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A thorough evaluation of the effect of plasma protein binding in the estimation of receptor concentration was performed for the paired-agents in this study. We are planning to evaluate ABY-029, an anti-epithelial growth factor receptor (EGFR) Affibody, and IRDye 700DX as a control agent. The plasma-dependent change in fluorescence intensity, percent binding, and in vivo distribution kinetics will be studied for each agent alone, and in combination. In this proceeding, the absorption, emission patterns for the targeted agent, ABY-029, measured by UV-Vis, fluorometer, and Pearl were shown. Initial studies indicate that binding to Bovine serum albumin (BSA), human serum albumin (HSA) and EGFR can introduce the Solvatochromic shift, which will change the absorption and emission pattern for ABY-029. Computational modeling will be performed to determine how each of these changes will affect the determined BP, and thus detection of tumors from normal tissue.
ABY-029 is an anti-EGFR Affibody molecule labeled with IRDye800CW that is currently under Phase 0 human trial for FGS. To date, several studies have been performed to evaluate ABY-029 signal intensity in untreated human sarcoma xenografts; however, many patients undergoing cancer surgery have received pre-operative radiation and/or chemotherapy, which can affect tissue properties and tumor molecule expression level. Determining the effects of radiation and chemotherapy exposure on fluorophore binding in sarcomas may influence best practices in implementing FGS for sarcoma.
In this project, fluorophore signal intensities in tumor and surrounding tissue were measured and compared to the receptor concentration determined by immunohistochemistry. Here, we report the result for one EGFR positive synovial sarcoma cell lines, SW982. Four groups of human dose equivalent therapies – control, radiation, chemotherapy (Doxorubicin) and radiation followed by chemotherapy – were given to the tumor-bearing mice. The difference between groups can be used to determine the effects of preoperative sarcoma therapies on EGFR expression, ABY-029 uptake, and optical properties of tissues.
Tumor implantation model for rapid testing of lymphatic dye uptake from paw to node in small animals
The goal of this work was to successfully deploy and test an intra-nodal cancer-cell injection model to enable planar fluorescence imaging of a clinically relevant blue dye, specifically methylene blue – used in the sentinel lymph node procedure – in normal and tumor-bearing animals, and subsequently segregate tumor-bearing from normal lymph nodes. This direct-injection based tumor model was employed in athymic rats (6 normal, 4 controls, 6 cancer-bearing), where luciferase-expressing breast cancer cells were injected into axillary lymph nodes. Tumor presence in nodes was confirmed by bioluminescence imaging before and after fluorescence imaging. Lymphatic uptake from the injection site (intradermal on forepaw) to lymph node was imaged at approximately 2 frames/minute. Large variability was observed within each cohort.
Quantifying receptor density in vivo using a dual probe approach with fluorescence molecular imaging
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