Second Harmonic Generation imaging and optical scattering probes of ovarian cancer
Significance and Importance of Project: Five year survival rates of ovarian cancer remain poor (~30%), where this arises in part due to inadequate screening technologies as well as poor understanding of how cancer cells interact with the host tissue. There remains a compelling need for new technologies that have both sufficient resolution and specificity to image microscopic malignant ovarian tumors or precursor lesions. It is also equally important to understand how these changes occur and how they evolve between early stage and advanced disease. This project will develop new microscope-based methods that address these needs. The methods to be developed will be applicable to other cancers as well.
It is important to understand the changes that occur in the extracellular matrix (ECM) in the tumor microenvironment in ovarian cancer and how these evolve between early stage and advanced disease and between disease grades (e.g. borderline vs high grade). In this project, we will implement multimodal forms of Second Harmonic Generation (SHG) microscopy with optical scattering measurements to address this need by characterizing macromolecular changes in collagen and also in the 3D fibrillar architecture. The methods will i) compliment histology which primarily focuses on cells, where a non-negligible fraction of tissues are mis-classified; ii) provide prognostic information based on ECM changes from other patients; and iii) afford the development of an ex vivo or ultimately in vivo imaging/screening device of either ovaries or fallopian tubes. SHG has already been shown to be a highly sensitive and specific imaging tool for quantitatively describing changes in the collagen organization in several diseases, including many cancers, fibroses, and connective tissue disorders. This success is due to the coherent nature of the process where detailed information on fibrillar assembly can be obtained through analysis of morphology, polarization properties, and emission directionality. Similarly, optical scattering measurements have proven to be powerful for delineating cancers from normal tissue in several organs. We will now leverage our previous results on ex vivo human ovarian tissues and develop new tools to further understand changes in the collagen architecture, both in terms of collagen macromolecular assembly and 3D fibrillar organization. These are both important considerations as: i) we have demonstrated that dramatic morphological changes occur between normal and high grade malignant tissues, and ii) it has been reported that several minor isoforms become up-regulated in ovarian cancer and these may be effective biomarkers. We will also interrogate the collagen fibrillar architecture using SHG and optical scattering probes, separately and in conjunction and will compare remodeling across a spectrum of ovarian cancer types (e.g. serous vs endometroid, high grade vs low grade, fallopian tube vs primary ovary) as well as primary vs metastatic tumors. Measuring the wavelength dependences of optical scattering and SHG will provide detailed structural information across different length scales and delineate different tumor types based on their structural characteristics. To this end, a classification system based on the collagen molecular changes and fibrillar architecture alterations will be developed. Key to this work is the development of a suite of new label free SHG polarization analyses to specifically probe changes in collagen molecular alignment within fibrils, the á-helix pitch angle, and the overall chirality of the collagen triple helix. All these properties can change due to increased protease activity or changes in collagen isoform incorporation (e.g. Col III) in carcinogenesis and tumor growth. These will be benchmarked in self-assembled fibrillar gels that mimic the collagen assembly in the ovarian stroma and extended to ex vivo human tissues. In sum, this project will develop tools that probe all levels of collagen organization and will serve as new label-free biomarkers for ovarian cancer. Moreover, these methods will be generally applicable to other cancers and diseases characterized by changes in collagen architecture.
