Recurrent Mutations of Chromatin-Remodeling Genes and Kinase Receptors in Pheochromocytomas and Paragangliomas

Abstract

Purpose Pheochromocytomas and paragangliomas (PPGLs) are genetically heterogeneous tumors of neural crest origin, but the molecular basis of most PPGLs is unknown. Experimental Design We performed exome or transcriptome sequencing of 43 samples from 41 patients. A validation set of 136 PPGLs was used for amplicon-specific resequencing. In addition, a subset of these tumors was used for microarray-based transcription, protein expression and histone methylation analysis by western blot or immunohistochemistry. In vitro analysis of mutants was performed in cell lines. Results We detected mutations in chromatin remodeling genes, including histone-methyltransferases, histone-demethylases and histones in 11 samples from 8 patients (20%). In particular, we characterized a new cancer syndrome involving PPGLs and giant cell tumors of bone (GCT) caused by a postzygotic G34W mutation of the histone 3.3 gene, H3F3A. Furthermore, mutations in kinase genes were detected in samples from 15 patients (37%). Among those, a novel germline kinase domain mutation of MERTK detected in a patient with PPGL and medullary thyroid carcinoma was found to activate signaling downstream of this receptor. Recurrent germline and somatic mutations were also detected in MET, including a familial case and sporadic PPGLs. Importantly, in each of these three genes mutations were also detected in the validation group. Additionally, a somatic oncogenic hotspot FGFR1 mutation was found in a sporadic tumor. Conclusions This study implicates chromatin-remodeling and kinase variants as frequent genetic events in PPGLs, many of which have no other known germline driver mutation. MERTK, MET, and H3F3A emerge as novel PPGL susceptibility genes.

Consensus Statement on next-generation-sequencing-based diagnostic testing of hereditary phaeochromocytomas and paragangliomas

“…Pituitary adenomas were reported in patients with PPGLs in familial settings in which an SDH mutation was detected 84,85 . Germ line FH mutations have been described in patients with phaeochromocytomas and uterine leiomyomas 14,20,24 , and, in the past year histone gene mutations (H3F3A) were found in a new syndrome of multiple paragangliomas and giant cell tumour of bone, which implicates chromatin remodelling defects in PPGL tumorigenesis and susceptibility 14 .…”

“…All participants have adopted, and reported on, NGS-based technologies in their research and/or clinical practice [14][15][16][17][18][19][20][21][22][23][24][25][26] . Discussions took place via conference calls, e-mail communications and file exchanges and one plenary session (at the 14th ENS@T Scientific Meeting on November 20th, 2015, Munich, Germany).…”

“…Both NF1 and VHL, which are established PPGL susceptibility genes, have been detected as mosaic, post-zygotic mutations in neurofibromatosis type 1 or von Hippel-Lindau syndrome; however, this finding has not been described in the specific setting of PPGLs 79,80 . In 2015, mosaic mutations leading to a syndrome involving PPGL and giant cell tumours of bone were reported in association with the H3F3A gene 14 . Therefore, mosaic transmission might occur more frequently in PPGLs than hitherto appreciated.…”

“…A possible limitation of this technique is the difficulty in identifying mutations that lead to decreased or absent transcription or very unstable mRNA of the target gene. RNA-seq has been performed in only a small number of PPGLs 14,26 . Although not tested for all known driver genes for PPGLs, at least in one report, truncating germ line mutations in SDHB and SDHD and a missense germ line mutation in VHL were promptly detected with evidence of LOH, and later validated by Sanger sequencing 14 , which supports the efficacy of RNA-based sequencing for screening of susceptibility mutations.…”

“…RNA-seq has been performed in only a small number of PPGLs 14,26 . Although not tested for all known driver genes for PPGLs, at least in one report, truncating germ line mutations in SDHB and SDHD and a missense germ line mutation in VHL were promptly detected with evidence of LOH, and later validated by Sanger sequencing 14 , which supports the efficacy of RNA-based sequencing for screening of susceptibility mutations. RNA-seq might also enable detection of fusion transcripts that cannot be identified by WES 26 .…”