Study On The Relationship Between HRP - 3 And The Development

Hepatocellular carcinoma (HCC) is one of the most malignant tumors in China and typically has poor prognosis, due to the fact that diagnosis is frequently made at an advanced stage. Heterogeneous phenotypic and genetic traits of affected individuals and a wide range of risk factors have classified it a complex disease. HCC is not amenable to standard chemotherapy and is resistant to radiotherapy. In most cases, surgical resection and liver transplantation remain the only curative treatment options. However, recurrence or metastasis is quite common in patients who have had a resection and survival rate is30%to40%at5years postoperatively. Therefore, development of novel, effective targets and therapies is of prime importance.HRP-3(hepatoma-derived growth factor related protein3) is a member of HDGF (hepatoma-derived growth factor) family. This family comprise6members, including human HDGF, murine HRP-1, murine HRP-2, murine HRP-3, bovine HRP-4, and lens epithelium-derived growth factor (LEDGF). These proteins share a common N-terminal HDGF amino terminus (HATH) homology domain, but the C-terminal regions show considerable variation in length and charge.HDGF is expressed in a variety of tissues. HRP-3, however, expressed mainly in human testis, brain and heart, but less in other tissues. In our lab, we found that HRP-3exhibited remarkable up-regulation in liver cancer samples on both mRNA and protein levels. Abrupt changes that occur in liver tissues due to either viral infection, genetic risks or exposure to hepatotoxic agents often alter gene expression and result in tumor formation. Therefore, HRP-3maybe is related to the incidence and development of liver cancer.In our work, we found that down-regulation of HRP-3resulted in no detectable effect on anchorage-dependent cell growth. In contrast, stable cell lines of SMMC-7721or SK-Hep-1infected with lentivirus carrying HRP-3-siRNA grew more slowly and formed significantly fewer colonies in soft agar than did cells infected with lentivirus harboring nonsilence control siRNA. Furthermore, down-regulation of HRP-3also induced the dephosphorylation of ERK. ERK signal pathway is often up-regulated in most kinds of tumors and over activation of ERK may result in tumor formation. At meanwhile, we found that recombinant HRP-3protein could also enhanced ERK pathway and made cells grew faster and formed more colonies in soft agar.To further determine the role of HRP-3in progression of HCC, we did an in vivo animal experiment. In nude mice model, stable cell lines SMMC-7721or SK-Hep-1infected with HRP-3-siRNA-lentivirus grew significantly slower than the cell lines infected with nonsilence-siRNA-lentivirus. And the tumor volume for mice of the former is much smaller compared with the latter. We did Western blot analysis of the tumor sample to compare ERK pathway in nude mice model. The results are consistent with or even better than the experiment in vitro. We also established stable cell.lines overexpressing exogenous HRP-3. The stable cell lines overexpressing exogenous HRP-3grew faster and formed bigger tumor than control stable cell lines. Stable cell lines of NIH3T3cells overexpressing HRP-3could neither form colonies in soft agar nor tumor in nude mice, which suggests us HRP-3can not induce tumorgenesis.In our work, HRP-3could be up-regulated by components of tumor microenvironment, including serum or glucose depletion and VEGF. It suggested that HRP-3may engage in angiogenesis and antagonism of apoptosis, not only cell proliferation.Remarkably, higher sensitivity to CPT was observed in stable cell lines infected with HRP-3-siRNA-lentivirus than control stable cell lines. HRP-3RNAi alone could significantly reduce the tumor volume, and whether a combination of CPT and HRP-3RNAi can reduce the tumor more effectively will be study in nude mice model.In summary, our work demonstrates the role of HRP-3in human HCC. We found that HRP-3exhibited remarkable up-regulation in HCC, promoted tumor proliferation through activating of ERK pathway and enhancing anchorage-independent growth of cells, and could be up-regulated by components of tumor microenvironment and effectively enhance in vitro sensitivity to chemotherapeutic agents. All these results suggest us that HRP-3could serve as a potential prognostic biomarker and therapeutic target for human HCC.