Hydrophobin-nanofibrillated cellulose stabilized emulsions for encapsulation and release of BCS class II drugs

The purpose of this study was to construct biopolymer-based oil-in-water emulsion formulations for encapsulation and release of poorly water soluble model compounds naproxen and ibuprofen. Class II hydrophobin protein HFBII from Trichoderma reesei was used as a surfactant to stabilize the oil/water interfaces of the emulsion droplets in the continuous aqueous phase. Nanofibrillated cellulose (NFC) was used as a viscosity modifier to further stabilize the emulsions and encapsulate protein coated oil droplets in NFC fiber network. The potential of both native and oxidized NFC were studied for this purpose. Various emulsion formulations were prepared and the abilities of different formulations to control the drug release rate of naproxen and ibuprofen, used as model compounds, were evaluated. The optimal formulation for sustained drug release consisted of 0.01% of drug, 0.1% HFBII, 0.15% oxidized NFC, 10% soybean oil and 90% water phase. By comparison, the use of native NFC in combination with HFBII resulted in an immediate drug release for both of the compounds. The results indicate that these NFC originated biopolymers are suitable for pharmaceutical emulsion formulations. The native and oxidized NFC grades can be used as emulsion stabilizers in sustained and immediate drug release applications. Furthermore, stabilization of the emulsions was achieved with low concentrations of both HFBII and NFC, which may be an advantage when compared to surfactant concentrations of conventional excipients traditionally used in pharmaceutical emulsion formulations. (C) 2017 Elsevier B.V. All rights reserved.Peer reviewe

Technetium-99m-labeled nanofibrillar cellulose hydrogel for in vivo drug release

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Infrared and Raman spectroscopy for purity assessment of extracellular vesicles

Extracellular vesicles (EVs) are a complex and heterogeneous population of nanoparticles involved in cell-to-cell communication. Recently, numerous studies have indicated the potential of EVs as therapeutic agents, drug carriers and diagnostic tools. However, the results of these studies are often difficult to evaluate, since different characterization methods are used to assess the purity, physical and biochemical characteristics of the EV samples. In this study, we compared four methods for the EV sample characterization and purity assessment: i) the particle-to-protein ratio based on particle analyses with nanoparticle tracking and protein concentration by bicinchoninic acid assay, ii) Western Blot analysis for specific EV biomarkers, iii) two spectroscopic lipid-to protein ratios by either the attenuated total reflection Fourier transform infrared (ATR-FTIR) or Raman spectroscopy. The results confirm the value of Raman and ATR-FTIR spectroscopy as robust, fast and operator independent tools that require only a few microliters of EV sample. We propose that the spectroscopic lipid-to protein (Li/Pr) ratios are reliable parameters for the purity assessment of EV preparations. Moreover, apart from determining protein concentrations, we show that ATR-FTIR spectroscopy can also be used for indirect measurements of EV concentrations. Nevertheless, the Li/Pr ratios do not represent full characterization of the EV preparations. For a complete characterization of selected EV preparations, we recommend also additional use of particle size distribution and EV biomarker analysis.Peer reviewe

New In Vitro-In Silico Approach for the Prediction of In Vivo Performance of Drug Combinations

Pharmacokinetic (PK) studies improve the design of dosing regimens in preclinical and clinical settings. In complex diseases like cancer, single-agent approaches are often insufficient for an effective treatment, and drug combination therapies can be implemented. In this work, in silico PK models were developed based on in vitro assays results, with the goal of predicting the in vivo performance of drug combinations in the context of cancer therapy. Combinations of reference drugs for cancer treatment, gemcitabine and 5-fluorouracil (5-FU), and repurposed drugs itraconazole, verapamil or tacrine, were evaluated in vitro. Then, two-compartment PK models were developed based on the previous in vitro studies and on the PK profile reported in the literature for human patients. Considering the quantification parameter area under the dose-response-time curve (AUCeffect) for the combinations effect, itraconazole was the most effective in combination with either reference anticancer drugs. In addition, cell growth inhibition was itraconazole-dose dependent and an increase in effect was predicted if itraconazole administration was continued (24-h dosing interval). This work demonstrates that in silico methods and AUCeffect are powerful tools to study relationships between tissue drug concentration and the percentage of cell growth inhibition over time

Microvesicle- and exosome- mediated drug delivery enhances the cytotoxicity of Paclitaxel in autologous prostate cancer cells

Background: Extracellular vesicles (EVs) are naturally occurring membrane particles that mediate intercellular communication by delivering molecular information between cells. In this study, we investigated the effectiveness of two different populations of EVs (microvesicle- and exosome-enriched) as carriers of Paclitaxel to autologous prostate cancer cells. Methods: EVs were isolated from LNCaP- and PC-3 prostate cancer cell cultures using differential centrifugation and characterized by electron microscopy, nanoparticle tracking analysis, and Western blot. The uptake of microvesicles and exosomes by the autologous prostate cancer cells was assessed by flow cytometry and confocal microscopy. The EVs were loaded with Paclitaxel and the effectiveness of EV-mediated drug delivery was assessed with viability assays. The distribution of EVs and EV-delivered Paclitaxel in cells was inspected by confocal microscopy. Results: Our main finding was that the loading of Paclitaxel to autologous prostate cancer cell-derived EVs increased its cytotoxic effect. This capacity was independent of the EV population and the cell line tested. Although the EVs without the drug increased cancer cell viability, the net effect of enhanced cytotoxicity remained. Both EV populations delivered Paclitaxel to the recipient cells through endocytosis, leading to the release of the drug from within the cells. The removal of EV surface proteins did not affect exosomes, while the drug delivery mediated by microvesicles was partially inhibited. Conclusions: Cancer cell-derived EVs can be used as effective carriers of Paclitaxel to their parental cells, bringing the drug into the cells through an endocytic pathway and increasing its cytotoxicity. However, due to the increased cell viability, the use of cancer cell-derived EVs must be further investigated before any clinical applications can be designed. (C) 2015 The Authors. Published by Elsevier B.V.Peer reviewe

Effectiveness and cost-effectiveness of a people-centred care model for community-living older people versus usual care ─ A randomised controlled trial

Background There is a need for effective and cost-effective interprofessional care models that support older people to maintain their quality of life (QoL) and physical performance to live longer independently in their own homes. Objectives The objectives were to evaluate effectiveness, QoL and physical performance, and cost-utility of a people-centred care model (PCCM), including the contribution of clinically trained pharmacists, compared with that of usual care in primary care. Methods A randomised controlled trial (RCT) with a two-year follow-up was conducted. The participants were multimorbid community-living older people, aged ≥75 years. The intervention comprised an at-home patient interview, health review, pharmacist-led clinical medication review, an interprofessional team meeting, and nurse-led care coordination and health support. At the baseline and at the 1-year and 2-year follow-ups, QoL (SF-36, 36-Item Short-Form Health Survey) and physical performance (SPPB, Short Performance Physical Battery) were measured. Additionally, a physical dimension component summary in the SF-36 was calculated. The SF-36 data were transformed into SF-6D scores to calculate quality-adjusted life-years (QALYs). Healthcare resource use were collected and transformed into costs. A healthcare payer perspective was adopted. Incremental cost-effectiveness ratio (ICER) was calculated, and one-way sensitivity analysis was performed. Results No statistically or clinically significant differences were observed between the usual care (n = 126) and intervention group (n = 151) patients in their QoL; at the 2-year follow-up the mean difference was −0.02, (95 % CI -0.07; 0.04,p = 0.56). While the mean difference between the groups in physical performance at the 2-year follow-up was −1.02, (−1.94;-0.10,p = 0.03), between the physical component summary scores it was −7.3, (−15.2; 0.6,p = 0.07). The ICER was −73 638€/QALY, hence, the developed PCCM dominated usual care, since it was more effective and less costly. Conclusions The cost-utility analysis showed that the PCCM including pharmacist-led medication review dominated usual care. However, it had no effect on QoL and the effect towards physical performance remained unclear.Peer reviewe

Preservation of biomaterials and cells by freeze-drying : Change of paradigm

Freeze-drying is the most widespread method to preserve protein drugs and vaccines in a dry form facilitating their storage and transportation without the laborious and expensive cold chain. Extending this method for the preservation of natural biomaterials and cells in a dry form would provide similar benefits, but most results in the domain are still below expectations. In this review, rather than consider freeze-drying as a traditional black box we "break it" through a detailed process thinking approach. We discuss freeze-drying from process thinking aspects, introduce the chemical, physical, and mechanical environments important in this process, and present advanced biophotonic process analytical technology. In the end, we review the state of the art in the freezedrying of the biomaterials, extracellular vesicles, and cells. We suggest that the rational design of the experiment and implementation of advanced biophotonic tools are required to successfully preserve the natural biomaterials and cells by freeze-drying. We discuss this change of paradigm with existing literature and elaborate on our perspective based on our new unpublished results.Peer reviewe

Differences in definitive endoderm induction approaches using growth factors and small molecules

Definitive endoderm (DE) is the first stage of human pluripotent stem cell (hPSC) differentiation into hepatocyte-like cells. Developing human liver cell models for pharmaceutical applications is highly demanding. Due to the vast number of existing protocols to generate DE cells from hPSCs, we aimed to compare the specificity and efficiency of selected published differentiation conditions. We differentiated two hPSC lines (induced PSC and embryonic stem cell) to DE cells on Matrigel matrix using growth factors (Activin A and Wnt-3a) and small molecules (sodium butyrate and IDE 1) in different combinations. By studying dynamic changes during 6 days in cell morphology and the expression of markers for pluripotency, DE, and other germ layer lineages, we found that Activin A is essential for DE differentiation, while Wnt-3a and sodium butyrate are dispensable. Although sodium butyrate exerted rapid DE differentiation kinetics, it caused massive cell death and could not generate sufficient cells for further differentiation and applications. We further discover that IDE 1 could not induce DE as reported previously. Hereby, we compared different conditions for DE induction and found an effective six day-protocol to obtain DE cells for the further differentiation and applications.Peer reviewe

Elucidating the Signal Responses of Multi-Parametric Surface Plasmon Resonance Living Cell Sensing: A Comparison between Optical Modeling and Drug-MDCKII Cell Interaction Measurements

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