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Background: GI stromal tumours (GISTs) are clinically heterogenous exhibiting varying degrees of disease aggressiveness in individual patients., Objectives: We sought to identify genetic alterations associated with high-risk GIST, explore their molecular consequences, and test their utility as prognostic markers., Designs: Exome sequencing of 18 GISTs was performed (9 patients with high-risk/metastatic and 5 patients with low/intermediate-risk), corresponding to 11 primary and 7 metastatic tumours. Candidate alterations were validated by prevalence screening in an independent patient cohort (n=120). Functional consequences of SETD2 mutations were investigated in primary tissues and cell lines. Transcriptomic profiles for 8 GISTs (4 SETD2 mutated, 4 SETD2 wild type) and DNA methylation profiles for 22 GISTs (10 SETD2 mutated, 12 SETD2 wild type) were analysed. Statistical associations between molecular, clinicopathological factors, and relapse-free survival were determined., Results: High-risk GISTs harboured increased numbers of somatic mutations compared with low-risk GISTs (25.2 mutations/high-risk cases vs 6.8 mutations/low-risk cases; two sample t test p=3.1×10 -5 ). Somatic alterations in the SETD2 histone modifier gene occurred in 3 out of 9 high-risk/metastatic cases but no low/intermediate-risk cases. Prevalence screening identified additional SETD2 mutations in 7 out of 80 high-risk/metastatic cases but no low/intermediate-risk cases (n=29). Combined, the frequency of SETD2 mutations was 11.2% (10/89) and 0% (0/34) in high-risk and low-risk GISTs respectively. SETD2 mutant GISTs exhibited decreased H3K36me3 expression while SETD2 silencing promoted DNA damage in GIST-T1 cells. In gastric GISTs, SETD2 mutations were associated with overexpression of HOXC cluster genes and a DNA methylation signature of hypomethylated heterochromatin. Gastric GISTs with SETD2 mutations, or GISTs with hypomethylated heterochromatin, showed significantly shorter relapse-free survival on univariate analysis (log rank p=4.1×10 -5 )., Conclusions: Our data suggest that SETD2 is a novel GIST tumour suppressor gene associated with disease progression. Assessing SETD2 genetic status and SETD2-associated epigenomic phenotypes may guide risk stratification and provide insights into mechanisms of GIST clinical aggressiveness., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.)

Objective: Gastric cancer (GC) is a deadly malignancy for which new therapeutic strategies are needed. Three transcription factors, KLF5, GATA4 and GATA6, have been previously reported to exhibit genomic amplification in GC. We sought to validate these findings, investigate how these factors function to promote GC, and identify potential treatment strategies for GCs harbouring these amplifications., Design: KLF5, GATA4 and GATA6 copy number and gene expression was examined in multiple GC cohorts. Chromatin immunoprecipitation with DNA sequencing was used to identify KLF5/GATA4/GATA6 genomic binding sites in GC cell lines, and integrated with transcriptomics to highlight direct target genes. Phenotypical assays were conducted to assess the function of these factors in GC cell lines and xenografts in nude mice., Results: KLF5, GATA4 and GATA6 amplifications were confirmed in independent GC cohorts. Although factor amplifications occurred in distinct sets of GCs, they exhibited significant mRNA coexpression in primary GCs, consistent with KLF5/GATA4/GATA6 cross-regulation. Chromatin immunoprecipitation with DNA sequencing revealed a large number of genomic sites co-occupied by KLF5 and GATA4/GATA6, primarily located at gene promoters and exhibiting higher binding strengths. KLF5 physically interacted with GATA factors, supporting KLF5/GATA4/GATA6 cooperative regulation on co-occupied genes. Depletion and overexpression of these factors, singly or in combination, reduced and promoted cancer proliferation, respectively, in vitro and in vivo. Among the KLF5/GATA4/GATA6 direct target genes relevant for cancer development, one target gene, HNF4α, was also required for GC proliferation and could be targeted by the antidiabetic drug metformin, revealing a therapeutic opportunity for KLF5/GATA4/GATA6 amplified GCs., Conclusions: KLF5/GATA4/GATA6 may promote GC development by engaging in mutual crosstalk, collaborating to maintain a pro-oncogenic transcriptional regulatory network in GC cells., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.)