Spatial heterogeneity of immune regulators drives dynamic changes of local immune responses, affecting disease outcomes in pancreatic cancer.

PURPOSE: Pancreatic ductal adenocarcinoma (PDAC) is considered a low immunogenic tumor with "cold" tumor microenvironment (TME) and is mostly unresponsive to immune checkpoint blockade therapies. Here we decipher the impact of intratumoral heterogeneity of immune determinants on antitumor response. EXPERIMENTAL DESIGN: We performed spatial proteomic and transcriptomic analyses and multiplexed immunofluorescence on multiple tumor regions, including tumor center (TC) and invasive front (IF), from 220 PDAC-patients, classified according to their transcriptomic immune signaling into high-immunogenic (HI-PDACs, n=54) and low-immunogenic tumors (LI-PDACs, n=166). Spatial compartments (tumor: Pancytokeratin+/CD45- and leukocytes: Pancytokeratin-/CD45+) were defined by fluorescent imaging. RESULTS: HI-PDACs exhibited higher densities of cytotoxic T lymphocytes with upregulation of T-cell priming-associated immune determinants, including CD40, ITGAM, GITR, CXCL10, GZMB, IFNG and HLA-DR, which was significantly more prominent at the IF than the TC. In contrast, LI-PDACs exhibited immune evasive TMEs with downregulation of immune determinants and a negative gradient from TC to IF. Patients with HI-PDACs had significantly better outcomes; however, they showed more frequently exhausted immune phenotypes. CONCLUSIONS: Our results indicate strategic differences in the regulation of immune determinants, which lead to different levels of effectiveness of antitumor responses between high- and low-immunogenic tumors and dynamic spatial changes, which affect the evolution of immune evasion and patient outcomes. This supports coevolution of tumor and immune cells and may help define therapeutic vulnerabilities to improve antitumor immunity and harness the responsiveness to immune checkpoint inhibitors in PDAC patients.

Resection margin status at the portomesenteric axis may not determine oncologic outcome after pancreaticoduodenectomy for lymph node-positive pancreatic ductal adenocarcinoma.

BACKGROUND: Lymph node and resection margin status are associated with oncologic outcomes after pancreaticoduodenectomy for pancreatic ductal adenocarcinoma. However, surgical radicality at the portomesenteric axis in case of suspected infiltration remains controversial. METHODS: Clinicopathological data of patients who underwent a partial or total pancreaticoduodenectomy for PDAC between 2012 to 2019 in 2 major hepato-pancreato-biliary centers in Germany and Switzerland were assessed. We evaluated the impact of positive resection margins at the vascular, parenchymal, and retropancreatic surfaces on overall survival in patients with and without lymph node involvement. Margin-positive vascular resection included both patients with positive margins at the vascular groove and the resected venous wall. RESULTS: During the study period, 217 patients underwent partial/total pancreaticoduodenectomy for pancreatic ductal adenocarcinoma. After excluding 7 patients suffering postoperative complications resulting in mortality within 90 days after surgery (3%), 169 patients had lymph node involvement (80%). In the entire study cohort, margin-positive resection (33%) was significantly associated with worse overall survival (3-year overall survival: margin-positive resection: 27% vs margin-negative resection: 43%, P = .014). Among patients with positive lymph nodes, margin-positive vascular resection (n = 48, 28%) was not significantly associated with impaired overall survival (3-year overall survival: margin-positive vascular resection: 28% vs margin-negative vascular resection: 36%, P = .065). On the contrary, margin-positive parenchymal resection (n = 7, 4%) (3-year overall survival: margin-positive parenchymal resection: 0% vs margin-negative parenchymal resection: 35%, P < .0001) and margin-positive retropancreatic resection (n = 21, 12%) (3-year overall survival: margin-positive retropancreatic resection: 6% vs margin-negative retropancreatic resection: 39%, P < .0001) significantly diminished overall survival in univariate and multivariate analysis in all patients. Among patients without lymph node involvement (n = 41, 20%), there were no margin-positive parenchymal or margin-positive retropancreatic resections. In contrast, only 5 patients had margin-positive vascular resection (12%), with overall survival compared to those with margin-negative vascular resection. CONCLUSIONS: In patients with pancreatic ductal adenocarcinoma and lymph nodal positivity, resection status at the parenchymal and retropancreatic surface but probably not at the portal and/or superior mesenteric vein is a determinant of survival. Therefore, margin-negative resection should be pursued during pancreaticoduodenectomy. However, radical venous resection and/or reconstruction for suspected tumor infiltration may not be necessary for patients with intraoperatively detected lymph node metastases.

Spatially restricted tumour-associated and host-associated immune drivers correlate with the recurrence sites of pancreatic cancer.

OBJECTIVE: Most patients with pancreatic ductal adenocarcinoma (PDAC) will experience recurrence after resection. Here, we investigate spatially organised immune determinants of PDAC recurrence. DESIGN: PDACs (n=284; discovery cohort) were classified according to recurrence site as liver (n=93/33%), lung (n=49/17%), local (n=31/11%), peritoneal (n=38/13%) and no-recurrence (n=73/26%). Spatial compartments were identified by fluorescent imaging as: pancytokeratin (PanCK)+CD45- (tumour cells); CD45+PanCK- (leucocytes) and PanCK-CD45- (stromal cells), followed by transcriptomic (72 genes) and proteomic analysis (51 proteins) for immune pathway targets. Results from next-generation sequencing (n=194) were integrated. Finally, 10 tumours from each group underwent immunophenotypic analysis by multiplex immunofluorescence. A validation cohort (n=109) was examined in parallel. RESULTS: No-recurrent PDACs show high immunogenicity, adaptive immune responses and are rich in pro-inflammatory chemokines, granzyme B and alpha-smooth muscle actin+ fibroblasts. PDACs with liver and/or peritoneal recurrences display low immunogenicity, stemness phenotype and innate immune responses, whereas those with peritoneal metastases are additionally rich in FAP+ fibroblasts. PDACs with local and/or lung recurrences display interferon-gamma signalling and mixed adaptive and innate immune responses, but with different leading immune cell population. Tumours with local recurrences overexpress dendritic cell markers whereas those with lung recurrences neutrophilic markers. Except the exclusive presence of RNF43 mutations in the no-recurrence group, no genetic differences were seen. The no-recurrence group exhibited the best, whereas liver and peritoneal recurrences the poorest prognosis. CONCLUSIONS: Our findings demonstrate distinct inflammatory/stromal responses in each recurrence group, which might affect dissemination patterns and patient outcomes. These findings may help to inform personalised adjuvant/neoadjuvant and surveillance strategies in PDAC, including immunotherapeutic modalities.

High tumor mutational burden (TMB) identifies a microsatellite stable pancreatic cancer subset with prolonged survival and strong anti-tumor immunity.

AIM: Tumor mutational burden (TMB: somatic mutations per megabase, mut/Mb) predicts the efficacy of immunotherapy. Here, we link TMB levels with the activation of immune pathways and intratumoral immune responses in pancreatic adenocarcinoma (PDAC) to explore immunoarchitectural patterns associated with high TMB. METHODS: We assessed TMB in 161 resected, microsatellite stable (MSS) PDACs, including 41 long-term survivors (LTS). Five microsatellite instable (MSI-high) cases were also assessed. Cases were classified into TMB-high (≥10 mut/Mb), TMB-intermediate (>5 < 10 mut/Mb), and TMB-low (≤5 mut/Mb) categories. Tumors additionally underwent mRNA in situ hybridization for immune pathway genes and were immunoprofiled by multiplex immunofluorescence followed by automated image analysis. RESULTS: We detected 12 TMB-high, 28 TMB-intermediate, and 121 TMB-low cases. TMB-high tumors comprised ten LTSs (10/41; 24%) and two conventional PDACs (2/120; 1.7%). They exhibited the highest T cell density with significantly increased CD3+CD4+T helper and CD208+dendritic cell (DC) counts, compared to all other cases. CD3+CD8+cytotoxic T cells were significantly closer to tumor cells and T helper cells closer to DCs in TMB-high PDACs. Immune pathways involved in T cell activation, immune cell adhesion/migration, antigen presentation, and cytokine signaling were upregulated in most TMB-high and many TMB-intermediate tumors. ARID1A and ERBB4 alterations were more frequent in TMB-high PDACs. All MSI-high PDACs were TMB-high. CONCLUSIONS: TMB-high cases frequently belong to specific PDAC subsets with prolonged survival such as LTSs and MSI-high PDACs. They display strong anti-tumor immune responses fueled by a T helper cell/DC-mediated priming of the cytotoxic T cells. Moreover, they frequently harbor further actionable alterations.

Preoperative Glucagon-like peptide-1 receptor imaging reduces surgical trauma and pancreatic tissue loss in insulinoma patients: a report of three cases.

BACKGROUND: Insulinomas are rare tumors, in the majority of cases best treated by surgical resection. Preoperative localization of insulinoma is challenging. The more precise the preoperative localization the less invasive and safer is the resection. The purpose of the study is to check the impact of a new technique to localize insulinoma on the surgical strategy. FINDINGS: We present exact preoperative localization with Glucagon-like peptide-1 receptor (GLP-1R) imaging. This allows a more precise resection thereby reducing surgical access trauma, loss of healthy pancreatic tissue and increasing safety and quality of the surgical intervention. CONCLUSION: With the help of precise preoperative localization of insulinoma with GLP-1R imaging the surgeon is able to minimize the amount of resected healthy pancreatic tissue. We hypothesize that GLP-1R imaging will become a preoperative diagnostic tool to be used for many patients scheduled for open or laparoscopic insulinoma resection.

How do we apply video-assisted retroperitoneal necrosectomy with minimal access?

BACKGROUND: The conservative treatment of acute necrotizing pancreatitis has greatly improved due to broad antibiotic treatment and improved organ support in intensive care units. Nevertheless, infected necrosis or persistent multi-organ dysfunction are predictors of poor outcome. In these patients, there is still a need to perform necrosectomy. Open surgery results in extensive operative trauma and is associated with high morbidity and mortality. Therefore, several minimally invasive techniques have been developed recently. Retroperitoneal necrosectomy has been shown to be safe and to reduce morbidity and mortality compared to the open procedure. METHODS AND RESULTS: In an instructive video, we show the technique of video-assisted retroperitoneal necrosectomy with minimal access, including the preoperative percutaneous drainage and several accesses to the necrosis. We discuss the indication for retroperitoneal necrosectomy as well as the optimal time point of the intervention. CONCLUSION: In the management of acute necrotizing pancreatitis, the multidisciplinary approach is crucial. The initial treatment by the intensive care units should be extended to intervention or surgery in case of infected necrosis or persistent multi-organ dysfunction. We show here a minimal access solution with the placement of a percutaneous drain followed by video-assisted retroperitoneal necrosectomy.