Living Donor Liver Transplantation as a Backup Procedure: Treatment Strategy for Hepatocellular Adenomas Requiring Complex Resections.

METHODS: We present a series of three patients with large hepatocellular adenoma lesions showing a central location, for which the living donor liver transplantation strategy was used as a backup procedure. RESULTS: Hepatocellular adenoma was confirmed by biopsy in all patients. Surgical resection was indicated because of the patients' symptoms and lesion size and growth. All patients had a lesion that was central or in close contact with major vessels. The final decision to proceed with the resection was made intraoperatively. A live donor was prepared for all three patients. Two patients underwent portal vein embolization associated with extended hepatectomy, and a total hepatectomy plus liver transplantation with a living donor was performed in one patient. All patients had good postoperative outcomes. CONCLUSIONS: In the treatment of hepatocellular adenomas for which complex resections are necessary and resectability can only be confirmed intraoperatively, surgical safety can be improved through the use of a living donor backup. Center expertise with living donor liver transplantation is paramount for the success of this approach.

Aberrant expression of RSK1 characterizes high-grade gliomas with immune infiltration.

The p90 ribosomal S6 kinase (RSK) family, a downstream target of Ras/extracellular signal-regulated kinase signaling, can mediate cross-talk with the mammalian target of rapamycin complex 1 pathway. As RSK connects two oncogenic pathways in gliomas, we investigated the protein levels of the RSK isoforms RSK1-4 in nontumoral brain (NB) and grade I-IV gliomas. When compared to NB or low-grade gliomas (LGG), a group of glioblastomas (GBMs) that excluded long-survivor cases expressed higher levels of RSK1 (RSK1hi ). No difference was observed in RSK2 median-expression levels among NB and gliomas; however, high levels of RSK2 in GBM (RSK2hi ) were associated with worse survival. RSK4 expression was not detected in any brain tissues, whereas RSK3 expression was very low, with GBM demonstrating the lowest RSK3 protein levels. RSK1hi and, to a lesser extent, RSK2hi GBMs showed higher levels of phosphorylated RSK, which reveals RSK activation. Transcriptome analysis indicated that most RSK1hi GBMs belonged to the mesenchymal subtype, and RSK1 expression strongly correlated with gene expression signature of immune infiltrates, in particular of activated natural killer cells and M2 macrophages. In an independent cohort, we confirmed that RSK1hi GBMs exclude long survivors, and RSK1 expression was associated with high protein levels of the mesenchymal subtype marker lysosomal protein transmembrane 5, as well as with high expression of CD68, which indicated the presence of infiltrating immune cells. An RSK1 signature was obtained based on differentially expressed mRNAs and validated in public glioma datasets. Enrichment of RSK1 signature followed glioma progression, recapitulating RSK1 protein expression, and was associated with worse survival not only in GBM but also in LGG. In conclusion, both RSK1 and RSK2 associate with glioma malignity, but displaying isoform-specific peculiarities. The progression-dependent expression and association with immune infiltration suggest RSK1 as a potential progression marker and therapeutic target for gliomas.

Mutation Detection in Tumor-Derived Cell Free DNA Anticipates Progression in a Patient With Metastatic Colorectal Cancer.

Background: The observation of tumor-derived cell-free DNA (ctDNA) in plasma brought new expectations to monitor treatment response in cancer patients. Case presentation: In an exploratory case of a 57-year-old man diagnosed with metastatic sigmoid adenocarcinoma, we used a hotspot panel of cancer-associated gene mutations to identify tumor-specific mutations in the primary tumor and metastasis. RESULTS: Five mutations were detected (KRAS, p.Gly12Val; TP53, p.Arg175His; RB1, p.Ile680Thr; ALK, p.Gly1184Glu; and ERBB2, p.Lys860Lys), of which three were detected in both tissue types (primary tumor and metastasis). All five mutations were monitored in the ctDNA of six serial plasma samples. Only KRAS and TP53 mutations were detected at a high frequency in the first plasma sample. After 1 month of chemotherapy the allele frequencies of both mutations fell below the detection limit. From the third month of systemic treatment onward, the allele frequencies of both mutations were detectable in plasma, displaying a continual increase thereafter. The remaining three mutations were not detected in plasma samples. Signs of disease progression in ctDNA during the treatment period were evident while computed tomography (CT) measurements suggested stable metastatic lesions throughout the treatment. Conclusions: Liquid biopsies revealed tumor heterogeneity and predicted tumor progression, demonstrating the potential of ctDNA analysis to be a sensitive and specific tool for monitoring treatment responsivity and for early identification of treatment resistance.

Could OX40 agonist antibody promote activation of the anti-tumor immune response in gastric cancer?

BACKGROUND AND OBJECTIVES: OX40, a membrane-bound molecule of the tumor-necrosis-factor-receptor superfamily, is a critical costimulatory receptor during the immune response, especially to T cells, but studies described their presence of OX-40 on neutrophils and monocytes, suggesting a potential role in the activation of immune response. Our aim was to characterize costimulatory receptors OX40 expression on circulating leukocytes in gastric cancer to identify novel targets for immunotherapy. METHODS: Peripheral blood mononuclear cells were isolated from 24 gastric cancer patients and 34 healthy controls and the expression of costimulatory (OX40) receptors were analyzed on T cells, neutrophil and monocyte using monoclonal antibodies by flow cytometry. RESULTS: We found that the higher levels of OX40 + T cells, monocytes/OX40+ and neutrophils/OX40+ from gastric cancer patients when compared to controls (P < 0.0001), and also higher levels of OX40+ T cells when compared to stages III and IV (P = 0.02). Percentage levels of total T cells were similar between patients and controls. CONCLUSIONS: OX40 as a therapeutic agent has been investigated in many preclinical tumor models. Our findings suggest that of levels of costimulatory in T cells in GC will direct future studies on the role that costimulatory receptors play in the failure of T cell-mediated immunity.

Tissue-Associated Bacterial Alterations in Rectal Carcinoma Patients Revealed by 16S rRNA Community Profiling.

Sporadic and inflammatory forms of colorectal cancer (CRC) account for more than 80% of cases. Recent publications have shown mechanistic evidence for the involvement of gut bacteria in the development of both CRC-forms. Whereas, colon and rectal cancer have been routinely studied together as CRC, increasing evidence show these to be distinct diseases. Also, the common use of fecal samples to study microbial communities may reflect disease state but possibly not the tumor microenvironment. We performed this study to evaluate differences in bacterial communities found in tissue samples of 18 rectal-cancer subjects when compared to 18 non-cancer controls. Samples were collected during exploratory colonoscopy (non-cancer group) or during surgery for tumor excision (rectal-cancer group). High throughput 16S rRNA amplicon sequencing of the V4-V5 region was conducted on the Ion PGM platform, reads were filtered using Qiime and clustered using UPARSE. We observed significant increases in species richness and diversity in rectal cancer samples, evidenced by the total number of OTUs and the Shannon and Simpson indexes. Enterotyping analysis divided our cohort into two groups, with the majority of rectal cancer samples clustering into one enterotype, characterized by a greater abundance of Bacteroides and Dorea. At the phylum level, rectal-cancer samples had increased abundance of candidate phylum OD1 (also known as Parcubacteria) whilst non-cancer samples had increased abundance of Planctomycetes. At the genera level, rectal-cancer samples had higher abundances of Bacteroides, Phascolarctobacterium, Parabacteroides, Desulfovibrio, and Odoribacter whereas non-cancer samples had higher abundances of Pseudomonas, Escherichia, Acinetobacter, Lactobacillus, and Bacillus. Two Bacteroides fragilis OTUs were more abundant among rectal-cancer patients seen through 16S rRNA amplicon sequencing, whose presence was confirmed by immunohistochemistry and enrichment verified by digital droplet PCR. Our findings point to increased bacterial richness and diversity in rectal cancer, along with several differences in microbial community composition. Our work is the first to present evidence for a possible role of bacteria such as B. fragilis and the phylum Parcubacteria in rectal cancer, emphasizing the need to study tissue-associated bacteria and specific regions of the gastrointestinal tract in order to better understand the possible links between the microbiota and rectal cancer.

Heterogeneous expression of A33 in colorectal cancer: possible explanation for A33 antibody treatment failure.

The A33 protein, expressed in colorectal tumors, is a target for improving treatment of patients with colorectal cancer. Over the last decade, studies have tested anti-A33 antibody as a therapeutic agent for these patients. Preclinical results were promising, but clinical trials did not confirm positive results. Here, immunohistochemistry in colorectal cancer tissue showed that samples from well-differentiated tumors presented a strong A33 membrane staining, whereas poorly differentiated tumors and mucinous adenocarcinomas showed weak cytoplasmic and nuclear staining. Moderately differentiated tumors presented variable staining. We suggest that in future clinical trials, patients should be selected on the basis of membrane expression of A33.

Identification of protein expression signatures in gastric carcinomas using clustering analysis.

BACKGROUND AND AIM: The identification of gastric carcinomas (GC) has traditionally been based on histomorphology. Recently, DNA microarrays have successfully been used to identify tumors through clustering of the expression profiles. Random forest clustering is widely used for tissue microarrays and other immunohistochemical data, because it handles highly-skewed tumor marker expressions well, and weighs the contribution of each marker according to its relatedness with other tumor markers. In the present study, we identified biologically- and clinically-meaningful groups of GC by hierarchical clustering analysis of immunohistochemical protein expression. METHODS: We selected 28 proteins (p16, p27, p21, cyclin D1, cyclin A, cyclin B1, pRb, p53, c-met, c-erbB-2, vascular endothelial growth factor, transforming growth factor [TGF]-ßI, TGF-ßII, MutS homolog-2, bcl-2, bax, bak, bcl-x, adenomatous polyposis coli, clathrin, E-cadherin, ß-catenin, mucin (MUC)1, MUC2, MUC5AC, MUC6, matrix metalloproteinase [MMP]-2, and MMP-9) to be investigated by immunohistochemistry in 482 GC. The analyses of the data were done using a random forest-clustering method. RESULTS: Proteins related to cell cycle, growth factor, cell motility, cell adhesion, apoptosis, and matrix remodeling were highly expressed in GC. We identified protein expressions associated with poor survival in diffuse-type GC. CONCLUSIONS: Based on the expression analysis of 28 proteins, we identified two groups of GC that could not be explained by any clinicopathological variables, and a subgroup of long-surviving diffuse-type GC patients with a distinct molecular profile. These results provide not only a new molecular basis for understanding the biological properties of GC, but also better prediction of survival than the classic pathological grouping.

GLUT1 expression in malignant tumors and its use as an immunodiagnostic marker.

OBJECTIVE: To analyze glucose transporter 1 expression patterns in malignant tumors of various cell types and evaluate their diagnostic value by immunohistochemistry. INTRODUCTION: Glucose is the major source of energy for cells, and glucose transporter 1 is the most common glucose transporter in humans. Glucose transporter 1 is aberrantly expressed in several tumor types. Studies have implicated glucose transporter 1 expression as a prognostic and diagnostic marker in tumors, primarily in conjunction with positron emission tomography scan data. METHODS: Immunohistochemistry for glucose transporter 1 was performed in tissue microarray slides, comprising 1955 samples of malignant neoplasm from different cell types. RESULTS: Sarcomas, lymphomas, melanomas and hepatoblastomas did not express glucose transporter 1. Forty-seven per cent of prostate adenocarcinomas were positive, as were 29% of thyroid, 10% of gastric and 5% of breast adenocarcinomas. Thirty-six per cent of squamous cell carcinomas of the head and neck were positive, as were 42% of uterine cervix squamous cell carcinomas. Glioblastomas and retinoblastomas showed membranous glucose transporter 1 staining in 18.6% and 9.4% of all cases, respectively. Squamous cell carcinomas displayed membranous expression, whereas adenocarcinomas showed cytoplasmic glucose transporter 1 expression. CONCLUSION: Glucose transporter 1 showed variable expression in various tumor types. Its absence in sarcomas, melanomas, hepatoblastomas and lymphomas suggests that other glucose transporters mediate the glycolytic pathway in these tumors. The data suggest that glucose transporter 1 is a valuable immunohistochemical marker that can be used to identify patients for evaluation by positron emission tomography scan. The function of cytoplasmic glucose transporter 1 in adenocarcinomas must be further examined.

Prognostic implications of altered human epidermal growth factor receptors (HERs) in gastric carcinomas: HER2 and HER3 are predictors of poor outcome.

PURPOSE: The human epidermal growth factor receptor (HER) family consists of four members: ErbB-1 (HER1), ErbB-2 (HER2), ErbB-3 (HER3), and ErbB-4 (HER4). These receptors activate numerous downstream pathways in response to extracellular ligands, regulating diverse processes that include differentiation, migration, proliferation, and survival. Alterations in these genes play a role in the development and progression of many human cancers. In gastric carcinomas (GCs), expression of HER1 and HER2 is thought to be a prognostic factor and target of novel biologic agents. The effect of HER3 or HER4 expression in GC has not been sufficiently studied. In this study, we explored the gene and protein expression of the HER family in GC to establish new potential prognostic factors. PATIENTS AND METHODS: Immunohistochemistry and fluorescence in situ hybridization were performed in 221 patients with GC using tissue microarray. Correlation between the expression or amplification of HER genes and the clinicopathologic parameters was statistically analyzed. RESULTS: Alterations of members of the HER family were significantly associated with the parameters involved in tumor progression, including depth of tumor invasion, involved lymph nodes, and tumor stage. In addition, HER2 amplification and HER3 expression were significantly related to worse survival. CONCLUSION: These results reveal that all members of the HER family are expressed in GC. Furthermore, expression of HER2 and HER3 is a significant predictor of poor survival in GC. Therefore, the development of HER-targeted agents and agents targeting downstream signaling pathways provides new possibilities in the treatment of GC.

GLUT1 expression in malignant tumors and its use as an immunodiagnostic marker

OBJECTIVE: To analyze glucose transporter 1 expression patterns in malignant tumors of various cell types and evaluate their diagnostic value by immunohistochemistry. INTRODUCTION: Glucose is the major source of energy for cells, and glucose transporter 1 is the most common glucose transporter in humans. Glucose transporter 1 is aberrantly expressed in several tumor types. Studies have implicated glucose transporter 1 expression as a prognostic and diagnostic marker in tumors, primarily in conjunction with positron emission tomography scan data. METHODS: Immunohistochemistry for glucose transporter 1 was performed in tissue microarray slides, comprising 1955 samples of malignant neoplasm from different cell types. RESULTS: Sarcomas, lymphomas, melanomas and hepatoblastomas did not express glucose transporter 1. Fortyseven per cent of prostate adenocarcinomas were positive, as were 29 percent of thyroid, 10 percent of gastric and 5 percent of breast adenocarcinomas. Thirty-six per cent of squamous cell carcinomas of the head and neck were positive, as were 42 percent of uterine cervix squamous cell carcinomas. Glioblastomas and retinoblastomas showed membranous glucose transporter 1 staining in 18.6 percent and 9.4 percent of all cases, respectively. Squamous cell carcinomas displayed membranous expression, whereas adenocarcinomas showed cytoplasmic glucose transporter 1 expression. CONCLUSION: Glucose transporter 1 showed variable expression in various tumor types. Its absence in sarcomas, melanomas, hepatoblastomas and lymphomas suggests that other glucose transporters mediate the glycolytic pathway in these tumors. The data suggest that glucose transporter 1 is a valuable immunohistochemical marker that can be used to identify patients for evaluation by positron emission tomography scan. The function of cytoplasmic glucose transporter 1 in adenocarcinomas must be further examined.

Evaluation of cell cycle protein expression in gastric cancer: cyclin B1 expression and its prognostic implication.

The cell cycle progression is regulated by interactions of specific cyclin-dependent kinases at the G1-S and G2-M checkpoints. In addition, the cell cycle dysregulation plays a major role in carcinogenesis of human cancers. To investigate the role of cell cycle regulators in the pathogenesis and progression of human gastric cancer, the expression of cyclin D1, A, B1, p16(INK4a), p21(CPI1), p27(KIP1), p53, and pRb was investigated in 482 gastric carcinomas using immunohistochemistry in terms of histologic type, tumor invasion, size, location, and metastatic behavior. The cyclin D1, A, and B1 expression (>10%) was observed in 49%, 69%, and 49% of the cases, respectively. Negative cases for p16(INK4a), p21(CPI1), and p27(KIP1) were detected in 90%, 86%, and 50.5%. There were 30% and 68% of the gastric tumors positive for p53 and pRb, respectively. Diffuse carcinomas frequently were positive for cyclin B1 and pRb, and negative for p21. A relationship between p53 expression and intestinal type carcinomas was found. In addition, the expression of cyclin B1 was associated with regional lymph node metastasis and poor prognosis. No relationship was noticed between any other cell cycle proteins expression and age, sex, tumor size, tumor location, and lymph node involvement. These findings have shown alterations in several cell cycle regulators, and it was suggested that cyclin B1 expression is closely associated with poor behavior in gastric cancer.

Cadherin-catenin adhesion system and mucin expression: a comparison between young and older patients with gastric carcinoma.

BACKGROUND: Young patients are thought to develop gastric carcinomas with a molecular genetic profile that is distinct from that of gastric carcinomas occurring at a later age. The aim of this study was to compare the clinicopathological features and expression patterns of the markers E-cadherin and beta-catenin, and mucins (MUC1, MUC2, MUC5AC, and MUC6) in young and older patients. METHODS: The clinicopathological features and overall survival data of 62 young patients (age 40 years). A tissue microarray method and immunohistochemistry were used in order to analyze marker expression in paraffin-embedded tissue blocks obtained from both groups. RESULTS: The young group presented a higher percentage of diffuse-type tumors in comparison to the older group (P<0.01). The rates of positivity for E-cadherin and beta-catenin membranous expression patterns and mucin (MUC2, MUC5AC and MUC6) positivity were higher in the young group (P<0.01). Although young patients showed a lower frequency of alterations in marker expression and had significantly better survival rates than the older patients, neither age nor the marker expression pattern were found to be independent prognostic factors of survival. Only stage, tumor size, and tumor location persisted as prognostic factors for patients with gastric cancer. CONCLUSION: Biological markers of cellular adhesion and gastric differentiation were differently expressed in young and older patients. Our findings support the hypothesis that young patients develop carcinomas with a different genetic pathway compared to the pathway of tumors occurring at a later age, and we suggest further investigations to assess the prognostic relevance of the markers to specific subgroups.

E-cadherin/beta-catenin expression pattern, clinicopathological parameters and prognosis in gastric carcinomas

Objectives: This study aimed to evaluate the expression pattern of some markers (E-cadherin and â-catenin) related to cellular adhesion and their relationship with histological tumor type according to Laurén’s system, clinicopathological features and patient survival. Material and Methods: We did immunohistochemical analysis in a retrospective series of 446 gastric carcinomas using tissue microarray method (TMA). Clinicopathological features and overall survival data of all patients were retrospectively reviewed from hospital records. For all statistical analyses, p<0.05 was considered significant. Results: The reduced/absent expression of E-cadherin occurred more frequently in diffuse than intestinal type tumors and it was correlated with worse biological behavior and poor prognosis for patients with diffuse type gastric carcinomas. The pattern of â-catenin expression was closely related to histological type and E-cadherin expression. Although patients with nuclear/absent â-catenin immunoreactivity showed worse survival index, no statistical correlation was found with overall survival. In multivariated analysis, only pTNM staging system persisted as independent prognostic marker. Conclusion: In the present study, alterations in E-cadherin/â-catenin complex expression showed significant correlations with clinicopathological parameters, as well as its implications for tumor progression and prognosis in gastric cancer. Our results indicate that markers expression pattern may be a useful marker of differentiation and suggest further investigations of their prognostic relevance to specific histological groups.

Differential expression of apoptosis related proteins and nitric oxide synthases in Epstein Barr associated gastric carcinomas.

AIM: To determine the incidence of Epstein Barr virus associated gastric carcinoma (GC) in Brazil and compare the expressions of apoptosis related proteins and nitric oxide synthases between EBV positive and negative gastric carcinoma. METHODS: In situ hybridization of EBV-encoded small RNA-1 (EBER-1) and PCR was performed to identify the presence of EBV in GCs. Immunohistochemistry was used to identify expressions of bcl-2, bcl-xl, bak, bax, p53, NOS-1, NOS-2, and NOS-3 proteins in 25 EBV positive GCs and in 103 EBV negative GCS. RESULTS: 12% of the cases of GC (25/208) showed EBER-1 and EBNA-1 expression. The cases were preferentially of diffuse type with intense lymphoid infiltrate in the stroma. EBV associated GCs showed higher expression of bcl-2 protein and lower expression of bak protein than in EBV negative GCs. Indeed, expressions of NOS-1 and NOS-3 were frequently observed in EBV associated GCs. CONCLUSION: Our data suggest that EBV infection may protect tumor cells from apoptosis, giving them the capacity for permanent cell cycling and proliferation. In addition, EBV positive GCs show high expression of constitutive NOS that could influence tumor progression and aggressiveness.

Molecular classifiers for gastric cancer and nonmalignant diseases of the gastric mucosa.

High incidence of gastric cancer-related death is mainly due to diagnosis at an advanced stage in addition to the lack of adequate neoadjuvant therapy. Hence, new tools aimed at early diagnosis would have a positive impact in the outcome of the disease. Using cDNA arrays having 376 genes either identified previously as altered in gastric tumors or known to be altered in human cancer, we determined expression signature of 99 tissue fragments representing normal gastric mucosa, gastritis, intestinal metaplasia, and adenocarcinomas. We first validated the array by identifying molecular markers that are associated with intestinal metaplasia, considered as a transition stage of gastric adenocarcinomas of the intestinal type as well as markers that are associated with diffuse type of gastric adenocarcinomas. Next, we applied Fisher's linear discriminant analysis in an exhaustive search of trios of genes that could be used to build classifiers for class distinction. Many classifiers could distinguish between normal and tumor samples, whereas, for the distinction of gastritis from tumor and for metaplasia from tumor, fewer classifiers were identified. Statistical validations showed that trios that discriminate between normal and tumor samples are powerful classifiers to distinguish between tumor and nontumor samples. More relevant, it was possible to identify samples of intestinal metaplasia that have expression signature resembling that of an adenocarcinoma and can now be used for follow-up of patients to determine their potential as a prognostic test for malignant transformation.