Sunday, 24 April 2016 16:57

Berlin: Panel discussion at Urania Featured

On the occasion of the International day for the Abolition of Animal Experiments, the alliance of animal welfare policy Berlin has organized a panel discussion at Urania Berlin with interesting personalities from academia and industry.

Under the question "Research on animals - do Germany lag behind?", Dr. Robert Landsiedel, head of the unit short-term tests at BASF SE, Prof. Dr. Roland Lauster, Head of Medical Biotechnology at the Technical University of Berlin and Dr. Mardas Daneshian, coordinator and Director of the Center for Alternatives to Animal Testing Europe (CAAT Europe) spoke and debated about about important issues. The event was moderated by André Tonn, economic journalist of info radio Berlin.

The question "do Germany lag behind?" was discussed by the panellists Professor Roland Lauster, Dr. Robert Landsiedel and Dr. Mardas Daneshian. Left side: Moderator André Tonn.
Photo: Christiane Hohensee

No recognition of test strategies so far

Dr. Robert Landsiedel from BASF explained that in most cases industry is bound to the use of animal testing according to EU regulations (e.g. REACH 1907/2006) by law. For many years, BASF's department has developed replacement methods to animal testing. Replacement methods to animal experiments are not necessarily cheaper, but faster and there less of the developed test substances would be used. It is important that the test results of new methods are reproducible and relevant. However, even if all the conditions are satisfied, it is not enough for a mandatory application, because the regulatory authorities must also be convinced and that's where the problem. From the development to the inclusion in the OECD Test Guidelines it would last 10 years.

Using the example of the eye and skin irritation tests he explained that not a single test, but a battery of several different tests would be needed to obtain the necessary test results in a required quality. In a recognition process (validation) not only whether the individual tests has to be are good, but also whether the sequence of the test battery is useful.

To investigate a substance for strong eye irritation, Dr. Landsiedel and his team have developed such a test battery as replacement to rabbit use, but it was not yet fully recognized. on one hand the team has the BCOP test developed (Bovine Corneal Opacity Test, OECD Test Guideline 437), wherein the strong eye irritation / eye corrosion is measured on corneas of slaughtered cattles in a specially designed apparatus, the Opacitometer.

If the substance is highly irritating or corrosive, this can be detected exactly by the turbidity of the cornea with the device. The device was developed by BASF and is now sold by Duratec. If the substance is not highly irritating, the test strategy developed by BASF comes into play in form of a subsequent test, the EpiOcular test. Therefore the investigator can determine whether the substance is mild or moderate irritant. Only when this test result is also negative, it is concluded that the substance is not irritating.

The BCOP test was recognized already at July 26, 2013. He has been admitted to the OECD Test Guidelines as a substitute method to test for corrosive and highly irritating properties, however not the EpiOcular as the second step within a test strategy that can distinguish between mild/moderate irritating and non-irritating substances. The piOcular test is a human tissue construct (upper cover layer of tissue of the human eye). The irritant effect is measured in the form of the cytotoxicity. The cells in a microtiter plate are exposed to the test substance. In a second step a color reagent is added. The cytotoxicity can be measured by a decrease in color intensity. The test as such is recognized by OECD Test Guideline 492, but not in its function within the test strategy, but as an alternative to BCOP for eye irritation test and strong corrosion of substances that must not be classified and labeled according to the Globally Harmonized System of the United Nations.

Dr. Landsiedel presents his test strategy for eye effects of substances.
Photo: Christiane Hohensee

On the basis of extensive studies the scientist has found out that the EpiOcular test is very well suited as a second step within the test strategy to be able to distinguish between non-irritating and mildly irritating.


In contrast to the Draizé test on rabbits the two alternative methods BCOP and EpiOcular cover the entire spectrum from extreme Corrosive up to very mild irritant. If a substance is not very mild irritant at the end, it is harmless (non-irritating). The test strategy is used both for industrial chemicals, as well as household products, cosmetics and toiletries. It is much more accurate.
Scheme: Curren, Harbell & Truba (o.J.) The piOcular model: Protocol, Performance and Experiences.

According to the regulator, however, the developed replacement does does not necessarily lead to a reduction in test animal numbers, because if the BCOP test (or EpiOcular test) is negative, the eye irritation test has to be carried out on rabbits, says Dr. Landsiedel. The regulatory authorities could not be convinced yet, despite the logic and the test results of BASF. According to the final stage of Chemicals being registered by REACh with means a consumption of countless rabbits - despite the presence of several single replacement tests.

The same issue occurs with another test strategy in the field of skin sensitization, which could save the lives of a large number of guinea pigs and mice: Again, the team led by Dr. Landsiedel developed a test strategy, with which a possible sensitizing substance property can be determined by several tiered testing steps. For this, the team has received the Animal Protection Research Award of the Federal Ministry of Food and Agriculture in 2013 (InVitro+Jobs reported here). Of the three graded individual tests the third (the so-called h-CLAT) as well as the testing strategy has still not been recognized by the regulatory authorities. The animal experiments on sensitization (so-called Local Lymph Node Assay (LLNA)) has only a prediction of 89%, the application of the tiered test strategy with the 3 tests has a prediction of 94% - an unbeatable value. As long as the test strategy is not recognized as a replacement method, the industry will continue to perform to the LLNA on mice.

A high demand for Organ-on-a-chip from industry

Professor Roland Lauster of the Technical University of Berlin informed about multi-organ reactors in chip format, which have the potential to replace animal experiments on a large scale. By the fact that the human organs are made up of countless smallest functional units, it is possible to breed them on a miniature scale 1:100,000 on the chip thus both to study the initiation of organogenesis and reaction on chemicals and phamaceuticals.

This smallest scale need also not a real blood vessel system, because the metabolism successes by diffusion. The heart is simulated by a three-diaphragm pump system to maintain the medium in the circuit. In future the researchers want to mimic all the major organs from induced pluripotent stem cells from one individual donor, thereby making it possible to integrate blood and lymph systems. The developers to whom also belong Dr. Uwe Marx and his team from TissUse (InVitro+Jobs reported here), are the European Leader with their organ-on-a-chip systems. There is a high demand for the miniatured systems from industry. Numerous well-known pharmaceutical companies have asked if it is possible to test well-characterized substances with the new chip systems. If the results are reliable, they want a changeover to the new, non-animal methods.

A controversial issue was presented by Dr. Mardas Daneshian from the Center for Alternatives to Animal Testing (CAAT). He feared that Germany is on the way to lose the connection to the development of alternative methods to animal testing. He presented examples that animal test results were not reproducible. The animal experiments are simply not safe and therefore Industry as well as Academia need global alternatives to animal testing. The pharmaceutical industry would have lost E 330 billions by high investments in the animal studies and subsequent failures. Therefore it would be a great interest in new methods to replace animal testing. The new vision of toxicology in the 21st century has the goal to evaluate questionable substances by in vitro and in silico methods thereby reducing the use of animals rapidly.

Future developments of human-on-a-chip models, analysis of Adverse Outcome Pathways and, for instance, system toxicological, virtual approaches: Such a complex knowledge need appropriate funding, which lacks in Germany. Compared to the budget for research on animals and the DFG budget of 1.92 billion the support of replacement method to animal experiments is comparatively low. UK and the United States care for much more funding. For stem cell research around 2 billion dollars are made available in the United States, for the human-on-a-chips, ToxCast and ToxCast Data is more than 305 million dollars avialable. In Europe, the Horizon 2020 to promote research into replacement methods to animal use in the field of long-term toxicity and developmental toxicity is only 30 million euros.

Possible causes were discussed and less the policy than then people in the authorities as a brakeman identified. Creativity would arise only when a certain amount of capital would be available, warned Dr. Daneshian.