Identification of tumour regression in neoadjuvantly treated pancreatic cancer is based on divergent and nonspecific criteria.

AIMS: Following the increased use of neoadjuvant therapy for pancreatic cancer, grading of tumour regression (TR) has become part of routine diagnostics. However, it suffers from marked interobserver variation, which is mainly ascribed to the subjectivity of the defining criteria of the categories in TR grading systems. We hypothesized that a further cause for the interobserver variation is the use of divergent and nonspecific morphological criteria to identify tumour regression. METHODS AND RESULTS: Twenty treatment-naïve pancreatic cancers and 20 pancreatic cancers treated with neoadjuvant chemotherapy were reviewed by three experienced pancreatic pathologists who, blinded for treatment status, categorized each tumour as treatment-naïve or neoadjuvantly treated, and annotated all tissue areas they considered showing tumour regression. Only 50%-65% of the cases were categorized correctly, and the annotated tissue areas were highly discrepant (only 3%-41% overlap). When the prevalence of various morphological features deemed to indicate TR was compared between treatment-naïve and neoadjuvantly treated tumours, only one pattern, characterized by reduced cancer cell density and prominent stroma affecting a large area of the tumour bed, occurred significantly more frequently, but not exclusively, in the neoadjuvantly treated group. Finally, stromal features, both morphological and biological, were investigated as possible markers for tumour regression, but failed to distinguish TR from native tumour stroma. CONCLUSION: There is considerable divergence in opinion between pathologists when it comes to the identification of tumour regression. Reliable identification of TR is only possible if it is extensive, while lesser degrees of treatment effect cannot be recognized with certainty.

Prognostic Impact of Resection Margin Status on Distal Pancreatectomy for Ductal Adenocarcinoma.

Pancreatic cancer is associated with a poor prognosis. While surgical resection is the only treatment option with curative intent, most patients die of locoregional and/or distant recurrence. The prognostic impact of the resection margin status has received much attention. However, the evidence is almost exclusively related to pancreatoduodenectomies, while corresponding data for distal pancreatectomy specimens are limited. The key data, such as the rate of microscopic margin involvement ("R1"), the site of margin involvement, and the impact of R1 on patient outcome, are divergent between studies and do not currently allow any general conclusions. The main reasons for the variability in the published data are the small size of the study cohorts and their heterogeneity, as well as the marked divergence in pathology examination practices. The latter is a consequence of the lack of concrete guidance, both for grossing and microscopic examination. The increasing administration of neoadjuvant chemo(radio)therapy introduces a further factor of uncertainty as the conventional definition of a tumour-free margin ("R0") based on 1 mm clearance is inadequate for these specimens. This review discusses the published data regarding the prognostic impact of margin status in distal pancreatectomy specimens along with the challenges and uncertainties that are related to the assessment of the margins.

Tumour Size and T-Stage in Pancreatic Cancer Resection Specimens Depend on the Pathology Examination Approach.

In the eighth edition of the TNM classification for pancreatic ductal adenocarcinoma (PDAC), stages T1 to T3 are defined by tumour size, size measurement being deemed objective and accurate. This study investigated whether various, currently used approaches to tumour measurement result in different tumour sizes and differences in T-stage assignment. In a series of 315 resected PDAC, tumour sizes were measured as follows: macroscopically in a single or in two perpendicular planes and with or without microscopic corroboration. Comparison of the resulting tumour sizes showed that both macroscopic measurement in two planes and microscopic corroboration gave significantly different results (p < 0.001). Compared to the most simple approach (macroscopic measurement in one plane), the comprehensive approach (macroscopic measurement in two planes with microscopic corroboration) resulted in a larger tumour size in 263 (83%) cases (mean absolute size difference: 10 mm; mean relative size change: 36%). T-stage assignment differed in 142 (45%) cases between the simple and comprehensive approach and affected 87%, 38% and 48% of the cases deemed to be stage T1, T2 and T3, respectively. In conclusion, tumour size and T-stage are highly approach-dependent. Consensus on an accurate method is required to ensure comparability of these basic data.

Gene expression profiling of morphologic subtypes of pancreatic ductal adenocarcinoma using surgical and EUS-FNB specimens.

BACKGROUND/OBJECTIVES: Various classifications of pancreatic ductal adenocarcinoma (PDAC) based on RNA profiling resulted in two main subtypes. Kalimuthu and coworkers proposed a morphology-based classification that concurred with these subtypes. Immune therapy approaches in PDAC were so far disappointing. Morphologic PDAC subtypes may differ regarding key immune-oncology pathways. We aimed to examine the reproducibility and prognostic value of Kalimuthu's morphologic classification, and to evaluate differences between subtypes regarding gene expression related to tumor biology and immune-oncology. METHODS: PDAC specimens from 196 patients were included, 108 consecutive chemotherapy-naïve surgical specimens and 88 endoscopic ultrasound-guided fine needle biopsies (EUS-FNBs). The specimens were evaluated as per Kalimuthu by two pancreatic pathologists, resulting in Group A and Group B tumors. Digital mRNA expression profiling was performed, on the surgical specimens using the NanoString IO360 panel of 770 key tumor biology related and 30 custom-genes, and on the EUS-FNBs using a targeted panel of 123 genes. RESULTS: Morphologic subtyping reached substantial interobserver agreement between the two pathologists. In the surgical and EUS-FNB cohorts, 44.4% and 38.6% were Group A tumors, which were associated with improved survival. Group A showed higher expression of immune-related genes and cytokine/chemokine/interleukin signaling and Group B of genes related to cancer cell proliferation and cell cycle regulation. Hierarchical clustering based on significant differences in gene expression levels between Groups A and B revealed clusters with prognostic value. CONCLUSIONS: Morphologic subtyping according to Kalimuthu is reproducible and holds prognostic value, in surgical as well as EUS-FNB specimens. As upregulation of immune-related genes was found in Group A, future studies should evaluate the potential of immune therapy approaches with special emphasis on this subtype of PDAC.

Cytokeratin-positive cells in the bone marrow from patients with pancreatic, periampullary malignancy and benign pancreatic disease show no prognostic information.

BACKGROUND: Pancreatic and periampullary carcinoma are aggressive tumours where preoperative assessment is challenging. Disseminated tumour cells (DTC) in the bone marrow (BM) are associated with impaired prognosis in a variety of epithelial cancers. In a cohort of patients with presumed resectable pancreatic and periampullary carcinoma, we evaluated the frequency and the potential prognostic impact of the preoperative presence of DTC, defined as cytokeratin-positive cells detected by immunocytochemistry (ICC). METHODS: Preoperative BM samples from 242 patients selected for surgical resection of presumed resectable pancreatic and periampullary carcinoma from 09/2009 to 12/2014, were analysed for presence of CK-positive cells by ICC. The median observation time was 21.5 months. Overall survival (OS) and disease-free survival (DFS) were calculated by Kaplan-Meier and Cox regression analysis. RESULTS: Successful resections of malignant tumours were performed in 179 of the cases, 30 patients resected had benign pancreatic disease based on postoperative histology, and 33 were deemed inoperable intraoperatively due to advanced disease. Overall survival for patients with resected carcinoma was 21.1 months (95% CI: 18.0-24.1), for those with benign disease OS was 101 months (95% CI: 69.4-132) and for those with advanced disease OS was 8.8 months (95% CI: 4.3-13.3). The proportion of patients with detected CK-positive cells was 6/168 (3.6%) in resected malignant cases, 2/31 (6.5%) in advanced disease and 4/29 (13.8%) in benign disease. The presence of CK-positive cells was not correlated to OS or DFS, neither in the entire cohort nor in the subgroup negative for circulating tumour cells (CTC). CONCLUSIONS: The results indicate that CK-positive cells may be present in both patients with malignant and benign diseases of the pancreas. Detection of CK-positive cells was not associated with differences in prognosis for the entire cohort or any of the subgroups analysed. TRIAL REGISTRATION: clinicaltrials.gov ( NCT01919151 ).

European Guideline on IgG4-related digestive disease - UEG and SGF evidence-based recommendations.

The overall objective of these guidelines is to provide evidence-based recommendations for the diagnosis and management of immunoglobulin G4 (IgG4)-related digestive disease in adults and children. IgG4-related digestive disease can be diagnosed only with a comprehensive work-up that includes histology, organ morphology at imaging, serology, search for other organ involvement, and response to glucocorticoid treatment. Indications for treatment are symptomatic patients with obstructive jaundice, abdominal pain, posterior pancreatic pain, and involvement of extra-pancreatic digestive organs, including IgG4-related cholangitis. Treatment with glucocorticoids should be weight-based and initiated at a dose of 0.6-0.8 mg/kg body weight/day orally (typical starting dose 30-40 mg/day prednisone equivalent) for 1 month to induce remission and then be tapered within two additional months. Response to initial treatment should be assessed at week 2-4 with clinical, biochemical and morphological markers. Maintenance treatment with glucocorticoids should be considered in multi-organ disease or history of relapse. If there is no change in disease activity and burden within 3 months, the diagnosis should be reconsidered. If the disease relapsed during the 3 months of treatment, immunosuppressive drugs should be added.

Genome-wide transcriptome profiling of ex-vivo precision-cut slices from human pancreatic ductal adenocarcinoma.

Ex-vivo tumor tissue culture systems are used as models to test specific anti-cancer drugs. Their main advantage is that they are closely comparable with the in vivo tumor in their host organism. We previously reported that precision-cut organotypic tissue slices of pancreatic ductal adenocarcinoma (PDAC) can be successfully cultured ex-vivo for at least 4 days. In order to study how culturing might affect transcription patterns, we now performed genome-wide transcriptome profiling of both baseline (0 h) and explanted tumors at daily intervals (24, 48 and 72 h) after start of culturing. The total-RNA from five samples of surgically resected human PDAC tumors at baseline and at different time points in culture was sequenced. Differential gene expression analysis of the whole transcriptome, testing 58,713 genes and over 206,000 transcripts, found that only a small number of genes showed significant changes in expression between baseline and cultured samples. The cultured tumor slices showed upregulation of a median of 12, 10 and 15 genes and downregulation of a median of 15, 12 and 25 genes at 24, 48 and 72 h in culture, respectively. One sample had morphologically increasing loss of tissue viability (range 0-18%). The vascular endothelial growth factor A (VEGFA) was significantly upregulated during the entire culture period in this case. Pathway over-representation analysis suggested that VEGFA together with the PTGS2 gene were upregulated at the same time as HIF-1-triggered cell apoptosis via NF-ĸB and the AP-1 activating factor was induced. Indeed, increased areas of apoptotic lesions were visible in this sample after 24 hours of culture. In conclusion, genome-wide transcriptome analysis supports that ex-vivo cultured tissue slices of PDAC may be a representative model of the original tumor. Transcriptome analysis was found to be a valuable complement to morphology for evaluation of ex-vivo cultures of PDAC.

Circulating Tumor Cells are an Independent Predictor of Shorter Survival in Patients Undergoing Resection for Pancreatic and Periampullary Adenocarcinoma.

OBJECTIVE: We evaluated the prognostic impact of circulating tumor cells (CTCs) for patients with presumed resectable pancreatic and periampullary cancers. SUMMARY OF BACKGROUND DATA: Initial treatment decisions for this group are currently taken without a reliable prognostic marker. The CellSearch system allows standardized CTC-testing and has shown excellent specificity and prognostic value in other applications. METHODS: Preoperative blood samples from 242 patients between September 2009 and December 2014 were analyzed. One hundred seventy-nine patients underwent tumor resection, of whom 30 with stage-I tumors and duodenal cancer were assigned to the low-risk group, and the others to the high-risk group. Further 33 had advanced disease, 30 benign histology. Observation ended in December 2016. Cancer-specific survival (CSS) and disease-free survival (DFS) were calculated by log-rank and Cox regression. RESULTS: CTCs (CTC-positive; ≥1 CTC/7.5 mL) were detected in 6.8% (10/147) of the high-risk patients and 6.2% (2/33) with advanced disease. No CTCs (CTC-negative) were detected in the low-risk patients or benign disease. In high-risk patients, median CSS for CTC-positive versus CTC-negative was 8.1 versus 20.0 months (P < 0.0001), and DFS 4.0 versus 10.5 months (P < 0.001). Median CSS in advanced disease was 7.7 months. Univariate hazard ratio (HR) of CTC-positivity was 3.4 (P < 0.001). In multivariable analysis, CTC-status remained independent (HR: 2.4, P = 0.009) when corrected for histological type (HR: 2.7, P = 0.030), nodal status (HR: 1.7, P = 0.016), and vascular infiltration (HR: 1.7, P = 0.001). CONCLUSION: Patients testing CTC-positive preoperatively showed a detrimental outcome despite successful tumor resections. Although the low CTC-rate seems a limiting factor, results indicate high specificity. Thus, preoperative analysis of CTCs by this test may guide treatment decisions and warrants further testing in clinical trials.

Kidney Involvement in Patients with Type 1 Autoimmune Pancreatitis.

INTRODUCTION: Autoimmune pancreatitis (AIP) type 1 is a special form of chronic pancreatitis with a strong lymphocytic infiltration as the pathological hallmark and other organ involvement (OOI). IgG4-related kidney disease (IgG4-RKD) was first reported as an extrapancreatic manifestation of AIP in 2004. The aim of the present study was to determine the frequency and clinical impact of kidney lesions observed in patients with AIP type 1. METHODS: We performed a single-centre retrospective study on a prospectively collected cohort of patients with a histologically proven or highly probable diagnosis of AIP according to the International Consensus Diagnostic Criteria (ICDC) classification. RESULTS: Seventy-one patients with AIP were evaluated. AIP type 1 was diagnosed in 62 (87%) patients. Kidney involvement was present in 17 (27.4%) patients with AIP type 1: 15 (88.2%) males and 2 (11.8%) females. Laboratory and/or imaging signs of kidney involvement were presented at the time of AIP diagnosis in eight (47.1%) patients. In other patients, the onset of kidney involvement occurred between four months and eight years following diagnosis. At the time of the diagnosis of kidney involvement, eight (47.1%) patients showed elevated creatinine, and nine (52.9%) patients showed normal serum creatinine. None of the patients were treated with dialysis. CONCLUSIONS: IgG4-RKD was present in 27.4% of patients with AIP type 1, with male gender predominance. In cases of early diagnosis and cortisone treatment, the clinical course was mild in most cases. Regular laboratory control of renal function should be a part of the follow-up of patients with AIP type 1.

Diagnosis, treatment and long-term outcome of autoimmune pancreatitis in Sweden.

INTRODUCTION: Autoimmune pancreatitis (AIP) is a pancreatic inflammatory process characterized by a strong inflammatory cell infiltration and two histopathologically distinct subtypes: type 1 and type 2. Diagnosis is often challenging and requires a combination of clinical, laboratory and imaging data. AIP can mimic pancreatic tumours leading to unnecessary resections if not correctly diagnosed. Short- and long-term outcomes of AIP have been poorly investigated so far and no large series have been previously reported from Sweden. METHODS: A single-centre, retrospective, cohort study of patients with histologically confirmed or highly probable diagnosis of AIP according to ICDC criteria. Demographic, clinical and radiological characteristics, type of treatment and its outcomes were collected and analysed. RESULTS: Seventy-one patients with AIP (87% with type 1), were evaluated at Karolinska University Hospital between 2004 and 2018; 49% males, mean age 49 years (range 44-53). Among them, 28% were histologically confirmed, 35% presented with jaundice, 22% with acute pancreatitis, 39% had non-specific symptoms such as weight loss or abdominal pain, 84% showed other organ involvement (OOI). Radiologically, 76% showed a focal pancreatic enlargement, 27% diffuse enlargement, 27% signs of acute pancreatitis and 10% of chronic pancreatitis. Overall, 58 patients (81%) underwent treatment with different medications: 46 (79%) cortisone, 7 (12%) azathioprine, 5 (8%) other immunosuppressive drugs. Twenty-six (36%) underwent biliary stenting and 12 (16%) were given surgery. In total, 47% of patients developed pancreatic exocrine insufficiency (PEI), of whom 76% had a severe form (faecal elastase-1 < 100 µg/g) and 21% of patients developed diabetes mellitus (pancreatic endocrine insufficiency), of whom 73% required insulin. CONCLUSIONS: AIP is a challenging disease for diagnosis and treatment. Cortisone treatment is generally successful and provides clinical remission in the large majority of patients (>90%). In the further course of the disease, a considerable number of patients develop PEI and diabetes. Only one-quarter of patients exhibit on imaging the characteristic "sausage-like" pancreas (diffuse enlargement), approximately three-quarters had a focal mass that could be misdiagnosed as pancreatic malignancy.

Stroma-regulated HMGA2 is an independent prognostic marker in PDAC and AAC.

BACKGROUND: The HMGA2 protein has experimentally been linked to EMT and cancer stemness. Recent studies imply that tumour-stroma interactions regulate these features and thereby contribute to tumour aggressiveness. METHODS: We analysed 253 cases of pancreatic ductal adenocarcinoma (PDAC) and 155 cases of ampullary adenocarcinoma (AAC) for HMGA2 expression by IHC. The data were correlated with stroma abundance and supplemented by experimental studies. RESULTS: HMGA2 acts as an independent prognostic marker associated with a significantly shorter overall survival in both tumour types. Overall, HMGA2-positivity was more frequent in patients with PDAC than with AAC. The HMGA2 status in tumour cells significantly correlated with the abundance of PDGFRß-defined stroma cells. In vivo co-injection of Panc-1 cancer cells with pancreatic stellate cells increased tumour growth in a manner associated with increased HMGA2 expression. Furthermore, in vitro treatment of Panc-1 with conditioned media from PDGF-BB-activated stellate cells increased their ability to form tumour spheroids. CONCLUSIONS: This study identifies HMGA2 expression in tumour cells as an independent prognostic marker in PDAC and AAC. Correlative data analysis gives novel tissue-based evidence for a heterotypic cross-talk with stroma cells as a possible mechanism for HMGA2 induction, which is further supported by experimental models.

Dissection of Pancreatic Resection Specimens.

Specimen grossing is a key step in the pathology examination of pancreatic resection specimens. Optimal display of pathologic changes and extensive tissue sampling are important determinants of the quality of pathology reporting. Divergence in macroscopic examination practice has led to considerable variation in the reporting of factors that are of clinical and prognostic significance. This article provides a detailed account of the macroscopic examination procedure with reference to current (inter-)national guidelines and recommendations.

Profile of MMP and TIMP Expression in Human Pancreatic Stellate Cells: Regulation by IL-1α and TGFß and Implications for Migration of Pancreatic Cancer Cells.

Pancreatic ductal adenocarcinoma is characterized by a prominent fibroinflammatory stroma with both tumor-promoting and tumor-suppressive functions. The pancreatic stellate cell (PSC) is the major cellular stromal component and the main producer of extracellular matrix proteins, including collagens, which are degraded by metalloproteinases (MMPs). PSCs interact with cancer cells through various factors, including transforming growth factor (TGF)ß and interleukin (IL)-1α. The role of TGFß in the dual nature of tumor stroma, i.e., protumorigenic or tumor suppressive, is not clear. We aimed to investigate the roles of TGFß and IL-1α in the regulation of MMP profiles in PSCs and the subsequent effects on cancer cell migration. Human PSCs isolated from surgically resected specimens were cultured in the presence of pancreatic cancer cell lines, as well as IL-1α or TGFß. MMP production and activities in PSCs were quantified by gene array transcripts, mRNA measurements, fluorescence resonance energy transfer-based activity assay, and zymography. PSC-conditioned media and pancreatic cancer cells were included in a collagen matrix cell migration model. We found that production of IL-1α by pancreatic cancer cells induced alterations in MMP and tissue inhibitors of matrix metalloproteinase (TIMP) profiles and activities in PSCs, upregulated expression and activation of MMP1 and MMP3, and enhanced migration of pancreatic cancer cells in the collagen matrix model. TGFß counteracted the effects of IL-1α on PSCs, reestablished PSC MMP and TIMP profiles and activities, and inhibited migration of cancer cells. This suggests that tumor TGFß has a role as a suppressor of stromal promotion of tumor progression through alterations in PSC MMP profiles with subsequent inhibition of pancreatic cancer cell migration.

Pathology and Surgical Treatment of High-Grade Pancreatic Neuroendocrine Carcinoma: an Evolving Landscape.

Pancreatic neuroendocrine neoplasms (PNENs) are rare, accounting for less than 5% of all pancreatic tumors. High-grade pancreatic neuroendocrine carcinomas (hgPNECs) represent about 5% of all PNENs. They show highly aggressive behavior with dismal prognosis. Throughout the last two decades, there has been a notable progress in basic and clinical research of PNENs and a therapeutic trend towards both more aggressive and minimally invasive surgery. Despite these advances, hgPNECs as a distinct clinical entity remains largely unexplored among surgeons. This review of current development in pathology reporting and surgical treatment of hgPNECs aims at increasing the awareness of an evolving field in pancreatic surgery.

Tumour growth is more dispersed in pancreatic head cancers than in rectal cancer: implications for resection margin assessment.

AIMS: The UK definition of microscopic resection margin involvement (R1) in pancreatic head cancer, based on tumour lying <1 mm from the margin, has been adopted from rectal cancer, but has never been validated. The aim of this study was to assess the adequacy of the R1 definition for pancreatic head cancers by comparing the growth patterns of rectal (RC), pancreatic (PC), ampullary (AC) and distal bile duct (DBC) adenocarcinomas. METHODS AND RESULTS: Distances between tumour cells and tumour cell density in the tumour centre and periphery were quantified by Minimum Spanning Tree (MST) analysis in 10 cases of the four cancer groups. In RC, the MST distance was similar throughout the entire width of the tumour, whereas in PC, DBC and AC it was significantly larger at the periphery than at the tumour centre (P ≤ 0.003). While results were similar for PC and DBC, however, distances at the centre and periphery of both cancers were larger compared to AC (P ≤ 0.046). Tumour cell density dropped at the periphery of PC to 31% of that at the centre, compared to 83% in RC (P < 0.0002). CONCLUSIONS: Tumour growth in pancreatic head cancers is more dispersed than in RC, particularly in the tumour periphery. Revision of the R1 definition for pancreatic head cancer may therefore need to be considered.