1. MiR-34c/CTNND1 Signaling Pathway Is An Important Target For The Inhibition Of Liver Cancer By Chemotherapeutic Drugs. 2. The Abnormal Expression Of RNA-binding Protein DCAF13/TDRKH In Hepatocellular Carcinoma And Its Clinical Significance

Hepatocellular carcinoma(HCC)is one of the most common malignancies worldwide.The majority of HCC patients are first diagnosed when the disease has reached an advanced stage at which curative treatment is no longer possible.Transcatheter arterial chemoembolization(TACE)and hepatic arterial infusion chemotherapy(HAIC)have an important role in the treatment for advanced HCC.However,the molecular mechanisms of the chemotherapeutic drugs used for HCC treatment are not well-established.In our study,we found that miR-34s were activated by 5-fluorouracil and pirarubicin in HCC cells,suggesting that chemotherapy may hold clinical promise for HCC treatment through regulating miR-34-mediated program.We then performed functional and mechanistic analysis of miR-34s in HCC.Re-introduction of miR-34s can inhibit proliferation,metastasis and invasion of HCC cell lines in vitro and in vivo.We predicted the targets of miR-34s bioinformatically,and further western blot,immunofluorescence and luciferase reporter assay demonstrated that CTNND1 is a novel target of miR-34s in HCC.Then,we found that silencing CTNND1 can also decrease HCC cell proliferation,migration and invasion.Moreover,rescue assay was conducted to evaluate the role of CTNND1 in the miR-34c-mediated suppression of HCC cell phenotypes,and the result indicates that miR-34c suppresses HCC cell proliferation,migration and invasion at least partly through targeting CTNND1.To investigate whether chemotherapeutic drugs exerted their suppressive effect on HCC via miR-34/CTNND1 axis,a series of rescue assays was conducted.CCK8 cell proliferation,Wound-healing and in vitro invasion assay indicate that miR-34c inhibition to prevent their induction by 5-fluorouracil and pirarubicin treatment led to increased HCC cell proliferation,migration and invasion.These findings suggest that the miR-34/CTNND1 axis is a critical target of chemotherapy drugs in HCC.DDB1 and CUL4 Associated Factor 13(DCAF13)is a protein coding gene located in chromosome 8q22.3,which is a hotspot amplified in various cancers.DCAF13 has been reported to be frequently amplified in breast cancer patients.However,the genetic alteration and potential role of DCAF13 in other cancers,including hepatocellular carcinoma(HCC),have not been investigated yet.In this study,we found that DCAF13 was amplified in 14.7%of the cases and its expression was up-regulated(P<0.001)in HCC samples in The Cancer Genome Atlas(TCGA)dataset.Increased expression of DCAF13 was also noticed in 40 paired HCC and adjacent non-tumor tissues both at mRNA and protein levels(P=0.0002 and 0.0016,respectively).A positive relationship was observed between augmented DCAF13 levels and poorer tumor grade(P=0.005),and we also found that HCC patients with increased DCAF13 expression in their tumors had significantly poorer survival compared with those with DCAF13 low expression(median survival time,45.73 and 70.53 months,respectively).Multivariate Cox regression analysis showed that DCAF13 was an independent prognostic predictor of survival in HCC patients.GO and KEGG analysis indicated the potential role of DCAF13 as a crucial cell cycle regulator.Collectively,our findings revealed that the overexpression of DCAF13 in HCC was significantly associated with poor survival and may participate in the regulation of cell cycle progression.TDRKH is a RNA binding protein,mainly involved in germ cell maturation and piRNA biogenesis.However,the expression and functional role of TDRKH in tumorigenesis is largely unknown.Herein,we tend to investigate the expression and clinical implication of TDRKH in HCC.In the TCGA HCC dataset,we found that TDRKH was significantly up-regulated in HCC(P<0.0001).Increased expression of TDRKH was also noticed in the fresh HCC tissues both at RNA and protein levels(P=0.0002).Augmented TDRKH expression was significantly associated with poorer tumor grade(P=0.0056)and increased AFP value(P=0.0078).Using Kaplan-Meier analysis,we found that HCC patients with increased TDRKH expression had significant poorer disease free survival compared with those with TDRKH low expression(P=0.002).Multivariate Cox regression analysis showed that TDRKH was an independent prognostic predictor of disease free survival in HCC patients(HR=2.16,P=0.003).GO and KEGG analysis indicated the potential role of TDRKH in the P53-mediated DNA repair pathway.