Dual drug delivery system of RAPTA-C and paclitaxel based on fructose coated nanoparticles for metastatic cancer treatment.

Ruthenium complexes have been widely studied as potential alternatives to platinum-type anticancer drugs due to their unique medical properties such as high selectivity, strong ability to inhibit solid tumour metastasis. However, non-specific biodistribution, and weak lethality of ruthenium to cancer cells limit its use in medical application. Drug delivery systems offer the ability to integrate multiple drugs in one system, which is particularly important to enhance the chemotherapeutic efficacy and to potentially achieve a synergistic effect of both drugs. Here, we report a dual drug nanocarrier that is based on a self-assembled biodegradable block copolymer, where the ruthenium complex (RAPTA-C) is chemically attached to the polymer chain, while another drug, paclitaxel (PTX), is entrapped in the core of the micelle. The dual drug delivery system was studied via in vitro tests using MDA-MB-231 breast cancer cells and it was observed that RAPTA-C in combination with PTX significantly enhanced anti-tumour and anti-metastasis activity.

Drug-Loading Content Influences Cellular Uptake of Polymer-Coated Nanocellulose.

While the effects of nanoparticle properties such as shape and size on cellular uptake are widely studied, influences exerted by drug loading have so far been ignored. In this work, nanocellulose (NC) coated by Passerini reaction with poly(2-hydroxy ethyl acrylate) (PHEA-g-NC) was loaded with various amounts of ellipticine (EPT) by electrostatic interactions. The drug-loading content was determined by UV-vis spectroscopy to range between 1.68 and 8.07 wt %. Dynamic light scattering and small-angle neutron scattering revealed an increased dehydration of the polymer shell with increasing drug-loading content, which led to higher protein adsorption and more aggregation. The nanoparticle with the highest drug-loading content, NC-EPT8.0, displayed reduced cellular uptake in U87MG glioma cells and MRC-5 fibroblasts. This also translated into reduced toxicity in these cell lines as well as the breast cancer MCF-7 and the macrophage RAW264.7 cell lines. Additionally, the toxicity in U87MG cancer spheroids was unfavorable. The nanoparticle with the best performance was found to have intermediate drug-loading content where the cellular uptake was adequately high while each nanoparticle was able to deliver a sufficiently toxic amount into the cells. Medium drug loading did not hinder uptake into cells while maintaining sufficiently toxic drug concentrations. It was concluded that while striving for a high drug-loading content is appropriate when designing clinically relevant nanoparticles, it needs to be considered that the drug can cause changes in the physicochemical properties of the nanoparticles that might cause unfavorable effects.

Functional groups in graphene oxide.

Graphene oxide has aroused significant interest for a range of applications owing to their outstanding physico-chemical properties. Specifically, the presence of a large number of reactive chemical moieties such as hydroxyl, carboxyl, epoxide, and sp2 carbon allows these novel materials to be tailored with additional functionalities with the purpose of tuning intrinsic properties. There has been a vivid discussion on the non-covalent modification of GO; however, a comprehensive summary of the chemical functionalization which enables forming a stable particle is still elusive. Hence, in this study, we summarize recently advanced methodologies used for designing the functional GO for their use in specific applications. Together with a brief discussion on the essential characterization techniques, this study will provide fundamental insight into the latest developments in the preparation of covalently modified GO derivatives, thereby leading to their broader utilization in future.

Photo-Induced Modification of Nanocellulose: The Design of Self-Fluorescent Drug Carriers.

Nanocellulose is an excellent carrier to deliver drugs, as the material is biocompatible and has a desirable non-spherical shape. However, nanocellulose displays low solubility in aqueous solution and needs to be modified with water-soluble polymers in order to achieve high colloidal stability. In this study, (2,2,6,6-tetramethylpiperidin-1-yl)oxyl or (2,2,6,6-tetramethylpiperidin-1-yl)oxidanyl (TEMPO)-oxidized cellulose nanofibers bearing carboxylic acid moieties (TOCNs) are modified by nitrile imine-mediated tetrazole/carboxylic acid ligation. The advantage of this reaction is that TOCNs do not need to be modified further and the polymer with tetrazole end-functionalities can be directly clicked onto the TOCNs forming fluorescent functional groups. Poly(2-hydroxyethyl acrylate) with a tetrazole end-functionality is prepared using RAFT polymerization. The polymer is mixed with TOCNs and after irradiation at λ = 326 nm for 10 h, fluorescent pHEA-g-TOCNs are obtained. The polymer-grafted nanocellulose is found to disperse well in water and has only limited albumin binding. The uptake of these nanoparticles by MCF-7 breast cancer cell lines can now be monitored by fluorescent microscopy without further modification. Excess negatively charged carboxylic groups of TOCNs allow doxorubicin loading by electrostatic interactions at various drug-loading capacities. Higher drug loading is more efficient in inhibiting the cell proliferation, highlighting the effect of drug loading on toxicity.

Sugar Concentration and Arrangement on the Surface of Glycopolymer Micelles Affect the Interaction with Cancer Cells.

Glycopolymer-coated nanoparticles have attracted significant interest over the past few years, because of their selective interaction with carbohydrate receptors found on the surface of cells. While the type of carbohydrate determines the strength of the ligand-receptor interaction, the presentation of the sugar can be highly influential as the carbohydrate needs to be accessible in order to display good binding. To shine more light on the relationship between nanoparticle structure and cell uptake, we have designed several micelles based on fructose containing block copolymers, which are selective to GLUT5 receptors found on breast cancer cell lines. The polymers were based on poly-d,l-lactide (PLA), poly(2-hydroxyethyl) acrylate (PHEA), and poly(1- O-acryloyl-ß-d-fructopyranose) (P[1- O-AFru]). A set of six micelles was synthesized based on four fructose containing micelles (PLA242- b-P[1- O-AFru]41, PLA242- b-P[1- O-AFru]179, PLA242- b-P[1- O-AFru46-c-HEA214], PLA242- b-PHEA280- b-P[1- O-AFru]41) and two neutral controls (PLA247- b-PHEA53 and PLA247- b-PHEA166). SAXS analysis revealed that longer hydrophilic polymers led to lower aggregation numbers and larger hydrophilic shells, suggesting more glycopolymer mobility. Cellular uptake studies via flow cytometry and confocal laser scanning microscopy (CLSM) confirmed that the micelles based on PLA242- b-P[1- O-AFru]179 show, by far, the highest uptake by MCF-7 and MDA-MB-231 breast cancer cell lines while the uptake of all micelles by RAW264.7 cell is negligible. The same micelle displayed was far superior in penetrating MCF-7 cancer spheroids (three-dimensional (3D) model). Taking the physicochemical characterization obtained by SAXS and the in vitro results together, it could be concluded that the glycopolymer chains on the surface of micelle must display high mobility. Moreover, a high density of fructose was found to be necessary to achieve good biological activity as lowering the epitope density led immediately to lower cellular uptake. This work showed that it is crucial to understand the micelle structure in order to maximize the biological activity of glycopolymer micelles.

Polyion complex micelle based on albumin-polymer conjugates: multifunctional oligonucleotide transfection vectors for anticancer chemotherapeutics.

Novel biocompatible polyion complex micelles, containing bovine serum albumin (BSA), polymer, and oligonucleotide, were synthesized as a generation of vectors for the gene transfection. Maleimide-terminated poly((N,N-dimethyl amino) ethyl methacrylate) (PDMAEMA) was prepared via reversible addition-fragmentation chain transfer (RAFT) polymerization and subsequently deprotected. Precise one to one albumin-PDMAEMA bioconjugates have been achieved via 1,4-addition with the free thiol group on Cys34 on the BSA protein. SDS-PAGE and GPC (water) confirmed and quantified the successful conjugation. The conjugation efficiency was found to be independent of the molecular weight of PDMAEMA. After careful pH adjustment, the conjugate could efficiently condense anticancer oligonucleotide, ISIS 5132, which resulted in particles of 15-35 nm with a negative zeta-potential. The size was easily controlled by the polymer chain length. The albumin corona provides complete protection of the cationic polymer and genetic drug, which gave rise to lower potential toxicity from the polymer and higher gene transfection efficiency. Although a control experiment with a traditional PEG-based polyion complex micelle could deliver the drug just as effectively, if not more so, to the ovarian cancer cell line OVCAR-3, this carrier had no selectivity toward cancerous cells and proved just as toxic to HS27 (fibroblast) cell line. In contrast, the albumin-coated particles demonstrated desirable selectivity toward cancerous cells and have been shown to have outstanding performance in the cytotoxicity tests of several carcinoma monolayer cell models. In addition, the complex micelles were able to destroy pancreatic multicellular tumor spheroids, while free ISIS 5132 could not penetrate the spheroid at all. Hence, albumin-coated/oligonucleotide complex micelles are far more promising than the most classical gene delivery vectors.

Fast-relaxing hydrogels with reversibly tunable mechanics for dynamic cancer cell culture.

The mechanics of the tumor microenvironment (TME) significantly impact disease progression and the efficacy of anti-cancer therapeutics. While it is recognized that advanced in vitro cancer models will benefit cancer research, none of the current engineered extracellular matrices (ECM) adequately recapitulate the highly dynamic TME. Through integrating reversible boronate-ester bonding and dithiolane ring-opening polymerization, we fabricated synthetic polymer hydrogels with tumor-mimetic fast relaxation and reversibly tunable elastic moduli. Importantly, the crosslinking and dynamic stiffening of matrix mechanics were achieved in the absence of a photoinitiator, often the source of cytotoxicity. Central to this strategy was Poly(PEGA-co-LAA-co-AAPBA) (PELA), a highly defined polymer synthesized by reversible addition-fragmentation chain transfer (RAFT) polymerization. PELA contains dithiolane for initiator-free gel crosslinking, stiffening, and softening, as well as boronic acid for complexation with diol-containing polymers to give rise to tunable viscoelasticity. PELA hydrogels were highly cytocompatible for dynamic culture of patient-derived pancreatic cancer cells. It was found that the fast-relaxing matrix induced mesenchymal phenotype of cancer cells, and dynamic matrix stiffening restricted tumor spheroid growth. Moreover, this new dynamic viscoelastic hydrogel system permitted sequential stiffening and softening to mimic the physical changes of TME.

WITHDRAWN: Conjugation of Ruthenium Drugs to Nanocellulose using Boronic Ester

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A new 3D organotypic model of ovarian cancer to help evaluate the antimetastatic activity of RAPTA-C conjugated micelles.

INTRODUCTION: Ovarian cancer is often diagnosed at a late stage, when disease has spread to extra-pelvic regions such as the omentum. There are limited treatment options available for women with extensive disease and tumours often relapse after current chemotherapy regimens. Therefore, novel drugs should be investigated for the treatment of ovarian cancer. A 3D organotypic model of ovarian cancer can provide a specific platform for the evaluation of nano-drugs. Using patient derived primary cells, the 3D model mimics the ovarian metastatic microenvironment allowing efficient and reproducible testing of many nanoparticles. Dichlororuthenium(ii) (p-cymene) (1,3,5-triaza-7-phosphaadamantane) (RAPTA-C) conjugated fructose-micelles have been used as the promising nano-drug for the treatment of metastatic cancer. Therefore we aimed to investigate the anti-metastatic properties of RAPTA-C conjugated micelles in ovarian cancer metastasis. METHODS: Ovarian cancer cell adhesion and invasion into a model of omentum were analyzed with and without RAPTA-C conjugated micelles in a range of conditions. RESULTS: We observed that RAPTA-C showed low general toxicity to both primary healthy and cancer cell lines. RAPTA-C loaded micelles significantly enhance the internalization of ruthenium inside the cells compared to free drugs. RAPTA-C did not affect adhesion of OVCAR4 ovarian cancer cells; however, it significantly inhibited invasion of these cells within the omentum model, either in its free form or as cargos inside the micelles. However, when OVCAR4 were treated prior to implantation, invasion was not inhibited. CONCLUSION: A 3D organotypic model provides a clinically relevant and simple method to evaluate the efficiency of nano-drug treatment of ovarian cancer. The ability to inhibit metastasis of RAPTA-C delivered in fructose coated nanoparticles was investigated for the first time via this model. These results provide a good basis to continue the development of this nano-drug in vivo.

Covalent Tethering of Temperature Responsive pNIPAm onto TEMPO-Oxidized Cellulose Nanofibrils via Three-Component Passerini Reaction.

A critical challenge in the application of functional cellulose fibrils is to perform efficient surface modification without disrupting the original properties. Three-component Passerini reaction (Passerini 3-CR) is regarded as an effective functionalization approach which can be carried out under mild and fast reaction condition. In this study, we investigated the application of Passerini 3-CR for the synthesis of thermoresponsive cellulose fibrils by covalently tethering poly(N-isopropylacrylamide) in aqueous condition at ambient temperature. The three components, a TEMPO-oxidized cellulose nanofiber bearing carboxylic acid moieties (TOCN-COOH), a functionalized polymer with aldehyde group (pNIPAm-COH) and a cyclohexyl isocyanide, were reacted in one pot resulting in 36% of grafting efficiency within 30 min. The chemical coupling was evidenced by improved aqueous dispersibility, which was further confirmed by FT-IR, TGA, UV-vis, and turbidity study. It was observed that the grafting efficiency is strongly dependent on the chain length of the polymer. Furthermore, AFM and X-ray diffraction measurements affirmed the suitability of the proposed method for chemical modification of cellulose nanofibers without significantly compromising the original morphology and structural integrity.

Dual-Responsive pH and Temperature Sensitive Nanoparticles Based on Methacrylic Acid and Di(ethylene glycol) Methyl Ether Methacrylate for the Triggered Release of Drugs.

A series of thermo-and pH-responsive poly(methyl methacrylate)-block-poly[methacrylic acid-co-di(ethylene glycol) methyl ether methacrylate] PMMA-b-P[MAA-co-DEGMA] block copolymers were synthesized by RAFT polymerization and self-assembled into micelles. The molar ratio of MAA was altered from 0-12% in order to modulate the lower critical solution temperature (LCST) of PDEGMA. The release of the drug albendazole from the micelle was strongly dependent on the temperature and the LCST value of the polymer. Systems below the LCST released the drug slowly while increasing the temperature above the LCST or decreasing the pH value to 5 resulted in the burst-like release of the drug. ABZ delivered in this pH-responsive drug carrier had a higher toxicity than the free drug or the drug delivered in a non-responsive drug carrier.