1. Inhibition of DCLK1 with DCLK1-IN-1 Suppresses Renal Cell Carcinoma Invasion and Stemness and Promotes Cytotoxic T-Cell-Mediated Anti-Tumor Immunity
Jian Du, Yuning Yang, Ling Ding, Sassan Hafizi, Zixing Yan, Nathaniel Weygant, Jiannan Yao, Xiaohui Xu, Xuzheng Chen, Jian Liu, Qin Lu, Zhiyun Cao, Yang Ge Cancers (Basel) . 2021 Nov 16;13(22):5729. doi: 10.3390/cancers13225729.
The approval of immune checkpoint inhibitors has expanded treatment options for renal cell carcinoma (RCC), but new therapies that target RCC stemness and promote anti-tumor immunity are needed. Previous findings demonstrate that doublecortin-like kinase 1 (DCLK1) regulates stemness and is associated with RCC disease progression. Herein, we demonstrate that small-molecule kinase inhibitor DCLK1-IN-1 strongly inhibits DCLK1 phosphorylation and downregulates pluripotency factors and cancer stem cell (CSC) or epithelial-mesenchymal transition (EMT)-associated markers including c-MET, c-MYC, and N-Cadherin in RCC cell lines. Functionally, DCLK1-IN-1 treatment resulted in significantly reduced colony formation, migration, and invasion. Additionally, assays using floating or Matrigel spheroid protocols demonstrated potent inhibition of stemness. An analysis of clinical populations showed that DCLK1 predicts RCC survival and that its expression is correlated with reduced CD8+ cytotoxic T-cell infiltration and increases in M2 immunosuppressive macrophage populations. The treatment of RCC cells with DCLK1-IN-1 significantly reduced the expression of immune checkpoint ligand PD-L1, and co-culture assays using peripheral blood monocytes (PBMCs) or T-cell expanded PBMCs demonstrated a significant increase in immune-mediated cytotoxicity alone or in combination with anti-PD1 therapy. Together, these findings demonstrate broad susceptibility to DCLK1 kinase inhibition in RCC using DCLK1-IN-1 and provide the first direct evidence for DCLK1-IN-1 as an immuno-oncology agent.
2. Discovery of a selective inhibitor of doublecortin like kinase 1
Brian M Wolpin, James D Vasta, Matthew B Robers, Srivatsan Raghavan, Sergio Espinosa, Kevin M Haigis, Lianbo Li, Yan Liu, Fleur M Ferguson, Nam Doo Kim, Ryoma Ohi, Emily J Poulin, Ling Huang, Senthil Muthaswamy, Nathanael S Gray, William C Hahn, Wayne Harshbarger, Charles Y Lin, Jinhua Wang, Cesear R Corona, Taebo Sim, Raymond W S Ng, Jose M Lizcano, Miljan Kuljanin, Radha L Kalekar, Nora Dieguez-Martinez, Jost Koren, Annan Yang, Shuning He, Joseph D Mancias, Zhiyang Zeng, Andrew J Aguirre, Kenneth D Westover, Alan L Leggett, Behnam Nabet, Rita Sulahian, A Thomas Look Nat Chem Biol . 2020 Jun;16(6):635-643. doi: 10.1038/s41589-020-0506-0.
Doublecortin like kinase 1 (DCLK1) is an understudied kinase that is upregulated in a wide range of cancers, including pancreatic ductal adenocarcinoma (PDAC). However, little is known about its potential as a therapeutic target. We used chemoproteomic profiling and structure-based design to develop a selective, in vivo-compatible chemical probe of the DCLK1 kinase domain, DCLK1-IN-1. We demonstrate activity of DCLK1-IN-1 against clinically relevant patient-derived PDAC organoid models and use a combination of RNA-sequencing, proteomics and phosphoproteomics analysis to reveal that DCLK1 inhibition modulates proteins and pathways associated with cell motility in this context. DCLK1-IN-1 will serve as a versatile tool to investigate DCLK1 biology and establish its role in cancer.
3. Synthesis and Structure-Activity Relationships of DCLK1 Kinase Inhibitors Based on a 5,11-Dihydro-6 H-benzo[ e]pyrimido[5,4- b][1,4]diazepin-6-one Scaffold
Ling Huang, Eugene N Muratov, Stephen J Capuzzi, Nathanael S Gray, Wayne Harshbarger, Jinhua Wang, Senthil Muthuswamy, Alexander Tropsha, Kenneth D Westover, Yan Liu, Fleur M Ferguson, Xianming Deng J Med Chem . 2020 Jul 23;63(14):7817-7826. doi: 10.1021/acs.jmedchem.0c00596.
Doublecortin-like kinase 1 (DCLK1) is a serine/threonine kinase that is overexpressed in gastrointestinal cancers, including esophageal, gastric, colorectal, and pancreatic cancers. DCLK1 is also used as a marker of tuft cells, which regulate type II immunity in the gut. However, the substrates and functions of DCLK1 are understudied. We recently described the first selective DCLK1/2 inhibitor, DCLK1-IN-1, developed to aid the functional characterization of this important kinase. Here we describe the synthesis and structure-activity relationships of 5,11-dihydro-6H-benzo[e]pyrimido[5,4-b][1,4]diazepin-6-one DCLK1 inhibitors, resulting in the identification of DCLK1-IN-1.
