Abstract A17: Gene expression changes associated with metastasis in mice with bladder cancer
Authors: Tham Sin Mun , Fung Stefanie Alicia , Esuvaranathan Kesavan , Mahendran Ratha 
Published: August 2020

Background: Prognosis for bladder cancer is dependent on both tumor stage and grade. In patients with non-muscle invasive bladder cancer, the tumor invariably recurs and may progress. Patients who subsequently develop metastatic disease or present with it have a worse prognosis. The purpose of this study was to detect and find differences in patterns of gene expression between the primary tumor and common metastatic sites in a mouse model of bladder cancer to identify possible triggering events that result in metastasis.

Methods: Five- to six-week-old female C57BL/6 mice were orthotopically implanted with MB49-PSA (murine bladder cancer cells secreting human prostate-specific antigen as a tumor marker). The institutional animal care and use committee approved the use of mice. At termination (up to 78 days after tumor implantation), bladders, kidneys, lymph nodes, and lungs were snap frozen and RNA was extracted by homogenizing the tissue in Trizol. Tumor presence in the tissues was confirmed by PSA gene expression, which was quantified by real-time PCR. Gene expression level profiling of the transcriptome was analyzed via RNA sequencing of samples from 4 bladders, 4 kidneys, 3 lymph nodes, and 2 lungs. Gene expression of a select group of genes was verified by real-time PCR with a bigger population of RNA samples.

Results: Common differentially expressed genes were found between bladder and kidney (42 genes), bladder and lung (54 genes), and bladder and lymph nodes (198 genes). Bioinformatics software Panther and String were used to assign the genes into 6 pathways. These were morphogenesis of a branching epithelium; positive regulation of transcription from RNA polymerase II promoter; cell differentiation; negative regulation of epithelial to mesenchymal transition; epithelial cell differentiation; and transcriptional mis-regulation in cancer. Expression of paired-box 8 (Pax8), a transcription factor involved in embryonic development, was significantly higher in the kidneys compared to the other tissues. This suggests that overexpression of Pax8 may facilitate malignant development but may not be required for later stages of cell differentiation. Nanog, also a transcription regulator, was significantly higher in the kidneys when compared to bladders and was elevated in the lungs as well. It promotes epithelial-mesenchymal transition, resulting in metastasis; thus, increased expression of Nanog in cancer cells may indicate tumor aggressiveness. In the literature, Pax8 is expressed in noninvasive bladder tumors but not in normal bladder tissue in humans and Nanog expression is high in bladder tumors.

Conclusion: Pax8 and Nanog expression indicates that the murine model of bladder cancer with metastatic disease is similar to that observed in patients. Thus, this model could be used to evaluate methods of blocking progression.

Citation Format: Sin Mun Tham, Stefanie Alicia Fung, Kesavan Esuvaranathan, Ratha Mahendran. Gene expression changes associated with metastasis in mice with bladder cancer [abstract]. In: Proceedings of the AACR Special Conference on Bladder Cancer: Transforming the Field; 2019 May 18-21; Denver, CO. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(15_Suppl):Abstract nr A17.