Candidate transcription factors involved in pancreatic endocrine development have been isolated using insulin gene regulation as a paradigm. The cell-type restricted basic helix–loop–helix (bHLH) gene, BETA2/NeuroD, expressed in pancreatic endocrine cells, the intestine, and the brain, activates insulin gene transcription and can induce neurons to differentiate. To understand the importance of BETA2 in pancreatic endocrine cell differentiation, mice lacking a functional BETA2 gene were generated by gene targeting experiments. Mice carrying a targeted disruption of theBETA2 gene developed severe diabetes and died perinatally. Homozygous BETA2 null mice had a striking reduction in the number of insulin-producing β cells and failed to develop mature islets. Islet morphogenesis appeared to be arrested between E14.5 and E17.5, a period characterized by major expansion of the β cell population. The presence of severe diabetes in these mice suggests that proper islet structure plays an important role in blood glucose homeostasis. In addition, secretin- and cholecystokinin-producing enteroendocrine cells failed to develop in the absence of BETA2. The absence of these two pancreatic secretagogs may explain the abnormal cellular polarity and inability to secrete zymogen granules in pancreatic acinar exocrine cells. The nervous system appeared to develop normally, despite abundant expression of BETA2 in differentiating neurons. Thus, BETA2 is critical for the normal development of several specialized cell types arising from the gut endoderm.

The BETA2 (neuroD) gene is expressed in endocrine cells during pancreas development and is essential for proper islet morphogenesis. The objective of this study is to identify potential upstream regulators of the BETA2 gene during pancreas development. We demonstrated that the expression of neurogenin 3 (ngn3), an islet- and neuron-specific basic-helix-loop-helix transcription factor, partially overlaps that of BETA2 during early mouse development. More importantly, overexpression of ngn3 can induce the ectopic expression of BETA2 in Xenopus embryos and stimulate the endogenous RNA of BETA2 in endocrine cell lines. Furthermore, overexpression of ngn3 could cause a dose-dependent activation on the 1.0-kb BETA2 promoter in islet-derived cell lines. Deletion and mutation analyses revealed that two proximal E box sequences, E1 and E3, could bind to ngn3-E47 heterodimer and mediate ngn3 activation. Based on these results, we hypothesize that ngn3 is involved in activating the expression of BETA2 at an early stage of islet cell differentiation through the E boxes in theBETA2 promoter.

Purpose: Recombinant tumor necrosis factor–related apoptosis-inducing ligand (TRAIL) is a promising antitumor agent. However, many hepatocellular carcinoma (HCC) cells show resistance to TRAIL-induced apoptosis. Here, we report that sorafenib improves the antitumor effect of TRAIL-related agents in resistant HCC. Experimental Design: HCC cell lines (PLC5, Huh-7, Hep3B, and Sk-Hep1) were treated with sorafenib and/or TRAIL-related agents (TRAIL or LBY135) and analyzed in terms of apoptosis and signal transduction. In vivo efficacy was determined in nude mice with PLC5 xenografts. Results: Sorafenib, the only approved drug for HCC, sensitizes resistant HCC cells to an agonistic DR5 antibody (LBY135) and TRAIL-induced apoptosis in TRAIL-resistant HCC cells. We found that STAT3 played a significant role in mediating TRAIL sensitization. Our data showed that sorafenib downregulated phospho-STAT3 (pSTAT3) and subsequently reduced the expression levels of STAT3-related proteins (Mcl-1, survivin, and cyclin D1) in a dose- and time-dependent manner in TRAIL-treated HCC cells. Knockdown of STAT3 by RNA interference overcame apoptotic resistance to TRAIL in HCC cells, and ectopic expression of STAT3 in HCC cells abolished the TRAIL-sensitizing effect of sorafenib. Moreover, SHP-1 inhibitor reversed downregulation of pSTAT3 and apoptosis induced by sorafenib, and silencing of SHP-1 by RNA interference abolished the effects of sorafenib on pSTAT3. Notably, sorafenib increased SHP-1 activity in PLC5 cells. Finally, sorafenib plus LBY135 significantly suppressed PLC5 xenograft tumor growth. Conclusions: Sorafenib sensitizes resistant HCC cells to TRAIL-induced apoptosis at clinical achievable concentrations, and this effect is mediated via the inhibition of STAT3. Clin Cancer Res; 16(21); 5189–99. ©2010 AACR.

Dysregulation of androgen signaling and pericellular proteolysis is necessary for prostate cancer progression, but the links between them are still obscure. In this study, we show how the membrane-anchored serine protease TMPRSS2 stimulates a proteolytic cascade that mediates androgen-induced prostate cancer cell invasion, tumor growth, and metastasis. We found that matriptase serves as a substrate for TMPRSS2 in mediating this proinvasive action of androgens in prostate cancer. Further, we determined that higher levels of TMPRSS2 expression correlate with higher levels of matriptase activation in prostate cancer tissues. Lastly, we found that the ability of TMPRSS2 to promote prostate cancer tumor growth and metastasis was associated with increased matriptase activation and enhanced degradation of extracellular matrix nidogen-1 and laminin β1 in tumor xenografts. In summary, our results establish that TMPRSS2 promotes the growth, invasion, and metastasis of prostate cancer cells via matriptase activation and extracellular matrix disruption, with implications to target these two proteases as a strategy to treat prostate cancer. Cancer Res; 75(14); 1–12. ©2015 AACR.

Pompe disease is caused by autosomal recessive mutations in the acid alpha-glucosidase (GAA) gene, which encodes GAA. Although enzyme replacement therapy has recently improved patient survival greatly, the results in skeletal muscles and for advanced disease are still not satisfactory. Here, we report the derivation of Pompe disease-induced pluripotent stem cells (PomD-iPSCs) from two patients with different GAA mutations and their potential for pathogenesis modeling, drug testing and disease marker identification. PomD-iPSCs maintained pluripotent features and had low GAA activity and high glycogen content. Cardiomyocyte-like cells (CMLCs) differentiated from PomD-iPSCs recapitulated the hallmark Pompe disease pathophysiological phenotypes, including high levels of glycogen and multiple ultrastructural aberrances. Drug rescue assessment showed that exposure of PomD-iPSC-derived CMLCs to recombinant human GAA reversed the major pathologic phenotypes. Furthermore, l-carnitine treatment reduced defective cellular respiration in the diseased cells. By comparative transcriptome analysis, we identified glycogen metabolism, lysosome and mitochondria-related marker genes whose expression robustly correlated with the therapeutic effect of drug treatment in PomD-iPSC-derived CMLCs. Collectively, these results demonstrate that PomD-iPSCs are a promising in vitro disease model for the development of novel therapeutic strategies for Pompe disease.