Aims: To determine interobserver variation in grading of dysplasia in Barrett's oesophagus (BO) between non‐expert general pathologists and expert gastrointestinal pathologists on the one hand and between expert pathologists on the other hand. Methods and results: In this prospective multicentre study, non‐expert and expert pathologists graded biopsy specimens of 920 patients with endoscopic BO, which were blindly reviewed by one member of a panel of expert pathologists (panel experts) and by a second panel expert in case of disagreement on dysplasia grade. Agreement between two of three pathologists was established as the final diagnosis. Analysis was performed by κ statistics. Due to absence of intestinal metaplasia, 127/920 (14%) patients were excluded. The interobserver agreement for dysplasia [no dysplasia (ND) versus indefinite for dysplasia/low‐grade dysplasia (IND/LGD) versus high‐grade dysplasia (HGD)/adenocarcinoma (AC)] between non‐experts and first panel experts and between initial experts and first panel experts was fair (κ = 0.24 and κ = 0.27, respectively), and substantial for differentiation of HGD/AC from ND/IND/LGD (κ = 0.62 and κ = 0.58, respectively). Conclusions: There was considerable interobserver variability in the interpretation of ND or IND/LGD in BO between non‐experts and experts, but also between expert pathologists. This suggests that less subjective markers are needed to determine the risk of developing AC in BO.
5686 Prostate cancer is the most common malignancy in men in developed countries. The molecular mechanisms underlying prostate cancer development and progressive growth are still poorly understood. To unravel these mechanisms we analyzed genomic DNA from eleven human prostate cancer xenografts by 1Mb spaced array-based comparative genomic hybridization (array CGH). Xenograft DNA lacks DNA from normal cells of human origin, allowing most accurate genomic studies. Xenografts reflect a variety of clinical stages of prostate cancer. The most frequent small region of chromosomal loss (approx. 3 Mb) was on 21q22.2–q22.3 (4 xenografts). This generates the recently described TMPRSS:ERG fusion gene, linking the TMPRSS2 promoter and exon 1 to exon 4 of the ERG gene. RT-PCR also showed variable levels of expression of the TMPRSS2:ERG fusion gene in three other xenografts. Further genomic analyses indicated two alternative mechanisms of TMPRSS2 to ERG gene fusion. Eleven homozygous deletions in eight different xenografts were detected. Most are predicted to be relevant for tumorigenesis. The most frequent homozygous deletion detected by array CGH was that of PTEN on 10q23.3 (3 xenografts). Novel homozygous deletions on 2q37.1 and 8p23.3-pter need further investigation, because obvious candidate tumor suppressor genes could not be indicated. A homozygous deletion close to fragile site FRA16D on 16q23.1 affected the WWOX gene. The homozygous deletion on 13q13.1 included at least three genes implicated in tumorigenesis: BRCA2, APRIN and DLC2; the one on 13q14.2 included RB1. Other homozygous deletions affected ATBF1 on 16q22.2–q22.3 and MKK4 on 17p12. A homozygous deletion on 17p11.2–p12 encompassed the N-COR transcription co-repressor gene. Detailed analysis of xenograft DNAs by PCR identified a second homozygous deletion of N-COR, which was not visualized by array CGH. Because N-COR has never been implicated in prostate cancer, it was subjected to further structural and functional investigation. Additionally to the two homozygous deletions we detected mutation or loss of one copy of N-COR in four other xenografts. Functional studies, utilizing N-COR specific siRNA, demonstrated stimulated in vitro growth of LNCaP prostate cancer cells. Our observations, as guided by array CGH, add ERG and N-COR to PTEN as novel important genes in prostate cancer.
<div>Abstract<p>In this study, we describe the properties of novel <i>ETV1</i> fusion genes, encoding N-truncated ETV1 (dETV1), and of full-length ETV1, overexpressed in clinical prostate cancer. We detected overexpression of novel <i>ETV1</i> fusion genes or of full-length <i>ETV1</i> in 10% of prostate cancers. Novel <i>ETV1</i> fusion partners included <i>FOXP1</i>, an EST (<i>EST14</i>), and an endogenous retroviral repeat sequence (<i>HERVK17</i>). Like <i>TMPRSS2, EST14</i> and <i>HERVK17</i> were prostate-specific and androgen-regulated expressed. This unique expression pattern of most <i>ETV1</i> fusion partners seems an important determinant in prostate cancer development. In transient reporter assays, full-length ETV1 was a strong transactivator, whereas dETV1 was not. However, several of the biological properties of dETV1 and full-length ETV1 were identical. On stable overexpression, both induced migration and invasion of immortalized nontumorigenic PNT2C2 prostate epithelial cells. In contrast to dETV1, full-length ETV1 also induced anchorage-independent growth of these cells. PNT2C2 cells stably transfected with dETV1 or full-length ETV1 expression constructs showed small differences in induced expression of target genes. Many genes involved in tumor invasion/metastasis, including <i>uPA/uPAR</i> and <i>MMPs</i>, were up-regulated in both cell types. Integrin β3 (<i>ITGB3</i>) was clearly up-regulated by full-length ETV1 but much less by dETV1. Based on the present data and on previous findings, a novel concept of the role of dETV1 and of full-length ETV1 overexpression in prostate cancer is proposed. [Cancer Res 2008;68(18):7541–9]</p></div>
Abstract Aim To report the long term results of the CROSS trial with a minimum follow-up of 10 years. Background Neoadjuvant chemoradiotherapy according to the Dutch randomised controlled ChemoRadiotherapy for Oesophageal cancer followed by Surgery Study (CROSS) has become standard of care for patients with cancer of the oesophagus or oesophagogastric junction. Methods Patients with locally advanced resectable squamous cell carcinoma or adenocarcinoma of the oesophagus or oesophagogastric junction were randomised between neoadjuvant chemoradiotherapy (five weekly cycles of intravenous carboplatin [AUC 2 mg/mL per min] and intravenous paclitaxel [50 mg/m² of body-surface area]) with concurrent 41.4 Gy radiotherapy given in 23 fractions of 1.8 Gy, 5 days per week) plus surgery versus surgery alone. Primary endpoint was overall survival, defined from date of randomisation to date of all-cause death or to last day of follow-up. Analysis was by intention-to-treat. Results Between March 2004 and 2008, eight centres enrolled 368 patients. Some 178 were analysed in the chemoradiotherapy plus surgery group and 188 in the surgery alone group. After a median follow-up for surviving patients of 146.6 months (IQR 133.5-156.6), median overall survival was 49.0 months (95%CI 34.7-76.6) in the neoadjuvant chemoradiotherapy plus surgery group compared to 25.1 months (95%CI 19.1-39.4) in the surgery alone group, which was significantly different (HR 0.71 [95%CI 0.55-0.90]; log-rank p=0·005). Ten-year overall survival was 38% (95%CI 31%-45%) in the neoadjuvant chemoradiotherapy followed by surgery group compared to 26% (95%CI 20%-33%) in the surgery alone group (HR 0.69 [95%CI 0.54-0.89]). For patients with squamous cell carcinoma ten-year overall survival was 46% (95%CI 33%-64%) in the neoadjuvant chemoradiotherapy plus surgery group compared to 23% (95%CI 13%-40%) in the surgery alone group. For patients with adenocarcinoma ten-year overall survival was 36% (95%CI 29%-45%) in the neoadjuvant chemoradiotherapy plus surgery group compared to 26% (95%CI 20%-35%) in the surgery alone group. Conclusion Survival benefit of patients with locally advanced resectable squamous cell carcinoma or adenocarcinoma of the oesophagus or oesophagogastric junction receiving neoadjuvant chemoradiotherapy persists for at least 10 years compared to patients undergoing surgery alone.
Decalcification is routinely performed for histological studies of bone-containing tissue. Although DNA in situ hybridization (ISH) and comparative genomic hybridization (CGH) have been successfully employed on archival material, little has been reported on the use of these techniques on archival decalcified bony material. In this study we compared the effects of two commonly used decalcifiers, i.e., one proprietary, acid-based agent (RDO) and one chelating agent (EDTA), in relation to subsequent DNA ISH and CGH to bony tissues (two normal vertebrae, six prostate tumor bone metastases with one sample decalcified by both EDTA and RDO). We found that RDO-decalcified tissue was not suited for DNA ISH in tissue sections with centromere-specific probes, whereas we were able to adequately determine the chromosomal status of EDTA-decalcified material of both control and tumor material. Gel electrophoresis revealed that no DNA could be successfully retrieved from RDO-treated material. Moreover, in contrast to RDO-decalcified tumor material, we detected several chromosomal imbalances in the EDTA-decalcified tumor tissue by CGH analysis. Furthermore, it was possible to determine the DNA ploidy status of EDTA-but not of RDO-decalcified material by DNA flow cytometry. Decalcification of bony samples by EDTA is highly recommended for application in DNA ISH and CGH techniques.
We have applied fluorescent in situ hybridization (FISH) to assess the presence of numerical chromosome aberrations in fresh specimens of human solid tumors of varying histology. For this purpose, a set of 12 biotinylated chromosome-specific, repetitive alpha-satellite DNA probes (for chromosomes 1, 6, 7, 9, 10, 11, 15, 16, 17, 18, X and Y) were hybridized directly to isolated interphase nuclei. Utilizing this approach, we found numerical chromosome changes in all tumors. FISH ploidy profiles were in accordance with flow cytometric DNA histograms of these tumor cells.
