Reanimating Pancreatic Grafts Subjected to Prolonged Cold Ischemic Injury Using Normothermic Ex Vivo Perfusion.

Background: Pancreas transplant volumes are limited because of poor utilization of "extended criteria grafts." Prolonged cold ischemia is a risk factor associated with poor allograft survival. We aimed to establish the feasibility of transplantation using grafts subjected to prolonged cold ischemia and determine whether these grafts could be optimized using normothermic ex vivo perfusion (NEVP) in a porcine model. Methods: The study population consisted of 35 to 40 kg male Yorkshire pigs in an allotransplantation model with a 3-d survival plan for recipients. Control grafts were subjected to cold storage (CS) in a University of Wisconsin solution for 21 to 24 h (n = 6), whereas the test group received an additional 3 h NEVP after CS of 21 h (n = 5). Results: The 3-d survival was 60% in the NEVP arm versus 0% in the control arm (P = 0.008; log rank). Graft parenchyma was 60% to 70% preserved in the NEVP arm at necropsy on gross appearance. In addition, the islet function was well preserved, and both the pancreas (including the islets) and the duodenal morphology were maintained histologically. The intravenous glucose tolerance test on the day of euthanasia was in the normoglycemic range for 80% of cases in the NEVP arm. Conclusions: Optimization of pancreas grafts exposed to extended CS with NEVP seems promising at rescuing and reanimating these grafts for transplantation, resulting in significantly improved survival in a porcine pancreas transplant model.

Normothermic Ex Vivo Machine Perfusion of Discarded Human Pancreas Allografts: A Feasibility Study.

Pancreas transplantation is the only curative treatment for patients with complicated diabetes, and organ shortage is a common and increasing problem. Strategies to expand the donor pool are needed, and normothermic ex vivo perfusion of the pancreas has the potential to test and repair grafts before implantation. Between January 2021 and April 2022, six human pancreases, declined for transplantation or islet isolation, were perfused using a previously established method by our group. All 6 cases were successfully perfused for 4 h, with minimal edema. The mean age of the donors was 44.16 ± 13.8 years. Five grafts were obtained from neurological death donors, and one was obtained from a donation after cardiac death. The mean glucose and lactate levels decreased throughout perfusion and insulin levels increased. All 6 grafts were metabolically active during perfusion and histopathology showed minimal tissue injury and no edema. Human normothermic ex vivo perfusion of the pancreas is feasible and safe and has the potential to expand the donor pool. Future studies will focus on tests and biomarkers for the assessment of grafts.

Barrett's Esophagus and Esophageal Adenocarcinoma: A Histopathological Perspective.

Esophageal adenocarcinoma (EAC) is increasing in prevalence. Barrett's esophagus (BE) has long been recognized as the putative precursor lesion for EAC, but much is still unknown regarding its cell of origin and what molecular factors influence its neoplastic progression. Accurate pathologic assessment of BE biopsies is important for identifying patients most at risk of progressing to EAC, whereas pathologic assessment of EAC specimens plays a major role in influencing therapeutic decision-making.

Normothermic ex situ pancreas perfusion for the preservation of porcine pancreas grafts.

Pancreas transplantation improves and extends the life of patients with insulin-dependent diabetes. Pancreata from extended criteria donors have been increasingly used due to the scarcity of available grafts. Normothermic ex situ pancreas perfusion (NESPP) can keep grafts metabolically active, potentially allowing for assessment and organ repair, and could improve outcomes of marginal grafts. A novel NESPP technique was developed and tested. Porcine pancreata were removed after a short period of warm ischemia and subjected to 6 h of NESPP. Perfusion parameters, potential graft assessment markers and graft injury were measured. Next, pancreata subjected to 3 h of NESPP were transplanted and animals were followed for up to 3 days. Graft function and injury post-transplantation were evaluated. Using this novel system of perfusion, pancreata were perfused for an extended period of time with minimal edema. Histology at the end of perfusion showed intact islet cells with only mild signs of tissue injury. NESPP transplanted grafts showed immediate function after transplantation, with glucose levels in normal range. NESPP maintains a physiologic environment and excellent graft function without causing significant graft injury. Porcine pancreas transplantation is feasible and allows for in vivo graft assessment of pancreas function and injury after NESPP.

Preoperative and Postoperative Approaches to Gastroesophageal Cancer: What is All the Fuss About.

Gastroesophageal cancers carry poor prognoses, and are a leading cause of cancer-related morbidity and mortality worldwide. Even in those with resectable disease, more than half of patients treated with surgery alone experience disease recurrence. Multimodality approaches using preoperative and postoperative chemotherapy and/or radiotherapy have been established, resulting in incremental improvements in outcomes. Globally, there is no standardized approach, and treatment varies with geographic location. The question remains of how to select the optimal perioperative treatment that will maximize benefit for patients while avoiding toxicities from unnecessary therapies. This article reviews currently available evidence supporting preoperative and postoperative therapy in gastroesophageal cancers, with an emphasis on recent practice-changing trials and ongoing areas of investigation, including the role of immune checkpoint inhibition and biomarker-guided treatment.

Loss of SMAD4 Is Sufficient to Promote Tumorigenesis in a Model of Dysplastic Barrett's Esophagus.

BACKGROUND & AIMS: Esophageal adenocarcinoma (EAC) develops from its precursor Barrett's esophagus through intermediate stages of low- and high-grade dysplasia. However, knowledge of genetic drivers and molecular mechanisms implicated in disease progression is limited. Herein, we investigated the effect of Mothers against decapentaplegic homolog 4 (SMAD4) loss on transforming growth factor ß (TGF-ß) signaling functionality and in vivo tumorigenicity in high-grade dysplastic Barrett's cells. METHODS: An in vivo xenograft model was used to test tumorigenicity of SMAD4 knockdown or knockout in CP-B high-grade dysplastic Barrett's cells. RT2 polymerase chain reaction arrays were used to analyze TGF-ß signaling functionality, and low-coverage whole-genome sequencing was performed to detect copy number alterations upon SMAD4 loss. RESULTS: We found that SMAD4 knockout significantly alters the TGF-ß pathway target gene expression profile. SMAD4 knockout positively regulates potential oncogenes such as CRYAB, ACTA2, and CDC6, whereas the CDKN2A/B tumor-suppressor locus was regulated negatively. We verified that SMAD4 in combination with CDC6-CDKN2A/B or CRYAB genetic alterations in patient tumors have significant predictive value for poor prognosis. Importantly, we investigated the effect of SMAD4 inactivation in Barrett's tumorigenesis. We found that genetic knockdown or knockout of SMAD4 was sufficient to promote tumorigenesis in dysplastic Barrett's esophagus cells in vivo. Progression to invasive EAC was accompanied by distinctive and consistent copy number alterations in SMAD4 knockdown or knockout xenografts. CONCLUSIONS: Altogether, up-regulation of oncogenes, down-regulation of tumor-suppressor genes, and chromosomal instability within the tumors after SMAD4 loss implicates SMAD4 as a protector of genome integrity in EAC development and progression. Foremost, SMAD4 loss promotes tumorigenesis from dysplastic Barrett's toward EAC.

Mutations in the RAS/MAPK Pathway Drive Replication Repair-Deficient Hypermutated Tumors and Confer Sensitivity to MEK Inhibition.

The RAS/MAPK pathway is an emerging targeted pathway across a spectrum of both adult and pediatric cancers. Typically, this is associated with a single, well-characterized point mutation in an oncogene. Hypermutant tumors that harbor many somatic mutations may obscure the interpretation of such targetable genomic events. We find that replication repair-deficient (RRD) cancers, which are universally hypermutant and affect children born with RRD cancer predisposition, are enriched for RAS/MAPK mutations (P = 10-8). These mutations are not random, exist in subclones, and increase in allelic frequency over time. The RAS/MAPK pathway is activated both transcriptionally and at the protein level in patient-derived RRD tumors, and these tumors responded to MEK inhibition in vitro and in vivo. Treatment of patients with RAS/MAPK hypermutant gliomas reveals durable responses to MEK inhibition. Our observations suggest that hypermutant tumors may be addicted to oncogenic pathways, resulting in favorable response to targeted therapies. SIGNIFICANCE: Tumors harboring a single RAS/MAPK driver mutation are targeted individually for therapeutic purposes. We find that in RRD hypermutant cancers, mutations in the RAS/MAPK pathway are enriched, highly expressed, and result in sensitivity to MEK inhibitors. Targeting an oncogenic pathway may provide therapeutic options for these hypermutant polyclonal cancers.This article is highlighted in the In This Issue feature, p. 1307.

Prognostic significance of nutritional markers in metastatic gastric and esophageal adenocarcinoma.

BACKGROUND: Malnutrition and sarcopenia are poor prognostic factors in many cancers. Studies in gastric and esophageal (GE) cancer have focused on curative intent patients. This study aims to evaluate the prognostic utility of malnutrition and sarcopenia in de novo metastatic GE adenocarcinoma. METHODS: Patients with de novo metastatic GE adenocarcinoma seen at the Princess Margaret Cancer Centre from 2010 to 2016 with an available pre-treatment abdominal computed tomography (CT) were included. Malnutrition was defined as nutritional risk index (NRI) <97.5. Skeletal muscle index (SMI) was measured at the L3 level (sarcopenia defined as SMI <34.4 cm2 /m2 in women and <45.4 cm2 /m2 in men). Patients receiving chemotherapy had NRI and SMI recalculated at the time of first restaging CT. RESULTS: Of 175 consecutive patients, 33% were malnourished and 39% were sarcopenic at baseline. Patients with pretreatment malnourishment had significantly shorter overall survival (OS; 5.8 vs. 10.9 months, p = 0.000475). Patients who became malnourished during chemotherapy had worse OS compared to those who maintained their nutrition (12.2 vs. 17.5 months p = 0.0484). On univariable analysis, ECOG (p < 0.001), number of metastatic sites (p = 0.029) and NRI (p < 0.001) were significant prognostic factors while BMI (p = 0.57) and sarcopenia (p = 0.19) were not. On multivariable analysis, ECOG (p < 0.001), baseline NRI (p = 0.025), and change in NRI during treatment (p < 0.001) were significant poor prognostic factors for OS. CONCLUSIONS: In de novo metastatic GE adenocarcinoma patients, ECOG, pretreatment NRI and change in NRI were significant prognostic factors for OS while sarcopenia was not. Use of NRI at baseline and during treatment can provide useful prognostic information.

Unsupervised Resolution of Histomorphologic Heterogeneity in Renal Cell Carcinoma Using a Brain Tumor-Educated Neural Network.

PURPOSE: Applications of deep learning to histopathology have proven capable of expert-level performance, but approaches have largely focused on supervised classification tasks requiring context-specific training and deployment. More generalizable workflows that can be easily shared across subspecialties could help accelerate and broaden adoption. Here, we hypothesized that histology-optimized feature representations, generated by a convolutional neural network (CNN) during supervised learning, are transferable and can resolve meaningful differences in large-scale, discovery-type unsupervised analyses. METHODS: We used a CNN, previously trained to recognize brain tumor histomorphologies, to extract 512 feature representations from > 550 digital whole-slide images (WSIs) of renal cell carcinomas (RCCs) from The Cancer Genome Atlas and other previously unencountered tumors. We use these extracted feature vectors to conduct unsupervised image-set clustering and analyze the clinical and biologic relevance of the intra- and interpatient subgroups generated. RESULTS: Within individual WSIs, feature-based clustering could reliably segment tumor regions and other relevant histopathologic subpatterns (eg, adenosquamous and poorly differentiated regions). Across the larger RCC cohorts, clustering extracted features generated subgroups enriched for clinically relevant subtypes (eg, papillary RCC) and outcomes (eg, survival). Importantly, individual feature activation mapping highlighted salient subtype-specific patterns and features of malignancies (eg, nuclear grade, sarcomatous change) contributing to subgroupings. Moreover, some proposed clusters were enriched for recurring, human-based RCC-subtype misclassifications. CONCLUSION: Our data support that CNNs, pretrained on large histologic datasets, can extend learned representations to novel scenarios and resolve clinically relevant intra- and interpatient tissue-pattern differences without explicit instruction or additional optimization. Repositioning of existing histology-educated networks could provide scalable approaches for image classification, quality assurance, and discovery of unappreciated patterns and subgroups of disease.

Impact of adjuvant therapy in patients with a microscopically positive margin after resection for gastric and esophageal cancers.

BACKGROUND: A microscopically positive (R1) resection margin following resection for gastric and esophageal cancers has been documented to be a poor prognostic factor. The optimal strategy and impact of different modalities of adjuvant treatment for an R1 resection margin remain unclear. METHODS: A retrospective analysis was performed for patients with gastric and esophageal adenocarcinoma treated at the Princess Margaret Cancer Centre (PMCC) from 2006-2016. Electronic medical records of all patients with an R1 resection margin were reviewed. Kaplan-Meier and Cox proportional hazards methods were used to analyze recurrence free survival (RFS) and overall survival (OS) with stage and neoadjuvant treatment as covariates in the multivariate analysis. RESULTS: We identified 69 gastric and esophageal adenocarcinoma patients with a R1 resection. Neoadjuvant chemoradiation was used in 13% of patients, neoadjuvant chemotherapy in 12%, surgery alone in 75%. Margins involved included proximal in 30%, distal in 14%, radial in 52% and multiple margins in 3% of patients. Pathological staging showed 3% with stage I disease, 20% stage II and 74% stage III. Adjuvant therapy was given in 52% of R1 pts (28% CRT, 20% chemotherapy alone, 3% radiation alone, 1% reoperation). Median RFS was 14.1 months [95% confidence interval (CI), 11.1-17.2]. The site of first recurrence was 72% distant, 12% mixed, 16% locoregional alone. Median OS was 34.5 months (95% CI, 23.3-57.9) for all patients. There was no significant difference in RFS (adjusted P=0.26) or OS (adjusted P=0.83) comparing modality of adjuvant therapy. CONCLUSIONS: Most patients with positive margins after resection for gastric and esophageal cancer had advanced pathologic stage and prognosis was poor. Our study did not find improved RFS or OS with adjuvant treatment and only one patient had reresection. The main failure pattern was distant recurrence, suggesting that patients being considered for adjuvant radiotherapy (RT) should be carefully selected. Further studies are required to determine factors to select patients with good prognosis despite a positive margin, or those who may benefit from adjuvant treatment.

Sclerosing Tumors of the Gastrointestinal Tract: A Systematic Approach.

Some lesions in the gastrointestinal tract have a propensity for sclerosis such that it may mask the actual true nature of the lesion. The purpose of this review is to highlight those lesions of the gastrointestinal tract that can be attended by sclerosis. The sclerosis can mask the cellularity of the lesion; hence, knowledge of the key lesions that are known to have sclerosis will be aid the diagnostic pathologist.

Primary inferior vena cava smooth muscle tumor with diffuse bizarre giant nuclei and low mitotic rate: a nomenclatural conundrum.

A male patient with obstructive jaundice was found to have an incidental nodule within the inferior vena cava (IVC), below the level of the renal vein, on abdominal imaging. At the time of the Whipple's procedure for pancreatic adenocarcinoma, the IVC mass measuring 3.4×2.7×2.2 cm was also removed. Histologically, the lesion was well circumscribed, composed focally of spindle-shaped cells with cigar-shaped nuclei reminiscent of smooth muscle and a dominant pervasive, pleomorphic, bizarre giant cell component. Two mitoses per 10 high-power fields were identified in the most mitotically active area of the entire tumor, with the vast majority of the tumor being mitotically inert. Additionally, no evidence of coagulative necrosis was noted. The bizarre giant cells had multi- and polylobated configurations, and several were replete with nuclear pseudoinclusions. Both the spindle cell and pleomorphic components displayed strong immunoreactivity for all smooth muscle markers. This lesion conformed morphologically to a smooth muscle tumor with bizarre nuclei or so-called symplastic/bizarre leiomyoma, as encountered in the uterus. However, current thinking based on location in the IVC and the presence of any mitotic activity with cellular atypia makes this lesion a leiomyosarcoma. Perhaps more pragmatic terminology would be smooth muscle tumor with bizarre nuclei and low malignant potential since the limited number of cases described thus far appear to have a more indolent course.

An unusual primary malignant tumor of the stomach: fetal gutlike gastric adenocarcinoma with "blastoma"-like component.

An unusual case of a polypoid, malignant gastric tumor in a 62-year-old man is presented. Endoscopy and subsequent polypectomy revealed an 8.5 × 6.5 × 4.5-cm lesion in the body of the stomach. Microscopy showed surface dysplasia with an invasive adenocarcinoma displaying prominent tubulopapillary areas composed of large vacuolated cells, pleomorphic nuclei, and occasional cytoplasmic hyaline globules. This component then blended with tubular structures lined by more primitive-appearing vacuolated cells embedded within a stroma made up of cellular primitive, high-grade blastemalike areas and less cellular, more pleomorphic foci with spindle and several bizarre, large cells. Immunohistochemistry showed the adenocarcinoma and primitive tubules to be strongly SALL4 and epithelial marker positive but with only focal expression of α-fetoprotein and glypican-3. The stromal component made up of blastemalike areas displayed strong immunoreactivity for glypican-3. The pleomorphic stromal areas were negative for all markers, including epithelial and muscle markers. The overall morphology and expression of primitive oncofetal proteins, especially SALL4 and glypican-3, are in keeping with this being a primitive adenocarcinoma showing fetal gutlike differentiation with an accompanying blastomalike component, a combination not previously described in a primary gastric cancer.

Association of Distinct Mutational Signatures With Correlates of Increased Immune Activity in Pancreatic Ductal Adenocarcinoma.

IMPORTANCE: Outcomes for patients with pancreatic ductal adenocarcinoma (PDAC) remain poor. Advances in next-generation sequencing provide a route to therapeutic approaches, and integrating DNA and RNA analysis with clinicopathologic data may be a crucial step toward personalized treatment strategies for this disease. OBJECTIVE: To classify PDAC according to distinct mutational processes, and explore their clinical significance. DESIGN, SETTING, AND PARTICIPANTS: We performed a retrospective cohort study of resected PDAC, using cases collected between 2008 and 2015 as part of the International Cancer Genome Consortium. The discovery cohort comprised 160 PDAC cases from 154 patients (148 primary; 12 metastases) that underwent tumor enrichment prior to whole-genome and RNA sequencing. The replication cohort comprised 95 primary PDAC cases that underwent whole-genome sequencing and expression microarray on bulk biospecimens. MAIN OUTCOMES AND MEASURES: Somatic mutations accumulate from sequence-specific processes creating signatures detectable by DNA sequencing. Using nonnegative matrix factorization, we measured the contribution of each signature to carcinogenesis, and used hierarchical clustering to subtype each cohort. We examined expression of antitumor immunity genes across subtypes to uncover biomarkers predictive of response to systemic therapies. RESULTS: The discovery cohort was 53% male (n = 79) and had a median age of 67 (interquartile range, 58-74) years. The replication cohort was 50% male (n = 48) and had a median age of 68 (interquartile range, 60-75) years. Five predominant mutational subtypes were identified that clustered PDAC into 4 major subtypes: age related, double-strand break repair, mismatch repair, and 1 with unknown etiology (signature 8). These were replicated and validated. Signatures were faithfully propagated from primaries to matched metastases, implying their stability during carcinogenesis. Twelve of 27 (45%) double-strand break repair cases lacked germline or somatic events in canonical homologous recombination genes-BRCA1, BRCA2, or PALB2. Double-strand break repair and mismatch repair subtypes were associated with increased expression of antitumor immunity, including activation of CD8-positive T lymphocytes (GZMA and PRF1) and overexpression of regulatory molecules (cytotoxic T-lymphocyte antigen 4, programmed cell death 1, and indolamine 2,3-dioxygenase 1), corresponding to higher frequency of somatic mutations and tumor-specific neoantigens. CONCLUSIONS AND RELEVANCE: Signature-based subtyping may guide personalized therapy of PDAC in the context of biomarker-driven prospective trials.

From traditional serrated adenoma to tubulovillous adenoma and beyond.

It is well established that colorectal cancer develops from a series of precursor epithelial polyps, including tubular adenomas, villous/tubulovillous adenomas (VA/TVA), sessile serrated adenomas (SSA) and traditional serrated adenomas (TSA). Of these, TSAs are least common and account for only 5% of all serrated polyps. TSAs are characterised by the presence of a "pinecone-like" architecture, granular eosinophilic cytoplasm, luminal serrations, ectopic crypt foci (ECF) and elongated, pencillate nuclei. However, the distinct slit-like luminal serrations, reminiscent of small bowel mucosa, appear to be the most unique and reproducible feature to distinguish TSAs from other polyps. There is a contention that TSAs are not inherently dysplastic and that the majority do not show cytological atypia. Two types of dysplasia are associated with TSA. Serrated dysplasia is less well recognised and less commonly encountered than adenomatous dysplasia. In addition, it is now becoming increasingly evident that TSAs can be admixed with HP, SSA and VA/TVA. At a genetic level, polyps may switch phenotype as they accumulate genetic changes, evolving from a serrated pathway to a more conventional one, which could be the basis for a spectrum theory starting out with a TSA with serration and ECF evolving into a TSA with conventional dysplasia and, eventually, to a well-developed conventional adenoma. Nevertheless, there is an exigency for future studies to provide further illumination and bridge the gaps in our present understanding.

Gene of the month: SMARCB1.

SMARCB1 is the core subunit of the SWI/sucrose non-fermenting ATP-dependent chromatin remodelling complex located on the long arm of chromosome 22 (22q11.2). Since discovering genetic alterations of the SMARCB1 gene in malignant rhabdoid tumours, the family of tumours harbouring loss of SMARCB1 expression has been steadily expanding. In this review, we give a general overview of SMARCB1, its role in various cancers including germline mutations, association with genetic syndromes and role in future targeted therapies.

The spectrum of histopathological changes encountered in pancreatectomy specimens after neoadjuvant chemoradiation, including subtle and less-well-recognised changes.

Preoperative (neoadjuvant) chemoradiation therapy/treatment (NCRT) is emerging as an important treatment modality in borderline resectable pancreatic ductal adenocarcinoma (PDAC). The constellation of histopathological changes secondary to chemoradiation is diverse and has been well documented, particularly in other gastrointestinal organs such as the oesophagus and colorectum. However, the histological changes specific to the pancreas have not been fully characterised and described. This review aims to provide a detailed catalogue of histological features associated with NCRT-treated PDAC and highlight any subtle, less-recognised changes.

Clinical outcome in patients with peripherally-sited atypical lipomatous tumours and dedifferentiated liposarcoma.

The reported incidence of local recurrence of peripheral atypical lipomatous tumours is highly variable and is likely to reflect the different inclusion criteria of cases, and the design of previous studies. We aimed to study the incidence of local recurrence of 90 cases of atypical lipomatous tumours and an additional 18 cases of de novo dedifferentiated liposarcoma. All tumours were diagnosed on the basis of MDM2 amplification: all patients had their first treatment in the same specialist sarcoma unit and were followed for a minimum of 60 months. The tumours were diagnosed between 1997 and 2009 and followed until the end of 2014. Seventy cases (78%) of atypical lipomatous tumours were located in the thigh (mean size 195 mm on presentation). Eight atypical lipomatous tumours (8.9%) recurred locally, of which 50% recurred after 60 months. The only two tumours with intralesional excisions recurred. Seven of the eight recurrent tumours were detected by the patient by self-examination. One case recurred a second time as a dedifferentiated liposarcoma. Seventeen per cent of the de novo dedifferentiated liposarcomas recurred within 60 months of presentation. Extending the study period revealed that atypical lipomatous tumour could recur up to 40 years after the first surgery. Furthermore, of 26 tumours that recurred in the extended study, 27% recurred more than once, and three of the seven that recurred more than once transformed into a dedifferentiated liposarcoma. We recommend that, following post-operative wound care, patients with atypical lipomatous tumour are referred back to their general practitioner for follow up, but that in the event of a suspected recurrence they have rapid access back to the specialist unit using a 'supported discharge' scheme. In the event of an intralesional excision and if a lesion recurs, patients are followed in a specialist unit at regular intervals: whether MRI scanning is a valuable means of monitoring such patients is unclear and requires an evidence base.