Loss of PSP94 expression is associated with early PSA recurrence and deteriorates outcome of PTEN deleted prostate cancers.

OBJECTIVE: Prostate secretory protein of 94 amino acids (PSP94) is a target gene of the EZH2 transcriptional repressor and is often downregulated in prostate cancer; however, its prognostic value is disputed. METHODS: Immunohistochemical analysis of a tissue microarray of 12, 432 prostate cancer specimens was performed to evaluate PSP94 expression. Correlation of PSP94 expression with tumor phenotype, patient prognosis, TMPRSS2:ERG fusion status, EZH2 expression and PTEN deletion was studied. RESULTS: PSP94 expression was increased in benign prostatic hyperplasia; however, it was downregulated in 48% and negative in 42% of the 9, 881 interpretable prostate cancer specimens. The loss of PSP94 expression was inversely correlated to EZH2 expression (P < 0.0001) and largely unrelated to the ERG status, but strongly correlated with high Gleason grade, advanced tumor stage, and nodal metastasis ( P <0.0001 each). The fraction of PSP94-negative cancer specimens increased from 40% in pT2 to 52% in pT3b-pT4 ( P < 0.0001) and from 40% in Gleason 3+3 = 6 to 46% in Gleason 4+3 = 7 and 60% in Gleason ≥4+4 = 8 ( P < 0.0001). Loss of PSP94 was linked to early prostate-specific antigen recurrence, but with little absolute effect ( P < 0.0001). However, it provided additional prognostic impact in cancer specimens with PTEN deletion. Loss of PSP94 deteriorated prognosis of cancer patients with PTEN deletion by more than 10% (P < 0.0001). The combination of PTEN deletion and PSP94 loss provided independent prognostic information that was observed in several subgroups defined by classical and quantitative Gleason grade. CONCLUSIONS: The results of our study suggest that combined PSP94/PTEN analysis can be potentially used in the clinical prognosis of prostate cancer.

Nuclear ELAC2 overexpression is associated with increased hazard for relapse after radical prostatectomy.

ELAC2 is a ubiquitously expressed enzyme potentially involved in tRNA processing and cell signaling pathways. Mutations of the ELAC2 gene have been found to confer increased prostate cancer susceptibility in families. ELAC2 protein expression was analyzed by immunohistochemistry in 9,262 patients and Kaplan-Meier curves of PSA recurrence-free survival were calculated in 8,513 patients treated with radical prostatectomy. Nuclear ELAC2 staining was observed in 60.8% of prostate cancers. It was weak in 26.3%, moderate in 26.6% and strong in 7.9%. Strong nuclear ELAC2 expression was associated with advanced tumor stage, nodal metastasis, higher Gleason grade, presence of TMPRSS2:ERG fusion, higher Ki67-labeling index and PTEN deletion. The difference in 1-, 5- and 10-year recurrence-free survival between strong and weak nuclear ELAC2 intensity is 7.2/13.8/17.6% in all cancers, 7.4/16.1/26.5% in the ERG negative subset, and 3.1/5.7/9.8% in the ERG positive subset. Regarding the univariate hazard ratio, PSA recurrence-free survival after prostatectomy for strong nuclear ELAC2 expression is 1.89 (1.64-2.10, p < 0.0001). It is independent of preoperative PSA-level, Gleason grade, pathological stage, surgical margin stage, and lymph node stage (multivariate hazard ratio 1.29 (1.11-1.49, p = 0.001). We conclude that nuclear ELAC2 expression is an independent prognostic marker for PSA recurrence-free survival after radical prostatectomy with a weak to moderate increase of the hazard ratio for biochemical relapse.

The independent prognostic impact of the GATA2 pioneering factor is restricted to ERG-negative prostate cancer.

GATA2 is a pioneering transcription factor governing androgen receptor expression and signaling in prostate cells. To understand the prognostic potential of GATA2 assessment in prostate cancer, we analyzed nuclear GATA2 expression on an annotated tissue microarray with 12,427 prostate cancer samples. Normal prostate glands were negative to weakly positive. GATA2 staining was found in almost all prostate cancers (95%). Strong GATA2 staining was linked to advanced tumor stage, high classical and quantitative Gleason grade (p < 0.0001 each), positive nodal stage (p = 0.0116), and early biochemical recurrence (p < 0.0001). GATA2 was linked to ERG-fusion-type cancers, with strong GATA2 staining in 29% of ERG-negative and 53% of ERG-positive cancers (p < 0.0001). Separate calculations in 3854 cancers with and 4768 cancers without TMPRSS2:ERG fusion revealed that these associations with tumor phenotype and patient outcome were largely driven by the subset of ERG-negative tumors. GATA2 expression was further linked to androgen receptor expression: Only 8% of androgen receptor-negative, but 56% of strongly androgen receptor expressing cancers had strong GATA2 expression (p < 0.0001). In conclusion, the results of our study demonstrate that increasing GATA2 levels are linked to prostate cancer progression and aggressiveness. The prognostic value of GATA2 is remarkable in ERG-negative cancers. However, the upregulation of GATA2 in ERG-positive cancers makes it unsuitable as a prognostic marker in this patient subset.

SNW1 is a prognostic biomarker in prostate cancer.

BACKGROUND: SNW1 is a nuclear receptor co-activator involved in splicing and transcription control, including androgen receptor signaling. Overexpression of SNW1 has been linked to adverse prognosis in different cancer types, but studies on the role of SNW1 in prostate cancer are lacking. METHODS: Using immunohistochemistry, we analyzed SNW1 expression in 10,310 prostate cancers in a tissue microarray (TMA) with attached clinical and molecular data. RESULTS: The comparison with normal prostate tissue revealed an up regulation of SNW1 in a subset of cancer samples. SNW1 staining was considered weak in 31.5%, moderate in 37.7% and strong in 14% of cancers. Strong SNW1 expression was markedly more frequent in prostate cancers harboring the TMPRSS2:ERG fusion (24%) than in ERG negative cancers (7%, p < 0.0001). Significant associations with Gleason grade, stage, nodal status and early biochemical recurrence were observed in the ERG negative and positive subset. Multivariable modeling revealed that the prognostic value of SNW1 up regulation was independent from the established preoperative histopathological and clinical parameters. CONCLUSION: These results demonstrate that SNW1 overexpression is an independent prognostic marker in prostate cancer with potential clinical utility.

Aberrant expression of the microtubule-associated protein tau is an independent prognostic feature in prostate cancer.

BACKGROUND: Microtubule-associated protein Tau (MAPT) overexpression has been linked to poor prognosis and decreased response to taxane-based therapies in several cancer types, but its relevance in prostate cancer is unknown. METHODS: In this study, MAPT expression was analyzed by immunohistochemistry on a tissue microarray containing 17,747 prostate cancers. RESULTS: MAPT was absent in normal prostate epithelial cells but detectable in 1004 (8.2%) of 12,313 interpretable cancers. Its expression was associated with advanced tumor stage, high Gleason grade, positive lymph nodes, and early biochemical recurrence (p < 0.0001 each). For example, MAPT was found in 3.6% of 2072 Gleason ≤3 + 3 cancers but in 14.4% of 704 Gleason ≥4 + 4 cancers. High-level MAPT staining was also linked to TMPRSS2:ERG fusions (p < 0.0001). MAPT staining was seen in 15.2 and 16% of cancers with TMPRSS2:ERG fusion detected by immunohistochemistry and fluorescence in-situ hybridization, but in only 3.5 and 3.9% of cancers without ERG staining or ERG rearrangements. Moreover, an association was found between MAPT expression and PTEN deletions, with 19% MAPT positivity in 948 PTEN deleted cancers but only 7% MAPT positivity in 3895 tumors with normal PTEN copy numbers (p < 0.0001). Multivariate analysis revealed that the prognostic value of MAPT was independent from established parameters. Conventional large section analyses showed intratumoral MAPT heterogeneity in all three analyzed cancers. CONCLUSIONS: The results of our study identify MAPT, as a moderate prognostic marker in prostate cancer, whose clinical impact, however, may be limited due to the rarity and heterogeneity of its expression.

High expression of class III ß-tubulin in upper gastrointestinal cancer types.

Class III ß-tubulin (TUBB3) is a component of microtubules of neuronal cells that is upregulated in various cancer entities. To better understand the role of TUBB3 in upper gastrointestinal tract cancer types, the present study assessed TUBB3 expression in tissue microarrays including 189 gastric and 428 esophageal cancer. TUBB3 expression was detected in 62.4% of gastric cancer, 73.8% of esophageal adenocarcinoma and 88.7% of esophageal squamous cell cancer, while control samples of normal esophageal and gastric epithelium were TUBB3-negative. TUBB3 positivity was not associated with the International Union Against Cancer classification, World Health Organization grading, lymph node involvement or distant metastasis in any entity. Of note, TUBB3 expression was associated with tumor localization and prognosis in gastric cancer, with the tumor stage in esophageal adenocarcinoma, and with the resection margin in esophageal squamous cell cancer. In conclusion, the substantial rate of positivity for TUBB3 already in early stages of gastric cancer in combination with the lack of a further increase in frequency with tumor stage, may suggest, that TUBB3 upregulation is rather relevant for cancer development than for cancer progression. TUBB3 might be a suitable prognostic biomarker in gastric cancer types.

High concordance of TMPRSS-ERG fusion between primary prostate cancer and its lymph node metastases.

Approximately 50% of prostate cancer types harbor the transmembrane protease, serine 2: Erythroblast transformation-specific-related gene (ERG) fusion, resulting in oncogenic expression of the ERG transcription factor. ERG represents an attractive target for potential future anticancer therapy in advanced and metastatic prostate cancer. To better understand whether the analysis of the primary cancer is sufficient to estimate the ERG expression status of the lymph node metastases, the present study examined patterns of immunohistochemical ERG expression in a tissue microarray created from multiple primary and metastatic sites of 77 prostate cancer tissues. Among the identified tumor types, 80% were either entirely ERG-positive (38%) or ERG-negative (42%) across all (at least 9) analyzed different tumor sites. The results were heterogeneous in 20% of the tumor types and typically resulted from small ERG-negative areas within otherwise ERG-positive tumor types. Comparison of the ERG expression status in 51 primary cancer types with at least three interpretable lymph node metastases revealed an entirely identical ERG status in all tumor sites in 75% of the cases, including 16 ERG-positive and 22 ERG-negative cancer types. The remaining 13 cancer types exhibited ERG heterogeneity within the primary tumor, while all metastases had an identical (12 positive and 1 negative) ERG status. The results of the present study revealed a high degree of concordance of the ERG expression status between primary prostate cancer types and their lymph node metastases. Therefore, potential anti-ERG therapy may also be effective against lymph node metastases in the majority of cases of ERG-positive metastatic prostate cancer.

Author Correction: Up regulation and nuclear translocation of Y-box binding protein 1 (YB-1) is linked to poor prognosis in ERG-negative prostate cancer.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

High BCAR1 expression is associated with early PSA recurrence in ERG negative prostate cancer.

BACKGROUND: Breast cancer anti-estrogen resistance 1 (BCAR1/p130cas) is a hub for diverse oncogenic signaling cascades and promotes tumor development and progression. METHODS: To understand the effect of BCAR1 in prostate cancer, we analyzed its expression on more than 11,000 prostate cancer samples. BCAR1 expression levels were compared with clinical characteristics, PSA recurrence, molecular subtype defined by ERG status and 3p, 5q, 6q and PTEN deletion. RESULTS: BCAR1 staining was barely detectable in normal prostate glands but seen in 77.6% of 9472 interpretable cancers, including strong expression in 38.5%, moderate in 23.2% and weak in 15.9% of cases. BCAR1 up regulation was associated with positive ERG status (p < 0.0001), high Gleason score (p < 0.0001), advanced pathological tumor stage (p = 0.0082), lower preoperative PSA level (p < 0.0001), increased cell proliferation (p < 0.0001), early PSA recurrence (p = 0.0008), and predicted prognosis independently from clinico-pathological parameters available at the time of the initial biopsy. However, subset analyses revealed that the prognostic impact of BCAR1 expression was limited to ERG-negative cancer. That BCAR1 up regulation was linked to almost all analyzed deletions (p < 0.0001 each for PTEN, 5q, 6q deletion) may suggest a functional link to genomic instability. CONCLUSION: The results of our study identify BCAR1 as a prognostic biomarker with potential clinical value for risk stratification of ERG-negative prostate cancer.

Up-regulation of Biglycan is Associated with Poor Prognosis and PTEN Deletion in Patients with Prostate Cancer.

Biglycan (BGN), a proteoglycan of the extracellular matrix, is included in mRNA signatures for prostate cancer aggressiveness. To understand the impact of BGN on prognosis and its relationship to molecularly defined subsets, we analyzed BGN expression by immunohistochemistry on a tissue microarray containing 12,427 prostate cancers. Seventy-eight percent of 11,050 interpretable cancers showed BGN expression, which was considered as low intensity in 47.7% and as high intensity in 31.1% of cancers. BGN protein expression rose with increasing pathological tumor stage, Gleason grade, lymph node metastasis and early PSA recurrence (P<.0001 each). Comparison with our molecular database attached to the TMA revealed that BGN expression was linked to presence of TMPRRS2:ERG fusion and PTEN deletion (P<.0001 each). In addition, BGN was strongly linked to androgen-receptor (AR) levels (P<.0001), suggesting a hormone-depending regulation of BGN. BGN up-regulation is a frequent feature of prostate cancer that parallels tumor progression and may be useful to estimate tumor aggressiveness particularly if combined with other molecular markers.

Apurinic/apyrimidinic endonuclease 1 (APE1/Ref-1) overexpression is an independent prognostic marker in prostate cancer without TMPRSS2:ERG fusion.

Polymorphisms of the base excision repair gene APE1 may be associated with an increased risk for developing prostate cancer. In other cancer types, altered APE1 protein expression is a candidate prognostic marker. Using immunohistochemistry, we thus analyzed APE1 expression in 9763 prostate cancers in a tissue microarray (TMA) with attached clinical and molecular data. The comparison with normal prostate tissue revealed an upregulation of APE1 in cancer samples. APE1 immunostaining was considered weak in 20.2%, moderate in 36.7%, and strong in 33.4% of cancers. Strong APE1 expression was markedly more frequent in prostate cancers harboring the TMPRSS2:ERG fusion (52.9%) than in ERG-negative cancers (19.1%, P < 0.0001). Significant associations with Gleason grade, tumor stage, tumor grade, and early biochemical recurrence (P < 0.0001 each) were largely limited to ERG-negative tumors. Multivariable analysis revealed that the prognostic value of APE1 upregulation in ERG-negative prostate cancers was independent from established histopathological and clinical parameters. In conclusion, the results of our study demonstrate that APE1 overexpression is an independent prognostic marker, but exclusively in ERG-negative prostate cancers.

Up regulation and nuclear translocation of Y-box binding protein 1 (YB-1) is linked to poor prognosis in ERG-negative prostate cancer.

Y-box binding protein 1 (YB-1) is an RNA and DNA binding factor with potential prognostic cancer. To evaluate the clinical impact of YB-1, a tissue microarray with 11,152 prostate cancers was analysed by immunohistochemistry. Cytoplasmic and nuclear staining was separately analysed. Cytoplasmic YB-1 was absent or weak in normal epithelium but seen in 86,3% of carcinomas. Cytoplasmic staining was weak, moderate, and strong in 29.6%, 43.7% and 13.0% of tumours and was accompanied by nuclear YB-1 staining in 32.1% of cases. Particularly nuclear staining was strongly linked to poor patient prognosis (p < 0.0001). YB-1 protein was more abundant in ERG positive (95.1%) than in ERG negative cancers (80.4%; p < 0.0001), but any prognostic impact of YB-1 staining was limited to the ERG-negative subset. Similarly, significant associations with pT stage and Gleason grade (p < 0.0001 each) were driven by the ERG negative subset. The significant association of YB-1 protein detection with deletions of PTEN, 5q21 and 6q15 fits well in the protein's role as an inhibitor of DNA damage dependent cell cycle arrest, a role that is likely to induce genomic instability. In summary, the data show, that the prognostic impact of YB-1 expression is limited to ERG negative prostate cancers.

High-Level γ-Glutamyl-Hydrolase (GGH) Expression is Linked to Poor Prognosis in ERG Negative Prostate Cancer.

γ-glutamyl-hydrolase (GGH) is a ubiquitously-expressed enzyme that regulates intracellular folate metabolism for cell proliferation, DNA synthesis, and repair. Employing GGH immunohistochemistry on a tissue microarray with 12,427 prostate cancers, we found that GGH expression was negative to low in normal prostate epithelium, whereas 88.3% of our 10,562 interpretable cancers showed GGH expression. GGH staining was considered as low intensity in 49.6% and as high intensity in 38.6% of cancers. High GGH expression was linked to the TMPRSS2:ERG-fusion positive subset of cancers (p < 0.0001), advanced pathological tumor stage, and high Gleason grade (p < 0.0001 each). Further analysis revealed that these associations were merely driven by the subset of ERG-negative cancers, High GGH expression was weakly linked to early biochemical recurrence in ERG negative cancers (p < 0.0001) and independent from established histo-pathological parameters. Moreover, GGH expression was linked to features of genetic instability, including presence of recurrent deletions at 3p, 5q, 6q, and 10q (PTEN, p ≤ 0.01 each), as well as to accelerated cell proliferation as measured by Ki67 immunohistochemistry (p < 0.0001). In conclusion, the results of our study identify GGH as an ERG subtype specific molecular marker with modest prognostic relevance, which may have clinical relevance if analyzed in combination with other molecular markers.

Endotoxin induces proliferation of NSCLC in vitro and in vivo: role of COX-2 and EGFR activation.

Lung cancer is frequently complicated by pulmonary infections which may impair prognosis of this disease. Therefore, we investigated the effect of bacterial lipopolysaccharides (LPS) on tumor proliferation in vitro in the non-small cell lung cancer (NSCLC) cell line A549, ex vivo in a tissue culture model using human NSCLC specimens and in vivo in the A549 adenocarcinoma mouse model. LPS induced a time- and dose-dependent increase in proliferation of A549 cells as quantified by MTS activity and cell counting. In parallel, an increased expression of the proliferation marker Ki-67 and cyclooxygenase (COX)-2 was detected both in A549 cells and in ex vivo human NSCLC tissue. Large amounts of COX-2-derived prostaglandin (PG)E(2) were secreted from LPS-stimulated A549 cells. Pharmacological interventions revealed that the proliferative effect of LPS was dependent on CD14 and Toll-like receptor (TLR)4. Moreover, blocking of the epidermal growth factor receptor (EGFR) also decreased LPS-induced proliferation of A549 cells. Inhibition of COX-2 activity in A549 cells severely attenuated both PGE(2) release and proliferation in response to LPS. Synthesis of PGE(2) was also reduced by inhibiting CD14, TLR4 and EGFR in A549 cells. The proliferative effect of LPS on A549 cells could be reproduced in the A549 adenocarcinoma mouse model with enhancement of tumor growth and Ki-67 expression in implanted tumors. In summary, LPS induces proliferation of NSCLC cells in vitro, ex vivo in human NSCLC specimen and in vivo in a mouse model of NSCLC. Pulmonary infection may thus directly induce tumor progression in NSCLC.

On the significance of Surfactant Protein-A within the human lungs.

Surfactant Protein-A (SP-A) is the most prominent among four proteins in the pulmonary surfactant-system. SP-A is expressed by alveolar epithelial cells type II as well as by a portion of non small cell lung carcinomas (NSCLC).The expression of SP-A is complexly regulated on the transcriptional and the chromosomal level. SP-A is a major player in the pulmonary cytokine-network and moreover has been described to act in the pulmonary host defense.By the use of cell culture or animal models the functional properties have been repeatedly shown in many aspects, often bearing surprising properties which strongly indicate the physiological importance of SP-A. To date SP-A is recognized as a molecule essential for pulmonary development, structure and function. An upcoming number of reports deals with the role of SP-A for pulmonary pathology. This article gives an overview about the state of knowledge on SP-A focused in applications for human pulmonary disorders and points out the importance for pathology-orientated research approaches using immunohistochemistry or in situ hybridization as promising methods to further elucidate the role of this molecule in adult lung diseases.

LED-FISH: Fluorescence microscopy based on light emitting diodes for the molecular analysis of Her-2/neu oncogene amplification.

Light emitting diodes (LED), which are available as small monochromatic light sources with characteristic features such as maximum illumination power combined with minimum energy consumption and extremely long lifespan have already proved as a highly potential low-cost alternative for specific diagnostic applications in clinical medicine such as tuberculosis fluorescence microscopy. Likewise, the most reliable evaluation of Her-2/neu (c-erbB2) gene amplification, which has been established in the last few years for routine diagnosis in clinical pathology as determinant towards Herceptin-based treatment of patients with breast cancer, is based on fluorescence in situ hybridization (FISH) and corresponding high priced fluorescence equipment. In order to test the possibility to utilize the advantages of low-cost LED technology on FISH analysis of c-erbB2 gene expression for routine diagnostic purposes, the applicability of a standard bright field Carl Zeiss Axiostar Plus microscope equipped with a Fraen AFTER (Amplified Fluorescence by Transmitted Excitation of Radiation) LED Fluorescence Microscope Kit for the detection of Her-2/neu gene signals was compared to an advanced Nikon Eclipse 80i fluorescence microscope in combination with a conventional 100W mercury vapor lamp. Both microscopes were fitted with the same Quicam FAST CCD digital camera to unequivocally compare the quality of the captured images. C-erbB2 gene expression was analyzed in 30 different human tissue samples of primary invasive breast cancer, following formalin fixation and subsequent paraffin-embedding. The Her2/neu gene signals (green) were identifiable in the tumor cells in all cases and images of equal quality were captured under almost identical conditions by 480 nm (blue) LED module equipped standard Axiostar microscope as compared to conventional fluorescence microscopy. In this first attempt, these monochromatic LED elements proved in principle to be suitable for the detection of Her-2/neu gene expression by FISH. Thus, our own experiences emphasize the high potential of this technology to provide a serious alternative to conventional fluorescence microscopy in routine pathology; representing a sustainable technological progress, this low-cost technology will clearly give direction also to the growing field of molecular pathology.