Abstract We have systematically dissected the microenvironmental hetereogeneity of lung adenocarcinomas, and provide cellular and molecular landscapes that contribute to carcinogenesis. RNA deep sequencing analysis of individually sorted stromal and tumor epithelial compartments from fresh clinical specimens and a mouse model of KrasG12D; p53-/- tumors identified cell type-specific differentially regulated genes, indicative of tumor “activated/reprogrammed” stroma. We developed a computational crosstalk discovery algorithm to model crosstalk signaling discovery based on ligand-receptor interactions and downstream signaling networks, and identified novel tumor-stroma paracrine and tumor autocrine crosstalk signaling pathways in the lung cancer microenvironment. We have explored the mechanistic basis of selected crosstalk pathways including HGF-MET, IL6-Stat3, Tsp-1-CD36, MMP14-HB-EGF in conferring lung adenocarcinoma progression and in mediating therapeutic resistance, and have used both genetic and pharmacological approaches to block these pathways. This study has the potential for development of therapeutic strategies that target tumor-stroma interactions, and may complement conventional anti-cancer treatments. Citation Format: Hyejin Choi, Jianting Shen, Anna Durrans, Tina El Rayes, Kari Fischer, Dingcheng Gao, Steve Wong, Nasser Altorki, Vivek Mittal. Genome-wide analysis of stroma-tumor crosstalk pathways in lung cancer: therapeutic implications. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3214. doi:10.1158/1538-7445.AM2015-3214