14. Characterizing response to checkpoint blockade treatment in mouse urothelial carcinoma model with scRNA

      While checkpoint blockade treatment has been shown to be highly effective in various tumor types, effectiveness for urothelial carcinoma has been minimal. To examine how urothelial carcinoma responds to combined PD-L1 and CTLA-4 inhibition, we used single cell RNA sequencing along with whole genome, exome, and bulk RNA sequencing derived from a mouse model with three condition groups: control, checkpoint treatment (Atezolizumab and Ipilimumab), and checkpoint treatment with CD4 depletion. The cell line used for analysis is an organoid-based line developed by inducing invasive urothelial carcinoma in mice through exposure to N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) in drinking water. We performed whole genome, exome, and bulk RNA sequencing of the cell line directly. To generate single cell RNA data, we injected organoid cells subcutaneously to establish tumors and pooled samples from three mice per condition to produce single cell samples for 10 × 5′ gene expression and T-cell receptor sequencing. Using exome data and comparing to sequence data from matched mouse tail, we detected a high number of somatic mutations (16,318) in our cell line and used a filtered list to characterize somatic variant expression in single cell data. Preliminary results show that we can detect a high number of variants and identify distinct tumor cell clusters based on the number of somatic variants detected. We also find the cell line has high immune infiltration based on numbers and sizes of immune cell populations detected. One cluster type of particular interest is Jchain-positive plasma cell clusters, which appear to expand with checkpoint blockade treatment.
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