Alexander S. Timin

Magnetic silica hybrids modified with guanidine containing co-polymers for drug delivery applications

Guanidine containing co-polymers grafted onto silica nanoparticles to form core-shell structure were prepared by sol-gel method in the presence of γ-Fe2O3 nanoparticles. The morphological features for uncoated and coated silica particles have been characterized with scanning electron microscopy. The results show that the polymer coated silicas exhibit spherical morphology with rough polymeric surface covered by γ-Fe2O3 nanoparticles. The grafting amount of guanidine containing co-polymers evaluated by thermogravimetric analysis was in the range from 17 to 30%. Then, the drug loading properties and cumulative release of silica hybrids modified with guanidine containing co-polymers were evaluated using molsidomine as a model drug. It was shown that after polymer grafting the loading content of molsidomine could reach up to 3.42±0.21 and 2.34±0.14mg/g respectively. The maximum drug release of molsidomine is achieved at pH1.6 (approximately 71-75% release at 37°C), whereas at pH7.4 drug release is lower (50.4-59.6% release at 37°C). These results have an important implication that our magneto-controlled silica hybrids modified with guanidine containing co-polymers are promising as drug carriers with controlled behaviour under influence of magnetic field.

Triple-responsive inorganic–organic hybrid microcapsules as a biocompatible smart platform for the delivery of small molecules

We designed novel hybrid inorganic/organic capsules with unique physicochemical features enabling multimodal triggering by physical (UV light, ultrasound) and chemical (enzymatic treatment) stimuli. Notably, the UV- and ultrasound response was achieved by a synergetic combination of TiO2 and SiO2 nanostructures which were in situ deposited into the polymer shell of microcapsules during sol-gel synthesis. This results in the formation of a composite hybrid shell with enhanced mechanical stability. Such sol-gel modification reduces the permeability of the capsule shell to allow for small molecule encapsulation. At the same time, these hybrid capsules consist of degradable polypeptides and polysaccharides and can be decomposed in response to enzymatic reaction. Upon employing different modes of treatment (UV-light, ultrasound or enzymatic degradation) we can stimulate different mechanisms of cargo release at desired times. Importantly, such capsules have been shown to be non-cytotoxic and can be internalized into human mesenchymal stem cells (MSCs) and cervical cancer cell lines (HeLa) revealing intracellular degradation. This work demonstrates that our hybrid capsules possess a triple stimuli-responsive effect, which is of capital importance for the future design and application of multimodal responsive platforms to improve externally stimulated release of bioactive compounds and their healthcare performance.

Mesenchymal Stem Cell Magnetization: Magnetic Multilayer Microcapsule Uptake, Toxicity, Impact on Functional Properties, and Perspectives for Magnetic Delivery

Mesenchymal stem cells (MSCs) are widely used in cell therapy due to their convenience, multiline differentiation potential, reproducible protocols, and biological properties. The potential of MSCs to impregnate magnetic microcapsules and their possible influence on cell function and ability to response to magnetic field have been explored. Interestingly, the cells suspended in media show much higher ability in internalization of microcapsules, then MSCs adhere into the surface. There is no significant effect of microcapsules on cell toxicity compared with other cell line-capsule internalization reported in literature. Due to internalization of magnetic capsules by the cells, such cell engineering platform is responsive to external magnetic field, which allows to manipulate MSC migration. Magnetically sorted MSCs are capable to differentiation as confirmed by their conversion to adipogenic and osteogenic cells using standard protocols. There is a minor effect of capsule internalization on cell adhesion, though MSCs are still able to form spheroid made by dozen of thousand MSCs. This work demonstrates the potential of use of microcapsule impregnated MSCs to carry internalized micron-sized vesicles and being navigated with external magnetic signaling.

Efficient gene editing via non-viral delivery of CRISPR–Cas9 system using polymeric and hybrid microcarriers

CRISPR-Cas9 is a revolutionary genome-editing technology that has enormous potential for the treatment of genetic diseases. However, the lack of efficient and safe, non-viral delivery systems has hindered its clinical application. Here, we report on the application of polymeric and hybrid microcarriers, made of degradable polymers such as polypeptides and polysaccharides and modified by silica shell, for delivery of all CRISPR-Cas9 components. We found that these microcarriers mediate more efficient transfection than a commercially available liposome-based transfection reagent (>70% vs. <50% for mRNA, >40% vs. 20% for plasmid DNA). For proof-of-concept, we delivered CRISPR-Cas9 components using our capsules to dTomato-expressing HEK293T cells-a model, in which loss of red fluorescence indicates successful gene editing. Notably, transfection of indicator cells translated in high-level dTomato knockout in approx. 70% of transfected cells. In conclusion, we have provided proof-of-principle that our micro-sized containers represent promising non-viral platforms for efficient and safe gene editing.

Multi-layer microcapsules: fresh insights and new applications

Delivery of biologically active compounds remains a topic of intensive research in the last decade. A number of delivery systems have been proposed to enable effective entrapment and targeting and to prolong circulation of drugs with a reduction of side effects [1]. In the late 1990s, an important development in colloidal engineering was to apply layer-by-layer (LbL) assembly of oppositely charged polyelectrolyte and other charged species which had typically been used on planar surfaces, to coat micronand submicron-sized colloidal particles. Coating of colloid particles evolved into fabrication of microcapsules with walls made of polyelectrolyte multilayers, and whose properties could be defined. Intensive research on multilayer microcapsules in the next decade underpinned the ability to bring different functionalities to this delivery system. Indeed, incorporation of responsive polymers or nanoparticles in the microcapsule wall can endow more functionality. There are now hundreds of publications on multilayer capsules demonstrating responsiveness to a range of stimuli including temperature, pH, enzyme activity, sugar, light, magnetic fields, and ultrasound [2]. The use of stimuli that are already utilized in clinical medicine, such as magnetic fields, ultrasound, and/or light to control delivery from microcapsules is a particularly attractive aspect of microcapsule use and could facilitate their development in biomedicine. Versatile use and ease of tailoring properties such as size, permeability, responsiveness, and encapsulated cargo are major advantages of these multilayer capsules. There are still some obstacles such as permeability to small molecules (e.g. anticancer drug – doxorubicin, fluorescent markers – rhodamine B or fluorescein, siRNAs) that limit the widespread application of multilayer capsules but recent developments have started to address these limitations. In this article, we will outline the potential of LbL microcapsules in biomedicine and review recent developments that overcome limitations.

Hybrid inorganic-organic capsules for efficient intracellular delivery of novel siRNAs against influenza A (H1N1) virus infection

The implementation of RNAi technology into the clinical practice has been significantly postponing due to the issues regarding to the delivery of naked siRNA predominantly to target cells. Here we report the approach to enhance the efficiency of siRNA delivery by encapsulating the siRNA into new carrier systems which are obtained via the combination of widely used layer-by-layer technique and in situ modification by sol-gel chemistry. We used three types of siRNAs (NP-717, NP-1155 and NP-1496) in encapsulated form as new therapeutic agents against H1N1 influenza virus infection. By employing the hybrid microcontainers for the siRNA encapsulation we demonstrate the reduction of viral nucleoprotein (NP) level and inhibition of influenza virus production in infected cell lines (MDCK and A549). The obtained hybrid carriers based on assembled biodegradable polyelectrolytes and sol-gel coating possess several advantages such as a high cell uptake efficiency, low toxicity, efficient intracellular delivery of siRNAs and the protection of siRNAs from premature degradation before reaching the target cells. These findings underpin a great potential of versatile microencapsulation technology for the development of anti-viral RNAi delivery systems against influenza virus infection.

A comparison study between electrospun polycaprolactone and piezoelectric poly(3-hydroxybutyrate-co-3-hydroxyvalerate) scaffolds for bone tissue engineering

In this study, bone scaffolds composed of polycaprolactone (PCL), piezoelectric poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) and a combination of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) and silicate containing hydroxyapatite (PHBV-SiHA) were successfully fabricated by a conventional electrospinning process. The morphological, chemical, wetting and biological properties of the scaffolds were examined. All fabricated scaffolds are composed of randomly oriented fibres with diameters from 800nm to 12μm. Fibre size increased with the addition of SiHA to PHBV scaffolds. Moreover, fibre surface roughness in the case of hybrid scaffolds was also increased. XRD, FTIR and Raman spectroscopy were used to analyse the chemical composition of the scaffolds, and contact angle measurements were performed to reveal the wetting behaviour of the synthesized materials. To determine the influence of the piezoelectric nature of PHBV in combination with SiHA nanoparticles on cell attachment and proliferation, PCL (non-piezoelectric), pure PHBV, and PHBV-SiHA scaffolds were seeded with human mesenchymal stem cells (hMSCs). In vitro study on hMSC adhesion, viability, spreading and osteogenic differentiation showed that the PHBV-SiHA scaffolds had the largest adhesion and differentiation abilities compared with other scaffolds. Moreover, the piezoelectric PHBV scaffolds have demonstrated better calcium deposition potential compared with non-piezoelectric PCL. The results of the study revealed pronounced advantages of hybrid PHBV-SiHA scaffolds to be used in bone tissue engineering.

Cell‐Based Drug Delivery and Use of Nano‐and Microcarriers for Cell Functionalization

Cell functionalization with recently developed various nano- and microcarriers for therapeutics has significantly expanded the application of cell therapy and targeted drug delivery for the effective treatment of a number of diseases. The aim of this progress report is to review the most recent advances in cell-based drug vehicles designed as biological transporter platforms for the targeted delivery of different drugs. For the design of cell-based drug vehicles, different pathways of cell functionalization, such as covalent and noncovalent surface modifications, internalization of carriers are considered in greater detail together with approaches for cell visualization in vivo. In addition, several animal models for the study of cell-assisted drug delivery are discussed. Finally, possible future developments and applications of cell-assisted drug vehicles toward targeted transport of drugs to a designated location with no or minimal immune response and toxicity are addressed in light of new pathways in the field of nanomedicine.

Analysis of binding ability of two tetramethylpyridylporphyrins to albumin and its complex with bilirubin

An interaction between 5,10,15,20-tetrakis-(N-methyl-x-pyridyl)porphyrins, x=2; 4 (TMPyPs) with bovine serum albumin (BSA) and its bilirubin (BR) complex was investigated by UV-Viz and fluorescence spectroscopy under imitated physiological conditions involving molecular docking studies. The parameters of forming intermolecular complexes (binding constants, quenching rate constants, quenching sphere radius etc.) were determined. It was showed that the interaction between proteins and TMPyPs occurs via static quenching of protein fluorescence and has predominantly hydrophobic and electrostatic character. It was revealed that obtained complexes are relatively stable, but in the case of TMPyP4 binding with proteins occurs better than TMPyP2. Nevertheless, both TMPyPs have better binding ability with free protein compared to BRBSA at the same time. The influence of TMPyPs on the conformational changes in protein molecules was studied using synchronous fluorescence spectroscopy. It was found that there is no competition of BR with TMPyPs for binging sites on protein molecule and BR displacement does not occur. Molecular docking calculations have showed that TMPyPs can bind with albumin via tryptophan residue in the hydrophilic binding site of protein molecule but it is not one possible interaction way.

Porous Inorganic Carriers Based on Silica, Calcium Carbonate and Calcium Phosphate for Controlled/Modulated Drug Delivery: Fresh Outlook and Future Perspectives

Porous inorganic nanostructured materials are widely used nowadays as drug delivery carriers due to their adventurous features: suitable architecture, large surface area and stability in the biological fluids. Among the different types of inorganic porous materials, silica, calcium carbonate, and calcium phosphate have received significant attention in the last decade. The use of porous inorganic materials as drug carriers for cancer therapy, gene delivery etc. has the potential to improve the life expectancy of the patients affected by the disease. The main goal of this review is to provide general information on the current state of the art of synthesis of the inorganic porous particles based on silica, calcium carbonate and calcium phosphate. Special focus is dedicated to the loading capacity, controllable release of drugs under internal biological stimuli (e.g., pH, redox, enzymes) and external noninvasive stimuli (e.g., light, magnetic field, and ultrasound). Moreover, the diverse compounds to deliver with silica, calcium carbonate and calcium phosphate particles, ranging from the commercial drugs to genetic materials are also discussed.