Metastasis is the primary cause of death in patients with lung cancer, and its genetic and biological bases are poorly understood. Progress in this area was hampered by the lack of in vivo models that faithfully recapitulate genetic and biochemical features of human lung cancer metastasis. To address this knowledge gap, we have developed a series of genetically-engineered mouse models of human lung adenocarcinoma initiated by K-rasG12D expression in which secondary oncogenic mutations, including Tp53R172H expression or inactivation of Pten or Map2k4, lead to more advanced disease but differ in the degree to which they promote disease advancement.
Our transcriptional profiling studies revealed that poor-prognosis human lung adenocarcinomas were highly enriched in genes differentially expressed between primary and metastatic tumors in mice that develop widely metastatic lung adenocarcinomas owing to expression of K-rasG12D and p53R172H (KP mice). We have shown that KP mice harbor disease whose progression closely mirrors that of poor-prognosis lung adenocarcinoma in patients and that lung adenocarcinoma cell lines derived from these mice provide a useful platform for the discovery of clinically relevant, pharmacologically actionable metastasis drivers. Metastatic tumor cells derived from KP mice switch reversibly between epithelial and mesenchymal states in response to extracellular cues; this plasticity is critical for metastasis and is driven by mutual antagonism between transcription factors that activate epithelial-to-mesenchymal transition (EMT) (e.g., ZEB, SNAIL, and TWIST family members) and microRNAs that target the EMT-activating transcription factors (e.g., miR-200 and miR34 family members). We are currently studying how the EMT regulatory axis governs tumor cell polarity and the formation of actin-based cytoplasmic protrusions (e.g., filopodia and lammellipodia) by controlling Golgi-regulated trafficking of secretory vesicles.
Epithelial tumor metastasis is preceded by an accumulation of collagen cross-links that heighten stromal stiffness and stimulate the invasive properties of tumor cells. In normal connective tissues, stiffness is regulated by changes in the biochemical types of collagen cross-links, leading us to postulate that metastasis is driven by a switch in the type of collagen cross-link in tumor stroma. Using mouse models of metastatic lung cancer and human lung cancers, we showed that metastatic tumor cells and cancer-associated fibroblasts (CAFs) have increased expression of lysyl hydroxylase 2 (LH2), an enzyme that hydroxylates telopeptidyl lysines on collagen. We showed that LH2 enhances the invasive and metastatic properties of tumor cells and functions as a regulatory switch, controlling the relative abundance of biochemically distinct types of collagen cross-links in the tumor stroma. To directly translate this finding to the clinic, we have developed a LH2 enzymatic assay, adapted it to high throughput screening, and are currently interrogating small molecule libraries to identify first-in-class LH2 antagonists. Such antagonists will be optimized through ongoing protein crystallographic studies that will elucidate structural features of the LH2 catalytic domain.
CAFs are mesenchymal cells of diverse origins that promote metastasis through physical interactions with tumor cells and a secretome that modulates the behaviors of diverse cell types in the tumor microenvironment. We have shown that, co-cultured in 3-dimensional matrices, CAFs and tumor cells form “leader-follower” cell structures in which CAFs lead the directional movement of invading tumor cells. We are currently examining the biochemical basis for how CAFs and tumor cells physically interact to form leader-follower cell structures and how CAFs promote invasive behavior by exerting biomechanical forces on tumor cells and the extracellular matrix.