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Abstract Background The Stanford Tissue Microarray Database (TMAD) is a repository of data serving a consortium of pathologists and biomedical researchers. The tissue samples in TMAD are annotated with multiple free-text fields, specifying the pathological diagnoses for each sample. These text annotations are not structured according to any ontology, making future integration of this resource with other biological and clinical data difficult. Results We developed methods to map these annotations to the NCI thesaurus. Using the NCI-T we can effectively represent annotations for about 86% of the samples. We demonstrate how this mapping enables ontology driven integration and querying of tissue microarray data. We have deployed the mapping and ontology driven querying tools at the TMAD site for general use. Conclusion We have demonstrated that we can effectively map the diagnosis-related terms describing a sample in TMAD to the NCI-T. The NCI thesaurus terms have a wide coverage and provide terms for about 86% of the samples. In our opinion the NCI thesaurus can facilitate integration of this resource with other biological data.

Background: MicroRNAs (miRNA) are 20~25 nucleotide non-coding RNAs that inhibit the translation of targeted mRNA, and they have been implicated in the development of human malignancies. High grade serous ovarian carcinomas, the most common and lethal subtype of ovarian cancer, can occur sporadically or in the setting of BRCA1/2 syndromes. Little is known regarding the miRNA expression profiles of high grade serous carcinoma in relation to BRCA1/2 status, and compared to normal tubal epithelium, the putative tissue of origin for high grade serous carcinomas. Methodology/Principal Findings: Global miRNA expression profiling was performed on a series of 33 high grade serous carcinomas, characterized with respect to BRCA1/2 status (mutation, epigenetic silencing with loss of expression or normal), and with clinical follow-up, together with 2 low grade serous carcinomas, 2 serous borderline tumors, and 3 normal fallopian tube samples, using miRNA microarrays (328 human miRNA). Unsupervised hierarchical clustering based on miRNA expression profiles showed no clear separation between the groups of carcinomas with different BRCA1/2 status. There were relatively few miRNAs that were differentially expressed between the genotypic subgroups. Comparison of 33 high grade serous carcinomas to 3 normal fallopian tube samples identified several dysregulated miRNAs (false discovery rate <5%), including miR-422b and miR-34c. Quantitative RT-PCR analysis performed on selected miRNAs confirmed the pattern of differential expression shown by microarray analysis. Prognostically, lower level miR-422b and miR-34c in high grade serous carcinomas were both associated with decreased disease-specific survival by Kaplan-Meier analysis (p<0.05). Conclusions/Significance: High grade serous ovarian carcinomas with and without BRCA1/2 abnormalities demonstrate very similar miRNA expression profiles. High grade serous carcinomas as a group exhibit significant miRNA dysregulation in comparison to tubal epithelium and the levels of miR-34c and miR-422b appear to be prognostically important. [ABSTRACT FROM AUTHOR]

Background: The Stanford Tissue Microarray Database (TMAD) is a repository of data serving a consortium of pathologists and biomedical researchers. The tissue samples in TMAD are annotated with multiple free-text fields, specifying the pathological diagnoses for each sample. These text annotations are not structured according to any ontology, making future integration of this resource with other biological and clinical data difficult. Results: We developed methods to map these annotations to the NCI thesaurus. Using the NCIT we can effectively represent annotations for about 86% of the samples. We demonstrate how this mapping enables ontology driven integration and querying of tissue microarray data. We have deployed the mapping and ontology driven querying tools at the TMAD site for general use. Conclusion: We have demonstrated that we can effectively map the diagnosis-related terms describing a sample in TMAD to the NCI-T. The NCI thesaurus terms have a wide coverage and provide terms for about 86% of the samples. In our opinion the NCI thesaurus can facilitate integration of this resource with other biological data. [ABSTRACT FROM AUTHOR]

Purpose: Leiomyosarcoma is a malignant neoplasm with smooth muscle differentiation. Little is known about its molecular heterogeneity and no targeted therapy currently exists for leiomyosarcoma. Recognition of different molecular subtypes is necessary to evaluate novel therapeutic options. In a previous study on 51 leiomyosarcomas, we identified three molecular subtypes in leiomyosarcoma. The current study was performed to determine whether the existence of these subtypes could be confirmed in independent cohorts., Experimental Design: Ninety-nine cases of leiomyosarcoma were expression profiled with 3'end RNA-Sequencing (3SEQ). Consensus clustering was conducted to determine the optimal number of subtypes., Results: We identified 3 leiomyosarcoma molecular subtypes and confirmed this finding by analyzing publically available data on 82 leiomyosarcoma from The Cancer Genome Atlas (TCGA). We identified two new formalin-fixed, paraffin-embedded tissue-compatible diagnostic immunohistochemical markers; LMOD1 for subtype I leiomyosarcoma and ARL4C for subtype II leiomyosarcoma. A leiomyosarcoma tissue microarray with known clinical outcome was used to show that subtype I leiomyosarcoma is associated with good outcome in extrauterine leiomyosarcoma while subtype II leiomyosarcoma is associated with poor prognosis in both uterine and extrauterine leiomyosarcoma. The leiomyosarcoma subtypes showed significant differences in expression levels for genes for which novel targeted therapies are being developed, suggesting that leiomyosarcoma subtypes may respond differentially to these targeted therapies., Conclusions: We confirm the existence of 3 molecular subtypes in leiomyosarcoma using two independent datasets and show that the different molecular subtypes are associated with distinct clinical outcomes. The findings offer an opportunity for treating leiomyosarcoma in a subtype-specific targeted approach., (©2015 American Association for Cancer Research.)

Endometrial stromal tumors may pose diagnostic challenges particularly when they exhibit variant histologic appearances, involve extrauterine sites, or present as metastatic disease. In such cases, use of immunohistochemical markers and identification of specific nonrandom chromosomal rearrangements may be helpful. Over the last decade, fluorescence in situ hybridization (FISH) has been progressively incorporated as a diagnostic tool for the evaluation of endometrial stromal tumors. The purpose of this study was to review a series of these tumors and compare the results of FISH analysis with the clinicopathologic characteristics. Three endometrial stromal nodules (ESNs), 13 endometrial stromal sarcomas (ESSs), and 7 undifferentiated endometrial sarcomas (UESs) were reviewed. Three metastases from 1 of the ESS cases were also analyzed. Nine of these tumors (1 ESN, 8 ESSs, and 1 UES) exhibited unusual histologic features, including smooth muscle (3), sex cord (7), epithelioid (1), fibromyxoid (1), and skeletal muscle (2) differentiation. A tissue microarray was prepared, and FISH analysis was performed using break-apart and fusion probes for JAZF1, SUZ12, EPC1, and PHF1 genes. FISH was successful in 22 cases, and rearrangements involving JAZF1, SUZ12, EPC1, and PHF1 genes were detected in 10 of the 22 (45%) uterine tumors, including 2 of the 3 ESNs and 8 of 12 ESSs. Genetic rearrangements were found neither in the 3 metastases of the ESS nor in any of the UESs. It is noteworthy that a correlation between sex cord differentiation and PHF1 rearrangement was encountered in ESSs (P=0.008). In our series, all ESSs showing sex cords had PHF1 genetic rearrangement, suggesting that such rearrangements may induce sex cord differentiation.

Introduction: High numbers of tumor-associated macrophages (TAMs) have been associated with poor outcome in several solid tumors. In 2 previous studies, we showed that colony stimulating factor-1 (CSF1) is secreted by leiomyosarcoma (LMS) and that the increase in macrophages and CSF1 associated proteins are markers for poor prognosis in both gynecologic and nongynecologic LMS in a multicentered study. The purpose of this study is to evaluate the outcome of patients with LMS from a single institution according to the number of TAMs evaluated through 3 CSF1 associated proteins., Methods: Patients with LMS treated at Stanford University with adequate archived tissue and clinical data were eligible for this retrospective study. Data from chart reviews included tumor site, size, grade, stage, treatment, and disease status at the time of last follow-up. The 3 CSF1 associated proteins (CD163, CD16, and cathepsin L) were evaluated by immunohistochemistry on tissue microarrays. Kaplan-Meier survival curves and univariate Cox proportional hazards models were fit to assess the association of clinical predictors as well as CSF1 associated proteins with overall survival., Results: A total of 52 patients diagnosed from 1983 to 2007 were evaluated. Univariate Cox proportional hazards models were fit to assess the significance of grade, size, stage, and the 3 CSF1 associated proteins in predicting OS. Grade, size, and stage were not significantly associated with survival in the full patient cohort, but grade and stage were significant predictors of survival in the gynecologic (GYN) LMS samples (P = 0.038 and P = 0.0164, respectively). Increased cathepsin L was associated with a worse outcome in GYN LMS (P = 0.049). Similar findings were seen with CD16 (P < 0.0001). In addition, CSF1 response enriched (all 3 stains positive) GYN LMS had a poor overall survival when compared with CSF1 response poor tumors (P = 0.001). These results were not seen in non-GYN LMS., Conclusions: Our data form an independent confirmation of the prognostic significance of TAMs and the CSF1 associated proteins in LMS. More aggressive or targeted therapies could be considered in the subset of LMS patients that highly express these markers.

Myometrial invasion is an independent prognostic parameter of the endometrioid carcinomas which correlates with the risk of metastasis to pelvic and/or paraaortic lymph nodes. Recognition of myometrial invasion is sometimes difficult. In fact, myoinvasion is overdiagnosed in routine practice in as many as 25% of the cases. Recently, it has been observed that tumor-associated macrophages stimulate angiogenesis and promote cancer dissemination. Tumor macrophages (CD163), microvessel density (CD31), and hypoxia inducible factor 1 α subunit (HIF1A) were investigated in 64 primary endometrioid carcinomas with (50 cases) and without (14 cases) myometrial invasion as well as in the corresponding regional lymph nodes metastases of 20 of the myoinvasive tumors. Endometrioid carcinomas with myometrial invasion showed higher number of CD163-tumor macrophages and greater microvessel density than endometrioid carcinomas without myometrial invasion (P=0.000 and P=0.000, respectively). In carcinomas confined to the corpus uteri (stage I), expression of HIF1A was associated with deep myoinvasion (stage IC) (P=0.006). There was a significant relationship between microvessel density and CD163-macrophages both in the myoinvasive and nonmyoinvasive tumors. On the other hand, high-grade endometrioid carcinomas had more macrophage infiltrates and microvessels than low-grade tumors (P=0.03 and P=0.07). Also, there was a positive correlation between CD163-macrophages and microvessel density in the primary tumors and their corresponding regional lymph node metastases. These findings link increased microvessel proliferation to stromal macrophage infiltrate and suggest that enhanced tumor angiogenesis, triggered by stromal macrophages, regulates the progression of endometrioid carcinomas. The identical stroma microenvironment found in the primary and the corresponding metastatic tumor suggests that tumor stroma response is determined by the intrinsic biology of the tumor.

DOG1 (discovered on GIST 1), known also as TMEM16A and ANO1, has emerged in recent years as a promising biomarker for gastrointestinal stromal tumors (GIST). It was originally discovered through microarray expression profiling analysis as gene that is highly expressed in GIST, and subsequent immunohistochemical studies have shown its use in its diagnosis. The results from several series have shown a high overall sensitivity and specificity for DOG1 in the detection of GISTs and about 6% of GISTs overall exhibiting a DOG1+/KIT-immunoprofile. DOG1 antibodies are more sensitive than KIT antibodies in detecting tumors of gastric origin, tumors with epithelioid morphology, and tumors harboring PDGFRA mutation. Furthermore, DOG1 immunoreactivity is rarely observed in other mesenchymal and nonmesenchymal tumor types. These results support the use of DOG1 as a diagnostic biomarker for GIST. When used in combination with KIT, this panel of diagnostic biomarkers can help pathologists and clinicians to identify more patients who may benefit from targeted therapies.

Background: MicroRNAs (miRNA) are 20 approximately 25 nucleotide non-coding RNAs that inhibit the translation of targeted mRNA, and they have been implicated in the development of human malignancies. High grade serous ovarian carcinomas, the most common and lethal subtype of ovarian cancer, can occur sporadically or in the setting of BRCA1/2 syndromes. Little is known regarding the miRNA expression profiles of high grade serous carcinoma in relation to BRCA1/2 status, and compared to normal tubal epithelium, the putative tissue of origin for high grade serous carcinomas., Methodology/principal Findings: Global miRNA expression profiling was performed on a series of 33 high grade serous carcinomas, characterized with respect to BRCA1/2 status (mutation, epigenetic silencing with loss of expression or normal), and with clinical follow-up, together with 2 low grade serous carcinomas, 2 serous borderline tumors, and 3 normal fallopian tube samples, using miRNA microarrays (328 human miRNA). Unsupervised hierarchical clustering based on miRNA expression profiles showed no clear separation between the groups of carcinomas with different BRCA1/2 status. There were relatively few miRNAs that were differentially expressed between the genotypic subgroups. Comparison of 33 high grade serous carcinomas to 3 normal fallopian tube samples identified several dysregulated miRNAs (false discovery rate <5%), including miR-422b and miR-34c. Quantitative RT-PCR analysis performed on selected miRNAs confirmed the pattern of differential expression shown by microarray analysis. Prognostically, lower level miR-422b and miR-34c in high grade serous carcinomas were both associated with decreased disease-specific survival by Kaplan-Meier analysis (p<0.05)., Conclusions/significance: High grade serous ovarian carcinomas with and without BRCA1/2 abnormalities demonstrate very similar miRNA expression profiles. High grade serous carcinomas as a group exhibit significant miRNA dysregulation in comparison to tubal epithelium and the levels of miR-34c and miR-422b appear to be prognostically important.

Previously, we showed that the presence of high numbers of macrophages correlates with poor prognosis in nongynecological leiomyosarcoma (LMS). In gynecological LMS, a similar trend was noted but did not reach statistical significance. Colony-stimulating factor-1 (CSF1) is a major chemoattractant for macrophages. Here we show that in a subset of LMS cases, CSF1 is expressed by the malignant cells. Previously, we found that CSF1 is translocated and highly expressed in tenosynovial giant cell tumors (TGCTs), and this observation allowed us to identify genes that showed a coordinate expression with CSF1. Here, we evaluated the expression of CSF1 and TGCT-associated proteins in 149 cases of LMS. The coordinate expression of CSF1 and three TGCT-associated proteins (CD163, FCGR3a, and CTSL1) identified cases with poor prognosis in both gynecological LMS (P = 0.00006) and nongynecological LMS (P = 0.03). In gynecological LMS, the coordinate expression of these four markers was the only independent prognosticator in multivariate analysis (hazard ratio, 4.2; 95% CI, 1.12 to 16; P = 0.03). Our findings indicate that CSF1 may play an important role in the clinical behavior of LMS that may open a window for new therapeutic reagents.

Purpose: Macrophages play an important role in breast carcinogenesis. The pathways that mediate the macrophage contribution to breast cancer and the heterogeneity that exists within macrophages are incompletely understood. Macrophage colony-stimulating factor 1 (CSF1) is the primary regulator of tissue macrophages. The purpose of this study was to define a novel CSF1 response signature and to evaluate its clinical and biological significance in breast cancer., Experimental Design: We defined the CSF1 response signature by identifying genes overexpressed in tenosynovial giant cell tumor and pigmented villonodular synovitis (tumors composed predominantly of macrophages recruited in response to the overexpression of CSF1) compared with desmoid-type fibromatosis and solitary fibrous tumor. To characterize the CSF1 response signature in breast cancer, we analyzed the expression of CSF1 response signature genes in eight published breast cancer gene expression data sets (n = 982) and did immunohistochemistry and in situ hybridization for CSF1 response genes on a breast cancer tissue microarray (n = 283)., Results: In both the gene microarray and tissue microarray analyses, a consistent subset (17-25%) of breast cancers shows the CSF1 response signature. The signature is associated with higher tumor grade, decreased expression of estrogen receptor, decreased expression of progesterone receptor, and increased TP53 mutations (P < 0.001)., Conclusions: Our data show that the CSF1 response signature is consistently seen in a subset of breast carcinomas and correlates with biological features of the tumor. Our findings provide insight into macrophage biology and may facilitate the development of personalized therapy for patients most likely to benefit from CSF1-targeted treatments.

Purpose: Macrophages are migratory cells that are frequently recruited to the site of tumors. Their presence is associated with poor clinical outcome in a variety of epithelial malignancies. The aim of this study is to examine the prognostic significance of tumor-associated macrophages in sarcomas., Experimental Design: Global gene expression profiling data of a series of soft tissue tumors were analyzed for macrophage-associated gene expression. Immunohistochemistry on tissue microarrays containing leiomyosarcoma cases with known clinical outcome was used to verify the presence of macrophages and to examine the relationship between tumor-associated macrophages and clinical outcome., Results: Gene expression profiling revealed high-level expression of several macrophage-associated genes such as CD163 and CD68 in a subset of leiomyosarcomas, indicating the presence of variable numbers of tumor-infiltrating macrophages. This was confirmed by CD68 and CD163 immunostaining of a tissue microarray containing 149 primary leiomyosarcomas. Kaplan-Meier survival analysis showed that high density of tumor-infiltrating macrophages as identified by CD163 or CD68 staining is associated with a significantly worse disease-specific survival in nongynecologic leiomyosarcomas, whereas leiomyosarcomas arising from the gynecologic tract showed no significant association between macrophage infiltration and survival. The presence of tumor necrosis did not correlate significantly with outcome., Conclusions: An increased density of CD163- or CD68-positive tumor-infiltrating macrophages is associated with poor outcome in nongynecologic leiomyosarcomas. This may help the clinical management of patients with leiomyosarcomas.

Gastrointestinal stromal tumors (GIST) occur primarily in the wall of the intestine and are characterized by activating mutations in the receptor tyrosine kinases genes KIT or PDGFRA. The diagnosis of GIST relies heavily on the demonstration of KIT/CD117 protein expression by immunohistochemistry. However, KIT expression is absent in approximately 4% to 15% of GIST and this can complicate the diagnosis of GIST in patients who may benefit from treatment with receptor tyrosine kinase inhibitors. We previously identified DOG1/TMEM16A as a novel marker for GIST using a conventional rabbit antipeptide antiserum and an in situ hybridization probe. Here, we describe 2 new monoclonal antibodies against DOG1 (DOG1.1 and DOG1.3) and compare their staining profiles with KIT and CD34 antibodies on 447 cases of GIST. These included 306 cases with known mutational status for KIT and PDGFRA from a molecular consultation service. In addition, 935 other mesenchymal tumors and 432 nonsarcomatous tumors were studied. Both DOG1 antibodies showed high sensitivity and specificity for GIST, with DOG1.1 showing some advantages. This antibody yielded positive staining in 370 of 425 (87%) scorable GIST, whereas CD117 was positive in 317 of 428 (74%) GIST and CD34 in 254 of 430 (59%) GIST. In GIST with mutations in PDGFRA, 79% (23/29) showed DOG1.1 immunoreactivity while only 9% (3/32) and 27% (9/33) stained for CD117 and CD34, respectively. Only 1 of 326 (0.3%) leiomyosarcomas and 1 of 39 (2.5%) synovial sarcomas among the 935 soft tissue tumors examined showed positive immunostaining for DOG1.1. In addition, DOG1.1 immunoreactivity was seen in fewer cases of carcinoma, melanoma, and seminoma as compared with KIT.