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The Hippo/MST signaling pathway is a critical player in controlling cell proliferation, self-renewal, differentiation, and apoptosis of most tissues and organs in diverse species. Previous studies have shown that Salvador homolog 1 (SAV1), a scaffolding protein which functions in the signaling system is expressed in mammalian ovaries and play a vital role in governing the follicle development. But the exact biological effects of chicken SAV1 in prehierarchical follicle development remain poorly understood. In the present study, we demonstrated that the SAV1 protein is predominantly expressed in the oocytes and undifferentiated granulosa cells in the various sized prehierarchical follicles of hen ovary, and the endogenous expression level of SAV1 mRNA appears down-regulated from the primordial follicles to the largest preovulatory follicles (F2-F1) by immunohistochemistry and real-time RT-PCR, respectively. Moreover, we found the intracellular SAV1 physically interacts with each of the pathway members, including STK4/MST1, STK3/MST2, LATS1 and MOB2 using western blotting. And SAV1 significantly promotes the phosphorylation of LATS1 induced by the kinase of STK4 or STK3 in vitro. Furthermore, SAV1 knockdown by small interfering RNA (siRNA) significantly increased proliferation of granulosa cells from the prehierarchical follicles (6–8 mm in diameter) by BrdU-incorporation assay, in which the expression levels of GDF9, StAR and FSHR mRNA was notably enhanced. Meanwhile, these findings were consolidated by the data of SAV1 overexpression. Taken together, the present results revealed that SAV1 can inhibit proliferation of the granulosa cells whereby the expression levels of GDF9, StAR and FSHR mRNA were negatively regulated. Accordingly, SAV1, as a member of the hippo/MST signaling pathway plays a suppressive role in ovarian follicle development by promoting phosphorylation and activity of the downstream LATS1, may consequently lead to prevention of the follicle selection during ovary development. [ABSTRACT FROM AUTHOR]

Forkhead box L2 (FOXL2) is a member of the forkhead nuclear factor 3 gene family and plays an essential role in ovarian growth and maturation in mammals. However, its potential effects and regulative mechanism in development of chicken ovarian prehierarchical follicles remain unexplored. In this study, the cooperative effects of FOXL2 with activin A, growth differentiation factor-9 (GDF9) and follistatin, three members of the transforming growth factor beta (TGF-β) superfamily that were previously suggested to exert a critical role in follicle development was investigated. We demonstrated herein, using in-situ hybridization, Northern blot and immunohistochemical analyses of oocytes and granulosa cells in various sizes of prehierarchical follicles that both FOXL2 transcripts and FOXL2 proteins are predominantly expressed in a highly similar expression pattern to that of GDF9 gene. In addition, the FOXL2 transcript was found at lower levels in theca cells in the absence of GDF9. Furthermore, culture of granulosa cells (GCs) from the prehierarchical follicles (6–8 mm) in conditioned medium revealed that in the pcDNA3.0-FOXL2 transfected GCs, there was a more dramatic increase in FSHR mRNA expression after treatment with activin A (10 ng/ml) or GDF9 (100 ng/ml) for 24 h which caused a stimulatory effect on the GC proliferation. In contrast, a significant decrease of FSHR mRNA was detected after treatment with follistatin (50 ng/ml) and resulted in an inhibitory effect on the cell proliferation. The results of this suggested that FOXL2 plays a bidirectional modulating role involved in the intracellular FSHR transcription and GC proliferation via an autocrine regulatory mechanism in a positive or negative manner through cooperation with activin A and/or GDF9, and follistatin in the hen follicle development. This cooperative action may be mediated by the examined Smad signals and simultaneously implicated in modulation of the StAR, CCND2, and CYP11A1 expression. [ABSTRACT FROM AUTHOR]

Cadherin-17 (CDH17), one member of 7D-cadherin superfamily, was overexpressed in gastric cancer (GC) and was associated with poor survival, tumor recurrence, metastasis, and advanced tumor stage. So far the cellular function and signaling mechanism of CDH17 in GC remains unclear. In this study, we showed that over 66% of GC cell lines (20/30) were CDH17 positive. Tissue microarray (TMA) assay showed that 73.6% Chinese GC tissues (159/216) were CDH17 positive, while 37% respective adjacent normal tissues were CDH17 positive. Knockdown of CDH17 inhibited cell proliferation, migration, adhesion and colony formation, and also induced a cell cycle arrest and apoptosis in AGS human GC cells. On the other side, overexpression of CDH17 facilitated MGC-803 GC tumor growth in nude mice. Antibody array and Western blotting assay demonstrated that knockdown of CDH17 in AGS cells down-regulated integrin β series proteins, further inactivated the Ras/Raf/MEK/ERK pathway and led to p53 and p21 accumulation, which resulted in proliferation inhibition, cell-cycle arrest and apoptosis induction. Collectively, our data firstly demonstrate the capacity of CDH17 to regulate the activity of Ras/Raf/MEK/ERK pathway for cell proliferation in GC, and suggest that CDH17 can serve as an attractive therapeutic target for future research. [ABSTRACT FROM AUTHOR]

Protein kinases play important roles in tumor development and progression. Lots of kinase inhibitors have entered into market and show promising clinical benefits. Here we report the discovery of a novel small molecule, well-tolerated, orally active kinase inhibitor, R1498, majorly targeting both angiogenic and mitotic pathways for the treatment of hepatocellular carcinoma (HCC) and gastric cancer (GC). A series of biochemical and cell-based assays indicated that the target kinase cluster of R1498 included Aurora kinases and VEGFR2 et al. R1498 showed moderate in vitro growth inhibition on a panel of tumor cells with IC50 of micromole range. The in vivo anti-tumor efficacy of R1498 was evaluated on a panel of GC and HCC xenografts in a parallel comparison with another multikinase inhibitor sorafenib. R1498 demonstrated superior efficacy and toxicity profile over sorafenib in all test models with >80% tumor growth inhibition and tumor regression in some xenogratfts. The therapeutic potential of R1498 was also highlighted by its efficacy on three human GC primary tumor derived xenograft models with 10–30% tumor regression rate. R1498 was shown to actively inhibit the Aurora A activity in vivo, and decrease the vascularization in tumors. Furthermore, R1498 presented good in vivo exposure and therapeutic window in the pharmacokinetic and dose range finding studies. Theses evidences indicate that R1498 is a potent, well-tolerated, orally active multitarget kinase inhibitor with a unique antiangiogenic and antiproliferative profile, and provide strong confidence for further development for HCC and GC therapy. [ABSTRACT FROM AUTHOR]

Hepatitis B virus (HBV) infection is a worldwide liver disease and nearly 25% of chronic HBV infections terminate in hepatocellular carcinoma (HCC). Currently, there is no effective therapy to inhibit HBV replication and to eliminate hepatoma cells, making it highly desired to develop novel therapies for these two stages of the HBV-caused detrimental disease. Recently, short hairpin RNA (shRNA) has emerged as a potential therapy for virus-infected disease and cancer. Here, we have generated a shRNA, pGenesil-siHBV4, which effectively inhibits HBV replication in the human hepatoma cell line HepG2.2.15. The inhibitory effects of pGenesil-siHBV4 are manifested by the decrease of both the HBV mRNA level and the protein levels of the secreted HBV surface antigen (HBsAg) and HBV e antigen (HBeAg), and by the reduction of secreted HBV DNA. Using mouse hydrodynamic tail vein injection, we demonstrate that pGenesil-siHBV4 is effective in inhibiting HBV replication in vivo. Because survivin plays a key role in cancer cell escape from apoptosis, we further generated pGenesil-siSurvivin, a survivin-silencing shRNA, and showed its effect of triggering apoptosis of HBV-containing hepatoma cells. To develop targeted shRNA therapy, we have identified that as a specific binder of the asialoglycoprotein receptor (ASGPR), jetPEI-Hepatocyte delivers pGenesil-siHBV4 and pGenesil-siSurvivin specifically to hepatocytes, not other types of cells. Finally, co-transfection of pGenesil-siHBV4 and pGenesil-siSurvivin exerts synergistic effects in inducing hepatoma cell apoptosis, a novel approach to eliminate hepatoma by downregulating survivin via multiple mechanisms. The application of these novel shRNAs with the jetPEI-Hepatocyte targeting strategy demonstrates the proof-of-principle for a promising approach to inhibit HBV replication and eliminate hepatoma cells with high specificity. [ABSTRACT FROM AUTHOR]