BackgroundChloroquine (CQ), the worldwide used anti-malarial drug, has recently being focused as a potential anti-cancer agent as well as a chemosensitizer when used in combination with anti-cancer drugs. It has been shown to inhibit cell growth and/or to induce cell death in various types of cancer. 5-Fluorouracil (5-FU) is the chemotherapeutic agent of first choice in colorectal cancer, but in most cases, resistance to 5-FU develops through various mechanisms. Here, we focused on the combination of CQ as a mechanism to potentiate the inhibitory effect of 5-FU on human colon cancer cells.MethodsHT-29 cells were treated with CQ and/or 5-FU, and their proliferative ability, apoptosis and autophagy induction effects, and the affection of the cell cycle were evaluated. The proliferative ability of HT-29 was analyzed by the MTS assay. Apoptosis was quantified by flow-cytometry after double-staining of the cells with AnnexinV/PI. The cell cycle was evaluated by flow-cytometry after staining of cells with PI. Autophagy was quantified by flow-cytometry and Western blot analysis. Finally, to evaluate the fate of the cells treated with CQ and/or 5-FU, the colony formation assay was performed.Results5-FU inhibited the proliferative activity of HT-29 cells, which was mostly dependent on the arrest of the cells to the G0/G1-phase but also partially on apoptosis induction, and the effect was potentiated by CQ pre-treatment. The potentiation of the inhibitory effect of 5-FU by CQ was dependent on the increase of p21Cip1 and p27Kip1 and the decrease of CDK2. Since CQ is reported to inhibit autophagy, the catabolic process necessary for cell survival under conditions of cell starvation or stress, which is induced by cancer cells as a protective mechanism against chemotherapeutic agents, we also analyzed the induction of autophagy in HT-29. HT-29 induced autophagy in response to 5-FU, and CQ inhibited this induction, a possible mechanism of the potentiation of the anti-cancer effect of 5-FU.ConclusionOur findings suggest that the combination therapy with CQ should be a novel therapeutic modality to improve efficacy of 5-FU-based chemotherapy, possibly by inhibiting autophagy-dependent resistance to chemotherapy.

Many long noncoding RNAs (lncRNAs) are reported to be dysregulated in human cancers and play critical roles in tumor development and progression. Furthermore, it has been reported that many lncRNAs regulate gene expression by recruiting chromatin remodeling complexes to specific genomic loci or by controlling transcriptional or posttranscriptional processes. Here we show that an lncRNA termed UPAT [ubiquitin-like plant homeodomain (PHD) and really interesting new gene (RING) finger domaincontaining protein 1 (UHRF1) Protein Associated Transcript] is required for the survival and tumorigenicity of colorectal cancer cells. UPAT interacts with and stabilizes the epigenetic factor UHRF1 by interfering with its β-transducin repeat-containing protein (TrCP)-mediated ubiquitination. Furthermore, we demonstrate that UHRF1 up-regulates Stearoyl-CoA desaturase 1 and Sprouty 4, which are required for the survival of colon tumor cells. Our study provides evidence for an lncRNA that regulates protein ubiquitination and degradation and thereby plays a critical role in the survival and tumorigenicity of tumor cells. Our results suggest that UPAT and UHRF1 may be promising molecular targets for the therapy of colon cancer.A mong the RNA products transcribed from the mammalian genome are numerous long noncoding RNAs (lncRNAs)-that is, RNAs longer than 200 nucleotides with little or no protein-coding potential (1, 2). Many lncRNAs are expressed in a developmentally regulated and cell type-dependent manner (3, 4). Increasing evidence suggests that lncRNAs play critical roles in a diverse set of biological processes, including proliferation, differentiation, embryogenesis, neurogenesis, and stem cell pluripotency (5, 6).It has been reported that many lncRNAs regulate gene expression by recruiting chromatin remodeling complexes to specific genomic regions (2). It has also been shown that many lncRNAs regulate transcription by modulating the activity of transcriptional regulators (1, 6-8). lncRNAs also regulate various posttranscriptional processes, including splicing, transport, translation, and degradation of mRNA. Furthermore, recent studies have shown that a number of lncRNAs play critical roles in tumor development and progression.UHRF1 [ubiquitin-like plant homeodomain (PHD) and really interesting new gene (RING) finger domain-containing protein 1] is an epigenetic factor that consists of multiple domains (9). UHRF1 regulates transcription by regulating DNA methylation and histone modification. UHRF1 also possesses E3 ubiquitin ligase activity and ubiquitinates histones and DNA methyltransferase 1 (DNMT1), thereby regulating the chromatin structure and stability of DNMT1 (10, 11). UHRF1 plays key roles in multiple biological processes, including proliferation and development. Furthermore, UHRF1 is overexpressed in various tumors, including colon, breast, bladder, prostate, and lung cancers, and plays a critical role in the proliferation and survival of tumor cells (9).In the present study, we attempted to identify lncRNAs criti...

Aberrant activation of Wnt/β-catenin signaling is a major driving force in colon cancer. Wnt/β-catenin signaling induces the expression of the transcription factor c-Myc, leading to cell proliferation and tumorigenesis. c-Myc regulates multiple biological processes through its ability to directly modulate gene expression. Here, we identify a direct target of c-Myc, termed MYU, and show that MYU is upregulated in most colon cancers and required for the tumorigenicity of colon cancer cells. Furthermore, we demonstrate that MYU associates with the RNA binding protein hnRNP-K to stabilize CDK6 expression and thereby promotes the G1-S transition of the cell cycle. These results suggest that the MYU/hnRNP-K/CDK6 pathway functions downstream of Wnt/c-Myc signaling and plays a critical role in the proliferation and tumorigenicity of colon cancer cells.

Aberrant activation of Wnt signalling results in colorectal tumours. Lgr5 is specifically expressed in stem cells of the intestine and has an essential role in maintaining tissue homeostasis. Lgr5-positive stem cells are responsible for the intestinal adenoma initiated by mutations in adenomatous polyposis coli. Furthermore, Lgr5 interacts with R-spondins and thereby activates Wnt signalling. However, the function of Lgr5 in colorectal tumourigenesis is unclear. Here we show that LGR5 is required for the tumourigenicity of colorectal cancer cells. We show that the transcription factor GATA6 directly enhances the expression of LGR5. We further demonstrate that GATA6 is upregulated in colorectal cancer cells due to the downregulation of miR-363, which directly targets GATA6. Moreover, we show that overexpression of miR-363 suppresses the tumourigenicity of colorectal cancer cells. These results suggest that the miR-363-GATA6-LGR5 pathway is critical for colorectal tumourigenesis and would be a promising target for the treatment of colorectal cancer.