The desire to develop and evaluate drugs as potential countermeasures for biological and chemical threats requires test systems that can also substitute for the clinical trials normally crucial for drug development. Animal models have limited predictivity for drug efficacy, as is well known from many disappointments in clinical trials. Traditional in vitro and in silico approaches are not really game changers here, but the substantial investment into novel tools now underway might bring about a second generation of alternative approaches. The avenue pursued focuses primarily on the development of a Human on a Chip, i.e., the combination of different three-dimensional (stem) cell-based organ equivalents combined with microfluidics. The prospects of such approaches, their impact on the field of alternative approaches, and necessary complementary activities are discussed. The need to adapt quality assurance measures and experiences from validation is stressed.

European Union (EU) legislation on the protection of animals used for scientific purposes requires that alternative methods must be used instead of animal tests wherever they are available. Unfortunately, this provision is not implemented to its full extent when it comes to risk assessment of chemicals and new products prior to their authorization and placing on the market in the EU. In this study, we screened data requirements of relevant EU law regarding chemicals (REACH), biocides, pesticides, and food safety (Novel Food) and found that data requirements as part of the risk assessment do not always reflect state-of-the-art science and technology. Most of the data requirements we investigated still include testing on animals for many toxicological endpoints, even though more than 40 alternative testing methods accepted at the level of the EU or the OECD are available. This may be due to a multitude of reasons, including a shortage of both manpower to implement existing knowledge and expertise in the field of alternative methods, as well as unclear and misleading statements on the applicability and state of validation of alternative methods. In conclusion, we strongly suggest a homogeneous EU-wide approach for all areas involving risk assessment of substances with the goal of better implementing the 3Rs and complying with Directive 2010/63/EU. This also would streamline data requirements, save costs on various levels, and enhance product safety for consumers.

The importance placed on risk avoidance in our society has resulted in a broad range of regulations intended to guarantee safety of products such as pharmaceuticals and chemicals. Many of these regulations rely on animal tests. As a result, about 25% of the animal experiments in Europe are done for regulatory purposes. There are many initiatives that aim to replace, reduce, or refine laboratory animal use, but the regulatory acceptance and use of 3R models lags behind. The central question of this study is: “Which variables influence the regulatory acceptance and use of 3R models and in what way?” Regulatory acceptance is seen as one of the biggest hurdles 3R models face, but the rationale behind this is still underexplored. This study is an approach to filling that gap by combining opinions from experts in the field with literature on technology acceptance and risk regulation, resulting in a model of the variables that determine the process of the regulatory acceptance and use of 3R models.

Drug formulation screenings for treatment of inflammatory bowel disease (IBD) are mostly conducted in chemically induced rodent models that represent acute injury-caused inflammation instead of a chronic condition. To accurately screen drug formulations for chronic IBD, a relevant model that mimics the chronic condition in vitro is urgently needed. In an effort to reduce and potentially replace this scientifically and ethically questionable animal testing for IBD drugs, our laboratory has developed an in vitro model for the inflamed intestinal mucosa observed in chronic IBD, which allows high-throughput screening of anti-inflammatory drugs and their formulations. The in vitro model consists of intestinal epithelial cells, human blood-derived macrophages, and dendritic cells that are stimulated by the inflammatory cytokine interleukin-1β. In this study, the model was utilized for evaluation of the efficacy and deposition of budesonide, an anti-inflammatory drug, in three different pharmaceutical formulations: (1) a free drug solution, (2) encapsulated into PLGA nanoparticles, and (3) encapsulated into liposomes. The in vitro model of the inflamed intestinal mucosa demonstrated its ability to differentiate therapeutic efficacy among the formulations while maintaining the convenience of conventional in vitro studies and adequately representing the complex pathophysiological changes observed in vivo.

Astrocytes are activated in most chronic neurodegenerative diseases associated with inflammatory events such as Parkinson’s disease or Alzheimer’s disease, but also in stroke. Due to an aging population worldwide, research efforts in these areas are likely to expand in the future. This will entail an increased demand for appropriate experimental models.

We introduce here the new immortalized mouse astrocyte cell line IMA 2.1 as an alternative to currently used primary astrocyte cultures. IMA 2.1 were directly compared with primary mouse astrocytes with respect to their response to proinflammatory stimuli, expression of typical astrocyte markers, and to the cell line’s capacity to metabolize the parkinsonian toxin MPTP to its toxic metabolite MPP+. Under inflammatory conditions, mimicked with the addition of a cytokine mix, IMA 2.1 responded similarly to primary astrocytes with mRNA upregulation, expression of iNOS and COX-2, and the release of various inflammatory mediators. Analysis of astrocytic markers indicated that IMA 2.1 represent a relatively early, GFAP-negative stage of astrocyte development. Moreover, conversion of MPTP by monoamine oxidase-B proceeded in IMA at least as quickly as in primary cells. For all endpoints investigated, the cell line IMA 2.1, derived from a single clone, delivered reproducible results over a period of several years and allowed upscaling of experiments due to its easy handling compared with primary cells.