Everolimus exerts anticancer effects through inhibiting the interaction of matrix metalloproteinase-7 with syndecan-2 in colon cancer cells.

Previous work showed that matrix metalloproteinase-7 (MMP-7) regulates colon cancer activities through an interaction with syndecan-2 (SDC-2) and SDC-2-derived peptide that disrupts this interaction and exhibits anticancer activity in colon cancer. Here, to identify potential anticancer agents, a library of 1,379 Food and Drug Administration (FDA)-approved drugs that interact with the MMP-7 prodomain were virtually screened by protein-ligand docking score analysis using the GalaxyDock3 program. Among five candidates selected based on their structures and total energy values for interacting with the MMP-7 prodomain, the known mechanistic target of rapamycin kinase (mTOR) inhibitor, everolimus, showed the highest binding affinity and the strongest ability to disrupt the interaction of the MMP-7 prodomain with the SDC-2 extracellular domain in vitro. Everolimus treatment of the HCT116 human colon cancer cell line did not affect the mRNA expression levels of MMP-7 and SDC-2 but reduced the adhesion of cells to MMP-7 prodomain-coated plates and the cell-surface localization of MMP-7. Thus, everolimus appears to inhibit the interaction between MMP-7 and SDC-2. Everolimus treatment of HCT116 cells also reduced their gelatin-degradation activity and anticancer activities, including colony formation. Interestingly, cells treated with sirolimus, another mTOR inhibitor, triggered less gelatin-degradation activity, suggesting that this inhibitory effect of everolimus was not due to inhibition of the mTOR pathway. Consistently, everolimus inhibited the colony-forming ability of mTOR-resistant HT29 cells. Together, these data suggest that, in addition to inhibiting mTOR signaling, everolimus exerts anticancer activity by interfering with the interaction of MMP-7 and SDC-2, and could be a useful therapeutic anticancer drug for colon cancer.NEW & NOTEWORTHY The utility of cancer therapeutics targeting the proteolytic activities of MMPs is limited because MMPs are widely distributed throughout the body and involved in many different aspects of cell functions. This work specifically targets the activation of MMP-7 through its interaction with syndecan-2. Notably, everolimus, a known mTOR inhibitor, blocked this interaction, demonstrating a novel role for everolimus in inhibiting mTOR signaling and impairing the interaction of MMP-7 with syndecan-2 in colon cancer.

Symmetry Breaking of Human Pluripotent Stem Cells (hPSCs) in Micropattern Generates a Polarized Spinal Cord-Like Organoid (pSCO) with Dorsoventral Organization.

Axis formation and related spatial patterning are initiated by symmetry breaking during development. A geometrically confined culture of human pluripotent stem cells (hPSCs) mimics symmetry breaking and cell patterning. Using this, polarized spinal cord organoids (pSCOs) with a self-organized dorsoventral (DV) organization are generated. The application of caudalization signals promoted regionalized cell differentiation along the radial axis and protrusion morphogenesis in confined hPSC colonies. These detached colonies grew into extended spinal cord-like organoids, which established self-ordered DV patterning along the long axis through the spontaneous expression of polarized DV patterning morphogens. The proportions of dorsal/ventral domains in the pSCOs can be controlled by the changes in the initial size of micropatterns, which altered the ratio of center-edge cells in 2D. In mature pSCOs, highly synchronized neural activity is separately detected in the dorsal and ventral side, indicating functional as well as structural patterning established in the organoids. This study provides a simple and precisely controllable method to generate spatially ordered organoids for the understanding of the biological principles of cell patterning and axis formation during neural development.

Production of human spinal-cord organoids recapitulating neural-tube morphogenesis.

Human spinal-cord-like tissues induced from human pluripotent stem cells are typically insufficiently mature and do not mimic the morphological features of neurulation. Here, we report a three-dimensional culture system and protocol for the production of human spinal-cord-like organoids (hSCOs) recapitulating the neurulation-like tube-forming morphogenesis of the early spinal cord. The hSCOs exhibited neurulation-like tube-forming morphogenesis, cellular differentiation into the major types of spinal-cord neurons as well as glial cells, and mature synaptic functional activities, among other features of the development of the spinal cord. We used the hSCOs to screen for antiepileptic drugs that can cause neural-tube defects. hSCOs may also facilitate the study of the development of the human spinal cord and the modelling of diseases associated with neural-tube defects.

Cisplatin Resistance in Epstein-Barr-Virus-Associated Gastric Carcinoma Acquired through ATM Methylation.

Epstein-Barr-virus-associated gastric carcinoma (EBVaGC), first reported in 1992, currently accounts for 10% of all gastric carcinoma worldwide. EBVaGC has unique DNA hypermethylation phenotypes that allow for higher proportions of DNA methylation than any other gastric cancer. CpG islands in the gene promoter region are one of the major regions in which DNA methylation controls gene transcription. Despite cisplatin-based chemotherapy being one of the standard treatment regimens for advanced gastric cancer, including EBVaGC, cisplatin alone or in combination with 5-fluorouracil has been limited by its less potent anticancer activity and the occurrence of cisplatin resistance. Accordingly, the current study evaluated the anticancer activities of a combination of cisplatin and 5-Azacytidine (5-AZA) against EBVaGC. Our findings showed that cisplatin upregulated the DNMT3A gene, whereas shRNA-targeted removal of DNMT3A mRNA contributed to cisplatin-mediated EBV lytic reactivation. Moreover, the removal of DNMT3A mRNA upregulated the ATM gene through DNA demethylation on the ATM promoter. Furthermore, CRISPR/Cas9-targeted removal of the ATM gene resulted in significantly reduced cell susceptibility and EBV lytic reactivation by a combination of cisplatin and DNMT3A inhibitor 5-AZA. Finally, 5-AZA exhibited a synergistic effect with cisplatin in anti-EBV and anti-EBVaGC activities by increasing drug susceptibility and EBV lytic reactivation. The aforementioned results suggest that cisplatin combined with DNA methylation inhibitors could be a novel therapeutic approach for EBVaGC.

ChimerDB 4.0: an updated and expanded database of fusion genes.

Fusion genes represent an important class of biomarkers and therapeutic targets in cancer. ChimerDB is a comprehensive database of fusion genes encompassing analysis of deep sequencing data (ChimerSeq) and text mining of publications (ChimerPub) with extensive manual annotations (ChimerKB). In this update, we present all three modules substantially enhanced by incorporating the recent flood of deep sequencing data and related publications. ChimerSeq now covers all 10 565 patients in the TCGA project, with compilation of computational results from two reliable programs of STAR-Fusion and FusionScan with several public resources. In sum, ChimerSeq includes 65 945 fusion candidates, 21 106 of which were predicted by multiple programs (ChimerSeq-Plus). ChimerPub has been upgraded by applying a deep learning method for text mining followed by extensive manual curation, which yielded 1257 fusion genes including 777 cases with experimental supports (ChimerPub-Plus). ChimerKB includes 1597 fusion genes with publication support, experimental evidences and breakpoint information. Importantly, we implemented several new features to aid estimation of functional significance, including the fusion structure viewer with domain information, gene expression plot of fusion positive versus negative patients and a STRING network viewer. The user interface also was greatly enhanced by applying responsive web design. ChimerDB 4.0 is available at http://www.kobic.re.kr/chimerdb/.