, it is also necessary to monitor the potential toxicity of such nanoparticles in different ecosystems. In this study, the effect of AgNO3 and AgNPs on germinated plants of Zea mays was studied. Effects on basic growth and physiological parameters were observed. There was a statistically significant difference between the variants tested

Zinc-modified nanotransporter of doxorubicin for targeted prostate cancer delivery.

This work investigated the preparation of chitosan nanoparticles used as carriers for doxorubicin for targeted cancer delivery. Prepared nanocarriers were stabilized and functionalized via zinc ions incorporated into the chitosan nanoparticle backbone. We took the advantage of high expression of sarcosine in the prostate cancer cells. The prostate cancer targeting was mediated by the AntiSar antibodies decorated surface of the nanocage. Formation of the chitosan nanoparticles was determined using a ninhydrin assay and differential pulse voltammetry. Obtained results showed the strong effect of tripolyphosphine on the nanoparticle formation. The zinc ions affected strong chitosan backbone coiling both in inner and outer chitosan nanoparticle structure. Zinc electrochemical signal depended on the level of the complex formation and the potential shift from -960 to -950 mV. Formed complex is suitable for doxorubicin delivery. It was observed the 20% entrapment efficiency of doxorubicin and strong dependence of drug release after 120 min in the blood environment. The functionality of the designed nanotransporter was proven. The purposed determination showed linear dependence in the concentration range of Anti-sarcosine IgG labeled gold nanoparticles from 0 to 1000 µg/mL and the regression equation was found to be y = 3.8x - 66.7 and R2 = 0.99. Performed ELISA confirmed the ability of Anti-sarcosine IgG labeled chitosan nanoparticles with loaded doxorubicin to bind to the sarcosine molecule. Observed hemolytic activity of the nanotransporter was 40%. Inhibition activity of our proposed nanotransporter was evaluated to be 0% on the experimental model of S. cerevisiae. Anti-sarcosine IgG labeled chitosan nanoparticles, with loaded doxorubicin stabilized by Zn ions, are a perspective type of nanocarrier for targeted drug therapy managed by specific interaction with sarcosine and metallothionein for prostate cancer