PROJECT SUMMARY Large-scale genomic studies have discovered numerous relevant alterations that activate or inactivate key signaling molecules in lung cancer, which led to significant improvement on patients' outcome and our therapeutic options. Despite these advances in the genome-based precision medicine, the 5-year survival rate of 18% remains less than adequate. This is attributable to eventual emergence of resistance and variability in response to the treatment, largely due to heterogeneity of lung cancers. To improve precision and efficacy, innovative strategies to distinguish different subgroups of lung cancer by additional features for future drug development are desperately needed. We hypothesize that our natural system to populate necessary cells to constitute the organ structure and its function is hijacked by cancer mimicking normal cell identities to maintain their survival. Our previous studies found lung cancer cells harboring amplification of lineage oncogenes are specifically dependent on their expression for survival. The need for lineage amplified genes in a subset of cancers suggests unique vulnerabilities in cancer cells that may be poised at the brink of survival. However, these amplifications represent only a minor subset of the two major subtypes of lung cancers. Therefore, this study will focus on lineage factors essential during the development and control the cell identity in adult, in the remaining subsets of lung cancers. Through genome-wide profiling of a histone mark, we identified `super- enhancers' (SE) on the lineage-defining transcription factor genes. Subsequent hierarchical clustering of lung cancer samples identified SEs specific to subsets of lung cancers. Therefore, we aim to understand the significance of our lead candidates defining the lineage state of the novel subsets in aim 1. We will test the subclass-specific local chromatin structure and then investigate the roles of novel lineage transcription factors specifically enriched in the two major subgroups of lung ADCs, whether they are essential for maintaining the cellular state of lung cancer cells and are dependent on the committed lineage for their survival. In aim 2, to determine the roles of a neural transcription factor Brn2 in a novel `neural' subtype of lung squamous cell cancers (SCC) as a cooperative partner of Sox2, by contrast to p63, a key squamous lineage factor we previously characterized as an important cooperative partner of Sox2 in classic lung SCC. In aim 3, to pursue inducing transdifferentiation as a therapeutic strategy, we seek to understand how lineage switch occurs within a tumor by determining heterogeneity and dynamics of lineage states. We will determine the mode of temporal dynamics of population shifts in lineage states induced by genetic perturbation in defined models via profiling genome-wide chromatin landscapes at a single cell level. We will then test our hypothesis that the degree of heterogeneity correlates with plasticity of the tumor and aggressive tumor behaviors using a mouse model and human specimens. The results will serve as a proof-of-principle how these lineage factors contribute to the formation of lung cancer and lead to new hypotheses how we can manipulate cell identity of lung cancers.