SGC-AAK1-1: A Chemical Probe Targeting AAK1 and BMP2K

Abstract

Inhibitors based on a 3-acylaminoindazole scaffold were synthesized to yield potent dual AAK1/BMP2K inhibitors. Optimization of this 3-acylaminoindazole scaffold furnished a small molecule chemical probe (SGC-AAK1-1, 25) that is potent and selective for AAK1/BMP2K over other NAK family members, demonstrates narrow activity in a kinome-wide screen, and is functionally active in cells. This inhibitor represents one of the best available small molecule tools to study the functions of AAK1 and BMP2K.The human protein Ser/Thr kinases Adaptor protein 2-Associated Kinase 1 (AAK1) and BMP-2-Inducible Kinase (BMP2K/BIKE) play critical roles in mediating endocytosis and other key signaling pathways. Both are broadly expressed and are members of the NAK family of human kinases, which also includes Cyclin G-Associated Kinase (GAK) and Myristoylated and Palmitoylated Serine/Threonine Kinase 1 (MPSK1/STK16). The family shares little homology outside of their kinase domains. 1 AAK1 and BMP2K are the most closely related, with overall sequence identity of 50% and kinase domain sequence identity of 74%. 2 A key function of AAK1 is regulation of receptor-mediated endocytosis via binding directly to clathrin and phosphorylating the medium subunit of AP2 (adaptor protein 2), which stimulates binding to cargo proteins. [3][4][5] AAK1 also modulates the Notch pathway, partially through its phosphorylation of Numb. 6, 7 BMP2K plays a role in osteoblast differentiation, is a clathrin-coated vesicle-associated protein, and, like AAK1, also associates with Numb. 8, 9 Due to their many functions, AAK1 and BMP2K have been implicated as potential drug targets for diverse conditions. AAK1 has been linked to diseases affecting the brain such as schizophrenia, Parkinson's disease and amyotrophic lateral sclerosis as well as implicated as a potential anti-viral target for the treatment of Hepatitis C. 5, 10, 11 BMP2K has been associated with myopia and evaluated as a potential treatment for HIV. 12, 13 A dual AAK1/BMP2K small molecule inhibitor was recently reported as a novel therapeutic to treat neuropathic pain. 14 X-ray crystal structures for the kinase domains of all NAK family members have been solved and reported. 2, 15, 16 Published and novel high-resolution crystal structures of AAK1 and BMP2K reveal target-specific structural features that have enabled our design of specific chemical probes and allowed further

PKIS deep dive yields a chemical starting point for dark kinases and a cell active BRSK2 inhibitor

“…Analogous cell culture conditions have been previously employed by our groups 12, 38 . nanoBRet measurements. NanoBRET assays were executed as described previously 38 . Constructs for BRSK2 NanoBRET measurements were provided in kind by Promega.…”

“…Cells were passaged regularly using 0.05% Trypsin/0.53 mM EDTA in Sodium Bicarbonate (Corning, 25-052-CI), and maintained in Dulbecco’s Modified Eagle Medium (DMEM) (Corning, 10–013-CV) supplemented with 10% fetal bovine serum (FBS). Analogous cell culture conditions have been previously employed by our groups 12 , 38 .…”

“…NanoBRET assays were executed as described previously 38 . Constructs for BRSK2 NanoBRET measurements were provided in kind by Promega.…”

Defining the Neural Kinome: Strategies and Opportunities for Small Molecule Drug Discovery to Target Neurodegenerative Diseases

“…SGC-AAK-1, in fact, was recently identified as a selective chemical probe for studying AAK1 biology and released alongside a structurally related negative control compound that does not inhibit AAK1. 42,43 AAK1 expression is increased in PD patient brains and was correlated with age of onset in a GWAS (Table 2). 56,57 The influence of AAK1 was suggested to rely on its essential role in endocytosis and lysosomal sorting, which, when aberrant, can contribute to the pathology of PD.…”

Strategy for Lead Identification for Understudied Kinases

“…ALS. 14,17,18 Figure 1 shows the structures and kinome-wide profiling data generated at DiscoverX (scanMAX or KINOMEscan) for three TBK1-targeting pyrimidines. 19 The data for TBK1 inhibitors MRT67307 and BX-912 (designed for PDK1 but potent inhibitor of TBK1) is already in the literature (LINCS database).…”