Abstract: FR-OR15
Characterization of Stepwise Transformation from Cysts to Carcinoma in Tsc1 Mutant Kidneys
Session Information
- Cystic Kidney Diseases: Basic and Translational Research
October 25, 2024 | Location: Room 23, Convention Center
Abstract Time: 05:20 PM - 05:30 PM
Category: Genetic Diseases of the Kidneys
- 1201 Genetic Diseases of the Kidneys: Cystic
Authors
- Clerici, Sara, IRCCS Ospedale San Raffaele, Milano, Italy
- Spies, Daniel, IRCCS Ospedale San Raffaele, Milano, Italy
- Boletta, Alessandra, IRCCS Ospedale San Raffaele, Milano, Italy
Background
A novel mouse model of Renal Cell Carcinoma (RCC) was generated by deletion of Tsc1 using the Cadherin16 Cre (KspCre). This results in cysts undergoing epithelial transformation to cancerous lesions with time. The peculiarity of Tsc1;kKO model is the time-driven progression from cysts to papillae, to cystadenomas, and carcinomas with full penetrance. As the malignant phenotype progresses canonical epithelial markers are lost. To overcome this problem, we generated a mTmG;Tsc1;kKO mouse in which KO cells express membrane GFP (mG) upon Cre expression.
Methods
RCC kidneys were characterized by IHC, IF analysis, and FACS. scRNA-seq was performed on early- (P30) and late-stage (P80) ctrl and RCC kidneys using 10X genomics. Multiplexed error-robust FISH (MERFISH) analysis on 500 genes panel is currently being conducted to spatially resolve the clusters identified.
Results
mTmG;Tsc1;kKO kidneys developed cysts and malignant structures at the same rate and time as the Tsc1;kKO. All the structures were mG+, with a considerable expansion of mG+ cells in P80 kidneys analyzed by FACS, in line with the increased proliferation of Tsc1 mutants. scRNA-seq on freshly dissociated kidneys at two time points (P30 and P80) identified clusters from all the major nephron segments in both ctrl and mutant kidneys. Egfp transcript was uniquely identified in several new epithelium-derived clusters, enriched at P80 in the mutant. We defined two distinct branches of progressive transformation originating from either principal or intercalated cells of the collecting duct. Interestingly, the two trajectories converge on a cluster with a papillary RCC signature. Initial pathway enrichment analysis of transcripts identified multiple metabolic pathways that are increasingly deranged during transformation. Novel, non-canonical markers were identified and used to design a MERFISH panel to spatially resolve stages of transformation in the murine kidneys and will be used to design a mini-panel to be validated on human specimens.
Conclusion
Our new RCC murine model allows to discriminate early cystogenic processes from subsequent transformation. The combination of scRNA-seq and spatial validation allows to identify new RCC populations, likely the cell of origin of the transforming clusters, and finally to define the mechanism causing the gradual transformation. Complete analysis will be presented.