en Cancer Cancer Original Article Original Article Colorectal cancer (CRC) is one of the most prevalent diagnosed cancers and a common cause of cancer-related mortality. Despite effective clinical responses, a large proportion of patients undergo resistance to radiation therapy. Therefore, the identification of efficient targeted therapy strategies would be beneficial to overcome cancer radioresistance. Doublecortin-like kinase 1 (DCLK1) is an intestinal and pancreatic stem cell marker that showed overexpression in a variety of cancers. The transfection of <em>DCLK1</em> siRNA to &lrm;normal HCT-116 cells was performed, and then cells were irradiated with X-rays. The effects of <em>DCLK1</em> inhibition on cell survival, apoptosis, cell cycle, DNA damage response (ATM and &gamma;H2AX proteins), epithelial- mesenchymal transition (EMT) related genes (vimentin, N‐cadherin, and E-cadherin), cancer stem cells markers (<em>CD44</em>, <em>CD133</em>, <em>ALDH1</em>, and <em>BMI1</em>), and <a name="OLE_LINK1">&beta;</a>‐catenin signaling pathway (&beta;‐catenin) were evaluated. <em>DCLK1</em>siRNA downregulated <em>DCLK1</em> expression in HCT-116 cells at both mRNA and protein levels <a name="_Hlk65092837">(P<em> <</em> 0.01)</a>. Colony formation assay showed a significantly reduced cell survival in the <em>DCLK1</em> siRNA transfected group in comparison with the control group following exposure to 4 and 6 Gy doses of irradiation (P < 0.01). Moreover, the expression of cancer stem cells markers (P < 0.01), EMT related genes (P < 0.01), and DNA repair proteins including pATM (P < 0.01) and &gamma;H2AX (P < 0.001) were significantly decreased in the transfected cells in comparison with the nontransfected group after radiation. Finally, the cell apoptosis rate (P < 0.01) and the number of cells in the G0/G1 phase in the silencing <em>DCLK1</em> group was increased (P < 0.01). These findings suggest that <em>DCLK1 </em>can be considered a promising therapeutic target for the treatment of radioresistant human CRC. DCLK1, ionizing radiation, colorectal cancer, radiosensitivity 23 33 http://ijmcmed.org/browse.php?a_code=A-10-942-2&slc_lang=en&sid=1 Chiman Mohammadi Chiman-mohamadi1985@gmail.com 100319475328460020501 100319475328460020501 No Research Center for Molecular Medicine, Hamadan University of Medical Sciences, Hamadan, Iran. Ali Mahdavinezhad ali-mahdavi@gmail.com 100319475328460020502 100319475328460020502 No Research Center for Molecular Medicine, Hamadan University of Medical Sciences, Hamadan, Iran. Massoud Saidijam s.jam110@yahoo.com 100319475328460020503 100319475328460020503 No Research Center for Molecular Medicine, Hamadan University of Medical Sciences, Hamadan, Iran. Fatemeh Bahreini fbahreini2824@gmail.com 100319475328460020504 100319475328460020504 No Research Center for Molecular Medicine, Hamadan University of Medical Sciences, Hamadan, Iran. Abdolazim Sedighi Pashaki sedighi.pashaki@gmail.com 100319475328460020505 100319475328460020505 No Mahdieh Radiotherapy and Brachytherapy Charitable Center, Hamadan, Iran. Mohammad Hadi Gholami hadigholami2@gmail.com 100319475328460020506 100319475328460020506 No Mahdieh Radiotherapy and Brachytherapy Charitable Center, Hamadan, Iran. Rezvan Najafi najafi2535@gmail.com 100319475328460020507 100319475328460020507 Yes Research Center for Molecular Medicine, Hamadan University of Medical Sciences, Hamadan, Iran.