We are interested in understanding both the heterogeneity within tumor cells that drives their capacity to resist therapy and metastasize as well as the contribution of the tumor microenvironment to tumor growth. Key to these efforts is the use of in vivo genetically engineered models of cancer. We previously identified a rare subpopulationof tumor cells characterized by surface expression ofCD24+/ITGB4+/Notch3+. We have shown that these cells have increased CSC-like characteristics in both the KRASLSLG12D model and in KRASLSLG12D;p53-/-mice. In addition, the percentage of these cells as a fraction of the bulk tumor increases after repeated doses of chemotherapy, suggesting that these cells are intrinsically chemoresistant (Zheng et al, Cancer Cell 2013). Our current efforts are directed at understanding the role of Notch3 in TPCs both in mouse models of lung cancer and in human lung cancer PDX models. We hypothesize that understanding how Notch3 signals may help identify novel approaches to therapy for NSCLC. We are also extending our initial characterization of TPCs in the KRASLSLG12D;p53-/- to other genetic backgrounds (loss of LKB1, loss of KEAP1, etc). Using advanced reporter assays and single-cell sequencing assays, we are also trying to further understand how intratumor heterogeneity evolves over time and how it is influenced by genetic context.

Tumor heterogeneity can occur not only within tumorcells but also with regards to the tumor stroma. We previouslyidentified a role for cancer-associated fibroblasts (CAFs) in promoting tumor development in both mouse and human lung cancer (Vicent et al, Cancer Research 2012). Using gene expression analysis, we identified a novel pro-oncogenic cytokine signaling axis in which CLCF1 secreted by CAFs signals to CNTFR on tumor cells to promote tumor growth. This work has led to a collaboration with a bioengineer at Stanford (Dr. Jennifer Cochran lab) with thegoal ofdevelopingtherapeutic strategies to block the interaction between CAF-expressed cytokines and their receptors on tumor cells. Our work on this novel targeted therapy demonstrates that it may be particularly helpful in patients with KRAS mutation, a currently difficult to treat subset of human lung cancer (Kim and Marquez et al, Nature Medicine, 2019). Current efforts are directed at further elucidating the mechanism of action of CNTFR signaling inhibition and expanding the potential utility of this therapy beyond lung cancer.