When light interacts with matter and the conditions are just right, light of one color can be converted to light of another color, in a process called second harmonic generation (SHG). SHG is very sensitive to the order, or organization, of matter. In tumors, we and others have shown that SHG is caused primarily by collagen, and is sensitive to the extent to which collagen is bundled into fibers, the way the fibers are bundled together, the thickness of the fibers, and other properties. SHG can be used in microscopes to make high-resolution images of tumor collagen that reveal many of these properties. This is interesting to people other than scientists because these images have revealed that tumor cells use "roads" of ordered (SHG-producing) collagen to move rapidly throughout a tumor, to approach and enter blood vessels, and to escape the tumor mass.

We propose to learn what cells in the tumor are contributing to the construction of these "roads" and what signaling molecules they are using to do this. In preliminary experiments, we were able to manipulate the state of the "roads" in tumors in mice, and these experiments, plus the literature, suggest that cells called macrophages and several types of T cells are probably responsible. In this project, we will determine if they are indeed responsible for this road construction, and what molecules they use to communicate their road building commands. This will allow us to figure out good targets for subsequent drug development that will disrupt these "roads" and hence prevent, or at least slow down, the tumor cells from metastasizing. In parallel with this work, we will also study patient biopsies from breast tumors and see if SHG imaging, making a "roadmap" of the tumors, can be used to predict metastatic ability: if a tumor has a poorly arranged road network or the quality of the roads themselves is poor, one might predict the tumor cells will not metastasize easily. A method to predict metastatic ability is an identified need in the clinic because currently most patients are "over-treated" with systemic chemotherapy after surgery while studies have shown that they really only needed local treatment. Eliminating unnecessary treatment after surgery will have a significant impact on many patients' quality of life. Hence, this study will attempt to improve a patient's treatment outcome as well as quality of life. The timescales of this proposal are encouraging: larger trials of the metastasis-prediction technology could start within 3 years, while development of the novel drugs could start within 3 to 5 years.

This proposal has a high likelihood of success because the projects within the proposal are highly interconnected, yet each is independently worthy of pursuit and stands upon its own merits: determining the role of macrophages (Aim 1) and CD4+ T cells (Aim 2) in collagen ordering are each worthwhile in order to develop future drug targets and improve treatment outcomes. Likewise, evaluating the ability of optical signatures of collagen ordering to predict metastasis (Aim 3), independent of understanding the underlying cells and signals, may provide a benefit to patients' quality of life. In summary, we hope to improve the outcome of breast cancer treatment, as well as the quality of life of breast cancer patients. We feel that this project has a high probability of success.