Genomic characterization of a well-differentiated grade 3 pancreatic neuroendocrine tumor

Abstract

Pancreatic neuroendocrine neoplasms (PanNENs) represent a minority of pancreatic neoplasms that exhibit variability in prognosis. Ongoing mutational analyses of PanNENs have found recurrent abnormalities in chromatin remodeling genes (e.g., DAXX and ATRX), and mTOR pathway genes (e.g., TSC2, PTEN PIK3CA, and MEN1), some of which have relevance to patients with related familial syndromes. Most recently, grade 3 PanNENs have been divided into two groups based on differentiation, creating a new group of well-differentiated grade 3 neuroendocrine tumors (PanNETs) that have had a limited whole-genome level characterization to date. In a patient with a metastatic well-differentiated grade 3 PanNET, our study utilized whole-genome sequencing of liver metastases for the comparative analysis and detection of single-nucleotide variants, insertions and deletions, structural variants, and copy-number variants, with their biologic relevance confirmed by RNA sequencing. We found that this tumor most notably exhibited a TSC1-disrupting fusion, showed a novel CHD7-BEND2 fusion, and lacked any somatic variants in ATRX, DAXX, and MEN1.

Trial registration: ClinicalTrials.gov NCT02155621.

Keywords: neoplasm of the pancreas; neuroendocrine neoplasm.

© 2019 Williamson et al.; Published by Cold Spring Harbor Laboratory Press.

Figures

Figure 1.

Tumor histopathology and Ki-67 immunohistochemistry. (A) Core biopsy of a liver metastasis stained for hematoxylin and eosin showing a metastatic well-differentiated neuroendocrine tumor clinicoradiologically consistent with pancreatic origin (magnification at 10× [left] and 40× [right]). (B) Paired Ki-67 immunohistochemistry showing a proliferative index of 30%, classifying the tumor as a well-differentiated grade 3 (G3) neuroendocrine tumor (magnification at 10× [left] and 40× [right]).

Figure 2.

Copy-number aberration and TSC1–TMEM71 structural rearrangement in the G3 PanNET. (A) Circos plot illustrating copy-number alterations and loss of heterozygosity (LOH) observed in the tumor sample. (B) Genomic sequencing read alignment of the genomic regions affected by the rearrangement. Green reads that aligned to TSC1 and purple reads that aligned to TMEM71 indicate split reads that support the rearrangement. (C) Illustration of TSC1–TMEM71 rearrangement generated by MAVIS (Reisle et al. 2018). The TMEM71 and TSC1 breakpoints are indicated at the chromosome, gene, and transcript level (B1 and B2, respectively, upper box). Protein-coding sequence associated with the respective transcripts are indicated by the black line and the amino acids included in the fusion product are indicated. Pfam (http://pfam.xfam.org) protein domains are indicated in the track below the transcript: Hamartin (PF04388). The predicted fusion transcript and protein-coding sequence are shown in the lower box (TSC1 exons are colored green; TMEM71 exons are colored blue).

Figure 3.

CHD7–BEND2 structural rearrangement and increased BEND2 expression. (A) Genomic sequencing read alignment of the genomic regions affected by the rearrangement. Gold reads that aligned to CHD7 and purple reads that aligned to BEND2 indicate reads that support the rearrangement. (B) Illustration of CHD7–BEND2 rearrangement. The CHD7 and BEND2 breakpoints are indicated at the chromosome, gene, and transcript level (B1 and B2, respectively, upper box). The protein-coding sequences associated with the respective transcripts are indicated by the black line, and the amino acids included in the fusion product are indicated. The predicted fusion transcript and protein-coding sequence are shown in the lower box (CHD7 exons are colored blue; BEND2 exons are colored green). Pfam protein domains are indicated by the tracks below the transcript: SNF2_N (F00176), helicase_C (PF00271), Chromo (PF00385), BRK (PF07533), BEN (PF10523). (C) Genomic sequencing reads (top) and RNA sequencing reads (bottom) aligned to the BEND2 gene. Genomic reads that are affected by the rearrangement are colored purple. The BEND2 fusion breakpoint is indicated by the dashed line. RNA-seq coverage downstream from the breakpoint (exons 5–14) is increased compared to upstream of the breakpoint (exons 1–4). (D) Boxplot of BEND2 RPKM (log10 transformed) across a panel of 40 TCGA data sets. BEND2 log10(RPKM) for this patient sample is indicated by the red horizontal line.

Figure 4.

Low expression of SSTR genes in the G3 PanNET. SSTR1, SSTR2, SSTR3, and SSTR5 RPKM normalized to the mean of the cohort were plotted for seven metastatic PanNET samples. Relative SSTR gene expression for this patient sample is highlighted in red.