No abstract

Studies by comparative genomic hybridization revealed that the 19q13 chromosomal region is frequently amplified in bladder cancer. The cyclin E gene (CCNE), coding for a regulatory subunit of cyclin-dependent kinase 2, has been mapped to 19q13. To investigate the role of cyclin E alterations in bladder cancer, a tissue microarray of 2,317 specimens from 1,842 bladder cancer patients was constructed and analyzed for CCNE amplification by fluorescence in situ hybridization and for cyclin-E protein overexpression by immunohistochemistry. Fluorescence in situ hybridization analysis showed amplification in only 30 of the 1,561 evaluable tumors (1.9%). Amplification was significantly associated with stage and grade (P: < 0.0005 each). Immunohistochemically detectable cyclin E expression was strong in 233 (12.4%), weak in 354 (18.9%), and negative in 1, 286 of the 1,873 interpretable tumors. The majority (62.1%) of CCNE-amplified tumors were strongly immunohistochemistry-positive (P: < 0.0001). The frequency of protein expression increased from stage pTa (22.2%) to pT1 (45.5%; P: < 0.0001) but then decreased for stage pT2-4 (29.4%; P: < 0.0001 for pT1 versus pT2-4). Low cyclin E expression was associated with poor overall survival in all patients (P: < 0.0001), but had no prognostic impact independent of stage. It is concluded that cyclin E overexpression is characteristic to a subset of bladder carcinomas, especially at the stage of early invasion. This analysis of the prognostic impact of CCNE gene amplification and protein expression in >1,500 arrayed bladder cancers was accomplished in a period of 2 weeks, illustrating how the tissue microarray technology remarkably facilitates the evaluation of the clinical relevance of molecular alterations in cancer.

The number of genes suggested to play a role in cancer biology is rapidly increasing. To be able to test a large number of molecular parameters in sufficiently large series of primary tumours, a tissue microarray (TMA) approach has been developed where samples from up to 1000 tumours can be simultaneously analysed on one glass slide. Because of the small size of the individual arrayed tissue samples (diameter 0.6 mm), the question arises of whether these specimens are representative of their donor tumours. To investigate how representative are the results obtained on TMAs, a set of 2317 bladder tumours that had been previously analysed for histological grade and Ki67 labelling index (LI) was used to construct four replica TMAs from different areas of each tumour. Clinical follow-up information was available from 1092 patients. The histological grade and the Ki67 LI were determined for every arrayed tumour sample (4x2317 analyses each). Despite discrepancies in individual cases, the grade and Ki67 information obtained on minute arrayed samples were highly similar to the data obtained on large sections (p<0.0001). Most importantly, every individual association between grade or Ki67 LI and tumour stage or prognosis (recurrence, progression, tumour-specific survival) that was observed in large section analysis could be fully reproduced on all four replica TMAs. These results show that intra-tumour heterogeneity does not significantly affect the ability to detect clinico-pathological correlations on TMAs, probably because of the large number of tumours that can be included in TMA studies. TMAs are a powerful tool for rapid identification of the biological or clinical significance of molecular alterations in bladder cancer and other tumour types.

No abstract

Plasma cell myelomas (PMs) have a poor prognosis. Cancer-testis (CT) antigens are immunogenic proteins, representing potential targets for tumor vaccination strategies. The expression of the CT antigens GAGE, MAGE-A4, MAGE-C1/CT-7, and NY-ESO-1 was investigated on paraffin-embedded bone marrow biopsies from 219 PM and 8 monoclonal gammopathy of undetermined significance (MGUS) patients. The frequency and prognostic impact of these CT antigens were compared with known morphological prognostic markers (i.e. Mib1 labeling index) and the presence of the translocations t(4;14)(p16.3; q32) and t(11;14)(q13;q32). We show that MAGE-C1/CT-7 is the most prevalent CT antigen, expressed in 57% of PMs in a high percentage of tumor cells. While MAGE-C1/CT-7 was absent in non-malignant plasma cells, plasma cells of patients with MGUS did express MAGE-C1/CT-7, but no other CT antigens. MAGE-C1/CT-7 was more frequently expressed in PMs with an elevated proliferation rate (Mib1 > > > >10%) compared to PMs with a low proliferation rate (Mib1 ≤ ≤ ≤ ≤10%, 71% versus 29%, P < < < < 0.001) and correlated with overall survival, depending on its subcellular distribution. PMs with pure cytoplasmic MAGE-C1/CT-7 expression showed a better prognosis (48 months versus 33 months, P < < < < 0.05) than PMs with combined nuclear-cytoplasmic or nuclear expression only. Thus, expression of MAGE-C1/CT-7 in patients with monoclonal gammopathies represents a predictor of outcome and overt malignant transformation. (Cancer Sci 2008; 99: 720-725)

A multicentre study of 51 cases of lymph-node infarction seen in the 30-year period 1956 to 1985 was conducted in order to assess both the short- and long-term prognostic implications of the condition. In 14 cases malignant lymphoma was found synchronously with the infarct. Of the remaining 37 patients with apparently 'benign' lymph-node infarction only six showed manifestations of malignant lymphoma in the follow-up time studied (mean = 48 months; range 1-156 months). These subsequent malignant lymphomas all occurred within 2 years of the lymph-node infarction. A postal enquiry and collation of other cases in the medical literature indicates that a minority (26 of 81) have developed malignant lymphoma, and that these lymphomas, too, have all appeared within 2 years. Thorough examination of both the infarcted lymph nodes and others resected at the same time is mandatory in order to exclude concomitant or underlying malignant lymphoma. Two years after lymph-node infarction the risk of malignant lymphoma is negligible.

Biopsies from 25 patients with primary malignant lymphoma in the salivary region were investigated morphologically and the clinical findings were analysed. Cases showing myoepithelial sialadenitis or Sjögren's syndrome were not included. The tumour was localized in the parotid region in 21 cases and to the submandibular region in four cases. Non-Hodgkin's lymphoma was diagnosed on 21 biopsies and Hodgkin's disease on four: all patients were of stages I or II. The most frequent type of malignant lymphoma was the centroblastic-centrocytic type; sclerosis was found in all but one of these 15 cases. Polymorphic immunocytoma was diagnosed in two cases, centroblastic lymphoma in two cases and immunoblastic lymphoma in two cases. In eight patients, the lymphomas definitely originated in intraglandular lymph nodes; in 10 other patients, the lymphomas might have developed in intraglandular lymph nodes. It was not possible to determine the origin of the lymphoma in the other seven cases. The prognosis was relatively favourable.

TAAs of the MAGE family are mostly studied as targets of specific immune responses. Their potential relevance as tumor markers has also been underlined. We used a MAb, 57B, recognizing MAGE-A4 protein in paraffin-embedded sections, to evaluate its expression in bladder cancers by employing TMA including 2,317 samples from 1,849 patients. In 2,090/2,317 cases (90.2%), immunostaining yielded interpretable results. Since for some patients more than 1 sample was available, only interpretable first biopsies (n ‫؍‬ 1,628) were considered. MAGE-A4 protein was expressed at significantly (p < 0.001) higher frequency in squamous (25/55, 45.5%) than in adeno (4/15, 26.7%), sarcomatoid (4/14, 28.6%), small cell (5/20, 25%) or transitional cell (281/1,522, 18.5%) carcinomas. In TCCs, overall MAGE-A4 positivity was significantly correlated with invasive phenotype (p < 0.001) and high tumor grade (p < 0.0001). Clinical data from 908 TCC patients were retrospectively evaluated, revealing that strong 57B staining was highly significantly associated with decreased tumorspecific survival (p < 0.0001). These data suggest that evaluation of MAGE-A4 protein expression is useful in the identification of groups of TCCs characterized by severe prognosis, thus possibly providing indications for early MAGE TAA-targeted immunotherapy. © 2002 Wiley-Liss, Inc. Key words: bladder cancer; 57B MAb; MAGE-A4; tissue microarray; tumor-specific survivalHuman TAAs of the MAGE family are recognized by CTLs from tumor patients. 1,2 Capitalizing on this background, clinical immunization trials are currently being implemented, showing promising preliminary results. [3][4][5][6][7] Concomitantly, the potential relevance of these proteins as tumor markers is also emerging. Clinical studies suggest that MAGE TAA expression is prevailingly detectable in highly aggressive, poorly differentiated tumors, 8 -10 thereby increasing their interest as therapeutic targets.In particular, MAGE gene expression has also been described in urinary bladder cancers, 8 but the number of cases was too low to allow evaluation of clinicopathologic or prognostic correlations.TMA technology offers the unique possibility to rapidly analyze large numbers of tumors. 11 Minute tissue cylinders (diameter 0.6 mm) are taken from hundreds of different primary tumor blocks and subsequently brought into 1 empty recipient paraffin block. Sections from such array blocks can then be used for simultaneous in situ analysis of hundreds or thousands of tumors at the DNA, RNA or protein level.In this study, we used a bladder cancer TMA containing 2,317 specimens to evaluate the frequency of MAGE-A4 protein expression as detected by a specific MAb (57B) and to assess its correlation with tumor phenotype and clinical outcome. MATERIAL AND METHODS MAb 57BReagent 57B was generated by our group using as immunogen recombinant MAGE-A3 protein. 12 Immunohistochemic studies carried out in different laboratories have emphasized that, although 57B identifies multiple MAGE gene products in transfected cells, ...