NIK Inhibitors cell-based screening assay

A method for identifying a compound that inhibits NIK activity, comprising:

• (a) providing HEK cells containing a DNA encoding for NIK and a DNA encoding for IKK1-KD (transfected HEK cells) in a culture medium;
• (b) incubating said transfected HEK cells with and without a candidate molecule to be tested at a temperature of 37° C.+/−2° C. in the presence of 5% CO2 for 24 hours;
• (c) removing the culture medium;
• (d) lysing the cells;
• (e) adding to the cell lysate an acceptor complex followed by a donor complex;
• (f) exciting the cell lysate with a laser illumination and
• (g) measuring the signal arising upon laser excitation of cell lysate using a bead-based luminescent proximity detection system,
• wherein a reduction or the absence of the signal obtained from a cell lysate deriving from transfected HEK cells cultured in the presence of a test compound is indicative of the inhibition of NIK-mediated phosphorylation of IKK1-KD, and wherein the acceptor complex comprises a molecule-coated bead and a binding partner which binds to the IKK1-KD protein and the donor complex comprises a molecule-coated bead and a binding partner which binds to the IKK1-KD protein.