Aberrant p16INK4A and DPC4/Smad4 expression in intraductal papillary mucinous tumours of the pancreas is associated with invasive ductal adenocarcinoma

Abstract

Background and aims: Intraductal papillary mucinous tumours (IPMT) of the pancreas constitute a unique pathological entity with an overall incidence of associated invasive malignancy of 20%. The malignant potential of an individual IPMT cannot be accurately predicted. Preoperative estimation of the risk of associated invasive malignancy with IPMT would be of significant clinical benefit. As aberrations in cell cycle regulatory genes are associated with the progression of precursor pancreatic ductal lesions to invasive adenocarcinoma, we examined expression of key cell cycle regulatory genes in the cyclin D1/retinoblastoma pathway and the transforming growth factor β/Smad4 signalling pathway in a cohort of patients with surgically resected IPMT. Methods: Sections of formalin fixed paraffin embedded pancreatic tissue from a cohort of 18 patients with IPMT were examined using immunohistochemistry for protein expression of cell cycle regulatory genes p16INK4A , p21 CIP1, p27 KIP1, cyclin D1, pRb, and p53, as well as the cell signalling molecule Smad4. A comparison of expression levels was made between adenoma/borderline IPMT (10 patients) and intraductal papillary mucinous carcinoma (IPMC) (eight patients, four of whom harboured invasive carcinoma). Statistical analysis was performed using the χ 2 and Fisher's exact tests. Results: Aberrant expression of the proteins examined increased in frequency from adenoma/ borderline IPMT to IPMC. Specifically, there was a significantly greater incidence of loss of p16 INK4A expression in IPMC: 8/8 lesions (100%) compared with 1/10 (10%) adenoma/borderline IPMT (p<0.001). Similarly, loss of Smad4 expression was associated with IPMC: 3/8 (38%) versus adenoma/borderline IPMT 0/10 (p<0.03). Loss of Smad4 expression within the IPMT was the best marker for the presence of invasive carcinoma (p<0.001). Conclusions: These data indicate that loss of p16 INK4A and Smad4 expression occur more frequently in IPMC alone, or with associated invasive carcinoma, compared with adenoma/borderline IPMT. Aberrant protein expression of these cell cycle regulatory genes in IPMT and pancreatic intraepithelial neoplasia in the current model of pancreatic cancer progression suggest similarities in their development and may also represent the subsequent risk of invasive carcinoma.

Targeted next-generation sequencing of cancer genes dissects the molecular profiles of intraductal papillary neoplasms of the pancreas.

“…The pathogenetic role of CDKN2A/P16 and SMAD4 was further investigated by immunohistochemical analysis. Our results demonstrated that 36% of IPMNs show loss of p16 and 24% of IPMN-related invasive carcinomas loss of Smad4, which is in accordance with previous studies [4,7].…”

“…Immunohistochemistry was used as a surrogate test to assess the involvement of CDKN2A/P16 and SMAD4 genes [7,24,25]. In fact, protein expression may more accurately reflect gene status, as both genes are mainly inactivated by homozygous deletion, and CDKN2A/P16 also by promoter methylation.…”

“…IPMNs commonly harbour activating mutations in KRAS and GNAS, and inactivating mutations in RNF43, CDKN2A/p16 and TP53, and less commonly mutations in BRAF, PIK3CA, STK11 and SMAD4 [5][6][7][8][9][10][11][12]. TP53, CDKN2A/p16 and SMAD4 mutations and/or loss of expression are generally found in higher-grade lesions [13][14][15].…”

Fascin overexpression in intraductal papillary mucinous neoplasms (adenomas, borderline neoplasms, and carcinomas) of the pancreas, correlated with increased histological grade

“…29,30 The fact that PanINs show fascin upregulation correlated with histological grade increased our interest in fascin expression in IPMNs, because PanINs and IPMNs share the following fundamental characteristics: 5,6 inherently intraductal; composed predominantly of columnar, mucin-producing cells that may grow in a flat configuration or may produce papillae; exhibit a range of cytologic and architectural atypia (mild, moderate and severe); recognized as precursors to Fascin expression in IPMNs of the pancreas H Yamaguchi et al invasive adenocarcinoma; and sequentially accumulate similar genetic alterations with increasing cytoarchitectural atypia. [43][44][45] We showed that fascin overexpression in IPMNs was correlated with increased histological grade by immunohistochemical analysis, followed by a supporting molecular experiment that showed upregulation of fascin mRNA. We consider that fascin upregulation would be a relatively early event in the progression of IPMN, because of the finding that fascin expression was significantly and almost equally greater in borderline neoplasms (86%) and carcinomas (88%) than in adenomas (51%).…”

The genetic classification of pancreatic neoplasia

“…Somatic TP53 mutations are almost exclusively found in areas of high-grade dysplasia or invasion, which indicates that this mutation is a relatively late event in carcinogenesis [88][89][90][91]. Most noninvasive IPMNs show intact expression of the tumor suppressor gene SMAD4, whereas SMAD4 loss can be seen in invasive carcinomas arising in association with IPMNs, again suggesting that SMAD4 is targeted late in neoplastic progression [92,93]. Similarly, loss of p16 is very common in invasive carcinoma arising in association with an IPMN, whereas p16 is preserved in most lower-grade IPMNs [93].…”