Targeting Stiff Tumor Microenvironment for Gastric Cancer

In many cancers, tumor progression is associated with increased tissue stiffness. Yet the mechanisms associating tissue stiffness with tumorigenesis and malignant transformation are unclear. We show that the stiffness of the extracellular matrix of gastric cancer reversibly regulates the DNA methylation of the promoter region of the oncogenic Yes-associated protein (YAP). This work offers insights into promising mechanotherapeutic strategies specifically targeting the mechanical properties of the ECM, to regulate epigenetic status and oncogenic transcription activity of malignant tumor cells.

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Gastric cancer (GC) is one of the leading causes of death in East Asia, and GC incidence is rising among younger generations of adults. Extracellular matrix (ECM) density is increased in GC, and it is widely known that a stiff cancer ECM helps cancer cells proliferate and invade nearby tissues; consequently, stiff tumor microenvironments correlate with a worse prognosis and decreased life expectancy.

Professor Pilnam Kim’s research lab in the Department of Bio and Brain Engineering at the Korea Advanced Institute of Science and Technology (KAIST) reported a study on matrix stiffness as a crucial determinant of Yes-associated-protein (YAP) transcriptional regulation and DNA methylation in gastric cancer (Nature Biomedical Engineering doi:10.1038/s41551-017-0158-x.).

Researchers first examined YAP activity in response to matrix stiffness. YAP, a well-known mechanotransducer, translocates from the cytoplasm to the nucleus when the cell is under mechanical stress. Clinically, patients with higher expression of nuclear YAP show poor survival rates, and some GC tissues confirm promoter hypomethylation and upregulation of YAP.

In order to confirm matrix stiffness-induced mechanotransduction, the research group constructed collagen and interpenetrating polymer network (IPN)-based 3D gel matrices, embedded with gastric cancer cells, to recapitulate the tumor microenvironment. The stiff matrix (IPN) successfully induced YAP activation and reduced circularity of the tumor aggregates (Figure 1a).

Next, the researchers softened the matrix and used mechanotransduction inhibitors to determine whether disrupting the mechanotransduction signaling cascade would reverse their previous observations. They found that YAP expression increasingly deactivated with time after matrix softening (Figure 1b). Total YAP expression was also reduced, indicating that lower transcriptional activity may epigenetically reduce total YAP gene expression.

Epigenetics can be simply described as changes in gene expressions without DNA sequence alterations. Methyl groups can attach to DNA and cause gene inactivity or silenced transcription; conversely, demethylation may activate silenced genes.

Researchers in Professor Kim’s lab found that DNA hypomethylation of YAP induced by a stiff matrix can be reversed by softening the matrix. The methylation index of YAP DNA recovered with time after matrix softening (Figure 1c). Upon a proliferation and drug resistance analysis, GC cells in the softened matrix were less proliferative and less resistant to drugs than those in the stiff matrix, confirming that the softening was effective in suppressing malignancy. Reversion by matrix softening recovered the methylation index the closest to that of the soft control, showing that external biophysical conditions are dominant in influencing cell behavior (Figures 1d, e). Transcriptome analysis by bulk RNA sequencing and Chromatin Immunoprecipitation-Atlas analysis identified DNA methylation-modifying genes that were recovered by matrix softening by comparing control and YAP-depleted cells. siRNA treatments of three genes (GRHL2, TET2, and KMT2A) confirmed the recovery of YAP DNA methylation comparable to the soft control (Figure 2). These findings suggest that mechanotherapy to soften tumor tissues and surrounding ECM may be an appropriate addition along with chemotherapy for better patient prognosis.