Probes for INS

Gastric cancer is the fifth most common cancer and third leading cause of cancer-related death worldwide. Several studies on angiogenic blocking agents in gastric cancer revealing promising results by the use of monoclonal antibodies against VEGFA or its receptor VEGFR2 or against VEGFA activating pathway. The validation of biomarkers useful to better organize the clinical trials involving anti-angiogenic therapies is crucial. Molecular markers such as RNA are increasingly used for cancer diagnosis, prognosis, and therapy guidance as in the case of the targeted therapies concerning the inhibition of angiogenesis. The aim of this study is to set the conditions for evaluating the expression of VEGFA and VEGFR2 in gastric cancer specimens and in healthy gastric mucosa by the use of RNAscope, a novel RNA in situ hybridization (ISH) method that allows the visualization of a specific gene expression in individual cells. We found the increased expression of VEGFA in the tubular glands and VEGFR2 in the endothelium of gastric cancer samples mainly in the T2, T3 and T4 stages of tumor progression as compared to the healthy controls. These results obtained by the application of this highly sensitive method for oligonucleotide detection the role of angiogenesis in gastric cancer progression already highlighted by conventional immunohistochemical methods, and offer significant promise as a new platform for developing and implementing RNA-based molecular diagnostics also in the conditions in which immunohistochemistry is not applicable.

Abstract

OBJECTIVE:

Cancer-associated fibroblasts (CAFs), a major component of cancer stroma, can confer aggressive properties to cancer cells by secreting multiple factors. Their phenotypes are stably maintained, but the mechanisms are not fully understood. We aimed to show the critical role of epigenetic changes in CAFs in maintaining their tumour-promoting capacity and to show the validity of the epigenomic approach in identifying therapeutic targets from CAFs to starve cancer cells.

DESIGN:

Twelve pairs of primary gastric CAFs and their corresponding non-CAFs (NCAFs) were established from surgical specimens. Genome-wide DNA methylation and H3K27me3 analyses were conducted by BeadArray 450K and ChIP-on-Chip, respectively. Functions of potential a therapeutic target were analysed by inhibiting it, and prognostic impact was assessed in a database.

RESULTS:

CAFs had diverse and distinct DNA methylation and H3K27me3 patterns compared with NCAFs. Loss of H3K27me3, but not DNA methylation, in CAFs was enriched for genes involved in stem cell niche, cell growth, tissue development and stromal-epithelial interactions, such as WNT5A, GREM1, NOG and IGF2. Among these, we revealed that WNT5A, which had been considered to be derived from cancer cells, was highly expressed in cancer stromal fibroblasts, and was associated with poor prognosis. Inhibition of secreted WNT5A from CAFs suppressed cancer cell growth and migration.

CONCLUSIONS:

H3K27me3 plays a crucial role in defining tumour-promoting capacities of CAFs, and multiple stem cell niche factors were secreted from CAFs due to loss of H3K27me3. The validity of the epigenetic approach to uncover therapeutic targets for cancer-starving therapy was demonstrated.

The epithelial lining of the Fallopian tube is vital for fertility, providing nutrition to gametes, and facilitating their transport. It is composed of two major cell types: secretory cells and ciliated cells. Interestingly, human ovarian cancer precursor lesions are primarily consisting of secretory cells. It is unclear why secretory cells are the dominant cell type in these lesions. Additionally, the underlying mechanisms governing Fallopian tube epithelial homoeostasis are currently unknown. In the present study, we showed that across the different developmental stages of mouse oviduct, secretory cells are the most frequently dividing cells of the oviductal epithelium. In vivo genetic cell lineage tracing showed that secretory cells not only self-renew, but also give rise to ciliated cells. Analysis of a Wnt reporter mouse model and different Wnt target genes showed that the Wnt signaling pathway is involved in oviductal epithelial homoeostasis. By developing two triple transgenic mouse models, we showed that Wnt/β-catenin signaling is essential for self-renewal as well as differentiation of secretory cells. In summary, our results provide mechanistic insight into oviductal epithelial homoeostasis.