Imaging Hypoxia-driven Signaling Pathways in the Breast Tumor Microenvironment (R01CA134695)

Tumor hypoxia has been associated with tumor progression, a higher risk of metastatic spread, and resistance to therapy, and has thus become a central issue in tumor physiology and cancer treatment. To date, very little is known about the molecular pathways that are affected by tumor hypoxia, and which eventually cause the disastrous clinical effects of poor cancer prognosis and poor treatment outcome. Hypoxia-inducible factor 1 alpha (HIF-1 alpha), whose levels increase under hypoxic conditions, plays an important role in tumor hypoxia as it affects the levels of other biomolecules. Because currently little is known about the key biomolecules in tumor hypoxia, we will seek to identify to date unknown molecules that are increased or decreased in hypoxic regions in breast tumors. We will use a unique model system to study hypoxia, which consists of human breast cancer cell lines and the corresponding tumor models grown in immune-compromised animals that were genetically engineered to contain a built-in hypoxia detector. This detector couples the natural hypoxia response of increased HIF-1 alpha to the production of a fluorescent marker that can be detected by optical imaging. We will combine optical hypoxia detection with in vivo magnetic resonance spectroscopic imaging (MRSI), cutting- edge mass spectrometry imaging (MSI) applications, and targeted proteomics strategies. In our first specific aim, we will discover, identify, and validate biomolecules that are decreased or increased due to hypoxia in breast cancer cell cultures. We will compare three human breast cell lines representing different degrees of aggressiveness and metastatic potential that have been made hypoxic in the laboratory. In our second specific aim, we will carry out parallel studies in actual breast tumor models grown from the same breast cancer cells lines, which contain the built-in hypoxia detector. We will analyze the hypoxic regions in these breast tumors using the same MS-based proteomics approach as in the cell lines. In the third specific aim, we will evaluate the hypoxia-related biomolecules initially identified in Aims 1 and 2 using a multimodal 3D molecular imaging approach, which will combine in vivo MRSI, optical imaging, and MSI methods. MRS, optical, and MS images will be acquired of the same breast tumor models containing the built-in hypoxia detector, which will enable us to assess the spatial relationship between hypoxia, already known hypoxia marker molecules, and our newly identified hypoxia-related molecules. Our studies will lead to a better understanding of the molecular pathways that are triggered by hypoxia in breast tumors. The proposed studies may eventually translate into new breast cancer therapies for patients that have hypoxic regions in their tumors. Future studies can explore possibilities to use these newly discovered hypoxia-related molecules as targets for treating tumor hypoxia, and hopefully improve the treatment outcome of cancer patients with hypoxic breast tumors. This project is in collaboration with Dr. Ron M.A. Heeren.