From bench to bedside: Mass spectrometric identification of an alternatively spliced fibronectin domain strongly expressed in neovasculature of liver metastases

摘要

The perfusion-based chemical proteomics approach used in this study led to the identification of more than one hundred proteins exclusively found in liver metastasis specimens. Many of these proteins were annotated to be localized in the plasma membrane or to be secreted to the extracellular matrix, including CD98 heavy chain, collagen XIV, vitronectin, clusterin, biglycan and endoglin. Numerous proteins identified in the liver metastases specimens had previously been reported to be associated with angiogenesis. In particular, our mass spectrometric analysis revealed that the alternatively spliced extra-domain A of fibronectin is strongly expressed in the neovasculature of liver metastases, while being undetectable in normal healthy liver. On the basis of these results, we developed high affinity human antibodies against EDA in several formats (scFv and SIP). These antibodies were used to show EDA expression in the neovasculature of metastatic lesions and primary tumors of a large set of human cancer specimens while no staining for EDA was observed in normal tissues. Furthermore, in vivo targeting experiments to liver metastases and subcutaneous tumors were performed.