Preparation and characterization of lapatinib-loaded PVP nanofiber amorphous solid dispersion by electrospinning

Lapatinib-loaded polyvinylpyrrolidone-based nanofibrous solid dispersions were prepared by electrospinning in order to enhance the aqueous solubility and dissolution rate of the anticancer drug. The prepared nanofibers were characterized by smooth-surfaced, homogenous filaments with average diameters of 462±160 nm determined by scanning electron microscopy. The crystalline to amorphous transition of the active ingredient was confirmed by differential scanning calorimetry, while Raman spectroscopy showed that amorphous lapatinib was uniformly distributed in the fibrous structures. Gas chromatographic analyses revealed that residual solvents in the nanofiber mats were below the ICH Guideline Q3C recommended limits, namely ethanol 10.9±2.3 ppm (recommended limit 5000 ppm) and dimethyl formamide 780±56 ppm (recommended limit 880 ppm). Determination of drug content and in vitro dissolution studies were performed in order to observe the influence of electrospinning on the drug release characteristics of the product obtained. The lapatinib content in the nanofibers were measured to be 16.76±0.11 w/w%, whereas the dissolution study at pH 6.8 indicated a rapid disintegration of the nanofibrous mats, releasing ~70% of the drug loading under 5 minutes compared to the ~0.05% dissolution of the neat lapatinib ditosylate. The results confirm the applicability of electrospinning for the improvement of physicochemical characteristics of the poorly bioavailable anticancer agent

Neutrophil extracellular traps in thrombi retrieved during interventional treatment of ischemic arterial diseases

IntroductionThe ultrastructure and cellular composition of thrombi has a profound effect on the outcome of acute ischemic stroke (AIS), coronary (CAD) and peripheral artery disease (PAD). Activated neutrophils release a web-like structure composed mainly of DNA and citrullinated histones, called neutrophil extracellular traps (NET) that modify the stability and lysability of fibrin. Here, we investigated the NET-related structural features of thrombi retrieved from different arterial localizations and their interrelations with routinely available clinical data.Patients and methodsThrombi extracted from AIS (n = 78), CAD (n = 66) or PAD (n = 64) patients were processed for scanning electron microscopy, (immune)stained for fibrin, citrullinated histone H3 (cH3) and extracellular DNA. Fibrin fiber diameter, cellular components, DNA and cH3 were measured and analyzed in relation to clinical parameters.ResultsDNA was least present in AIS thrombi showing a 2.5-fold lower DNA/fibrin ratio than PAD, whereas cH3 antigen was unvaryingly present at all locations. The NET content of thrombi correlated parabolically with systemic inflammatory markers and positively with patients' age. The median platelet content was lower in PAD (2.2%) than in either AIS (3.9%) or CAD (3.1%) and thrombi from smokers contained less platelets than non-smokers. Fibrin fibers were significantly thicker in male patients with CAD (median fiber diameter 76.3 nm) compared to AIS (64.1 nm) or PAD (62.1 nm) and their diameter correlated parabolically with systemic inflammatory markers.ConclusionsThe observed NET-related variations in thrombus structure shed light on novel determinants of thrombus stability that eventually affect both the spontaneous progress and therapeutic outcome of ischemic arterial diseases

Mutations at the Subunit Interface of Yeast Proliferating Cell Nuclear Antigen Reveal a Versatile Regulatory Domain

Acknowledgments We thank Szilvia Minorits for technical assistance. I.U. conceived and designed the project and wrote the manuscript. All authors participated in designing and performing the experiments, and analyzing the results. The authors declare no competing financial interests. This work was also supported by a grant from the National Research, Development and Innovation Office GINOP-2.3.2-15-2016-00001. Funding: This work was supported by Hungarian Science Foundation Grant OTKA 109521 and National Research Development and Innovation Office GINOP-2.3.2-15-2016-00001. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Peer reviewedPublisher PD

Ribonucleoprotein-masked nicks at 50-kbp intervals in the eukaryotic genomic DNA

By using a microscopic approach, field inversion single-cell gel electrophoresis, we show that preformed single-strand discontinuities are present in the chromatin of resting and proliferating mammalian and yeast cells. These single-strand breaks are primarily nicks positioned at ≈50-kbp intervals throughout the entire genome that could be efficiently labeled in situ by DNA polymerase I holoenzyme but not by Klenow fragment and terminal transferase unless after ribonucleolytic treatments. The RNA molecules involved appear to comprise R-loops, recognized by the S9.6 RNA/DNA hybrid-specific antibody. By using the breakpoint cluster region of the Mixed Lineage Leukemia (MLL) gene as a model, we have found that the number of manifest nicks detected by FISH performed after field inversion single-cell gel electrophoresis depends on epigenetic context, but the difference between germ-line and translocated MLL alleles is abolished by protease treatment. Our data imply that the double-stranded genomic DNA is composed of contiguous rather than continuous single strands and reveal an aspect of higher-order chromatin organization with ribonucleoprotein-associated persistent nicks defining ≈50-kbp domains