Determination of a cellular method to identify inhibitors of direct protein interactions

Abstract

Proteins are essential elements in biological functions. They interact with each other and can form complexes. The mapping of the human interactome aims at gathering all the interactions between proteins in the human organism in order to better understand biological mechanisms and processes. The number of protein-protein interactions (PPIs) is estimated to be up to 650 000. At present, about 53 000 protein interaction pairs have been detected. Since these PPIs are involved in many biological functions, aberrant PPIs can lead to diseases such as autoimmune diseases, cancers and neurodegenerative diseases. Therefore, they are interesting targets for the development of new therapeutic treatments. This project aims to determine a cellular method to identify inhibitors of direct interaction between proteins. Some recent methods based on luminescence detection allow to identify these interactions at high throughput. However, there is no similar method to identify modulators of these interactions at high throughput and in cells. These inhibitors would be interesting in understanding the molecular mechanisms and as development tools for new drugs. This is why this project uses 4 biosensors based on the complementation of luciferases: mN2H, yN2H, GPCA and NanoBiT. The first objective is to select PPIs for which inhibitors already exist from the iPPI-DB and 2P2i databases. Then, the detection of 7 PPIs will be evaluated using the 4 biosensors mentioned above. Finally, inhibitors already described in the literature for some of these PPIs will be tested. For one of the BCL2L1+BAK1 interaction inhibitors, ABT-737, the results obtained appear to be promising for the GPCA and NanoBiT biosensors. Although the concentration-response curves observed for the mN2H and yN2H assays correspond to sigmoids, the IC50 values obtained for GPCA and NanoBiT are lower and show a good sensitivity and a certain reversibility of these 2 biosensors