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Colorectal Cancer Cell Surface Protein Profiling Using an Antibody Microarray and Fluorescence Multiplexing
Published on: September 25, 2011
Targeting EpCAM expression via near-infrared fluorescent antibodies enables microscopic delineation of primary and
Bernd Uhl1,2,3,4,5, Dennis Eggert6, Bojan Smiljanov7,8
1Department of Otorhinolaryngology, TUM University Hospital, Ismaninger Str. 22, Munich, 81675, Germany. Bernd.Uhl@mri.tum.de.
Background:
In surgical treatment of head and neck squamous cell carcinoma (HNSCC), resection with adequate tumor-free margins is a fundamental challenge. Epithelial cell adhesion molecule (EpCAM/CD326) is substantially overexpressed in HNSCC representing a potential target for fluorescence-guided delineation of HNSCC.
Materials And Methods:
EpCAM expression was assessed in vitro on squamous cell carcinomas of the upper aerodigestive tract (SCC-UADT) and fibroblasts employing immunostaining with anti-EpCAM antibodies VU1D9 and MT201/adecatumumab (clinically validated). Whole tumor analyses of EpCAM expression were conducted in patient samples with primary and recurrent HNSCC. Confocal microscopy was used for EpCAM expression analyses upon immunostaining in cultured patient HNSCC resection samples using IRDye800CW-labeled MT201.
Results:
Immunostaining with VU1D9 and MT201 revealed a high, consistent, and specific expression of EpCAM on SCC-UADT in vitro. Human whole tumor analyses showed high and consistent EpCAM expression (primary: 9/9 > = 80%; recurrent: 6/9 > = 80%) and significantly elevated near-infrared fluorescence intensities for IRDye800CW-VU1D9 and IRDye800CW-MT201 in HNSCC as compared to non-malignant tissue (2-7 fold vs. mucosa; 6-20 fold vs. HNSCC-associated stroma). Translational postoperative immunostaining of live cultured patient HNSCC samples using the clinically validated IRDye800CW-MT201/adecatumumab enabled microscopical differentiation of HNSCC from adjacent non-malignant tissue (fluorescence intensity ratios HNSCC vs. mucosa: 7,66 ± 1,26; and vs. HNSCC-associated stroma: 50,97 ± 8,24)).
Conclusions:
IRDye800CW-labeled anti-EpCAM antibodies allow experimental microscopic delineation of HNSCC from non-malignant tissue. As a future outlook, these probes might improve tumor resection in near-infrared fluorescence-guided surgery of primary and recurrent HNSCC.
Insights
Epithelial cell adhesion molecule (EpCAM) is highly expressed in head and neck squamous cell carcinoma (HNSCC). IRDye800CW-labeled anti-EpCAM antibodies enable microscopic differentiation of HNSCC, potentially improving tumor resection in fluorescence-guided surgery.
Area of Science:
- Oncology
- Surgical Oncology
- Molecular Imaging
Background:
- Head and neck squamous cell carcinoma (HNSCC) surgical resection requires adequate tumor-free margins.
- Epithelial cell adhesion molecule (EpCAM/CD326) is overexpressed in HNSCC, making it a target for fluorescence-guided surgery.
Purpose of the Study:
- To evaluate the utility of EpCAM expression for delineating HNSCC.
- To assess the efficacy of IRDye800CW-labeled anti-EpCAM antibodies for microscopic visualization of HNSCC.
Main Methods:
- EpCAM expression was analyzed in vitro and in patient samples using immunostaining with anti-EpCAM antibodies (VU1D9, MT201/adecatumumab).
- Confocal microscopy and near-infrared (NIR) fluorescence imaging were employed on cultured HNSCC samples labeled with IRDye800CW-conjugated antibodies.
Main Results:
- Immunostaining confirmed high and specific EpCAM expression on HNSCC cells.
- Whole tumor analyses revealed consistent EpCAM expression in primary and recurrent HNSCC.
- Significantly elevated NIR fluorescence was observed in HNSCC compared to adjacent non-malignant tissues (mucosa and stroma).
- Microscopic differentiation of HNSCC from non-malignant tissue was achieved using IRDye800CW-MT201, with high fluorescence intensity ratios.
Conclusions:
- IRDye800CW-labeled anti-EpCAM antibodies facilitate experimental microscopic delineation of HNSCC.
- These probes hold potential for improving tumor resection margins in near-infrared fluorescence-guided surgery for HNSCC.
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