Discover how immunohistochemistry (IHC) markers from Atlas Antibodies, built on the legacy of the Human Protein Atlas, advance cancer research and improve diagnosis of neuroendocrine neoplasms (NENs).
What are neuroendocrine neoplasms?
Neuroendocrine neoplasms (NENs) are a diverse group of tumors that develop from neuroendocrine cells — specialized sensory and secretory cells that release hormones into the bloodstream and act as a link between the nervous and endocrine systems.
These cells are distributed throughout the body, which is why NENs can arise in almost any organ. However, they most frequently occur in the gastrointestinal tract (around 61%) and the bronchopulmonary system (about 25%), followed by the pancreas (around 8%), reflecting the high density of neuroendocrine cells in these regions.
Neuroendocrine neoplasms can either be classified as functioning (producing excess hormones causing symptoms) or non-functioning (not producing excess hormones or not enough to cause symptoms).
How are NENs diagnosed?
A correct diagnosis is imperative for the patient to obtain the most efficient treatment. However, diagnosing NENs is challenging because symptoms are often nonspecific, such as chronic fatigue or pain unrelated to a specific injury. Often, NENs are found incidentally while testing for other conditions.
The ultimate NEN diagnosis can only be established with tissue biopsy and further tumor grading based on combining the proliferative index with cell mitotic rate.
Standard laboratory diagnosis assessment relies mainly on markers such as Chromogranin A (CHGA), neuron-specific enolase (NSE), synaptophysin (SYP), and neural cell adhesion molecule 1 (NCAM1 or CD56). However, while virtually all carcinoids are positive for SYP and CHGA, their expression is highly variable in NENs.
Therefore, the immunohistochemical assessment of the complex biomarker expression patterns in targeted biopsies is fundamental and instrumental in all phases of the diagnostic process, such as differentiation (neuroendocrine or epithelial origin) and proliferation (grading and staging)
Step 1: Confirm the Neuroendocrine Nature Using First-Generation IHC Markers
Objective: Verify that the tumor exhibits neuroendocrine differentiation.
Key Markers:
- Chromogranin A (CHGA): A secretory granule protein that serves as a gold-standard neuroendocrine marker. Strong CHGA expression correlates with well-differentiated neuroendocrine tumors (NETs).
Evidence: Studies show CHGA sensitivity of ~85–90% for well-differentiated NETs, but reduced expression in high-grade NECs. - Synaptophysin (SYP): A synaptic vesicle glycoprotein; highly sensitive across the spectrum of NENs.
Evidence: SYP remains expressed even when CHGA is lost in poorly differentiated NECs, providing complementary diagnostic value. - CD56 (NCAM), CD57 (B3GAT1), UCHL1, and NSE (Neuron-Specific Enolase): Supportive markers with variable sensitivity and specificity.
Evidence: While NSE and CD56 are highly sensitive, they lack specificity, making them adjunct markers rather than definitive indicators.
Step 2: Confirm Epithelial Origin
Objective: Differentiate epithelial-derived NENs from non-epithelial mimics (e.g., paragangliomas, melanomas, lymphomas).
Key Markers:
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AE1/AE3 (Pan-cytokeratin cocktail):
AE1 targets acidic (type I) cytokeratins — mainly CK10, CK14, CK15, CK16, and CK19.
AE3 targets basic (type II) cytokeratins — CK1, CK2, CK3, CK4, CK5, CK6, CK7, and CK8.
When combined, AE1/AE3 detects a broad spectrum of cytokeratins across both squamous and glandular epithelia, making it a pan-epithelial marker.
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CAM5.2 (Low–molecular weight cytokeratin marker):
Recognizes CK8 (KRT8) and CK18 (KRT18), which are characteristic of simple and glandular epithelia.
Provides confirmatory evidence of epithelial lineage, especially in neuroendocrine tumors, adenocarcinomas, and ductal epithelia.
Evidence and diagnostic relevance: Almost all epithelial neuroendocrine neoplasms (NENs) express cytokeratins (detectable by AE1/AE3 or CAM5.2). Paragangliomas, in contrast, are typically cytokeratin-negative but synaptophysin-positive, helping pathologists differentiate them from epithelial NENs.
Step 3: Determine Tumor Grade and Proliferation Index
Objective: Establish proliferative activity for grading and prognostic assessment.
Key Marker:
- Ki-67 (MIB-1): Nuclear proliferation marker used to assign WHO grades (G1–G3).
Evidence: Ki-67 index correlates strongly with mitotic rate and clinical outcomes in both gastrointestinal and pancreatic NENs (WHO Classification of Endocrine and Neuroendocrine Tumors, 5th Ed., 2022).
Step 4: Refine Neuroendocrine Differentiation Using Second-Generation IHC Markers
Objective: Identify poorly differentiated or dedifferentiated NENs where classical markers may be downregulated.
These markers target transcriptional regulators and calcium-binding proteins critical for neuroendocrine differentiation:
- Anti-ISL1 (Isl Lim homeobox1, AMAb91729)
- Anti-INSM (Insm transcriptional repressor1, AMAb91727)
- Anti-SCGN (Secretagogin, AMAb90630, and AMAb90632)
- Anti-OTP (Orthopedia homeobox, AMAb91695, and AMAb91696)
Evidence:
INSM1: Demonstrated sensitivity >90% across NEN types, with superior nuclear staining clarity compared to CHGA or SYP.
ISL1 and OTP: Useful for site determination (e.g., OTP positivity in pulmonary carcinoids, ISL1 in pancreatic NETs).
SCGN: Expressed in various endocrine tissues and stable even in high-grade NECs.
Interpretation:
Unlike first-generation secretory markers, these second-generation transcriptional markers retain expression in dedifferentiated tumors that have lost their secretory machinery, enhancing diagnostic confidence in challenging cases.
Step 5: Integrate Classical and Modern Markers for Diagnostic and Clinical Utility
Objective: Combine marker generations to improve accuracy, prognostication, and therapy stratification.
Clinical Application:
A dual-panel approach (e.g., CHGA/SYP + INSM1/ISL1) ensures sensitivity across the NEN spectrum. This improves tumor classification, guides therapeutic decisions (e.g., somatostatin analog eligibility), and supports longitudinal follow-up through reliable biomarker monitoring.
Scientific Consensus:
Pathology guidelines now recommend combined use of classical and second-generation IHC markers for optimal sensitivity and specificity in NEN diagnosis (WHO 2022; Bellizzi, Modern Pathology, 2020).
In conclusion, first-generation markers such as Chromogranin A (CHGA) and Synaptophysin (SYP) remain the gold standard for confirming neuroendocrine differentiation in routine endocrine pathology.
However, integrating second-generation markers into immunohistochemical panels offers enhanced sensitivity and specificity, particularly in cases where classical markers are weak or absent. This combined IHC approach represents a promising diagnostic strategy for more accurately detecting tumors with neuroendocrine differentiation, improving both diagnostic precision and clinical management of NENs.
