Structure-Based Design of Selective Salt-Inducible Kinase Inhibitors

Abstract

Salt-inducible kinases (SIKs) are key metabolic regulators. The imbalance in SIK function is associated with the development of diverse cancers, including breast, gastric, and ovarian cancers. Chemical tools to clarify the roles of SIK in different diseases are, however, sparse and are generally characterized by poor kinome-wide selectivity. Here, we have adapted the pyrido[2,3-d]pyrimidin-7-one-based p21-activated kinase (PAK) inhibitor G-5555 for the targeting of SIK, by exploiting differences in the back-pocket region of these kinases. Optimization was supported by high-resolution crystal structures of G-5555 bound to the known off-targets, MST3 and MST4, leading to a chemical probe, MRIA9, with dual SIK/ PAK activity and excellent selectivity over other kinases. Furthermore, we show that MRIA9 sensitizes ovarian cancer cells to treatment with the mitotic agent paclitaxel, confirming earlier data from genetic knockdown studies and suggesting a combination therapy with SIK inhibitors and paclitaxel for the treatment of paclitaxel-resistant ovarian cancer.

The Small-Molecule Inhibitor MRIA9 Reveals Novel Insights into the Cell Cycle Roles of SIK2 in Ovarian Cancer Cells

“…Due to the lack of crystal structure of SIK proteins, we based our development of the new SIK inhibitor on the scaffold of the pyrido [2,3-d]pyrimidin-7-one PAK inhibitor G-5555 [33]. By exploiting the differences in the back-pocket region of the SIK protein family, we developed a G-5555 derivative, MRIA9, that potently and selectively inhibits SIK2 [15]. We observed that the specific MRIA9-induced inhibition of SIK2 reduced its enzymatic activity in ovarian cancer cells monitored by measuring the autophosphorylation on Ser385.…”

“…Recently, we developed a potent pan-SIK inhibitor, MRIA9, which displayed high selectivity against SIK kinases [15]. MRIA9 is an ATP-competitive inhibitor with an in vitro IC 50 of 180 nM determined by nanoBRET (Bioluminescence Resonance Energy Transfer) assays for the kinase isoform SIK2.…”

“…MRIA9 is an ATP-competitive inhibitor with an in vitro IC 50 of 180 nM determined by nanoBRET (Bioluminescence Resonance Energy Transfer) assays for the kinase isoform SIK2. Moreover, at a concentration of 1 µM, MRIA9 showed strong SIK2 selectivity against 443 kinases in radiometric enzyme kinetic assay (PanQinase ® ) and evaluation of all potential off-targets in cell-based on-target assays identified only PAK1-3 as off-targets suggesting that MRIA9 can be used as a chemical probe to investigate SIK function in cellular systems [15].…”

“…All these findings argue for SIK2 as a promising target in suppressing metastatic ovarian cancer. Although several SIK inhibitors have already been investigated [7,10,13,14], all these reagents exhibit a common problem: the lack of specificity and significant off-target activity limiting their use as mechanistic tools for studying the role of SIK kinases in cancer and other diseases [15]. Hence, there has been a need to optimize small molecules to achieve better inhibitor selectivity.…”

“…Hence, there has been a need to optimize small molecules to achieve better inhibitor selectivity. Recently, we addressed this issue by developing a small molecule with potent panSIK activity [15]. Kinome-wide selectivity screening and cellular on-target assays identified only the p21-activated kinase family members 1-3 (PAK1-3) as off-targets of the lead compound MRIA9.…”