Abstract LB-355: Image-guided genomics reveals a symbiotic relationship between heterogeneous phenotypes in collective cancer invasion

During tumor metastasis, cancer cells invade as a phenotypically heterogeneous collective pack to navigate the tumor microenvironment. Leader cells pioneer invasion into the microenvironment whereas follower cells immediately attach to and follow the leaders. To dissect the molecular mechanisms underlying this phenotypic heterogeneity, we developed a technique termed spatiotemporal genomic and cellular analysis (SaGA), which uses image-guided genomics to precisely select living, rare cell subpopulations that are maintained within a physiologically relevant environment for downstream genomic and molecular analyses. In this manner, we can precisely select, isolate, and amplify any living cell based upon phenotypic criteria. Using H1299 lung cancer cells, we precisely selected as few as 10 leader cells using the SaGA technique and extracted them from the bulk of a multicellular cancer spheroid embedded within a 3-D matrix. These isolated cells were used to create the first leader and follower purified cell cultures. The purified leader cell cultures continually show dynamic invasive patterns over time, while follower cells alone have limited invasive capabilities. Reintroducing limited numbers of leader cells, or even leader cell conditioned media, into follower cell cultures promoted invasion and motility in the follower cells. Gene expression analysis comparing leader versus follower cell lines showed significant enrichment in cell adhesion and vascular endothelial growth factor (VEGF) signaling pathways in leader cells. This was confirmed by protein analysis showing that leader cells secrete high levels of VEGF, and VEGF inhibition abolished leader-follower collective invasion, suggesting vascularity sprouting mimicry during chain formation. Additional analysis confirmed that leader cells utilize focal adhesion kinase-fibronectin signaling to create tension force to promote chain motility. While leader cells provide an escape mechanism for followers, follower cells in turn provide leaders with increased proliferation and survival. These data support a symbiotic model of collective invasion where different cell subtypes cooperate to promote successful escape. Overall, our data demonstrate that SaGA can precisely select living cells based upon dynamic behaviors for genomic analysis and can amplify rare cell populations for subsequent molecular, cellular, and proteomic analyses. Therefore, this image-guided method has the potential to impact the field of tumor heterogeneity by uncovering genomic signatures of rare yet dynamic subpopulations within a heterogeneous cancer population. Citation Format: Jessica Konen, Emily Summerbell, Kornelia Galior, Bhakti Dwivedi, Khalid Salaita, Jeanne Kowalski, Adam Marcus. Image-guided genomics reveals a symbiotic relationship between heterogeneous phenotypes in collective cancer invasion. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr LB-355.