In vitro characterization of a biodegradable chitosan/bioactive glass coating for Mg alloys

Magnesium and its alloys have already been proposed for biomedical applications in 1878. However, up to date no extended, successful medical Mg product is commercially available. The drawbacks of permanent implants, like stress-shielding or possible release of metal ions through wear, can be avoided with the use of biodegradable metals. Temporary implants as such could make a second surgical process to remove the implant unnecessary, not only decreasing the healthcare costs and associated risks of a surgery, but also reducing the trauma to the patient. Mg is an abundant cation in the human body and in part physiologically beneficial as the surrounding tissue can absorb and consume the ions. The main problems related to the usage of Mg and its alloys is its high chemical reactivity, a related low corrosion resistance, especially in chloride-containing environments and the accompanying fast hydrogen gas production. [1] In order to overcome these problems in this study a coating of a natural polymer/bioactive glass composite is applied using electrophoretic deposition. Additionally, functional properties like drug delivery characteristics and antibacterial capacity are added to these coatings. As natural, cationic polymer chitosan is taken which is the supporting material in the exoskeleton of crustaceans and insects (crab, butterfly) and in cell walls of fungi. It combines biodegradability and biocompatibility with the ability to promote cell adhesion. [1] To avoid dissolution of the Mg alloy substrate during the deposition, a pretreatment is used. The immersion in DMEM for 24 h is increasing the corrosion resistance to a level that the acidic, aqueous electrolyte during deposition is not corroding the Mg substrate. [2] A comparative study was performed on replacing part of the bioactive glass as ceramic part with silica particles in order to maintain a topography during dissolution of the glass. A constant solid content of 1 g/l was chosen, with 0.5 g/l chitosan in 1 vol% acetic acid, 20 vol% water and 79 vol% ethanol following previous studies. [3] For the cathodic deposition process 0.5 cm electrode distance with stainless steel as the counter electrode was used. The deposition was performed under constant current (50 V) and constant voltage (35 mA) with varying processing times. [1] Heise S, Virtanen S, Boccaccini AR. 2016. Tackling Mg alloy corrosion by natural polymer coatings—A review. J Biomed Mater Res Part A 2016:104A:2628–2641 [2] Wagener V, Virtanen S. 2016 Protective layer formation on magnesium in cell culture medium. Mater. Sci. Eng. C 63, 341–351 [3] Cordero-Arias, L. et al. 2013 Electrophoretic deposition of nanostructured-TiO2/Chitosan composite coatings on stainless steel. R. Soc. Chem. 3, 11247-11254 Acknowledgements: This study is supported by the German Science Foundation (DFG)

DEVELOPMENT OF A BIODEGRADABLE NATURAL POLYMER/CERAMIC COATING FOR MG ALLOYS USING ELECTROPHORETIC DEPOSITION

Magnesium and its alloys have already been proposed for biomedical applications in 1878. However, up to date no extended, successful medical Mg product is commercially available. The drawbacks of permanent implants, like stress-shielding or possible release of metal ions through wear, can be avoided with the use of biodegradable metals. Temporary implants as such could make a second surgical process to remove the implant unnecessary, not only decreasing the healthcare costs and associated risks of a surgery, but also reducing the trauma to the patient. Mg is an abundant cation in the human body and in part physiologically beneficial as the surrounding tissue can absorb and consume the ions. The main problems related to the usage of Mg and its alloys is its high chemical reactivity, a related low corrosion resistance, especially in chloride-containing environments and the accompanying fast hydrogen gas production. [1] In order to overcome these problems in this study a coating of a natural polymer/bioactive glass composite is applied using electrophoretic deposition. Additionally, functional properties like drug delivery characteristics and antibacterial capacity are added to these coatings. As natural, cationic polymer chitosan is taken which is the supporting material in the exoskeleton of crustaceans and insects (crab, butterfly) and in cell walls of fungi. It combines biodegradability and biocompatibility with the ability to promote cell adhesion. [1] To avoid dissolution of the Mg alloy substrate during the deposition, a pretreatment is used. The immersion in DMEM for 24 h is increasing the corrosion resistance to a level that the acidic, aqueous electrolyte during deposition is not corroding the Mg substrate. [2] A comparative study was performed on replacing part of the bioactive glass as ceramic part with silica particles in order to maintain a topography during dissolution of the glass. A constant solid content of 1 g/l was chosen, with 0.5 g/l chitosan in 1 vol% acetic acid, 20 vol% water and 79 vol% ethanol following previous studies. [3] For the cathodic deposition process 0.5 cm electrode distance with stainless steel as the counter electrode was used. The deposition was performed under constant current (50 V) and constant voltage (35 mA) with varying processing times. [1] Heise S, Virtanen S, Boccaccini AR. 2016. Tackling Mg alloy corrosion by natural polymer coatings—A review. J Biomed Mater Res Part A 2016:104A:2628–2641 [2] Wagener V, Virtanen S. 2016 Protective layer formation on magnesium in cell culture medium. Mater. Sci. Eng. C 63, 341–351 [3] Cordero-Arias, L. et al. 2013 Electrophoretic deposition of nanostructured-TiO2/Chitosan composite coatings on stainless steel. R. Soc. Chem. 3, 11247-11254 Acknowledgements: This study is supported by the German Science Foundation (DFG)

Biomarkers for Homologous Recombination Deficiency in Cancer

DNA double-strand breaks foster tumorigenesis and cell death. Two distinct mechanisms can be activated by the cell for DNA repair: the accurate mechanism of homologous recombination repair or the error-prone non-homologous end joining. Homologous Recombination Deficiency (HRD) is associated with sensitivity towards PARP inhibitors (PARPi) and its determination is used as a biomarker for therapy decision making. Nevertheless, the biology of HRD is rather complex and the application, as well as the benefit of the different HRD biomarker assays, is controversial. Acquiring knowledge of the underlying molecular mechanisms is the main prerequisite for integration of new biomarker tests. This study presents an overview of the major DNA repair mechanisms and defines the concepts of HRR, HRD and BRCAness. Moreover, currently available biomarker assays are described and discussed with respect to their application for routine clinical diagnostics. Since patient stratification for efficient PARP inhibitor therapy requires determination of the BRCA mutation status and genomic instability, both should be established comprehensively. For this purpose, a broad spectrum of distinct assays to determine such combined HRD scores is already available. Nevertheless, all tests require careful validation using clinical samples to meet the criteria for their establishment in clinical testing

Immune-phenotyping of pleomorphic dermal sarcomas suggests this entity as a potential candidate for immunotherapy

BackgroundPleomorphic dermal sarcomas (PDS) are sarcomas of the skin with local recurrences in up to 28% of cases, and distant metastases in up to 20%. Although recent evidence provides a strong rational to explore immunotherapeutics in solid tumors, nothing is known about the immune environment of PDS.MethodsIn the current study, a comprehensive immune-phenotyping of 14 PDS using RNA and protein expression analyses, as well as quantitative assessment of immune cells using an image-analysis tool was performed.ResultsThree out of 14 PDS revealed high levels of CD8-positive tumor-infiltrating T-lymphocytes (TILs), also showing elevated levels of immune-related cytokines such as IL1A, IL2, as well as markers that were very recently linked to enhanced response of immunotherapy in malignant melanoma, including CD27, and CD40L. Using a multivariate analysis, we found a number of differentially expressed genes in the CD8-high group including: CD74, LYZ and HLA-B, while the remaining cases revealed enhanced levels of immune-suppressive cytokines including CXCL14. The CD8-high PDS showed strong MHC-I expression and revealed infiltration by PD-L1-, PD-1- and LAG-3-expressing immune cells. Tumor-associated macrophages (TAMs) predominantly consisted of CD68+, CD163+, and CD204+M2 macrophages showing an accentuation at the tumor invasion front.ConclusionsTogether, we provide first explorative evidence about the immune-environment of PDS tumors that may guide future decisions whether individuals presenting with advanced PDS could qualify for immunotherapeutic options

Advance of theragnosis biomarkers in lung cancer: from clinical to molecular pathology and biology

One distinct molecular subtype of non-small cell lung cancer (NSCLC) is defined by rearrangement of the anaplastic lymphoma kinase (ALK). The increasing knowledge over the last years has enabled the continuous improvement of ALK inhibitors; however, resistance in these patients remains a major concern. In this review, we summarize recent findings in ALK+-adenocarcinoma of the lung, highlighting the role of TP53 mutations in this specific cancer type and suggest new diagnostic strategies for the future, in order to improve patient's outcome

First report on two cases of pleomorphic dermal sarcoma successfully treated with immune checkpoint inhibitors

Pleomorphic dermal sarcoma (PDS) is one of the most common sarcoma of the skin. Currently, limited treatment options exist for advanced stages of the disease. While immune checkpoint inhibitors (CPIs) have revolutionized cancer treatment options-their efficacy in PDS has not been explored yet. Here, we present two advanced PDS cases that showed response to anti-PD-1 therapy. Patient A had a locally metastasized PDS and reached a complete remission of the disease after eight cycles of Pembrolizumab. Patient B developed an inoperable relapse of PDS with a complete remission of the disease 4 months after treatment with Pembrolizumab in combination with radiotherapy. To our knowledge, this is the first report of two individuals with advanced PDS that successfully underwent anti-PD1 treatment. By comparing the immune micromilieu to a previously published cohort, we show that the two cases are representative for PDS tumors - potentially making these results more generalizable

Comparison of in situ and extraction-based methods for the detection of MET amplifications in solid tumors

In EGFR-treatment naive NSCLC patients, high-level MET amplification is detected in approximately 2-3% and is considered as adverse prognostic factor. Currently, clinical trials with two different inhibitors, capmatinib and tepotinib, are under way both defining different inclusion criteria regarding MET amplification from proven amplification only to defining an exact MET copy number. Here, 45 patient samples, including 10 samples without MET amplification, 5 samples showing a low-level MET amplification, 10 samples with an intermediate-level MET amplification, 10 samples having a high-level MET amplification by a MET/CEN7 ratio >= 2.0 and 10 samples showing a high-level MET amplification with GCN >= 6, were evaluated by MET FISH, MET IHC, a ddPCR copy number assay, a NanoString nCounter copy number assay and an amplicon-based parallel sequencing. The MET IHC had the best concordance with MET FISH followed by the NanoString copy number assay, the ddPCR copy number assay and the custom ampliconbased parallel sequencing assays. The concordance was higher in the high-level amplified cohorts than in the low- and intermediate-level amplified cohorts. In summary, currently extraction-based methods cannot replace the MET FISH for the detection of low-level, intermediate-level and high-level MET amplifications, as the number of false negative results is very high. Only for the detection of high-level amplified samples with a gene copy number >= 6 extraction-based methods are a reliable alternative. (C) 2019 The Authors. Published by Elsevier B.V. on behalf of Research Network of Computational and Structural Biotechnology

Comparison of Blood Collection Tubes from Three Different Manufacturers for the Collection of Cell-Free DNA for Liquid Biopsy Mutation Testing

The improvement in sensitive techniques has allowed the detection of tumor-specific aberrations in circulating tumor (ct) DNA. Amplification-refractory mutation system PCR has been used for the analysis of ctDNA to detect resistance-causing mutations in the epidermal growth factor receptor gene (eg, EGFR T790M) in lung cancer patients. So far, Streck tubes have been widely used as standard blood collection tubes. Here, we compared blood collection tubes manufactured by Streck with tubes from Roche and Qiagen regarding their utility in stabilizing ctDNA in plasma samples. Venous blood from healthy donors was collected in tubes from Streck, Roche, and Qiagen. Samples were spiked with artificially fragmented EGFR T790M-mutated DNA and stored for different periods of time or spiked with different DNA amounts before plasma preparation. Extracted ctDNA was analyzed by amplification-refractory mutation system PCR. Mutant DNA, spiked into blood samples from healthy donors at quantities of 1 or 3 ng, was still reliably detectable in all samples after 7 days. EGFR T790M could be detected when spiking was performed with an amount of artificial ctDNA of 0.5 ng when tubes from Roche and Qiagen were used. Blood collection tubes from Roche and Qiagen are highly suitable for ctDNA stabilization and subsequent liquid biopsy testing. Even low ctDNA concentrations allow the detection of somatic mutations