The Role Of Dapper1 On The Progression Of Hepatocellular Carcinoma And Its Underlying Mechanism

Part I The Role of Dapper1 on the Progression of Hepatocellular Carcinoma and Its MechanismHepatocellular carcinoma(HCC)is one of the most common malignancies known for its poor prognosis and high mortality,causing the third cancer-related death worldwide.There are more than 600,000 newly diagnosed cases every year globally,of which nearly 55% are found in China.The prognosis of HCC is far from desirable due to high heterogeneity of HCC and a lack of effective chemotherapy or biotherapy.Therefore,elucidating essential pathways involved in tumorigenesis and tumor progression is of importance for potential personalized targeted therapy.Of many activated pathways associated with HCC progression,Wnt/β-catenin pathway plays important roles in regulating cancer cell differentiation,proliferation,epithelial-mensenchymal transition and stem cell renewal.Of all the gene mutations accumulated in tumor development,β-catenin ranks as the second most frequently,observed in 20%-40% of HCC samples.Therefore,the activation of Wnt/β-catenin pathway is considered one of the important features for the clinical classification of HCC.In the canonical Wnt signaling pathway,the soluble form of the core factorb-catenin is mainly targeted for ubiquitination and proteasomal degradation,which is promoted by the destruction complex of Axin,adenomatous polyposis coli(APC)2,and glycogen synthase kinase 3b.The binding of Wnt to its cell surface receptors,Frizzled(Fz)and low density lipoprotein receptor-related proteins 5 and 6(LRP5/6)leads to the recruitment of Dishevelled(Dvl)to Fz and Axin to LRP5/6,resulting in the disruption of the destruction complex and consequently the accumulation of b-catenin.The accumulated b-catenin moves into the nucleus and activates the transcription of target genes through interacting with T-cell factor(TCF)/lymphoid enhancer binding factor(LEF)transcription factors.Dapper(Dpr),which was first identified as an interacting protein of Dvl,a central mediator in Wnt signaling,functioning as a general antagonist of Dvl to modulate the b-catenin-dependent and JNK-dependent Wnt signaling.Several Dpr orthologs,including Dpr1,Dpr2 and Dpr3,have been identified.Dpr family members positively regulate Wnt signaling through distinct mechanisms during embryo development.Recently Dpr has also been implicated to be associated with tumorigenesis.Researchers found that human Dpr1 was down-regulated in hepatocellular carcinoma,which was correlated with the cytoplasm accumulation of b-catenin.However,the function of Dpr1 in HCC tumorigenesis and the underlying mechanism are still unclear.Autophagy is a catabolic process with crucial roles in maintaining intracellular metabolic homeostasis,and could be also induced in response to starvation,protein aggregation,and other forms of stress such as oxidative and endoplasmic reticulum(ER)stress.Abundant evidence has revealed that autophagy is involved in the pathogenesis of various liver diseases,such as viral hepatitis,non-alcoholic fatty liver disease,alcoholic liver disease,fibrosis,cirrhosis,and HCC.The mechanism of autophagy has been extensively investigated and suggests a dual role of autophagy in the development and promotion of HCC.Early studies have demonstrated a tumor suppressive function of autophagy,while more recent studies have uncovered the dark side of it in tumorigenesis.Due to the above understanding of autophagy in the progression of HCC,it is indicated that autophagy-modulating compounds may bear potential value in the treatment of HCC in the near future.p62/SQSTM1 is a multifunctional signaling hub and autophagy adaptor with many binding partners,which allows it to activate mTORC1-dependent nutrient sensing,NF-kB-mediated inflammatory responses,and NRF2-activated antioxidant defense.p62 recognizes polyubiquitin chains and binds to LC3 via its LIR motif,thereby promoting the autophagic degradation of ubiquitinated cargos through autophagy.p62 accumulates when autophagy is compromised,however,by activating NRF2 and mTORC1 pathways,a positive feedback loop could be established,resulting in massive p62 accumulation.p62 upregulation is observed in many human liver diseases,including nonalcoholic steatohepatitis(NASH)and hepatocellular carcinoma(HCC),where it is a component of Mallory–Denk bodies and intracellular hyaline bodies.Chronic p62 elevation contributes to HCC development by preventing oncogene-induced senescence and death of cancer-initiating cells as well as enhancing their proliferation.Therefore,elucidating p62-mediated signaling pathways and their roles in liver pathophysiology is beneficial to the development of autophagy or p62 based therapies in the treatment of HCC.Besides its function in regulating Wnt signaling pathway,Dpr1 also plays a role in autophagy promoting.In a recent study,it has revealed that Dpr1 directly interacts with Beclin1 and Atg14 L,enhancing Beclin1-Vps34 interaction and Vps34 activity.As Beclin1-Vps34-Atg14 L complex is essential for autophagosome formation,Dpr1 could significantly enhances autophagy in the central nervous system as proved in the conditional Dpr1-knockout mice model.To explore the role of Dpr1 in HCC development and progression,we have found deprivation of Dpr1 is associated with declined formation of autophagy flux and could notably promote tumorigenesis in DEN-induced HCC mice model in our preliminary study.In order to examine the function of Dpr1 in adult mouse tissues,we generated hepatocyte Dpr1-knockout(Dact1hep-/-)mice and tried to elucidate its role in HCC tumorigenesis and if autophagy or Wnt signaling pathway activation is involved in the progress.In seek of the mechanism of Dpr1 function,we explored its value in evaluating the occurrence of HCC in population with high risks and its indication in the prognosis of HCC patients,possibly extending its function in personalized treatment of HCC.Part 2 Hepatitis B Virus x Protein Promotes Metastasis of Hepatocellular Carcinoma via Induced Secretion of HMGB1Hepatocellular carcinoma is one of the most common cancers worldwide,ranked as the third leading cause of cancer-related death.Chronic hepatitis B virus(HBV)infection has been identified as a major risk factor for HCC.HBV infection could develop into chronic liver diseases(CLD),which appears as hepatitis,fibrosis,cirrhosis and finally the onset of HCC.The poor prognosis of HCC is mainly caused by the high frequency of metastasis and de novo tumor formation in the diseased liver.In addition,HBV reactivation is common after partial hepatectomy for HBV related HCC and might influence postoperative survival.Sustained HBV suppression is associated with long-term survival.Postoperative antiviral treatment is expected to reduce viral load and relieve hepatic inflammation,thus improving postoperative prognosis.Among the proteins coded by four partially overlapping open reading frames in HBV genomic DNA,the hepatitis B virus X protein(HBx),a small 17-k Da soluble protein in the nucleus and cytoplasm of host cells,is known to be essential for HBV induced carcinogenesis.HBx appears to play a role on carcinogenesis and progression of HCC by promoting cell proliferation,and invasive potential.Several pioneering studies reported that HBx interacts with p53 and disrupts its function as a transcription factor.HBx modulates the transcriptional activation of AP-1,AP-2,and NF-k B.It can also upregulate the Ras/Raf/MAPK,PI3K/Akt,and JAK/STAT signaling pathways.However,the molecular mechanism involved in the association of HBx with HCC development and progression remains to be completely understood.HMGB1 was discovered as an evolutionarily conserved,chromatin-binding protein acting as a DNA chaperone.Besides acting as a late mediator in the systemic inflammation of sepsis,extracellular HMGB1 also plays a role as an early mediator in liver ischemia perfusion injury.Also HMGB1 has been reported to play a role in human malignancies,including liver,prostate,breast,colon cancers and malignant mesothelioma,linking to poor prognosis.During hypoxia,HMGB1 could be released from HCC cells followed by subsequent production of multiple inflammatory mediators,to promote cancer invasion and metastasis.As tumor microenvironment has recently been considered as the emerging hallmark of cancer,the role of HMGB1 in affecting tumor microenvironment by regulating inflammation to promote tumor progression should be reexamined.In this study,we provided evidence that HBx enhanced the secretion of HMGB1 from HCC cell lines in vitro.By blocking the function of HMGB1,the invasiveness and metastasis of tumor cells expressing HBX was significantly impaired.In addition,we showed that HBx induced secretion of HMGB1 was cytoplasmic calcium dependent,via modulating calcium/Ca MKK/CAMKIV pathway.Both serum HMGB1 level and cytoplasmic HMGB1 level were positively correlated with HBV load in HCC patients.Our data suggests that HMGB1 could be used as a potential marker for prognosis and therapeutic target of HBV-related HCC.