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The objective of this study was to elucidate the association between neoplastic progression and somatic copy number alterations (SCNAs) occurring within the same colorectal cancer (CRC) tumor.We investigated SCNAs to identify the progression from a high-grade intramucosal lesion (HGIL) to an invasive front lesion (IFL), via an invasive submucosal lesion (ISL), within the same tumor using a crypt isolation method combined with a SNP array. Immunohistochemistry was also performed.We identified 51 amplified genes that potentially promote progression from HGIL to ISL and 6 amplified genes involved in the progression from ISL to IFL. Of the 51 genes involved in HGIL to ISL progression, TORC1, MSLN, and STUB1, which are closely associated with CRC, were identified as candidate markers of submucosal invasion. However, no candidate genes were identified among the six genes associated with ISL to IFL progression. In addition, the number of total SCNAs and the number of gains were correlated with cancer progression (from HGIL to IFL). Finally, immunohistochemistry revealed higher expression of TORC1, MSLN, and STUB1 in ISL than in HGIL.These results suggest that specific SCNAs are required for acquisition of invasive ability in CRC, and the affected genes are potential markers of invasion.

Fibroblast-activating protein (FAP) is expressed in cancer-associated fibroblasts (CAFs) in many human carcinomas and in some types of carcinoma cells. Here, we examined the proportion of FAP protein expression in non-small cell lung carcinoma (NSCLC) and investigated the correlation of FAP expression with clinicopathological background.In total, 344 NSCLC tissues were examined. Tissue microarrays were constructed, and FAP expression was analyzed using immunohistochemistry. The status of FAP expression in tumor cells and CAFs was correlated with clinicopathological background, molecular features, and patient outcomes.A total of 280 patients (81.4%) had low FAP expression, and 64 patients (18.6%) had high FAP expression in tumor cells. In CAFs, 230 patients (66.9%) had low FAP expression, and 114 patients (33.1%) had high FAP expression. In multivariate analyses, high FAP expression in tumor cells was an independent predictive factor of both overall survival (OS; hazard ratio [HR] = 2.57, 95% confidence interval [CI]: 1.49-4.42, p 0.001) and recurrence-free survival (RFS; HR = 2.13, 95% CI: 1.38-3.29, p 0.001). Based on combinations of FAP expression in tumor cells and CAFs, patients with LowOverall, FAP expression in tumor cells and the combination FAP expression in tumor cells and CAFs were strongly associated with patient survival and may be useful predictive biomarkers for patient outcomes in NSCLC.

No effective early diagnostic biomarkers are available for colorectal cancer (CRC). Therefore, we sought to identify new biomarkers that could identify CRC from progression as a pre-cancerous lesion to its invasive form. Recent studies have shown that microRNAs (miRs) are associated with the onset of cancer invasion and progression.We hypothesized that the identification of miRs associated with CRC might be useful to detect this disease at early stages.We conducted an integrated analysis of 79 isolated colorectal tumor glands, including adenomas, intramucosal cancers, and invasive CRCs that showed a microsatellite stable phenotype using GeneChip miRNA 4.0 microarray assays. The colorectal tumors we examined were divided into 2 cohorts (42 in the first cohort and 37 in the second cohort).First, cluster analysis was performed to stratify expression patterns of multiple miRs that were pooled according to the following criteria: fold change in expression ( -2.0 or 2.0), p 0.05, and mature miRs. As a result, the expression patterns of pooled miRs were subdivided into 3 subgroups that were correlated with tumor grade. Each subgroup was characterized by specific miRs. In addition, we found that specific miRs, including miR-140-3p and miR-378i, were closely associated with cancer invasion. Finally, we analyzed paired dysregulated miRs between adenomatous and cancerous components present within the same tumor.We showed that several miRs were dysregulated during progression from adenoma to intramucosal cancer. Specific miRs may have key roles in progression from intramucosal tumor to invasive CRC.

BACKGROUND: Squamous cell carcinoma (SCC) is the major histological type in lung cancer (LC). The tumor microenvironment (TME) drives tumor progression and metastasis. In the TME, cancer-associated fibroblasts (CAFs) play key roles in carcinogenesis. However, the roles of CAFs in lung SCC remain unknown. In this study, we evaluated whether the CAF phenotype was determined by various CAF-related proteins and whether CAF-related protein expression contributed to clinical outcomes in patients with lung SCC. METHODS: We examined the associations of CAF- and epithelial-mesenchymal transition (EMT)-related markers expressed in CAFs, including α-smooth muscle actin (α-SMA), CD10, podoplanin, fibroblast-specific protein 1 (FSP1), platelet-derived growth factor receptor (PDGFR) α, PDGFRβ, adipocyte enhancer-binding protein 1 (AEBP1), fibroblast activation protein (FAP), tenascin-C, Zinc finger E-box binding homeobox 1 (ZEB1), and twist homolog 1 gene (TWIST1), in 108 lung SCC tissues using immunohistochemistry. In addition, cluster analysis was used to identify objective expression patterns of immunohistochemical markers. Finally, the CD3/CD8 ratio was evaluated in order to identify the associations of CAF-related proteins with the CD3/CD8 ratio using immunohistochemistry. RESULTS: SCC samples were classified into two subgroups (CAF-phenotype), which were significantly correlated with disease-free and overall survival using univariate and multivariate analyses. Moreover, high AEBP1 expression was identified as an independent prognostic marker in this cohort by univariate and multivariate analyses. The CD3/CD8 ratio was not correlated with the CAF-phenotype. CONCLUSIONS: The presence of a specific subgroup defined by multiple markers could be used for prediction of prognosis in patients with lung SCC. In addition, AEBP1 overexpression played key roles in prediction of a poor prognosis in patients with lung SCC.

Background Gastroblastoma is a rare gastric tumor composed of epithelial and spindle cell components. The characteristic MALAT–GLI1 fusion gene has only been identified in 5 reported cases. We report the morphological characterization of gastroblastoma with the MALAT1–GLI1 fusion gene in a young Japanese woman. Case presentation A 29-year-old Japanese woman visited Iwate Medical University Hospital with upper abdominal pain. Computed tomography revealed a tumor in expansive lesions involving the gastric antrum. Histologically, we observed a biphasic morphology composed of epithelial and spindle cell components. The epithelial components appeared as slit-like glandular structures with tubular or rosette-like differentiation. The spindle cell components consisted of short spindle-shaped oval cells. Immunohistochemical (IHC) analysis revealed that the spindle cell component was positive for vimentin, CD10, CD56, GLI1, and HDAC2, and focally positive for PD-L1. The epithelial component was positive for CK AE1/AE3, CAM5.2, and CK7, and negative for CK20 and EMA. Both components were negative for KIT, CD34, DOG1, SMA, desmin, S100 protein, chromogranin A, synaptophysin, CDX2, and SS18-SSX. The MALAT-GLI1 fusion gene was detected molecularly. Conclusions We report the following new findings with this case: (i) gastric tumors mimic the gastrointestinal mesenchyme in the embryonic period; (ii) nuclear expression of PD-L1 and HDAC2 were observed in the spindle cell component of a gastroblastoma. We speculate that histone deacetylase (HDAC) inhibitors may offer a promising treatment option for gastroblastoma.

Serous carcinoma of the uterus (USC) is a pathological subtype of high-grade endometrial cancers, with no effective treatment for advanced cases. Since such refractory tumors frequently harbor antitumor immune tolerance, many immunotherapies have been investigated for various malignant tumors using immuno-competent animal models mimicking their local immunities. In this study, we established an orthotopic mouse model of high-grade endometrial cancer and evaluated the local tumor immunity to explore the efficacy of immunotherapies against USC. A multivariate analysis of 62 human USC cases revealed that the tumor-infiltrating cell status, few CD8+ cells and abundant myeloid-derived suppressor cells (MDSCs), was an independent prognostic factor (P < 0.005). A murine endometrial cancer cell (mECC) was obtained from C57BL/6 mice via endometrium-specific deletion of Pten and Tp53, and another high-grade cell (HPmECC) was established by further overexpressing Myc in mECCs. HPmECCs exhibited higher capacities of migration and anchorage-independent proliferation than mECCs (P < 0.01, P < 0.0001), and when both types of cells were inoculated into the uterus of C57BL/6 mice, the prognosis of mice bearing HPmECC-derived tumors was significantly poorer (P < 0.001). Histopathological analysis of HPmECC orthotopic tumors showed serous carcinoma-like features with prominent tumor infiltration of MDSCs (P < 0.05), and anti-Gr-1 antibody treatment significantly prolonged the prognosis of HPmECC-derived tumor-bearing mice (P < 0.05). High CCL7 expression was observed in human USC and HPmECC, and MDSCs migration was promoted in a CCL7 concentration-dependent manner. These results indicate that antitumor immunity is suppressed in USC due to increased number of tumor-infiltrating MDSCs via CCL signal.

mRNA expression varies in human cancers. Such altered mRNA expression is negatively regulated by the expression of microRNAs (miRNAs), which play an important role in human tumorigenesis. According to this theory, inverse mRNA/miRNA expression may be a direct driver of cancer development, and certain genetic events may occur prior to the development of any discernible histological abnormalities. We examined the inverse expression between mRNAs and their corresponding miRNAs in colorectal cancer (CRC) and adjacent normal mucosa and performed pathway analysis to identify mRNA/miRNA networks. The cancer samples were divided into first (20 cases) and second (24 cases) cohorts, and 48 samples were obtained from two sections of the normal mucosa adjacent to the tumors from the second cohort. We investigated mRNAs with commonly altered expression in CRC and adjacent normal mucosa using isolated cancer glands and normal crypts from the first cohort, compared with that of distal normal crypts, using an array-based method. As a result, significant inverse correlations between CEACAM1 and miRNA-7114-5p and between AK1 and miRNA-6780-5p were found in CRC and adjacent normal mucosa. We validated these correlations in the second cohort using RT-PCR. To confirm these findings, transfection and immunohistochemical assays were also performed, which verified the inverse correlation between CEACAM1 and miRNA-7114-5p. Our findings suggest that the inverse correlations between the CEACAM1/miRNA-7114-5p and possibly AK1/miRNA-6780-5p pairs play an important role in early CRC development, and may help identify potential molecular targets for early detection of CRC.

MicroRNA (miRNA) expression is dysregulated in human tumors, thereby contributing to tumorigenesis through altered expression of mRNA. Thus, identification of the relationships between miRNAs and mRNAs is important for evaluating the molecular mechanisms of tumors. Additionally, elucidation of the molecular features of serrated lesions is essential in colorectal tumorigenesis. Here, we examined the relationships of miRNA and mRNA expressed in serrated lesions, including 26 sessile serrated lesions (SSLs), 12 traditional serrated adenomas (TSAs), and 11 colorectal cancers (CRCs) with a microsatellite instability (MSI) phenotype using crypt isolation. We divided the samples into the first and second cohorts for validation. Array-based expression analyses were used to evaluate miRNAs and mRNAs with opposite expression patterns in isolated tumor glands. In addition, we validated the relationships of miRNA/mRNA pairs in the second cohort using real-time polymerase chain reaction. We found that the expression of miRNA-5787 was correlated with reciprocal expression of 2 mRNAs, i.e., SRRM2 and POLR2J3, in SSL samples. In TSA samples, 2 pairs of miRNAs/mRNAs showing opposite expression patterns, i.e., miRNA-182-5p/ETF1 and miRNA-200b-3p/MYB, were identified. Ultimately, three pairs of miRNAs/mRNAs with opposite expression patterns, including miRNA-222-3p/SLC26A3, miRNA-6753-3p/FABP1, and miRNA-222-3p/OLFM4, were retained in CRC with an MSI phenotype. Finally, we performed transfection with an miR-222-3p mimic to confirm the expression of SLC26A3 and OLFM4; the results showed that ectopic expression of miR-222-3p moderately suppressed OLFM4 and downregulated SLC26A3 to some extent. Overall, our results provided basic insights into the evaluation of colorectal tumorigenesis of serrated lesions and CRC with an MSI phenotype. This article is protected by copyright. All rights reserved.

Background Gastroblastoma is a rare gastric tumor composed of epithelial and spindle cell components. Identification of the characteristic MALAT–GLI1 fusion gene has only been reported in 5 cases. We report the morphological characterization of gastroblastoma with the MALAT1–GLI1 fusion gene in a young Japanese woman. Case presentation A 29-year-old Japanese woman visited Iwate Medical University Hospital with upper abdominal pain. Computed tomography revealed a tumor in the expansive lesions involving the gastric antrum. Histologically, we observed a biphasic morphology composed of epithelial and spindle cell components. The epithelial components appeared as slit-like glandular structures and tubular or rosette-like differentiation. The spindle cell components consisted of short spindle-shaped oval cells. Immunohistochemical (IHC) analysis revealed that the spindle cell component was positive for vimentin, CD10, CD56, GLI1, HDAC2 and focally positive for PD-L1. The epithelial component was positive for CK AE1AE3, CAM5.2, CK7, and negative for CK20 and EMA. Both components were negative for KIT, CD34, DOG1, SMA, desmin, S100 protein, chromogranin A, synaptophysin, CDX2, and SS18-SSX. Finally, MALAT-GLI1 fusion gene was molecularly detected. Conclusions We report the following new findings with this case: (i) gastric tumors mimic the gastrointestinal mesenchyme in the embryonic period; (ii) nuclear expression of PD-L1 and HDAC2 were observed in the spindle cell component of gastroblastoma. We speculate that HDAC (Histone Deacetylase) inhibitors may offer a promising treatment option for gastroblastoma.

Cancer-associated fibroblasts (CAFs) are a prominent component in the tumor microenvironment (TME), which plays an important role in lung carcinogenesis. Here, we investigated microenvironmental markers expressed by CAFs, including α-smooth muscle actin, CD10, podoplanin, fibroblast-specific protein 1, platelet-derived growth factor α and β, fibroblast-associated protein, tenascin-C, zinc finger E-box binding homeobox 1 (ZEB1), and twist-related protein 1 expression levels. We evaluated samples from 257 patients with lung adenocarcinoma (LAD) to assess the associations of CAF-related protein expression patterns with prognosis. LAD cases were stratified using cluster analysis. To determine the utility of prognostic markers in LAD, univariate and multivariate analyses were performed. LAD cases were classified into subgroups 1 and 2. Subgroup 2 was shown to be significantly correlated with disease-free and overall survival using univariate and multivariate analyses in this group. Upregulation of podoplanin was identified as a single prognostic marker in this study by univariate and multivariate analyses. In addition, ZEB1 overexpression was correlated with disease-free survival. Our current results suggested that the specific CAF phenotype (e.g., the expression pattern of CAF-related proteins) could predict outcomes in patients with LAD. In addition, podoplanin upregulation may predict outcomes in these patients.

Macrophage polarization is an important pathogenetic factor in neoplastic diseases and is regulated by transcription factors, i.e., phosphorylated signal transducer and activator of transcription 1 (phospho-STAT1) for the M1 phenotype and c-Maf for the M2 phenotype. However, the role of macrophage phenotype in lung adenocarcinoma (LAD) remains unclear. Here, we examined whether the density of M1 and M2 macrophages was associated with prognosis in patients with LAD using double-labeling immunohistochemistry for the detection of macrophage markers. Additionally, programmed death ligand 1 (PD-L1) expression was investigated. Immune cells co-expressing CD68 and phospho-STAT1 were considered M1 macrophages, whereas those co-expressing CD68 and c-Maf were recognized as M2 macrophages. Three hundred seven patients with LAD were divided into two cohorts (N = 100 and 207 cases, respectively) to evaluate the associations of M1 and M2 phenotypes with prognosis in patients with LAD. We determined the cut-off values of CD68/phospho-STAT1-positive cells (5 or less) and CD68/c-Maf-positive cells (more than 11) to assess correlations with survival using receiver operating characteristic curve analysis in the first cohort. According to the cut-off values, high expression of CD68/c-Maf was identified as an independent prognostic marker, whereas CD68/Phospho-STAT1 expression was inversely correlated with patient outcomes. Moreover, the M1/M2 ratio (0.19 or less) was a poor prognostic factor for LAD. However, PD-L1 expression was not correlated with patient outcomes. Overall, these findings suggested that double staining of markers identifying M1 and M2 macrophages, including phospho-STAT1 and c-Maf, can be used as prognostic indicators for patients with LAD.

Identification of molecular alterations occurring in the adenomatous and carcinomatous components within the same tumor would greatly enhance understanding of the neoplastic progression of colorectal cancer. We examined somatic copy number alterations (SCNAs) and mRNA expression at the corresponding loci involved in the adenoma–carcinoma sequence in the isolated adenomatous and cancer glands of the same tumor in 15 cases of microsatellite‐stable “carcinoma in adenoma,” using genome‐wide SNP and global gene expression arrays. Multiple copy‐neutral loss of heterozygosity events were detected at 4q13.2, 15q15.1, and 14q24.3 in the adenomatous component and at 4q13.2, 15q15.1, and 14q24.3 in the carcinomatous component. There were significant differences in the copy number (CN) gain frequencies at 20q11.21–q13.33, 8q13.3, 8p23.1, and 8q21.2–q22.2 between the adenomatous and carcinomatous components. Finally, we found a high frequency of five genotypes involving CN gain with upregulated expression of the corresponding gene (RPS21, MIR3654, RSP20, SNORD54, or ASPH) in the carcinomatous component, whereas none of these genotypes were detected in the adenomatous component. This finding is interesting in that CN gain with upregulated gene expression may enhance gene function and play a crucial role in the progression of an adenoma into a carcinomatous lesion.

BackgroundAlthough MicroRNAs (miRNAs) play important roles in various biological processes, the biological functions of miRNAs are achieved through mRNAs. The aim of this study is to identify dysregulated miRNA/mRNA expression patterns in colorectal tumors.MethodsWe examined 42 colorectal tumors [15 adenomas, 8 intramucosal cancers (IMCs), and 19 invasive colorectal cancers (CRCs)] with the microsatellite stable (MSS) phenotype (first cohort). The first cohort was used for genome-wide miRNA and mRNA expression arrays, whereas the second cohort (37 colorectal neoplasias) was used for validation analyses. Finally, we used 15 cases of “adenoma in/with carcinoma” to identify network patterns of miRNAs/mRNAs that were directly associated with neoplastic progression. In addition, simple regression analysis for array-based and RT-PCR analyses was performed to select candidate miRNA–mRNA pairs. Transfection of miRNA mimics was also performed to confirm whether target mRNA expression is affected by specific miRNAs.ResultsSpecific paired miRNA/mRNA networks, including hsa-miR-34a-5p/SLC12A2, hsa-miR-15b-5p/SLC12A2, hsa-miR-195-5p/SLC12A2, hsa-miRNA-502-3p/OLFM4, hsa-miRNA-6807-5p/ZG16, and hsa-miRNA 3064-5p/SH3BGRL3, were identified in samples of adenoma, IMC, and CRC with the MSS phenotype. In adenomatous lesions obtained from the same tumor with a carcinomatous lesion, we identified pairs of miRNA-130a-3p/HSPA8 and miRNA-22-3p/RP53 that were linked to multiple pathways. On the other hand, 2 pairs of miRNA/mRNA (miRNA-660-5p and miRNA-664a-5p/APP) were found in isolated carcinomatous glands. Ectopic expression of miRNA 3064-5p suppressed SH3BGRL3 expression.ConclusionsWe found that networks based on specific pairs of miRNAs/mRNAs contribute to progression from adenomatous and carcinomatous lesions. Our results provide insights into the molecular tumorigenesis of colorectal tumors.

Background Metastasis of colorectal cancer (CRC) is a major cause of CRC-related mortality. However, the detailed molecular mechanism of CRC metastasis remains unknown. A recent study showed that the tumor microenvironment, which includes cancer cells and the surrounding stromal cells, plays a major role in tumor invasion and metastasis. Identification of altered messenger RNA (mRNA) expression in the tumor microenvironment is essential to elucidation of the mechanisms responsible for tumor progression. This study investigated the mRNA expression of genes closely associated with metastatic CRC compared with non-metastatic CRC. Methods The samples examined were divided into cancer tissue and isolated cancer stromal tissue. The study examined altered mRNA expression in the cancer tissues using The Cancer Genome Atlas (TCGA) (377cases) and in 17 stromal tissues obtained from our laboratory via stromal isolation using an array-based analysis. In addition, 259 patients with CRC were enrolled to identify the association of the candidate markers identified with the prognosis of patients with stage 2 or 3 CRC. The study examined the enriched pathways identified by gene set enrichment analysis (GSEA) based on the Kyoto Encyclopedia of Genes and Genomes (KEGG) module in both the TCGA dataset and isolated stromal tissue. Results As a result, whereas tenascin-C, secreted phosphoprotein 1 and laminin were expressed in metastatic CRC cells, olfactory receptors (ORs) 11H1 and OR11H4 were expressed in stromal tissue cells isolated from metastatic CRC cases. Finally, upregulated expression of tenascin-C and OR11H4 was correlated with the outcome for CRC patients. Conclusion The authors suggest that upregulated expression levels of tenascin-C and OR11H1 play an important role in CRC progression.

The International Association for the Study of Lung Cancer (IASLC) Pathology Committee recently proposed a new histological grading system for invasive lung adenocarcinoma (ADC). This study evaluated the usefulness of this grading system.A total of 395 patients with ADC were examined. ADCs were reclassified based on comprehensive histological subtyping according to the IASLC grading system. We evaluated the following histological grading systems for invasive ADC: the architectural (Arch), Sica's grading, and IASLC grading systems. Multivariate analyses of overall and recurrence-free survival (RFS) based on these three grading systems were performed using Cox proportional hazards models.Multivariate analysis showed that all three grading systems were useful for predicting the outcomes of patients at all stages. However, the IASLC grading system was superior to the Arch and Sica's grading systems in differentiating grade 3 from grade 1 ADCs in terms of both overall survivals (IASLC vs. Arch vs. Sica's grading systems: hazard ratio [HR] = 3.77 vs. 3.03 vs. 2.63) and RFS (HR = 4.25 vs. 2.69 vs. 2.4).The newly proposed IASLC grading system was useful for predicting patient outcomes and was superior to the other grading systems in detecting high-grade malignancy.

Introduction. Carcinoma with sarcomatoid components is a highly malignant tumor exhibiting both epithelial and stromal malignant differentiation. Its tumorigenesis is associated with epithelial–mesenchymal transition (EMT), and phenotypic changes from carcinoma to sarcoma are associated with TP53 mutations. Case presentation. A 73-year-old female with bloody stool was diagnosed with rectal adenocarcinoma. She underwent trans-anal mucosal resection. Histopathologically, the tumor cells showed 2 morphologically distinct populations. One was composed of well-formed to fused glands or cribriform glands and was considered a moderately differentiated adenocarcinoma. The other consisted of pleomorphic discohesive atypical tumor cells with spindle and/or giant cell features, which was considered a sarcomatous tumor. Immunohistochemistry analysis showed that E-cadherin expression changed from positive to negative in the sarcomatous component. On the other hand, ZEB1 and SLUG were positive. Finally, she was diagnosed with carcinoma with a sarcomatoid component. We performed a mutation analysis by next genome sequencing and found KRAS and TP53 mutations in both carcinomatous and sarcomatous components. Conclusions. Immunohistochemistry and mutation analyses revealed tumorigenesis of rectal carcinoma with sarcomatoid components correlated with EMT and TP53 mutations.

A 49 years old male, who had had postprandial dysphagia during the preceding 10 years, was referred to our hospital. Esophagogastroduodenoscopy (EGD) revealed longitudinal furrows and concentric rings in the mid to lower esophagus and stenosis in the lower esophagus. Histologic findings from esophageal biopsies showed eosinophilic infiltration (> 15 per high-power field). Under a diagnosis of eosinophilic esophagitis, an endoscopic bougie was performed, which resulted in symptomatic relief. Follow-up EGD revealed that the stenosis had improved, but histologic findings of eosinophilic esophagitis were remaining. Our case suggests that although rare, esophageal stenosis occurs in Japanese patients with EoE, and that the complication may be a consequence of prolonged disease. Other risks and the appropriate treatment for the prevention of stenosis need to be elucidated further.

Colorectal adenocarcinoma (CRA) is characterized by marked heterogeneity and may be composed of an admixture of various histologic patterns, including well-formed gland and cribriform types. Although tumors displaying a prominent or predominant cribriform feature are frequently found in CRA, this type may contain specific histologic variants with a characteristic molecular alteration. We investigated the molecular features of 51 primary CRAs with a predominant cribriform histology using array-based analyses [somatic copy number alterations (SCNAs); mRNA expression]. Mutations (TP53, KRAS, PIK3CA and BRAF) and DNA methylation status were also analyzed. The crypt isolation method was used to obtain isolated tumor glands of each type separately. All patients were classified by their CRA histologic subtype into two groups: well-formed gland and cribriform. Next, we performed cluster analysis to stratify SCNA and mRNA expression patterns between the two subtypes. Two distinctive subgroups were stratified based on patterns of SCNA and mRNA expression and were correlated with each histologic subtype. The cribriform type was characterized by a high frequency of SCNA compared with that of the well-formed gland type and was closely associated with the expression of specific mRNAs. In addition, the frequency of KRAS mutation was significantly higher in the cribriform type than in the well-formed gland type. Finally, there was no difference in DNA methylation status between the two subtypes. Overall, these data suggest that the cribriform type provides important insights into colorectal carcinogenesis, suggesting specific potential histologic implications based on the molecular profile.

Aim: Biliary tract carcinoma (BTC), including gall bladder carcinoma (GBC) and biliary duct carcinoma (BDC), has a poor prognosis. Comprehensive genomic profiling has important roles in evaluation of the carcinogenesis of BTC. Materials & methods: We examined somatic copy number alterations (SCNAs) using a single nucleotide polymorphism array system to analyze 36 BTC samples (11 GBCs and 25 BDCs). Results: In hierarchical cluster analysis, two clusters were identified (subgroup 1 with low SCNAs and subgroup 2 with high SCNAs). GBC was predominant in subgroup 1, whereas BDC was predominant in subgroup 2, suggesting that GBC and BDC had different genetic backgrounds in terms of SCNAs. Conclusion: These findings could be helpful for establishing the molecular carcinogenesis of BTCs., Lay abstract Biliary tract carcinoma, including gall bladder carcinoma (GBC) and biliary duct carcinoma (BDC), has a poor prognosis. Comprehensive genomic (single nucleotide polymorphism-array) profiling plays important roles in evaluation of the carcinogenesis of biliary tract carcinoma. In the hierarchical cluster analysis, two clusters were identified (subgroup 1 with low somatic copy number alterations [SCNAs] and subgroup 2 with high SCNAs); GBC was found to be predominant in subgroup 1, whereas BDC was predominant in subgroup 2. These findings suggested that GBC and BDC had different genetic backgrounds in terms of SCNAs.

Background Low-grade fetal adenocarcinoma of the lung (L-FLAC) is a rare pulmonary tumor resembling fetal lung histologically. Due to its rarity, there is limited information about L-FLAC pathogenesis and biological characteristics. Here, we describe two cases of L-FLAC treated at our hospital and summarize L-FLAC cases reported in the literature. Case presentation: We examined one woman and one man who were 30-years-old and 67-years-old, respectively. Histologically, tumor tissue from both cases had a complex glandular component with clear cuboidal and columnar cells that resembled histological features of fetal lung. In some areas, squamous morules were prominent. Immunohistochemically, nuclear/cytoplasmic expression of β-catenin was detected in both cases. Mutation analysis revealed a CTNNB1 mutation in both cases and a DICER1 mutation in 1 case. No mutations in EGFR, BRAF, KRAS, PIK3CA mutation were found. Conclusions L-FLAC showed a high frequency of CTNNB1 mutation and low frequency of EGFR, KRAS, BRAF and PIK3CA mutation in our examined cases and in previous studies. This rare tumor has unique clinicopathological characteristics with specific genetic aberrations involving the Wnt pathway. These results provide a molecular basis for development of new therapies to treat these tumors.

IntroductionDNA mismatch repair (MMR) deficiency leads to changes in the length of nucleotide repeat sequences of tumor DNA. In that situation, DNA replicational errors occur and accumulate during DNA replication. As a result, this mechanism frequently affects the coding regions of oncogenes and tumor suppressor genes and causes carcinogenesis. Recently, DNA MMR deficiency has been recognized as a predictive biomarker for immunotherapy. The aim of this study is to examine the frequency of DNA MMR deficiency and clinicopathological characteristics in surgically resected lung carcinoma (LC) and their correlation.MethodsA total of 1153 LCs were examined. Tissue microarrays were constructed. The status of MMR deficiency was evaluated by immunohistochemical analysis of MMR protein expression (hMLH1, hMSH2, hMSH6, and hPMS2). Microsatellite instability analysis, BRAF mutation, and MLH1 methylation analysis were performed for cases that showed MMR deficiency.ResultsOnly 2 of the 1153 cases (0.17%) showed a loss of hMLH1/hPMS2 protein expression. They also had high levels of microsatellite instability (MSI-H), had neither MLH1 promoter methylation nor BRAF mutation, and were male smokers. Histopathologically, one was a squamous cell carcinoma, and the other was combined small cell carcinoma with squamous cell carcinoma. Regarding PD-L1 protein expression, one had high expression, and the other had none.ConclusionThe frequency of MMR deficiency was very low in LC. However, our two cases were non-adenocarcinoma and differed from previous studies. Because of its very low frequency, MMR deficiency is not a practical biomarker to predict the effect of immune checkpoint inhibitors in LC.

Precursor lesions that progress into colorectal cancer (CRC) could be largely classified into sessile serrated lesions (SSLs), traditional serrated adenoma (TSA), and tubular adenoma (TA). We aimed to determine whether high expression of trefoil factor 1 (TFF1) is closely associated with serrated lesions, particularly SSLs. The samples were divided into the first (12 SSLs, 5 TSAs, and 15 TAs) and second cohorts (15 SSLs, 9 TSAs, and 15 TAs). First, we investigated TFF1 expression in isolated gland samples using array-based and reverse-transcription PCR. Second, we performed immunohistochemical analysis of TFF1 expression in paraffin-embedded tissues obtained from SSL, TSA, TA, and hyperplastic polyp (HP) samples. In addition, we compared TFF1 mRNA levels between SSLs and HPs. TFF1 expression was significantly higher in SSLs than in TSA and TA in both cohorts. Additionally, immunohistochemical staining of TFF1 in the HP, SSL, TSA, and TA samples revealed significant differences in the immunohistochemical scores of TFF1 among the four types of lesions (higher expression in SSLs than in the other three lesions). Finally, there were significant differences in TFF1 mRNA expression levels between SSLs and HPs in paraffin-embedded tissues. However, there was considerable overlap in the immunohistochemical scores and expression levels of TFF1 transcripts between SSLs and HPs. The current findings may help elucidate the molecular mechanisms involved in serrated lesion development. In addition, we suggest that despite the limited practical application, upregulation of TFF1 transcripts may help differentiate SSLs from other lesions.

Background/Aims: To evaluate gastric early differentiate-type carcinogenesis, we attempted to identify clinicopathological and biological differences in differentiated-type minute intramucosal neoplasia (MIMN), which was defined as a tumor with a diameter of < 5 mm. Methods: We examined clinicopathological findings and biological factors, including TP53 overexpression, mucin phenotype, Ki-67-positive rate, MLH1, intranuclear accumulation of β-catenin, and DNA methylation status (low methylation epigenotype [LME], intermediate methylation epigenotype, and high methylation epigenotype [HME]) in MIMNs. In addition, non-MIMNs were also analyzed. In the present study, MIMN and non-MIMN were also examined based on low-grade dysplasia, high-grade dysplasia, and intramucosal cancer (IMC). Results: In clinicopathological findings, there were significant differences in sex ratios and tumor locations between MIMNs and non-MIMNs. Among the examined biological factors, no significant differences in the frequencies of biological factors were observed between the 2 intramucosal neoplasia types. However, the frequency of intranuclear accumulation of β-catenin was higher in non-MIMNs than in MIMNs. Finally, although the frequency of HME was significantly lower in MIMNs than in non-MIMNs, the opposite was observed for LME. Conclusions: The current finding suggested that DNA methylation and accumulation of β-catenin were closely associated with tumor development from MIMN to non-MIMN.

ObjectiveUterine carcinosarcoma (UCS) is a highly aggressive neoplasm that is composed of an intricate admixture of carcinomatous and sarcomatous elements. The relationship between UCS and the epithelial to mesenchymal transition (EMT) has been reported. In this study, we examined how expression of E-cadherin was associated with the expression of EMT-related proteins in UCS.MethodsUCS samples were histologically divided into three components: carcinomatous, transitional, and sarcomatous regions. Next, we examined the expression of E-cadherin and EMT-related proteins, including SNAI2, ZEB1, and TWIST1, in each component of the UCS using immunohistochemistry. The expression score was determined by combining the staining intensity and staining area of the target cells.ResultsThe expression score of E-cadherin was significantly lower in transitional and sarcomatous components than in the carcinomatous component. In addition, a significant difference in the low expression score of E-cadherin between transitional and sarcomatous components (transitional > sarcomatous components) was found. There were significant differences between the expression scores of ZEB1 in the three components (sarcomatous > transitional > carcinomatous components). However, no difference in the expression of TWIST1 between the components was found. Conversely, the expression level of SNAI2 was higher in sarcomatous or transitional components than in the carcinomatous component. However, a significant difference between the transitional and sarcomatous components was not detected.ConclusionThese results suggest that the EMT plays an essential role in the pathogenesis of UCS.

Supplemental digital content is available in the text., Objectives The aim of this study was to identify an association of pancreatic anaplastic carcinoma (APC) with the epithelial-mesenchymal transition (EMT). Methods Resected APCs (n = 24) were examined to assess components of APCs, including carcinomatous, transitional, and sarcomatous regions. Analysis was performed based on the immunoreactivity of E-cadherin and 3 EMT-related proteins: Slug (zinc finger protein SNAI2), Twist (Twist-related protein 1), and Zeb1 (zinc finger E-box–binding homeobox 1). Expression score was determined based on staining intensity and stained area of the target cells. Finally, we performed a hierarchical clustering based on the expression pattern of E-cadherin and EMT-related proteins of the sarcomatous component. Results The expression score of E-cadherin decreased in the order of sarcomatous > transitional > carcinomatous components (P < 0.01). Although there were significant differences in the immunohistochemical scores of Slug, Twist, and Zeb1 between carcinomatous and transitional components (P < 0.01), the significant difference in immunohistochemical score of Zeb1 between transitional and sarcomatous components was found (P < 0.05). Furthermore, APCs were divided into 2 subgroups based on the expression patterns of E-cadherin and EMT-related proteins (hierarchical clustering analysis). Consequently, these subgroups were distinguished by Twist expression. Conclusions Epithelial-mesenchymal transition plays an essential role in the pathogenesis of APC.

Background. Biological markers expressed in cancer cells and the surrounding cancer-associated fibroblasts (CAF) can be used for prediction of patient prognosis in colorectal cancer (CRC). Here, we used immunohistochemical techniques to evaluate cancer cells’ expression of specific biomarkers that are closely associated with neoplastic progression. Patients and Methods. Immunohistochemical markers included Ki-67, p53, β-catenin, MMP7, E-cadherin and HIF1-α. We also characterized microenvironmental markers expressed by CAF, including expression of α-smooth muscle actin, CD10, podoplanin, fibroblast specific protein 1, platelet derived growth factor β, fibroblast association protein, tenascin-C (TNC), ZEB1 and TWIST1. The study population consisted of 286 CRC patients with stage II and III disease. Stage II and III CRC were divided into a first and a second cohort (for validation). The CRCs were stratified using cluster analysis. To identify the utility of prognostic markers in stage II and III CRC, univariate and multivariate analyses were performed in both cohorts. Results. Stage II and III CRCs were stratified into 3 subgroups. Specific subgroups were significantly correlated to disease-free survival using univariate and multivariate analyses in the first cohort. High expression of TNC was identified as a single prognostic marker in both cohorts by univariate and multivariate analyses. Conclusions. We suggest that the presence of a specific subgroup defined by multiple markers can be used for prediction of CRC outcome in stages II and III. In addition, we showed that high expression of TNC was correlated with a poorer prognosis in stages II and III of CRC.

Programmed death ligand 1 (PD-L1) protein expression is a proposed predictive biomarker of immunotherapy; thus, identification of the clinicopathological and molecular characteristics associated with PD-L1 expression is important and necessary. We examined PD-L1 immunohistochemical expression and its relationships with the clinicopathological and molecular characteristics of patients with surgically resected nonsmall cell lung carcinoma. PD-L1 expression differed according to the histological subtype. Among 633 patients with adenocarcinoma, 523 (82.6%) had no PD-L1 expression, 78 (12.3%) low expression, and 32 (5.1%) high expression. PD-L1 expression was more common in men (p 0.001), in smokers (p = 0.002), and in patients with a more advanced stage (p = 0.002), the solid predominant subtype (p 0.001), no epidermal growth factor receptor(EGFR) mutations (p 0.001), a high MIB-1 labeling index (p 0.001), and positive p53 immunohistochemical expression (p 0.001). In a multivariate logistic regression analysis, the solid predominant subtype (odds ratio [OR] = 4.92, 95% confidence interval [CI]: 2.72-8.89, p 0.001), no EGFR mutations (OR = 2.27, 95% CI: 1.35-2.7, p = 0.002), a high MIB-1 labeling index (OR = 2.78, 95% CI: 1.72-4.55, p 0.001), and p53 positivity (OR = 2.13, 95% CI: 1.34-4.36, p = 0.042) were significantly and independently associated with PD-L1 expression. The combination of the solid predominant subtype with a high MIB-1 labeling index was strongly associated with positive expression of PD-L1. In the 193 patients with squamous cell carcinoma, 92 (47.7%) had no PD-L1 expression, 57 (29.5%) low expression, and 44 (22.8%) high expression. There were no significant correlations between PD-L1 expression and the evaluated clinicopathological or molecular characteristics of these patients. These results, indicating associations of PD-L1 with various clinicopathological or molecular characteristics in adenocarcinoma but not squamous cell carcinoma, may be useful for selecting patients with a good response to immune checkpoint inhibitors.

Background: Pulmonary epithelial–myoepithelial carcinoma is a rare subtype of lung cancer. Because of its rarity, the molecular information on this carcinoma is insufficient.Case presentation: We report a case of pulmonary epithelial–myoepithelial carcinoma without AKT1, HRAS or PIK3CA mutations in a 76-year-old woman. Computed tomography revealed a tumor located in the left lower lung. Thoracoscopic left lower lobectomy was performed. Histopathologically, the tumor consisted of duct-like structures and polygonal-to-spindle-cell features. The duct-like structures were composed of two distinct cell layers. Immunohistochemically, the inner layer consisted of cuboidal cells that were positive for pan-cytokeratin and negative for p63, whereas the outer layer consisted of polygonal and spindle cells that were positive for p63 and weakly positive for pan-cytokeratin. We evaluated mutations in AKT1, BRAF, CTNNB1, HRAS, KRAS and PIK3CA but did not detect any mutations.Conclusion: Pulmonary epithelial–myoepithelial carcinoma is a rare subtype of lung cancer, with only 56 previous cases reported in the English literature. The genetic alterations in pulmonary epithelial–myoepithelial carcinoma are still unclear. To our knowledge, only one study described HRAS mutations in pulmonary epithelial–myoepithelial carcinoma, detected in all three tumors evaluated. However, our case did not show any mutations in any of the above-examined genes.

MicroRNAs (miRNAs) are potential biomarkers of neoplastic lesions, but additional information on dysregulated miRNA expression during progression of the adenoma-adenocarcinoma sequence may be helpful to identify the role of miRNAs in this sequence. We examined the expression levels of 13 miRNAs (hsa-miRNA-19a-3p, hsa-miRNA-21-5p, hsa-miRNA-27a-3p, hsa-miRNA-27b-3p, hsa-miRNA-31-5p, hsa-miRNA-34b-3p, hsa-miRNA-125b-5p, hsa-miRNA-143-3p, miRNA-191-5p, hsa-miRNA-193b-3p, hsa-miRNA-195-5p, hsa-miRNA-206 and hsa-let-7a-5p) that are closely associated with colorectal carcinogenesis in 40 conventional adenomas (tubular and tubulovillous adenomas), 20 intramucosal carcinomas (IMCs) and 60 invasive colorectal cancers (iCRCs) using reverse-transcription polymerase chain reaction. These 120 tumors were divided into two cohorts, that is, cohort 1 (60 cases) and cohort 2 (for validation; 60 cases). We analyzed the expression levels of these miRNAs in the first step (adenoma→IMC) and second step IMC→iCRC) of the adenoma-carcinoma sequence in both cohorts. Although no significant differences in the expression of any of the 13 miRNAs were found between adenomas and IMCs consistently in both cohorts, the expression levels of hsa-miRNA-125b-5p, hsa-miRNA-143-3p, and hsa-miRNA-206 were significantly upregulated in iCRC in both cohorts compared with those in IMC. The current results suggest that certain miRNAs, including hsa-miRNA-125b-5p, hsa-miRNA-143-3p and hsa-miRNA-206, are candidate markers that play critical roles in the progression of IMC to iCRC.

Gastric intraepithelial foveolar type neoplasia (IEFN) is not well defined. In addition, atrophic mucosa (AM) is an important issue to consider when evaluating gastric tumorigenesis. Here, we assessed the clinicopathological characteristics and molecular alterations contributing to the development of IEFN compared with intestinal type neoplasia. We examined the clinicopathological and molecular features of 42 cases of IEFN with low-grade dysplasia (LGD) and those of 77 cases of intraepithelial intestinal type neoplasia (IEIN) with LGD. The clinicopathological and molecular features examined included the AM status, mucin phenotype expression, CDX2 expression, p53 overexpression, β-catenin intranuclear accumulation, microsatellite instability (MSI), DNA methylation status (low methylation epigenotype [LME], intermediate ME, or high ME), allelic imbalances (AIs), and APC promoter 1B mutations. There were no differences in the frequencies of AM and rates of CDX2 expression between IEFN and IEIN cases. Although no differences in the frequencies of p53 overexpression and MSI were observed between the two histological types, intranuclear expression of β-catenin was significantly higher in IEIN than in IEFN. In addition, although the rate of LME was significantly higher in IEFN cases than in IEIN cases, IEFN was characterized by AIs at multiple foci. Finally, mutation of the APC promoter 1B, which is a characteristic of gastric adenocarcinoma and proximal polyposis of the stomach (potentially resembling IEFN), was detected in only one IEFN case. These findings suggested that IEFN may be an independent entity in terms of molecular alterations including the presence of multiple AIs and LME. Electronic supplementary material The online version of this article (10.1007/s00428-020-02846-0) contains supplementary material, which is available to authorized users.

Activated cancer-associated fibroblasts (CAFs) and fibroblasts that have undergone the epithelial-mesenchymal transition (EMT) in cancer stroma contribute to tumor progression and metastasis. However, no reports have investigated the CAF phenotype and its clinicopathological relevance in cutaneous malignant tumors, including basal cell carcinoma (BCC), squamous cell carcinoma (SCC), and malignant melanoma (MM). Here, we investigated the CAF phenotype in cutaneous malignant tumors based on their histology and immunohistochemical expression of CAF-related markers, including adipocyte enhancer-binding protein 1 (AEBP1), podoplanin, platelet-derived growth factor receptor α (PDGFRα), PDGFRβ, fibroblast activating protein (FAP), CD10, S100A4, α-smooth muscle actin (α-SMA), and EMT-related markers (Zeb1, Slug, and Twist). In addition, we assessed the role of the CAF phenotype in cutaneous malignant cancers using hierarchical cluster analysis. Consequently, 3 subgroups were stratified based on the expression pattern of CAF- and EMT-related markers. Subgroup 1 was characterized by low expression of AEBP1, PDGFRα, PDGFRβ, FAP and Slug, whereas subgroup 2 was closely associated with high expression of PDGFRβ, S100A4 and Twist. In addition, high expression levels of podoplanin, PDGFRβ, CD10, S100A4, α-SMA, Zeb1, Slug and Twist were observed in subgroup 3. High expression of CD10 was commonly found in all 3 subgroups. These subgroups were correlated with histologic subtypes, that is, subgroup 1, MM; subgroup 2, BCC; and subgroup 3, SCC. We suggest that the expression pattern of CAF- and EMT-related proteins plays crucial roles in the progression of BCC, SCC, and MM.

Background Hemorrhage at presentation in primary central nervous system (CNS) lymphoma is rare. We encountered a case of primary CNS lymphoma presenting as a growing intracerebral hemorrhage. Case Description An 80-year-old man presented with mild dysarthria. Computed tomography demonstrated a round, high-density mass with surrounding vasogenic edema in the left frontal lobe. Although the patient was placed on antihypertensive therapy for suspected subacute subcortical hemorrhage, neurologic symptoms gradually worsened. Computed tomography after 2 weeks revealed that the high-density lesion and surrounding edema had increased in size compared with previous images. The patient had been transferred to our hospital 14 days after admission to another institution. Magnetic resonance imaging demonstrated a mass lesion comprising hemorrhage of different phases in the left frontal lobe. Contrast-enhanced T1-weighted imaging demonstrated a mass lesion with heterogeneous enhancement in the left frontal lobe. The patient underwent craniotomy with gross total removal of the hemorrhagic lesion. The histopathologic diagnosis was diffuse large-cell non-Hodgkin lymphoma, and immunohistochemistry showed high immunoreactivity for vascular endothelial growth factor. Conclusion Although exceedingly rare, primary central nervous system lymphoma can present as growing intracerebral hemorrhage due to repeated intratumoral hemorrhages. High expression of vascular endothelial growth factor and the mass effects of hemorrhage could be associated with the onset and growth of intracerebral hemorrhage. Early evaluation and meticulous observation are important to avoid progressive, life-threatening situations in such cases.

Summary The molecular alterations and pathological features of gastric papillary adenocarcinoma (GPA) remain unknown. We examined GPA samples and compared their molecular and pathological characteristics with those of gastric tubular adenocarcinoma (GTA). Additionally, we identified pathological and molecular features of GPA that vary with microsatellite stability. In the present study, samples from 63 GPA patients and 47 GTA patients were examined using a combination of polymerase chain reaction (PCR)-microsatellite assays and PCR-pyrosequencing in order to detect microsatellite instability (microsatellite instability, MSI; microsatellite stable, MSS), methylation status (low methylation, intermediate methylation and high methylation level), and chromosomal AI in multiple cancer-related loci. Additionally, the expression levels of TP53 and Her2 were evaluated using immunohistochemistry. GTA and GPA are statistically different in their frequency of pathological features, including mucinous, poorly differentiated and invasive micropapillary components. Clear genetic patterns differentiating GPA and GTA could not be identified with a hierarchical cluster analysis, but microsatellite stability was linked with TP53 and Her2 overexpression. Methylation status in GPA was also associated with the development of high microsatellite instability. However, no pathological differences were associated with microsatellite stability. We suggest that although molecular alterations in a subset of GPAs are closely associated with microsatellite stability, they play a minor role in GPA carcinogenesis.

Tumor budding is thought to reflect the epithelial-mesenchymal transition (EMT). However, the molecular mechanism linking tumor buds and the EMT remains unclear. Here, we examined the induction of tumor budding and EMT and their association with EMT-related proteins (ZEB1, TWIST, SNAIL, and SLUG) in colorectal cancer (CRC). Immunohistochemical expression of pan-cytokeratin was examined for identification of tumor budding in 101 CRCs. Grading of tumor budding was classified into low- and high-grade groups. Tissue microarray was conducted to identify tumor budding sites. The expression of E-cadherin, ZEB1, TWIST, SNAIL, and SLUG was examined in areas of tumor budding and the surrounding tumor stroma using a double-immunostaining method. Specifically, pan-cytokeratin and EMT-related proteins were assessed by double immunostaining. Low or no expression of E-cadherin was found in areas of tumor budding. Moreover, ZEB1, TWIST, SNAIL, and SLUG were not expressed in regions of tumor budding. However, the expression level of ZEB1 in the stromal cells surrounding tumor budding was significantly more frequent than that of TWIST, SNAI, and SLUG. In addition, the expression of EMT-related proteins in surrounding stromal cells was significantly greater in areas of high-grade tumor budding than in low-grade areas. Our present results suggest that EMT-related proteins play a minor role in forming tumor buds. In addition, our findings suggest the existence of subtypes of stromal cells in CRC with phenotypical and functional heterogeneity.

Object: The aim of the present study was to investigate the clinicopathological characteristics and prognostic factors associated with sporadic colorectal cancer (CRC). We examined the clinicopathological findings and immunohistochemical expression of tumor prognostic markers at tumor budding sites to determine their predictive value for patient prognosis. Materials and Methods: Immunohistochemical examination was performed by tissue microarray (TMA) of specimens from 106 patients with CRC. On hematoxylin and eosin (H&E)-stained tumor tissue slides, a representative area of tumor budding at the invasive front was selected for the construction of a TMA. Immunostaining for matrix metalloproteinase-7 (MMP7), the laminin-5 (ln-5) γ2 chain and S100A4 was performed to determine the association between patient survival and these markers. Results: Clinicopathological variables were also assessed. Tumor location, histological type, degree of lymphatic invasion and vascular invasion, tumor stage, epithelial expression of S100A4, stromal cell expression of S100A4 and expression of the ln-5γ2 chain were associated with an increased risk of mortality. Five factors were retained in the multivariate logistic regression analysis. Specifically, the tumor location, degree of lymphatic invasion and vascular invasion, tumor stage and stromal cell expression of S100A4 remained significant predictors of patient survival after controlling for the other variables. Conclusion: Vascular invasion and stromal expression of S100A4 in the tumor budding areas correlated with patient survival. Stromal immunostaining of S100A4 may be useful for identifying high-risk patients with advanced CRC.

Since the new adenocarcinoma (ADC) classification was presented in 2011, several authors have reported that patients with solid (S) and/or micropapillary (MP) predominant patterns showed a worse prognosis. On the other hand, there are several patients who have S and/or MP patterns even if their patterns are not predominant. However, the evaluation of these patients is uncertain.A total of 531 ADCs were examined. We classified the patients into five subgroups according to the proportion of S and/or MP patterns: (1) both patterns absent (S-/MP-), (2) S predominant (S pre), (3) MP predominant (MP pre), (4) S pattern present although not predominant and MP pattern absent (S+ not pre/MP-), and (5) MP pattern present although not predominant (MP+ not pre).Of the 531 ADCs, 384 (72.3%) were classified as S-/MP-, 55 (10.4%) as S pre, 11 (2.1%) as MP pre, 42 (7.9%) as S+ not pre/MP-, and 39 (7.3%) as MP+ not pre. In a univariate analysis, the recurrence-free survival (RFS) and overall survival differed significantly among the five subgroups (p0.01 and p0.01, respectively). In a multivariate analysis, patients with S-/MP- had significantly higher RFS rates than did those with other subgroups. On the other hand, patients with MP pre had lower RFS rates than did those with other subgroups.Patients with S and/or MP patterns have a poorer prognosis even if their patterns are not predominant. The S and/or MP patterns must be treated at the time of diagnosis.

Somatic copy number alterations (SCNAs) are important biological characteristics that can identify genome‐wide alterations in renal cell carcinoma (RCC). Recent studies have shown that SCNAs have potential value for determining the prognosis of RCC. We examined SCNAs using the Affymetrix platform to analyze samples from 59 patients with clear cell RCCs (ccRCCs) including first cohort (30 cases) and second cohort (validation cohort, 29 cases). We stratified SCNAs in the ccRCCs using a hierarchical cluster analysis based on SCNA types, including gain, loss of heterozygosity (LOH), copy neutral LOH, mosaic, and mixed types. In this way, the examined two cohorts were categorized into two subgroups (1 and 2). Although the frequency of mixed type was higher in subgroup 1 than in subgroup 2 in the two cohorts, the association did not reach statistical significance. There was a significant difference in the frequency of metachronous metastasis between subgroups 1 and 2 (subgroup 2 > 1). In addition, subgroup 2 was retained in multivariate analysis of both cohorts. We examined whether there were specific alleles differing between subgroups 1 and 2 in both cohorts. We found that there was indeed a statistically significant difference in the 3p mixed types. Among the 3p mixed type, we found that 3p24.3 mixed type was inversely correlated with the presence of metachronous metastasis in ccRCC. The association was also retained in multivariate analysis in second cohort. We suggest that the 3p24.3 mixed type may be a novel marker to predict a favorable prognosis in ccRCC.

Summary The expression of gastrointestinal differentiation markers is associated with the tumorigenesis and prognosis of digestive cancers. However, little is known about the significance of gastrointestinal differentiation marker profiles in patients with extrahepatic cholangiocarcinoma (CCA), which is classified as perihilar and distal CCA. The purpose of this study was to clarify the role of gastrointestinal differentiation marker expression in extrahepatic CCA based on tumor location. We examined the expression of gastrointestinal differentiation markers in resected perihilar (n = 30) and distal (n = 54) CCAs based on the immunohistochemical expression of the following markers: MUC2, MUC5AC, MUC6, CD10, CDX-2, and cytokeratin 20. Expression scores were determined semiquantitatively based on the rate of positively stained cells. Furthermore, we performed hierarchical clustering of the CCAs based on the immunohistochemical expression scores to evaluate differences in the expression patterns of the 6 gastrointestinal differentiation markers. Consequently, perihilar and distal CCAs were stratified into 2 subgroups each. Among the perihilar CCAs, subgroup 1 was characterized by lower expression of MUC5AC and MUC6, a larger median tumor size, and a significantly worse prognosis compared with subgroup 2. Furthermore, the immunohistological subgroup (subgroup 1 versus 2) and TNM stage (stage III versus II) were independent predictors of patient survival. Among the distal CCAs, subgroup 1 was characterized by lower expression of MUC5AC compared with subgroup 2. We suggest that gastrointestinal differentiation marker profiles are useful for stratifying perihilar and distal CCAs. In addition, gastrointestinal differentiation markers play a crucial role in tumor development, particularly in perihilar CCA.

We report the case of a patient with spinal immature teratoma and cerebrospinal fluid leakage from the congenital dermal sinus tract. A 0-day-old female infant presented with a subcutaneous soft mass with a dimple in the lumbosacral region at birth. Magnetic resonance imaging revealed a mixed low-intensity mass located in the extraspinal and intraspinal canal with a sinus tract. The reconstructed three-dimensional spinal computed tomography image showed spina bifida and ectopic ossification at the dorsal aspect of the sacrum. Urgent removal of the tumor and dermal sinus tract was then performed under evoked electromyography monitoring. The resected tumor was histopathologically diagnosed as immature teratoma.

Background: MicroRNAs (miRNA) are frequently dysregulated in clear cell renal cell carcinoma (ccRCC). Objective: This study aimed to elucidate the role of miRNA expression patterns in renal carcinogenesis and to identify the specific miRNAs that exhibit expression patterns closely associated with patient outcomes. Methods: We examined the expression patterns of selected miRNAs, including miRNA-155-5p, miRNA-122-5p, miRNA-21-5p, miRNA-185-5p, miRNA-106a-5p, miRNA-106b-3p, miRNA-34b-3p, miRNA-210-3p, miRNA-141-3p, miRNA-200c-3p, miRNA-135a-5p, miRNA-30a-5p, miRNA-218-5p, miRNA-429, miRNA-200a-3p and miRNA-200b-3p, in 96 samples of ccRCCs using the TaqMan real-time PCR method. In addition, cluster analysis was performed to stratify expression patterns of multiple miRNAs. Results: In the present study, three distinct subgroups could be clearly stratified in ccRCCs. Subgroup 1 was characterized by upregulation of miRNA-155-5p, miRNA-122-5p, miRNA-21-5p, miRNA-185-5p, miRNA-106a-5p, miRNA-106b-3p, miRNA-34b-3p and miRNA-210-3p. Subgroup 2 was closely associated with downregulation of miRNA-141-3p, miRNA200c-3p, miRNA-30a-5p, miRNA-218-5p, miRNA-429, miRNA-200a-3p and miRNA-200b-3p. Moreover, significant lower expression of miRNA-135a-5p was a distinctive feature of subgroup 3, which was correlated with metachronous metastasis. Among the individual markers in subgroup 3, miRNA-135a-5p was retained in multivariate analysis. The cutoff value of miRNA-135a-5p expression to identify the association of an altered level of miRNA-135a-5p with metachronous metastasis in ccRCCs was determined and showed excellent specificity. Conclusion: We suggest that the expression pattern of the chosen miRNAs is useful to identify renal carcinogenesis and to help identify the association of such expression patterns with metachronous metastasis in ccRCCs. In addition, miRNA-135a-5p was an excellent marker for prediction of metachronous metastasis.

Immunoglobulin A (IgA) nephropathy (IgAN) is characterized by high serum IgA levels and IgA deposition in the renal mesangium. Previous studies suggest that elevated serum IgA partly originates from the tonsils. Here, we investigated the mechanisms of IgA production in the tonsils of patients with IgAN. Immunohistochemistry revealed that the number and relative percentage of IgA-bearing cells were significantly increased in the tonsils of IgAN patients. Compared with non-IgAN patients, enhanced IgA class switching and overexpression of thymic stromal lymphopoietin (TSLP), TSLP receptor (TSLPR), activation-induced cytidine deaminase (AID), transforming growth factor-β1 (TGF-β1), B cell-activating factor of the tumor necrosis factor family (BAFF), and a proliferation-inducing ligand (APRIL) were detected in follicular dendritic cells (FDCs) of tonsillar germinal centers from IgAN patients. Importantly, TSLP correlated with IgA production in isolated FDC-associated clusters. Serum TSLP levels were increased and correlated with IgA overexpression in the tonsils and serum of IgAN patients. These data indicated that TSLP overexpression in tonsillar FDCs may promote IgA class switching in IgAN patients through the cooperative roles of AID, TGF-β1, BAFF, and APRIL. Therefore, interactions between TSLP in FDCs and IgA production in tonsils may be an important mechanism contributing to the pathogenesis of IgAN.

The prognosis for myelodysplastic syndrome with bone marrow fibrosis (MDS-F) is worse than the prognosis of MDS without fibrosis. Hematopoietic stem cell transplantation (HSCT) is the only curative therapy; however, the indications and the procedures involved in HSCT remain unclear. We herein describe a 69-year-old Japanese man with MDS-F who received haploidentical HSCT and post-transplantation cyclophosphamide. Although the first HSCT resulted in secondary graft failure, the second HSCT using PTCy led to successful engraftment after early improvement in fibrosis. Since the incidence of graft failure is high in myelofibrosis patients, a secondary HSCT using PTCy may be successful if employed.