Single-Cell Profiling of Murine Bladder Cancer Identifies Sex Specific Transcriptional Signatures with Prognostic Relevance. Single-Cell Profiling of Murine Bladder Cancer Identifies Sex Specific Transcriptional Signatures with Prognostic Relevance

Bladder cancer (BLCA) is more common in men but more aggressive in women. A common model used to study sex-based differences in cancer biology is murine BLCA generated by N-butyl-N-(4-hydroxybutyl)-nitrosamine (BBN). While tumors in this model have been profiled, these profiles provided limited information on the tumor microenvironment. Hence, we applied single-cell RNA sequencing to characterize cell type specific transcriptional differences between male and female BBN induced tumors and then studied their human relevance. We found proportion and gene expression differences in the epithelial and non-epithelial subpopulations between male and female tumors. Of translational importance, expression of several genes was found to predict sex specific survival in several human BLCA datasets. We also noted a male biased CD8 T cell exhaustion program in this model that correlates highly with androgen receptor activity. Our study identified novel and clinically relevant sex specific transcriptional signatures and validates the relevance of the BBN model for studying sex differences in human BLCA. This work highlights the importance of considering sex as a biological variable in cancer marker development. Overall design: Single cell RNA-sequencing of mouse bladder tissue following BBN treatment for 6 months (n=20) pooled into 10 samples

膀胱癌（BLCA）在男性中发病率更高，但女性患者的肿瘤侵袭性更强。用于研究癌症生物学中性别差异的经典模型之一，是由N-丁基-N-(4-羟丁基)亚硝胺（BBN）诱导构建的小鼠膀胱癌模型。尽管已有针对该模型中肿瘤的组学表征，但此类表征对肿瘤微环境的解析较为有限。因此，本研究采用单细胞RNA测序技术，解析BBN诱导的雌雄小鼠膀胱癌的细胞类型特异性转录组差异，并进一步探究其与人类膀胱癌的相关性。研究发现，雌雄肿瘤的上皮及非上皮亚群在细胞比例与基因表达层面均存在差异。具有转化研究价值的是，在多个人类膀胱癌数据集内，多个基因的表达水平可预测性别特异性的患者生存情况。此外，本研究还在该模型中观察到一种雄性偏向性的CD8 T细胞耗竭程序，其与雄激素受体活性高度相关。本研究鉴定出了全新且具有临床相关性的性别特异性转录特征，并验证了BBN模型在研究人类膀胱癌性别差异中的适用性。本研究凸显了将性别作为癌症标志物开发过程中的生物学变量的重要性。实验设计概要：对经BBN处理6个月的小鼠膀胱组织进行单细胞RNA测序（共20只小鼠，样本混合为10个测序样本）