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Jackson Lab, GIS Team IDs Recurrent Gene Fusions in Gastric Cancer

NEW YORK (GenomeWeb) – Researchers from the Jackson Laboratory, the Genome Institute of Singapore, and elsewhere have sequenced the genomes of 15 Southeast Asian patients with gastric cancer, identifying recurrent gene fusions.

The researchers used a sequencing method known as paired-end tag sequencing, or DNA-PET, a technique for detecting structural rearrangements that was developed by the Jackson Lab's Yijun Ruan when he was at GIS. They published their results this month in Cell Reports.

Nearly 1 million new gastric cancer cases were diagnosed in 2008, and nearly 70 percent of new cases and deaths came from developing countries, with the highest incidence in Eastern Asia. Although a few recurrently mutated genes have been identified, very little is known about structural rearrangements, highlighting both the need to study the cancer genome in Asian populations as well as the need to develop "new sophisticated genomic analysis approaches which will ultimately provide critical insights into cancer progression," senior author Ruan said in a statement.

The key to the DNA-PET method, developed by researchers at GIS and Thermo Fisher's Life Technologies in 2009, is the creation of a 10-kb insert fragment sequencing library. The long pieces of DNA in combination with paired-end tags enable researchers to better identify large rearrangements and map across repetitive regions.

The team sequenced 14 primary gastric tumors along with 10 matched normal samples and a gastric cancer cell line. They identified 1,945 somatic structural variations. From the data, the team predicted 136 fusion genes, 97 of which they validated using PCR and Sanger sequencing. Of those, they identified 44 that were expressed and 15 that were expressed and in frame. Because "active oncogenic fusion genes are usually in-frame fusions," the authors focused on that category to screen an additional 85 tumor/normal pairs.

Five fusions were recurrent at overall frequencies of 2 percent to 5 percent. Notably, they identified the gene fusion CLDN18-ARHGAP26. CLDN18 is an "essential tight junction component" in the stomach. The gene ARHGAP26, is known to regulate RHOA, a gene that has recently been found to be mutated in gastric cancer. The researchers found that the fusion leads to a loss of function of CLDN18 and a gain of ARHGAP26.

Next, they looked at the function of the fusion in cell lines, finding that it results in in impaired epithelial barrier properties and wound healing. The authors concluded that the fusion gene "mediates epithelial disintegration, possibly leading to stomach H+ leakage." In addition, the fusion could "lead to invasiveness once a cell is transformed."

"CLDN18 is a critical component of the gastric epithelial barrier," Walter Hunziker, deputy director of the Institute of Molecular and Cell Biology at GIS, said in a statement. "Fusion of ARHGAP26 to CLDN18 not only interferes with the tethering of CLDN18 to the actin cytoskeleton, but could also affect the actin cytoskeleton by inhibiting RHOA at the wrong location, thereby compromising barrier integrity. The resulting inflammation and gastritis are well known risk factors for gastric cancer."

The researchers next plan to continue to study the functional impact of the other four recurrent fusions. Network analysis predicted that CLDN18-ARHGAP26 along with the SNYX2-PRDM6 and MLL3-PRKAG2 fusions were drivers. In addition, an initial functional study in cell lines showed evidence that MLL3-PRKAG2 and CLEC16A-EMP2 supported proliferation.