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Glycoproteomic Identification of Prognostic Serum Biomarkers
Detection, Diagnosis and Prognosis
Background: Effective biomarkers capable of serving as prognostic indicators of disease recurrence or therapeutic response for breast cancers are still needed. The identification and simultaneous analysis of a panel of biomarkers by proteomic technologies offers the potential for improving prognostic assays for breast cancers. Changes in the glycosylation constituents of serum glycoproteins are known to occur as breast cancers progress towards metastasis, thus monitoring post-treatment changes in serum glycoprotein patterns could provide new candidate biomarkers. Objective/Hypothesis: The objective is to assess altered patterns of glycosylation on serum glycoproteins to develop new breast cancer prognosis and treatment monitoring assays. The hypothesis to be tested is that proteomic technologies can be used to identify and monitor changes in specific serum glycoproteins that will better indicate therapy outcomes and disease recurrences. Specific Aims: 1. Use multiple lectins to capture differentially expressed glycoproteins in serum from pre- and post-treatment breast cancer patients . The protein and oligosaccharide sequences of these captured serum glycoproteins will be determined. 2. Characterize other prognostic serum markers from pre- and post-treatment samples . Extensive proteomic mass spectrometry and gel electrophoresis studies will be done to identify other new serum biomarkers. Study Design: Archived longitudinal sera samples from individual breast cancer patients with invasive ductal carcinoma that underwent surgery and chemotherapy or radiation will be used. Different lectin conjugates will be used to capture differentially expressed serum glycoproteins. Gel electrophoresis coupled to a MALDI TOF/TOF instrument for rapid protein sequence identification will be used. In Aim 2, affinity bead separations and dual fluorescence gels will be used prior to tandem mass spectrometry sequencing. Potential Outcomes and Benefits: We anticipate detecting serum glycosylation changes that reflect a more non-cancer state in patients with known favorable treatment outcomes. Conversely, detection of unchanged serum glycosylation patterns following treatments will indicate the opposite, a poor prognosis. This type of assay could be used to monitor responses in real time during the treatment regimen, allowing faster determination of success or failure, thus giving clinicians greater flexibility for individualizing treatment to each patient.
Background: Proteomic technologies are a new approach that have been successfully applied to profiling and identifying tumor associated proteins in the blood and tissues of breast cancer patients. These tumor associated proteins are called biomarkers, and characterization of these proteins can be used to develop new diagnostic cancer assays. Glycoproteomics is an emerging branch of proteomics that focuses on characterizing the sugar residues (carbohydrates) that are commonly attached to proteins as they are being made in cells, a process termed glycosylation. This glycosylation process can be altered during development of breast cancers, and using new proteomic technologies, we propose to characterize the changes in glycosylation patterns of blood proteins to develop improved therapy monitoring and surveillance assays. Objective/Hypothesis: The objective is to use stored blood obtained from women undergoing breast cancer therapies to look for altered patterns of glycosylation. The hypothesis to be tested is that we can use proteomic technologies to identify and monitor specific changes in blood glycoproteins for use as predictors of therapy outcome and disease recurrence. Specific Aims: 1. Use glycoprotein capture reagents, termed lectins, to pull out glycoproteins from stored blood samples of pre- and post-treatment breast cancer patients. 2. Characterize other predictive blood protein biomarkers from these pre- and post-treatment samples. Study Design: Stored blood samples collected from breast cancer patients before, during and after their surgeries and other treatments will be used. Different lectins will be used to capture differentially expressed blood glycoproteins. Gel electrophoresis and protein mass spectrometry will be used to accomplish the objectives. Other affinity capture methods and new dual labeling gel methods will also be used. Potential Outcomes and Benefits: We expect to develop an assay that will detect changes in blood glycoproteins that will reflect how effective a particular therapy regimen has worked. Return of a non-cancer pattern over time will be expected in patients with known favorable treatment outcomes. However, an unchanged blood glycoprotein pattern would indicate tumor relapses. This type of assay could be used to monitor responses in real time during treatments, allowing faster determination of success or failure, giving doctors more options to individualize treatment plans for each patient.