Ki-67 MIB1 labelling index and the prognosis of primary TaT1 urothelial cell carcinoma of the bladder.

AIMS: To evaluate whether ki-67 labelling index (LI) has independent prognostic value for survival of patients with bladder urothelial tumours graded according to the 2004 World Health Organisation classification. METHODS: Ki-67 LI was evaluated in 164 cases using the grid counting method. Non-invasive (stage Ta) tumours were: papilloma (n = 5), papillary urothelial neoplasia of low malignant potential (PUNLMP; n = 26), and low (LG; n = 34) or high grade (HG; n = 15) papillary urothelial carcinoma. Early invasive (stage T1) tumours were: LG (n = 58) and HG (n = 26) carcinoma. Statistical analysis included Fisher and chi2 tests, and mean comparisons by ANOVA and t test. Univariate and multivariate survival analyses were performed according to the Kaplan-Meier method with log rank test and Cox's proportional hazard method. RESULTS: Mean ki-67 LI increased from papilloma to PUNLMP, LG, and HG in stage Ta (p<0.0001) and from LG to HG in stage T1 (p = 0.013) tumours. High tumour proliferation (>13%) was related to greater tumour size (p = 0.036), recurrence (p = 0.036), progression (p = 0.035), survival (p = 0.054), and high p53 accumulation (p = 0.015). Ki-67 LI and tumour size were independent predictors of disease free survival (DFS), but only ki-67 LI was related to progression free survival (PFS). Cancer specific overall survival (OS) was related to ki-67 LI, tumour size, and p27kip1 downregulation. Ki-67 LI was the main independent predictor of DFS (p = 0.0005), PFS (p = 0.0162), and cancer specific OS (p = 00195). CONCLUSION: Tumour proliferation measured by Ki-67 LI is related to tumour recurrence, stage progression, and is an independent predictor of DFS, PFS, and cancer specific OS in TaT1 bladder urothelial cell carcinoma.

Changes produced in the urothelium by traditional and newer therapeutic procedures for bladder cancer.

A handful of traditional and newer therapeutic procedures, such as chemotherapy, immunotherapy, radiotherapy, photodynamic and laser treatment, and gene therapy, are used to treat epithelial malignancies of bladder origin. These treatment modalities, used either intravesically or systemically, produce morphological changes in the urothelial mucosa that can be mistaken for carcinoma. The pathologist must be able to separate toxic and drug related alterations from tumour related changes. The clinical history is usually invaluable in this assessment.

The pagetoid variant of bladder urothelial carcinoma in situ A clinicopathological study of 11 cases.

Pagetoid urothelial carcinoma in situ (CIS) is a rare variant of bladder cancer that is characterized by an intraepithelial proliferation of large cells arranged singly or in clusters and randomly distributed. These neoplasms deserve recognition and attention, chiefly because they may be overlooked or misdiagnosed as urothelial dysplasia, then causing unsuspected tumor recurrence after surgery. We report on the clinicopathological features and immunohistochemical findings of 11 (14.86%) cases of pagetoid CIS in a retrospective study of 74 cases of conventional carcinoma in situ. Most patients were male ( n=10). Their ages ranged from 31 years to 78 years. The lesion can be present with primary ( n=2) or secondary ( n=9) CIS. Pagetoid CIS is usually a focal lesion occurring in a clinical and histological setting of conventional CIS, and these patients essentially have the same progression and survival rates as patients without pagetoid changes and are treated in the same way. In cases with extensive urothelial denudation, pagetoid CIS may be focally present in otherwise normal-looking urothelium, thus alerting the pathologist to search for additional CIS elsewhere in the bladder. Given that primary extramammary Paget disease of the external genitalia and of the anal canal may extend to the bladder and, conversely, some bladder cases of pagetoid CIS may extend to the urethra, ureter, and beyond to the external genitalia, the differential diagnoses between these two entities represent an important therapeutic consideration. Our data suggest that a panel of immunostains including CK7+/CK20+/TM+ may assist in differentiating urothelial pagetoid CIS from extramammary Paget disease which is known to be CK7+/CK20-.

Hepatoid adenocarcinoma of the urinary bladder.

Hepatoid adenocarcinoma is rare in the urinary bladder with only three well-illustrated previously reported cases. Pathological diagnosis is based on a combination of histological features resembling hepatocellular carcinoma and the positive immunostaining for alpha-fetoprotein. We present the clinicopathological features of four additional cases. The patients were males 66, 85, 61 and 68 years old. Hematuria was the initial symptom in all four patients. Two cases were treated by cystoprostatectomy and the remaining two by transurethral resection of the bladder. On histology, the cases showed a mixture of cells growing in a solid fashion and sheets or anastomosing trabeculae of hepatoid cells merging focally with a secondary glandular pattern of adenocarcinoma. Intracytoplasmic hyaline globules in all and bile production in three of the cases also supported the impression of hepatocytic differentiation. Immunoreactivity for alpha-fetoprotein, low molecular weight cytokeratin, alpha-1-antitrypsin, albumin, epithelial membrane antigen and a striking canalicular pattern when stained against polyclonal carcinoembryonic antigen (CEA), all indicate hepatocellular differentiation. The hepatic nature of the cells was further assessed by detecting the recently incorporated marker hepatocyte paraffin 1, by means of immunohistochemistry and albumin gene mRNA non-isotopic in situ hybridization, both of which had positive signals in all four cases. Three patients died 12, 14 and 19 months after diagnosis. The fourth patient was alive with disease at 26 months of follow-up. In conclusion, hepatoid adenocarcinoma seems to be an aggressive malignant neoplasm that is rare in the bladder whose correct diagnosis may need appropriate immunohistochemical and in situ hybridization means in addition to a complete patient clinical and pathological evaluation. The exact histogenesis and classification of these tumors remains to be established.

Lymphoepithelioma-like carcinoma of the urinary bladder: a clinicopathologic study of 13 cases.

Lymphoepithelioma-like carcinoma (LELCA) of the urinary bladder is a rare variant of bladder cancer characterized by a malignant epithelial component densely infiltrated by lymphoid cells. It is characterized by indistinct cytoplasmic borders and a syncytial growth pattern. These neoplasms deserve recognition and attention, chiefly because they may be responsive to chemotherapy. We report on the clinicopathologic features of 13 cases of LELCA recorded since 1981. The chief complaint in all 13 patients was hematuria. Their ages ranged from 58 years to 82 years. All tumors were muscle invasive. A significant lymphocytic reaction was present in all of these tumors. There were three pure LELCA and six predominant LELCA with a concurrent transitional cell carcinoma (TCC). The remainder four cases had a focal LELCA component admixed with TCC. Immunohistochemistry showed LELCA to be reactive against epithelial membrane antigen and several cytokeratins (CKs; AE1/AE3, AE1, AE3, CK7, and CK8). CK20 and CD44v6 stained focally. The lymphocytic component was composed of a mixture of T and B cells intermingled with some dendritic cells and histiocytes. Latent membrane protein 1 (LMP1) immunostaining and in situ hybridization for Epstein-Barr virus were negative in all 13 cases. DNA ploidy of these tumors gave DNA histograms with diploid peaks (n=7) or non-diploid peaks (aneuploid or tetraploid; n=6). All patients with pure and 66% with predominant LELCA were alive, while all patients having focal LELCA died of disease. Our data suggest that pure and predominant LELCA of the bladder appear to be morphologically and clinically different from other bladder (undifferentiated and poorly differentiated conventional TCC) carcinomas and should be recognized as separate clinicopathological variants of TCC with heavy lymphocytic reaction relevant in patient management.

Loss of heterozygosity at 9q32-33 (DBC1 locus) in primary non-invasive papillary urothelial neoplasm of low malignant potential and low-grade urothelial carcinoma of the bladder and their associated normal urothelium.

Tumour recurrence has a major impact on patients with non-invasive papillary urothelial tumours of the bladder. To explore the role of DBC1 (deleted in bladder cancer 1 locus), a candidate tumour suppressor gene located at 9q32-33, as prognostic marker we have performed loss of heterozygosity (LOH) testing in 49 patients with primary papillary urothelial tumours and associated normal urothelium. Data from the 38 tumours and 11 specimens of normal urothelium that were informative in the LOH study (D9S195 marker) showed that LOH in urothelium (45.4%) but not in non-invasive tumours (60.5%) was associated with tumour recurrence (p = 0.026) but not to grade or progression. Also, tumours whose normal urothelium had LOH were larger (p = 0.020) and showed cyclin D1 over-expression (p = 0.032). Non-significant increased expression of p53, p21Waf1, apoptotic index and tumour proliferation, and decreased expression of p27Kip1 or cyclin D3 also characterized tumours whose normal urothelium had LOH. The expression of these G1-S modulators, apoptotic index and tumour proliferation was more heterogeneous in papillary urothelial tumours, irrespective of having retained heterozygosity or LOH. Also, Bax expression decreased in papillary urothelial tumours having LOH (p = 0.0473), but Bcl-2 was unrelated to LOH status. In addition, FGFR3 protein expression decreased in LOH tumours (p = 0.036) and in those having LOH in their normal urothelium (p = 0.022). FGFR3 immunohistochemical expression was validated by western blot in selected cases. The survival analysis selected LOH in normal urothelium as a marker of disease-free survival (log-rank 5.32, p = 0.021), progression-free survival (log-rank 3.97, p = 0.046) and overall survival (log-rank 4.26, p = 0.038); LOH in tumours was significant in progression-free survival (log-rank 3.83, p = 0.042). It is concluded that LOH at the DBC1 locus in normal urothelium seems to be relevant in the prognosis of non-invasive papillary tumours of the bladder via selecting cases with increased proliferation, frequent alterations of the G1-S phase modulators, and decreased FGFR3 protein expression.

Cyclin D3 expression in primary Ta/T1 bladder cancer.

Cyclin D3 deregulation has recently been reported in bladder cancer but its prognostic significance remains uncertain. A cohort of 159 patients with stage Ta or T1 primary bladder tumours was investigated to determine the significance of cyclin D3 expression in association with other G1-S phase regulators of the cell cycle (p53, p21Waf1, p27kip1, cyclin D1), including tumour proliferation (ki67-MIB1); its association with conventional clinicopathological parameters; and the relationship between cyclin D3 and loss of heterozygosity (LOH) at the 9p21 (p16INK4a locus) chromosome region. The end point of the study was progression-free survival. Cyclin D3, other G1-S phase regulators, and tumour proliferation were investigated by immunohistochemistry and measured by the grid-counting method. To validate the immunohistochemical expression, cyclin D3 was additionally assessed by western blotting in selected cases. LOH at the 9p21 chromosome region (marker D9S171) was assessed in 125 cases using an AB Prism 310 genetic analyser and a set of microsatellite fluorescence-labelled primers. Cyclin D3 overexpression was related to larger tumour size (>5 cm; p < 0.0001) and high tumour proliferation (>10%; p = 0.025). Mean cyclin D3 expression increased with 2004 WHO grading categories in stage Ta (p = 0.035, ANOVA) and stage T1 (p = 0.047, t test) tumours. Cyclin D3 was not related to other clinicopathological parameters, G1-S phase modulators, or 9p21 LOH. Cox's multivariate analysis selected cyclin D3 as an independent predictor of progression-free survival (p = 0.0012, relative risk (RR) = 5.2366) together with tumour size (p = 0.0115, RR = 4.4442) and cyclin D1 (p = 0.0065, RR = 3.3023). Cyclin D3 expression had the highest risk ratio. Our results suggest that expression of cyclin D3 is relevant to the progression-free survival of patients with Ta/T1 bladder carcinomas.

A practical approach to bladder sampling and diagnostic reporting of pathological findings.

The most common bladder specimens are obtained from endoscopic biopsies and transurethral resections (TURB), both of which sample subepithelial tissue of varying depth. Other specimens can be obtained from cystectomy, cystoprostatectomy, pelvic exenteration ("en bloc" resection), and partial cystectomy including resection of diverticulae and surgical excision of a urachal carcinoma. The correct assessment of bladder specimens may provide clinically relevant diagnostic and prognostic data. This protocol is intended to assist pathologists in providing clinically useful information as a result of examination of surgical specimens.

Prognostic factors in stage T1 grade 3 bladder cancer survival: the role of G1-S modulators (p53, p21Waf1, p27kip1, Cyclin D1, and Cyclin D3) and proliferation index (ki67-MIB1).

OBJECTIVE: To determine the prognostic value of a subset of regulators of the transition from G1-to-S phase of cell cycle in stage T1 grade 3 bladder cancers. METHODS: Fifty-one such cases were investigated to determine the significance on patient's survival of p53, p21Waf1, p27Kip1, Cyclin D1, Cyclin D3, and ki67-MIB1 (proliferation index). The statistical analysis included Kaplan-Meier methodology with Log-rank test and Cox' proportional hazard analysis. RESULTS: Tumor size (p=0.0034), and the labeling index of ki67-MIB1 (p=0.0034), p53 (p=0.0332), p27kip1 (p=0.0059) and Cyclin D1 (p=0.0103) were associated to disease-free survival. Progression-free survival was related to tumor size (p<0.0001), ki67 (p=0.0163), p53 (p=0.0041), p27kip1 (p=0.0161), Cyclin D1 (p<0.0001) and Cyclin D3 (p<0.0001). Patient's overall survival was associated to Cyclin D3 (p<0.0001), p53 (p=0.0017), p21Waf1 (p=0.0142), Cyclin D1 (p<0.0001), ki67-MIB1 (p=0.0450), and tumor size (p=0.0296). Down-regulation of p27kip1 and Cyclin D3 over-expression (disease-free), over-expression of p53, Cyclin D1 and Cyclin D3 (progression-free), and over-expression of Cyclin D3 (overall survival) were independent predictors by Cox's multivariate analysis. Down-regulation of p27kip1 (p<0.001, R.R. 0.997, 95%C.I. 0.995-0.999), and over-expression of Cyclin D1 (p<0.001, R.R. 1.009, 95%C.I. 1.004-1.014) and Cyclin D3 (p=0.005, R.R. 1.013, 95%C.I. 1.004-1.022) were the main independent predictors. CONCLUSION: Down-regulation of p27kip1 and over-expression of Cyclin D1 and Cyclin D3 might be relevant predictors of survival in stage T1 grade 3 bladder cancers, thus selecting a group of patients at higher risk of malignant behavior.