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In the present study, we attempted to develop a lecithin-stabilized micellar drug delivery system (LsbMDDs) for loading docetaxel (DTX) to enhance its therapeutic efficacy and minimize systemic toxicity. A novel DTX-loaded LsbMDDs was optimally prepared by a thin-film hydration method and then hydrated with a lecithin nanosuspension while being subjected to ultrasonication. Physical characteristics of the optimized DTX-loaded LsbMDDs formulations were examined and found to have a mean size of 90%, and drug loading of >6% with stability at room temperature and at 4 °C being longer than 2 and 7 days, respectively. The in vitro release of DTX from the DTX-loaded LsbMDDs was slower than that from the generic product of DTX (Tynen®). A cell viability assay demonstrated that the LsbMDDs showed better cytotoxicity than Tynen® against CT26 cancer cells. The in vivo antitumor efficacy of the DTX-loaded LsbMDDs was observed to be better than that of Tynen® in a CT26 tumor-bearing mice model. A high-dose regimen of the DTX-loaded LsbMDDs formulation showed greater inhibition of DU145 tumor growth than did Tynen®, but with less to similar systemic toxicity. An in vivo study also showed that a greater amount of drug was able to accumulate in the tumor site with the DTX-loaded LsbMDDs, and its maximal tolerable doses for single and repeated injections were 2–2.5-fold higher than those of Tynen®. In conclusion, the LsbMDDs could be a promising high drug-loaded nanocarrier for delivering hydrophobic chemotherapeutic agents that can enhance the efficacy of chemotherapy and reduce systemic toxicity.

KLF10 is a transforming growth factor (TGF)-β/Smad downstream regulated gene. KLF10 binds to the promoter of target genes and mimics the effects of TGF-β as a transcriptional factor. In our laboratory, we noted that Klf10 deficiency in mice is associated with significant inflammation of the lungs. However, the precise mechanism of this association remains unknown. We previously identified NPRA as a target gene potentially regulated by KLF10 through direct binding; NPRA knockout have known that prevented lung inflammation in a mouse model of allergic asthma. Here, we further explored the regulatory association between KLF10 and NPRA on the basis of the aforementioned findings. Our results demonstrated that KLF10 acts as a transcriptional repressor of NPRA and that KLF10 binding reduces NPRA expression in vitro. Compared with wild-type mice, Klf10-deficient mice were more sensitive to lipopolysaccharide or ovalbumin challenge and showed more severe inflammatory histological changes in the lungs. Moreover, Klf10-deficient mice showed pulmonary neutrophil accumulation. These findings collectively reveal the precise site where KLF10 signaling affects pulmonary inflammation by attenuating NPRA expression. They also verify the importance of KLF10 and atrial natriuretic peptide/NPRA in exerting influences on chronic pulmonary disease pathogenesis.

Background: Proton pump inhibitors (PPIs) are used for the long-term treatment of gastroesophageal disorders and the non-prescription medicines for acid reflux. However, there is growing concerns about PPI misuse, overuse and abuse. This study aimed to develop an animal model to examine the effects of long-term use of PPI in vivo. Methods: Twenty-one Wistar rats were given omeprazole orally for 30 days. After euthanization, the serum and stool were collected to perform MS-based quantitative analysis of metabolites. We carried out 16S rRNA gene profiling of fecal microbiota, assessed the expression of bile acid metabolism regulators and examined the immunopathological characteristics of bile ducts. Findings: After long-term PPI exposure, the fecal microbial and metabolites profile was altered and showed similarity to those observed in high-fat diet studies. Surprisingly, morphological changes were observed in the bile duct, including ductal epithelial proliferation, micropapillary growth of biliary epithelium, focal bile duct stricture formation and bile duct obstruction. These are characteristics of precancerous lesions of bile duct. FXR and RXRα expressions were significantly reduced, which were similar to that observed in cholangiocarcinoma in TCGA and Oncomine databases. Interpretation: We established a novel animal model to examine the effects of long-term use of omeprazole. The gut microbes, metabolic change and biliary epithelial hyperplasia are consequences of long-term PPI exposure. And the results showed the environment in vivo tends to a high-fat diet. Funding Statement: This work was financially supported in part by a grant from Taipei Medical University, Taiwan (TMU105-AE1-B41), a grant from Ministry of Science and Technology, Taiwan (MOST106-2320-B-038-005) and the “TMU Research Center of Cancer Translational Medicine” from The Featured Areas Research Center Program within the framework of the Higher Education Sprout Project by the Ministry of Education (MOE) in Taiwan. Declaration of Interests: The authors declare that they have no competing interests. Ethics Approval Statement: All animals were maintained and handled in an accredited facility in accordance with IACUC (Institutional Animal Care and Use Committee) guidelines and institutional ethics review board approval.

Downregulation of multiple cell cycle-regulatory molecules is a dominant event in TGF-β1-mediated growth inhibition of human carcinoma cells. It is known that KLF10 mimics the anti-proliferative and apoptotic effects that TGF-β1 has on epithelial cell growth and the growth of various tumor cells; based on these findings it is considered as a tumor suppressor. KLF10 protein expression is tightly associated with cell cycle-dependent events. However, the regulatory mechanism and its biological meaning have not been identified. In this study, we have demonstrated that KLF10 is a substrate of CDK2/cyclin E and can be phosphorylated. We also have shown that KLF10 efficiently binds to CDK2, while binding much less to CDK4, and displaying no binding to Cdk6. Using mass spectrometry, site direct mutagenesis, in vitro kinase assays and depletion assays, we have established that CDK2 phosphorylates Ser206, which subsequently affects the steady state level of KLF10 in cells. Our studies have also proved that CDK2 up-regulates the protein level of KLF10 through reducing its association with SIAH1, a KLF10 E3-ubiqutin ligase involved in proteasomal degradation. Taken all together, these findings indicate that CDK2-dependent phosphorylation regulates KLF10 stability and that this affects the role of KLF10 in cell.

Deregulation of transforming growth factor (TGF)-β function is a common feature of pancreatic cancer, rendering these cancers unresponsive to TGF-β–stimulated growth inhibition. Recent findings have supported a primary role for Kruppel-like factor 10 (KLF10) as an important transcription factor involved in mediating TGF-β1 signaling. The aim of this study was to evaluate the correlation between KLF10 expression and the clinical and pathologic features of pancreatic cancer. Tissue specimens from patients with pancreatic adenocarcinoma were retrospectively collected for immunohistochemical analysis. To demonstrate that Klf10 expression was primarily regulated by methylation status, the Klf10 promoter was examined by methylation-specific PCR using a pancreatic cancer cell line (Panc-1). DNA methyltransferase (DNMT) inhibitor and small-interfering RNA depletion of DNMT genes were used to reverse KLF10 expression in the Panc-1 cells. In parallel, DNMT1 expression was evaluated in the pancreatic cancer tissue specimens. In 95 pancreatic cancer tissue specimens, KLF10 expression was inversely correlated with pancreatic cancer stage ( P = 0.01). Multivariable analysis revealed that, in addition to the presence of distant metastasis at diagnosis ( P = 0.001 and 0.001, respectively), KLF10 was another independent prognostic factor related to progression-free and overall survival ( P = 0.018 and 0.037, respectively). The loss of KLF10 expression in advanced pancreatic cancer is correlated with altered methylation status, which seems to be regulated by DNMT1. Our results suggest that KLF10 is a potential clinical predictor for progression of pancreatic cancer.

Aim: Deregulation of transforming growth factor (TGF)-beta function is a common feature of pancreatic cancer, rendering it unresponsive to radiotherapy. Kruppel-like factor 10 (KLF10) is an important transcription factor mediating TGF-beta 1 signaling. The aim of this study is to evaluate the mechanisms of Klf10 in regulating radio-sensitivity of pancreatic cancer. Methods: The expression of Klf10 and radio-sensitivity of genetically modulated and radiation selected pancreatic cancer cells were demonstrated by immunoblots and clonogenic assays. Chip-PCR and luciferase promoter assay were used to demonstrate down-stream targets interaction with Klf10. The level of autophagy, apoptosis and DNA damage after radiation was evaluated. Klf10 modulated tumor response to radiation was measured in orthotopic model. Klf10 expression was correlated with pathologic tumor regression in pancreatic cancer patients. Results: Decreasing Klf10 expression was found in Panc-1 cells with increasing radiation resistance. Over-expressing Klf10 suppressed surviving fraction of MiaPaCa cells after radiation. The observation was associated with increased apoptosis, DNA damage, and decreased autophagy. The expression of ultraviolet irradiation resistance-associated gene (UVRAG) was demonstrated to be transcriptionally suppressed by Klf10. In vitro studies demonstrated Klf10 regulated pancreatic cancer radiation sensitivity via UVRAG. In the orthotopic model, up-regulating Klf10 genetically or pharmacologically sensitized pancreatic tumors to radiation. Analysis from pancreatic cancer patients, receiving neoadjuvant chemo-radiotherapy before surgery, showed a significant correlation of high Klf10 expression with good pathologic tumor regression. Conclusions: Our results suggest Klf10 regulates radio-sensitivity of pancreatic cancer cells by transcriptionally suppressing UVRAG expression which modulates autophagy, apoptosis and DNA damage repair after radiation. Klf10 is a potential biomarker of tumor response to radiotherapy and a therapeutic target to pancreatic cancer. Disclosure: All authors have declared no conflicts of interest.