Intravital Imaging of Tumor Cell Motility in the Tumor Microenvironment Context.

Cancer cell motility and invasion are key features of metastatic tumors. Both are highly linked to tumor microenvironmental parameters, such as collagen architecture or macrophage density. However, due to the genetic, epigenetic and microenvironmental heterogeneities, only a small portion of tumor cells in the primary tumor are motile and furthermore, only a small portion of those will metastasize. This creates a challenge in predicting metastatic fate of single cells based on the phenotype they exhibit in the primary tumor. To overcome this challenge, tumor cell subpopulations need to be monitored at several timescales, mapping their phenotype in primary tumor as well as their potential homing to the secondary tumor site. Additionally, to address the spatial heterogeneity of the tumor microenvironment and how it relates to tumor cell phenotypes, large numbers of images need to be obtained from the same tumor. Finally, as the microenvironment complexity results in nonlinear relationships between tumor cell phenotype and its surroundings, advanced statistical models are required to interpret the imaging data. Toward improving our understanding of the relationship between cancer cell motility, the tumor microenvironment context and successful metastasis, we have developed several intravital approaches for continuous and longitudinal imaging, as well as data classification via support vector machine (SVM) algorithm. We also describe methods that extend the capabilities of intravital imaging by postsacrificial microscopy of the lung as well as correlative immunofluorescence in the primary tumor.