Discovery of a potent and selective covalent threonine tyrosine kinase (TTK) inhibitor.

Threonine tyrosine kinase (TTK) is a critical component of the spindle assembly checkpoint and plays a pivotal role in mitosis. TTK has been identified as a potential therapeutic target for human cancers. Here, we describe our design, synthesis and evaluation of a class of covalent TTK inhibitors, exemplified by 16 (SYL1073). Compound 16 potently inhibits TTK kinase with an IC50 of 0.016 µM and displays improved selectivity in a panel of kinases. Mass spectrometry analysis reveals that 16 covalently binds to the C604 cysteine residue in the hinge region of the TTK kinase domain. Furthermore, 16 achieves strong potency in inhibiting the growth of various human cancer cell lines, outperforming its relative reversible inhibitor, and eliciting robust downstream effects. Taken together, compound 16 provides a valuable lead compound for further optimization toward the development of drug for treatment of human cancers.

Discovery of Pyrido[2,3-d]pyrimidin-7-one Derivatives as Highly Potent and Efficacious Ectonucleotide Pyrophosphatase/Phosphodiesterase 1 (ENPP1) Inhibitors for Cancer Treatment.

Ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1) is an extracellular enzyme responsible for hydrolyzing cyclic guanosine monophosphate-adenosine monophosphate (cGAMP), the endogenous agonist for the stimulator of interferon genes (STING) pathway. Inhibition of ENPP1 can trigger STING and promote antitumor immunity, offering an attractive therapeutic target for cancer immunotherapy. Despite progress in the discovery of ENPP1 inhibitors, the diversity in chemical structures and the efficacy of the agents are far from desirable, emphasizing the demand for novel inhibitors. Herein, we describe the design, synthesis, and biological evaluation of a series of ENPP1 inhibitors based on the pyrido[2,3-d]pyrimidin-7-one scaffold. Optimization efforts led to compound 31 with significant potency in both ENPP1 inhibition and STING pathway stimulation in vitro. Notably, 31 demonstrated in vivo efficacy in a syngeneic 4T1 mouse triple negative breast cancer model. These findings provide a promising lead compound with a novel scaffold for further drug development in cancer immunotherapy.

Discovery of a PROTAC degrader for METTL3-METTL14 complex.

N6-methyladenosine (m6A) methylation is the most abundant type of RNA modification that is mainly catalyzed by the METTL3-METTL14 methyltransferase complex. This complex has been linked to multiple cancers and is considered a promising therapeutic target for acute myeloid leukemia (AML). However, only a few METTL3 inhibitors targeting the catalytic activity were developed recently. Here, we present the discovery of WD6305 as the potent and selective proteolysis-targeting chimera (PROTAC) degrader of METTL3-METTL14 complex. WD6305 suppresses m6A modification and the proliferation of AML cells, and promotes apoptosis much more effectively than its parent inhibitor. WD6305 also affects a variety of signaling pathways related to the development and proliferation of AML. Collectively, our study reveals PROTAC degradation of METTL3-METTL14 complex as a potential anti-leukemic strategy and provides desirable chemical tool for further understanding METTL3-METTL14 protein functions.