Patients with recurrent or metastatic squamous cell head and neck cancer (R/M SCHNC) exhibit a poor prognosis with a median overall survival (OS) time of <1 year. Platinum-based chemotherapy with or ...without cetuximab has been the standard of care in the last decade. The aim of the current retrospective study was to evaluate the outcome and tolerability of treatment in patients with R/M SCHNC receiving platinum/5-fluorouracil/cetuximab (PFE) chemotherapy compared with platinum/5-fluorouracil (PF) chemotherapy in daily clinical practice. A retrospective analysis was performed using the data of patients treated at the Institute of Oncology Ljubljana between April 2008 and May 2018. Progression-free survival (PFS) and OS were calculated with the Kaplan-Meier method and compared with the log-rank test. Multivariate regression Cox analysis was used to determine independent prognostic factors. A total of 67 patients were treated at the aforementioned Institute: 34 patients received the PF and 33 the PFE regimen. The mean age of patients was 54.6 years and 91% of patients were male. Median PFS time was 6.6 vs. 7.1 months for the PF vs. PFE groups, respectively (P=0.852). Median OS time was 9.6 vs. 11.5 months for the PF vs. PFE groups, respectively (P=0.029). The prognostic factor for PFS was partial remission hazard ratio (HR), 0.32; 95% CI, 0.15-0.70; P=0.004. Prognostic factors for OS were partial remission (HR, 0.15; 95% CI, 0.06-0.38; P<0.001) or stable disease (HR, 0.28; 95% CI, 0.13-0.64; P=0.002), and a subsequent line of treatment upon progression (HR, 0.28; 95% CI, 0.15-0.52; P<0.001). In the PFE group, 15.4% of patients had a grade >2 infusion reaction to cetuximab and 27.3% had grade 3 skin rash. There were no differences in diarrhoea, hypomagnesaemia, infections and febrile neutropenia; however, the mortality on active treatment was high (13.4%). In conclusion, patients treated with PFE had similar PFS, but improved OS compared with patients treated with the PF protocol. The proportion of patients who died under treatment due to disease progression and toxicity was high in both treatment arms. A thorough selection of patients for this treatment is crucial. Key words: cetuximab, platinum-based chemotherapy, overall survival, progression-free survival, prognostic factors, safety
V Ljubljani sta med 6. in 8. 11. 2019 potekala 54. srečanje in javni posvet ALTE (Association of Language Testers in Europe). Srečanje na temo Enojezično testiranje v večjezični realnosti: jezikovne ...ideologije in njihov vpliv na jezikovno testiranje sta organizirala Univerza v Ljubljani, Filozofska fakulteta in njen Center za slovenščino kot drugi in tuji jezik pri Oddelku za slovenistiko. V tem okviru je 8. 11. 2019 potekala okrogla miza (Bližnja) srečanja oblikovalcev jezikovne politike. Objavljamo zapis posnetka pogovora sodelujočih na dogodku.
The aim of our prospective study was to quantify phosphate removal during long nocturnal high‐flux hemodialysis or hemodiafiltration (HD) with total dialysate collection. Eight patients (two women) ...were studied for the first‐in‐the‐week HD session that lasted 7–8 h. Total dialysate was collected. Serum and collected dialysate phosphate concentrations were measured every hour. Phosphate removal as assessed from the serum concentration was most important during the first 2 h of HD, and then a plateau was reached. The highest average phosphate concentration in the total dialysate was in the 1st hour, thereafter the concentration decreased but remained stable. The average total removed mass of phosphate quantified from hourly collected dialysate was 5195.7 ± 1898 mg. Phosphate had been removed in a consistent manner during the whole duration of nocturnal HD as assessed through dialysate (despite stable serum phosphate concentration). This could indicate phosphate transfer from intracellular space. The total removed phosphate quantified from the total dialysate collection was higher than previously reported and exceeded the normal phosphate food intake.
•Caco-2 cell model was used for assessing transepithelial permeability to proteins.•Similar Papp values of around 4×10−10cm/s were measured for four model proteins.•Addition of ...N-decyl-β-d-maltopyranoside considerably increased permeability to erythropoietin.•Paracellular route of permeation is suggested.•We showed suitability of Caco-2 model for enhanced protein permeability studies.
Appropriate assessment of transepithelial permeability in vitro is needed to estimate and model trans-mucosal bioavailability to achieve oral delivery of protein biopharmaceuticals. The Caco-2 cell-based intestinal epithelium model is widely used for this purpose for low molecular mass drugs. The aim of this study was to test the suitability of the Caco-2 model for assessing enhanced transepithelial permeability to proteins. Four unrelated proteins were chosen to test the permeability of Caco-2 monolayers. It was found that proteins could cross the epithelium model, in spite of their size. All tested proteins had similar very low apparent permeability coefficients (Papp) of around 4×10−10cm/s. Protein stability over three-hour exposure to Caco-2 was also confirmed. Their crossing rate in a cell-free setup was also measured, to determine the upper limit of permeability to proteins. An epithelium permeability enhancer N-decyl-β-d-maltopyranoside (MP C10) was used to demonstrate accelerated permeability conditions. Papp values could be increased dose dependently up to about 1×10−7cm/s, close to the level in the cell-free setup, indicating distinctive potential of the model. This along with enhancing effect of known specific route permeators suggests involvement of the paracellular route in protein transport. Our results thus indicate that the Caco-2 model is a suitable tool for in vitro assessment of enhanced permeability to proteins.
Keratin 8 and 18 (K8/K18) mutations have been implicated in the aetiology of certain pathogenic processes of the liver and pancreas. While some K8 mutations (K8 G62C, K8 K464N) are also presumed ...susceptibility factors for inflammatory bowel disease (IBD), the only K18 mutation (K18 S230T) discovered so far in an IBD patient is thought to be a polymorphism. The aim of our study was to demonstrate that these mutations might also directly affect intestinal cell barrier function. Cell monolayers of genetically engineered human colonocytes expressing these mutations were tested for permeability, growth rate and resistance to heat-stress. We also calculated the change in dissociation constant (Kd, measure of affinity) each of these mutations introduces into the keratin protein, and present the first model of a keratin dimer L12 region with in silico clues to how the K18 S230T mutation may affect keratin function. Physiologically, these mutations cause up to 30% increase in paracellular permeability in vitro. Heat-stress induces little keratin clumping but instead cell monolayers peel off the surface suggesting a problem with cell junctions. K18 S230T has pronounced pathological effects in vitro marked by high Kd, low growth rate and increased permeability. The latter may be due to the altered distribution of tight junction components claudin-4 and ZO-1. This is the first time intestinal cells have been suggested also functionally impaired by K8/K18 mutations. Although an in vitro colonocyte model system does not completely mimic the epithelial lining of the intestine, nevertheless the data suggest that K8/K18 mutations may be also able to produce a phenotype in vivo.
We present the potential of inclusion bodies (IBs) as a protein delivery method for polymeric filamentous proteins. We used as cell factory a strain of E. coli, a conventional host organism, and ...keratin 14 (K14) as an example of a complex protein. Keratins build the intermediate filament cytoskeleton of all epithelial cells. In order to build filaments, monomeric K14 needs first to dimerize with its binding partner (keratin 5, K5), which is then followed by heterodimer assembly into filaments.
K14 IBs were electroporated into SW13 cells grown in culture together with a "reporter" plasmid containing EYFP labeled keratin 5 (K5) cDNA. As SW13 cells do not normally express keratins, and keratin filaments are built exclusively of keratin heterodimers (i.e. K5/K14), the short filamentous structures we obtained in this study can only be the result of: a) if both IBs and plasmid DNA are transfected simultaneously into the cell(s); b) once inside the cells, K14 protein is being released from IBs; c) released K14 is functional, able to form heterodimers with EYFP-K5.
Soluble IBs may be also developed for complex cytoskeletal proteins and used as nanoparticles for their delivery into epithelial cells.
The keratin filament cytoskeleton is vital to the normal function of epithelial cells. It provides structural support and regulates different aspects of cell metabolism. Mutations in keratins 5 and ...14 cause a skin fragility disorder, epidermolysis bullosa simplex (EBS). Patients with severe EBS have an increased cumulative risk for basal cell carcinoma. In this study, we tested how keratin 5 and 14 mutant EBS patient-derived keratinocytes behave in the face of two different types of stressors that are able to induce cell death: ionizing radiation and cytokines TNF-α and TRAIL. The data point out to a substantial difference between how normal and keratin mutant keratinocytes deal with such stresses. When case of DNA damage, the ATM/Chk2-pathway is one of the two main tracks that can prevent the progression of mitosis and so allow repair. This was altered in all investigated keratin mutants with a particular down-regulation of the activated form of checkpoint kinase 2 (pChk2). Keratin mutants also appear less sensitive than normal cells to treatment with TNF-α or TRAIL, and this may be linked to the up-regulation of two pro-survival proteins, Bcl-2 and FLIP. Such changes are likely to have a profound effect on mutant keratinocytes ability to survive and withstand stress, and in theory this may be also a contributing factor to cell transformation.
As an in vitro model system, patient-derived epidermolysis bullosa simplex keratinocytes have had an immense impact on what we know today about keratin filament function and their role in disease ...development. In the absence of gene therapy, screening compound libraries for new or better drugs is another approach to improve existing treatments for genodermatoses. However in this study, we report of the potential pitfalls when using this type of cell lines as a “reporter” system. When cell lines with different genetic backgrounds are being used in cell-based assays, the greatest obstacle is to determine the most appropriate culture conditions (i.e., the composition of medium, number of cells plated and number of days in culture). We demonstrate how culture conditions can greatly interfere with the cellular response in cell-based assays (cell proliferation, metabolic activity and migration), potentially also giving rise to misleading data.
Microplastics in the environment occur in different sizes and shapes and are made of various polymers. Therefore, they also considerably differ in their properties and ecotoxicity. However, the ...majority of microplastics research uses pre-made spherical microplastics, which practically do not exist in the environment. Our work focused on a comprehensive study of six different types of microplastic that were prepared to simulate common microplastics found in the environment. All types of microplastics where chemically and physically characterized using Fourier-transform infrared spectroscopy, thermal analysis, field-emission scanning electron microscopy, optical microscopy and laser diffraction analysis. The specific surface area was determined using the BET method. Furthermore, effects of microplastics and microplastic leachates on a common duckweed (Lemna minor) were evaluated. All tested microplastics did not affect specific growth rate and chlorophyll a content in duckweed, while microplastics with a rough surface and sharp edges caused a significant reduction of duckweed root length. Microplastics made of Bakelite also showed an intensive leaching, which increased their ecotoxicity potential. Natural particles used as a control did not have any negative effect on duckweed. Overall, microplastic particles have significantly different ecotoxicity profiles depending on their physico-chemical properties. Therefore, the testing of environmentally relevant particles and their proper characterization, as well as the testing of microplastic leaching properties, is crucial for understanding of microplastics ecotoxicological potential.
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•MPs with rough surface and sharp edges had negative impact on duckweed's root length.•MPs made of Bakelite had negative impact on root length due to leaching of chemicals.•Natural particles, used as control, showed no negative impact on duckweed.
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