Identification of the DEAD box RNA helicase DDX3 as a therapeutic target in colorectal cancer

// Marise R. Heerma van Voss 1, 2 , Farhad Vesuna 1 , Kari Trumpi 3 , Justin Brilliant 1 , Cynthia Berlinicke 4 , Wendy de Leng 2 , Onno Kranenburg 3 , Johan G. Offerhaus 2 , Horst Bürger 5 , Elsken van der Wall 6 , Paul J. van Diest 2 , Venu Raman 1, 2, 7 1 Department of Radiology and Radiological Science, Johns Hopkins University, School of Medicine, Baltimore, MD, USA 2 Department of Pathology, University Medical Center Utrecht, Utrecht, The Netherlands 3 Department of Surgery, University Medical Center Utrecht, Utrecht, The Netherlands 4 Wilmer Eye Institute, Johns Hopkins University, School of Medicine, Baltimore, MD, USA 5 Institute of Pathology, Paderborn, Germany 6 Department of Internal Medicine, University Medical Center Utrecht, Utrecht, The Netherlands 7 Department of Oncology, Johns Hopkins University, School of Medicine, Baltimore, MD, USA Correspondence to: Venu Raman, e-mail: vraman2@jhmi.edu Keywords: colorectal cancer, DEAD-box RNA helicases, Wnt signaling, small molecule inhibitors, β-catenin Received: May 01, 2015 Accepted: July 09, 2015 Published: August 01, 2015 ABSTRACT Identifying druggable targets in the Wnt-signaling pathway can optimize colorectal cancer treatment. Recent studies have identified a member of the RNA helicase family DDX3 ( DDX3X ) as a multilevel activator of Wnt signaling in cells without activating mutations in the Wnt-signaling pathway. In this study, we evaluated whether DDX3 plays a role in the constitutively active Wnt pathway that drives colorectal cancer. We determined DDX3 expression levels in 303 colorectal cancers by immunohistochemistry. 39% of tumors overexpressed DDX3. High cytoplasmic DDX3 expression correlated with nuclear β-catenin expression, a marker of activated Wnt signaling. Functionally, we validated this finding in vitro and found that inhibition of DDX3 with siRNA resulted in reduced TCF4-reporter activity and lowered the mRNA expression levels of downstream TCF4-regulated genes. In addition, DDX3 knockdown in colorectal cancer cell lines reduced proliferation and caused a G1 arrest, supporting a potential oncogenic role of DDX3 in colorectal cancer. RK-33 is a small molecule inhibitor designed to bind to the ATP-binding site of DDX3. Treatment of colorectal cancer cell lines and patient-derived 3D cultures with RK-33 inhibited growth and promoted cell death with IC50 values ranging from 2.5 to 8 μM. The highest RK-33 sensitivity was observed in tumors with wild-type APC -status and a mutation in CTNNB1 . Based on these results, we conclude that DDX3 has an oncogenic role in colorectal cancer. Inhibition of DDX3 with the small molecule inhibitor RK-33 causes inhibition of Wnt signaling and may therefore be a promising future treatment strategy for a subset of colorectal cancers.