Elsevier

European Urology

Volume 72, Issue 4, October 2017, Pages 641-649
European Urology

Urothelial Cancer
Comprehensive Genomic Characterization of Upper Tract Urothelial Carcinoma

https://doi.org/10.1016/j.eururo.2017.05.048Get rights and content

Abstract

Background

Upper urinary tract urothelial cancer (UTUC) may have unique etiologic and genomic factors compared to bladder cancer.

Objective

To characterize the genomic landscape of UTUC and provide insights into its biology using comprehensive integrated genomic analyses.

Design, setting, and participants

We collected 31 untreated snap-frozen UTUC samples from two institutions and carried out whole-exome sequencing (WES) of DNA, RNA sequencing (RNAseq), and protein analysis.

Outcome measurements and statistical analysis

Adjusting for batch effects, consensus mutation calls from independent pipelines identified DNA mutations, gene expression clusters using unsupervised consensus hierarchical clustering (UCHC), and protein expression levels that were correlated with relevant clinical variables, The Cancer Genome Atlas, and other published data.

Results and limitations

WES identified mutations in FGFR3 (74.1%; 92% low-grade, 60% high-grade), KMT2D (44.4%), PIK3CA (25.9%), and TP53 (22.2%). APOBEC and CpG were the most common mutational signatures. UCHC of RNAseq data segregated samples into four molecular subtypes with the following characteristics. Cluster 1: no PIK3CA mutations, nonsmokers, high-grade <pT2 tumors, high recurrences. Cluster 2: 100% FGFR3 mutations, low-grade tumors, tobacco use, noninvasive disease, no bladder recurrences. Cluster 3: 100% FGFR3 mutations, 71% PIK3CA, no TP53 mutations, five bladder recurrences, tobacco use, tumors all <pT2. Cluster 4: KMT2D (62.5%), FGFR3 (50%), TP53 (50%) mutations, no PIK3CA mutations, high-grade pT2+ disease, tobacco use, carcinoma in situ, shorter survival. We identified a novel SH3KBP1-CNTNAP5 fusion.

Conclusions

Mutations in UTUC occur at differing frequencies from bladder cancer, with four unique molecular and clinical subtypes. A novel SH3KBP1 fusion regulates RTK signaling. Further studies are needed to validate the described subtypes, explore their responses to therapy, and better define the novel fusion mutation.

Patient summary

We conducted a comprehensive study of the genetics of upper urinary tract urothelial cancer by evaluating DNA, RNA and protein expression in 31 tumors. We identified four molecular subtypes with distinct behaviors. Future studies will determine if these subtypes appear to have different responses to treatments.

Introduction

Upper tract urothelial carcinoma (UTUC) is histologically similar to urothelial bladder cancer yet several clinical, biological and molecular features are unique to UTUC, prompting the term disparate twins when considering the similarities and differences between bladder urothelial cancer and UTUC [1]. Major knowledge gaps remain in our understanding of the biology and genomic landscape of UTUC, a rare disease in Western countries but of potentially epidemic proportions in the Far East [2]. Biologically interesting features along the environmental-genetic spectrum include the strong association between known exposure to agents such as tobacco [3] and aristolochic acid [2] and genetic predisposition in patients with Lynch syndrome [4].

The Cancer Genome Atlas (TGCA) reported the mutation landscape in muscle-invasive bladder cancer, which has a high somatic mutation frequency among adult solid tumors, similar to melanoma and lung adenomas and squamous carcinomas [5]. The most common mutation was TP53, with frequent alterations in chromatin modifier genes (MLL2, ARID1A, KDM6A) [6], [7]. Four expression-based subtypes were described, and were shown to be associated with overall survival and response to immune checkpoint inhibition and potentially cisplatin-based chemotherapy [8], [9], [10].

The largest targeted genomic study of UTUC to date evaluated 300 cancer-associated genes in 83 patients using next-generation sequencing [11]. Mutations of FGFR3, CREBBP, and STAG2 were commonly found in low-grade tumors, while TP53 mutations were more common in high-grade tumors. FGFR3 mutations were observed at a similar rate in high- and low-grade tumors [6].

In this study we report the first integrated comprehensive genomic analysis of UTUC using whole exome sequencing (WES), gene expression profiling, and protein expression analysis to further characterize the genomic landscape of UTUC and provide deeper insights into the biology of this rare cancer.

Section snippets

Materials and methods

UTUC samples were obtained from 31 patients under protocols approved by institutional review boards using endoscopic biopsy or surgical resection, and were stored frozen at −80 °C. Ten samples were primary ureter and 21 were renal pelvis in origin. Histology slides were reviewed by genitourinary pathology experts at each respective institution (M.I., C.G.). All tumors were composed of conventional urothelial carcinoma, and no variant histology was present. Microdissection was not performed, as

Patient demographic and clinical data

Patient demographic and clinical data are shown in Table 1. The male/female ratio of 2:1 is similar to that in previous reports for UTUC [21]. The majority of patients were white and former or current smokers. The majority had high-grade tumors and 32.3% had muscle-invasive or higher-stage disease (pT2+). Recurrences that were local, distant, or in the bladder were detected in approximately half of patients during median follow-up of 20 mo (range 3–66) for living patients. Median overall

Discussion

This is the first comprehensive, integrated genomic analysis of UTUC. Procurement of sufficient banked samples of this rare disease required pooling from two different institutions, and harmonization of clinical and genomic data. From the merged WES data set we identified 2784 mutations, with FGFR3 the most commonly mutated gene (74%) in both low-grade (92%) and high-grade (60%) tumors. The relatively high rate of FGFR3 mutations in high-grade tumors is nearly double the rate previously

Conclusions

Comprehensive genomic characterization of UTUC revealed novel mutations and mutation frequencies in comparison to bladder cancer, and identified four expression subtypes with unique molecular profiles and clinical correlates. WES analyses demonstrated that DNA repair and chromatin-modifying genes play a critical role in this disease. Expression analyses confirmed the additional critical role of FGFR3 in both low- and high-grade tumors, and validate this as a rational therapeutic target, along

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