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Comparative Analysis of the Membrane Proteome of Closely Related Metastatic and Nonmetastatic Tumor Cells

¹ Department of Chemistry and Applied Biosciences, ETH Zurich, Zurich, Switzerland and ² Laboratory of Biology and Treatment of Metastasis, Department of Oncology, Mario Negri Institute for Pharmacological Research, Bergamo, Italy

Requests for reprints: Dario Neri, Department of Chemistry and Applied Biosciences, ETH Zurich, Institute of Pharmaceutical Sciences, Wolfgang-Pauli-Strasse 10, HCI G396.4, 8093 Zurich, Switzerland. Phone: 41-44-633-74-01; Fax: 41-44-633-13-58; E-mail: neri{at}pharma.ethz.ch.

Key Words: metastasis markers • chemical proteomics • mass spectrometry • subcellular protein localization • tumor targeting

The identification of proteins that are preferentially expressed on the membrane of metastatic tumor cells is of fundamental importance in cancer research. Here, we report the systematic comparison of the membrane proteome of two closely related murine teratocarcinoma cell lines (F9B9 and F9DR), of which only one (F9DR) is capable of forming liver metastases in vivo. The proteomic methodology used in this study featured the surface protein biotinylation on tumor cells followed by protein purification on streptavidin resin and relative quantification of corresponding tryptic peptides by mass spectrometric procedures. The study allowed the identification of 998 proteins and the determination of their relative abundance. Proteins previously known to be associated with metastatic spread were found to be either up-regulated (e.g., synaptojanin-2) or down-regulated (e.g., Ceacam1) in F9DR cells. A dramatic increase in abundance at the cell membrane was observed for a broad variety of proteins (e.g., high-mobility group protein B1), which were mainly thought to reside in intracellular compartments, a finding that was confirmed using confocal laser scanning microscopy and immunochemical analysis of cell cultures. Furthermore, we showed by microautoradiographic analysis that certain target proteins can readily be reached by intravenously administered radiolabeled antibodies. Finally, we showed that the most promising antigens for antibody-based pharmacodelivery approaches are strongly and selectively expressed on the surface of tumor cells in three different syngeneic mouse models of liver metastases. Taken together, our results indicate that the expression of intracellular proteins on the membrane of metastatic cells is a feature much more common than previously expected. [Cancer Res 2009;69(13):5406–14]