Enzymatic Tools for 2D Tissue Localized and Deeper Proteomic Sequencing of Cancer Stromal Proteins

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Molecular & Cellular Analysis Technologies
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Not Applicable
Determining the role of stromal proteins in cancer biology has remained difficult due to a lack of tools. Currenttools are qualitative and cannot report on protein sequence variation that is critical to understanding how stromalproteins help or impede cancer growth and metastasis. Our previous work has shown that bacterial collagenasescan be used to produce stromal proteomes detected by mass spectrometry as 2D distributions on cancer tissues.We still lack tools that can focus down on target proteins for detailed studies of sequence variation and post-translational modification. We propose using matrix metalloproteinases (MMPs) as enzymatic tools for targeteddetection of stromal protein sequences in the cancer tissue microenvironment. Aim 1, we will develop andoptimize a suite of MMPs for analysis of stromal sequences in tumor tissue sections. We will produce andoptimize recombinant MMPs specifically for use on 2D solid substrates of tissue sections. We will integratesequence information with current qualitative tools, expanding capabilities for stromal protein sequence detectiondown to a single cell niche. In Aim 2, we will focus on developing standards for quantitative detection of targetstromal proteins from the tissue microenvironment. We will test the developed technology against a small butrelevant cohort of emergent hepatocellular carcinoma from cirrhosis. Our research team has a uniquecombination of expertise for the production, development, and future commercialization of MMPs. The proposalis transformative since it will allow detection of stromal protein sequence variation from fixed clinical specimenswhich is impossible with conventional methods. A long-term objective is to significantly improve the community'sability to target stromal proteins within the tumor tissue microenvironment for use in basic research, directingtherapies, or prognostic or diagnostic biomarkers.