Abstract 441: Dynamic biochemical tissue analysis of selectin ligands present on tissue derived from solid tumors

Abstract

Abstract Identification and characterization of molecular markers on cancerous tissue can lead to novel diagnostics and prognostics for cancer. Markers of interest include sialofucosylated molecules such as sialyl Lewis X and sialyl Lewis A (CA19-9), which are expressed on both lipids and proteins, and are often upregulated on certain cancerous tissues. Mounting evidence suggests that circulating cancer cells use these molecules to bind E-, P- and L- selectin (adhesion molecules present on leukocytes, platelets, and endothelial cells), to facilitate dissemination indicating that the presence of selectin ligands may correlate with metastatic potential. The hallmark of selectin-mediated binding is “rolling,” i.e. adhesion governed by the unique kinetic and tensile properties of selectin-selectin ligand bonds occurring under dynamic (flow) conditions. At present, the analysis of pathological tissue (e.g. immunohistochemistry) is performed under static conditions that only reveal high affinity interactions between the probing molecule and the tissue. Traditional biochemical tissue analysis is woefully inadequate for investigating selectin ligands on cancerous tissue, as it does not allow a detailed exploration of ligand chemistry in situ under biophysical conditions fit for selectin binding. Recently, we established a new assay for probing tissue, termed dynamic biochemical tissue analysis (DBTA), in which the probing molecule is conjugated to microspheres that are contacted with tissue samples under fluid shear. In our current investigation, E-, P- and L-selectin microspheres rolled on colon cancer tissue microarrays in DBTA. The adhesive interactions appeared to be specific, as the microspheres were released from the tissue samples and no further adhesion events were observed when EDTA was perfused through the system. Additionally, the adhesion of the microspheres with a given tissue section was dependent on the selectin. E-selectin microspheres were the most promiscuous, binding at high levels to many of the tissue samples, while the adhesion of the P- and L-selectin microspheres was more select. For a given type of selectin microsphere, the observed adhesion was not homogeneous; it was localized on discrete regions of the tissue sample. Notably, HECA-452 staining (an antibody that recognizes sialyl Lewis X and related moieties) in conjunction with selectin microsphere rolling revealed that HECA-452 reactivity only partially correlated with selectin binding. This observation strongly suggests that DBTA yields tissue characterization that is distinct compared to a high affinity immunohistochemistry assay. In conclusion, DBTA provides a controlled environment in which the shear-dependent selectin-selectin ligand interactions may be observed, providing a unique characterization of cancerous tissue applicable to developing novel diagnostic and prognostic strategies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 441. doi:1538-7445.AM2012-441