Shuai Zhao

Prohibitin overexpression predicts poor prognosis and promotes cell proliferation and invasion through ERK pathway activation in gallbladder cancer

AbstractBackgroundProhibitin (PHB), a pleiotropic protein overexpressed in several tumor types, has been implicated in the regulation of cell proliferation, invasive migration and survival. However, PHB expression and its biological function in gallbladder cancer (GBC) remain largely unknown.MethodsPHB and p-ERK protein expressions were determined in human GBC tissues by immunohistochemistry (IHC). The effects of PHB knockdown on GBC cell proliferation and invasiveness were evaluated using Cell Counting Kit-8 (CCK-8) cell viability, cell cycle analysis, transwell invasion and gelatin zymography assays. Subcutaneous xenograft and tail vein-lung metastasis tumor models in nude mice were employed to further substantiate the role of PHB in GBC progression.ResultsPHB protein was overexpressed in GBC tissues and was significantly associated with histological grade, tumor stage and perineural invasion. Furthermore, PHB expression was negatively associated with overall survival in GBC patients. In vitro experimental studies demonstrated that the downregulation of PHB expression by lentivirus-mediated shRNA interference not only inhibited the ERK pathway activation but also reduced the proliferative and invasive capacities of GBC cells. Moreover, PD0325901, a specific inhibitor of MEK, markedly impaired PHB- mediated phosphorylation of ERK protein. IHC statistical analyses further validated that PHB expression was positively correlated with ERK protein phosphorylation levels in GBC tissue samples. In vivo, PHB depletion also resulted in dramatic reductions in the growth and metastasis of GBC cells.ConclusionOur findings demonstrate that PHB overexpression predicts poor survival in GBC patients. PHB could serve as a novel prognostic biomarker and a potential therapeutic target for GBCs.

A novel PI3K/AKT signaling axis mediates Nectin-4-induced gallbladder cancer cell proliferation, metastasis and tumor growth

Nectin-4 is a Ca(2+)-independent immunoglobulin-like cell adhesion molecule which has diverse functions in cell-cell adhesion via homophilic and heterophilic interactions. Cell-cell adhesive processes are central to cell polarization, differentiation, proliferation, survival and movement. Here we report that Nectin-4 is substantially overexpressed in gallbladder cancer (GBC), the most common biliary tract malignancy with a high risk of local tumor spread and invasion. Further, Nectin-4 high expression in GBC patients was associated with pathologic T stage and lymph node metastasis status, and the expression level of the downstream target Rac1 and poor prognoses were also correlated with Nectin-4. Ectopic expression of Nectin-4 promoted GBC cell growth, motility and tumor growth in a mouse model. The depletion of Nectin-4 inhibited GBC cell proliferation and migration both in cell culture and in mice. Our data suggest that activation of the PI3K/AKT pathway was involved in the oncogenic function of Nectin-4 to activate Rac1 in GBC. Inhibition of PI3K/AKT with LY294002 and/or Rac1 with NSC23766 impaired Nectin-4-mediated GBC cell proliferation and motility. We hypothesize that Nectin-4 is critical for GBC progression via PI3K/AKT pathway activation of Rac1. Nectin-4 may be a novel prognostic factor and therapeutic target in GBC patients.

Zinc finger X-chromosomal protein (ZFX) is a significant prognostic indicator and promotes cellular malignant potential in gallbladder cancer

The Zinc finger X-chromosomal protein (ZFX), a novel member of the Krueppel C2H2-type zinc finger protein family, has been implicated in multiple human cancers. However, the clinical significance of ZFX expression in gallbladder cancer (GBC) remains largely unknown. In this study, we focused on the clinical significance, biological function and mechanism of ZFX in GBC, and found that ZFX protein overexpression was frequently detected in GBC tissues. The expression of ZFX was significantly correlated with histological grade, perineural invasion, and margin status and lead to a significantly poorer prognosis in GBC patients(P <0.001). Furthermore, knockdown of ZFX result in significant inhibition of proliferation, migration, invasion and cause cell cycle arrest in GBC-SD cells, while over-expression of ZFX in NOZ shows the opposite results. Activation of PI3K/AKT pathway maybe the potential mechanism behind these effects. In conclusion, ZFX may serve as a oncogene and could be used as a potential prognostic marker and genetic treatment target for GBC patients.

LASP-1 induces proliferation, metastasis and cell cycle arrest at the G2/M phase in gallbladder cancer by down-regulating S100P via the PI3K/AKT pathway.

LASP-1 is an actin-binding protein that regulates cytoskeletal dynamics and cell migration. LASP-1 was previously identified in a cDNA library from metastatic breast cancer samples. This protein has since been detected in multiple human cancers, including liver cancer, gastric cancer and pancreatic cancer. S100P is a small calcium-binding protein in the S100 protein family that regulates cellular, physiological and pathological processes in various cancers. However, the clinical significance of LASP-1 and S100P expression in gallbladder cancer (GBC) is not yet clear. In our study, we focused on the clinical significance, biological function and mechanism of LASP-1 in gallbladder cancer and detected LASP-1 and S100P overexpression in GBC tissues. The expression of LASP-1 was significantly correlated with poor prognosis in GBC patients (P < 0.05). Furthermore, down-regulation of LASP-1 expression resulted in the obvious inhibition of proliferation and migration and caused cell cycle arrest by down-regulating S100P via the PI3K/AKT pathway; in mice, tumor volume was significantly decreased. In conclusion, LASP-1 may act as an oncogene to regulate the expression of S100P to influence cellular functions in GBC. LASP-1 could serve as a genetic treatment target in GBC patients.

miR-223 increases gallbladder cancer cell sensitivity to docetaxel by downregulating STMN1

Background MicroRNAs (miRs) are involved in cancer carcinogenesis, and certain regulatory miRs could provide promising therapeutic methods for refractory malignancies, such as gallbladder cancer (GBC). miR-223 was found to play a pivotal role in enhancing chemotherapeutic effects, therefore evoking interest in the role of miR-223 in GBC. Results miR-223 was decreased in GBC tissues and cell lines, and ectopic miR- 223 expression exhibited multiple anti-tumorigenic effects in GBC cells, including decreased proliferation, migration and invasion in vitro. However, treatment with a miR-223 inhibitor increased cell viability. We determined that STMN1 was negatively correlated with and regulated by miR-223 in GBC. miR-223 increased GBC sensitivity to docetaxel in vitro and in vivo, and the induced sensitivity to docetaxel was suppressed by the restoration of STMN1 expression. Methods We examined miR-223 expression in GBC tissue and GBC cell lines using qRT-PCR. The effects of modulated miR-223 expression in GBC cells were assayed using Cell Counting Kit-8 (CCK8), flow cytometry, and wound-healing and invasion assays. Susceptibility to docetaxel was evaluated in miR-223/STMN1-modulated GBC cells and xenograft tumor models. The protein expression of relevant genes was examined by Western blotting. Conclusions These findings indicated that miR-223 might serve as an onco-suppressor that enhances susceptibility to docetaxel by downregulating STMN1 in GBC, highlighting its promising therapeutic value.

Upregulated LASP-1 correlates with a malignant phenotype and its potential therapeutic role in human cholangiocarcinoma.

LIM and SH3 protein 1 (LASP-1) is demonstrated to play a key role in occurrence and development of tumors. However, the expression and function of LASP-1 in cholangiocarcinoma (CCA) remain largely unexplored. This study aimed to investigate the effect of regulated LASP-1 expression on migration, invasion, proliferation, and apoptosis of CCA cells and on tumorigenesis in vivo, and to examine clinico-oncological correlates of LASP-1 expression. Expression of LASP-1 by immunohistochemistry was evaluated in CCA tissue samples. HCCC-9810 and RBE cells were transfected with the LASP-1 small interfering RNA (siRNA), and the effect of knocking down LASP-1 gene expression on cell migration, invasion, proliferation, and apoptosis were examined by wound healing, transwell assays, CCK-8 assays, colony formation, and flow cytometry assays, respectively. Xenograft tumor model was used to validate the effect of downregulated LASP-1 in vivo. Our results demonstrated that LASP-1 was over-expressed in CCA tissues, positively correlating with larger tumors, poor histological differentiation, lymph node metastasis, advanced TNM stage, and poor prognosis in CCA patients (P < 0.05). Downregulation of LASP-1 in HCCC-9810 and RBE cell lines significantly increased cell apoptosis and suppressed cell migration, invasion, and proliferation in vitro and tumorigenesis in vivo. Our results indicate that LASP-1 may essentially involve in the metastasis and growth of CCA and clinical significance of LASP-1 may reside in function as a biomarker to predict prognosis and as a promising therapeutic strategy for CCA patients by the inhibition of LASP-1 expression.

A three-step method for modular lymphadenectomy in gastric cancer surgery: The ability to retrieve sufficient lymph nodes and improve survival

BACKGROUND Systematic lymphadenectomy for the resection of sufficient lymph nodes is the most important part of curative resection in gastric cancer surgery. Here, we explore the outcomes of the three-step method for modular lymphadenectomy (TSMML) and determine its safety and efficacy, compared with the conventional method for lymphadenectomy (CML). METHODS From 2008 to 2011, 270 patients with gastric cancer were divided into 2 subgroups: the TSMML group and the CML group. RESULTS Patients in the TSMML group had a significantly higher median number of retrieved lymph nodes (rLNs), lower median metastatic lymph node ratios (MLRs), and superior 5-year relapse-free survival (RFS) than the CML group. Moreover, the use of the TSMML procedure was an independent protective factor for RFS. No significant intergroup differences were found in morbidity or mortality in these two groups. CONCLUSION The TSMML procedure is safe and effective and is easy to learn.

Whole-genome sequencing of an advanced case of small-cell gallbladder neuroendocrine carcinoma

The majority of gallbladder cancer cases are discovered at later stages, which frequently leads to poor prognoses. Small-cell gallbladder neuroendocrine carcinoma (GB-SCNEC) is a relatively rare histological type of gallbladder cancer, and its survival rate is exceptionally low because of its greater malignant potential. In addition, the genomic landscape of GB-SCNEC is rarely considered in treatment decisions. We performed whole-genome sequencing on an advanced case of GB-SCNEC. By analyzing the whole-genome sequencing data of the primary cancer tissue (76.29X coverage), lymphatic metastatic cancer tissue (73.92X coverage) and matched non-cancerous tissue (35.73X coverage), we identified approximately 900 high-quality somatic single nucleotide variants (SNVs), 109 of which were shared by both the primary and metastatic tumor tissues. Somatic non-synonymous coding variations with damaging impact in HMCN1 and CDH10 were observed in both the primary and metastatic tissue specimens. A pathway analysis of the genes mapped to the SNVs revealed gene enrichment associated with axon guidance, ERBB signaling, sulfur metabolism and calcium signaling. Furthermore, we identified 20 chromosomal rearrangements that included 11 deletions, 4 tandem duplications and 5 inversions that mapped to known genes. Two gene fusions, NCAM2-SGCZ and BTG3-CCDC40 were also discovered and validated by Sanger sequencing. Additionally, we identified genome-wide copy number variations and microsatellite instability. In this study, we identified novel biological markers of GB-SCNEC that may serve as valuable prognostic factors or indicators of treatment response in patients with GB-SCNEC with lymphatic metastasis.