NTRC-0066-0

TTK is an essential spindle assembly checkpoint enzyme in many organisms. It plays a central role in tumor cell proliferation and is aberrantly overexpressed in a wide range of tumor types. We recently reported on a series of potent and selective TTK inhibitors with strong antiproliferative activity in triple negative breast cancer (TNBC) cell lines (8: TTK IC₅₀ = 3.0 nM; CAL-51 IC₅₀ = 84.0 nM). Inspired by previously described potent tricyclic TTK inhibitor 6 (TTK IC₅₀ = 0.9 nM), we embarked on a structure-enabled design and optimization campaign to identify an improved series with excellent potency, TTK selectivity, solubility, CYP inhibition profile, and in vivo efficacy in a TNBC xenograft model. These efforts culminated in the discovery of 25 (TTK IC₅₀ = 3.0 nM; CAL-51 IC₅₀ = 16.0 nM), which showed significant single-agent efficacy when dosed iv in a TNBC xenograft model without body weight loss.

Protein kinase inhibitors disrupt phosphorylation of the target kinases, which are an important class of drug for treating cancer and other diseases. Conventional structure-based design methods (such as molecular docking) focus on the static binding mode of the kinase inhibitor with its target. However, dissociation kinetic properties of a drug molecule are found to correlate with its residence time in vivo and thus have drawn the attention of drug designers in recent years. In this study, we have applied the local-scaled molecular dynamics (MD) simulation enabled in GROMACS software to explore the unbinding mechanism of a total of 41 type I and type II kinase inhibitors. Our simulation considered multiple starting configurations as well as possible protonation states of kinase inhibitors. Based on our local-scaled MD results, we discovered that the integrals of the favorable binding energy during dissociation correlated well (R² = 0.64) with the experimental dissociation rate constants of those kinase inhibitors on the entire data set. Given its accuracy and technical advantage, this method may serve as a practical option for estimating this important property in reality. Our simulation also provided a reasonable explanation of the dynamic properties of kinase and its inhibitor as well as the role of relevant water molecules in dissociation.