Molecular Profiling of Digestive Grade 3 Neuroendocrine Tumors Reveals a Shared Molecular Framework with Lower-Grade Tumors, Marked Heterogeneity, and Therapeutic Opportunities

: Well-differentiated neuroendocrine tumors, grade 3 (NETs G3) of the digestive system are high-grade neuroendocrine neoplasms characterized by well-differentiated neuroendocrine morphology and high proliferative activity. Their molecular identity and therapeutic vulnerabilities remain incompletely defined. We performed a transcriptomic analysis and a comprehensive genomic profiling of 40 cases, including 26 NETs G3, 8 NETs G1/G2, and 6 neuroendocrine carcinomas (NECs). Targeted sequencing assessed alterations at both the DNA and RNA level, and gene expression profiling was conducted using a targeted immune-oncology panel. NETs G3 retained recurrent alterations in chromatin-regulatory genes characteristic of well-differentiated tumors and largely lacked the consistent TP53 and RB1 inactivation typical of NECs. Transcriptomic and genomic analysis demonstrated separation of NECs from NETs, with NETs G3 partially overlapping with NETs G1/G2 supporting a model of molecular relatedness. However, when compared with NETs G1/G2, NETs G3 showed upregulation of extracellular matrix remodeling, metabolic reprogramming, and cytokine signaling pathways. NETs G3 were heterogeneous in terms of number of genomic alterations per case and involved genes. A distinctive genomic feature was the predominance of large-segment copy-number losses, frequently involving chromatin-regulatory regions on chromosomes X and 10. Potentially targetable alterations were detected in up to 25% of NETs G3. Analysis of immune microenvironment supported a poorly inflamed tumor status, with individual cases harboring microsatellite instability or enrichment in tumor-infiltrating lymphocytes potentially amenable to immunotherapy.