Oers, J.M.M. van

Prediction of Progression of Non-Muscle-Invasive Bladder Cancer by WHO 1973 and 2004 Grading and by FGFR3 Mutation Status: A Prospective Study (Article)

2008-10-01T00:00:00Z

Objectives: The clinical management of non-muscle-invasive urothelial cell carcinoma of the bladder (UCC) is challenging, as it has a marked tendency to recur and to progress. Aim of this study was to investigate the prognostic value of the WHO 1973 and 2004 grading systems and biomarkers FGFR3, CK20 and Ki-67. Methods: In a prospective study, tumours from 221 patients were studied for the expression of CK20 and Ki-67 by immunohistochemistry, and FGFR3 status by SNaPshot mutation detection. Staging and grading were performed according to the WHO classification systems of 1973 and 2004. Results: : Median follow-up was 35 mo. Recurrence occurred in 72 of 221 patients. None of the parameters was able to predict disease recurrence. CK20, Ki-67, FGFR3 mutation, molecular grade using FGFR3 mutation analysis and Ki-67, and histological grading and staging were significantly associated with disease progression in stage. In multivariable analyses, WHO 1973 and 2004 grading systems remained statistically significant and independent predictors of progression, with p = 0.005 for WHO 1973 and p = 0.004 for 2004. FGFR3 status was able to discriminate progressors from nonprogressors in a subset of patients with high-grade UCC (p = 0.009). Conclusions: This is the first prospective study comparing the WHO 1973 and 2004 grading systems. We show that both grading systems contribute valuable independent information. Therefore, it should be considered whether a better grading system could be developed that incorporates essential elements from both. The combination of WHO 2004 grading with FGFR3 status allows a better risk stratification for patients with high-grade non-muscle-invasive UCC.

Activation of the Fibroblast Growth Factor Receptor 3 in Bladder Cancer (Doctoral Thesis)

2007-09-05T00:00:00Z

The identification of frequent FGFR3 mutations in superficial bladder cancer suggests that mutation of the FGFR3 gene is a key genetic event in the development of noninvasive bladder tumors. Furthermore, FGFR3 mutations were associated with a good prognosis, suggesting that the activation of FGFR3 has a beneficial effect during urothelial tumor formation. In this thesis, two aspects of FGFR3 mutations in bladder cancer have been investigated. First, the potential use of FGFR3 mutations in bladder cancer diagnosis, prognosis, and in surveillance of patients with bladder cancer has been explored. The second, more important aim was to get insight in the functional role of mutant FGFR3 in bladder carcinogenesis. For clinical studies, a simple and fast method for FGFR3 mutation detection was developed that is fast, easy, more sensitive and less laborious than previous techniques. With this technique, several studies described in this thesis were done on the role of FGFR3 in bladder carcinogenesis and patient outcome. We analyzed FGFR3 mutations in urothelial hyperplasia, a precursor of low-grade papillary carcinoma, and found that FGFR3 mutations are already present in hyperplastic lesions. Furthermore, we reported a low number of chromosomal aberrations in bladder tumors with an FGFR3 mutation, which suggests that FGFR3 mutated tumors are genetically stable, in contrast to most tumors that do not carry this mutation, and we showed that this genetic stability of FGFR3 mutant tumors was paralleled by a normal differentiation of urothelial cells, since a normal staining pattern for cytokeratin 20, a marker for terminal differentiation of urothelium, was correlated with bladder cancers carrying the FGFR3 mutation. We also described the relation of molecular markers to bladder cancer patient outcome. The subset of FGFR3 mutated tumors with a normal CK20 staining pattern rarely progressed, providing a combination of two molecular markers that is able to define the group of prognostically favorable low-grade noninvasive papillary tumors. Upper urinary tract tumors (i.e. in the ureter and renal pelvis) may be genetically different from bladder tumors. The FGFR3 mutation frequency was, however, equally high (~50%) in all urothelial tract tumors, and FGFR3 mutation status was an independent predictor for progression and disease-specific survival in tumors from the ureter. Finally, we report the results of an experimental study analyzing the expression of a mutant FGFR3 receptor transfected in a human bladder cancer cell line. The most striking effect was that cells expressing the mutant FGFR3 receptor display both loss of integrin expression and increased apoptosis when cultured in Matrigel. This would suggest that interaction of bladder cancer cells expressing mutant FGFR3 with Matrigel (i.e. basement membrane substances) does not permit their survival. The latter would also explain the clinical finding that FGFR3 mutant bladder cancers have a comparatively low tendency to become invasive. In conclusion, this thesis describes both clinical and functional aspects of the activation of FGFR3 in bladder cancer. The results confirm the association of mutant FGFR3 with well-differentiated, noninvasive bladder tumors with few genetic alterations and a good prognosis.

FGFR3 Mutations and a Normal CK20 Staining Pattern Define Low-Grade Noninvasive Urothelial Bladder Tumours (Article)

2007-09-01T00:00:00Z

Objectives: Molecular markers superior to conventional clinicopathologic parameters are needed to predict disease courses in bladder cancer patients. In this study, we investigated four markers (Ki-67, TP53, CK20, FGFR3) in primary urothelial bladder tumours and compared them with traditional pathologic features. Methods: Tissue microarrays were used to analyse CK20, TP53, and Ki-67 expression immunohistochemically in 255 unselected patients. FGFR3 mutations were detected by SNaPshot analysis. Results: Abnormal CK20 expression was strongly associated with higher tumour grades and stages (p < 0.001); however, 65% of pTa tumours revealed an abnormal CK20 pattern. In the group of pTaG1 tumours, 59% presented with an abnormal CK20 pattern, whereas 82% carried the FGFR3 mutation. In the group of bladder tumours with normal CK20 pattern, the FGFR3 gene was mutated in 89%, whereas a mutated FGFR3 gene was found in only 37% of cases with abnormal CK20 expression (p < 0.001). All markers proved to be strong predictors of disease-specific survival in univariate studies. However, in multivariate analyses they were not independent from classical pathologic parameters. None of the molecular markers was significantly associated with tumour recurrence. Conclusions: Dysregulation of CK20 expression is an early event in the carcinogenesis of papillary noninvasive bladder cancer, but occurs later than FGFR3 mutations. The group of low-grade noninvasive papillary tumours is defined by the presence of an FGFR3 mutation and a normal CK20 expression pattern.

Low frequency of molecular changes and tumor recurrence in inverted papillomas of the urinary tract (Article)

2007-06-01T00:00:00Z

AIM: Inverted papilloma (IP) of the urinary tract can be difficult to distinguish from noninvasive urothelial carcinoma with prominent inverted growth pattern (invNIUC). Ancillary markers may help to resolve such cases and clarify the reported malignant potential of some IPs. METHODS: Eighty-nine urothelial lesions initially diagnosed as IP were reviewed by 4 experienced urologic pathologists and studied immunohistochemically (Ki67, p53, CK20, MSH2, MLH1, and MSH6). Mutations of the FGFR3 gene, deletions (loss of heterozygosity) of 9p, 9q, and 17p, microsatellite instability, and elevated microsatellite instability at selected tetranucleotides were also analyzed. RESULTS: Considerable interobserver variability in histopathologic diagnoses was noticed. Only 62 (69.7%) initial diagnoses were confirmed by the review pathologists whereas 23 tumors (25.8%) were redefined as invNIUC. Molecular analyses revealed infrequent alterations in IPs, including microsatellite instability (1.8%), elevated microsatellite instability at selected tetranucleotides (13.2%), FGFR3 mutations (9.8%), 9p deletions (3.9%), 9q deletions (13.2%), 17p deletions (5.1%), nuclear p53 accumulation (18.9%), and aberrant immunostaining for MSH2 (5.8%), MLH1 (11.8%), and MSH6 (3.8%). IP and invNIUC differed in FGFR3 mutations and Ki-67 labeling index (P<0.001 each), and 9q loss of heterozygosity (P=0.03). There were fewer recurrences in IP (5.4%) compared with invNIUC (40.9%; P<0.0001). CONCLUSIONS: IP is a benign lesion that lacks specific genetic alterations found in exophytic noninvasive papillary urothelial tumors. These lesions could be reactive in nature, perhaps secondary to chronic inflammation or a neoplastic process that lack specific genetic alterations. Nevertheless given the clinical and molecular data of this study a conservative clinical approach is appropriate.

A simple and fast method for the simultaneous detection of nine fibroblast growth factor receptor 3 mutations in bladder cancer and voided urine (Article)

2005-11-01T00:00:00Z

Purpose:Mutations in the fibroblast growth factor receptor 3 (FGFR3) occur in 50% of primary bladder tumors.An FGFR3 mutationis associatedwith goodprognosis, illustrated by significantly lower percentage of patients with progression and disease-specific mortality. FGFR3 mutations are especially prevalent in low grade/stage tumors, with pTa tumors harboring mutations in 85% of the cases.These tumors recur in 70% of patients. Efficient FGFR3 mutation detection for prognostic purposes and for detectionof recurrences inurine is animportant clinical issue. Inthis paper, we describe a simple assay for the simultaneous detection of nine different FGFR3 mutations. Experimental Design: The assay consists of one multiplex PCR, followed by extension of primers for each mutation with a labeled dideoxynucleotide.The extended primers are separated by capillary electrophoresis, and the identity of the incorporated nucleotide indicates the presence or absence of amutation. Results: The assay was found to be more sensitive than single-strand conformation polymorphism analysis.Mutations could still be detected with an input of only 1ng of genomic DNA and in a 20-fold excess of wild-type DNA. Moreover, in urine samples from patients with a mutant tumor, the sensitivity of mutation detectionwas 62%. Conclusions:We have developed a fast, easy to use assay for the simultaneous detection of FGFR3 mutations, which can be of assistance in clinical decision-making and as an alternative for the follow-up of patients by invasive cystoscopy for the detection of recurrences in urine.