IGF1-mediated HOXA13 overexpression promotes colorectal cancer metastasis through upregulating ACLY and IGF1R

Metastasis is the major reason for the high mortality of colorectal cancer (CRC) patients and its molecular mechanism remains unclear. Here, we report a novel role of Homeobox A13 (HOXA13), a member of the Homeobox (HOX) family, in promoting CRC metastasis. The elevated expression of HOXA13 was positively correlated with distant metastasis, higher AJCC stage, and poor prognosis in two independent CRC cohorts. Overexpression of HOXA13 promoted CRC metastasis whereas downregulation of HOXA13 suppressed CRC metastasis. Mechanistically, HOXA13 facilitated CRC metastasis by transactivating ATP-citrate lyase (ACLY) and insulin-like growth factor 1 receptor (IGF1R). Knockdown of ACLY and IGFIR inhibited HOXA13-medicated CRC metastasis, whereas ectopic overexpression of ACLY and IGFIR rescued the decreased CRC metastasis induced by HOXA13 knockdown. Furthermore, Insulin-like growth factor 1 (IGF1), the ligand of IGF1R, upregulated HOXA13 expression through the PI3K/AKT/HIF1α pathway. Knockdown of HOXA13 decreased IGF1-mediated CRC metastasis. In addition, the combined treatment of ACLY inhibitor ETC-1002 and IGF1R inhibitor Linsitinib dramatically suppressed HOXA13-mediated CRC metastasis. In conclusion, HOXA13 is a prognostic biomarker in CRC patients. Targeting the IGF1-HOXA13-IGF1R positive feedback loop may provide a potential therapeutic strategy for the treatment of HOXA13-driven CRC metastasis.Subject terms: Metastasis, Colorectal cancer     

FBXW11 contributes to stem-cell-like features and liver metastasis through regulating HIC1-mediated SIRT1 transcription in colorectal cancer

Colorectal tumorigenesis is a heterogeneous disease driven by multiple genetic and epigenetic alterations. F-box and WD repeat domain containing 11 (FBXW11) is a member of the F-box protein family that regulates the ubiquitination of key factors associated with tumor growth and aggressiveness. Our study aimed to explore the role of FBXW11 in the development and metastasis of colorectal cancer (CRC). FBXW11 was overexpressed in colorectal tumor tissues and its overexpression was associated with a poor prognosis of CRC patients. The upregulation of FBXW11 not only promoted cell proliferation, invasion, and migration, but also contributed to maintaining stem-cell features in colorectal tumor cells. Further analysis revealed that FBXW11 targeted hypermethylated in cancer 1 (HIC1) and reduced its stability in CRC cells through ubiquitination. Moreover, the expression of sirtuin 1 (SIRT1), a deacetylase in tumor cells was upregulated by FBXW11 via regulating HIC1 expression. The mouse xenograft models of CRC confirmed that FBXW11 knockdown impeded colorectal tumor growth and liver metastasis in vivo. In summary, our study identified FBXW11 as an oncogenic factor that contributed to stem-cell-like properties and liver metastasis in CRC via regulating HIC1-mediated SIRT1 expression. These results provide a rationale for the development of FBXW11-targeting drugs for CRC patients.Subject terms: Cancer, Endocrine system and metabolic diseases     

Matrix metalloproteinase 2 promotes cell growth and invasion in colorectal cancer

Colorectal cancer (CRC) is the third leading cause of cancer-related death in the western world. In this study, we evaluated the expression of matrix metalloproteinase 2 gene (MMP2) in CRC and analyzed its correlation with clinicopathological features. We found that the expression of MMP2 was significantly higher in CRC tissues than in the colorectal tissues. In addition, high levels of MMP2 protein were positively correlated with the status of tumor size, lymph node metastasis, distant metastasis, Dukes' stage, and tumor invasion. Moreover, patients with higher MMP2 levels had markedly shorter overall survivals than those with low MMP2 levels. Multivariate analysis results suggested that the level of MMP2 expression is an independent prognostic indicator for the survival of patients with CRC. Silencing MMP2 expression in CRC cell lines with lentiviral-mediated shRNA markedly suppressed cell proliferation, colony formation, and invasion. Furthermore, we observed that vascular endothelial growth factor (VEGF) and membrane type 1 (MT1)-MMP protein levels were decreased in MMP2-down-regulated colorectal cells. Therefore, our study demonstrated that MMP2 is an important factor related to carcinogenesis and metastasis of CRC, and MMP2 promotes CRC cell growth and invasion by up-regulating VEGF and MT1-MMP expression, which makes this pathway a potential target for cancer treatment.     

Colorectal cancer-associated fibroblasts promote metastasis by up-regulating LRG1 through stromal IL-6/STAT3 signaling

Cancer-associated fibroblasts (CAFs) have been shown to play a strong role in colorectal cancer metastasis, yet the underlying mechanism remains to be fully elucidated. Using CRC clinical samples together with ex vivo CAFs-CRC co-culture models, we found that CAFs induce expression of Leucine Rich Alpha-2-Glycoprotein 1(LRG1) in CRC, where it shows markedly higher expression in metastatic CRC tissues compared to primary tumors. We further show that CAFs-induced LRG1 promotes CRC migration and invasion that is concomitant with EMT (epithelial-mesenchymal transition) induction. In addition, this signaling axis has also been confirmed in the liver metastatic mouse model which displayed CAFs-induced LRG1 substantially accelerates metastasis. Mechanistically, we demonstrate that CAFs-secreted IL-6 (interleukin-6) is responsible for LRG1 up-regulation in CRC, which occurs through a direct transactivation by STAT3 following JAK2 activation. In clinical CRC tumor samples, LRG1 expression was positively correlated with CAFs-specific marker, α-SMA, and a higher LRG1 expression predicted poor clinical outcomes especially distant metastasis free survival, supporting the role of LRG1 in CRC progression. Collectively, this study provided a novel insight into CAFs-mediated metastasis in CRC and indicated that therapeutic targeting of CAFs-mediated IL-6-STAT3-LRG1 axis might be a potential strategy to mitigate metastasis in CRC.Subject terms: Colon cancer, Cancer microenvironment     

Role of HGF/c‐Met in the treatment of colorectal cancer with liver metastasis

The system of hepatocyte growth factor (HGF) and its receptor c‐Met plays a critical role in tumor invasive growth and metastasis. The mortality rate of colorectal cancer (CRC), one of the most commonly diagnosed malignancies, is increased by it gradual development into metastasis, most frequently in the liver. Overexpression of c‐Met, the protein tyrosine kinase receptor for the HCF/scatter factor, has been implicated in the progression and metastasis of human colorectal carcinoma. In this study, we aimed to investigate the role of c‐Met in CRC liver metastasis and illustrate the clinical impact of regulating HGF/c‐Met signaling in patients with CRC liver metastasis. We found that (I) higher levels of c‐Met expression (mRNA and Protein) in CRC liver metastasis than primary CRC by assessing the patient tissue samples; (II) a positive correlation of c‐Met expression with tumor stages of CRC liver metastasis, as well as c‐Met expression in CRC, live metastasis concurred with regional lymph node metastasis; (III) the clinical impact of downregulation of HGF/c‐Met signaling on the reduction of proliferation and invasion in CRC liver metastasis. Therefore, we demonstrate that the regulation of HGF/c‐Met pathways may be a promising strategy in the treatment of patients with CRC liver metastasis.     

MYH9-dependent polarization of ATG9B promotes colorectal cancer metastasis by accelerating focal adhesion assembly

Tumour metastasis is a major reason accounting for the poor prognosis of colorectal cancer (CRC), and the discovery of targets in the primary tumours that can predict the risk of CRC metastasis is now urgently needed. In this study, we identified autophagy-related protein 9B (ATG9B) as a key potential target gene for CRC metastasis. High expression of ATG9B in tumour significantly increased the risk of metastasis and poor prognosis of CRC. Mechanistically, we further find that ATG9B promoted CRC invasion mainly through autophagy-independent manner. MYH9 is the pivotal interacting protein for ATG9B functioning, which directly binds to cytoplasmic peptide segments aa368–411 of ATG9B by its head domain. Furthermore, the combination of ATG9B and MYH9 enhance the stability of each other by decreasing their binding to E3 ubiquitin ligase STUB1, therefore preventing them from ubiquitin-mediated degradation, which further amplified the effect of ATG9B and MYH9 in CRC cells. During CRC cell invasion, ATG9B is transported to the cell edge with the assistance of MYH9 and accelerates focal adhesion (FA) assembly through mediating the interaction of endocytosed integrin β1 and Talin-1, which facilitated to integrin β1 activation. Clinically, upregulated expression of ATG9B in human CRC tissue is always accompanied with highly elevated expression of MYH9 and associated with advanced CRC stage and poor prognosis. Taken together, this study highlighted the important role of ATG9B in CRC metastasis by promoting focal adhesion assembly, and ATG9B together with MYH9 can provide a pair of potential therapeutic targets for preventing CRC progression.Subject terms: Metastasis, Oncogenes     

Long noncoding RNA lncBRM promotes proliferation and invasion of colorectal cancer by sponging miR-204-3p and upregulating TPT1

Long noncoding RNAs (lncRNAs) are characterized as a type of noncoding RNAs over 200 nucleotides with little or none protein-coding potential. In the past years, lncRNAs have been proved to participant in many physiological and pathological processes. However, the role of lncRNAs in colorectal cancer (CRC) still needs more attentions. In our study, we found that lncBRM was highly expressed in CRC samples and the expression level of lncBRM was correlated with metastasis and advanced stage in CRC patients. And also, we showed that high expression of lncBRM predicted poor prognosis. Furthermore, we found that knockdown of lncBRM impaired the proliferation, migration and invasion of CRC cells while overexpressing of lncBRM promotes the proliferation, migration and invasion of CRC cells. Mechanically, we found that lncBRM served as a sponge of miR-204-3p that targeted TPT1. Highly expressed TPT1 can promote the proliferation, migration and invasion of CRC cells. In conclusion, we found that lncBRM was highly expressed in CRC and sponged miR-204-3p to modulate the expression of TPT1.     

SIRT2, a direct target of miR‐212‐5p,suppresses the proliferation and metastasis of colorectal cancer cells

The aberrant expression of human sirtuin 2 (SIRT2) has been detected in various types of cancer; however, the biological roles, underlying mechanisms and clinical significance of SIRT2 dysregulation in human colorectal cancer (CRC) remain unclear. The results of this study demonstrate that compared with paired normal tissues, SIRT2 expression is significantly decreased in CRC tissues. SIRT2 loss has been correlated with clinicopathological characteristics, including distant metastasis, lymph node metastasis and American Joint Committee on Cancer (AJCC) stage; this loss serves as an independent factor that indicates a poor prognosis for patients with CRC. Further gain‐ and loss‐of‐function analyses have demonstrated that SIRT2 suppresses CRC cell proliferation and metastasis both in vivo and in vitro. Mechanistically, miR‐212‐5p was identified to directly target the SIRT2 3′‐untranslated region (3′‐UTR), leading to SIRT2 down‐regulation. The ectopic expression of SIRT2 reverses the effect of miR‐212‐5p overexpression on CRC cell colony formation, invasion, migration and proliferation. Clinically, an inverse correlation was found between miR‐212‐5p and SIRT2 expression. High miR‐212‐5p expression has been found to result in a poor prognosis and aggressive clinicopathological characteristics in patients with CRC. Taken together, these results suggest that SIRT2, targeted by miR‐212‐5p, acts as a tumour suppressor in CRC and that the miR‐212‐5p/SIRT2 axis is a promising prognostic factor and potential therapeutic target in CRC.     

PTEN Loss Increases PD-L1 Protein Expression and Affects the Correlation between PD-L1 Expression and Clinical Parameters in Colorectal Cancer

Background

Programmed death ligand-1 (PD-L1) has been identified as a factor associated with poor prognosis in a range of cancers, and was reported to be mainly induced by PTEN loss in gliomas. However, the clinical effect of PD-L1 and its regulation by PTEN has not yet been determined in colorectal cancer (CRC). In the present study, we verified the regulation of PTEN on PD-L1 and further determined the effect of PTEN on the correlation between PD-L1 expression and clinical parameters in CRC.

Methods/Results

RNA interference approach was used to down-regulate PTEN expression in SW480, SW620 and HCT116 cells. It was showed that PD-L1 protein, but not mRNA, was significantly increased in cells transfected with siRNA PTEN compared with the negative control. Moreover, the capacity of PTEN to regulate PD-L1 expression was not obviously affected by IFN-γ, the main inducer of PD-L1. Tissue microarray immunohistochemistry was used to detect PD-L1 and PTEN in 404 CRC patient samples. Overexpression of PD-L1 was significantly correlated with distant metastasis (P<0.001), TNM stage (P<0.01), metastatic progression (P<0.01) and PTEN expression (P<0.001). Univariate analysis revealed that patients with high PD-L1 expression had a poor overall survival (P<0.001). However, multivariate analysis did not support PD-L1 as an independent prognostic factor (P = 0.548). Univariate (P<0.001) and multivariate survival (P<0.001) analysis of 310 located CRC patients revealed that high level of PD-L1 expression was associated with increased risks of metastatic progression. Furthermore, the clinical effect of PD-L1 on CRC was not statistically significant in a subset of 39 patients with no PTEN expression (distant metastasis: P = 0.102; TNM stage: P = 0.634, overall survival: P = 0.482).

Conclusions

PD-L1 can be used to identify CRC patients with high risk of metastasis and poor prognosis. This clinical manifestation may be partly associated with PTEN expression.     

Long non-coding RNA ZFAS1 sponges miR-484 to promote cell proliferation and invasion in colorectal cancer

The incidence and mortality rate of colorectal cancer (CRC) have been significantly increasing. However, mechanisms involved in CRC progression are still unclear. LncRNA ZFAS1 has been verified as oncogenic molecular in a series of tumors, including CRC. However, the underlying mechanism of ZFAS1 in CRC carcinogenesis remains unclear. In the present study, our data showed that ZFAS1 expression was significantly upregulated in CRC tissues and cell lines. Correlation analysis showed that high ZFAS1 expression was significantly associated with Helicobacter pylori infection, lymph nodes metastasis, advanced TNM stage and poor overall survival of CRC patients. Loss-of-function experiments revealed that ZFAS1 inhibition could markedly suppress CRC cells proliferation and invasion both in vitro and in vivo. Bioinformatics analysis and luciferase reporter assay revealed that ZFAS1 directly interacted with miR-484. Rescue experiments showed that miR-484 inhibitor reversed the tumor suppressing roles of ZFAS1 knockdown on CRC cells. Therefore, our study suggested that ZFAS1 could act as an oncogene in CRC tumorigenesis, and discovered the functional regulatory pathway of ZFAS1 sponging miR-484.     

CHD4 as an important mediator in regulating the malignant behaviors of colorectal cancer

Colorectal cancer (CRC) has ranked first in terms of incidence in Taiwan. Surgical resection combined with chemo-, radio-, or targeted-therapies are the main treatments for CRC patients in current clinical practice. However, many CRC patients still respond poorly to these treatments, leading to tumor recurrence and an unacceptably high incidence of metastasis and death. Therefore, appropriate diagnosis, treatment, and drug selection are pressing issues in clinical practice.The Mi-2/nucleosome remodeling and deacetylase complex is an important epigenetic regulator of chromatin structure and gene expression. An important component of this complex is chromodomain-helicase-DNA-binding protein 4 (CHD4), which is involved in DNA repair after injury. Recent studies have indicated that CHD4 has oncogenic functions that inhibit multiple tumor suppressor genes through epigenetic regulation. However, the role of CHD4 in CRC has not yet been well investigated.In this study, we compared CHD4 expression in CRC patients from The Cancer Genome Atlas database. We found higher levels of CHD4 expression in CRC patients. In a series of in vitro experiments, we found that CHD4 affected cell motility and drug sensitivity in CRC cells. In animal models, the depletion of CHD4 affected CRC tumor growth, and the combination of a histone deacetylase 1 (HDAC1) inhibitor and platinum drugs inhibited CHD4 expression and increased the cytotoxicity of platinum drugs. Moreover, CHD4 expression was also a prognostic biomarker in CRC patients.Based on the above results, we believe that CHD4 expression is a viable biomarker for predicting metastasis CRC patients, and it has the potential to become a target for drug development.     

Prognostic Significance of CD26 in Patients with Colorectal Cancer

Background

CD26, dipeptidyl peptidase IV, was discovered firstly as a membrane-associated peptidase on the surface of leukocyte. We previously demonstrated that a subpopulation of CD26⁺ cells were associated with the development of distant metastasis, enhanced invasiveness and chemoresistance in colorectal cancer (CRC). In order to understand the clinical impact of CD26, the expression was investigated in CRC patient''s specimens. This study investigated the prognostic significance of tumour CD26 expression in patients with CRC. Examination of CD26⁺ cells has significant clinical impact for the prediction of distant metastasis development in colorectal cancer, and could be used as a selection criterion for further therapy.

Methods

Tumour CD26 expression levels were studied by immunohistochemistry using Formalin-fixed paraffin embedded (FFPE) tissues in 143 patients with CRC. Tumour CD26 expression levels were correlated with clinicopathological features of the CRC patients. The prognostic significance of tumour tissue CD26 expression levels was assessed by univariate and multivariate analyses.

Result

CD26 expression levels in CRC patients with distant metastasis were significantly higher than those in non-metastatic. High expression levels of CD26 were significantly associated with advanced tumour staging. Patients with a high CD26 expression level had significantly worse overall survival than those with a lower level (p<0.001).

Conclusions

The expression of CD26 was positively associated with clinicopathological correlation such as TNM staging, degree of differentiation and development of metastasis. A high CD26 expression level is a predictor of poor outcome after resection of CRC. CD26 may be a useful prognostic marker in patients with CRC.     

The Metastasis Suppressor,N-myc Downregulated Gene 1 (NDRG1), Is a Prognostic Biomarker for Human Colorectal Cancer

Metastasis remains to be one of the most prevalent causes leading to poor long-term survival of colorectal cancer (CRC) patients. The clinical significances of tumor metastatic suppressor, N-myc downregulated gene 1 (NDRG1), have been inconsistently reported in a variety of cancerous diseases. In this study with 240 CRC clinical specimens, we showed that NDRG1 expression was significantly decreased in most of CRC tissues compared to the paired non-tumor counterparts. Statistical analysis revealed a significant inverse correlation of NDRG1 expression with tumor stage, differentiation status and metastasis. Compared with NDRG1-negative group, NDRG1-positve group had better disease-free/overall survival (p = 0.000) over 5 years’ follow-up. Furthermore, NDRG1 was considered to be an independent prognostic factor for overall survival (p = 0.001) and recurrence (p = 0.003). Our study concludes that NDRG1 is a novel favorable predictor for the prognosis in CRC patients.     

A tale of two multi‐focal therapies for glioblastoma: An antibody targeting ELTD1 and nitrone‐based OKN‐007

Glioblastoma (GBM) is the most common primary malignant brain tumour in adults. Despite a multimodal treatment response, survival for GBM patients remains between 12 and 15 months. Anti‐ELTD1 antibody therapy is effective in decreasing tumour volumes and increasing animal survival in an orthotopic GBM xenograft. OKN‐007 is a promising chemotherapeutic agent that is effective in various GBM animal models and is currently in two clinical trials. In this study, we sought to compare anti‐ELTD1 and OKN‐007 therapies, as single agents and combined, against bevacizumab, a commonly used therapeutic agent against GBM, in a human G55 xenograft mouse model. MRI was used to monitor tumour growth, and immunohistochemistry (IHC) was used to assess tumour markers for angiogenesis, cell migration and proliferation in the various treatment groups. OKN and anti‐ELTD1 treatments significantly increased animal survival, reduced tumour volumes and normalized the vasculature. Additionally, anti‐ELTD1 was also shown to significantly affect other pro‐angiogenic factors such as Notch1 and VEGFR2. Unlike bevacizumab, anti‐ELTD1 and OKN treatments did not induce a pro‐migratory phenotype within the tumours. Anti‐ELTD1 treatment was shown to be as effective as OKN therapy. Both OKN and anti‐ELTD1 therapies show promise as potential single‐agent multi‐focal therapies for GBM patients.