Unveiling the Genomic Landscape of Intraductal Carcinoma of the Prostate Using Spatial Gene Expression Analysis.

Intraductal carcinoma of the prostate (IDCP) has recently attracted increasing interest owing to its unfavorable prognoses. To effectively identify the IDCP-specific gene expression profile, we took a novel approach of characterizing a typical IDCP case using spatial gene expression analysis. A formalin-fixed, paraffin-embedded sample was subjected to Visium CytAssist Spatial Gene Expression analysis. IDCP within invasive prostate cancer sites was recognized as a distinct cluster separate from other invasive cancer clusters. Highly expressed genes defining the IDCP cluster, such as MUC6, MYO16, NPY, and KLK12, reflected the aggressive nature of high-grade prostate cancer. IDCP sites also showed increased hypoxia markers HIF1A, BNIP3L, PDK1, and POGLUT1; decreased fibroblast markers COL1A2, DCN, and LUM; and decreased immune cell markers CCR5 and FCGR3A. Overall, these findings indicate that the hypoxic tumor microenvironment and reduced recruitment of fibroblasts and immune cells, which reflect morphological features of IDCP, may influence the aggressiveness of high-grade prostate cancer.

The cyto-linker and scaffolding protein "plectin" mis-localization leads to softening of cancer cells.

Discovering tools to prevent cancer progression requires understanding the fundamental differences between normal and cancer cells. More than a decade ago, atomic force microscopy (AFM) revealed cancer cells' softer body compared to their healthy counterparts. Here, we investigated the mechanism underlying the softening of cancerous cells in comparison with their healthy counterparts based on AFM high resolution stiffness tomography and 3D confocal microscopy. We showed microtubules (MTs) network in invasive ductal carcinoma cell cytoskeleton is basally located and segmented for around 400 nm from the cell periphery. Additionally, the cytoskeleton scaffolding protein plectin exhibits a mis-localization from the cytoplasm to the surface of cells in the carcinoma which justifies the dissociation of the MT network from the cell's cortex. Furthermore, the assessment of MTs' persistence length using a worm-like-chain (WLC) model in high resolution AFM images showed lower persistence length of the single MTs in ductal carcinoma compared to that in the normal state. Overall, these tuned mechanics support the invasive cells to ascertain more flexibility under compressive forces in small deformations. These data provide new insights into the structural origins of cancer aids in progression.

TCGA RNA-Seq and Tumor-Infiltrating Lymphocyte Imaging Data Reveal Cold Tumor Signatures of Invasive Ductal Carcinomas and Estrogen Receptor-Positive Human Breast Tumors.

Comparative studies of immune-active hot and immune-deserted cold tumors are critical for identifying therapeutic targets and strategies to improve immunotherapy outcomes in cancer patients. Tumors with high tumor-infiltrating lymphocytes (TILs) are likely to respond to immunotherapy. We used the human breast cancer RNA-seq data from the cancer genome atlas (TCGA) and classified them into hot and cold tumors based on their lymphocyte infiltration scores. We compared the immune profiles of hot and cold tumors, their corresponding normal tissue adjacent to the tumor (NAT), and normal breast tissues from healthy individuals from the Genotype-Tissue Expression (GTEx) database. Cold tumors showed a significantly lower effector T cells, lower levels of antigen presentation, higher pro-tumorigenic M2 macrophages, and higher expression of extracellular matrix (ECM) stiffness-associated genes. Hot/cold dichotomy was further tested using TIL maps and H&E whole-slide pathology images from the cancer imaging archive (TCIA). Analysis of both datasets revealed that infiltrating ductal carcinoma and estrogen receptor ER-positive tumors were significantly associated with cold features. However, only TIL map analysis indicated lobular carcinomas as cold tumors and triple-negative breast cancers (TNBC) as hot tumors. Thus, RNA-seq data may be clinically relevant to tumor immune signatures when the results are supported by pathological evidence.

Effect of HER2-targeted therapy on PDX and PDX-derived organoids generated from HER2-positive salivary duct carcinoma.

BACKGROUND: We previously established a patient-derived xenograft (PDX) model, patient-derived organoids (PDOs), and PDX-derived organoids (PDXOs) for salivary duct carcinoma (SDC). Using these models, this study examined the therapeutic effect of human epidermal growth factor receptor 2 (HER2) blockade on HER2-positive SDC. METHODS: The therapeutic effect of lapatinib was assessed in SDC PDXOs with regards to cell growth, receptor/downstream signaling molecule expression, phosphorylation levels, and apoptosis. Effect of lapatinib treatment was evaluated in vivo in SDC PDX mice. RESULTS: The siRNA knockdown of HER2 and lapatinib suppressed cell proliferation in SDC PDXOs. Lapatinib inhibited the phosphorylation of HER2 and its downstream targets, and induced apoptosis in SDC PDXOs. Lapatinib also significantly reduced tumor volumes compared with that of the control in SDC PDX mice. CONCLUSION: For the first time, we demonstrated the efficacy of anti-HER2 therapy in HER2-positive SDC using preclinical models of SDC PDX and PDXO.

High-Throughput Single-Cell Mass Spectrometry Reveals Abnormal Lipid Metabolism in Pancreatic Ductal Adenocarcinoma.

Even populations of clonal cells are heterogeneous, which requires high-throughput analysis methods with single-cell sensitivity. Here, we propose a rapid, label-free single-cell analytical method based on active capillary dielectric barrier discharge ionization mass spectrometry, which can analyze multiple metabolites in single cells at a rate of 38 cells/minute. Multiple cell types (HEK-293T, PANC-1, CFPAC-1, H6c7, HeLa and iBAs) were discriminated successfully. We found evidence for abnormal lipid metabolism in pancreatic cancer cells. We also analyzed gene expression in a cancer genome atlas dataset and found that the mRNA level of a critical enzyme of lipid synthesis (ATP citrate lyase, ACLY) was upregulated in human pancreatic ductal adenocarcinoma (PDAC). Moreover, both an ACLY chemical inhibitor and a siRNA approach targeting ACLY could suppress the viability of PDAC cells. A significant reduction in lipid content in treated cells indicates that ACLY could be a potential target for treating pancreatic cancer.

Altered expression of ERK, Cytochrome-c, and HSP70 triggers apoptosis in Quinacrine-exposed human invasive ductal carcinoma cells.

Invasive ductal carcinoma (IDC) is the most recurrent cancer, accounting for 80% of all breast cancers worldwide. Originating from the milk duct, it eventually invades the fibrous tissue of the breast outside the duct, proliferation takes 1-2 months for each division. Quinacrine (QC), an FDA-approved small molecule, has been shown to have anti-cancer activity in numerous cancerous cell lines through diverse pathways; ultimately leading to cell death. Here, we have investigated the mode of action of QC in MCF7 cells. This study demonstrated the modulation of cellular cytoskeleton, such as the formation of distinct filopodial and lamellipodial structures and spikes, through the regulation of small-GTPases. We also observed that QC induces a signaling cascade by inducing apoptotic cell death by increasing ROS generation and altering HSP70 expression; which presumably involves ERK regulation. Our findings show that QC could be an attractive chemotherapeutic agent having a "shotgun" nature with potential of inducing different signaling pathways leading to apoptotic cell death. This opens new avenues for research on developing QC as an effective therapeutic agent for the treatment of invasive ductal carcinomas.

PAX-8 Expression in Salivary Duct Carcinoma.

Salivary duct carcinoma (SDC) is a high-grade adenocarcinoma resembling breast ductal carcinoma. It accounts for ~10% of malignant tumors of the salivary glands. Most cases show expression of CK7 and androgen receptor. PAX-8 is a transcription factor, with expression reported in renal, Müllerian, and thyroid carcinomas. Previous studies have described an absence of PAX-8 immunostaining in most primary salivary gland neoplasms, including SDCs. However, PAX-8 expression is frequently found in neoplasms that can metastasize to salivary glands, suggesting the possibility that this protein can be used to differentiate SDC from secondary neoplastic involvement of the salivary gland. We evaluated the expression of PAX-8 in 14 cases of SDC from our institution. One case showed diffuse moderate to strong PAX-8 positivity, while 2 tumors showed focal weak staining. Therefore, we conclude that although the majority of SDC are negative for PAX-8, rare diffuse positivity can be seen in these primary salivary gland tumors. This could potentially pose difficulty in ruling out metastatic disease from another PAX-8-positive primary neoplasm.

A Mouse Model of Cholangiocarcinoma Uncovers a Role for Tensin-4 in Tumor Progression.

BACKGROUND AND AIMS: Earlier diagnosis and treatment of intrahepatic cholangiocarcinoma (iCCA) are necessary to improve therapy, yet limited information is available about initiation and evolution of iCCA precursor lesions. Therefore, there is a need to identify mechanisms driving formation of precancerous lesions and their progression toward invasive tumors using experimental models that faithfully recapitulate human tumorigenesis. APPROACH AND RESULTS: To this end, we generated a mouse model which combines cholangiocyte-specific expression of KrasG12D with 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) diet-induced inflammation to mimic iCCA development in patients with cholangitis. Histological and transcriptomic analyses of the mouse precursor lesions and iCCA were performed and compared with human analyses. The function of genes overexpressed during tumorigenesis was investigated in human cell lines. We found that mice expressing KrasG12D in cholangiocytes and fed a DDC diet developed cholangitis, ductular proliferations, intraductal papillary neoplasms of bile ducts (IPNBs), and, eventually, iCCAs. The histology of mouse and human IPNBs was similar, and mouse iCCAs displayed histological characteristics of human mucin-producing, large-duct-type iCCA. Signaling pathways activated in human iCCA were also activated in mice. The identification of transition zones between IPNB and iCCA on tissue sections, combined with RNA-sequencing analyses of the lesions supported that iCCAs derive from IPNBs. We further provide evidence that tensin-4 (TNS4), which is stimulated by KRASG12D and SRY-related HMG box transcription factor 17, promotes tumor progression. CONCLUSIONS: We developed a mouse model that faithfully recapitulates human iCCA tumorigenesis and identified a gene cascade which involves TNS4 and promotes tumor progression.

Prediction of tumor size in patients with invasive ductal carcinoma using FT-IR spectroscopy combined with chemometrics: a preliminary study.

Precise detection of tumor size is essential for early diagnosis, treatment, and evaluation of the prognosis of breast cancer. However, there are some errors between the tumor size of breast cancer measured by conventional imaging methods and the pathological tumor size. Invasive ductal carcinoma (IDC) is a common pathological type of breast cancer. In this study, serum Fourier transform infrared spectroscopy (FT-IR) combined with chemometric methods was used to predict the maximum diameter and maximum vertical diameter of tumors in IDC patients. Three models were evaluated based on the pathological tumor size measured after surgery and included grid search support vector machine regression (GS-SVR), back propagation neural network optimized by genetic algorithm (GA-BP-ANN), and back propagation neural network optimized by particle swarm optimization (PSO-BP-ANN). The results show that three models can accurately predict tumor size. The GA-BP-ANN model provided the best fitting quality of the largest tumor diameter with the determination coefficients of 0.984 in test set. And the GS-SVR model provided the best fitting quality of the largest vertical tumor diameter with the determination coefficients of 0.982 in test set. The GS-SVR model had the highest prediction efficiency and the lowest time complexity of the models. The results indicate that serum FT-IR spectroscopy combined with chemometric methods can predict tumor size in IDC patients. In addition, compared with traditional imaging methods, we found that the experimental results of the three models are better than traditional imaging methods in terms of correlation and fitting degree. And the average fitting error of PSO-BP-ANN and GA-BP-ANN models was less than 0.3 mm. The minimally invasive detection method is expected to be developed into a new clinical diagnostic method for tumor size estimation to reduce the diagnostic trauma of patients and provide new diagnostic experience for patients. Graphical Abstract.

The significance of tyrosine kinase receptor B and brain-derived neurotrophic factor expression in salivary duct carcinoma.

Salivary duct carcinoma (SDC) is a high-grade salivary gland neoplasm. It may occur de novo or secondarily from pleomorphic adenoma (ex-PA), with secondary development accounting for more than 50% of the cases. In recent years, the expression of tyrosine kinase receptor B (TrkB), which is in the same family as HER2, has been confirmed in various types of carcinomas. However, there are a few studies on SDC. In order to examine the expression and role of TrkB in SDC, we investigated it. Immunohistochemistry was used to detect the expression of TrkB and its ligands, brain-derived neurotrophic factor (BDNF) and neurotrophin-4 (NT-4) in 20 patients with SDC. The mRNA levels of TrkB, BDNF, and NT-4 were analyzed using quantitative polymerase chain reaction. TrkB was negative in 10 cases and positive in 10 cases, BDNF was negative in 11 cases and positive in 9 cases, and NT-4 was positive in all cases. There was a high number of TrkB-positive cases in the pT4 group and The H-score of TrkB was also significantly higher in the stage III and IV groups. There was a high number of BDNF-positive cases in the ex-PA group and Histo-score of BDNF had a trend of high expression in ex-PA. There were no significant differences or correlations in mRNA expression. Our results suggest that TrkB may be involved in SDC tumor growth.

Primary De novo ductal adenocarcinoma of the lacrimal gland.

BACKGROUND: Primary ductal adenocarcinoma of the lacrimal gland is a rare and aggressive malignant epithelial lacrimal gland neoplasm, morphologically and phenotypically resembles salivary duct carcinoma, and both strongly resemble infiltrating ductal carcinoma of breast. METHOD: Retrospective Chart review of cases of malignant lacrimal gland tumors from 2013 July to 2020 July. Authors describe the clinico radiological, morphological and immunohistochemical features of primary ductal adenocarcinoma (PDA) of lacrimal gland. Extensive review of literature of PDA of lacrimal gland and salivary gland ductal carcinoma has been performed. RESULTS: Retrospective chart review of the last 7 years yielded 22 malignant lacrimal gland neoplasms of which 4 cases demonstrated features of primary ductal adenocarcinoma of lacrimal gland, 2/4 cases showed an evidence of a pre existing pleomorphic adenoma and 2 were found to be de novo ductal adenocarcinomas. PDA of lacrimal gland showed expression of CK7, CK19, AR, HER2, cyclin D1 and were negative for CK5/14, CK 20, ER, PR, PSA, TTF-1, S-100 and SMA. Expression of GCDFP-15 was noted in one case. The presence of multiple events of loco-regional recurrences and/or distant metastasis necessitated a multidisciplinary approach. CONCLUSIONS: Authors have expressed the need of clinical correlation; thorough tissue sampling and extensive immunohistochemical work up in identification of de novo PDA's and their molecular subtypes. A multi-institutional study might help in formulating the diagnostic criteria, identification of actionable targets, and thus study the role of targeted therapy in this rare and aggressive tumor which may result in better patient outcomes.

Systemic therapy in the management of recurrent or metastatic salivary duct carcinoma: A systematic review.

BACKGROUND: Salivary duct carcinoma (SDC) is an aggressive subtype of salivary gland cancer. Approximately half of SDC patients will develop recurrences or metastases. Therapeutic palliative therapy is therefore often needed. The majority of SDC tumors expresses the androgen receptor (AR) and one-third expresses human epidermal growth factor receptor 2 (HER2), both are potential therapeutic targets. The aim of this paper is to systematically review and summarize the evidence on systemic palliative therapy for SDC and to provide treatment recommendations. MATERIALS AND METHODS: Electronic libraries were systematically searched with a broad search strategy to identify studies where SDC patients received systemic therapy. Due to the rarity of SDC no restrictions were placed on study designs. RESULTS: The search resulted in 2014 articles of which 153 were full-text analyzed. Forty-five studies were included in the analysis, which included in total 256 SDC patients receiving systemic therapy. Two phase 2 trials primarily including SDC patients were identified. The majority of the studies were case series or case reports, resulting in an overall low quality of available evidence. Based on studies including ≥ 5 SDC patients, objective responses to HER2 targeting agents were observed in 60-70%, to AR pathway agents in 18-53% and to chemotherapy in 10-50%. CONCLUSION: For AR or HER2 positive SDC, agents targeting these pathways are the cornerstone for palliative treatment. Regarding chemotherapy, the combination of carboplatin combined with a taxane is best studied. Regarding other targeted agents and immunotherapy evidence is anecdotal, limiting formulation of treatment recommendations for these antineoplastic agents.

Histologic diagnosis of a case of anal duct carcinoma with cytological correlation and differential diagnoses.

Anal duct carcinoma is an uncommon malignancy of the glands of the anal duct. This entity poses a diagnostic challenge, both clinically and histologically. This article describes histopathologic findings in a case of anal duct carcinoma, including the initial diagnosis on biopsy and subsequent cytology specimens. Additionally, differential diagnoses of this neoplasm are discussed. With a high index of suspicion, and attention to histological and immunohistochemical features, anal duct carcinoma can be accurately diagnosed both on biopsy and on cytology.

Multiplatform Investigation of Plasma and Tissue Lipid Signatures of Breast Cancer Using Mass Spectrometry Tools.

Plasma and tissue from breast cancer patients are valuable for diagnostic/prognostic purposes and are accessible by multiple mass spectrometry (MS) tools. Liquid chromatography-mass spectrometry (LC-MS) and ambient mass spectrometry imaging (MSI) were shown to be robust and reproducible technologies for breast cancer diagnosis. Here, we investigated whether there is a correspondence between lipid cancer features observed by desorption electrospray ionization (DESI)-MSI in tissue and those detected by LC-MS in plasma samples. The study included 28 tissues and 20 plasma samples from 24 women with ductal breast carcinomas of both special and no special type (NST) along with 22 plasma samples from healthy women. The comparison of plasma and tissue lipid signatures revealed that each one of the studied matrices (i.e., blood or tumor) has its own specific molecular signature and the full interposition of their discriminant ions is not possible. This comparison also revealed that the molecular indicators of tissue injury, characteristic of the breast cancer tissue profile obtained by DESI-MSI, do not persist as cancer discriminators in peripheral blood even though some of them could be found in plasma samples.

68Ga-PSMA-HBED-CC PET/CT imaging for adenoid cystic carcinoma and salivary duct carcinoma: a phase 2 imaging study.

Rationale: Treatment options for recurrent and/or metastatic (R/M) adenoid cystic carcinoma (ACC) and salivary duct carcinoma (SDC), major subtypes of salivary gland cancer, are limited. Both tumors often show overexpression of prostate-specific membrane antigen (PSMA). In prostate cancer, PSMA-ligands labeled with 68Ga or 177Lu are used for imaging and therapy, respectively. Primary aim of this study in R/M ACC and SDC patients was to systematically investigate 68Ga-PSMA-uptake by PET/CT imaging to determine if PSMA radionuclide therapy could be a treatment option. Methods: In a prospective phase II study, PET/CT imaging was performed 1 h post injection of 68Ga-PSMA-HBED-CC in 15 ACC patients and 10 SDC patients. Maximum standardized uptake values (SUV) were determined in tumor lesions. Immunohistochemical PSMA expression was scored in primary tumors and metastatic tissue. Standard imaging (MRI or CT) was performed for comparison. Results: In ACC patients, SUVmax ranged from 1.1 to 30.2 with a tumor/liver-ratio >1 in 13 out of 14 evaluable patients (93%). In SDC patients, SUVmax ranged from 0.3 to 25.9 with a tumor/liver-ratio >1 in 4 out of 10 patients (40%). We found a large intra-patient inter-metastatic variation in uptake of 68Ga-PSMA, and immunohistochemistry did not predict ligand uptake in ACC and SDC. Finally, PSMA-PET detected additional bone metastases compared to CT in 2 ACC patients with unexplained pain. Conclusion: In 93% of ACC patients and 40% of SDC patients we detected relevant PSMA-ligand uptake, which warrants to study PSMA radionuclide therapy in these patients. Additionally, our data provide arguments for patient selection and treatment timing. Finally, PSMA-PET imaging has added diagnostic value compared to CT in selected patients.

Comparative proteomic analysis of different stages of breast cancer tissues using ultra high performance liquid chromatography tandem mass spectrometer.

BACKGROUND: Breast cancer is the fifth most prevalent cause of death among women worldwide. It is also one of the most common types of cancer among Malaysian women. This study aimed to characterize and differentiate the proteomics profiles of different stages of breast cancer and its matched adjacent normal tissues in Malaysian breast cancer patients. Also, this study aimed to construct a pertinent protein pathway involved in each stage of cancer. METHODS: In total, 80 samples of tumor and matched adjacent normal tissues were collected from breast cancer patients at Seberang Jaya Hospital (SJH) and Kepala Batas Hospital (KBH), both in Penang, Malaysia. The protein expression profiles of breast cancer and normal tissues were mapped by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The Gel-Eluted Liquid Fractionation Entrapment Electrophoresis (GELFREE) Technology System was used for the separation and fractionation of extracted proteins, which also were analyzed to maximize protein detection. The protein fractions were then analyzed by tandem mass spectrometry (LC-MS/MS) analysis using LC/MS LTQ-Orbitrap Fusion and Elite. This study identified the proteins contained within the tissue samples using de novo sequencing and database matching via PEAKS software. We performed two different pathway analyses, DAVID and STRING, in the sets of proteins from stage 2 and stage 3 breast cancer samples. The lists of molecules were generated by the REACTOME-FI plugin, part of the CYTOSCAPE tool, and linker nodes were added in order to generate a connected network. Then, pathway enrichment was obtained, and a graphical model was created to depict the participation of the input proteins as well as the linker nodes. RESULTS: This study identified 12 proteins that were detected in stage 2 tumor tissues, and 17 proteins that were detected in stage 3 tumor tissues, related to their normal counterparts. It also identified some proteins that were present in stage 2 but not stage 3 and vice versa. Based on these results, this study clarified unique proteins pathways involved in carcinogenesis within stage 2 and stage 3 breast cancers. CONCLUSIONS: This study provided some useful insights about the proteins associated with breast cancer carcinogenesis and could establish an important foundation for future cancer-related discoveries using differential proteomics profiling. Beyond protein identification, this study considered the interaction, function, network, signaling pathway, and protein pathway involved in each profile. These results suggest that knowledge of protein expression, especially in stage 2 and stage 3 breast cancer, can provide important clues that may enable the discovery of novel biomarkers in carcinogenesis.

Aurora B induces epithelial-mesenchymal transition by stabilizing Snail1 to promote basal-like breast cancer metastasis.

Aurora B is a serine/threonine kinase that has been implicated in regulating cell proliferation in distinct cancers, including breast cancer. Here we show that Aurora B expression is elevated in basal-like breast cancer (BLBC) compared with other breast cancer subtypes. This high level of expression seems to correlate with poor metastasis-free survival and relapse-free survival in affected patients. Mechanistically, we show that elevated Aurora B expression in breast cancer cells activates AKT/GSK3ß to stabilize Snail1 protein, a master regulator of epithelial-mesenchymal transition (EMT), leading to EMT induction in a kinase-dependent manner. Conversely, Aurora B knock down by short-hairpin RNAs (shRNAs) suppresses AKT/GSK3ß/Snail1 signaling, reverses EMT and reduces breast cancer metastatic potential in vitro and in vivo. Finally, we identified a specific OCT4 phosphorylation site (T343) responsible for mediating Aurora B-induced AKT/GSK3ß/Snail1 signaling and EMT that could be attenuated by Aurora B kinase inhibitor treatment. These findings support that Aurora B induces EMT to promote breast cancer metastasis via OCT4/AKT/GSK3ß/Snail1 signaling. Pharmacologic Aurora B inhibition might be a potential effective treatment for breast cancer patients with metastatic disease.

Estrogen Receptor-α Quantification in Breast Cancer: Concordance Between Immunohistochemical Assays and mRNA-In Situ Hybridization for ESR1 Gene.

Immunohistochemical (IHC) quantification of estrogen receptor-α (ER) is used for assessment of treatment regimen in breast cancer. Different ER IHC assays may produce diverging results, because of different antibody clones, protocols, and stainer platforms. Objective tissue-based techniques to assess sensitivity and specificity of IHC assays are therefore needed. We tested the usability of ER mRNA-in situ hybridization (mRNA-ISH) in comparison with assays based on clones SP1 and 6F11. We selected 56 archival specimens according to their reported ER IHC positivity, representing a wide spectrum from negative to strongly positive cases. The specimens were used to prepare 4 TMAs with 112 cores. Serial sections of each TMA were stained for ER and pan-cytokeratin (PCK) by IHC and ESR1 (ER gene) by mRNA-ISH. Digital image analysis (DIA) was used to determine ER IHC H-score. ESR1 mRNA-ISH was scored both manually and by DIA. DIA showed a nonlinear correlation between IHC and ESR1 mRNA-ISH with R-values of 0.80 and 0.78 for the ER antibody clones SP1 and 6F11, respectively. Comparison of manual mRNA-ISH scoring categories and SP1 and 6F11 IHC H-scores showed a highly significant relationship (P<0.001). In conclusion, the study showed good correlation between mRNA-ISH and IHC, suggesting that mRNA-ISH can be a valuable tool in the assessment of the sensitivity and specificity of ER IHC assays.

Reciprocal and Autonomous Glucocorticoid and Androgen Receptor Activation in Salivary Duct Carcinoma.

PURPOSE: To determine the expression of glucocorticoid receptor (GR) and androgen receptor (AR) in salivary duct carcinoma (SDC) and to analyze the role of these proteins in the development and management of this disease entity. EXPERIMENTAL DESIGN: We performed a phenotypic assessment of GR and AR localization and expression, and determined their association with clinicopathologic factors in 67 primary SDCs. In vitro functional and response analysis of SDC cell lines was also performed. RESULTS: Of the 67 primary tumors, 12 (18%) overexpressed GR protein, 30 (45%) had constitutive expression, and 25 (37%) had complete loss of expression. Reciprocal GR and AR expression was found in 32 (48%) tumors, concurrent constitutive GR and AR expression in 23 (34%), and simultaneous loss of both receptors and high GR with AR expressions were found in 12 (18%). GR overexpression was significantly associated with worse clinical outcomes. In vitro ligand-independent AR activation was observed in both male- and female-derived cell lines. GR antagonist treatment resulted in decreased cell proliferation and survival in GR-overexpressing cells, irrespective of AR status. Reciprocal GR- and AR-knockdown experiments revealed an independent interaction. CONCLUSIONS: Our study, for the first time, demonstrates differential GR and AR expressions, autonomous GR and AR activation, and ligand-independent AR expression and activation in SDC cells. The findings provide critical information on the roles of GR and AR steroid receptors in SDC tumorigenesis and development of biomarkers to guide targeted steroid receptor therapy trials in patients with these tumors.

The contribution of dietary and plasma folate and cobalamin to levels of angiopoietin-1, angiopoietin-2 and Tie-2 receptors depend on vascular endothelial growth factor status of primary breast cancer patients.

The aim of this study was to determine the association of dietary folate and cobalamin with plasma levels of Angiopoietins (ANG), vascular endothelial growth factor-C (VEGF-C) and tyrosine kinase receptor-2 (Tie-2) of primary breast cancer patients. Women (n = 177), aged 30 to 75 years diagnosed with breast cancer were recruited from an ongoing case series study. Dietary intake of nutrients was estimated by using a validated food frequency questionnaire. Enzyme-linked immunosorbent assay was applied to measure biomarkers. MCF-7 cell cultures were supplemented with folic acid (0-40 µM) for 24 h to measure cell viability and fold change of expression by the real-time reverse transcriptase-polymerase chain reaction. Structural equation modeling was applied to analyze the structural relationships between the measured variables of nutrients and Angiopoietins. Dietary intake of folate and cobalamin showed a significant inverse correlation with plasma ANG-1 and ANG-2 (P < 0.05), particularly in subjects with estrogen-receptor positive tumors or low plasma VEGF-C. Plasma folate was positively associated with the ratio of ANG-1/ANG-2 (P < 0.05). Residual intake levels of total cobalamin were inversely associated with plasma ANG-1 when plasma stratum of VEGF-C was high (P < 0.05). Structural equation modeling identified a significant inverse contribution of folate profiles on the latent variable of Angiopoietins (coefficient ß = -0.99, P < 0.05). Folic acid treatment resulted in dose-dependent down-regulations on ANGPT1 and ANGPT1/ANGPT2 ratio but VEGF and ANGPT2/VEGF were upregulated at folic acid >20 µM. Studying the contributing role of dietary folate to pro-angiogenic biomarkers in breast cancer patients can infer the preventive role of folate in the ANGs/VEGF-C-dependent cascade of tumor metastasis. By contrast, high concentrations of folic acid in vitro supported VEGF-C-dependent ANGPT2 overexpression might potentiate micro-lymphatic vessel development to support malignant cell dissemination.