Presentation Authors: Bernhard Kiss*, Aaron Kershner, Stanford, CA, Lolita Penland, San Francisco, CA, Edward Diaz, Joseph Liao, Philip Beachy, Stanford, CA
Introduction: Invasive bladder cancer has been analyzed extensively using various &[prime]bulk&[prime] genomic and transcriptomic approaches, such as mRNA sequencing. While these studies have been useful for elucidation of bladder cancer molecular subtypes, these bulk approaches have the inherent disadvantage that non-tumor (i.e., non-urothelial) cell types may contaminate sample preparation. Indeed, contaminating, non-tumor cell types have caused mischaracterization of bladder cancer subtypes. To avoid this problem associated with bulk sequencing approaches, we sought to characterize bladder transcriptome at single cell resolution, in both disease-free and tumor-bearing bladders.
Methods: Bladder specimen (n=12) were obtained during TURBT or radical cystectomy from chemo naÃ¯ve human bladders, and samples were isolated from the tumor as well as from normal-looking tissue located at sites distant from the tumor. Other bladder samples (n=4) were isolated from tumor-free bladders in otherwise healthy donors. Bladder mucosa was minced, enzymatically dissociated into single cell suspensions, and then subjected to single cell sequencing using the emulsion-based 10X Genomics platform and analysis using the Seurat single cell mRNA sequencing analysis pipeline.
Results: The predominant cell types in our samples included epithelial, mesenchymal, and immune lineages. Whereas tumor-free bladder samples contained few KRT14+ UPK3A low basal epithelial cells, invasive bladder tumors contained much more KRT14+ UPK3A low cells. In addition, the tumor-free bladder derived basal cells uniformly expressed SHH, but SHH was lost in invasive cancer, consistent with the known loss of SHH in aggressive invasive cancer. Interestingly, normal-looking tissue from invasive tumor-bearing bladders also contained a population of KRT14+ UPK3A low cells lacking SHH, suggesting that these sites distant from the tumor contained aggressively growing basal cells similar in molecular phenotype to the invasive tumor. Non-invasive tumor epithelial cells expressed UPK3A and also retained SHH expression.
Conclusions: Bladder mucosa contains a variety of cell types. The population of bladder epithelial cells in normal looking tissue from cancer-bearing bladder is substantially different from tissue from disease free bladder, containing cells with molecular characteristics typical of tumors.
Source of Funding: BK was funded by Swiss National Foundation(P300PB 167793/1) and Bern Cancer League.Sequencing data was obtained with the support of the Chen Zuckerberg Initiative and the Biohub