Analysis of genetic events in 17p13 and 9p21 regions supports predominant monoclonal origin of multifocal and recurrent bladder cancer

Summary

Clonality was tested in 86 tumours from 25 patients with recurrent and multifocal superficial bladder transitional cell carcinomas (TCCs) using the analysis of TP53 mutations and of LOH in the 17p13 and 9p21 regions. Tumours from the majority of individuals showed either absence or presence of the same TP53 mutation and/or an identical LOH pattern, with the same allele lost in all tumours. Only two pairs of tumours from two patients had discordant findings, which were incompatible with monoclonality. Therefore, our results rather support the monoclonal model of development of highly recurrent superficial bladder TCCs.

Introduction

Bladder cancer is the second most common malignancy of the genitourinary tract with a male to female ratio of 3:1 [1]. About 90% of all bladder cancers are transitional cell carcinomas (TCCs) derived from the urothelium, and more than 80% of them are superficial, non-muscle-invasive tumours (classified as Ta, T1 or Tis). Approximately 70% of all patients with superficial bladder TCCs develop recurrences after transurethral resection (TUR), and in 10–20% of patients the progression to muscle invasion (T2–T4) occurs [2]. Nearly 30% of all patients present at diagnosis with multifocal disease—simultaneous occurrence of several spatially distinct tumours at different sites of the bladder wall [3].

Simultaneous or metachronous development of multiple superficial bladder TCCs evokes the question of possible monoclonal nature of these tumours. During the last years, two hypotheses have been proposed: the monoclonal hypothesis, and the field cancerization hypothesis. According to the monoclonal model, the progeny of a single malignant cell proliferates and spreads throughout the urothelium either via intraluminal seeding (when the transformed cell is fully released from the primary tumour), or via intraepithelial migration of the malignant cell. On the contrary, the field cancerization model proposes independent transformation of numerous urothelial cells at multiple sites as a result of accumulation of carcinogenic events, leading to the growth of multiple unrelated tumours [4].

Various molecular genetic methods or their combinations can be used to determine the clonality of multiple synchronous or recurrent tumours in one patient including X-chromosome inactivation analysis, molecular cytogenetic techniques (FISH, CGH), loss of heterozygosity (LOH) analysis, and single-gene mutation analysis [5], [6], [7], [8], [9], [10], [11]. Several studies published up to now presented evidence both for and against the monoclonal model of bladder cancer, but the majority of molecular studies favoured the monoclonal origin of multiple tumours in one patient. Most of these studies concentrated on advanced-stage invasive carcinomas [5], [6], [7], [8]. On the other hand, several studies, which found evidence for the existence of more than one tumour clone, particularly in the early stages of bladder carcinoma, supported the field cancerization hypothesis [9], [10], [11].

We attempted to address this question by the analysis of genetic events in the 17p13 and 9p21 regions of the human genome, harbouring the TP53 and CDKN2A genes known to play a role in bladder tumorigenesis [12], in a large series of well characterized multifocal and recurrent superficial bladder TCCs. TP53 gene mutations and LOH at several TP53 and CDKN2A intragenic and extragenic polymorphic DNA markers were analysed in a total of 86 tumours from 25 patients sampled during a period of 5 years. The patients were selected for highly recurrent disease, and the study was particularly focused on the analysis of the genetic profile of the early stages of the disease. Our results support the monoclonal model of development of multifocal and recurrent superficial bladder TCCs.