The molecular characteristics of high-grade gastroenteropancreatic neuroendocrine neoplasms

Abstract

High-grade (HG) gastroenteropancreatic (GEP) neuroendocrine neoplasms (NEN) are rare but have a very poor prognosis and represent a severely understudied class of tumours. Molecular data for HG GEP-NEN are limited, and treatment strategies for the carcinoma subgroup (HG GEP-NEC) are extrapolated from small-cell lung cancer (SCLC). After pathological re-evaluation, we analysed DNA from tumours and matched blood samples from 181 HG GEP-NEN patients; 152 neuroendocrine carcinomas (NEC) and 29 neuroendocrine tumours (NET G3). Based on the sequencing of 360 cancer-related genes, we assessed mutations and copy number alterations (CNA). For NEC, frequently mutated genes were TP53 (64%), APC (28%), KRAS (22%) and BRAF (20%). RB1 was only mutated in 14%, but CNAs affecting RB1 were seen in 34%. Other frequent copy number losses were ARID1A (35%), ESR1 (25%) and ATM (31%). Frequent amplifications/gains were found in MYC (51%) and KDM5A (45%). While these molecular features had limited similarities with SCLC, we found potentially targetable alterations in 66% of the NEC samples. Mutations and CNA varied according to primary tumour site with BRAF mutations mainly seen in colon (49%), and FBXW7 mutations mainly seen in rectal cancers (25%). Eight out of 152 (5.3%) NEC were microsatellite instable (MSI). NET G3 had frequent mutations in MEN1 (21%), ATRX (17%), DAXX, SETD2 and TP53 (each 14%). We show molecular differences in HG GEP-NEN, related to morphological differentiation and site of origin. Limited similarities to SCLC and a high fraction of targetable alterations indicate a high potential for better-personalized treatments.

Keywords: gastroenteropancreatic; genetic alterations; high-grade; molecular markers; neuroendocrine carcinoma; neuroendocrine neoplasms.

Figure 1

(A) Distribution of primary tumour site for the included cases of neuroendocrine carcinomas (NECs) cohort. (B) Oncoplot showing the top 50 most frequently altered genes (rows) among 152 NEC patients (columns). Upper panel shows the mutational burden per sample. Percentages on the right represent mutations frequency per gene. Genes with potentially targetable alterations are highlighted in red font (additional targetable mutations were observed among genes less frequently altered. The plot shows both potentially targetable and non-targetable alterations for these genes; for example, deletions of ESR1 and non-V600E mutations of BRAF are not considered as targetable). The panel under the oncoplot area is composed of four single row heatmaps showing in order, from top to bottom, primary tumour site, cell type, MSI status and presence of one or more potentially targetable mutation. Stacked barplot (at the bottom) shows the fraction of nucleotide changes in each sample. 'Multi_hit' indicates that more than one mutation occurs in the same gene, in the same patient. (C) Co-occurrence and mutual exclusiveness of mutations in NEC patients. Co-occurring mutations are indicated by green squares and mutually exclusive mutations between gene pairs in purple. The color intensity is proportionate to the –log₁₀ (P -value). P -values were determined using Fisher’s exact test.

Figure 2

(A) Stacked barplot illustrating the average ploidy in each of the NEC samples. (B) Frequency of copy number aberrations in the NEC cohort (n = 152). Y-axis indicates the fraction of patients with copy number losses (blue) and gains (red) across the genome. Chromosome numbers are indicated on the x-axis. Chromosomes and chromosome arms are separated by vertical lines. (C) Heatmap representing the copy number alterations for each segment, relative to average genome ploidy for each sample. Unsupervised hierarchical clustering of patients on the y-axis and chromosomes on the x-axis. (D) Significant copy-number gains (red) and losses (blue) identified by two-sided hypothesis testing using GISTIC2.0, corrected for multiple hypothesis testing. Significant regions (chromosome locus and focal copy number changes) for known cancer-associated genes are labeled.

Figure 3

Barplots indicating mutation frequency for the top 16 frequently altered genes in NEC patients, stratified for the six primary tumour sites (left colon, right colon, esophagus, gastric, pancreas, rectum). Y-axis shows the frequency (in percentage) of the alteration for each site.

Figure 4

Co-bar plots illustrating the differences in mutation frequencies for the most frequently mutated genes, between (A) large cell NEC (n = 87) vs small cell NEC (n = 65), (B) NEC (n = 152) vs NET G3 (n = 29) and (C) large cell NEC (n = 87) vs NET G3 (n = 29).