DDR1 regulates RUNX1-CBFβ to control breast stem cell differentiation

Understanding epithelial stem cell differentiation and morphogenesis during breast tissue development is essential, as disruption in these processes underlie breast cancer formation. Recent work has shown that epithelial stem and progenitor cells use the collagen receptor Discoidin Domain Receptor 1 (DDR1) for differentiation into both basal and luminal cell lineages, which together are necessary for complex ductal-lobular morphogenesis. Here, we used a next-generation single cell derived organoid model that generates miniaturized breast tissue, to study how single stem cells can give rise to multiple cell types and compound tissue structures. We show that DDR1inhibition traps cells in a bipotent state, blocking alveolar morphogenesis and luminal cell expansion necessary for complex epithelium formation. Disrupting RUNX1 function produced nearly identical phenotypes, underscoring its critical role downstream of DDR1. Mechanistically, DDR1 affects the interaction and expression of RUNX1 and its cofactor CBFβ, thereby regulating its activity. Mutational analyses in breast cancer patients reveal frequent alterations in the DDR1-RUNX1 signaling axis, particularly co-occurring mutations. Together, these findings uncover DDR1-RUNX1 as a central signaling pathway driving breast epithelial differentiation, whose dysregulation may contribute fundamentally to breast cancer pathogenesis. Three primary patient samples were cultured in 3D hydrogels for 7 days bafore receiving chronic treatment (Vehicle, DDR1i, AI-10-104 (RUNXi) for an additional 14-21 days until control was fully developed. Bulk RNA-seq was performed on these samples, followed by gene expression analysis comparison of differential gene sets from RUNXi and DDR1i treated samples compared to the control.

解析乳腺组织发育过程中的上皮干细胞分化与形态发生机制至关重要，因为这些过程的紊乱是乳腺癌发生的根源。已有研究表明，上皮干细胞与祖细胞借助胶原受体盘状结构域受体1（Discoidin Domain Receptor 1，DDR1）完成向基底细胞系与腔细胞系的分化，这两类细胞谱系共同参与复杂导管-小叶形态发生过程，是其必要组分。本研究采用可生成微型化乳腺组织的新一代单细胞来源类器官模型，探究单个干细胞如何分化为多种细胞类型并形成复合组织结构。研究结果显示，抑制DDR1会将细胞阻滞于双潜能状态，阻断复杂上皮形成所需的肺泡形态发生与腔细胞扩增过程。破坏RUNX1功能可产生几乎一致的表型，凸显了其作为DDR1下游关键调控因子的核心作用。从机制层面来看，DDR1可调控RUNX1及其辅因子CBFβ的表达与相互作用，进而调节RUNX1的转录活性。乳腺癌患者的突变分析显示，DDR1-RUNX1信号轴存在高频突变，尤以共突变现象最为显著。综上，本研究揭示DDR1-RUNX1信号轴是驱动乳腺上皮分化的核心信号通路，其功能紊乱可能从根本上参与乳腺癌的发病进程。本研究将3例原代患者样本置于三维水凝胶中培养7天，随后分别给予长效处理：溶媒对照（Vehicle）、DDR1抑制剂（DDR1i）、AI-10-104（RUNX抑制剂，RUNXi），持续处理14-21天直至对照组样本完全成熟。随后对所有样本进行批量RNA测序（Bulk RNA-seq），并对比分析RUNXi与DDR1i处理组相较于对照组的差异基因集。