Mechanism of BRG1 silencing in primary cancers

Abstract

BRG1 (SMARCA4) is a documented tumor suppressor and a key subunit of the SWI/SNF chromatin remodeling complex that is silenced in many cancer types. Studies have shown that BRG1 is mutated in cancer-derived cell lines, which led to the assertion that BRG1 is also mutated in primary human tumors. However, the sequencing of BRG1-deficient tumors has revealed a paucity of mutations; hence, the cause of BRG1 silencing in tumors remains an enigma. We conducted immunohistochemistry (IHC) on a number of tumor microarrays to characterize the frequency of BRG1 loss in different tumor types. We also analyzed BRG1-deficient tumors by sequencing the genomic DNA and the mRNA. We then tested if BRG1 expression could be induced in BRG1-negative cell lines (i.e., that lack mutations in BRG1) after the application of several different epigenetic agents, including drugs that inhibit the AKT pathway. We found that a subset of BRG1-negative cell lines also demonstrated aberrant splicing of BRG1, and in at least 30% of BRG1-deficient tumors, BRG1 expression appeared to be suppressed due to aberrant BRG1 splicing. As the majority of BRG1-deficient tumors lack mutations or splicing defects that could drive BRG1 loss of expression, this suggests that other mechanisms underlie BRG1 silencing. To this end, we analyzed 3 BRG1-deficient nonmutated cancer cell lines and found that BRG1 was inducible in these cell lines upon inhibition of the AKT pathway. We show that the loss of BRG1 is associated with the loss of E-cadherin and up-regulation of Vimentin in primary tumors, which explains why BRG1 loss is associated with a poor prognosis in multiple tumor types.