Multi-colour lineage tracing reveals intra-stage proliferative heterogeneity during mammary tumour progression. Multi-colour lineage tracing reveals intra-stage proliferative heterogeneity during mammary tumour progression

We have assessed clonal heterogeneity within individual primary tumours and metastasis and also during the distinct stages of malignant tumour progression using the Confetti lineage reporter in the background of the MMTV-PyMT mouse model of metastatic breast cancer. Comparative gene expression analysis of the clonal populations reveals a substantial level of heterogeneity across and also within the various stages of breast carcinogenesis. This intra-stage heterogeneity is manifested by differences in cell proliferation, where the fast-proliferating subclass is further enriched in oxidative phosphorylation and cell death. Overall design: Outgrowth of clonal cell populations was analysed when the maximum tumour volume comprising all tumour stages (normal mammary gland, hyperplasia, adenoma, carcinoma, pulmonary metastases) had been reached in colour-induced MMTV-PyMT; R26-CBW; K8-CreERT2 mice. Zonal patches of clonal populations of the primary tumour as well as metastases were isolated by Laser Capture Microdissection (encircled regions) and samples obtained were subjected to RNA sequencing. n = 3.

本研究以转移性乳腺癌的MMTV-PyMT小鼠模型为遗传背景，借助Confetti谱系报告基因（Confetti lineage reporter），对个体原发性肿瘤与转移灶内的克隆异质性（clonal heterogeneity）以及恶性肿瘤进展的不同阶段的克隆异质性进行了评估。对克隆群体开展的比较基因表达分析结果显示，乳腺癌发生的各阶段之间以及各阶段内部均存在显著的异质性。该阶段内异质性可通过细胞增殖差异得以体现：快速增殖的细胞亚群在氧化磷酸化（oxidative phosphorylation）与细胞死亡相关通路中呈现进一步富集的特征。整体实验设计：在经颜色诱导的MMTV-PyMT; R26-CBW; K8-CreERT2小鼠中，当包含所有肿瘤阶段（正常乳腺组织、增生、腺瘤、癌、肺转移灶）的最大肿瘤体积形成时，对克隆细胞群体的增殖情况进行分析。通过激光捕获显微切割（Laser Capture Microdissection，圈定区域）分离原发性肿瘤及转移灶的克隆群体区域斑块，将获取的样本进行RNA测序（RNA sequencing）。本实验重复数n=3。