Target Residence Time-Guided Optimization on TTK Kinase Results in Inhibitors with Potent Anti-Proliferative Activity

Area of Science:

• Biochemistry
• Molecular Biology
• Oncology

Background:

• Threonine tyrosine kinase (TTK), also known as Mps1, is crucial for the spindle assembly checkpoint.
• TTK is a promising drug target for aggressive cancers like triple-negative breast cancer.
• Understanding how TTK inhibitors affect cellular activity is vital as they enter clinical trials.

Purpose of the Study:

• To investigate the relationship between TTK inhibitor properties and cellular activity.
• To elucidate the mechanism of action for potent TTK inhibitors.
• To develop novel TTK inhibitors with enhanced efficacy.

Main Methods:

• Comparative analysis of 12 TTK inhibitors, including a novel laboratory-developed compound (NTRC 0066-0).
• In vitro cellular assays measuring binding affinity and target residence time.
• X-ray crystallography and thermal stability experiments to determine binding modes.
• Development and testing of novel TTK inhibitors based on a (5,6-dihydro)pyrimido[4,5-e]indolizine scaffold.

Main Results:

• Cellular activity of TTK inhibitors strongly correlates with target residence time.
• Potent inhibitors bind to TTK, causing a glycine-rich loop shift and forming a 'lysine trap' at Lys553.
• Novel inhibitors with extended target residence times exhibit potent anti-proliferative activity.
• A new hybrid Type I/Type III binding mode was identified for the developed inhibitors.

Conclusions:

• Target residence time is a critical determinant of TTK inhibitor efficacy in cellular assays.
• The 'lysine trap' mechanism provides a novel strategy for kinase inhibition.
• Developed TTK inhibitors demonstrate significant anti-proliferative potential, comparable to cytotoxic therapies.
• These findings open new possibilities for TTK inhibitor applications in cancer treatment.