Purpose: GEP-NENs are rare malignancies with increasing incidence. Their molecular characteristics are still undefined. We explored the underlying biology of GEP-NENs and the differences between gastrointestinal (GI) and pancreatic (PNET), high grade (HG) and low grade (LG) tumors.

Experimental design: GEP-NENs were analyzed using next-generation sequencing (NGS; MiSeq on 47 genes, NextSeq on 592 genes), immunohistochemistry, and in-situ hybridization. Tumor mutational burden (TMB) was calculated based on somatic nonsynonymous missense mutations, and microsatellite instability (MSI) was evaluated by NGS of known MSI loci.

Results: In total, 724 GEP-NENs were examined: GI (N=469), PNEN (N=255), HG (N=135), and LG (N=335). Forty-nine% were female, median age was 59. Among LG tumors, the most frequently mutated genes were ATRX (13%), ARID1A (10%) and MEN1 (10%). HG tumors showed TP53 (51%), KRAS (30%), APC (27%), ARID1A (23%). Immune-related biomarkers yielded a lower prevalence in LG tumors compared to HG: MSI-H 0% vs 4% (P=.04), PD-L1 overexpression 1% vs 6% (P=.03) TMB-high 1% vs 7% (P=.05). Compared to LG, HG NETs showed a higher mutation rate in BRAF (5.4% v 0%, P<.0001), KRAS (29.4% v 2.6%, P<.0001) and PI3KCA (7% v 0.3%, P<.0001). When compared to GI, PNEN carried higher frequency of MEN1 (25.9% v 0.0%, P<.0001), FOXO3 (8.6% v 0.8%, P=.005), ATRX (20.6% v 2.0%, P=.007), and TSC2 (6.3% vs 0.0%, P=.007), but lower frequency of mutations in APC (1.0% v 13.8%, P<.0001).

Conclusions: Significant molecular differences were observed in GEP-NENs by tumor location and grade, indicating differences in carcinogenic pathways and biology.