Low frequency of epigenetic events in urothelial tumors in young patients.

PURPOSE: Bladder urothelial cell carcinoma is uncommon in young patients. We recently reported a series of tumors in patients younger than 20 years at diagnosis and performed exhaustive genetic screening for molecular alterations. Few events typical of bladder urothelial cell carcinoma were detected. Since many carcinogenic events occur at the epigenetic rather than the genetic level, we analyzed the same tumors for alterations in DNA hypermethylation. We compared our findings with those in tumors in older patients with similar pathological profiles. MATERIALS AND METHODS: We analyzed 76 bladder urothelial cell carcinomas from 3 groups stratified by age at diagnosis, including less than 19, 20 to 45 and greater than 46 years (median 78), and matched for low grade and nonmuscle invasive stage. We used quantified methyl specific polymerase chain reaction to investigate promoter methylation for 8 tumor suppressor genes implicated in urothelial carcinogenesis. RESULTS: Tumors in the youngest age group had the lowest incidence of global hypermethylation compared to the other tumors with a methyl index of 37.5% vs 62.5% and 50%, respectively (ANOVA p = 0.009). When individual loci were analyzed, younger patients had a significantly lower rate and concentration of methylation at APC, Bcl2, MGMT and E-cadherin promoters than in the older groups (p <0.05). Few differences were present between the 2 older cohorts but the APC and MGMT methylation concentration increased with age. CONCLUSIONS: Urothelial tumors in patients younger than 19 years have a low rate of epigenetic alteration. Tumors in patients older than 20 years have epigenetic profiles similar to those of tumors in patients within the typical bladder urothelial cell carcinoma age range.

Expression of the lipid transporters ABCA3 and ABCA1 is diminished in human breast cancer tissue.

ATP-binding cassette transporters ABCA3 and ABCA1 are related to a differentiated, lipid-secreting phenotype of type II pneumocytes. Since mammary gland epithelial cells also show pronounced lipid metabolism and secretion, we investigated the expression of these proteins in normal as well as in neoplastic breast tissue. Normal human breast tissue, breast cancer cell lines, and 162 tumor samples of patients with primary unilateral invasive breast cancer were analyzed for ABCA3 and ABCA1 protein expression by immunohistochemistry using tissue microarrays. Strong ABCA3 and ABCA1 expression was found in the inner layer of normal mammary gland epithelium. Concurrent cytoplasmic ABCA3 and ABCA1 immunoreactivity was found in 9 of 11 breast cancer cell lines. ABCA3 and ABCA1 were shown to be differentially expressed in human breast cancer. Loss of ABCA3 staining was significantly associated with positive nodal status and negative progesterone receptor expression. In multivariate analysis, diminished ABCA3 expression proved to be a significant, independent and adverse risk factor for tumor recurrence. ABCA1 expression was associated with positive lymph nodes, but not significantly associated with tumor recurrence or breast cancer-specific survival. ABCA3 and ABCA1 are strongly expressed in normal mammary gland epithelium. Decreased ABCA3 expression in breast cancer seems to be associated with poor prognosis.

The number of lymph nodes in colon cancer specimens: what do the numbers really mean?

AIM: Stage-specific survival for colon cancer is improved when more lymph nodes are identified in the surgical specimen. This association is typically attributed to staging effect, but may instead be a surrogate for tumour biology. METHOD: We retrospectively studied a cohort of 48 consecutively treated patients with Stage II colon cancer who underwent complete resection between January 2000 and December 2002. Archived H&E slides were reviewed for lymphocytic infiltration at the leading edge, presence and degree of sinus histiocytosis in the largest node and the presence of lymph node hyperplasia. RESULTS: The mean number of lymph nodes identified was 14.1 +/- 9.4. T stage was strongly associated with the number of nodes identified (P = 0.01) and the presence of a significant degree of sinus histiocytosis approached statistical significance (P = 0.077). No statistically significant relationship existed between number of lymph nodes in a specimen and tumour location (P = 0.44), grade (P = 0.56) or lymphovascular invasion (P = 0.64). CONCLUSIONS: T stage is highly associated with the number of nodes found in a colon cancer specimen; a significant degree of sinus histiocytosis may also be predictive. Finding more nodes may be a surrogate for tumour or host-related factors that impact prognosis.

The molecular epidemiology of p53 gene mutations in human breast cancer.

The P53 tumor-suppressor gene is an advantageous tool for analyzing the molecular epidemiology of cancer. We describe the utility of the P53 gene as a 'mutagen test' and a prognostic indicator in breast cancer. Aspects of study design and methodology are discussed. Two major conclusions emerge: (1) there is an extraordinary diversity of mutational patterns among cohorts, hinting that the unique biology of mammary cells results in exposure to high doses of a diversity of ingested lipophilic mutagens; and (2) mutations in the P53 gene predict poor outcome in breast cancer.

Novel pattern of P53 mutation in breast cancers from Austrian women.

Since mutagens produce an extraordinary diversity of mutational patterns, differential mutational exposures among populations are expected to produce different patterns of mutation. Classical epidemiological methods have been successful in implicating specific mutagens in cancers such as those of lung and skin in which one mutagen predominates. In breast cancer, however, no mutagens have been implicated in an unequivocal manner. In an attempt to facilitate epidemiological studies, we have been studying the pattern of p53 gene mutations in breast cancers from multiple populations with high and low breast cancer incidences. We previously reported that breast cancers from Midwest United States, predominantly rural Caucasian women, have a different pattern of p53 gene mutation from populations of Western European women. Herein, we analyze patterns of p53 mutations from Graz, Austria, another population with a high incidence of breast cancer. Among the 60 Austrian breast cancers analyzed, 14 (23%) have a p53 gene mutation in exons 5-9 or in adjacent splice junctions. Analysis of the patterns of mutation shows differences between the "Western European" profile and the Austrian and Midwest United States groups (P = 0.027 and 0.024, respectively). The Austrian pattern is characterized by a high frequency of A:T-->T:A transversions (P = 0.006). The presence of distinct patterns of mutation among the limited number of analyzed populations of Western European origin supports the idea that differential mutagenic exposure and/or genetic differences contribute to breast cancer mutagenesis among geographically distinct Caucasians of Western European origin.

Pseudoactinomycotic radiate granules of the gynaecological tract: review of a diagnostic pitfall.

The filamentous bacterium actinomyces can cause serious gynaecological tract infections, including pelvic inflammatory disease (PID) and tubo-ovarian abscess. Thus, definitive diagnosis of actinomycotic granules (AMGs) in gynaecological specimens is clinically important. Non-infectious pseudoactinomycotic radiate granules (PAMRAGs) can mimic the microscopic appearance of AMGs. PAMRAGs may be more common than actinomycotic infections in specimens from patients using intrauterine devices and may be seen in patients with PID. Although the composition and aetiology of PAMRAGs is unclear and variable, a panel of histochemical stains can aid in diagnosis. On haematoxylin and eosin (H&E) stained sections, AMGs show as distinct granules with basophilic peripheral radiating filaments and a dense central eosinophilic core, whereas H&E stained sections of PAMRAGs feature refractile granules with irregular club-like peripheral projections and no central dense core. The filaments of AMGs are Gram positive on Brown and Brenn (B&B) stain and are highlighted with Gomori methenamine silver stain (GMS). They stain negatively with a modified acid fast bacillus (AFB) stain, aiding in the distinction of actinomyces from nocardia. PAMRAGs show negative or non-specific staining with B&B, GMS, and AFB stains. Therefore, knowledge of these staining properties and the distinguishing characteristics of PAMRAGs and AMGs enables recognition of this important diagnostic pitfall.

Evidence for oligoclonality and tumor spread by intraluminal seeding in multifocal urothelial carcinomas of the upper and lower urinary tract.

Multifocality and recurrence of urothelial carcinoma may result from either the field effect of carcinogens leading to oligoclonal tumors or monoclonal tumor spread. Previous molecular studies, favoring the monoclonality hypothesis, are mostly limited to the urinary bladder. We investigated genetic alterations in a total of 94 synchronous or metachronous multifocal tumors from 19 patients with at least one tumor both in the upper and lower urinary tract. Loss of heterozygosity (LOH) was determined using eight markers on chromosome 9 and one marker on 17p13 (p53). Microsatellite instability was investigated at six loci and protein expression of MSH2 and MLH1 was evaluated by immunohistochemistry. In addition, exons 5-9 of the p53 gene were sequenced. Deletions at chromosome 9 were found in 73% of tumors and at 17p13 in 18% of tumors. There was no significant difference in the frequency of LOH in the upper and lower urinary tract. Deletions at 9p21 were significantly correlated with invasive tumor growth. The pattern of deletion revealed monoclonality of all tumors in nine patients. In five patients there were at least two tumor clones with different genetic alterations. In four of these patients the different clones occurred in the bladder and subsequently in the ureter and renal pelvis. All four patients with p53 mutations revealed identical mutations in all tumors. Thus, multifocal urothelial carcinomas are frequently monoclonal, whereas others show oligoclonality, providing molecular evidence for field cancerization. Intraluminal tumor cell seeding appears to be an important mechanism of multifocal occurrence and recurrence of urothelial carcinomas.

p53 gene mutations inside and outside of exons 5-8: the patterns differ in breast and other cancers.

Most studies of mutations in the p53 tumor suppressor gene in tumors have examined only exons 5-8. Our laboratory previously found 64 mutations in exons 5-8 of the p53 gene in 194 primary breast cancers. Herein, we report 18 additional mutations found outside of exons 5-8. Mutations are present in exons 4, 9 and 10, and flanking splice junctions, but not in the promotor region or in exons 1, 2, 3 and 11. No missense mutations are found outside of exons 5-8. Instead, there is a predominance of frameshift mutations with lesser numbers of nonsense and splice site mutations. In contrast, the majority of mutations in exons 5-8 in this sample are missense changes and all of these are at amino acids that are identical in the 11 known p53 sequences that represent about 1.6 billion years of evolutionary divergence. The difference in mutational pattern between these two regions of the p53 gene is due to a lack of missense mutations and inframe microdeletions outside of exons 5-8. A review of our database of p53 mutations (De Vries et al., in preparation) shows that the patterns of mutation inside and outside of exons 5-8 differ in other types of cancers as well. The paucity of missense mutations in exons 2-4 and 9-11 in breast and other cancers (even at amino acids identical throughout p53 gene evolution) suggest that at least some missense mutations result in a phenotype other than malignant transformation. These data also illustrate the importance of examining identical exons when comparing the pattern of p53 gene mutations in different populations.

p53 gene mutations in breast cancers in midwestern US women: null as well as missense-type mutations are associated with poor prognosis.

We determined the pattern of mutations in exons 2-11 and adjacent intronic regions in breast cancers from Midwestern US white women. Twenty-one mutations were detected in 53 tumors (39.6%). Comparisons of the pattern of mutations within exons 5-9 showed that the frequency of missense mutations (44%) was lower in breast cancers of US Midwestern women than in most tumor types including breast cancers in other populations. Compared to breast cancers reported in a Scottish population, US women had a high frequency of G:C-->T:A transversions (P = 0.046). These findings suggest that environmental or endogenous factors contribute to p53 mutagenesis in mammary tissue to different extents among different populations. With a median follow-up of 19 months, the presence of a mutation was associated with shorter time to disease recurrence (P = 0.05) and shorter survival (P = 0.003). Putative dominant negative missense-type mutations (missense and in-frame microdeletions; P = 0.001) and null mutations (hemizygous nonsense and frameshift mutations; P = 0.007) were equally ominous. Thus, tumors with missense p53 mutations resulting in over-expression of a dysfunctional but otherwise intact protein have a clinical outcome similar to tumors with null mutations resulting in a truncated or garbled protein.

Molecular epidemiology of breast cancers in northern and southern Japan: the frequency, clustering, and patterns of p53 gene mutations differ among these two low-risk populations.

Comparison of acquired mutations in the p53 tumor suppressor gene can illuminate factors contributing to carcinogenesis among cancer cohorts. Japan has an ethnically homogeneous population with a low incidence of breast cancer. Previously we reported an unusual frequency, allelic status, and clustering of mutations in breast cancers from the northern part of the main Japanese island. To extend these findings, exons 2-11 and adjacent intronic sequences were analysed in tumors of women from northern (Hokkaido) and southern (Tokushima) Japan. The frequency of breast cancers with p53 gene mutations in the Hokkaido group is the highest reported (81%) while that in Tokushima (28%) is similar to most other populations. Thirteen of the 19 mutations (68.4%) in the Hokkaido cohort were heterozygous, an unusually high frequency for p53 mutations in any tumor type. There were three missense mutations at codon 175, a known hotspot for alterations in the p53 gene, and three missense mutations at codon 179, a rare site for p53 changes. In addition, the patterns of p53 gene mutation differed between the two Japanese cohorts (P=0.04). The multiple differences in acquired p53 mutations suggest unsuspected biological differences among breast cancers in northern and southern Japan. In addition, the high frequency of p53 mutations in breast cancers from Hokkaido predicted a poorer prognosis for this population which was confirmed on examination of mortality data.

Indiscriminate loss of myenteric neurones in the TNBS-inflamed guinea-pig distal colon.

This investigation was conducted to establish whether guinea-pig trinitrobenzene sulfonic acid (TNBS)-colitis was associated with a change in the number of neurones of the myenteric plexus, and, if so, whether select subpopulations of neurones were affected. Total neurones were quantified with human (Hu) antiserum, and subpopulations were evaluated with antisera directed against choline acetyltransferase, nitric oxide synthase, calretinin, neuronal nuclear protein or vasoactive intestinal peptide (VIP). Colitis was associated with a loss of 20% of the myenteric neurones, most of which occurred during the first 12 h past-TNBS administration. During this period, myenteric ganglia were infiltrated with neutrophils while lymphocytes appeared at a later time-point. The neuronal loss persisted at a 56-day time-point, when inflammation had resolved. The decrease in myenteric neurones was not associated with a decrease in any given subpopulation of neurones, but the proportion of VIP-immunoreactive neurones increased 6 days following TNBS administration and returned to the control range at the 56 days. These findings indicate that there is an indiscriminant loss of myenteric neurones that occurs during the onset of TNBS-colitis, and the loss of neurones may be associated with the appearance of neutrophils in the region.

Rapid and efficient screening for p53 gene mutations by dideoxy fingerprinting.

Dideoxy fingerprinting (ddF) is an efficient method for detecting single base and other sequence changes in PCR-amplified DNA segments. This screening method is a hybrid between single-strand conformation polymorphism analysis (SSCP) and Sanger dideoxy sequencing. It involves a Sanger sequencing reaction with one dideoxynucleotide followed by non-denaturing gel electrophoresis. We are using ddF to screen for mutations in the p53 tumor suppressor gene in primary breast cancers. ddF detected more than 100 mutations in different regions of the gene, including all types of single-base mutations and microdeletions/microinsertions of various sizes. Furthermore, ddF reliably detected heterozygous mutations, if the region of interest was screened in both directions. In a blinded, prospective study, ddF detected all 25 mutations within exons 4-10 and adjacent flanking intronic regions previously found by direct sequencing. ddF was also useful in scoring two common polymorphisms within the p53 gene. Guidelines for preventing false-positive and false-negative results are summarized.

Persistent proliferation of normal hepatocytes and promotion of preneoplastic development by N-nitrosodibenzylamine in rats.

In a traditional long-term study N-nitrosodibenzylamine (NDBzA) was proven to be noncarcinogenic, but recently the substance was found to produce genotoxic lesions in hepatocytes. Our own experiments have shown that relatively low single doses of NDBzA cause liver hypertrophy and additive proliferation of hepatocytes in rats. Both effects are known from well-documented promoters and non-genotoxic carcinogens, respectively, in rodents. Investigation of NDBzA in an initiation-promotion assay (IP assay) showed it to cause an increase in the number and size of preneoplastic liver cell foci. This occurred only after initiation with diethylnitrosamine, but not when 2-acetylaminofluorene was used. Another property of NDBzA is its sustained mitotic stimulation of extrafocal hepatocytes. This is inconsistent with their adaptive loss of susceptibility to mitogens in IP assays using other promoters of hepatocarcinogenesis. The following conclusions can be drawn. First, "differential inhibition" of the proliferation of extrafocal hepatocytes, in contrast to the selective mitostimulation of preneoplastic cells, is obviously no prerequisite for cancer development. Second, primary mitogenicity of a compound in short-term studies can be a useful indicator for tumorigenic potential. In the case of NDBzA the data available at present are still insufficient to classify it unequivocally in terms of genotoxic or nongenotoxic carcinogenicity.

Acute aortic regurgitation due to spontaneous rupture of a fenestrated cusp: report in a 65-year-old man and review of seven additional cases.

A 65-year-old man with chronic hypertension developed dyspnea, a cough, and a new diastolic murmur. Two-dimensional echocardiography showed severe aortic regurgitation. No valvular vegetations were identified and blood cultures were negative. Surgical intervention was recommended, but the patient died of an acute intracranial hemorrhage two weeks later. At autopsy, the posterior aortic cusp was flail, due to rupture of the residual cord above two large fenestrations. There was no acute or healed endocarditis. To our knowledge, this is the eighth reported case of aortic valve incompetence due to spontaneous rupture of a fenestrated cusp. Patients ranged in age from 31-67 years (mean, 54), and 4 (50%) were older than 60 years. Seven (88%) of the 8 were men, and 4 (57%) of 7 had chronic hypertension. Analogously, in another four reported cases, aortic insufficiency developed following spontaneous rupture of the fenestrated raphe of an atypical congenitally bicuspid aortic valve. Noninfective and nontraumatic rupture of cord-like aortic valve structures may result in severe acute aortic regurgitation, particularly in men with chronic hypertension.

Death from pulmonary thromboembolism in severe obesity: lack of association with established genetic and clinical risk factors.

Several clinical and environmental conditions are causally related to sudden death from acute pulmonary thromboembolism (APT). Morbid obesity, despite its frequency and association with adverse health effects, is usually considered at most only an additive risk factor for APT. We reviewed protocols and histories from 7227 consecutive autopsies performed between 1985 and 1996 at the Mayo Clinic, including all deaths from APT where no clinical or environmental risk factor could be identified in the study. Body mass indices (BMI) were calculated and compared with those of age- and sex-matched controls who had died suddenly and naturally without evidence of APT. Resistance to activated protein C is the most common molecular clotting defect predisposing to APT, and it is caused by a point mutation in the factor V gene (R506Q). Genomic DNA was extracted from archival tissues of all cases and controls, and the R506Q status was determined by polymerase chain reaction amplification, restriction endonuclease digestion, and direct sequencing. APT was found as the immediate cause of death in 433 patients, with 36 (8%) having no previously established risk factors. Twenty-four of these persons (67%) were morbidly obese (BMI >30 kg/m2). compared with only five controls (14%, P<0.0001). Four patients in both groups, each with a BMI <30 kg/m2. had at least one allele positive for R506Q. Morbid obesity is an independent risk factor in cases of sudden death from APT after the exclusion of previously established clinical, environmental, and molecular risk factors.

Factor V leiden and morbid obesity in fatal postoperative pulmonary embolism.

HYPOTHESIS: Currently, the risk for postoperative acute pulmonary embolism (APE) is assessed clinically. We hypothesize that the expensive screening for the most common genetic thrombophilic clotting defect (factor V Leiden; R(506)Q) after exclusion of established clinical risk factors does not offer additional benefit to surgical patients. DESIGN: We reviewed protocols and histories from 8249 consecutive autopsies performed at the Mayo Clinic, Rochester, Minn. All patients who died of APE after routine surgery and who lacked any other clinical risk factors for APE were included and compared with matched controls. Genomic DNA was extracted from archival tissues and examined for R(506)Q by polymerase chain reaction amplification, restriction enzyme digestion, and direct sequencing. RESULTS: Acute pulmonary embolism was the immediate cause of death in 454 patients (5.5%). Of those, 32 (7.0%) had undergone routine surgery. These patients represent less than 0.07% of all case-adjusted surgical procedures in the same period. The rate of postoperative death from APE was higher after neurosurgical procedures (0.3%) than all other procedures (0.04%). Sixteen patients (50.0%) were morbidly obese. Only 1 patient was heterozygous and none were homozygous for R(506)Q. CONCLUSIONS: (1) Fatal APE is uncommon in surgical patients lacking clinically apparent risk factors for venous thromboembolism. (2) Neurosurgical patients are at increased risk for postoperative APE. (3) Morbid obesity is a major independent risk factor in cases of sudden death from APE postoperatively. (4) Routine preoperative screening for R(506)Q in the factor V gene does not appear to offer additional benefit in surgical patients without clinically recognizable thromboembolic risk factor(s).

Mitogenic short-term effects on hepatocytes and adrenocortical cells: phenobarbital and reserpine compared to carcinogenic and non-carcinogenic fluorene derivatives.

The adrenal cortex has a low physiologic cell renewal and shows only a moderate cell replication even after contralateral adrenalectomy. Although rather unsusceptible to the malignancy-inducing action of carcinogens, a single oral dose of various tumorigenic xenobiotics induced an additive mitotic response of adrenocortical cells studied after 48 h. Presently we report on three different response patterns in rats. First, a selective mitostimulation of the zona glomerulosa occured after reserpine associated with a loss of body weight, thymus and liver weight. These are unspecific stress effects and occur also after exogenous ACTH. Second, hepatomitogenic and liver-enlarging congeners, e.g. fluorene (FEN), fluorenone (FON) and 4-benzoyl-FON, but also the genotoxic 2-acetylaminofluorene (2-AAF) and 2,4,7-trinitro-FON induced a selective mitotic response of the zona fasciculata (ZF). After the lowest effective dose of FEN or FON the afore-mentioned effects occured simultaneously, but were absent in the high dose group (only studied with fluorene). The 2-benzyl and 2-benzoyl-substituted derivatives were ineffective at all. Third, a bizonal response was found only after phenobarbital (PB) or the lowest effective FEN dose. The preventive action of a low PB dose on the 2-AAF-induced ZF response indicates a modified metabolism. We conclude that the rapid mitotic ZF response is an endogenously mediated net effect of interactions between metabolic and various adaptive mechanisms. The latter are reported to be activated in a stressor-dependent manner and converge in the adrenals. In this way the early mitotic ZF response could reflect indirectly 'specific' proliferation-prone properties of xenobiotics.

Mutational activation of FGFR3: no involvement in the development of renal cell carcinoma.

BACKGROUND: Somatic point mutations in the fibroblast growth factor receptor 3 (FGFR3) gene have been identified in certain types of urological cancers, especially urothelial carcinoma of the bladder and the renal pelvis, and could be correlated with a favourable outcome. However, comprehensive data on the FGFR3 mutation status in renal cell carcinoma (RCC) are still missing. METHODS: In order to investigate a possible role for FGFR3 mutations in renal cell carcinogenesis, we performed a sequence-based mutational analysis of FGFR3 in 238 primary RCC. The cohort obtained the common RCC subtypes including 101 clear cell, 50 papillary and 68 chromophobe RCC specimens. The analysed regions encompassed all FGFR3 point mutations previously described in epithelial tumours and other noncutaneous epithelial malignancies. RESULTS: No mutations were detected in any renal tumour type examined, and all cases showed wild-type sequence. CONCLUSION: Our results argue against an involvement of mutational activation of FGFR3 in the development of RCC. A recently described cystic renal dysplasia in a patient with thanatophoric dysplasia type 1 due to a germ line FGFR3 mutation might portend to an involvement of mutational FGFR3 activation in renal cyst formation, but this speculation needs further evaluation.

Value of multicolour fluorescence in situ hybridisation (UroVysion) in the differential diagnosis of flat urothelial lesions.

AIMS: During the past 10 years, multitarget fluorescence in situ hybridisation has been established as a valuable adjunct in the cytological diagnosis of precancerous and malignant lesions of the urinary tract. The aim of the present study was to define its value in detecting chromosomal imbalances in patients with various flat urothelial lesions in routine paraffin-embedded bladder biopsy samples. In addition, the HER2 gene amplification and HER2 expression pattern were examined, since alterations of the HER2 expression patterns have been demonstrated in invasive bladder cancer. METHODS: 29 samples of normal urothelium and 86 flat urothelial lesions (hyperplasia, reactive atypia, dysplasia and carcinoma in situ (CIS)) from 73 patients were analysed patients using tissue microarrays and centromeric probes for chromosomes 3, 7 and 17, and gene-specific probes for 9p21/P16 and HER2 (UroVysion, PathVysion). The expression of HER2 was studied by immunohistochemistry. RESULTS: Polysomy of at least one of the chromosomes was found in about half of the dysplastic cells, and in more than 90% of cells in CIS or cells in invasive bladder tumours. Polysomic cells were found in only 17% of urothelial hyperplasia, reactive atypia and normal urothelium of healthy patients, whereas about 30% of non-neoplastic lesions in patients with concomitant urothelial carcinoma showed polysomy of at least one chromosome. These alterations indicate a field effect and are associated with synchronous development of dysplastic lesions of a higher grade. Deletion of the P16 locus was most frequently observed in aneuploid lesions, whereas overexpression of HER2 was found in 10-20% of invasive urothelial carcinomas, and only occasionally in CIS (5%). An altered HER2 expression pattern was present in non-neoplastic lesions (25%). CONCLUSIONS: UroVysion fluorescence in situ hybridisation is a valuable tool for the detection of genetically unstable flat urothelial lesions, and can help to resolve difficult cases, particularly the differential diagnosis of reactive atypia and dysplasia.

Genomic aberrations are rare in urothelial neoplasms of patients 19 years or younger.

Urothelial neoplasms in patients 19 years of age or younger are rare, and the data regarding clinical outcome are conflicting. Molecular data are not available. Urothelial tumours from 14 patients aged 4 to 19 years were analysed, including FGFR3 and TP53 mutation screening, comparative genomic hybridization (CGH), UroVysion FISH analysis, polymerase chain reaction for human papillomavirus (HPV), microsatellite analysis using the NIH consensus panel for detection of microsatellite instability (MSI) and six markers for loss of heterozygosity on chromosome arms 9p, 9q, and 17p and immunohistochemistry for TP53, Ki-67, CK20 and the mismatch repair proteins (MRPs) hMSH2, hMLH1, and hMSH6. Based on the 2004 WHO classification, one urothelial papilloma, seven papillary urothelial neoplasms of low malignant potential (PUNLMPs), five low-grade, and one high-grade papillary urothelial carcinoma were included. No multifocal tumours were found and recurrence was seen in only one patient with a urothelial papilloma. All patients were alive with no evidence of disease at a median follow-up of 3.0 years. We found no mutations in FGFR3, deletions of chromosome arms 9p, 9q or 17p, MSI or MRP loss, or HPV positivity in any of the patients. Three cases showed chromosome alterations in CGH analyses, urothelial dedifferentiation with CK20 overexpression, or aneuploidy, and one TP53 mutation with TP53 overexpression was found. Urothelial neoplasms in people younger than 20 years are predominantly low grade and are associated with a favourable clinical outcome. Genetic alterations frequently seen in older adults are extremely rare in young patients. Urothelial neoplasms in children and young adults appear to be biologically distinct and lack genetic instability in most cases.