Intrinsic subtypes of high-grade bladder cancer reflect the hallmarks of breast cancer biology

Abstract

We sought to define whether there are intrinsic molecular subtypes of high-grade bladder cancer. Consensus clustering performed on gene expression data from a meta-dataset of high-grade, muscle-invasive bladder tumors identified two intrinsic, molecular subsets of high-grade bladder cancer, termed "luminal" and "basal-like," which have characteristics of different stages of urothelial differentiation, reflect the luminal and basal-like molecular subtypes of breast cancer, and have clinically meaningful differences in outcome. A gene set predictor, bladder cancer analysis of subtypes by gene expression (BASE47) was defined by prediction analysis of microarrays (PAM) and accurately classifies the subtypes. Our data demonstrate that there are at least two molecularly and clinically distinct subtypes of high-grade bladder cancer and validate the BASE47 as a subtype predictor. Future studies exploring the predictive value of the BASE47 subtypes for standard of care bladder cancer therapies, as well as novel subtype-specific therapies, will be of interest.

Fig. 1.

Consensus clustering defines two distinct molecular subtypes of MIBC. (A) Consensus clustering was performed on 262 muscle-invasive tumors curated from four publicly available datasets (meta-dataset), yielding two subtypes. (B) Consensus clustering was performed independently on a dataset of high-grade bladder tumors obtained from MSKCC (n = 49). (C) The median gene expression of all common genes between the datasets were compared, and the Pearson correlation was plotted (yellow, correlation; blue, anticorrelation). Numerical values represent the Pearson correlation. (D) Gene expression of epithelial and urothelial markers were visualized by heatmap, supervised by ConsensusClusterPlus calls in the meta-dataset. (E) Significantly differentially expressed genes between K1 and K2 from the meta-dataset and their respective fold changes, as determined by two-class SAM (2,393 genes; FDR, 0) were analyzed for predicted pathway enrichment by IPA. Selected significant pathways enriched in K1 are shown.

Fig. 2.

Generation of the BASE47 subtype predictor. PAM was performed using the basal-like and luminal subtype calls generated by ConsensusClusterPlus. A predictor consisting of 47 genes was generated that accurately predicted the subtypes from the meta-dataset training set (P < 0.001), as well as a MSKCC validation dataset (45 of 47 genes present; P < 0.001).

Fig. 3.

Luminal and basal-like bladder cancer have differential survival and are associated with distinct genomic alterations. (A and B) Kaplan–Meier plots for muscle-invasive tumors from the MSKCC dataset (≥pT2) were generated for disease-specific survival (A) and overall survival (B). Basal-like tumors (n = 22) had a significantly decreased disease-free and overall survival compared with luminal tumors (n = 16) (P = 0.0194 and 0.0081, respectively). (C) Superficial tumors not included in the generation of BASE47 were subjected to BASE47 subtype prediction. (D) Sequencing and copy number analysis was performed on common mutations in bladder cancer. FGFR3 and TSC1 alterations were significantly enriched in luminal bladder cancer, whereas alterations of the RB1 pathway were enriched in basal-like bladder cancer. TP53 alterations were distributed evenly in both subtypes. (E) Basal-like and luminal tumors from the meta-dataset were annotated for sex (two of four datasets). The prevalence of basal-like bladder cancer was significantly greater in female patients (P value = 0.0203, χ² test).

Fig. 4.

Basal-like and luminal bladder cancers are correlated with the intrinsic molecular subtypes of breast cancer. (A) Median gene expression of genes present in the breast cancer-specific intrinsic gene list were determined for each bladder and breast subtype, and 1-Pearson correlation was calculated comparing bladder subtypes (bladder tumors) with breast subtypes (breast tumors) in both the TCGA and UNC337 breast datasets. (B) The meta-dataset tumors were run against previously published breast cancer-related gene sets. The resulting pathway scores were clustered by hierarchical clustering, and heatmaps were generated for visualization.

Fig. 5.

A subset of basal-like bladder tumors are claudin-low. The meta-dataset was clustered hierarchically using representative genes known to define claudin-low breast tumors. Claudin-low subtype designation was assigned using a previously defined 807-gene signature.

Fig. 6.

Proposed model of urothelial tumorigenesis and relationships with intrinsic subtypes of breast cancer. (A) Low-grade (LG) and high-grade (HG) UCs are associated with specific genetic alterations. Low-grade papillary tumors often incur FGFR3, RAS, and receptor tyrosine kinase alterations, whereas high-grade tumors are characterized by loss of tumor-suppressor genes, such as PTEN, TP53, and RB1 pathway alterations. Whereas most low-grade tumors recur as low-grade, a small proportion progress to high-grade tumors in association with PTEN, TP53, and RB1 pathway alterations. We propose that low-grade tumors that progress are likely to be high-grade papillary tumors of the luminal molecular subtype, and that de novo high-grade tumors are likely to be of the basal-like expression subtype. Whether luminal, nonpapillary tumors arise from low-grade tumors is unclear. (B) Diagram showing the proposed relationship between intrinsic subtypes of breast and bladder cancers.