Search

Background Recurrent glioblastoma multiforme (GBM) is a highly aggressive primary malignant brain tumor that is resistant to existing treatments. Recently, we reported that activated autologous natural killer (NK) cell therapeutics induced a marked increase in survival of some patients with recurrent GBM. Methods To identify biomarkers that predict responsiveness to NK cell therapeutics, we examined immune profiles in tumor tissues using NanoString nCounter analysis and compared the profiles between 5 responders and 7 non-responders. Through a three-step data analysis, we identified three candidate biomarkers (TNFRSF18, TNFSF4, and IL12RB2) and performed validation with qRT-PCR. We also performed immunohistochemistry and a NK cell migration assay to assess the function of these genes. Results Responders had higher expression of many immune-signaling genes compared with non-responders, which suggests an immune-active tumor microenvironment in responders. The random forest model that identified TNFRSF18, TNFSF4, and IL12RB2 showed a 100% accuracy (95% CI 73.5–100%) for predicting the response to NK cell therapeutics. The expression levels of these three genes by qRT-PCR were highly correlated with the NanoString levels, with high Pearson’s correlation coefficients (0.419 (TNFRSF18), 0.700 (TNFSF4), and 0.502 (IL12RB2)); their prediction performance also showed 100% accuracy (95% CI 73.54–100%) by logistic regression modeling. We also demonstrated that these genes were related to cytotoxic T cell infiltration and NK cell migration in the tumor microenvironment. Conclusion We identified TNFRSF18, TNFSF4, and IL12RB2 as biomarkers that predict response to NK cell therapeutics in recurrent GBM, which might provide a new treatment strategy for this highly aggressive tumor.

With the advent of checkpoint inhibitors, it has opened up opportunities for numerous cancer patients. However, as is the case with every treatment, complications need to be weighed. Gastrointestinal adverse effects, such as diarrhea and colitis are well-known complications for checkpoint inhibitors. In severe cases, colitis-induced colonic perforation may occur with an estimation of 1.0% to 1.5% in anti-CTLA-4 antibodies. However, only a handful of cases of such devastating complications have been reported in anti-PD-1 antibodies such as pembrolizumab and nivolumab. We here report a case of intestinal perforation in a patient treated with nivolumab.

Histologically poor differentiation is associated with lymph node metastasis. Thus, pathological evaluation of biopsy specimens is crucial when treating stomach cancers. Deep learning of WSIs is challenging because the images are enormous. Given the computing limitations, patch-level supervised learning methods have been proposed. However, valuable information is lost when dividing WSIs into smaller patches. Another drawback is the need for pixel-level annotation by a pathologist. It is acceptable to differentiate, i.e., grade, gastric cancer at the holistic tissue level (i.e., under low magnification). We developed a weakly supervised learning technique for tissue-level gastric adenocarcinoma histological differentiation (well-to-moderately or poorly differentiated) and applied global reasoning to tissue-level features. The tissue-level AUROCs of the histological differentiation classifiers were 0.953, 0.969, and 0.943, respectively when data from five hospitals were subjected to threefold cross-validation. Comparison of the Grad-CAM heatmaps of the trained classifier and the pathologists’ annotations confirmed that our weakly supervised model exhibited performed well.

Alteration in expression of miRNAs can cause various malignant changes and the metastatic process. Our aim was to identify the miRNAs involved in cervical squamous cell carcinoma (SqCC) and metastasis, and to test their utility as indicators of metastasis and survival. Using microarray technology, we performed miRNA expression profiling on primary cervical SqCC tissue (n = 6) compared with normal control (NC) tissue and compared SqCC that had (SqC-M; n = 3) and had not (SqC-NM; n = 3) metastasized. Four miRNAs were selected for validation by qRT-PCR on 29 SqC-NM and 27 SqC-M samples, and nine metastatic lesions (ML-SqC), from a total of 56 patients. Correlation of miRNA expression and clinicopathological parameters was analyzed to evaluate the clinical impact of candidate miRNAs. We found 40 miRNAs differentially altered in cervical SqCC tissue: 21 miRNAs were upregulated and 19 were downregulated (≥2-fold, p < 0.05). Eight were differentially altered in SqC-M compared with SqC-NM samples: four were upregulated (miR-494, miR-92a-3p, miR-205-5p, and miR-221-3p), and four were downregulated (miR-574-3p, miR-4769-3p, miR-1281, and miR-1825) (≥1.5-fold, p < 0.05). MiR-22-3p might be a metastamiR, which was gradually further downregulated in SqC-NM > SqC-M > ML-SqC. Downregulation of miR-30e-5p significantly correlated with high stage, lymph node metastasis, and low survival rate, suggesting an independent poor prognostic factor.

PurposeIn this study, we investigated the frequencies of mutations in DNA damage repair genes including BRCA1, BRCA2, homologous recombination genes and TP53 gene in ovarian high-grade serous carcinoma, alongside those of germline and somatic BRCA mutations, with the aim of improving the identification of patients suitable for treatment with poly(ADP-ribose) polymerase inhibitors.Materials and MethodsTissue samples from 77 Korean patients with ovarian high-grade serous carcinoma were subjected to next-generation sequencing. Pathogenic alterations of 38 DNA damage repair genes and TP53 gene and their relationships with patient survival were examined. Additionally, we analyzed BRCA germline variants in blood samples from 47 of the patients for comparison.ResultsBRCA1, BRCA2, and TP53 mutations were detected in 28.6%, 5.2%, and 80.5% of the 77 patients, respectively. Alterations in RAD50, ATR, MSH6, MSH2, and FANCA were also identified. At least one mutation in a DNA damage repair gene was detected in 40.3% of patients (31/77). Germline and somatic BRCA mutations were found in 20 of 47 patients (42.6%), and four patients had only somatic mutations without germline mutations (8.5%, 4/47). Patients with DNA damage repair gene alterations with or without TP53mutation, exhibited better disease-free survival than those with TP53 mutation alone.ConclusionDNA damage repair genes were mutated in 40.3% of patients with high-grade serous carcinoma, with somatic BRCA mutations in the absence of germline mutation in 8.5%. Somatic variant examination, along with germline testing of DNA damage repair genes, has potential to detect additional candidates for PARP inhibitor treatment.

Ovarian cancer is one of the leading causes of deaths among patients with gynecological malignancies worldwide. In order to identify prognostic markers for ovarian cancer, we performed RNA-sequencing and analyzed the transcriptome data from 51 patients who received conventional therapies for high-grade serous ovarian carcinoma (HGSC). Patients with early-stage (I or II) HGSC exhibited higher immune gene expression than patients with advanced stage (III or IV) HGSC. In order to predict the prognosis of patients with HGSC, we created machine learning-based models and identified USP19 and RPL23 as candidate prognostic markers. Specifically, patients with lower USP19 mRNA levels and those with higher RPL23 mRNA levels had worse prognoses. This model was then used to analyze the data of patients with HGSC hosted on The Cancer Genome Atlas, this analysis validated the prognostic abilities of these two genes with respect to patient survival. Taken together, the transcriptome profiles of USP19 and RPL23 determined using a machine-learning model could serve as prognostic markers for patients with HGSC receiving conventional therapy.

Immune checkpoint blockade is promising for treating non-small-cell lung cancer (NSCLC). We used multipanel markers to predict the response to immune checkpoint inhibitors (ICIs) by characterizing gene expression signatures or individual genes in patients who showed durable clinical benefit to ICIs. Twenty-one patients with NSCLC treated with single-agent anti-programmed cell death protein (PD)-1 antibody were analyzed and their clinicopathological characteristics and response to ICIs were characterized. Nine (43%) showed a durable clinical benefit (DCB), while the remaining 12 (57%) patients showed non-durable benefit (NDB). The M1 and peripheral T cell signatures showed the best performance for discriminating DCB from NDB (sensitivity, specificity, accuracy = 0.89, 1.0, 0.95, respectively). Progression-free survival (PFS) was significantly longer in patients with high M1 signature or high peripheral T cell signature scores. CD137 and PSMB9 mRNA expression was higher in the DCB group than in the NDB group. Patients with high PSMB9 expression showed longer PFS. M1 signature, peripheral T cell signature and high mRNA expression level of CD137 and PSMB9 showed better predictive performance than known biomarkers, such as PD-L1 immunohistochemistry, tumor mutation burden, or tumor-infiltrating lymphocytes.

Background: Fascin is an actin-bundling protein that promotes cancer cell migration and invasion. By contrast, breast cancer metastasis suppressor 1 (BRMS1) inhibits cancer metastasis by targeting multiple steps of the metastatic cascade. We evaluated whether expression patterns of fascin and BRMS1 correlate with clinicopathological features and patient outcome. Methods: Immunohistochemistry for fascin and BRMS1 was performed using a tissue microarray constructed from 183 human breast cancer tissues. Fascin expression determined by the proportion of stained tumor cells (0: 0-5%, 1: 6-25%, 2: 26-50%, 3: 51-75%, or 4: >75%) and staining intensity (0: negative, 1: weak, 2: moderate, or 3: strong) were multiplied and defined as negative (0-3) or positive (4-12). BRMS1 expression was scored separately based on nuclear and cytoplasmic staining intensity (0: negative, 1: weak, 2: moderate, 3: strong). We obtained the BRMS1 H score by summing the nuclear and cytoplasmic scores and defined it as negative (0-2) or positive (3-6). Results: Expression of BRMS1 showed a significant inverse correlation with that of fascin. Fascin+ tumors were significantly associated with no lymph node metastasis, higher histological and higher nuclear grade, ER/PR/HER2 negativity, and triple-negative subtype (all ps < 0.05). These clinicopathological differences showed the same trend in a comparison of fascin-/BRMS1+ and fascin+/BRMS1- tumors. Negative or weak BRMS1 cytoplasmic expression was significantly associated with shorter disease-free survival (DFS; p = 0.043). Fascin positivity was significantly associated with shorter DFS (p = 0.005) and overall survival (p = 0.020) when analyses were confined to node-negative patients. Conclusions: This study confirms an inverse correlation between expression of fascin and expression of BRMS1 using a quite large cohort of human breast cancer tissues. Fascin alone or combined with BRMS1 was a worse prognostic marker, particularly in node-negative breast cancer patients.

What happens in the early and still undetectable human malignancy is unknown because direct observations are impractical. Here we present a “Big Bang” model, whereby a tumor grows predominantly as a single expansion producing numerous intermixed sub-clones, which are not subject to stringent clonal selection. In this model, both public and most detectable private mutations arise during the earliest phase of tumor growth. Multi-scale genomic profiling of 349 individual glands sampled from 15 colorectal tumors revealed the absence of selective sweeps, uniformly high intra-tumor heterogeneity, and sub-clone mixing in distant tumor regions, as postulated by the Big Bang. By integrating the data in a spatial model of tumor growth and statistical inference framework we also verified the most striking prediction of our model, namely that most detectable intra-tumor heterogeneity originates from private alterations acquired early during growth, and not from the later expansion of selected sub-clones. Hence, early sub-clones define the genomic profile of colorectal carcinomas and advanced adenomas, whereas potentially dangerous late-arising sub-clones will go undetected. Moreover, our results suggest that sub-clone mixing may be a biomarker of malignant potential. This new model provides a quantitative framework that explains the origins of intra-tumor heterogeneity and tumor growth dynamics with significant clinical implications. Citation Format: Andrea Sottoriva, Haeyoun Kang, Zhicheng Ma, Trevor A. Graham, Matthew Salomon, Junsong Zhao, Paul Marjoram, Kim Siegmund, Michael F. Press, Darryl Shibata, Christina Curtis. A Big Bang model of human colorectal tumor growth. [abstract]. In: Proceedings of the AACR Special Conference on Computational and Systems Biology of Cancer; Feb 8-11 2015; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(22 Suppl 2):Abstract nr B2-23.

Fibulin-5 is an extracellular matrix (ECM) glycoprotein which has a role in the organisation and stabilisation of ECM structures and regulating cell proliferation and tumourigenesis. Here, the expression of fibulin-5 and its functional effects on the migration and invasion of ovarian cancer cells were assessed.Expression of fibulin-5 was detected in 44 ovarian tumour tissues by qRT-PCR, Western blotting and immunohistochemistry. We performed cell migration and invasion assays, and cell cycle analysis in fibulin-5 transfected SKOV3 (SKOV3-FBLN5) cells and the parental SKOV3 cells. We further examined the expression of three tissue inhibitors of metalloproteinases (TIMPs) and seven matrix metalloproteinases (MMPs) by RT-PCR.mRNA and protein expression of fibulin-5 were down-regulated (0.05-fold and 0.1-fold) in ovarian carcinomas compared with control tissues (p0.01 and p=0.022). In wound-healing and invasion assays, significantly fewer SKOV3-FBLN5 cells than SKOV3 control cells migrated and invaded (39.1%, p=0.046 and 70%, p=0.03, respectively), which was reversed by siRNA-treatment. Overexpression of fibulin-5 induced G2/M arrest and increased cyclin B1, CDC2 and CDC25C. Expression of TIMP-2 (0.56-fold), MMP-3 (0.43-fold) and MMP-13 (0.18-fold) was lower and MMP-9 expression (2.20-fold) was higher in SKOV3-FBLN5 cells than in control cells.Fibulin-5 is significantly down-regulated in ovarian carcinoma and acts as a tumour suppressor by inhibiting the migration and invasion of ovarian cancer cells.

Intratumoural mutational heterogeneity (ITH) or the presence of different private mutations in different parts of the same tumour is commonly observed in human tumours. The mechanisms generating such ITH are uncertain. Here we find that ITH can be remarkably well structured by measuring point mutations, chromosome copy numbers, and DNA passenger methylation from opposite sides and individual glands of a 6cm human colorectal adenoma. ITH was present between tumour sides and individual glands, but the private mutations were side-specific and subdivided the adenoma into two major subclones. Furthermore, ITH disappeared within individual glands because the glands were clonal populations composed of cells with identical mutant genotypes. Despite mutation clonality, the glands were relatively old, diverse populations when their individual cells were compared for passenger methylation and by FISH. These observations can be organized into an expanding star-like ancestral tree with co-clonal expansion, where many private mutations and multiple related clones arise during the first few divisions. As a consequence, most detectable mutational ITH in the final tumour originates from the first few divisions. Much of the early history of a tumour, especially the first few divisions, may be embedded within the detectable ITH of tumour genomes. Copyright (c) 2015 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Intrahepatic cholangiocarcinoma (ICC) is a malignant tumor with biliary epithelial differentiation. Malignant transformation of von Meyenburg complex (VMC) to ICC has been reported [1,2], and a new subtype of ICC with ductal plate malformation (DPM) pattern has been suggested [3]. However, bile duct adenoma (BDA) is a rare entity and is not as well known as DPM. Moreover, it has not been determined whether BDA is a risk factor of ICC. We present a rare case of ICC with DPM-like features associated with BDA.

Notch signaling plays an important role in ovarian cancer chemoresistance, which is responsible for recurrence. Gamma-secretase inhibitor (GSI) is a broad-spectrum Notch inhibitor, but it has serious side effects. The efficacy of Notch3-specific inhibition in paclitaxel-resistant ovarian cancers was assessed in this study, which has not yet been evaluated relative to GSI. To analyze the effect of Notch3-specific inhibition on paclitaxel-resistant ovarian cancers, we compared cell viability, apoptosis, cell migration, angiogenesis, cell cycle, and spheroid formation after treatment with either Notch3 siRNA or GSI in paclitaxel-resistant SKpac cells and parental SKOV3 cells. Expression levels of survival, cell cycle, and apoptosis-related proteins were measured and compared between groups. Notch3 was significantly overexpressed in chemoresistant cancer tissues and cell lines relative to chemosensitive group. In paclitaxel-resistant cancer cells, Notch inhibition significantly reduced viability, migration, and angiogenesis and increased apoptosis, thereby boosting sensitivity to paclitaxel. Spheroid formation was also significantly reduced. Both Notch3 siRNA-treated cells and GSI-treated cells arrested in the G2/M phase of the cell cycle. Proteins of cell survival, cyclin D1 and cyclin D3 were reduced, whereas p21 and p27 were elevated. Both GSI and Notch3 siRNA treatment reduced expression of anti-apoptotic proteins (BCL-W, BCL2, and BCL-XL) and increased expression of pro-apoptotic proteins (Bad, Bak, Bim, Bid, and Bax). These results indicate that Notch3-specific inhibition sensitizes paclitaxel-resistant cancer cells to paclitaxel treatment, with an efficacy comparable to that of GSI. This approach would be likely to avoid the side effects of broad-spectrum GSI treatment. © 2015 Wiley Periodicals, Inc.

Ovarian carcinoma is a highly lethal malignancy due to frequent relapse and drug resistance. Cancer stem cells (CSCs) are thought to contribute significantly to disease relapse and drug resistance. In this study, a subpopulation of CSCs of ovarian carcinoma was isolated and the genes differentially expressed in these cells were identified to characterize CSCs and to find candidate biomarkers. Ovarian carcinoma cells from patients were primarily cultured, and spheroid-forming cells (SFCs) were isolated. The characteristic genes of SFCs were identified through cDNA microarray and validation by quantitative real-time polymerase chain reaction and immunohistochemistry, and the association of their expression with clinicopathologic parameters was analyzed. GSC (4.26-fold), VAV3 (7.05-fold), FOXA2 (12.06-fold), LEF1 (17.26-fold), COMP (21.33-fold), GRIN2A (9.36-fold), CD86 (23.14-fold), PYY (4.18-fold), NKX3-2 (10.35-fold), and PDK4 (74.26-fold) were significantly upregulated in SFCs compared with parental cancer cells. With validation for human ovarian carcinomas, LEF1, PYY, NKX3-2, and WNT3A were significantly upregulated in chemoresistant cancers compared with chemosensitive cancers. Overexpression of LEF1, VAV3, and NKX3-2 was significantly associated with distant metastasis by immunohistochemistry. VAV3 overexpression was an independent poor survival indicator (hazard ratio=15.27, P

MicroRNA-145 (miR-145) expression is downregulated in several human cancers, but its clinical and functional relevance to ovarian carcinoma has not yet been elucidated. This study addressed the hypothesis that miR-145 serves as a prognostic biomarker and a tumor suppressor that regulates the expression of high-mobility group A2 (HMGA2) oncoprotein in ovarian cancer. Here, we found that low miR-145 expression and HMGA2 overexpression determined by qRT-PCR and immunohistochemistry significantly correlated with advanced stage, lymph node involvement, and distant metastasis in 74 ovarian carcinomas. Low miR-145 expression significantly correlated with tumor recurrence and worse overall survival (HR=8.62, P = 0.039). Transfection of pre-miR-145 resulted in reduced cell growth and migration, and increased apoptosis of ovarian cancer cells by TUNEL, colony forming, and cell migration assays. MiR-145 was found to directly target HMGA2 by luciferase assay and Western blotting. Our findings suggest that miR-145 functions as a tumor suppressor in ovarian cancer and directly targets HMGA2 oncoprotein. Low miR-145 and high HMGA2 expressions are potential biomarkers of poor prognosis of ovarian carcinoma and miR-145 is the more powerful predictor of patient outcome.

Calcifying aponeurotic fibroma is a rare, benign fibroblastic tumor that typically occurs in the palms of the hands and soles of the feet in children and adolescents. We report an unusual case of a calcifying aponeurotic fibroma with diffuse intra-articular involvement of the carpal joints in a 59-year-old female. Radiographs and computed tomography scans revealed a large lobulated soft tissue mass with multiple stippled calcifications around the carpal joints and numerous erosions of the second to fifth carpometacarpal and intercarpal joints. Magnetic resonance imaging showed diffuse multinodular synovial proliferation with inhomogeneous hypo- to isointense signal intensity on T1-weighted images, inhomogeneous hypointense to hyperintense signal intensity on T2-weighted images, and inhomogeneous intense enhancement on fat-suppressed contrast-enhanced T1-weighted images. Radiologic diagnosis included gout, calcium pyrophosphate dihydrate deposition disease, and tenosynovial giant cell tumor. Surgical excision was performed, and the mass was diagnosed on pathologic examination as a calcifying aponeurotic fibroma. There has been no reported case of a calcifying aponeurotic fibroma with diffuse intra-articular involvement of the carpal joints in the literature.

Angioleiomyoma is a benign, vascular smooth muscle tumor originating from the tunica media of the vessel wall. In general, it typically arises in the cutaneous, subcutaneous tissue or fascia of the lower extremities in middle-aged women and is less than 2 cm in diameter. We report an unusual case of an angioleiomyoma of the sacral foramina in an 82-year-old man. MRI revealed a well-defined irregular-shaped deep-seated mass in the sacral foramina, showing branching pattern of growth associated with pressure bony erosion of the adjacent bones, with isointense to hypointense signal on T2-weighted images. Surgical excision was performed and the mass was diagnosed as angioleiomyoma on pathological examination. To the best of our knowledge, there has been no report of an angioleiomyoma involving the sacral foramina.

// Tae Hoen Kim 1, 2 , Ju-Yeon Jeong 2 , Ju-Yeon Park 2 , Se-Wha Kim 1, 2 , Jin Hyung Heo 1 , Haeyoun Kang 1, 2 , Gwangil Kim 1, 2 and Hee Jung An 1, 2 1 Department of Pathology, CHA Bundang Medical Center, CHA University, Gyeonggi-do, Korea 2 Institute for Clinical Research, CHA Bundang Medical Center, CHA University, Gyeonggi-do, Korea Correspondence to: Hee Jung An, email: hjahn@cha.ac.kr Keywords: ovarian cancer, miR-150, Notch3, chemoresistance, sensitization Received: March 26, 2017 Accepted: July 19, 2017 Published: August 18, 2017 ABSTRACT Tumor recurrence by obtaining chemoresistance is a major obstacle to treating ovarian cancer. By TargetScan database and a luciferase reporter assay, we identified miR-150 directly targets Notch3 , which is a key oncogene in ovarian cancer. We, therefore, investigated the role of miR-150 in ovarian cancer cells, and the usefulness of miR-150 as a therapeutic target in chemoresistant ovarian cancer, through examining miR-150 expression by qRT-PCR in ovarian cancer cell lines and tissues, and assessing the gain-of-function effect by WST, colony forming, TUNEL, wound healing and angiogenesis assays. Western blotting was performed to evaluate its downstream targets. The miR-150 expression was significantly downregulated in ovarian cancers. Treatment with pre-miR-150 significantly inhibited cancer cell proliferation, and induced apoptosis in PTX (paclitaxel) -resistant SKpac cells, which was not seen by PTX only treatment. On spheroid forming assay, an additional pre-miR-150 treatment with PTX decreased cancer stem cell activation in PTX-resistant SKpac cells. An experimental upregulation of miR-150 also decreased cancer cell migration and angiogenesis in SKpac cells. The Notch3 downstream proteins(NICD3 and HEY2), and cell cycle-related proteins (cyclinD3, pS6, and NF-kB), and apoptosis-related proteins (BCL-2 and BCL-W) were significantly downregulated by pre-miR-150 transfection. Taken together, miR-150 is related with PTX-resistance in ovarian cancer, and treatment with pre-miR-150 resensitizes cancer cells to PTX. Therefore, it may be a promising treatment strategy in chemoresistant and recurrent ovarian cancer.

The neoplastic cells in classical Hodgkin lymphoma (HL) have been difficult to analyze because ReedSternberg cells are relatively rare in involved tissues. HL infrequently presents in composite or metachronous settings with other lymphomas, offering opportunities to characterize HL genomes if both lymphomas follow similar evolutionary pathways. In composite Hodgkin and non-Hodgkin B-cell lymphomas, it has been generally assumed that the 2 lymphomas frequently share a common clonal origin. To our knowledge, the clonal relationships between composite or metachronous classical Hodgkin and non-HL have not previously been studied in a comprehensive comparison of their shared somatic mutations. We exome-sequenced a diffuse large B-cell lymphoma, and subsequently assessed for the presence of 93 somatic mutations in a metachronous HL in the same patient. Surprisingly, we did not detect any common mutations, suggesting clonal unrelatedness in this case. Similar to how lymphomas in different individuals accumulate unique sets of mutations, metachronous lymphomas might evolve separately even in the same individual. More generally, with the routine availability of next-generation sequencing techniques, such studies can provide novel insights into the pathobiology and mechanisms of composite lymphomas.

Desmoid type fibromatosis is a benign fibroblastic tumor arising from the fascia or musculoaponeurosis. It may occur in various locations, but most commonly in the shoulder girdle and neck; to our knowledge, there has been no reported case originating from a facet joint of the spine. We report CT and MR imaging findings of a desmoid type fibromatosis, involving the facet joint of the L3-4 spine with bone involvement.

To identify microRNAs (miRNAs) regulating Notch3 expression in association with paclitaxel resistance, candidate miRNAs targeting Notch3 were predicted using TargetScan. We found that miR-136 directly targets Notch3, and miR-136 was significantly downregulated in OSC tissues relative to normal control tissues, and low expression of miR-136 correlated with poor overall in ovarian cancer patients. Artificial miR-136 overexpression significantly reduced cell viability, proliferation, Cancer stem cell (CSC) spheroid formation, and angiogenesis, and increased apoptosis in paclitaxel-resistant SKpac cells compared with the effects of paclitaxel alone. miR-136 overexpression downregulated cell survival- (survivin, DNA-PK, pS6, S6) and cell cycle- (Cyclin D1, NF-κB) related proteins, and anti-apoptotic proteins (BCL2, and BCL-XL), and upregulated pro-apoptotic proteins (Bim, Bid, and Bax). Taken together, miR-136 targets the Notch3 oncogene and functions as a tumor suppressor. miR-136 overexpression resensitized paclitaxel-resistant ovarian cancer cells and reduced CSC activities, suggesting a promising new target for the treatment of chemoresistant ovarian cancers.

Kang H, An H-J, Song J-Y, Kim T-H, Heo J-H, Ahn D-H & Kim G (2012) Histopathology 61, 576–586 Notch3 and Jagged2 contribute to gastric cancer development and to glandular differentiation associated with MUC2 and MUC5AC expression Aims: Notch signalling plays diverse roles in malignant tumours as well as in normal tissue development. In this study we investigated the expression of Notch signalling pathway genes and their clinicopathological significance in gastric carcinomas. Methods and results: Notch1, Notch3, Jagged1, Jagged2 and Hes1 expression were analysed by quantitative real-time polymerase chain reaction (qRT–PCR) (n = 81) and immunohistochemistry (n = 103) in gastric carcinomas. MUC2 and MUC5AC expression were also assessed, using immunohistochemistry only. With qRT–PCR, Notch1, Notch3, Jagged1 and Jagged2 expression were increased significantly in tumour compared to normal tissue (P

Paragangliomas occur extremely rarely in the spermatic cord. A 40-year-old man presented with a scrotal mass that was diagnosed as spermatic cord paraganglioma with malignant histological features. To our knowledge, this is the first case of spermatic cord paraganglioma presented with malignant histological features evaluated by histological scoring. Careful evaluation of histological features and systematic evaluation should be considered for patients with spermatic cord paragangliomas.

The principal objective of this study was to comprehensively assess the clinicopathologic and prognostic impacts of the expression of various cell cycle regulatory proteins in patients with ovarian epithelial tumors. The tissue microarrays were constructed from formalin-fixed, paraffin-embedded tissues of 205 ovarian epithelial tumors. We investigated 55 benign (20 serous cystadenomas, 17 mucinous cystadenomas, and 18 endometriotic cysts), 72 borderline (26 serous borderline tumors and 46 mucinous borderline tumors), and 78 malignant tumors (45 serous carcinomas, 10 mucinous adenocarcinomas, 15 endometrioid adenocarcinomas, and 8 clear cell carcinomas). Immunohistochemical staining was performed using antibodies to p16(Ink4a), p53, p21(Waf1/Cip1), p27(Kip1), Cyclin D1, Cyclin E, Cyclin A, Cyclin B1, and Cyclin-dependent Kinase2. We noted significantly different levels of p53 and Cyclin B1 expressions between malignant and borderline tumors and between borderline and benign tumors excepting the mucinous type. The p21(Waf1/Cip1) and Cyclin A were significantly overexpressed in malignant tumors compared with borderline tumors. Cyclin A, Cyclin E, and p16(Ink4a) were more pronounced proteins, showing differential expression patterns among the histologic types of ovarian carcinomas. We determined that the overexpression of p16(Ink4a) (P=0.031), Cyclin E (P=0.009), and Cyclin-dependent Kinase2 (P=0.004) were significantly associated with higher tumor grades. Overexpression of p16(Ink4a) was correlated with both lymph node metastasis (P=0.030) and distant metastasis (P=0.034). Overexpression of Cyclin E was associated with advanced stage (P=0.004). Higher tumor grade (P=0.008), advanced stage (P=0.001), mucinous histologic type (P=0.012), low expression levels of p16(Ink4a) (P=0.032), and overexpression of p53 (P=0.032) were associated with poor overall survival on multivariate analysis in patients with ovarian cancer. In serous carcinomas, old age (P=0.005), distant metastasis (P=0.020), low expression of p16(Ink4a) (P=0.047), and overexpression of cytoplasmic p27(Kip1) (P=0.007) and Cyclin A (P=0.031) were all independent predictors of worse overall survival. Our data indicate that the overexpression of p16 and Cyclin E are valuable factors predicting disease progression and metastatic potential. Low p16(Ink4a) expression, overexpression of p53, cytoplasmic p27(Kip1), and Cyclin A are predictive markers for shorter overall survival in ovarian carcinomas.

Endometriosis is a common gynaecologic disease involving the ectopic growth of endometrial glands and stroma outside the uterus. The most commonly affected organs are the ovaries and other pelvic s...

Kim T H, Kim Y K, Kwon Y, Heo J H, Kang H, Kim G & An H J (2010) Histopathology57, 734–743 Deregulation of miR-519a, 153, and 485-5p and its clinicopathological relevance in ovarian epithelial tumours Aims: The pathological and clinical significance of aberrant miRNA expression in ovarian tumours has yet to be adequately documented. The aim of this study was to assess the differences in miRNA expression of human ovarian tumours according to histological subtype, and to determine whether miRNAs are potential diagnostic and prognostic markers in ovarian cancers. Method and results: The miRNA expression profiles of 103 human ovarian tumours were evaluated. Via a bead-based miRNA microarray, five aberrant miRNAs were selected which were expressed differentially in malignant serous tumours from borderline and benign ovarian tumours, including miRNA (miR)-519a, miR-18b (up-regulation) and miR-153, miR-511 and miR-485-5p (down-regulation). We conducted quantitative real-time reverse transcription–polymerase chain reaction (qRT–PCR) in order to confirm that these miRNAs are differentially expressed in different histological subtypes of ovarian tumours, and compared the expression profiles of these miRNAs between different clinical subsets. The expression of these miRNAs was correlated with clinicopathological parameters. miR-519a, miR-153 and miR-485-5p were differentially expressed in four major histotypes of ovarian cancers (P

Ovarian cancer is a gynecological malignancy with the highest mortality. Chemoresistance is an important subject for the treatment of ovarian cancer, because obtaining significant drug resistance to the first line chemotherapy, paclitaxel, causes major therapeutic obstacles. It is essential to improve the survival rate of ovarian cancer patients by mining the biomarkers indicating the drug resistance and prognosis, and by further understanding underlying mechanisms of drug resistance. In the present study, we established paclitaxel-resistant subline (SKpac) from human epithelial ovarian cancer cell line, SKOV3, and performed comparative analysis of whole proteomes between paclitaxel-resistant SKpac sublines and paclitaxel-sensitive parental SKOV3 cells to identify differentially expressed proteins and useful biomarkers indicating chemoresistance. Proteins related to chemoresistant process were identified by two-dimensional gel electrophoresis (2DE) with mass spectrometry (MALDI-TOF and LC-MS/MS). Eighteen spots were differentially expressed and were identified in SKpac chemoresistant cells compared to SKOV3. The expressions of ALDH 1A1, annexin A1, hnRNP A2, and GDI 2 proteins were validated by Western blot, which was consistent with proteomic analysis. Among the selected proteins, downregulation of hnRNP A2 and GDI 2 was found to be the most significant finding in SKpac cells and chemoresistant ovarian cancer tissues. Our results suggest that hnRNP A2 and GDI 2 may represent potential biomarkers of the paclitaxel-resistant ovarian cancers for tailored cancer therapy.

With the recent development of molecular markers, it has become possible to characterize colorectal carcinomas beyond clinical and histologic aspects. When considered together with tumor stage, molecular markers will allow us further insight into individual tumor biologies and prognoses. To investigate the expression and prognostic implications of the molecular markers, p53, bcl-2, Rb, hMLH1, hMSH2, β-catenin, E-cadherin, and MUC-2. We analyzed the clinical, histologic, and molecular factors for 229 patients with colorectal carcinoma of stage II and III and compared their prognoses. We used tissue microarrays to analyze the expressions of molecular markers and to assess their correlations with prognosis. Semiquantitative expressions of molecular markers and clinicopathologic parameters were analyzed with respect to prognosis. Among the clinicopathologic parameters, left-sided location, age older than 70 years, higher preoperative serum carcinoembryonic antigen (CEA) level (≤ 5 ng/mL), irregular growth pattern, and perineural invasion were significantly related to poor prognosis in stage II and III patients. For molecular factors, loss of expression of E-cadherin and MUC-2 showed significant correlation with poor overall survival in both cancer stages. Multivariate analysis showed that higher TNM stage, higher preoperative serum CEA level (≤ 5 ng/mL), perineural tumor cell invasion, and loss of E-cadherin and MUC-2 expressions were independently correlated with poor overall survival. Our results indicated that of the analyzed molecular markers, MUC-2 and E-cadherin might be useful in predicting prognosis and planning for adjuvant therapy in patients with stage II and III colorectal carcinomas.

Background and aims: The aim of the present study is to evaluate the difference of immunophenotypes of cyclin A, skp2 and p27 in ovarian epithelial tumors and to determine the potential role of these molecules in ovarian cancer development and progression. Methods: Immunohistochemical staining for cyclin A, skp2 and p27 was performed using tissue microarray blocks that contained 211 ovarian epithelial tumors including 78 carcinomas (43 serous, 10 mucinous, 17 endometrioid, 8 clear cell), 73 borderline tumors (26 serous, 47 mucinous) and 60 benign tumors. Results: Cyclin A and skp2 overexpression were more frequently detected in malignant tumors of all histological subtype than in borderline and benign tumors except clear cell type (cyclin A: serous 77%; P < 0.0001, mucinous 40%; P < 0.0001, and endometrioid 59%; P < 0.0001, Skp2: serous 42%; P= 0.0004, mucinous 50%; P= 0.003, and endometrioid 59%; P= 0.0003). In addition, cyclin A expression was significantly associated with higher tumor grade (P= 0.034) and advanced clinical stages (P= 0.011). Loss of p27 expression was observed more frequently in the benign and borderline mucinous tuomors, but there is no statistically significant differences. Conclusions: Cyclin A and skp2 expression is related to the development of ovarian carcinomas. However, we could not find a significant alteration of p27 expression in ovarian adenocarcinomas.

Aims: Claudin 2 (CLDN2) is a family of integral membrane tight junctions. The aim was to determine the influence of CLDN2 expression on tumour behaviour and its role in breast carcinogenesis. Method and results: Thirty-seven invasive breast carcinomas and corresponding normal breast tissues were examined for CLDN2 protein and mRNA expression using Western blotting and semiquantitative reverse transcriptase-polymerase chain reaction. The expression of CLDN2 protein in 118 cases of breast carcinoma was further studied with immunohistochemistry and related to various clinicopathological parameters. CLDN2 protein expression was significantly down-regulated (0.4-fold) in tumours compared with corresponding normal breast tissue (P

Sox10 is a transcription factor regulating the development of several cell lineages and is involved in tumor development. However, the clinicopathological relevance of Sox10 expression in ovarian cancer has not been examined. We assessed expression of Sox10 in ovarian epithelial tumors by immunohistochemistry and assessed its prognostic value by analyzing the correlation between its expression and clinicopathological factors. We used tissue microarrays including 244 ovarian epithelial tumors. Sox10 staining was found in the cytoplasm or nucleus of tumor cells. Malignant serous, mucinous, and endometrioid tumors were significantly more likely to express Sox10 than benign and borderline tumors. Expression patterns in adenocarcinomas were different for histologic subtypes: nuclear Sox10 staining was common in clear-cell adenocarcinomas and serous adenocarcinomas, whereas all cases of mucinous and endometrioid tumors were negative for nuclear staining. Nuclear Sox10 staining was also associated with chemoresistance and shorter overall survival in ovarian adenocarcinomas, notably in high-grade serous adenocarcinoma. Sox10 is expressed in many ovarian carcinomas, suggesting that it might be involved in oncogenesis of ovarian carcinoma. Expression pattern of Sox10 differs between histological subtypes. Nuclear Sox10 expression is an independent indicator of poor prognosis in ovarian adenocarcinomas, notably in high-grade serous adenocarcinomas.

Intratumoural mutational heterogeneity (ITH) or the presence of different private mutations in different parts of the same tumour is commonly observed in human tumours. The mechanisms generating such ITH are uncertain. Here we find that ITH can be remarkably well structured by measuring point mutations, chromosome copy numbers, and DNA passenger methylation from opposite sides and individual glands of a 6 cm human colorectal adenoma. ITH was present between tumour sides and individual glands, but the private mutations were side-specific and subdivided the adenoma into two major subclones. Furthermore, ITH disappeared within individual glands because the glands were clonal populations composed of cells with identical mutant genotypes. Despite mutation clonality, the glands were relatively old, diverse populations when their individual cells were compared for passenger methylation and by FISH. These observations can be organized into an expanding star-like ancestral tree with co-clonal expansion, where many private mutations and multiple related clones arise during the first few divisions. As a consequence, most detectable mutational ITH in the final tumour originates from the first few divisions. Much of the early history of a tumour, especially the first few divisions, may be embedded within the detectable ITH of tumour genomes.

Urachal anomalies are relatively uncommon and result from incomplete obliteration of the urachus perinatally. In children, most urachal diseases including urachal cysts and sinuses are benign, and these can sometimes become secondarily infected. Malignant involvement of the urachus is rarely reported, one in 5 million people, accounting for 0.35% to 0.7% of all bladder cancers. There are only five cases of urachal cancer diagnosed at the age of twenties in English written literature. Age at the diagnosis of urachal carcinoma is important to understand pathogenetic transition from benign to malignancy. A 26-year-old man visited our clinic with gross hematuria starting a few months before. CT scan showed a 4.0×6.8 cm sized lobulated cystic mass over the bladder dome. Cystoscopy showed a ball-shaped extrinsic mass from the bladder dome with intact bladder mucosa. With an impression of urachal cancer, laparoscopic partial cystectomy with wide excision of urachus was performed. Final diagnosis was well differentiated mucinous urachal adenocarcinoma invading bladder muscle, staged as pT3a based on Sheldon’s staging system. To our best knowledge, this case is the youngest Korean case of urachal carcinoma (the fourth youngest ever in English written literature).

Notch signaling plays an important role in ovarian cancer chemoresistance, which is responsible for recurrence. Gamma-secretase inhibitor (GSI) is a broad-spectrum Notch inhibitor, but it has serious side effects. The efficacy of Notch3-specific inhibition in paclitaxel-resistant ovarian cancers was assessed in this study, which has not yet been evaluated relative to GSI. To analyze the effect of Notch3-specific inhibition on paclitaxel-resistant ovarian cancers, we compared cell viability, apoptosis, cell migration, angiogenesis, cell cycle, and spheroid formation after treatment with either Notch3 siRNA or GSI in paclitaxel-resistant SKpac cells and parental SKOV3 cells. Expression levels of survival, cell cycle, and apoptosis-related proteins were measured and compared between groups. Notch3 was significantly overexpressed in chemoresistant cancer tissues and cell lines relative to chemosensitive group. In paclitaxel-resistant cancer cells, Notch inhibition significantly reduced viability, migration, and angiogenesis and increased apoptosis, thereby boosting sensitivity to paclitaxel. Spheroid formation was also significantly reduced. Both Notch3 siRNA-treated cells and GSI-treated cells arrested in the G2/M phase of the cell cycle. Proteins of cell survival, cyclin D1 and cyclin D3 were reduced, whereas p21 and p27 were elevated. Both GSI and Notch3 siRNA treatment reduced expression of anti-apoptotic proteins (BCL-W, BCL2, and BCL-XL) and increased expression of pro-apoptotic proteins (Bad, Bak, Bim, Bid, and Bax). These results indicate that Notch3-specific inhibition sensitizes paclitaxel-resistant cancer cells to paclitaxel treatment, with an efficacy comparable to that of GSI. This approach would be likely to avoid the side effects of broad-spectrum GSI treatment. © 2015 Wiley Periodicals, Inc.

Purpose: Although most stage II colon cancers are potentially curable by surgery alone, ∼20% of patients relapse, suggesting a need for establishing prognostic markers that can identify patients who may benefit from adjuvant chemotherapy. We tested the hypothesis that differences in expression of apoptosis-regulating proteins account for differences in clinical outcome among patients with early-stage colorectal cancer. Experimental Design: Tissue microarray technology was employed to assay the expression of apoptosis-regulating proteins by immunohistochemistry in 106 archival stage II colorectal cancers, making correlations with disease-specific survival. The influence of microsatellite instability (MSI), tumor location (left versus right side), patient age, and gender was also examined. Results: Elevated expression of several apoptosis regulators significantly correlated with either shorter (cIAP2; TUCAN) or longer (Apaf1; Bcl-2) overall survival in univariate and multivariate analyses. These biomarkers retained prognostic significance when adjusting for MSI, tumor location, patient age, and gender. Moreover, certain combinations of apoptosis biomarkers were highly predictive of death risk from cancer. For example, 97% of patients with favorable tumor phenotype of cIAP2low plus TUCANlow were alive at 5 years compared with 60% of other patients (P = 0.00003). In contrast, only 37% of patients with adverse biomarkers (Apaf1low plus TUCANhigh) survived compared with 83% of others at 5 years after diagnosis (P< 0.0001). Conclusions: Immunohistochemical assays directed at detection of certain combinations of apoptosis proteins may provide prognostic information for patients with early-stage colorectal cancer, and therefore could help to identify patients who might benefit from adjuvant chemotherapy or who should be spared it.

The amplification and overexpression of Her2 proto-oncogene have been found to be associated with the development and progression of human breast cancer. A polymorphic valine allele at codon 655 of the Her2 gene (Her2(V655)) was suggested by some authors to be a susceptible genetic factor for the development of breast cancer. The Her2 polymorphism at codon 655 was investigated in 304 Korean women including 177 patients with breast cancer. The association between Her2 genotype and Her2 protein overexpression was also examined in breast cancers by immunohistochemistry. Her2(V655) was not associated with a significant breast cancer risk (odds ratio (OR), 1.792; 95% confidence interval (CI), 0.459-6.991). The frequency of homozygous or heterozygous valine allele increased in stage 2 patients (OR, 1.67; 95% CI, 0.67-4.19), and patients in stages 3 and 4 (OR, 3.36; 95% CI, 0.85-13.42) compared to patients in stage 0. However, an association between the presence of the valine allele and the overexpression of Her2 protein could not be demonstrated. These results suggest that Her2 polymorphism at codon 655 is not associated with the development of breast cancer in Korean women. However, there is a possibility that the valine allele at codon 655 might be related to increased risk of breast cancer progression.

Background Solitary myofibroma of the spine is extremely rare, particularly among adults. To the best of our knowledge, only 3 cases affecting lumbar vertebrae have been reported in the English language literature. Of them, only 1 case was an adult case of solitary myofibroma affecting the L1 vertebra. Methods We report a case of solitary myofibroma affecting the L5 vertebra in an 18-year-old man and the postoperative imaging of solitary myofibroma for the first time. Conventional radiographs demonstrated an expansile osteolytic lesion with thinned cortex and marginal sclerosis. Computed tomography (CT) showed a purely osteolytic expansile lesion with partial disappearance of thinned cortex. MRI of the lesion revealed an isointense signal on T1-weighted images, an inhomogeneous slightly hyperintense signal on T2-weighed images, and homogeneous avid enhancement with gadolinium. Results Surgical excision was performed and the lesion was diagnosed as solitary myofibroma on pathological examination. One-year follow-up postoperative CT demonstrated decreased size of the osteolytic lesion with sclerotic change. Four-year follow-up postoperative MRI revealed complete resolution of the lesion replaced by normal fatty marrow. Conclusion If a benign-looking expansile osteolytic lesion reveals a homogeneously isointense signal on T1-weighted image, inhomogeneous slightly hyperintense signal on T2-weighted image, and homogeneous avid enhancement with gadolinium, solitary myofibroma should be considered in the differential diagnosis of spine bone tumors. It can be resolved completely.

Subcutaneous bronchogenic cysts have been described rarely, particularly among adolescents. Only a few reports have described the ultrasonographic features of bronchogenic cysts, characterizing them as nonspecific cystic masses with or without internal echogenic foci or debris. Therefore, it is hard to differentiate subcutaneous bronchogenic cysts from other subcutaneous cystic tumors ultrasonographically.We report a case of presternal subcutaneous bronchogenic cyst in an 18-year-old man with unusual ultrasonographic findings. Ultrasonography revealed a small, oval, cystic mass containing a well-circumscribed, heterogeneously hypoechoic, egg-shaped lesion in the dependent portion of the mass within the subcutaneous fat layer overlying the sternum.Surgical excision was performed, and the cystic mass was diagnosed as a bronchogenic cyst. On pathological examination, the internal, heterogeneously hypoechoic, ball-like lesion was found to be mucous material within the cyst. To our knowledge, this is the first reported case of a presternal subcutaneous bronchogenic cyst presenting with a ball-like lesion inside of the cyst. This unusual ultrasonographic feature can be a clue to the diagnosis of subcutaneous bronchogenic cyst.In conclusion, if an anechoic cyst containing an internal, well-circumscribed, hypoechoic ball-like lesion is seen in the presternal subcutaneous fat layer, subcutaneous bronchogenic cyst should be considered in the differential diagnosis of subcutaneous cystic masses.

Chondromyxoid fibroma is a rare benign bone tumor, which represents less than 1% of primary bone tumors. However, chondromyxoid fibroma developing in the soft tissue is extremely rare. We report the sonographic findings in a case of soft tissue chondromyxoid fibroma in the foot confirmed pathologically.

What happens in the early, still undetectable human malignancy is unknown because direct observations are impractical. Here we present and validate a “Big Bang” model, whereby tumors grow predominantly as a single expansion producing numerous intermixed sub-clones that are not subject to stringent selection, and where both public (clonal) and most detectable private (subclonal) alterations arise early during growth. Genomic profiling of 349 individual glands from 15 colorectal tumors revealed the absence of selective sweeps, uniformly high intra-tumor heterogeneity (ITH), and sub-clone mixing in distant regions, as postulated by our model. We also verified the prediction that most detectable ITH originates from early private alterations, and not from later clonal expansions, thus exposing the profile of the primordial tumor. Moreover, some tumors appear born-to-be-bad, with sub-clone mixing indicative of early malignant potential. This new model provides a quantitative framework to interpret tumor growth dynamics and the origins of ITH with significant clinical implications.