Cytopathology in focus—Low-grade pancreatic neuroendocrine tumors in small samples: Grading challenges

Christopher O’Conor, MD, PhD
M. Lisa Zhang, MD

August 2024—Two to five percent of pancreatic tumors are pancreatic neuroendocrine neoplasms, of which more than 90 percent are well-differentiated pancreatic neuroendocrine tumors (PanNETs). The current World Health Organization classification stratifies PanNETs into three histological grades based on mitotic count and Ki-67 proliferation index (Table 1). Low-grade PanNETs (grades one and two) are morphologically indistinguishable and have characteristic neuroendocrine cytomorphologic features: a dispersed/loosely cohesive smear pattern composed of small-to-medium monomorphic cells with occasional plasmacytoid appearance; amphophilic, granular, or vacuolated cytoplasm; round nuclei; and coarse salt-and-pepper chromatin.

The grading criteria for PanNETs are based on examination of resection specimens and therefore can pose challenges for grading on small biopsy and fine-needle aspiration samples. For mitotic rate, the most recent CAP protocol for PanNET (December 2023) recommends that at least 10 mm² be evaluated in the most mitotically active part of the tumor, which corresponds to 42–50 high-power fields at 40× magnification on commonly used microscopes with ocular field diameters of 0.50–0.55 mm. Given that FNA samples typically do not contain at least 42 HPF of tumors, small biopsy/cell block samples are generally insufficient for grading based on mitotic count.

For evaluation of the Ki-67 proliferation index, the CAP protocol recommends that a minimum of 500 tumor cells be counted, with some experts recommending at least 2,000 cells, which may again not be possible on FNA samples. Two studies^1,2 have investigated grading PanNETs by Ki-67 immunohistochemistry in FNA cell blocks. Both studies included cell blocks with more than 100 tumor cells and found that compared with the final grade established on surgical specimens, grading on cell blocks using a Ki-67 manual count had 69 percent and 73 percent concordance in each study, respectively. In particular, Jin, et al.,¹ found that approximately 40 percent of grade two tumors were undergraded on cell block, 78 percent of which contained more than 500 tumor cells, highlighting the fact that small biopsies may not be representative of proliferative “hot spots” even with “adequate” cellularity. Abi-Raad, et al.,² found that all grade two PanNETs were undergraded in specimens with fewer than 1,000 tumor cells while cell blocks with 1,000 or more tumor cells demonstrated significantly improved categorization of grade two PanNETs (64 percent concordance). As one might expect, the concordance rate of cytology/biopsy and resection grading appears to be higher in patients with smaller tumors. A multi-institutional study found that small tumors (<2 cm) had significantly higher rates of concordance compared with larger tumors (98 percent versus 76 percent).³ Finally, different methods of counting the Ki-67 proliferation index in small samples were also examined in these studies, including “eyeballing,” hot spot counting, complete counting, and digital image analysis; manual counting of the hot spot demonstrated the highest concordance with surgical specimens.^1,2

In summary, cytopathologists are encountering increasing numbers of FNA specimens for the diagnosis and grading of PanNETs. Undergrading of grade two PanNETs based on Ki-67 evaluation of limited FNA material is a common pitfall; therefore, caution is recommended in interpretation and reporting (Fig. 1 and example report). Based on the available evidence, grading is most reliable on specimens containing 1,000 or more tumor cells and from tumors less than 2 cm in size.

1. Jin M, Roth R, Gayetsky V, Niederberger N, Lehman A, Wakely PE Jr. Grading pancreatic neuroendocrine neoplasms by Ki-67 staining on cytology cell blocks: manual count and digital image analysis of 58 cases. J Am Soc Cytopathol. 2016;5(5):286–295.
2. Abi-Raad R, Lavik JP, Barbieri AL, Zhang X, Adeniran AJ, Cai G. Grading pancreatic neuroendocrine tumors by Ki-67 index evaluated on fine-needle aspiration cell block material. Am J Clin Pathol. 2020;153(1):74–81.
3. Javed AA, Pulvirenti A, Razi S, et al.; Pancreatic Neuroendocrine Disease Alliance (PANDA). Grading pancreatic neuroendocrine tumors via endoscopic ultrasound-guided fine needle aspiration: a multi-institutional study. Ann Surg. 2023;277(6):e1284–e1290.

Dr. O’Conor is assistant professor and associate director of quality in cytopathology, Vanderbilt University Medical Center, and director of cytology, VA Tennessee Valley Healthcare System. Dr. Zhang is a cytopathologist and gastrointestinal pathologist, Massachusetts General Hospital, and instructor in pathology, Harvard Medical School. Both are members of the CAP Cytopathology Committee.