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. 2021 Dec 15;9(6):416-434.
eCollection 2021.

Patient derived models of bladder cancer enrich the signal of the tumor cell transcriptome facilitating the analysis of the tumor cell compartment

Michalis Mastri  1 , Swathi Ramakrishnan  2 , Shruti D Shah  2 , Ellen Karasik  2 , Bryan M Gillard  2 , Michael T Moser  2 , Bailey K Farmer  3 , Gissou Azabdaftari  4 , Gurkamal S Chatta  3 , Anna Woloszynska  2 , Kevin H Eng  1   5 , Barbara A Foster  2 , Wendy J Huss  6   7
Affiliations

Patient derived models of bladder cancer enrich the signal of the tumor cell transcriptome facilitating the analysis of the tumor cell compartment

Michalis Mastri et al. Am J Clin Exp Urol. .

Abstract

The evolving paradigm of the molecular classification of bladder cancer requires models that represent the classifications with less heterogeneity. Robust transcriptome based molecular classifications are essential to address tumor heterogeneity. Patient derived models (PDMs) are a powerful preclinical tool to study specific tumor compartments. We tested if the consensus molecular subtype analysis was applicable to PDMs and evaluated the tumor compartment each model represents. PDMs derived from surgical specimens were established as xenografts (PDX), organoids (PDO), and spheroids (PDS). The surgical specimens and PDMs were molecularly characterized by RNA sequencing. PDMs that were established in immune deficient mice or in vitro significantly downregulated transcripts related to the immune and stromal compartments compared to the surgical specimens. However, PDMs upregulate a patient-specific bladder cancer cell signal which allowed for analysis of cancer cell pathways independent of the tumor microenvironment. Based on transcriptomic signatures, PDMs are more similar to their surgical specimen than the model type; indicating that the PDMs retained unique features of the tumor from which the PDM was derived. When comparing models, PDX models were the most similar to the surgical specimen, while PDO and PDS models were most similar to each other. When the consensus molecular subtype classification system was applied to both the surgical samples and the three PDMs, good concordance was found between all samples indicating that this system of classification can be applied to PDO and PDS models. PDMs reduce tumor heterogeneity and allow analysis of tumor cells while maintaining the gene expression profile representative of the original tumor.

Keywords: Bladder cancer; epithelial to mesenchymal transition (EMT); organoid; patient derived models; spheroid; xenograft.

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Conflict of interest statement

None.

Figures

Figure 1
Figure 1
PDMs are a faithful representation of their original tumor. (A-D) Euclidean distance was visualized with t-SNE for (A) all samples, (B) surgical and PDX samples, (C) only PDM samples, and (D) PDO and PDS samples. (E) Euclidean distance of each PDM to their corresponding surgical specimen. (F) Heatmap of differentially expressed genes identified by paired comparison of PDM compared to their corresponding surgical specimen.
Figure 2
Figure 2
PDMs represent the tumor but not the tumor microenvironment. A. Volcano plots of differentially expressed genes comparing surgical specimens to PDM emphasizing the selected genes for tumor and tumor microenvironment. B. Heatmap of gene expression corresponding to tumor, bladder cancer stem cell (BSC), stem cell (SC), immune, and stroma genes.
Figure 3
Figure 3
Hallmark pathway analysis RNAseq. (A) Gene set enrichment analysis for all the hallmark gene sets for 3 different comparisons (SURG vs. PDM, PDX vs. 3D, PDS vs. PDO). Gene sets were categorized based on processes. (B-D) Enrichment of processes based on fisher exact test for (B) SURG vs. PDM, (C) PDX vs. 3D, and (D) PDS vs. PDO.
Figure 4
Figure 4
EMT and Ba/Sq molecular subtypes are enriched in PDMs. (A) Gene set enrichment analysis for the hallmark epithelial to mesenchymal transition gene set for 3 different comparisons (PDM vs. SURG, PDX vs. 3D, PDS vs. PDO). (B, C) Expression of the hallmark epithelial to mesenchymal transition genes were used for (B) t-SNE plot and (C) heatmap. (D) Correlation heatmap based on the consensus molecular subtype scores for each sample derived from RNA sequencing data.
Figure 5
Figure 5
Pathology and differentiation marker analysis of PDMs with conflicting phenotypes. H&E staining of surgical specimens and PDX models derived from the surgical specimens PDX models RP-BL005, -003, and 022. IHC analysis of PDX models for E-Cadherin, CK5, CK20, Vimentin, and Synaptophysin. Slides were imaged at 20×, Scale Bar =100 µm
Figure 6
Figure 6
Pathology and differentiation marker analysis of PDMs. H&E staining of surgical specimens and PDX models derived from the surgical specimens PDX models RP-BL0051, 0050, 040, 054 and 019. IHC analysis of PDX models for E-Cadherin, CK5, CK20, and Vimentin. Slides were imaged at 20×, Scale Bar =100 µm.
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