Mechanism Of Tob1 And Berberine In Colorectal Cancer Development

Colorectal cancer(CRC)is one of the third most common cancers in the world,and is seriously threatening the health of people all around the world.Various internal and external factors are responsible for the tumorigenesis of CRC,including chronic malignant progression of inflammation,long-term stimulation of various cancer-promoting factors,as well as a series of molecular events of a multi-stage,multi-step and multi-genes involvement,including abnormally activation of oncogenes,inactivation of tumor suppressor genes,etc.,which ultimately result in colorectal cancer development.In this paper,we used Tobl,a tumor suppressor,and Berberine,a traditional Chinese medicine monomer with anti-cancer activity as our research objects,and by studying two important aspects of colorectal cancer,i.e.the molecular development and the inflammation-carcinoma transformation of colorectal cancer,we try to get insights of the regulation mechanism of some key processes during the development of colorectal cancer.In many signaling pathways involved in the development of colorectal cancer,over-activation of Wnt signaling pathway is considered to be the most critical initiation event.While TOB1 is a widely-reported tumor suppressor gene,it can cross-talk with Wnt signaling pathway in colorectal cancer and jointly promote the cancer development.As a member of the TOB/BTG anti-proliferative family,Tobl has been proven by multiple studies to have an anti-proliferative function and is down-regulated or inactivated in various cancers.However,our study showed that Tobl has an abnormally elevated expression in colorectal cancer,in which Tobl has not been thoroughly studied before.So we first started with the clinical tissue samples,and verified that Tobl expression has been up-regulated indeed by the analysis of the tissue microarray.This conclusion was validated in vitro in multiple colorectal cancer cell lines.We then used colorectal cancer cell lines as a material to study the role of Tobl in colorectal cancer under in vitro conditions.We found that Tobl in colorectal cancer is mainly localized to the cytoplasm,which is different from the nuclear localization in other cancers,and that Tob1 can bind to p-catenin,the core component of Wnt signaling,enhance its stability,and ultimately activate the Wnt/β-catenin signaling pathway.The activated Wnt/α-catenin signaling pathway can increase the expression of Tobl in turn,so that a positive feedback loop is formed between these two parties,which up-regulating each other and ultimately promoting the proliferation of colorectal cancer cells.We further confirmed this conclusion in an in vivo mouse model and found that Tobl deficiency can reduce tumor formation and cancer cell proliferation in AOM/DSS chemical tumor-inducing model and ApcMin/+ genetic-disrupted mouse model;whereas in ApcMin/+ mice,in which Wnt/β-catenin signaling pathway was over-activated,we also detected an increasement in Tob1 expression by in situ hybridization.In summary,we have elucidated the unique pro-proliferation role of Tobl in colorectal cancer from clinical,in vitro and in vivo aspects,which can provide insights in better understanding of the molecular mechanism of colorectal cancer and finding new potential therapy targets for the treatment of colorectal cancer.Another special feature of colorectal cancer is that it is a typical example for inflammatory micro-environment of tumorigenesis.Inflammation is considered to be the seventh hallmarks of cancer,and in the colorectal system,inflammation is involved in every step of the tumorigenesis and development.Colitis-associated colorectal cancer(CACRC)is a subtype of colorectal cancer that has a tight connection with inflammatory bowel disease(IBD),which is difficult to treat and has a high mortality rate.As a traditional Chinese herbal medicines monomer,Berberine has been used to treat intestinal parasitic infections and bacterial diarrhea for a long time.The anti-inflammatory and anti-cancer functions of Berberine are receiving more and more attention in recent years.In a previous study,we established a CACRC mouse model by DSS treatment of ApcMin/+ mice,showing the advantages of Berberine in CACRC treatment.Both in vivo and in vitro studies demonstrated that Berberine can reduce pro-inflammatory cytokine production by regulating macrophages and inhibit inflammatory signaling-driven EGFR signaling in tumor cell growth.In order to prove with more evidences that Berberine does inhibit the production of pro-inflammatory cytokines in colonic macrophages,rather than solely targeting the cancer cells itself to inhibit CACRC,we established an advanced DSS-treating ApcMin/+ mouse model based on previous experiments,by supplementing an repeatedly intraperitoneal injection of pro-inflammatory cytokine(IL-6).The results showed that IL-6 treatment significantly attenuated the inhibitory effect of Berberine on CACRC.Therefore we came to the conclusion that Berberine inhibits CACRC at least partially through inhibiting the production of pro-inflammatory cytokines by colon macrophages.This result further clarified the detailed mechanism of the anti-tumor function of Berberine,and also suggested that Berberine can exert its anti-tumor function through its anti-inflammatory function in CACRC,making it a promising drug for treating CACRC.