title: SMARCB1 protein and mRNA loss is not caused by promoter and histone hypermethylation in epithelioid sarcoma.
abstract: About 10% of epithelioid sarcomas have biallelic mutation of the SMARCB1 (SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily b, member 1) gene resulting in a lack of this nuclear protein. It has been suggested that SMARCB1 may be silenced by epigenetic changes in the remaining 90% of tumors. Thus, we hypothesized that the promoter of SMARCB1 is hypermethylated. We also examined SMARCB1 mRNA level to determine if a post-translational change was possible. Thirty-six cases of epithelioid sarcomas were studied. Immunohistochemistry and mutation analysis of the SMARCB1 gene were performed to select appropriate cases. Methylation status was assessed by methylation-specific PCR. Laser capture microdissection of tumor cells followed by real-time PCR was applied to examine the expression of SMARCB1 mRNA. Of 36 epithelioid sarcomas, 31 (86%) displayed a lack of SMARCB1 nuclear protein. In all, 4 (13%) of 31 SMARCB1-negative cases harbored biallelic deletion while 9 (33%) cases showed single-allelic deletion. One (4%) frameshift deletion of exon 3 and one point mutation of exon 7 were also found. In 16 (59%) cases, both alleles were intact. Altogether, 25/31 (81%) SMARCB1-negative cases had at least one intact allele. None of these cases demonstrated promoter hypermethylation. Low levels of SMARCB1 mRNA were found in all cases with tumor tissue extracted RNA (because of the minimal normal cell contamination) but no mRNA could be detected in laser dissected cases (containing only tumor cells). Enhancer of zeste homolog 2 (EZH2) overexpression was not characteristic of epithelioid sarcoma. Thus, loss of SMARCB1 expression in epithelioid sarcoma is caused neither by DNA hypermethylation nor by post-translational modifications. Most likely it is the microRNA destruction of SMARCB1 mRNA but further investigations are needed to elucidate this issue.
authors.affiliation: First Department of Pathology and Experimental Cancer Research, Semmelweis University, Budapest, Hungary.
authors.forename: Gergő
authors.initials: G
authors.lastname: Papp
authors.forename: Yi-Che
authors.initials: YC
authors.lastname: Changchien
authors.forename: Bálint
authors.initials: B
authors.lastname: Péterfia
authors.forename: Loránd
authors.initials: L
authors.lastname: Pecsenka
authors.forename: Thomas
authors.initials: T
authors.lastname: Krausz
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Stricker
authors.forename: Andras
authors.initials: A
authors.lastname: Khoor
authors.forename: Ludvik
authors.initials: L
authors.lastname: Donner
authors.forename: Zoltán
authors.initials: Z
authors.lastname: Sápi
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D002648:Child; D002868:Chromosomal Proteins, Non-Histone / Q000032:analysis / Q000235:genetics*; D019175:DNA Methylation; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000032:analysis / Q000235:genetics*; D015536:Down-Regulation; D000071221:Enhancer of Zeste Homolog 2 Protein; D005260:Female; D017353:Gene Deletion; D015972:Gene Expression Regulation, Neoplastic; D020022:Genetic Predisposition to Disease; D006657:Histones / Q000032:analysis*; D006801:Humans; D007150:Immunohistochemistry; D017404:In Situ Hybridization, Fluorescence; D061346:Laser Capture Microdissection; D008297:Male; D008875:Middle Aged; D009154:Mutation; D010641:Phenotype; D011044:Poland; D063151:Polycomb Repressive Complex 2 / Q000032:analysis; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000032:analysis*; D060888:Real-Time Polymerase Chain Reaction; D012189:Retrospective Studies; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000071796:SMARCB1 Protein; D012509:Sarcoma / Q000737:chemistry / Q000235:genetics* / Q000473:pathology; D012983:Soft Tissue Neoplasms / Q000737:chemistry / Q000235:genetics* / Q000473:pathology; D014157:Transcription Factors / Q000032:analysis / Q000235:genetics*; D014481:United States; D055815:Young Adult
chemical_list: D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; D006657:Histones; D012333:RNA, Messenger; D000071796:SMARCB1 Protein; C513266:SMARCB1 protein, human; D014157:Transcription Factors; C498409:EZH2 protein, human; D000071221:Enhancer of Zeste Homolog 2 Protein; D063151:Polycomb Repressive Complex 2