Single-cell RNA sequencing of chemotherapy-resistant muscle-invasive urothelial bladder cancer. Single-cell RNA sequencing of chemotherapy-resistant muscle-invasive urothelial bladder cancer

To identify the therapeutic targets in a treatment-refractory cancer patient, we performed single-cell RNA sequencing for 3,115 cells from primary bladder cancer (BC159-T#3) and patient-derived xenografts (BC159-T#3-PDX-vehicle and BC159-T#3-PDX-tipifarnib). Matched time-series bulk tumor tissues were also sequenced using whole exome target probe (WES) and whole transcriptome target probe (WTS). Overall design: All single-cell mRNA sequencing data were acquired from a single patient (BC159-T#3) and patient-derived xenografts (BC159-T#3-PDX-vehicle and BC159-T#3-PDX-tipifarnib). The PDX samples were mapped to the human and mouse genome, respectively. To remove the cells with low sequencing quality, we applied filtering criteria using nUMI, nGene, proportion of mitochondiral genes, and averaged expression of housekeeping genes. Additional filtering criteria was applied to the PDX samples to eliminate human-mouse mixed cells. Total 3,934 cells met the criteria. After identification of cell type, the strict criteria were applied to tumor cells using nGene and inferred copy number variations. Thereafter, we removed outliers in nGene to eliminate potential multiplets in nontumor cell population. The raw data at EGA archive due to patient privacy concerns [EGAD00001005978].

为鉴定难治性癌症患者的治疗靶点，我们对来自原发性膀胱癌患者（BC159-T#3）以及患者来源异种移植模型（BC159-T#3-PDX-溶剂组和BC159-T#3-PDX-替吡法尼组）的3115个细胞实施了单细胞RNA测序。匹配的时序性实体瘤整体组织（bulk）样本还通过全外显子组靶向探针（whole exome target probe, WES）与全转录组靶向探针（whole transcriptome target probe, WTS）完成了测序。整体实验设计如下：所有单细胞mRNA测序数据均来自同一名患者（BC159-T#3）及其对应的患者来源异种移植模型（BC159-T#3-PDX-溶剂组和BC159-T#3-PDX-替吡法尼组）。我们分别将PDX样本比对至人类和小鼠基因组。为去除测序质量较低的细胞，我们基于nUMI、nGene、线粒体基因占比以及管家基因平均表达量设置了过滤标准。针对PDX样本，我们额外增设过滤规则以去除人鼠混合细胞。最终共有3934个细胞符合所有过滤标准。完成细胞类型注释后，我们针对肿瘤细胞群体基于nGene数值与推断的拷贝数变异设置了严格的筛选标准。随后，我们对非肿瘤细胞群体中的nGene数值进行异常值剔除，以去除潜在的多重细胞（multiplets）。由于患者隐私保护要求，原始数据已归档至EGA数据库中[EGAD00001005978]。