Doublecotin-like kinase 1 increases chemoresistance of colorectal cancer cells through the anti-apoptosis pathway

Abstract

Colorectal cancer (CRC) is the third most common cancer diagnosed and the second leading cause of cancer-related deaths in the United States. About 50% of CRC patients relapsed after surgical resection and ultimately died of metastatic disease. Cancer stem cells (CSCs) are believed to be the primary reason for the recurrence of CRC. Specific stem cell marker, doublecortin-like kinase 1 (DCLK1) plays critical roles in initiating tumorigenesis, facilitating tumor progression, and promoting metastasis of CRC. It is up-regulated in CRC and upregulation of DCLK1 indicates poor prognosis. Whether DCLK1 is correlated with enhanced chemoresistance of CRC cells is unclear. Our research aims to reveal association of DCLK1 with chemoresistance of CRC cells and the underlying molecular mechanisms. In order to achieve our goal, we established stable DCLK1 over-expression cells (DCLK1+) using the HCT116 cells (WT). DCLK1+ and WT cells were treated with 5-Fluorouracil (5-Fu) at different doses for 24 or 48 hours. MTT assay was used to evaluate cell viability and IC 50 of 5-Fu was determined. Quantitative real time PCR was applied to determine gene expression of caspase-3 (casp-3), caspase-4 (casp-4), and caspase-10 (casp-10). Cleaved casp-3 expression was investigated using Western blot and immunofluorescence. Our results demonstrated that IC 50 of 5-Fu for the DCLK1+ cells was significantly higher than that of the WT cells for both 24 and 48hour treatment (P=0.002 and 0.048 respectively), indicating increased chemoresistance of the DCLK1+ cells. Gene expression of casp-3, casp-4, and casp-10 were significantly inhibited in the DCLK1+ cells after 5-Fu treatment compared to the WT cells (P=7.616e-08, 1.575e-05 and 5.307e-08, respectively). Cleaved casp-3 amount and casp-3 positive cells were significantly decreased in the DCLK1+ cells after 5-Fu treatment compared to the WT cells (P=0.015). In conclusion, our results demonstrated that DCLK1 overexpression enhanced the chemoresistance of CRC cells to 5-Fu treatment by suppressing gene expression of key caspases in the apoptosis pathway and activation of apoptosis pathway. DCLK1 can be an intriguing therapeutic target for the effective treatment of CRC patients.

STK35 Is Ubiquitinated by NEDD4L and Promotes Glycolysis and Inhibits Apoptosis Through Regulating the AKT Signaling Pathway, Influencing Chemoresistance of Colorectal Cancer

“…However, the presence of tumor stem cells possessing chemoresistance, which accounts for approximately 0.05–0.1% of the total tumor cell mass, and harbor unlimited self-renewal competency, is the main cause of failure of cancer chemotherapy ( Zeuner et al, 2014 ). As a matter of fact, a great number of human kinases have been identified as coexistent putative markers of tumor stem cells, which contribute to minimizing 5-FU chemosensitivity in human cancers ( Li et al, 2019 ; Hou et al, 2020 ). In the current study, we observed the anti-CRC functions of 5-FU in a dose-dependent manner as expected, whereas the overexpression of STK35 in CRC cells partially reversed these effects, such as the promotion of apoptosis, tumor growth inhibition, and survival improvement in an in vivo mouse model.…”

Single Cell Transcriptome in Colorectal Cancer—Current Updates on Its Application in Metastasis, Chemoresistance and the Roles of Circulating Tumor Cells

“…Moreover, current chemotherapy does not possess the strength to fully eradicate solid tumors, resulting in secondary tumor and relapse. Owing to this, numerous efforts have been made to dissect the chemoresistant cancer cells based on the genes expressions, epigenetics, pathways signatures and therapeutic responses (Datta et al, 2016;Baharudin et al, 2017;Abu et al, 2019;Li et al, 2019). Although bulk transcriptomics is adequate to study the average gene expression signatures related to chemoresistance, they generally involve bulk tissue with assumption that all the cells obtained are of homogeneous material, thereby ignoring the stochasticity of gene expression (Raj and van Oudenaarden, 2008;Stegle et al, 2015).…”

Application of RNA-sequencing to identify transcriptome modification by DCLK1 in colorectal cancer cells

“…Control of multiply critical biological pathways which are involved in the regulation of cell proliferation might be one of the critical underlying molecular mechanisms for DCLK1 to play its function during oncogenesis, progression, invasion and metastasis of CRC. Moreover, our previous findings also demonstrated that DCLK1 significantly increased the chemoresistance of CRC cells to 5-Fu treatment 26 . Therefore, DCLK1 can be developed as a novel therapeutic target for the treatment of colorectal cancer.…”