In-depth article: De-risk your drug development at an early stage

In preclinical safety studies, investigating whether a compound activates the complement system may prevent unexpected adverse effects at a later stage of the development process. The use of whole human blood yields the most comprehensive picture, but in practice it involves some challenges. Thanks to a specially developed protocol, Sanquin PBS is able to conduct whole blood assays, including assessment of the complement activity. This helps pharmaceutical companies optimize their lead selection process.

When a compound is tested for safety reasons, one aim is to rule out undesirable effects of the immune system upon interaction with the compound. If this is assessed early on in the development phase, optimization of the lead can prevent adverse events later on, thereby saving time and money. The most common approach is to examine cytokine levels in the blood as a measure for immune activation. Cytokine release is, however, only one part of the equation, reflecting cellular activation. A compound may also activate the complement system: a vital, humoral part of the immune system with an activation response within seconds. Complement activation may contribute to a compound’s adverse effects. Sanquin PBS is one of the few partners worldwide that can screen compounds on a large scale in the discovery and early preclinical phases for cytokine production as well as  complement activation.

Dr. Annelies Turksma heads the Sanquin Immunomonitoring lab. “What makes immunology such an interesting field of research is that all the different cells, proteins and other substances in our immune system are working together.” Likewise, she is working together with complement experts Dr. Kyra Gelderman and Dr. Richard Pouw. “Because we can also study the complement system in the preclinical phase, we can detect additional potential adverse effects of a new compound at an early stage. Particularly since we can simultaneously look at cytokine production and compare the results, we are able to conduct a valuable overall risk assessment.”

Whole blood assays, mimicking real life

For the performance of both cytokine release assays and complement assays, whole human blood is strongly preferred. The reason for this? In-vitro whole blood assays mimic the biological processes as closely as ex-vivo. As the term ‘whole blood’ indicates, because all cells and proteins in whole blood remain in their original environment, it is possible to study their interactions – something we cannot do when performing assays with isolated cells, plasma or serum.

Bridge between the innate and the adaptive immune system 
The complement system is part of the innate immune system and consists of a group of proteins that are either circulating in plasma or attached to a cell. The complement system can be activated quickly and can cause a trigger in both innate and adaptive immune cells. For example, the complement plays a major role in removing bacteria by marking them for phagocytosis, while also attracting immune cells to the infection site.

The challenges

The performance of whole blood assays involves many challenges, which demand highly specific knowledge and experience. For example, the time of incubation is vital to ensure reliable results, as is good knowledge of the relevant positive and negative controls. Since all people are different, compounds may interact differently with blood samples of different individuals. At Sanquin PBS we have in-house expertise in this area, as well as access to a large and diverse supply of blood samples. And if we notice something of interest, we can zoom in. Thanks to the fact that all specialized areas of immunology are represented in our team, we can investigate specific cells and processes in depth to unravel the mechanisms of action. 

The need for an adjusted blood collection protocol 

The complement system reacts rapidly and amplifies itself. Therefore, it is vital that the samples are handled correctly immediately after the blood draw to prevent false-positive results. Complement activation markers are usually measured in plasma, but in order to include the effect of the cellular compartment, whole blood should be used. However, the anticoagulant additives commonly used in blood draws for cytokine production assays block activation of the complement system. Fortunately, there are special anticoagulants that do not inhibit complement activation. 

What it brings to Pharma

We recently acquired data to support the safety studies of a large pharmaceutical company for their vaccine compound that is based on a common cold virus. “Our client is particularly interested in whether the administration of their viral compound can have undesirable effects on the immune system if people have already been infected with the common cold virus and have produced antibodies,“ Turksma says. “They were looking specifically for a partner that not only has the necessary knowledge of white blood cells and antibodies, but also of the complement system and whole blood assays. Furthermore, they did not want to choose from a catalogue of assays, but were looking for a partner that could create a solution tailored to their requirements. We were able to offer this, and together with the client our in-house experts on the complement system developed a new assay.”

“We were able to show the presence and the influence on complement activation of virus-specific antibodies and gain insight into virus-induced cytokine production,” Turksma concludes. “This information offers valuable starting points for the general risk assessment regarding the therapeutic use of viral-based therapies that are currently being investigated by many pharmaceutical companies.”

If you would like to know more about complement or whole blood assays, please contact our experts Dr. Annelies Turksma, Dr. Kyra Gelderman and Dr. Richard Pouw via [email protected]