Dengue; A tropical disease
Dengue fever is caused by the dengue virus. All over the tropics and subtropics, hospitals are flooded with infected patients during the rainy season.
Dengue fever
The rainy season is a time with high mosquito activity. It is not surprising, therefore, that most dengue infections occur during this time of the year. Although there is no adequate treatment or medication for dengue fever, proper medical care increases the odds of survival. Every year, an estimated 500,000 people are hospitalised with severe cases of dengue fever, sometimes with life-threatening complications. As a result, local hospitals are full of dengue patients during an epidemic. A medicine to treat dengue fever would be a giant leap forward for medical care in the tropics. Since there are no animal-free techniques available, new drugs are tested in animals at some point during the experimental phase.
Second dengue virus infection leads to increased risk of complications
There are four sub-types of the dengue virus, which are all quite similar. Being infected with one particular sub-type prevents a subsequent infection with that same sub-type. However, a second infection with a different sub-type leads to an increased risk of illness and dengue-related complications. Scientists refer to this phenomenon as 'antibody-mediated enhancement'.
Vaccine to prevent infection
A vaccine is the most effective way to prevent infection and spreading of an infectious disease. Usually, a vaccine teaches the immune system to recognise and attack a particular pathogen. For dengue virus, however, things are a bit different.
There is, in fact, a commercially available dengue virus vaccine, but it is not advised for everyone. In extreme cases the vaccine can actually aggravate dengue virus infection due to the antibody-mediated enhancement phenomenon mentioned above. It is for that reason that the World Health Organization (WHO) only recommends the vaccine to be used by people who live in an area where dengue virus is commonly present. In that case the vaccine protects from a second infection. The vaccine is not recommended for young people or tourists. Scientists from all over the world are working to develop better and safer dengue vaccines that can be used by everyone.
Antiviral drugs
Another way of preventing serious complications caused by dengue virus is to specifically inhibit the virus – for instance, with antiviral drugs that prevent viral reproduction. So far there is no antiviral drug to inhibit dengue virus.
BPRC research
BPRC evaluates experimental dengue virus vaccines and medication in what is called a dengue virus infection model.
We vaccinate monkeys with the dengue vaccine. After a few weeks, we draw a little blood to test it on dengue-specific antibodies. The presence of these antibodies is a first indication the vaccine may be effective. To test if the antibodies indeed prevent dengue virus infection, we expose the animals to the virus. This is done under controlled conditions to ensure reproducibility.
Rhesus macaques typically do not show clinical symptoms after dengue virus infection. As such, the amount of virus in the blood is an important indicator for the severity of infection. To determine the viral load over time, we draw a small blood sample at regular time points after exposure.
Just like any other scientific experiment, an animal study requires proper controls. Since a virus consists of genetic material surrounded by a protein shell, it cannot exist on its own. Although a virus is not a living being, it can die. In order to prevent drawing false conclusions from an animal study – for example based on dead virus instead of live virus – we include 'control animals' in each study. These animals are not vaccinated before being exposed to the virus.
The efficacy of an experimental vaccine is determined by comparing the viral load in vaccinated animals and control animals. The results of these ‘proof of principle’ studies largely determine if a vaccine candidate will be further developed for human use, or not.
Potential new medication is tested in a similar way, and here too the viral load in the blood and organs is an important factor. The difference between treated and untreated animals may indicate how well a particular medicine works.
The 3Rs in dengue virus research
Anyone who works with laboratory animals in Europe is required to explain why their research cannot be performed in any other way. Can it be done without using laboratory animals? Or can it be done using fewer animals? Which techniques are available to minimise the discomfort for the laboratory animals? In other words: are the 3Rs implemented?
Replacement
A vaccine is designed to activate an immunological memory. To evaluate an experimental vaccine, a fully functional immune system is required. In the case of the dengue virus, we currently do not know how this memory is generated and what mechanisms in the immune system are responsible. As such, there is currently no animal-free method to study the interaction between a dengue virus vaccine and the immune system.
We can only test experimental medication for dengue virus using animals that are susceptible to the virus. We know that the virus is able to reproduce in rhesus monkey blood, which is why we are using these animals in order to test experimental medication.
Reduction
Only the most promising vaccine candidates are evaluated in animals. Numerous animal-free techniques are used to prevent ineffective or less effective vaccines from being tested in animals. Combining several vaccine candidates in one study also reduces the number of animals needed.
Refinement
The Netherlands National Committee for the protection of animals used for scientific purposes uses the following definition for the refinement of animal testing: the refinement of animal testing aims to reduce the discomfort of laboratory animals and/or to optimise laboratory animal welfare.
Good living conditions are a key part of our animal welfare policy. At BPRC, animals are always housed in social groups and never alone, and we have an extensive environmental enrichment programme. Furthermore, the animals are trained to reduce stress during experiments.
The rise of HIV and the unprecedented progress in biomedical science go hand in hand, both in terms of technology and animal welfare. As a result, much more data can be collected per animal and their discomfort is minimised.