Metastatic Tumor Evolution and Organoid Modeling Implicate TGFBR2 as a Cancer Driver in Diffuse Gastric Cancer

Background: Gastric cancer is the second-leading cause of global cancer deaths, with metastatic disease representing the primary cause of mortality. To identify candidate drivers involved in oncogenesis and tumor evolution, we conduct an extensive genome sequencing analysis of metastatic progression in a diffuse gastric cancer. This involves a comparison between a primary tumor from a hereditary diffuse gastric cancer syndrome proband and its recurrence as an ovarian metastasis. Results: Both the primary tumor and ovarian metastasis have common biallelic loss-of-function of both the CDH1 and TP53 tumor suppressors, indicating a common genetic origin. While the primary tumor exhibits amplification of the Fibroblast growth factor receptor 2 (FGFR2) gene, the metastasis notably lacks FGFR2 amplification but rather possesses unique biallelic alterations of Transforming growth factor-beta receptor 2 (TGFBR2), indicating the divergent in vivo evolution of a TGFBR2-mutant metastatic clonal population in this patient. As TGFBR2 mutations have not previously been functionally validated in gastric cancer, we modeled the metastatic potential of TGFBR2 loss in a murine three-dimensional primary gastric organoid culture. The Tgfbr2 shRNA knockdown within Cdh1-/-; Tp53-/- organoids generates invasion in vitro and robust metastatic tumorigenicity in vivo, confirming Tgfbr2 metastasis suppressor activity. Conclusions: We document the metastatic differentiation and genetic heterogeneity of diffuse gastric cancer and reveal the potential metastatic role of TGFBR2 loss-of-function. In support of this study, we apply a murine primary organoid culture method capable of recapitulating in vivo metastatic gastric cancer. Overall, we describe an integrated approach to identify and functionally validate putative cancer drivers involved in metastasi

An International Consensus to Standardize Integration of Histopathology in Ulcerative Colitis Clinical Trials

Background & Aims: Histopathology is an emerging treatment target in ulcerative colitis (UC) clinical trials. Our aim was to provide guidance on standardizing biopsy collection protocols, identifying optimal evaluative indices, and defining thresholds for histologic response and remission after treatment. Methods: An international, interdisciplinary expert panel of 19 gastroenterologists and gastrointestinal pathologists was assembled. A modified RAND/University of California, Los Angeles appropriateness methodology was used to address relevant issues. A total of 138 statements were derived from a systematic review of the literature and expert opinion. Each statement was anonymously rated as appropriate, uncertain, or inappropriate using a 9-point scale. Survey results were reviewed and discussed before a second round of voting. Results: Histologic measurements collected using a uniform biopsy strategy are important for assessing disease activity and determining therapeutic efficacy in UC clinical trials. Multiple biopsy strategies were deemed acceptable, including segmental biopsies collected according to the endoscopic appearance. Biopsies should be scored for architectural change, lamina propria chronic inflammation, basal plasmacytosis, lamina propria and epithelial neutrophils, epithelial damage, and erosions/ulcerations. The Geboes score, Robarts Histopathology Index, and Nancy Index were considered appropriate for assessing histologic activity; use of the modified Riley score and Harpaz Index were uncertain. Histologic activity at baseline should be required for enrollment, recognizing this carries operational implications. Achievement of histologic improvement or remission was considered an appropriate and realistic therapeutic target. Current histologic indices require validation for pediatric populations. Conclusions: These recommendations provide a framework for standardized implementation of histopathology in UC trials. Additional work is required to address operational considerations and areas of uncertainty

Diagnostic accuracy of cyst fluid amphiregulin in pancreatic cysts

<p>Abstract</p> <p>Background</p> <p>Accurate tests to diagnose adenocarcinoma and high-grade dysplasia among mucinous pancreatic cysts are clinically needed. This study evaluated the diagnostic utility of amphiregulin (AREG) as a pancreatic cyst fluid biomarker to differentiate non-mucinous, benign mucinous, and malignant mucinous cysts.</p> <p>Methods</p> <p>A single-center retrospective study to evaluate AREG levels in pancreatic cyst fluid by ELISA from 33 patients with a histological gold standard was performed.</p> <p>Results</p> <p>Among the cyst fluid samples, the median (IQR) AREG levels for non-mucinous (n = 6), benign mucinous (n = 15), and cancerous cysts (n = 15) were 85 pg/ml (47-168), 63 pg/ml (30-847), and 986 pg/ml (417-3160), respectively. A significant difference between benign mucinous and malignant mucinous cysts was observed (<it>p </it>= 0.025). AREG levels greater than 300 pg/ml possessed a diagnostic accuracy for cancer or high-grade dysplasia of 78% (sensitivity 83%, specificity 73%).</p> <p>Conclusion</p> <p>Cyst fluid AREG levels are significantly higher in cancerous and high-grade dysplastic cysts compared to benign mucinous cysts. Thus AREG exhibits potential clinical utility in the evaluation of pancreatic cysts.</p

Utilizing Deep Learning to Analyze Whole Slide Images of Colonic Biopsies for Associations Between Eosinophil Density and Clinicopathologic Features in Active Ulcerative Colitis

Background: Eosinophils have been implicated in the pathogenesis of ulcerative colitis and have been associated with disease course and therapeutic response. However, associations between eosinophil density, histologic activity, and clinical features have not been rigorously studied. Methods: A deep learning algorithm was trained to identify eosinophils in colonic biopsies and validated against pathologists' interpretations. The algorithm was applied to sigmoid colon biopsies from a cross-sectional cohort of 88 ulcerative colitis patients with histologically active disease as measured by the Geboes score and Robarts histopathology index (RHI). Associations between eosinophil density, histologic activity, and clinical features were determined. Results: The eosinophil deep learning algorithm demonstrated almost perfect agreement with manual eosinophil counts determined by 4 pathologists (interclass correlation coefficients: 0.805-0.917). Eosinophil density varied widely across patients (median 113.5 cells per mm2, interquartile range 108.9). There was no association between eosinophil density and RHI (P=0.5). Significant differences in eosinophil density were seen between patients with Montreal E3 vs E2 disease (146.2 cells per mm2 vs 88.2 cells per mm2, P=0.005). Patients on corticosteroids had significantly lower eosinophil density (62.9 cells per mm2 vs 124.1 cells per mm2, P=0.006). No association between eosinophil density and biologic use was observed (P=0.5). Conclusions: We developed a deep learning algorithm to quantify eosinophils in colonic biopsies. Eosinophil density did not correlate with histologic activity but did correlate with disease extent and corticosteroid use. Future studies applying this algorithm in larger cohorts with longitudinal follow-up are needed to further elucidate the role of eosinophils in ulcerative colitis

BRAF-mutated, Microsatellite-stable Adenocarcinoma of the Proximal Colon: An Aggressive Adenocarcinoma With Poor Survival, Mucinous Differentiation, and Adverse Morphologic Features

The association of BRAF V600E mutation and the presence of the CpG island methylator phenotype (CIMP) and microsatellite instability (MSI) often confound analysis of BRAF mutation status and survival in colorectal carcinoma. We evaluated a consecutive series of proximal colonic ad-enocarcinomas for mismatch repair protein abnormalities/MSI, BRAF V600E mutation, and KRAS mutations in an attempt to determine the prognostic significance of these abnormalities and to correlate histopathologic features with molecular alterations. Of the 259 proximal colon adenocarcinomas analyzed for mismatch repair protein abnormalities and/or MSI, 181 proximal colonic adenocarcinomas demonstrated proficient DNA mismatch repair using either MSI PCR (n = 78), mismatch repair protein immunohistochemistry (n = 91), or both MSI PCR and mismatch repair immunohistochemistry (n = 12); these were tested for the BRAF V600E mutation and KRAS mutations. Compared with BRAF wild-type adenocarcinomas, BRAF -mutated adenocarcinomas more frequently demonstrated adverse histologic features such as lymphatic invasion (16/20, 80% vs. 75/161, 47%; P = 0.008), mean number of lymph node metastases (4.5 vs. 2.2; P = 0.01), perineural invasion (8/20, 40% vs. 13/161, 8%; P = 0.0004), and high tumor budding (16/20, 80% vs. 83/161, 52%; P = 0.02). BRAF -mutated adenocarcinomas frequently contained areas with mucinous histology ( P = 0.0002) and signet ring histology ( P = 0.03), compared with KRAS -mutated and KRAS/BRAF wild-type adenocarcinomas. Clinical follow-up data were available for 173 proximal colonic adenocarcinomas with proficient DNA mismatch repair. Patients with BRAF -mutated adenocarcinomas had a median survival of 12.3 months with a 1-year probability of survival of 54% and a 1-year disease-free survival of 56%. Patients with KRAS -mutated and KRAS/BRAF wild-type adenocarcinomas had significantly improved overall survival (unadjusted log-rank P = 0.03 and unadjusted log-rank P = 0.0002, respectively) and disease-free survival (unadjusted log-rank P = 0.02 and unadjusted log-rank P = 0.02, respectively) compared with patients with BRAF -mutated adenocarcinomas. When adjusting for tumor stage, survival analysis demonstrated that patients with BRAF -mutated adenocarcinoma had a significantly poor overall survival and disease-free survival (hazard ratios 6.63, 95% CI, 2.60–16.94; and 6.08, 95% CI, 2.11–17.56, respectively) compared with patients with KRAS/BRAF wild-type adenocarcinomas. No significant difference in overall or disease-free survival was identified between patients with KRAS -mutated and KRAS/BRAF wild-type adenocarcinomas. Our results demonstrate that BRAF -mutated proximal colon adenocarcinomas with proficient DNA mismatch repair have a dismal prognosis with an aggressive clinical course and often display mucinous differentiation, focal signet ring histology, and other adverse histologic features such as lymphatic and perineural invasion and high tumor budding

DNA mismatch repair protein deficient non-neoplastic colonic crypts: a novel indicator of Lynch syndrome

Lynch syndrome is the most common form of hereditary colorectal carcinoma. However, establishing the diagnosis of Lynch syndrome is challenging, and ancillary studies that distinguish between sporadic DNA mismatch repair (MMR) protein deficiency and Lynch syndrome are needed, particularly when germline mutation studies are inconclusive. The aim of this study was to determine if MMR protein-deficient non-neoplastic intestinal crypts can help distinguish between patients with and without Lynch syndrome. We evaluated the expression of MMR proteins in non-neoplastic intestinal mucosa obtained from colorectal surgical resection specimens from patients with Lynch syndrome-associated colorectal carcinoma (n = 52) and patients with colorectal carcinoma without evidence of Lynch syndrome (n = 70), including sporadic MMR protein-deficient colorectal carcinoma (n = 30), MMR protein proficient colorectal carcinoma (n = 30), and "Lynch-like" syndrome (n = 10). MMR protein-deficient non-neoplastic colonic crypts were identified in 19 of 122 (16%) patients. MMR protein-deficient colonic crypts were identified in 18 of 52 (35%) patients with Lynch syndrome compared to only 1 of 70 (1%) patients without Lynch syndrome (p

Variation in Pathologist Classification of Colorectal Adenomas and Serrated Polyps

OBJECTIVES: Endoscopist quality measures such as adenoma detection rate (ADR) and serrated polyp detection rates (SPDRs) depend on pathologist classification of histology. Although variation in pathologic interpretation is recognized, we add to the literature by quantifying the impact of pathologic variability on endoscopist performance. METHODS: We used natural language processing to abstract relevant data from colonoscopy and related pathology reports performed over 2 years at four clinical sites. We quantified each pathologist's likelihood of classifying polyp specimens as adenomas or serrated polyps. We estimated the impact on endoscopists' ADR and SPDR of sending their specimens to pathologists with higher or lower classification rates. RESULTS: We observed 85,526 colonoscopies performed by 119 endoscopists; 50,453 had a polyp specimen, which were analyzed by 48 pathologists. There was greater variation across pathologists in classification of serrated polyps than in classification of adenomas. We estimate the endoscopist's average SPDR would be 0.5% if all their specimens were analyzed by the pathologist in our sample with the lowest classification rate and 12.0% if all their specimens were analyzed by the pathologist with the highest classification rate. In contrast, the endoscopist's average ADR would be 28.5% and 42.4% if their specimens were analyzed by the pathologist with lowest and highest classification rate, respectively. CONCLUSIONS: There is significant variation in pathologic interpretation, which more substantially affects endoscopist SPDR than ADR