Abstract B12: Carcinogen exposure alters keratin 5 expression and K5-Cre recombination in transgenic mouse urothelium

Abstract

Abstract In the urinary bladder, the normal urothelium is divided into three subpopulations of cells: superficial umbrella cells, intermediate cells, and a basal layer, each with distinct expression patterns of uroplakins (UP) and keratin 5 (K5). Many current transgenic mouse models (Tg) for urinary bladder cancer rely on uroplakin or keratin promoters to drive inducible Cre-lox recombination in basal or luminal bladder urothelial cells. Further, many models utilize the chemical carcinogen, N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN), to drive or accelerate tumorigenesis. A major limitation in Tg model systems where genes are knocked out is the requirement for highly penetrant and reproducible Cre activity and subsequent gene deletion in order to see a phenotype. This problem is compounded in long-term experiments such as BBN carcinogenesis that can run for up to six months after Cre induction. We have previously shown that Trim29 (tripartite motif containing 29) is sufficient to drive invasive bladder cancer formation in Tg models and in human bladder tumors. We have now developed a Tg system utilizing K5-Cre-ERT2 to target Trim29 for deletion in K5-expressing basal urothelial cells. In this model, tamoxifen exposure results in Cre expression and Trim29 deletion in K5 basal cells. Whether this would translate into complete loss of Trim29 in the urothelium over time and what effect BBN might have on tamoxifen induction of K5-Cre and loss of Trim29 were unclear. We found that exposure of K5-Cre-ERT2, Trim29 flox/flox mice to tamoxifen for six weeks resulted in loss of Trim29 expression only in the K5+ basal urothelium and that these K5+ Trim29 -/- basal cells did not give rise to intermediate cells or umbrella cells during the six months of follow-up. In contrast, mice treated with 0.05% BBN in drinking water for six months after induction of Cre produced full thickness of Trim29 knockout (KO) in 50% of mice (8/16 mice). Our results indicate that in a nonstressed state, K5 basal cells give rise to other basal cells but do not repopulate intermediate or umbrella cell layers. Furthermore, exposure to BBN resulted in the proliferation of basal cells and the replacement of intermediate cells by basal precursors in the urothelium. These results suggest that K5-Cre promoters may have different effects on transgene recombination in the bladder in the presence or absence of BBN, which induces more universal expression of Cre and its subsequent recombination events. These findings should be considered when interpreting and developing BBN-induced bladder cancer Tg models. Citation Format: Alan Kelleher, Yin Wang, Luke Broses, Sylvia Zelenka-Wang, Phillip L. Palmbos. Carcinogen exposure alters keratin 5 expression and K5-Cre recombination in transgenic mouse urothelium [abstract]. In: Proceedings of the AACR Special Conference on the Evolving Landscape of Cancer Modeling; 2020 Mar 2-5; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2020;80(11 Suppl):Abstract nr B12.