A Practical Approach to the Classification of WHO Grade 3 (G3) Well-differentiated Neuroendocrine Tumor (WD-NET) and Poorly Differentiated Neuroendocrine Carcinoma (PD-NEC) of the Pancreas

Abstract

High-grade neuroendocrine neoplasms (World Health Organization [WHO] G3) of the pancreas include both well-differentiated neuroendocrine tumor (WD-NET) and poorly differentiated neuroendocrine carcinoma (PD-NEC). According to the WHO classification scheme, the diagnosis of this group of tumors is based on both the histopathology of the tumor and the assessment of proliferation fraction. However, the former can be challenging due to the lack of well-defined histologic criteria, and the latter alone (ie, >20 mitoses/10 high-power fields or Ki67>20%) may not sufficiently distinguish WD-NETs from PD-NECs. Given the considerable differences in treatment strategies and clinical outcome, additional practical modalities are required to facilitate the accurate diagnosis of high-grade pancreatic neuroendocrine neoplasms. We examined 33 cases of WHO G3 neuroendocrine neoplasms of the pancreas and attempted to classify them into WD-NET, small cell PD-NEC (PD-NEC-SCC), and large cell PD-NEC (PD-NEC-LCC) or to designate them as "ambiguous" when an uncertain diagnosis was rendered by any of the observers or there was any disagreement in classification among the 3 observers. To simplify the interpretation, both PD-NEC-SCC and PD-NEC-LCC were considered together as PD-NECs in the final analysis. The initial approach was to assess microscopically a single morphologically challenging hematoxylin and eosin section from each case without the knowledge of Ki67 values, performed independently by 3 pathologists to assess the degree of diagnostic concordance, and then evaluate immunohistochemical staining for surrogate biomarkers of known genotypes of WD-NET and PD-NEC, respectively, and, lastly, complete a clinicopathologic review to establish a final definitive classification. Loss of DAXX or ATRX protein expression defined WD-NET, and abnormal p53, Rb, SMAD4 expression signified PD-NEC. When the chosen section displayed an element of WD histopathology, or other tumor sections contained WHO G1/G2 components, or there had been a prior established diagnosis of a primary WD-NET, the final diagnosis was rendered as a WD-NET with high-grade (G3) progression. If a component of conventional adenocarcinoma was present (in slides not seen in the initial review), the diagnosis was established as a combined adenocarcinoma and PD-NEC. All 3 pathologists agreed on the morphologic classification of 33% of the cases (6 WD-NET, 3 PD-NEC-SCC, and 2 PD-NEC-LCC), were conflicted on 2 cases between PD-NEC-SCC and PD-NEC-LCC, and disagreed or were uncertain on the classification for the remaining 20 cases (61%), which were therefore categorized as ambiguous. In the group of cases in which all pathologists agreed on the classification, the 6 WD-NET cases had either loss of DAXX or ATRX or had evidence of a WD-NET based on additional or prior pathology slides. The 7 PD-NEC cases had abnormal expression of p53, Rb, and/or SMAD4 or a coexisting adenocarcinoma. In the ambiguous group (n=20), 14 cases were established as WD-NETs, based upon loss of DAXX or ATRX in 7 cases and additional pathology evidence of high-grade progression from WD-NET in the other 7 cases; 5 cases were established as PD-NEC based upon abnormal expression of p53, Rb, and/or SMAD4; 1 case remained undetermined with normal expression of all markers and no evidence of entity-defining histologic findings in other slides. On the basis of the final pathologic classifications, the disease-specific survival was 75 and 11 months for the WD-NET and PD-NEC groups, respectively. Thus, we conclude that morphologic diagnosis of high-grade pancreatic neuroendocrine neoplasms is challenging, especially when limited pathologic materials are available, and necessitates better defined criteria. The analysis of both additional sections and prior material, along with an immunohistochemical evaluation, can facilitate accurate diagnosis in the majority of cases and guide the appropriate clinical management and prognosis.

Figures

Figure 1. Typical Morphologic Features of Pancreatic WD-NET, PD-NEC-SCC, and PD-NEC-LCC

WD-NETs (A-B) revealed nested/organoid and trabecular architecture, a regular intratumoral vascular pattern, abundant granular cytoplasm, and stippled nuclei with inconspicuous nucleoli. PD-NEC-SCC (C-D) demonstrated stromal desmoplasia (C), tumor necrosis (C), fusiform (“oat cell”) nuclei lacking nucleoli, and nuclear molding. PD-NEC-LCC (E-F) displayed tumor necrosis, expansile and irregular nests with peripheral palisading, and rosettes/tubular structures within the large nests.

Figure 2. Morphologically Ambiguous Pancreatic Neuroendocrine Neoplasms

Two cases of WD-NET (A-B) were initially considered as morphologically ambiguous due to an infiltrative growth pattern with irregular architecture, significant intratumoral fibrosis, single cell (A) and punctate (B) tumor necrosis, and brisk mitotic activity. However, upon retrospective review, the tumors appeared to retain some morphologic features of WD-NETs such as a hyalinized type of fibrosis (A), delicate vascular pattern (B), and abundant granular cytoplasm and low nuclear to cytoplasm (N/C) ratio (B). The two cases of PD-NEC (C-D) shared some morphologic features of WD-NET such as the vascular patterns and nested or trabecular architecture. Although the cytologic features such as large nuclei, high N/C ratio, and minimal cytoplasm might have suggested PD-NEC. The distinction between WD-NET (E) and PD-NEC (F) was especially difficult in small biopsies where the architecture of the tumor could not be fully appreciated. While the cytologic features of the tumor (abundant cytoplasm and low N/C ratio) were suggestive of a WD-NET (E), the presence of extensive tumor necrosis rendered the tumor as ambiguous. Similarly, the small nested structures in a PD-NEC (F) without the context of the global architecture of the tumor could culminate in an incorrect classification of this neuroendocrine neoplasm.

Figure 3. Abnormal p53, Rb, SMAD4, and DAXX expression by Immunohistochemistry in High Grade Pancreatic WD-NET and PD-NEC

The expression of abnormal p53 served as a surrogate biomarker of TP53 gene mutation, which was observed in 67% of PD-NECs (A). Similarly, loss of Rb (B) and SMAD4 (C) protein expression were associated with PD-NEC. In contrast, loss of DAXX (D) or ATRX (data not shown) expression was seen in 40-50% WD-NET, but not in PD-NEC.

Figure 4. Disease Specific Survival of High Grade Pancreatic Neuroendocrine Neoplasm

Disease specific survival of the entire cohort (A) and disease specific survival of morphologically ambiguous cases (B)

Figure 5. Recommended Diagnostic Algorithm for Pancreatic High Grade Neuroendocrine Neoplasms