Identification of tumorigenic cells in Kras(G12D)-induced lung adenocarcinoma.

We established an inducible Kras(G12D)-driven lung adenocarcinoma in CCSP-rtTA/TetO-Cre/LSL-Kras(G12D) mice that enable pursuits of the cellular and molecular processes involved in Kras-induced tumorigenesis. To investigate the cellular origin of this cancer, we first report a strategy using fluorescence-activated cell sorting fractionation that could highly enrich bronchiolar Clara and alveolar type II cells, respectively. The EpCAM(+)MHCII(-) cells (bronchiolar origin) were more enriched with tumorigenic cells in generating secondary tumors than EpCAM(+)MHCII(+) cells (alveolar origin) in primary tumors that had been already initiated with oncogenic Kras activation. In addition, secondary tumors derived from EpCAM(+)MHCII(-) cells showed diversity of tumor locations compared with those derived from EpCAM(+)MHCII(+) cells. In the alveolar region, secondary tumors from EpCAM(+)MHCII(-) cells expressed not only bronchiolar epithelial marker, panCK, but also differentiation marker, proSPC, consistent with the notion that cancer-initiating cells display not only the abilities for self-renewal but also the features of differentiation to generate heterogeneous tumors with phenotypic diversity. Furthermore, high level of ERK1/2 activation and colony-forming ability as well as lack of Sprouty-2 expression were also observed in EpCAM(+)MHCII(-) cells. Therefore, these results suggest that bronchiolar Clara cells are the origin of cells and tumorigenesis for Kras(G12D)-induced neoplasia in the lungs.