Abstract 3121: Characterization of immune profiling of pancreatic intraductal papillary mucinous neoplasm using multiplex immunofluorescence and image analysis approaches

Pancreatic intraductal papillary mucinous neoplasms (IPMN) are cystic mucin-secreting tumors that arise within the pancreatic ductal system and are considered precursor lesions of pancreatic ductal adenocarcinoma. Profiling the immune infiltrate of IPMNs is essential to understand the immune mechanisms associated with the progression of IPMN. The aim of this study was to characterize the immune profiling of IPMN. We examined 31 formalin-fixed and paraffin-embedded tumor samples from surgically resected IPMN. The diagnostic slides were histologically reviewed by 2 expert pathologists in pancreas and classified as low-grade IPMN (low to intermediate grade dysplasia, N=11), high-grade IPMN (high grade dysplasia, N=18) and IPMN with invasive adenocarcinoma (N=2). Multiplex immunofluorescence (mIF) was used to characterize the immune profiling using two panels: Panel 1: PD-L1, PD1, CD3 (pan T cells), CD8 (T-cytotoxic), CD68 (macrophages), CK (AE1/AE3), DAPI. Panel 2: CD20 (B cells), CD45RO (T-memory), CD57 (NK), Granzyme B (NK and cytotoxic), FOXP3 (T reg), CK (AE1/AE3), DAPI. The slides were scanned using the Vectra multispectral microscope and analyzed by the InForm software (PerkinElmer). We quantified and co-localized immune phenotypes (epithelial cells PD-L1+; Total T-cell lymphocytes; T-cells antigen-experienced; Cytotoxic T-cells; Total macrophages; Macrophages PD-L1+; Total B-cell lymphocytes; Memory T cells; Granzyme B+ cells; Regulatory T cells; NK cells) in low grade dysplasia (LGD), high grade dysplasia (HGD), and invasive carcinoma areas in epithelial and stromal compartments. The Wilcoxon rank sum test or the Wilcoxon singed-rank test was used to test the equality of medians between two independent groups or paired samples, respectively. In all patients, HGD areas showed significantly higher density of macrophages (P= 0.041) and lower density of B-cell lymphocytes (P= 0.028) compared with LGD areas in the epithelial/stromal and epithelial compartments, respectively. Similar differences were observed when patients with high-grade IPMN (n=18) were analyzed for immune infiltrates in the HGD areas compared with LGD areas in the same tumor samples: HGD areas showed significantly higher density of macrophages (P= 0.035) and lower density of B-cell lymphocytes (P= 0.031). Patients with high-grade IPMN had significantly lower densities of B-cell lymphocytes (P= 0.011), T-cell lymphocytes (P= 0.034), and memory T-cells (P= 0.031) in the HGD areas when comp