Downregulated F-Box/LRR-Repeat Protein 7 Facilitates Pancreatic Cancer Metastasis by Regulating Snail1 for Proteasomal Degradation

Pancreatic cancer (PCa) is one of the most aggressive lethal malignancies, and cancer metastasis is the major cause of PCa-associated death. F-box/LRR-repeat protein 7 (FBXL7) regulates cancer metastasis and the chemosensitivity of human pancreatic cancer. However, the clinical significance and biological role of FBXL7 in PCa have been rarely studied. In this study, we found that the expression of FBXL7 was down-regulated in PCa tissues compared with tumor-adjacent tissues, and the low expression of FBXL7 was positively associated with cancer metastasis. Functionally, overexpression of FBXL7 attenuated PANC1 cell invasion, whereas FBXL7 silencing promoted BxPC-3 cell invasion. Forced expression of FBXL7 upregulated the expression of epithelial markers (e.g., E-cadherin) and repressed the expression of mesenchymal markers (e.g., N-cadherin and Vimentin), indicating that FBXL7 negatively regulated the epithelial-mesenchymal transition (EMT) of PCa cells. Furthermore, we identified that FBXL7 repressed the expression of Snail1, a crucial transcription factor of EMT. Mechanistically, FBXL7 bound to Snail1 and promoted its ubiquitination and proteasomal degradation. In vivo studies demonstrated that FBXL7 inhibition promotes PCa metastasis. Taken together, our findings demonstrate that FBXL7 knockdown could efficiently enhance PCa metastasis by regulating Snail1-dependent EMT.

Metabolomic Detection Between Pancreatic Cancer and Liver Metastasis Nude Mouse Models Constructed by Using the PANC1-KAI1/CD82 Cell Line

Background: Pancreatic cancer (PC) has a poor prognosis and is prone to liver metastasis. The KAI1/CD82 gene inhibits PC metastasis. This study aimed to explore differential metabolites and enrich the pathways in serum samples between PC and liver metastasis nude mouse models stably expressing KAI1/CD82. Methods: KAI1/CD82-PLV-EF1α-MCS-IRES-Puro vector and PANC1 cell line stably expressing KAI1/CD82 were constructed for the first time. This cell line was used to construct 3 PC nude mouse models and 3 liver metastasis nude mouse models. The different metabolites and Kyoto encyclopedia of genes and genomes (KEGG) and human metabolome database (HMDB) enrichment pathways were analyzed using the serum samples of the 2 groups of nude mouse models on the basis of untargeted ultra-performance liquid chromatography-tandem mass spectrometry platform. Results: KAI1/CD82-PLV-EF1α-MCS-IRES-Puro vector and PANC1 cell line stably expressing KAI1/CD82 were constructed successfully, and all nude mouse models survived and developed cancers. Among the 1233 metabolites detected, 18 metabolites (9 upregulated and 9 downregulated) showed differences. In agreement with the literature data, the most significant differences between both groups were found in the levels of bile acids (taurocholic acid, chenodeoxycholic acid), glycine, prostaglandin E2, vitamin D, guanosine monophosphate, and inosine. Bile recreation, primary bile acid biosynthesis, and purine metabolism KEGG pathways and a series of HMDB pathways ( P < .05) contained differential metabolites that may be associated with liver metastasis from PC. However, the importance of these metabolites on PC liver metastases remains to be elucidated. Conclusions: Our findings suggested that the metabolomic approach may be a useful method to detect potential biomarkers in PC.

Extracellular Matrix Composition Modulates the Responsiveness of Differentiated and Stem Pancreatic Cancer Cells to Lipophilic Derivate of Gemcitabine

Background: Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal disease. Gemcitabine (GEM) is used as the gold standard drug in PDAC treatment. However, due to its poor efficacy, it remains urgent to identify novel strategies to overcome resistance issues. In this context, an intense stroma reaction and the presence of cancer stem cells (CSCs) have been shown to influence PDAC aggressiveness, metastatic potential, and chemoresistance. Methods: We used three-dimensional (3D) organotypic cultures grown on an extracellular matrix composed of Matrigel or collagen I to test the effect of the new potential therapeutic prodrug 4-(N)-stearoyl-GEM, called C18GEM. We analyzed C18GEM cytotoxic activity, intracellular uptake, apoptosis, necrosis, and autophagy induction in both Panc1 cell line (P) and their derived CSCs. Results: PDAC CSCs show higher sensitivity to C18GEM treatment when cultured in both two-dimensional (2D) and 3D conditions, especially on collagen I, in comparison to GEM. The intracellular uptake mechanisms of C18GEM are mainly due to membrane nucleoside transporters’ expression and fatty acid translocase CD36 in Panc1 P cells and to clathrin-mediated endocytosis and CD36 in Panc1 CSCs. Furthermore, C18GEM induces an increase in cell death compared to GEM in both cell lines grown on 2D and 3D cultures. Finally, C18GEM stimulated protective autophagy in Panc1 P and CSCs cultured on 3D conditions. Conclusion: We propose C18GEM together with autophagy inhibitors as a valid alternative therapeutic approach in PDAC treatment.

Mutant p53, Stabilized by Its Interplay with HSP90, Activates a Positive Feed-Back Loop Between NRF2 and p62 that Induces Chemo-Resistance to Apigenin in Pancreatic Cancer Cells

Pancreatic cancer is one of the most aggressive cancers whose prognosis is worsened by the poor response to the current chemotherapies. In this study, we investigated the cytotoxic effect of Apigenin, against two pancreatic cell lines, namely Panc1 and PaCa44, harboring different p53 mutations. Apigenin is a flavonoid widely distributed in nature that displays anti-inflammatory and anticancer properties against a variety of cancers. Here we observed that Apigenin exerted a stronger cytotoxic effect against Panc1 cell line in comparison to PaCa44. Searching for mechanisms responsible for such different effect, we found that the higher cytotoxicity of Apigenin correlated with induction of higher level of intracellular ROS, reduction of mutant (mut) p53 and HSP90 expression and mTORC1 inhibition. Interestingly, we found that mutp53 was stabilized by its interplay with HSP90 and activates a positive feed-back loop between NRF2 and p62, up-regulating the antioxidant response and reducing the cytotoxicity of Apigenin. These results suggest that targeting the molecules involved in the mTOR-HSP90-mutp53-p62-NRF2-antioxidant response axis could help to overcome the chemo-resistance of pancreatic cancer to Apigenin.

The MicroRNA3686 Inhibits the Proliferation of Pancreas Carcinoma Cell Line by Targeting the Polo-Like Kinase 1

The Polo-like kinase 1 (PLK1) is one member of the so-called Polo-like kinase family which plays an important role in tumorigenesis. By analyzing the potential complementary microRNA (miRNA) targeting sequence of PLK1, we identified that miRNA-3686 (hereby and thereafter mir3696) could be the potential regulator for PLK1. Real-time PCR demonstrated that the mir3686 has a relatively higher expression in the immortalized pancreas cell HPDE6C7 than pancreas carcinoma derived cell line PANC1. The upregulation of mir3686 in HPDE6C7 cell corresponded with the low expression of PLK1 as well. Both luciferase based reporter assay and evaluation of endogenous PLK1 expression demonstrated that mir3686 regulated PLK1, which confirms our speculation. Moreover, we found that transfection of mir3686 in PANC1 cell could lead to proliferation inhibition and promote apoptosis. Further analysis demonstrated that mir3686 transfection in PANC1 cell also inhibited cell invasion, and clone formation in cell invasion assay and clonogenic cell survival assay, respectively. In contrast, inhibition of mir3686 expression in HPDE6C7 cell enhanced the capability of proliferation, cell invasion and clone formation. Taken together, our results indicated that mir3686 could target PLK1 to inhibit the cell proliferation in pancreas cancer derived cell line and mir3686 could be a new therapeutic target for pancreas cancer treatment.