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The separation of bio-suspensions, such as blood, is of great interest for many biomedical applications, such as the extraction of leucocytes from whole blood, the laboratory analysis of specific cells or a mixed cell suspensions from bioreactors or the preparation of cellular products for cellbased therapy. In particular, mechanical separation processes have been established for the rapid separation of biosuspensions. The principle of mechanical separation is based on the specific density of the particles or the particle size. Thus, the corresponding processes are based on centrifugation and/or filtration. The standard process to prepare blood products from whole blood donations contains several steps. First the blood is separated into different phases by centrifugation and then further separated by filters containing specific filtration layers. The shortcomings of this process is the low yield due to bio-particles remaining in the filter and the high costs for equipment due to the use of multiple devices for centrifugation and filtration. Here we present the proof of principle of an adaptive, size-selective filter that addresses the above mentioned shortcomings. The filter is designed to combine the filtration and centrifugation process within one step. Therefore a novel filtration device was designed that can be used within a standard centrifugation tube. During centrifugation, the centrifugal force changes the filtration coefficient of the filter. This can be used to separate different sized of bio-particles at different centrifugation speeds. Moreover, the cells that were hold back in the filter can be removed afterwards by increasing the centrifugation speed. In this way, the filter is created to gradually release cells of a specific size yielding in a high purity of different cell fractions.

Radiopharmaceuticals can be used for targetspecific functional diagnostics, such as PET or SPECT imaging, or radionuclide therapy of diseased tissue, depending on the incorporated radionuclide. Following initial in vitro testing, radiopharmaceutical candidates are usually further characterized in small animals. Since reduction, replacement and refinement (3R) of animal testing is a central precept in preclinical research it would be beneficial to replace at least some of these tests by alternative methods. Using micro physiological system technology, various organ-on-chip models can be created with human cell systems/organoids, which are operated in a circulatory system under defined physiological conditions. Here we present first attempts to introduce micro physiological systems for evaluating radiopharmaceuticals using the radiolabeled anti-EGFR antibody cetuximab as reference compound. In a micro physiological system equipped with six 96-well plate-like microwells in a flow chamber, binding of 64Cu and 68Ga-labeled cetuximab to cells and spheroids grown from A431 (EGFR-positive) and MDA-MB435S (EGFR-negative) cells was measured and compared to conventional microplates. Specific saturation binding of radiolabeled cetuximab at increasing concentrations was analyzed using a phosphor imaging system. The affinity of radiolabeled cetuximab towards A431 spheroids measured in the micro physiological system was in the same range as that of the spheroids in conventional microplates. Within the assays in micro physiological systems, the results showed a trend towards increased affinity for A431 monolayers compared to the spheroids. The values of binding capacity for radiolabeled cetuximab on 2D and 3D A431 cell culture models were in the same order of magnitude when measured in micro physiological systems or in microplates. Building on these first promising results, the work will continue on MPS modules containing advanced human spheroid/ organoid models.

Antibody-mediated rejection after kidney transplantation is a major limitation for long-term survival of donated kidneys within the recipient allograft. With the existing HLA cross-matching test only complement activating donor specific antibodies are evaluated. Other measurements are more sensitive, but cannot assert the clinical relevance of the donor specific antibodies. Thus, novel methods that address the more complex nature of cellular interactions with serum components are needed to improve antibody-mediated rejection prediction. Recent studies suggest, that the interaction of natural killer cells of the recipient with endothelial cells of the donor is of major interest. Here we present a microfluidic cell culture device and a corresponding assay to test for the relevance of natural killer cell activated antibody-mediated rejection in vitro. Therefore, a protocol to co-cultivate both cell types and track natural killer cell activated killing within a fluorescence-based assay was established. To automate this approach, the prototype of a well plate sized microfluidic device was developed, that enables staining of the two cell types and repeatable and timedependent co-cultivation of the cells with serum. This automated approach enables further evaluation of the clinical relevance of the established assay paving the way to additional information on HLA cross-matching that helps to quickly estimate the probability of Antibody-mediated rejection and the clinical relevance of donor specific antibodies.

Newly developed materials for blood-contacting devices need to undergo hemocompatibility testing to prove compliance with clinical requirements. However, many current in vitro models disregard the influence of flow conditions and blood exchange as it occurs in vivo. Here, we present a flow model which allows testing of blood-surface interactions under more physiological conditions. This modular platform consists of a triple-pump-chip and a microchannel-chip with a customizable surface. Flow conditions can be adjusted individually within the physiological range. A performance test with whole blood confirmed the hemocompatibility of our modular platform. Hemolysis was negligible, inflammation and hemostasis parameters were comparable to those detected in a previously established quasi-static whole blood screening chamber. The steady supply of fresh blood avoids secondary effects by nonphysiological accumulation of activation products. Experiments with three subsequently tested biomaterials showed results similar to literature and our own experience. The reported results suggest that our developed flow model allows the evaluation of blood-contacting materials under physiological flow conditions. By adjusting the occurring wall shear stress, the model can be adapted for selected test conditions.

Current in vitro and in vivo disease models have been reported to lack sufficient translation to human. Precision-Cut Lung Slices (PCLS) are viable sections of lung tissue and have been described to be a translational model for the ex vivo assessment of pharmacological and toxicological compounds. In most studies PCLS were cultured under static conditions. These lung sections, however, suffer from the limited viability. Here we present a novel modular microphysiological system (MPS) to prolong the cultivation of ex vivo lung tissue. A tailored MPS setup was designed using the PDMS free modular plug&play MPS construction kit. PCLS from mice were cultivated for up to one week under static versus perfused conditions. Using the MPS technology enabled a prolonged culture period with improved viability as shown by lowered lactate dehydrogenase release and improved membrane integrity. Using this technology might allow us to use PCLS for longer culture periods such as e.g. repeated dose toxicity or pharmacology studies.

Aortic valve (AV) stenosis is characterized by tissue fibrosis and calcification. Fibrous thickening can result in reduced tissue oxygen supply leading to pathological valvular interstitial cell (VIC) differentiation and calcification. Static 2D VIC cultures and animal models are limited in the ability to reflect human AV calcification. Culturing of VICs in micro-physiological-systems (MPS) in a pulsatile flow and the establishment of a modular AV tissue incubation chamber (TIC) are new approaches to evaluate pathophysiological processes of AV disease. Therefore, a MPS able to adjust hypoxic conditions was applied for VIC culture. A significant increase of mRNA-expression of EGLN1 and HIF1α- regulated LDHA and HIF1α nuclear localisation were proven under hypoxia. AV tissue culture was established within a TIC and viability was monitored by Resazurin-reduction in the incubation medium and visualized by LDH-activity in tissue cryosections. Viability was compared between fluid and static incubated tissues revealing an advantageous effect of the fluidic assay condition. Consecutively, the application of MPS in AV research allows i) the investigation of VIC cultures with efficient oxygen regulation and ii) the culture of porcine or human AV tissues preserving viability and specifically reflecting in vivo parameters. These methods open up new possibilities beyond static 2D culture and facilitate a reduction of animal experiments in AV research.

Within the human blood immune cells assume specific functions to modulate and control the human immune response. Consequently, they are used for the diagnosis and treatment of autoimmune and hematologic diseases. Traceless Affinity Cell Selection Technology (Fab-TACS® technology) is a new method for selective isolation of specific immune cells from blood samples. Currently, the technology is used in a 60 mL syringe-like cartridge, which is operated by a control unit (Fabian®). Nevertheless, diagnostic applications require the reduction of the sample volume to several microliters. Therefore, we developed a cartridge for the purification of small blood volumes with the aid of suitable scaling and simulation methods. By implementing a multi domain physical systems model of the established cartridge we could simulate the separation process. Based on that a microfluidic cartridge was designed and several prototypes were manufactured. To operate the microfluidic cartridge a setup and software for fluidic control were created. Afterwards the prototype cartridge and the microfluidic control were used to isolate lymphocytes from a buffy coat. By flow cytometric analysis we could show that it is possible to isolate lymphocytes with a purity that is comparable to the established 60 mL syringe-like cartridge.

Despite disadvantages, such as high cost and their poor predictive value, animal experiments are still the state of the art for pharmaceutical substance testing. One reason for this problem is the inability of standard cell culture methods to emulate the physiological environment necessary to recapitulate in vivo processes. Microphysiological systems offer the opportunity to close this gap. In this study, we utilize a previously employed microphysiological system to examine the impact of pressure and flow on the transportation of substances mediated by multidrug resistance protein 1 (MDR1) across an artificial cell-based tubular barrier. By using a miniaturized fluorescence measurement device, we could continuously track the MDR1-mediated transport of rhodamine 123 above the artificial barrier over 48 h. We proved that applying pressure and flow affects both active and passive transport of rhodamine 123. Using experimental results and curve fittings, the kinetics of MDR1-mediated transport as well as passive transport were investigated; thus, a kinetic model that explains this transport above an artificial tubular barrier was identified. This kinetic model demonstrates that the simple Michaelis–Menten model is not an appropriate model to explain the MDR1-mediated transport; instead, Hill kinetics, with Hill slope of n = 2, is a better fit. The kinetic values, Km, Vmax, and apparent permeability (Papp), obtained in this study are comparable with other in vivo and in vitro studies. Finally, the presented proximal tubule-on-a-chip can be used for pharmaceutical substance testing and to investigate pharmacokinetics of the renal transporter MDR1. [ABSTRACT FROM AUTHOR]

Background: The efficacy and safety of opicapone, a once-daily catechol-O-methyltransferase inhibitor, have been established in two large randomized, placebo-controlled, multinational pivotal trials. Still, clinical evidence from routine practice is needed to complement the data from the pivotal trials. Methods: OPTIPARK (NCT02847442) was a prospective, open-label, single-arm trial conducted in Germany and the UK under clinical practice conditions. Patients with Parkinson's disease and motor fluctuations were treated with opicapone 50 mg for 3 (Germany) or 6 (UK) months in addition to their current levodopa and other antiparkinsonian treatments. The primary endpoint was the Clinician's Global Impression of Change (CGI-C) after 3 months. Secondary assessments included Patient Global Impressions of Change (PGI-C), the Unified Parkinson's Disease Rating Scale (UPDRS), Parkinson's Disease Questionnaire (PDQ-8), and the Non-Motor Symptoms Scale (NMSS). Safety assessments included evaluation of treatment-emergent adverse events (TEAEs) and serious adverse events (SAEs). Results: Of the 506 patients enrolled, 495 (97.8%) took at least one dose of opicapone. Of these, 393 (79.4%) patients completed 3 months of treatment. Overall, 71.3 and 76.9% of patients experienced any improvement on CGI-C and PGI-C after 3 months, respectively (full analysis set). At 6 months, for UK subgroup only (n = 95), 85.3% of patients were judged by investigators as improved since commencing treatment. UPDRS scores at 3 months showed statistically significant improvements in activities of daily living during OFF (mean ± SD change from baseline: − 3.0 ± 4.6, p < 0.0001) and motor scores during ON (− 4.6 ± 8.1, p < 0.0001). The mean ± SD improvements of − 3.4 ± 12.8 points for PDQ-8 and -6.8 ± 19.7 points for NMSS were statistically significant versus baseline (both p < 0.0001). Most of TEAEs (94.8% of events) were of mild or moderate intensity. TEAEs considered to be at least possibly related to opicapone were reported for 45.1% of patients, with dyskinesia (11.5%) and dry mouth (6.5%) being the most frequently reported. Serious TEAEs considered at least possibly related to opicapone were reported for 1.4% of patients. Conclusions: Opicapone 50 mg was effective and generally well-tolerated in PD patients with motor fluctuations treated in clinical practice. Trial registration: Registered in July 2016 at clinicaltrials.gov (NCT02847442). [ABSTRACT FROM AUTHOR]

Introduction: Effects of antiepileptic drug (AED) load changes in patients with focal seizures have not been well evaluated. Methods: SP1065 (NCT01673282) was a noninterventional, prospective, observational study conducted in a clinical practice setting. Patients (aged ≥18 years) with focal seizures were enrolled within 7 days of being prescribed adjunctive lacosamide. Observation period was ~6 months. Drug load was assessed using percentage change in ratio of actual prescribed dose and World Health Organization defined daily dose (DDD) for concomitant AEDs and all AEDs (including lacosamide). Subgroups were defined for patients with at least one concomitant sodium channel–blocking AED (SCB [+]) and those without (SCB [−]). Results: A total of 311 patients were assessed for safety, 302 for measurement of drug load, and 240 for effectiveness. Ratio of AED dose to DDD decreased for concomitant AEDs (−9.6%) and increased for all AEDs (including lacosamide; 15.5%). Median reduction in focal seizure frequency per 28 days was 100% (range: −100, 2275.8). 70.4% and 61.7% of patients had a ≥50% or ≥75% reduction in seizure frequency, respectively; 50.8% became seizure‐free. In the SCB (+) subgroup (n = 149), ratio of AED dose to DDD decreased for concomitant AEDs (−15.0%) and increased for all AEDs (10.7%). In the SCB (−) subgroup (n = 153), ratio of AED dose to DDD decreased for concomitant AEDs (−4.4%) and increased for all AEDs (20.2%). Fifty‐seven patients (18.3%) reported ADRs, most commonly dose >400 mg/d (7.1%). Seventeen patients (5.5%) had ADRs leading to discontinuation. Significance: Addition of lacosamide resulted in reduction of concomitant AED drug load regardless of whether concomitant AEDs were SCB (+) or SCB (−). These results indicate that addition of lacosamide in patients with focal seizures could allow clinicians to withdraw or reduce the dose of less well‐tolerated or less effective AEDs. [ABSTRACT FROM AUTHOR]

A series of diphosphoramidites has been synthetized with a piperazine, homopiperazine, and an acyclic 1,2-diamine unit in the backbone. New compounds were tested alongside related N-acyl phosphoramidites as ligands in the Rh-catalyzed hydroformylation of n-octenes to investigate their influence on the activity and regioselectivity. A subsequent study of their hydrolysis stability revealed that the most stable ligands induced the highest activity in the catalytic reaction. [ABSTRACT FROM AUTHOR]

This symposium reviewed the issues of non-oral therapy in the late stage Parkinson's disease (PD). The accepted standard treatment of PD is oral levodopa or oral dopamine agonists. However, the long-term complications and limitations of this treatment might be improved by changing therapy from the present pulsatile stimulation to a more constant stimulation of central dopamine receptors. Stimulation of these receptors may be possible with non-oral drug delivery treatments. Many of these non-oral options have been evaluated during the last few decades to find a more continuous drug delivery. The non-oral treatment options include invasive measures such as intraduodenal levodopa, subcutaneous apomorphin and most recently, the non-invasive transdermal (patch) delivery system, with the novel dopamine agonist rotigotine (Aderis Pharmaceuticals Inc.). The benefits of the non-oral, more continuous dopaminergic treatment of PD needs to be demonstrated in clinical trials and long-term clinical practice, before they can be considered as potential replacements of the standard oral therapy.