Evaluating breast cancer stem cell response to antiangiogenic therapy
Abstract
Angiogenic inhibitors function by blocking tumor cell signals used to recruit host tissue vasculature to the tumor site, thereby depriving the cancer of the nutriment needed for further expansion. The development and implementation of angiogenic inhibitors in conjunction with standard chemotherapy agents has increased progression-free survival but not overall patient survival. It is hypothesized that chronic exposure to large doses of AAT drugs worsens hypoxic conditions within the tumor mass, selectively stimulating aggressive cancer stem cell populations to grow and proliferate. In this study, the expression of the CSC biomarkers ALDH1, DLL1, and EpCAM were evaluated in breast cancer tumors grown in mouse models for varying doses of angiogenic inhibitor DC101 using a threshold analysis technique developed in-house. SUM149 triple-negative breast cancer cells were grown in athymic nude mice and administered either 10mg/kg, 40mg/kg, or 120mg/kg DC101, corresponding to “Low Dose”, “Medium Dose”, and “High Dose”, respectively. Following a period of several days, the tumors were harvested, sectioned into slices at specific depths, and stained for one of each biomarker. Stained sections were scanned into a computer, where images were subjected to a series of coded protocols written in-house for Matlab or IDL. Images were segmented to remove non-target background pixels and co-registered to a series of static images to allow for comparative analysis. Biomarker-specific thresholds were applied to separate biomarker-positive image pixels from those pixels deficient in the biomarker stain. Pixel values were counted for both the control tumor slides and those having received DC101 exposure. Values were compared to evaluate changes in biomarker expression with variations in dose concentration. Pixel values were also compared between corresponding slices stained for different biomarkers in order to determine the prevalence of spatially-overlapping regions. Experimental results demonstrate a significant (p < 0.05) decrease in ALDH1 and DLL1 biomarker expression for the MD groups compared to controls, and elevated expression in the HD group compared to the LD and MD groups for the same biomarkers. Changes were most dramatic in the expression of the DLL1 biomarker. EpCAM expression did not vary significantly (p < 0.05) with dose. Variations in overlap between ALDH1 and EpCAM suggest that expression of the two biomarkers may be linked, while DLL1 overlap data suggests that DLL1 expression is independent of ALDH1 and EpCAM. Through a combination of perfusion imaging and biomarker expression, our data suggest that “medium dose” concentrations of 40mg/kg of DC101 can affect normalization of tumor vasculature within our mouse models, thereby alleviating hypoxia within the tumor microenvironment. While our ALDH1 quantification results provide some validation for our technique, future goals should focus on further validation through additional quantification of known biomarkers, the incorporation of PCA, and the use of the variance factor in the co-registration script to assess accuracy of the co-registration process.
Degree
M.S.
Advisors
Stantz, Purdue University.
Subject Area
Health sciences|Physics|Biophysics
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