Expression of the membrane tetraspanin claudin 18 on cancer cells promotes T lymphocyte infiltration and antitumor immunity in pancreatic cancer.

Tumors weakly infiltrated by T lymphocytes poorly respond to immunotherapy. We aimed to unveil malignancy-associated programs regulating T cell entrance, arrest, and activation in the tumor environment. Differential expression of cell adhesion and tissue architecture programs, particularly the presence of the membrane tetraspanin claudin (CLDN)18 as a signature gene, demarcated immune-infiltrated from immune-depleted mouse pancreatic tumors. In human pancreatic ductal adenocarcinoma (PDAC) and non-small cell lung cancer, CLDN18 expression positively correlated with more differentiated histology and favorable prognosis. CLDN18 on the cell surface promoted accrual of cytotoxic T lymphocytes (CTLs), facilitating direct CTL contacts with tumor cells by driving the mobilization of the adhesion protein ALCAM to the lipid rafts of the tumor cell membrane through actin. This process favored the formation of robust immunological synapses (ISs) between CTLs and CLDN18-positive cancer cells, resulting in increased T cell activation. Our data reveal an immune role for CLDN18 in orchestrating T cell infiltration and shaping the tumor immune contexture.

CLDN6-specific CAR-T cells plus amplifying RNA vaccine in relapsed or refractory solid tumors: the phase 1 BNT211-01 trial.

The oncofetal antigen Claudin 6 (CLDN6) is highly and specifically expressed in many solid tumors, and could be a promising treatment target. We report dose escalation results from the ongoing phase 1/2 BNT211-01 trial evaluating the safety and feasibility of chimeric antigen receptor (CAR) T cells targeting the CLDN6 with or without a CAR-T cell-amplifying RNA vaccine (CARVac) at two dose levels (DLs) in relapsed/refractory CLDN6-positive solid tumors. The primary endpoints were safety and tolerability, maximum tolerated dose and recommended phase 2 dose (RP2D). Secondary endpoints included objective response rate (ORR) and disease control rate. We observed manageable toxicity, with 10 out of 22 patients (46%) experiencing cytokine release syndrome including one grade 3 event and 1 out of 22 (5%) with grade 1 immune effector cell-associated neurotoxicity syndrome. Dose-limiting toxicities occurred in two patients at the higher DL, resolving without sequelae. CAR-T cell engraftment was robust, and the addition of CARVac was well tolerated. The unconfirmed ORR in 21 evaluable patients was 33% (7 of 21), including one complete response. The disease control rate was 67% (14 of 21), with stable disease in seven patients. Patients with germ cell tumors treated at the higher DL exhibited the highest response rate (ORR 57% (4 of 7)). The maximum tolerated dose and RP2D were not established as the trial has been amended to utilize an automated manufacturing process. A repeat of the dose escalation is ongoing and will identify a RP2D for pivotal trials. ClinicalTrials.gov Identifier: NCT04503278 .

GATA4 and GATA6 loss-of-expression is associated with extinction of the classical programme and poor outcome in pancreatic ductal adenocarcinoma.

OBJECTIVE: GATA6 is a key regulator of the classical phenotype in pancreatic ductal adenocarcinoma (PDAC). Low GATA6 expression associates with poor patient outcome. GATA4 is the second most expressed GATA factor in the pancreas. We assessed whether, and how, GATA4 contributes to PDAC phenotype and analysed the association of expression with outcome and response to chemotherapy. DESIGN: We analysed PDAC transcriptomic data, stratifying cases according to GATA4 and GATA6 expression and identified differentially expressed genes and pathways. The genome-wide distribution of GATA4 was assessed, as well as the effects of GATA4 knockdown. A multicentre tissue microarray study to assess GATA4 and GATA6 expression in samples (n=745) from patients with resectable was performed. GATA4 and GATA6 levels were dichotomised into high/low categorical variables; association with outcome was assessed using univariable and multivariable Cox regression models. RESULTS: GATA4 messenger RNA is enriched in classical, compared with basal-like tumours. We classified samples in 4 groups as high/low for GATA4 and GATA6. Reduced expression of GATA4 had a minor transcriptional impact but low expression of GATA4 enhanced the effects of GATA6 low expression. GATA4 and GATA6 display a partially overlapping genome-wide distribution, mainly at promoters. Reduced expression of both proteins in tumours was associated with the worst patient survival. GATA4 and GATA6 expression significantly decreased in metastases and negatively correlated with basal markers. CONCLUSIONS: GATA4 and GATA6 cooperate to maintain the classical phenotype. Our findings provide compelling rationale to assess their expression as biomarkers of poor prognosis and therapeutic response.

Molecular heterogeneity and commonalities in pancreatic cancer precursors with gastric and intestinal phenotype.

OBJECTIVE: Due to the limited number of modifiable risk factors, secondary prevention strategies based on early diagnosis represent the preferred route to improve the prognosis of pancreatic ductal adenocarcinoma (PDAC). Here, we provide a comparative morphogenetic analysis of PDAC precursors aiming at dissecting the process of carcinogenesis and tackling the heterogeneity of preinvasive lesions. DESIGN: Targeted and whole-genome low-coverage sequencing, genome-wide methylation and transcriptome analyses were applied on a final collective of 122 morphologically well-characterised low-grade and high-grade PDAC precursors, including intestinal and gastric intraductal papillary mucinous neoplasms (IPMN) and pancreatic intraepithelial neoplasias (PanIN). RESULTS: Epigenetic regulation of mucin genes determines the phenotype of PDAC precursors. PanIN and gastric IPMN display a ductal molecular profile and numerous similarly regulated pathways, including the Notch pathway, but can be distinguished by recurrent deletions and differential methylation and, in part, by the expression of mucin-like 3. Intestinal IPMN are clearly distinct lesions at the molecular level with a more instable genotype and are possibly related to a different ductal cell compartment. CONCLUSIONS: PDAC precursors with gastric and intestinal phenotype are heterogeneous in terms of morphology, genetic and epigenetic profile. This heterogeneity is related to a different cell identity and, possibly, to a different aetiology.

Pathological Reporting of Radical Prostatectomy Specimens Following ICCR Recommendation: Impact of Electronic Reporting Tool Implementation on Quality and Interdisciplinary Communication in a Large University Hospital.

Prostate cancer represents one of the most common malignant tumors in male patients in Germany. The pathological reporting of radical prostatectomy specimens following a structured process constitutes an excellent prototype for the introduction of software-based standardized structured reporting in pathology. This can lead to reports of higher quality and could create a fundamental improvement for future AI applications. A software-based reporting template was used to generate standardized structured pathological reports of radical prostatectomy specimens of patients treated at the University Hospital Klinikum rechts der Isar of Technische Universität München, Germany. Narrative reports (NR) and standardized structured reports (SSR) were analyzed with regard to completeness, and clinicians' satisfaction with each report type was evaluated. SSR show considerably higher completeness than NR. A total of 10 categories out of 32 were significantly more complete in SSR than in NR (p < 0.05). Clinicians awarded overall high scores in NR and SSR reports. One rater acknowledged a significantly higher level of clarity and time saving when comparing SSR to NR. Our findings highlight that the standardized structured reporting of radical prostatectomy specimens, qualifying as level 5 reports, significantly increases objectively measured content quality and the level of completeness. The implementation of nationwide SSR in Germany, particularly in oncologic pathology, can serve pathologists, clinicians, and patients.

MALDI-MSI: A Powerful Approach to Understand Primary Pancreatic Ductal Adenocarcinoma and Metastases.

Cancer-related deaths are very commonly attributed to complications from metastases to neighboring as well as distant organs. Dissociate response in the treatment of pancreatic adenocarcinoma is one of the main causes of low treatment success and low survival rates. This behavior could not be explained by transcriptomics or genomics; however, differences in the composition at the protein level could be observed. We have characterized the proteomic composition of primary pancreatic adenocarcinoma and distant metastasis directly in human tissue samples, utilizing mass spectrometry imaging. The mass spectrometry data was used to train and validate machine learning models that could distinguish both tissue entities with an accuracy above 90%. Model validation on samples from another collection yielded a correct classification of both entities. Tentative identification of the discriminative molecular features showed that collagen fragments (COL1A1, COL1A2, and COL3A1) play a fundamental role in tumor development. From the analysis of the receiver operating characteristic, we could further advance some potential targets, such as histone and histone variations, that could provide a better understanding of tumor development, and consequently, more effective treatments.

A Modified Burn Comb Model With a New Dorsal Frame That Allows for Local Treatment in Partial-Thickness Burns in Rats.

Burn wound progression (BWP) leads to vertical and horizontal injury extension. The "burn comb model" is commonly used, in which a full-thickness burn with intercalated unburned interspaces is induced. We aimed to establish an injury progressing to the intermediate dermis, allowing repeated wound evaluation. Furthermore, we present a new dorsal frame that enables topical drug application. Eight burn fields and six interspaces were induced on each of 17 rats' dorsa with a 10-second burn comb application. A developed 8-panel aluminum frame was sutured onto 12 animals and combined with an Elizabethan collar. Over 14 days, macroscopic and histologic wound assessment and laser speckle contrast imaging (LSCI) were performed besides evaluation of frame durability. The 10-second group was compared with nine animals injured with a full-thickness 60-second model. Frame durability was sufficient up to day 4 with 8 of the 12 frames (67%) still mounted. The 60-second burn led to an increased extent of interspace necrosis (P = .002). The extent of necrosis increased between days 1 and 2 (P = .001), following the 10-second burn (24% ± SEM 8% to 40% ± SEM 6%) and the 60-second burn (57% ± SEM 6% to 76% ± SEM 4%). Interspace LSCI perfusion was higher than burn field perfusion. It earlier reached baseline levels in the 10-second group (on day 1: 142% ± SEM 9% vs 60% ± SEM 5%; P < .001). Within day 1, the 10-second burn showed histological progression to the intermediate dermis, both in interspaces and burn fields. This burn comb model with its newly developed fixed dorsal frame allows investigation of topical agents to treat BWP in partial-thickness burns.

Integrative analysis reveals early and distinct genetic and epigenetic changes in intraductal papillary and tubulopapillary cholangiocarcinogenesis.

OBJECTIVE: A detailed understanding of the molecular alterations in different forms of cholangiocarcinogenesis is crucial for a better understanding of cholangiocarcinoma (CCA) and may pave the way to early diagnosis and better treatment options. DESIGN: We analysed a clinicopathologically well-characterised patient cohort (n=54) with high-grade intraductal papillary (IPNB) or tubulopapillary (ITPN) neoplastic precursor lesions of the biliary tract and correlated the results with an independent non-IPNB/ITPN associated CCA cohort (n=294). The triplet sample set of non-neoplastic biliary epithelium, precursor and invasive CCA was analysed by next generation sequencing, DNA copy number and genome-wide methylation profiling. RESULTS: Patients with invasive CCA arising from IPNB/ITPN had better prognosis than patients with CCA not associated with IPNB/ITPN. ITPN was localised mostly intrahepatic, whereas IPNB was mostly of extrahepatic origin. IPNB/ITPN were equally associated with small-duct and large-duct type intrahepatic CCA. IPNB exhibited mutational profiles of extrahepatic CCA, while ITPN had significantly fewer mutations. Most mutations were shared between precursor lesions and corresponding invasive CCA but ROBO2 mutations occurred exclusively in invasive CCA and CTNNB1 mutations were mainly present in precursor lesions. In addition, IPNB and ITPN differed in their DNA methylation profiles and analyses of latent methylation components suggested that IPNB and ITPN may have different cells-of-origin. CONCLUSION: Integrative analysis revealed that IPNB and ITPN harbour distinct early genetic alterations, IPNB are enriched in mutations typical for extrahepatic CCA, whereas ITPN exhibited few genetic alterations and showed distinct epigenetic profiles. In conclusion, IPNB/ITPN may represent a distinctive, intermediate form of intrahepatic and extrahepatic cholangiocarcinogenesis.

An analysis of 130 neuroendocrine tumors G3 regarding prevalence, origin, metastasis, and diagnostic features.

Limited data exist on high-grade neuroendocrine tumors (NETs G3) which represent a new category among neuroendocrine neoplasms (NEN). We analyzed NETs G3 in a consultation series regarding prevalence, origin, metastasis, and diagnostic problems. Based on the WHO classification of digestive system tumors, 130 NETs G3 (9%) were identified in 1513 NENs. NET G3 samples were more often obtained from metastatic sites (69%) than NET G1/G2 samples (24%). NET G3 metastases presented most frequently in the liver (74%) and originated from the pancreas (38/90, 42%), followed by the lung (9%), ileum (7%), stomach (3%), rectum (1%), and rare sites (2%) such as the prostate and breast. The primaries remained unknown in 15%. NETs G3 had a median Ki67 of 30% that distinguished them from NECs (60%), though with great overlap. The expression of site-specific markers, p53, Rb1, and SST2 was similar in NETs G3 and NETs G1/G2, except for p53 and Rb1 which were abnormally expressed in 8% and 7% of liver metastases from NET G3 but not from NET G1/G2. NETs G3 were frequently referred as NECs (39%) but could be well distinguished from NECs by normal p53 (92% versus 21%) and Rb1 expression (93% versus 41%) expression. In conclusion, NETs G3 are frequently discovered as liver metastases from pancreatic or pulmonary primaries and are often misinterpreted as NEC. p53 and Rb1 are powerful markers in the distinction of NET G3 from NEC. Rarely, carcinomas from non-digestive, non-pulmonary organs with neuroendocrine features may present as NET G3.

Native glycan fragments detected by MALDI mass spectrometry imaging are independent prognostic factors in pancreatic ductal adenocarcinoma.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) remains one of the deadliest malignancies to date. The impressively developed stroma that surrounds and modulates the behavior of cancer cells is one of the main factors regulating the PDAC growth, metastasis and therapy resistance. Here, we postulate that stromal and cancer cell compartments differentiate in protein/lipid glycosylation patterns and analyze differences in glycan fragments in those compartments with clinicopathologic correlates. RESULTS: We analyzed native glycan fragments in 109 human FFPE PDAC samples using high mass resolution matrix-assisted laser desorption/ionization Fourier-transform ion cyclotron resonance mass spectrometric imaging (MALDI-FT-ICR-MSI). Our method allows detection of native glycan fragments without previous digestion with PNGase or any other biochemical reaction. With this method, 8 and 18 native glycans were identified as uniquely expressed in only stromal or only cancer cell compartment, respectively. Kaplan-Meier survival model identified glycan fragments that are expressed in cancer cell or stromal compartment and significantly associated with patient outcome. Among cancer cell region-specific glycans, 10 predicted better and 6 worse patient survival. In the stroma, 1 glycan predicted good and 4 poor patient survival. Using factor analysis as a dimension reduction method, we were able to group the identified glycans in 2 factors. Multivariate analysis revealed that these factors can be used as independent survival prognostic elements with regard to the established Union for International Cancer Control (UICC) classification both in tumor and stroma regions. CONCLUSION: Our method allows in situ detection of naturally occurring glycans in FFPE samples of human PDAC tissue and highlights the differences among glycans found in stromal and cancer cell compartment offering a basis for further exploration on the role of specific glycans in cancer-stroma communication.

The Impact of Histological Annotations for Accurate Tissue Classification Using Mass Spectrometry Imaging.

Knowing the precise location of analytes in the tissue has the potential to provide information about the organs' function and predict its behavior. It is especially powerful when used in diagnosis and prognosis prediction of pathologies, such as cancer. Spatial proteomics, in particular mass spectrometry imaging, together with machine learning approaches, has been proven to be a very helpful tool in answering some histopathology conundrums. To gain accurate information about the tissue, there is a need to build robust classification models. We have investigated the impact of histological annotation on the classification accuracy of different tumor tissues. Intrinsic tissue heterogeneity directly impacts the efficacy of the annotations, having a more pronounced effect on more heterogeneous tissues, as pancreatic ductal adenocarcinoma, where the impact is over 20% in accuracy. On the other hand, in more homogeneous samples, such as kidney tumors, histological annotations have a slenderer impact on the classification accuracy.

Loss of CDX2 in colorectal cancer is associated with histopathologic subtypes and microsatellite instability but is prognostically inferior to hematoxylin-eosin-based morphologic parameters from the WHO classification.

BACKGROUND: Immunohistochemical loss of CDX2 has been proposed as a biomarker of dismal survival in colorectal carcinoma (CRC), especially in UICC Stage II/III. However, it remains unclear, how CDX2 expression is related to central hematoxylin-eosin (HE)-based morphologic parameters defined by 2019 WHO classification and how its prognostic relevance is compared to these parameters. METHODS: We evaluated CDX2 expression in 1003 CRCs and explored its prognostic relevance compared to CRC subtypes, tumour budding and WHO grade in the overall cohort and in specific subgroups. RESULTS: CDX2-low/absent CRCs were enriched in specific morphologic subtypes, right-sided and microsatellite-instable (MSI-H) CRCs (P < 0.001) and showed worse survival characteristics in the overall cohort/UICC Stage II/III (e.g. DFS: P = 0.005) and in microsatellite stable and left-sided CRCs, but not in MSI-H or right-sided CRCs. Compared with CDX2, all HE-based markers showed a significantly better prognostic discrimination in all scenarios. In multivariate analyses including all morphologic parameters, CDX2 was not an independent prognostic factor. CONCLUSION: CDX2 loss has some prognostic impact in univariate analyses, but its prognostic relevance is considerably lower compared to central HE-based morphologic parameters defined by the WHO classification and vanishes in multivariate analyses incorporating these factors.

[Ductal adenocarcinoma of the pancreas: subtypes and molecular pathology]. / Das duktale Adenokarzinom des Pankreas: Subtypen und Molekularpathologie.

Ductal adenocarcinoma is the most common tumor of the pancreas. Although relatively rare, it poses one of the greatest oncological challenges because of its poor prognosis, which has so far only slightly improved. Progress has been made in the more precise classification and standardization of the morphological assessment. In the current WHO classification, prognostically relevant subtypes are clearly delimited among themselves and from ductal adenocarcinoma not otherwise specified (NOS). In the recent TNM classification, a size-based T­category was introduced. Diagnostically, the histological assessment of the resection specimen is relatively easy; on the other hand, assessment of the fine-needle biopsy from the primary tumor or a liver metastasis is still difficult. The molecular stratification of ductal adenocarcinoma and the other pancreatic neoplasms has made great progress. This not only defined the genetics of tumor entities, but also identified the prognosis and biology of tumor groups on the basis of RNA expression patterns. The range of treatment could be expanded by targeted molecular therapies (especially for patients with BRCA1/2 germline mutations, NTRK- or NRG1-fusions, or oncogenic BRAF and PIK3CA mutations as well as tumors with microsatellite instability (MSI)), even if targeted therapies are currently only available for a minority of patients (<10%).

Mesenchymal/non-epithelial mimickers of neuroendocrine neoplasms with a focus on fusion gene-associated and SWI/SNF-deficient tumors.

Mimickers of neuroendocrine neoplasms (NEN) include a number of important pitfall tumors. Here, we describe our experience with mesenchymal mimics of NENs to illustrate their spectrum and draw the attention particularly to a group of mesenchymal/non-epithelial neoplasms (MN) that combine epithelioid histology with neuroendocrine (NE-) features and peculiar genetic abnormalities. In a consultation series of 4498 cases collected between 2009 and 2021, 2099 neoplasms expressing synaptophysin and/or chromograninA were reviewed and analyzed. A total of 364 (18%) were diagnosed as non-NENs, while the remaining tumors were NEN. The group of mesenchymal/non-epithelial neoplasms with NE-features (MN-NE) included 31/364 (8%) cases. These mostly malignant neoplasms showed an epithelioid morphology. While all but one tumor expressed synaptophysin, mostly patchy, only 10/29 (34%) co-expressed chromograninA. A total of 13/31 (42%) of the MN-NE showed EWSR1-related gene fusions (6 Ewing sarcomas, 5 clear cell sarcomas, and 1 desmoplastic small round cell tumor, 1 neoplasm with FUS-CREM gene fusion) and 7 (23%) were SWI/SNF (SMARCB1 or SMARCA4)-deficient neoplasms. The remaining MN-NE included synovial sarcoma, sclerosing epithelioid mesenchymal neoplasm, melanoma, alveolar soft part sarcoma, solitary fibrous tumor, and chordoma. A total of 27/31 MN-NE were from the last 8 years, and 6 of them were located in the pancreas. Eleven MN-NE were initially diagnosed as neuroendocrine carcinomas (NECs). MN-NE with epithelioid features play an increasing role as mimickers of NECs. They mostly belong to tumors with gene fusions involving the EWSR1 gene, or with SWI/SNF complex deficiency. Synaptophysin expression is mostly patchy and chromograninA expression is infrequent in MN-NE of this series and data extracted from literature.

[Standardized diagnosis of pancreatic head carcinoma]. / Standardisierte Befundung des Pankreaskopfkarzinoms.

Most pancreatic ductal adenocarcinomas are localized in the pancreatic head. Due to the complex anatomic relationships with the surrounding organs and vascular structures in the retroperitoneal space and to the presence of numerous transection margins and dissection planes, pancreatic head resections belong to the most complex specimens concerning grossing and sampling for histopathologic analysis.Here we discuss current guidelines for standardized grossing and reporting of pancreatic cancer, with special reference to the assessment of the resection margin status. The importance of standardized reporting for the sake of completeness, comprehensibility, comparability, and quality control as well as for the integration of pathology reports in interdisciplinary digital workflows and artificial intelligence applications will be emphasized.

Whole Exome Sequencing of Biliary Tubulopapillary Neoplasms Reveals Common Mutations in Chromatin Remodeling Genes.

The molecular carcinogenesis of intraductal tubulopapillary neoplasms (ITPN), recently described as rare neoplasms in the pancreato-biliary tract with a favorable prognosis despite a high incidence of associated pancreato-biliary adenocarcinoma, is still poorly understood. To identify driver genes, chromosomal gains and losses, mutational signatures, key signaling pathways, and potential therapeutic targets, the molecular profile of 11 biliary and 6 pancreatic ITPNs, associated with invasive adenocarcinoma in 14/17 cases, are studied by whole exome sequencing (WES). The WES of 17 ITPNs reveals common copy number variants (CNVs) broadly distributed across the genome, with recurrent chromosomal deletions primarily in 1p36 and 9p21 affecting the tumor suppressors CHD5 and CDKN2A, respectively, and gains in 1q affecting the prominent oncogene AKT3. The identified somatic nucleotide variants (SNVs) involve few core signaling pathways despite high genetic heterogeneity with diverse mutational spectra: Chromatin remodeling, the cell cycle, and DNA damage/repair. An OncoKB search identifies putative actionable genomic targets in 35% of the cases (6/17), including recurrent missense mutations of the FGFR2 gene in biliary ITPNs (2/11, 18%). Our results show that somatic SNV in classical cancer genes, typically associated with pancreato-biliary carcinogenesis, were absent (KRAS, IDH1/2, GNAS, and others) to rare (TP53 and SMAD4, 6%, respectively) in ITPNs. Mutational signature pattern analysis reveals a predominance of an age-related pattern. Our findings highlight that biliary ITPN and classical cholangiocarcinoma display commonalities, in particular mutations in genes of the chromatin remodeling pathway, and appear, therefore, more closely related than pancreatic ITPN and classical pancreatic ductal adenocarcinoma.

Bcl3 Couples Cancer Stem Cell Enrichment With Pancreatic Cancer Molecular Subtypes.

BACKGROUND & AIMS: The existence of different subtypes of pancreatic ductal adenocarcinoma (PDAC) and their correlation with patient outcome have shifted the emphasis on patient classification for better decision-making algorithms and personalized therapy. The contribution of mechanisms regulating the cancer stem cell (CSC) population in different subtypes remains unknown. METHODS: Using RNA-seq, we identified B-cell CLL/lymphoma 3 (BCL3), an atypical nf-κb signaling member, as differing in pancreatic CSCs. To determine the biological consequences of BCL3 silencing in vivo and in vitro, we generated bcl3-deficient preclinical mouse models as well as murine cell lines and correlated our findings with human cell lines, PDX models, and 2 independent patient cohorts. We assessed the correlation of bcl3 expression pattern with clinical parameters and subtypes. RESULTS: Bcl3 was significantly down-regulated in human CSCs. Recapitulating this phenotype in preclinical mouse models of PDAC via BCL3 genetic knockout enhanced tumor burden, metastasis, epithelial to mesenchymal transition, and reduced overall survival. Fluorescence-activated cell sorting analyses, together with oxygen consumption, sphere formation, and tumorigenicity assays, all indicated that BCL3 loss resulted in CSC compartment expansion promoting cellular dedifferentiation. Overexpression of BCL3 in human PDXs diminished tumor growth by significantly reducing the CSC population and promoting differentiation. Human PDACs with low BCL3 expression correlated with increased metastasis, and BCL3-negative tumors correlated with lower survival and nonclassical subtypes. CONCLUSIONS: We demonstrate that bcl3 impacts pancreatic carcinogenesis by restraining CSC expansion and by curtailing an aggressive and metastatic tumor burden in PDAC across species. Levels of BCL3 expression are a useful stratification marker for predicting subtype characterization in PDAC, thereby allowing for personalized therapeutic approaches.

Stroma composition and proliferative activity are related to therapy response in neoadjuvant treated pancreatic ductal adenocarcinoma.

BACKGROUND: Tumor regression grading (TRG) based on histopathology is the main tool to assess therapy effects after neoadjuvant therapy (NAT) of pancreatic ductal adenocarcinoma (PDAC). However, reliable markers to distinguish therapy effects from pre-existing tumor features are lacking. The aim of this study was the characterization of PDAC after NAT, focusing on the stroma. MATERIAL AND METHODS: Tissue samples from patients resected for PDAC after NAT (n=27) were analyzed. TRG was assessed using the Royal North Shore (RNS) system. Stromal composition was evaluated by Movat's stain. Immunohistochemistry (IH) for Ki-67 and five previously established stroma markers (alpha-Crystallin B, alpha-Smooth muscle actin (alpha-SMA), Neurotrophin-3 (NT-3), SPARC and Tenascin C) was also performed. Results were compared with therapy-naïve PDACs (n=10). RESULTS: Most cases showed a moderate response (RNS 2; 74%), while 15% displayed a poor response (RNS 3), and 11% a good response (RNS 1). No complete response was observed. Poor regression was associated with mucin-rich stroma, while good regression was associated with collagen-rich stroma. Cases with poorer therapy response had significantly higher proliferation. Higher peritumoral staining intensity for alpha-SMA and Tenascin C also showed a trend towards an association with poor regression. CONCLUSIONS: Similar to the stroma in therapy-naïve PDAC, the stroma of PDAC after NAT is heterogeneous. Distinguishing between desmoplastic stroma and therapy-induced fibrosis by single markers is not possible. Movat's pentachrome stain, IH for Ki-67, and to some extent for Tenascin C and alpha-SMA, can help detect poor histopathological response to NAT.

mTORC1 and mTORC2 Converge on the Arp2/3 Complex to Promote KrasG12D-Induced Acinar-to-Ductal Metaplasia and Early Pancreatic Carcinogenesis.

BACKGROUND & AIMS: Oncogenic KrasG12D induces neoplastic transformation of pancreatic acinar cells through acinar-to-ductal metaplasia (ADM), an actin-based morphogenetic process, and drives pancreatic ductal adenocarcinoma (PDAC). mTOR (mechanistic target of rapamycin kinase) complex 1 (mTORC1) and 2 (mTORC2) contain Rptor and Rictor, respectively, and are activated downstream of KrasG12D, thereby contributing to PDAC. Yet, whether and how mTORC1 and mTORC2 impact on ADM and the identity of the actin nucleator(s) mediating such actin rearrangements remain unknown. METHODS: A mouse model of inflammation-accelerated KrasG12D-driven early pancreatic carcinogenesis was used. Rptor, Rictor, and Arpc4 (actin-related protein 2/3 complex subunit 4) were conditionally ablated in acinar cells to deactivate the function of mTORC1, mTORC2 and the actin-related protein (Arp) 2/3 complex, respectively. RESULTS: We found that mTORC1 and mTORC2 are markedly activated in human and mouse ADM lesions, and cooperate to promote KrasG12D-driven ADM in mice and in vitro. They use the Arp2/3 complex as a common downstream effector to induce the remodeling the actin cytoskeleton leading to ADM. In particular, mTORC1 regulates the translation of Rac1 (Rac family small GTPase 1) and the Arp2/3-complex subunit Arp3, whereas mTORC2 activates the Arp2/3 complex by promoting Akt/Rac1 signaling. Consistently, genetic ablation of the Arp2/3 complex prevents KrasG12D-driven ADM in vivo. In acinar cells, the Arp2/3 complex and its actin-nucleation activity mediated the formation of a basolateral actin cortex, which is indispensable for ADM and pre-neoplastic transformation. CONCLUSIONS: Here, we show that mTORC1 and mTORC2 attain a dual, yet nonredundant regulatory role in ADM and early pancreatic carcinogenesis by promoting Arp2/3 complex function. The role of Arp2/3 complex as a common effector of mTORC1 and mTORC2 fills the gap between oncogenic signals and actin dynamics underlying PDAC initiation.

Multiparametric Modelling of Survival in Pancreatic Ductal Adenocarcinoma Using Clinical, Histomorphological, Genetic and Image-Derived Parameters.

RATIONALE: Pancreatic ductal adenocarcinoma (PDAC) remains a tumor entity of exceptionally poor prognosis, and several biomarkers are under current investigation for the prediction of patient prognosis. Many studies focus on promoting newly developed imaging biomarkers without a rigorous comparison to other established parameters. To assess the true value and leverage the potential of all efforts in this field, a multi-parametric evaluation of the available biomarkers for PDAC survival prediction is warranted. Here we present a multiparametric analysis to assess the predictive value of established parameters and the added contribution of newly developed imaging features such as biomarkers for overall PDAC patient survival. METHODS: 103 patients with resectable PDAC were retrospectively enrolled. Clinical and histopathological data (age, sex, chemotherapy regimens, tumor size, lymph node status, grading and resection status), morpho-molecular and genetic data (tumor morphology, molecular subtype, tp53, kras, smad4 and p16 genetics), image-derived features and the combination of all parameters were tested for their prognostic strength based on the concordance index (CI) of multivariate Cox proportional hazards survival modelling after unsupervised machine learning preprocessing. RESULTS: The average CIs of the out-of-sample data were: 0.63 for the clinical and histopathological features, 0.53 for the morpho-molecular and genetic features, 0.65 for the imaging features and 0.65 for the combined model including all parameters. CONCLUSIONS: Imaging-derived features represent an independent survival predictor in PDAC and enable the multiparametric, machine learning-assisted modelling of postoperative overall survival with a high performance compared to clinical and morpho-molecular/genetic parameters. We propose that future studies systematically include imaging-derived features to benchmark their additive value when evaluating biomarker-based model performance.