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.
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KINOMEscan and KiNativ data are provided in Supplementary Datasets 1 and 2. Cell line RNA-sequencing data has been deposited to the NCBI GEO (accession number GSE140490). Cell line and deidentified patient-derived organoid RNA-sequencing analyzed data files are provided in Supplementary Dataset 3. Cell line and deidentified patient-derived organoid mass spectrometry-based proteomics and phosphoproteomics analyzed data files are provided in Supplementary Datasets 4 and 5.
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We thank M. Kostic for critical reading of the manuscript and S. Nabet and members of the Gray laboratory for helpful discussions. We gratefully acknowledge S. Gygi for use of CORE for mass spectrometry data analysis software. This work was supported by an American Cancer Society Postdoctoral Fellowship PF-17-010-01-CDD (B.N.), Claudia Adams Barr Program in Innovative Basic Cancer Research Award (B.N.), Katherine L. and Steven C. Pinard Research Fund (N.S.G. and B.N.), Hope Funds for Cancer Research Postdoctoral Fellowship (S.R.), Harvard Catalyst KL2/CMeRIT Fellowship (S.R.), Perry Levy Fellowship (S.R.), the Lustgarten Foundation (S.R., B.M.W., A.J.A. and W.C.H.), NCI HCMI program (A.J.A. and S.R.), American Cancer Society 129089-PF-16-088-01-TBG (E.J.P.), KU-KIST Graduate School of Converging Science and Technology Program (T.S.), Spanish Ministerio de Economia y Competitividad grant no. SAF2015-60268R, cofunded by Fondo Europeo de Desarrollo Regional funds (J.M.L.), Pancreatic Cancer Action Network Catalyst Award (A.J.A.), Doris Duke Charitable Foundation Clinician Scientist Development Award (A.J.A.), NCI K08 CA218420 (A.J.A.), NCI U01 CA176058 (W.C.H.), NCI U01 CA199253 (W.C.H.) and NCI U01 CA224146 (W.C.H.), American Cancer Society Award 132205-RSG-18-039-01-DMC (K.D.W.), Welch Foundation grant no. I1829 (K.D.W.), 2017 AACR-Bayer Innovation and Discovery grant no. 17-80-44-GRAY (N.S.G.), DF/HCC GI SPORE Developmental Research Project Award P50CA127003 (N.S.G. and K.M.H.) and Hale Center for Pancreatic Research (J.D.M., B.M.W., A.J.A., W.C.H. and N.S.G.).
F.M.F. and N.S.G. are inventors on a patent application related to the DCLK1 inhibitors described in this manuscript (WO/2018/075608). B.N. is an inventor on patent applications related to the dTAG system described in this manuscript (WO/2017/024318, WO/2017/024319, WO/2018/148443, WO/2018/148440). Z.Z., C.R.C., J.D.V. and M.B.R. are employees of Promega Corporation. B.M.W. receives research funding from Celgene, Inc, and is a consultant for G1 Therapeutics, BioLineRx and GRAIL. A.J.A. has consulted for Oncorus, Inc. W.C.H. is a consultant for Thermo Fisher, AjuIB, MPM Capital, iTeos and Paraxel and is a Scientific Founder and serves on the Scientific Advisory Board (SAB) for KSQ Therapeutics. K.D.W. is a member of the SAB for Vibliome Therapeutics. N.S.G. is a Scientific Founder, member of the SAB and equity holder in C4 Therapeutics, Syros, Soltego, B2S, Gatekeeper and Petra Pharmaceuticals. The Gray laboratory receives or has received research funding from Novartis, Takeda, Astellas, Taiho, Janssen, Kinogen, Voroni, Her2llc, Deerfield and Sanofi.
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Supplementary Tables 1 and 2, Figs. 1–22 and Note.
Full KINOMEscan dataset.
Full KiNativ dataset from lysate and live cell format.
RNA-sequencing dataset for PANFR0172_T2, PANFR0172_T3 and PATU-8988T on comparison of DMSO and 2.5 µM DCLK1-IN-1 treatments for 24 h.
Mass spectrometry-based proteomics dataset for PANFR0172_T2, PANFR0172_T3 and PATU-8988T on comparison of DMSO and 2.5 µM DCLK1-IN-1 treatments for 24 h.
Mass spectrometry-based phosphoproteomics dataset for PANFR0172_T2, PANFR0172_T3 and PATU-8988T on comparison of DMSO and 2.5 µM DCLK1-IN-1 treatments for 24 h.
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Ferguson, F.M., Nabet, B., Raghavan, S. et al. Discovery of a selective inhibitor of doublecortin like kinase 1. Nat Chem Biol 16, 635–643 (2020). https://doi.org/10.1038/s41589-020-0506-0
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