Histo- and immunohistochemistry-based estimation of the TCGA and ACRG molecular subtypes for gastric carcinoma and their prognostic significance: A single-institution study.

Gastric cancers comprise molecularly heterogeneous diseases; four molecular subtypes were identified in the cancer genome atlas (TCGA) study, with implications in patient management. In our efforts to devise a clinically feasible means of subtyping, we devised an algorithm based on histology and five stains available in most academic pathology laboratories. This algorithm was used to subtype our cohort of 107 gastric cancer patients from a single institution (St. Michael's Hospital, Toronto, Canada), which was divided into 3 cases of EBV-positive, 23 of MSI, 27 of GS and 54 of CIN tumours. 87% of the tumours with diffuse histology were classified as GS subtype, which was notable for younger age. Examining for characteristic molecular features, aberrant p53 immunostaining was seen most frequently in the CIN subtype (43% in CIN vs. 6% in others), whereas ARID1A loss was rarely seen (6% vs. 35% in others). HER2 overexpression was seen exclusively in CIN tumours (17% of CIN tumours). PD-L1 positivity was seen predominantly in the EBV and MSI tumours. As with the TCGA study, no survival differences were seen between the subtypes. A similar strategy was employed to approximate the Asian Cancer Research Group (ACRG) molecular subtyping, with the addition of p53 IHC to the algorithm. We observed rates of ARID1A loss and HER2 overexpression that were comparable to the ACRG study. In summary, our algorithm allowed for clinically feasible means of subtyping gastric carcinoma that recapitulated the key molecular features reported in the large scale studies.

Prognostic significance of human tissue kallikrein-related peptidases 11 and 15 in gastric cancer.

Human tissue kallikrein-related peptidases (KLK) are a group of 15 serine proteases which have been investigated as potential cancer biomarkers. This study determined the prognostic significance of KLK 11 and 15 expression levels in gastric carcinoma specimens. Expression of KLK11 and KLK15 was assessed by immunohistochemistry staining on a tissue microarray constructed from 113 gastrectomy specimens from patients with gastric carcinoma. To minimize inter-observer variability, expression levels were quantified using an automated algorithm. Epithelial and stromal staining were assessed separately. Both KLK11 and KLK15 were expressed in gastric carcinoma. There was no significant correlation between either KLK11 or KLK15 expression and the presence of lymph node metastases or Lauren classification (intestinal vs. diffuse). Higher levels of KLK11 expression in gastric carcinoma were associated with significantly worse overall survival (p = 0.008), and a multivariate analysis showed that it had prognostic value independent of tumor stage and differentiation (p = 0.004). Variations in KLK15 expression were not significantly associated with prognosis. KLK11 shows promise as a potential independent prognostic marker for gastric carcinoma.

Inflammatory pseudotumours of the oesophagus--histological and immunohistochemical findings.

AIMS: Inflammatory oesophageal pseudotumours are rare lesions, thought to be reactive. Due to marked atypia of the stromal cells, these can be misdiagnosed as malignancies. The objective of this study was to characterize histological and immunohistochemical features of a series of inflammatory pseudotumours of the oesophagus. METHODS AND RESULTS: We present 12 cases of inflammatory oesophageal pseudotumours, occurring in seven females and five males, with a mean age of 57.3 years. Clinical presentations were variable; dysphagia, abdominal pain and weight loss and upper gastrointestinal bleed. In a majority of the cases, nodules or masses in the distal oesophagus were identified at endoscopy. Microscopically, the lamina propria in all 12 cases contained inflammation and granulation tissue. Ten of 12 cases showed mucosal ulceration and 11 of 12 cases had acutely inflamed epithelium. Markedly atypical pleomorphic stromal cells with prominent nucleoli were identified in all 12 cases. Immunohistochemistry showed uniform positivity for vimentin in 11 of 11 cases, and two of seven cases demonstrated weak focal positivity for smooth muscle actin. The cells were negative for all other markers. CONCLUSIONS: Reactive oesophageal lesions can show marked nuclear atypia in stromal fibroblasts/myofibroblasts, which are easily mistaken for malignancies. Pathologists must consider the diagnosis of an inflammatory pseudotumour if stromal atypia is present in an inflammatory background.

Prognostic significance of human tissue kallikrein-related peptidases 6 and 10 in gastric cancer.

The prognosis of patients following surgery for gastric cancer is often poor and is estimated using traditional clinicopathological parameters, which can be inaccurate predictors of future survival. Kallikreins are a group of serine proteases, which are differentially expressed in many human tumors and are being investigated as potential cancer biomarkers. This study assessed the prognostic utility of human tissue kallikrein-like peptidases 6 and 10 (KLK6 and KLK10) and correlated their expression with histopathological and clinical parameters in gastric cancer. We constructed a gastric tumor tissue microarray from 113 gastrectomy specimens and quantified KLK6 and KLK10 expression using immunohistochemistry. To overcome the problem of inter-observer variability and subjectivity in immunohistochemistry interpretation, a whole-slide scanned image of the tissue microarray was analyzed using an automated algorithm to quantify staining intensity. KLK6 expression was positively correlated with nodal involvement (p=0.002) and was predictive of advanced-stage disease (p<0.05). Kaplan-Meier survival curves revealed that tumors expressing high levels of KLK6 were significantly associated with significantly lower overall survival (p=0.04). KLK10 overexpression was also a predictor of advanced-stage disease (p<0.01), but was not significantly correlated with lymph node involvement or survival period. Our results show the potential ability of KLK6 as a prognostic marker for gastric cancer.

The histomorphology of Lynch syndrome-associated ovarian carcinomas: toward a subtype-specific screening strategy.

Women with Lynch syndrome (LS) are at increased risk for the development of epithelial ovarian cancer (OC). Analogous to previous studies on BRCA1/2 mutation carriers, there is evidence to suggest a histotype-specific association in LS-associated OCs (LS-OC). Whereas the diagnosis of high-grade serous carcinoma is an indication for BRCA1/2 germline testing, in contrast, there are no screening guidelines in place for triaging OC patients for LS testing based on histotype. We performed a centralized pathology review of tumor subtype on 20 germline mutation-confirmed LS-OCs, on the basis of morphologic assessment of hematoxylin and eosin-stained slides, with confirmation by immunohistochemistry when necessary. Results from mismatch-repair immunohistochemistry (MMR-IHC) and microsatellite instability (MSI) phenotype status were documented, and detailed pedigrees were analyzed to determine whether previously proposed clinical criteria would have selected these patients for genetic testing. Review of pathology revealed all LS-OCs to be either pure endometrioid carcinoma (14 cases), mixed carcinoma with an endometrioid component (4 cases), or clear cell carcinoma (2 cases). No high-grade or low-grade serous carcinomas or mucinous carcinomas of intestinal type were identified. Tumor-infiltrating lymphocytes were prominent (≥40 per 10 high-powered fields) in 2 cases only. With the exception of 1 case, all tumors tested for MMR-IHC or MSI had an MMR-deficient phenotype. Within this cohort, 50%, 55%, 65%, and 85% of patients would have been selected for genetic workup by Amsterdam II, revised Bethesda Guidelines, SGO 10% to 25%, and SGO 5% to 10% criteria, respectively, with <60% of index or sentinel cases detected by any of these schemas. To further support a subtype-driven screening strategy, MMR-IHC reflex testing was performed on all consecutive non-serous OCs diagnosed at 1 academic hospital over a 2-year period; MMR deficiency was identified in 10/48 (21%) cases, all with endometrioid or clear cell histology. We conclude that there is a strong association between endometrioid and clear cell ovarian carcinomas and hereditary predisposition due to MMR gene mutation. These findings have implications for the role of tumor subtype in screening patients with OC for further genetic testing and support reflex MMR-IHC and/or MSI testing for newly diagnosed cases of endometrioid or clear cell ovarian carcinoma.

ARID1A/BAF250a as a prognostic marker for gastric carcinoma: a study of 2 cohorts.

ARID1A/BAF250a has been recently implicated as a tumor suppressor in gastric cancer. We sought to clarify the clinical significance of BAF250a/ARID1A in relation to other clinical parameters and relevant biomarkers in gastric carcinoma. Cases from 2 separate cohorts of patients with gastric carcinoma from Vancouver (n = 173) and Toronto (n = 80) were selected for the construction of tissue microarrays, which were used to assess the immunohistochemical status of BAF250a (anti-ARID1A), mismatch repair proteins and p53, as well as in situ hybridization for HER2 amplification and Epstein-Barr virus infection. The Toronto cohort contained a higher proportion of early stage cases (P = .019) and a smaller proportion of cases from the proximal stomach (P < .001). Overall, immunohistochemical loss of BAF250a was observed in 22.5% of gastric adenocarcinomas from the Vancouver group and 20% from Toronto. In both cohorts, loss of BAF250a was positively associated with loss of mismatch repair protein expression (P < .0001 and P = .035, respectively). Loss of BAF250a expression was independently associated with poor overall survival in the Toronto cohort (P = .0015), whereas no significant association with survival was observed in the Vancouver cohort. BAF250a loss was not significantly associated with any additional clinical parameters in either cohort. HER2 amplification was confirmed as a negative prognostic factor in both cohorts. These findings suggest that ARID1A/BAF250a may be of prognostic significance in a subset of patients with early stage gastric cancer and that pathological assessment should increasingly use a multimarker approach.

Review of findings in prophylactic gynaecological specimens in Lynch syndrome with literature review and recommendations for grossing.

AIMS: Prophylactic hysterectomy with bilateral salpingo-oophorectomy is being increasingly undertaken in patients with Lynch syndrome (LS). The pathological features in such specimens are not well described and, unlike the SEE-FIM protocol for salpingo-oophorectomy specimens in BRCA1/2 mutation carriers and the gastrectomy grossing protocols for patients with CDH1 (E-cadherin) mutations, guidelines have not been devised for the grossing of prophylactic gynaecological specimens from LS patients. We aimed to review the pathological findings in a series of prophylactic gynaecological specimens from LS patients and develop guidelines for the grossing of these specimens. METHODS AND RESULTS: We reviewed the pathological findings in 25 prophylactic gynaecological specimens from LS patients and audited the grossing protocols in different centres across Ontario, Canada. We found a 32% incidence of endometrial carcinoma or a precursor lesion; the two endometrial cancers identified were low-grade, low-stage endometrioid adenocarcinomas. To address the absence of guidelines for pathological examination, we undertook a literature review of gynaecological malignancies and incidental findings in prophylactic specimens in LS patients. CONCLUSION: We provide recommendations regarding the grossing of such specimens which includes in-toto examination of the lower uterine segment, endometrium, ovaries and fallopian tubes with representative sampling of the cervix.

Prevalence of loss of expression of DNA mismatch repair proteins in primary epithelial ovarian tumors.

Although different histologic subtypes of epithelial ovarian tumors have long been recognized, their molecular abnormalities have not been fully defined. We examined the prevalence of DNA mismatch repair (MMR) protein loss in these tumors. Tissue microarrays (TMA) of suspected ovarian carcinomas were stained for hMLH1, hMSH2, hMSH6, and hPMS2 and scored separately by 2 groups of investigators. Loss of staining (negative) or discrepant staining results on TMA were verified on whole-section slides. Intact (positive) staining results were also verified for an additional 25 randomly selected cases. Clinical data for cases demonstrating MMR protein loss were collected. A second set of TMA composed purely of mucinous tumors was also stained for antibodies to MMR proteins and scored by 1 group of investigators. TMA was an effective method for screening a large number of ovarian tumors for MMR protein expression, with a sensitivity of 100% for all 4 MMR proteins, and a specificity of 22.2%-53.8% for different MMR proteins. Of the primary epithelial tumors of the ovary, loss of expression of MMR proteins was significantly more common in the endometriosis-associated carcinomas (7/69; 10.1%) than in high-grade serous carcinomas (2/182; 1.1%): P=0.0021. The former group also showed more frequent loss of MMR proteins compared with mucinous intestinal-type carcinomas (0/32; P=0.0940). Cases within the group of endometriosis-associated carcinomas were endometrioid (2/29 cases), clear cell (1/27 cases), undifferentiated (1/8 cases), and mixed carcinomas with an endometrioid, clear cell, and/or undifferentiated component (3/5 cases). No loss of MMR protein expression was identified in epithelial tumors of other histologic subtypes. Our study demonstrated the loss of MMR protein expression in 10.1% of endometriosis-associated ovarian carcinomas. These results raise the possibility of selective screening for Lynch syndrome in patients with these types of ovarian carcinoma.

Safety and effectiveness of radiologic percutaneous gastrostomy and gastro jejunostomy in children with cardiac disease.

OBJECTIVE: The purpose of our study was to evaluate the safety and effectiveness of radiologic percutaneous gastrostomy and gastrojejunostomy for providing nutritional support in children with cardiac disease. MATERIALS AND METHODS: Retrospective chart review of 58 children with cardiac disease who underwent radiologic percutaneous gastrostomy from November 2001 to June 2005 was conducted. Patient data were collected until January 2007. The patients' weights were collected at the time of insertion and 6, 12, 18, and 24 months after insertion, and weight-for-age z-scores were calculated. RESULTS: The mean weight-for-age z-score increased from -2.79 at the time of radiologic percutaneous gastrostomy insertion to -2.33 (p = 0.05) at 6 months after insertion, -1.89 (p = 0.001) at 12 months, -1.65 (p = 0.0002) at 18 months, and -1.40 (p = 0.0004) at 24 months. Repeated measures regression analysis showed a significant increase in weight-for-age z-score over time (p < 0.0001), with an estimated mean increase in weight-for-age z-score of 0.055 per month. No mortality was associated with the insertion or usage of radiologic percutaneous gastrostomy. Major complications included intestinal perforation (3.4%) and aspiration pneumonia (12.1%). CONCLUSION: Radiologic percutaneous gastrostomy is a safe method for providing long-term nutritional support in children with cardiac disease and is effective for improving growth and nutrition in this group of patients.

Disruption of pancreatic beta-cell lipid rafts modifies Kv2.1 channel gating and insulin exocytosis.

In pancreatic beta-cells, the predominant voltage-gated Ca(2+) channel (Ca(V)1.2) and K(+) channel (K(V)2.1) are directly coupled to SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP) receptor) proteins. These SNARE proteins modulate channel expression and gating and closely associate these channels with the insulin secretory vesicles. We show that K(V)2.1 and Ca(V)1.2, but not K(V)1.4, SUR1, or Kir6.2, target to specialized cholesterol-rich lipid raft domains on beta-cell plasma membranes. Similarly, the SNARE proteins syntaxin 1A, SNAP-25, and VAMP-2, but not Munc-13-1 or n-Sec1, are associated with lipid rafts. Disruption of the lipid rafts by depleting membrane cholesterol with methyl-beta-cyclodextrin shunts K(V)2.1, Ca(V)1.2, and SNARE proteins out of lipid rafts. Furthermore, methyl-beta-cyclodextrin inhibits K(V)2.1 but not Ca(V)1.2 channel activity and enhances single-cell exocytic events and insulin secretion. Membrane compartmentalization of ion channels and SNARE proteins in lipid rafts may be critical for the temporal and spatial coordination of insulin release, forming what has been described as the excitosome complex.