Phenotypic evaluation of the basal-like subtype of invasive breast carcinoma

Microarray profiling of invasive breast carcinomas has identified five distinct subtypes of tumors (luminal A, luminal B, normal breast-like, HER2 overexpressing, and basal-like) that are associated with different clinical outcomes. The basal-like subtype is associated with poor clinical outcomes and is the subtype observed in BRCA1-related breast cancers. The aim of this study was to characterize the histologic and immunophenotypic properties of breast basal-like carcinomas that were first positively identified using DNA microarray analysis. Detailed histologic review was performed on 56 tumors with known microarray profiles (23 basal-like, 23 luminal, and 12 HER2+). Immunohistochemistry for estrogen receptor (ER), HER2, EGFR, smooth muscle actin (SMA), p63, CD10, cytokeratin 5/6, cytokeratin 8/18, and vimentin was performed on 18 basal-like, 16 luminal, and 12 HER2+ tumors. The basal-like tumors were grade 3 ductal/NOS (21/23) or metaplastic (2/23) carcinomas that frequently showed geographic necrosis (17/23), a pushing border of invasion (14/23), and a stromal lymphocytic response (13/23). Most basal-like tumors showed immunoreactivity for vimentin (17/18), luminal cytokeratin 8/18 (15/18), EGFR (13/18), and cytokeratin 5/6 (11/18), while positivity for the myoepithelial markers SMA (4/18), p63 (4/18) and CD10 (2/18) was infrequent. All basal-like tumors tested were ER− and HER2−. Morphologic features significantly associated with the basal-like subtype included markedly elevated mitotic count (P less than 0.0001), geographic tumor necrosis (P=0.0003), pushing margin of invasion (P=0.0001), and stromal lymphocytic response (P=0.01). The most consistent immunophenotype seen in the basal-like tumors was negativity for ER and HER2, and positivity for vimentin, EGFR, cytokeratin 8/18, and cytokeratin 5/6. The infrequent expression of myoepithelial markers in basal-like carcinomas does not support a direct myoepithelial cell derivation of these tumors. These findings should further assist in the identification of basal-like carcinomas in clinical specimens, facilitating treatment and epidemiologic studies of this tumor subtype

Clear Cell Renal Cell Carcinoma With a Poorly-Differentiated Component: A Novel Variant Causing Potential Diagnostic Difficulty

Background: Several variant histologic patterns of clear cell renal cell carcinoma (RCC) are well known, especially those with sarcomatoid and rhabdoid features. However, we have encountered rare cases in which a high-grade adenocarcinoma or urothelial carcinoma-like component would be difficult to appreciate as clear cell RCC. DesignWe retrieved 26 tumors with histologically typical clear cell RCC juxtaposed to a high-grade non-clear cell component.High grade non-clear cell component was defined as non-sarcomatoid, non-rhabdoid areas that would be difficult to assign as renal cell in origin if viewed in isolation. Tumors were studied with immunohistochemistry and fluorescence in situ hybridization (FISH) or sequencing.ResultsMedian percentage of poorly differentiated component: 50%(IQR20-70). All tumors showed abrupt transition from clear cell carcinoma to poorly-differentiated (non-sarcomatoid/non-rhabdoid) areas, which showed micropapillary (7/26; 27%), urothelial-like (10/26; 39%), and adenocarcinoma NOS features (9/26; 35%). 19 tumors had necrosis. Carbonic anhydrase IX (CA-IX) was uniformly positive in well-differentiated component (20/20); poorly differentiated component showed a median positivity of 82.5% (IQR 65-100). Poorly differentiated component was positive for CK7 (5/19; 26%), CK20 (3/12; 25%), AMACR (7/12; 58%), PAX8 (12/15; 80%), and showed intact FH (6/6; 100%). CDX2 was uniformly negative. Chromosome 3p loss or VHL mutation was present in 8/13 (62%), tested with either FISH (n = 9) or sequencing (n = 4). All tested cases were negative for TFE3 (0/11) and TFEB (0/9) rearrangements on FISH. 5/21 (24%) patients were alive with metastatic disease and 5/21 (24%) had died of disease on follow up. One metastasis was composed only of the poorly-differentiated component and was near-negative for CA-IX. Conclusion: Clear cell RCC with a poorly differentiated component resembling adenocarcinoma or urothelial carcinoma is a novel source of morphologic heterogeneity that has not been previously well characterized. Potential pitfalls include decreased or absent CA-IX staining the high-grade component and aberrant positivity for cytokeratin 7 or 20. With the increasing use of renal mass biopsy and biopsies of metastatic sites for targeted therapy, pathologists should be aware of this entity and consider the possibility of clear cell RCC even for morphologically unusual tumors.https://scholarlycommons.henryford.com/merf2019caserpt/1069/thumbnail.jp

Genomic analysis of estrogen cascade reveals histone variant H2A.Z associated with breast cancer progression

We demonstrate an integrated approach to the study of a transcriptional regulatory cascade involved in the progression of breast cancer and we identify a protein associated with disease progression. Using chromatin immunoprecipitation and genome tiling arrays, whole genome mapping of transcription factor-binding sites was combined with gene expression profiling to identify genes involved in the proliferative response to estrogen (E2). Using RNA interference, selected ERα and c-MYC gene targets were knocked down to identify mediators of E2-stimulated cell proliferation. Tissue microarray screening revealed that high expression of an epigenetic factor, the E2-inducible histone variant H2A.Z, is significantly associated with lymph node metastasis and decreased breast cancer survival. Detection of H2A.Z levels independently increased the prognostic power of biomarkers currently in clinical use. This integrated approach has accelerated the identification of a molecule linked to breast cancer progression, has implications for diagnostic and therapeutic interventions, and can be applied to a wide range of cancers

The EphB4 Receptor Tyrosine Kinase Promotes Lung Cancer Growth: A Potential Novel Therapeutic Target

Despite progress in locoregional and systemic therapies, patient survival from lung cancer remains a challenge. Receptor tyrosine kinases are frequently implicated in lung cancer pathogenesis, and some tyrosine kinase inhibition strategies have been effective clinically. The EphB4 receptor tyrosine kinase has recently emerged as a potential target in several other cancers. We sought to systematically study the role of EphB4 in lung cancer. Here, we demonstrate that EphB4 is overexpressed 3-fold in lung tumors compared to paired normal tissues and frequently exhibits gene copy number increases in lung cancer. We also show that overexpression of EphB4 promotes cellular proliferation, colony formation, and motility, while EphB4 inhibition reduces cellular viability in vitro, halts the growth of established tumors in mouse xenograft models when used as a single-target strategy, and causes near-complete regression of established tumors when used in combination with paclitaxel. Taken together, these data suggest an important role for EphB4 as a potential novel therapeutic target in lung cancer. Clinical trials investigating the efficacy of anti-EphB4 therapies as well as combination therapy involving EphB4 inhibition may be warranted.</p

The molecular portraits of breast tumors are conserved acress microarray platforms

Background Validation of a novel gene expression signature in independent data sets is a critical step in the development of a clinically useful test for cancer patient risk-stratification. However, validation is often unconvincing because the size of the test set is typically small. To overcome this problem we used publicly available breast cancer gene expression data sets and a novel approach to data fusion, in order to validate a new breast tumor intrinsic list. Results A 105-tumor training set containing 26 sample pairs was used to derive a new breast tumor intrinsic gene list. This intrinsic list contained 1300 genes and a proliferation signature that was not present in previous breast intrinsic gene sets. We tested this list as a survival predictor on a data set of 311 tumors compiled from three independent microarray studies that were fused into a single data set using Distance Weighted Discrimination. When the new intrinsic gene set was used to hierarchically cluster this combined test set, tumors were grouped into LumA, LumB, Basal-like, HER2+/ER-, and Normal Breast-like tumor subtypes that we demonstrated in previous datasets. These subtypes were associated with significant differences in Relapse-Free and Overall Survival. Multivariate Cox analysis of the combined test set showed that the intrinsic subtype classifications added significant prognostic information that was independent of standard clinical predictors. From the combined test set, we developed an objective and unchanging classifier based upon five intrinsic subtype mean expression profiles (i.e. centroids), which is designed for single sample predictions (SSP). The SSP approach was applied to two additional independent data sets and consistently predicted survival in both systemically treated and untreated patient groups. Conclusion This study validates the breast tumor intrinsic subtype classification as an objective means of tumor classification that should be translated into a clinical assay for further retrospective and prospective validation. In addition, our method of combining existing data sets can be used to robustly validate the potential clinical value of any new gene expression profile

The molecular portraits of breast tumors are conserved across microarray platforms

BACKGROUND: Validation of a novel gene expression signature in independent data sets is a critical step in the development of a clinically useful test for cancer patient risk-stratification. However, validation is often unconvincing because the size of the test set is typically small. To overcome this problem we used publicly available breast cancer gene expression data sets and a novel approach to data fusion, in order to validate a new breast tumor intrinsic list. RESULTS: A 105-tumor training set containing 26 sample pairs was used to derive a new breast tumor intrinsic gene list. This intrinsic list contained 1300 genes and a proliferation signature that was not present in previous breast intrinsic gene sets. We tested this list as a survival predictor on a data set of 311 tumors compiled from three independent microarray studies that were fused into a single data set using Distance Weighted Discrimination. When the new intrinsic gene set was used to hierarchically cluster this combined test set, tumors were grouped into LumA, LumB, Basal-like, HER2+/ER-, and Normal Breast-like tumor subtypes that we demonstrated in previous datasets. These subtypes were associated with significant differences in Relapse-Free and Overall Survival. Multivariate Cox analysis of the combined test set showed that the intrinsic subtype classifications added significant prognostic information that was independent of standard clinical predictors. From the combined test set, we developed an objective and unchanging classifier based upon five intrinsic subtype mean expression profiles (i.e. centroids), which is designed for single sample predictions (SSP). The SSP approach was applied to two additional independent data sets and consistently predicted survival in both systemically treated and untreated patient groups. CONCLUSION: This study validates the "breast tumor intrinsic" subtype classification as an objective means of tumor classification that should be translated into a clinical assay for further retrospective and prospective validation. In addition, our method of combining existing data sets can be used to robustly validate the potential clinical value of any new gene expression profile

ERG finally has something to YAP about in prostate cancer

SummaryThe significance of ERG in human prostate cancer is unclear because mouse prostate is resistant to ERG-mediated transformation. We determined that ERG activates the transcriptional program regulated by YAP1 of the Hippo signaling pathway and found that prostate-specific activation of either ERG or YAP1 in mice induces similar transcriptional changes and results in age-related prostate tumors. ERG binds to chromatin regions occupied by TEAD/YAP1 and transactivates Hippo target genes. In addition, in human luminal-type prostate cancer cells, ERG binds to the promoter of YAP1 and is necessary for YAP1 expression. These results provide direct genetic evidence of a causal role for ERG in prostate cancer and reveal a connection between ERG and the Hippo signaling pathway

The growth pattern of transplanted normal and nodular hepatocytes

Overt neoplasia is often the end result of a long biological process beginning with the appearance of focal lesions of altered tissue morphology. While the putative clonal nature of focal lesions has often been emphasized, increasing attention is being devoted to the possible role of an altered growth pattern in the evolution of carcinogenesis. Here we compare the growth patterns of normal and nodular hepatocytes in a transplantation system that allows their selective clonal proliferation in vivo. Rats were pre-treated with retrorsine, which blocks the growth of resident hepatocytes, and were then transplanted with hepatocytes isolated from either normal liver or hepatocyte nodules. Both cell types were able to proliferate extensively in the recipient liver, as expected. However, their growth pattern was remarkably different. Clusters of normal hepatocytes integrated in the host liver, displaying a normal histology; however, transplanted nodular hepatocytes formed new hepatocyte nodules, with altered morphology and sharp demarcation from surrounding host liver. Both the expression and distribution of proteins involved in cell polarity, cell communication, and cell adhesion, including connexin 32, E-cadherin, and matrix metalloproteinase-2, were altered in clusters of nodular hepatocytes. Furthermore, we were able to show that down-regulation of connexin 32 and E-cadherin in nodular hepatocyte clusters was independent of growth rate. These results support the concept that a dominant pathway towards neoplastic disease in several organs involves defect(s) in tissue pattern formation

Cancer Biomarker Discovery: The Entropic Hallmark

Background: It is a commonly accepted belief that cancer cells modify their transcriptional state during the progression of the disease. We propose that the progression of cancer cells towards malignant phenotypes can be efficiently tracked using high-throughput technologies that follow the gradual changes observed in the gene expression profiles by employing Shannon's mathematical theory of communication. Methods based on Information Theory can then quantify the divergence of cancer cells' transcriptional profiles from those of normally appearing cells of the originating tissues. The relevance of the proposed methods can be evaluated using microarray datasets available in the public domain but the method is in principle applicable to other high-throughput methods. Methodology/Principal Findings: Using melanoma and prostate cancer datasets we illustrate how it is possible to employ Shannon Entropy and the Jensen-Shannon divergence to trace the transcriptional changes progression of the disease. We establish how the variations of these two measures correlate with established biomarkers of cancer progression. The Information Theory measures allow us to identify novel biomarkers for both progressive and relatively more sudden transcriptional changes leading to malignant phenotypes. At the same time, the methodology was able to validate a large number of genes and processes that seem to be implicated in the progression of melanoma and prostate cancer. Conclusions/Significance: We thus present a quantitative guiding rule, a new unifying hallmark of cancer: the cancer cell's transcriptome changes lead to measurable observed transitions of Normalized Shannon Entropy values (as measured by high-throughput technologies). At the same time, tumor cells increment their divergence from the normal tissue profile increasing their disorder via creation of states that we might not directly measure. This unifying hallmark allows, via the the Jensen-Shannon divergence, to identify the arrow of time of the processes from the gene expression profiles, and helps to map the phenotypical and molecular hallmarks of specific cancer subtypes. The deep mathematical basis of the approach allows us to suggest that this principle is, hopefully, of general applicability for other diseases

P504S immunostaining boosts diagnostic resolution of suspicious foci in prostatic needle biopsy specimens

From 1.5% to 9.0% of prostatic needle biopsy specimens disclose atypical small acinar proliferations (ASAPs) suggestive of malignancy, carrying an approximate 45% predictive value for cancer. We applied keratin 34 beta E12 and P504S monoclonal immunostains to 93 cases that were judged as ASAP after HandE staining alone. Forty-one ASAP foci survived recutting for both immunostains. Three urologic pathologists independently assigned post-keratin 34 beta E12 diagnoses of cancer, ASAP, high-grade prostatic intraepithelial neoplasia, or benign and then reviewed P504S slides and assigned final diagnoses. Eight foci (20%) were resolved unanimously after keratin 34 beta E12 staining; 18 (44%) were resolved by 1 or 2 evaluators and 29 (71%) by at least 1. According to whether post-keratin 34 beta E12 ASAP designation was given by 3, 2, or 1 evaluator(s), P504S immunostaining unanimously resolved an additional 5 (12%), 10 (24%), or 23 (56%) of 41 ASAP foci and cumulatively, 31 foci (76%). Among 35 men (excluding 6 with cancer in other cores of the original biopsy), these immunostains could have permitted cancer diagnosis in 11 (31%), without repeated biopsy. Thus, the consensus diagnosis rate improved from poor to good after supplementing 34 beta E12 immunostaining with P504S