TROP2 is a novel, potent stimulator of tumor growth and of metastatic spreading of human cancer

Background: Trop-2 is a transmembrane calcium signal transducer, that plays a role in cell-cell and cell-substrate adhesion. Material and methods: We investigated the expression pattern of Trop-2 by DNA array, SAGE, Northern and Western blotting, flow cytometry, confocal microscopy and IHC analysis in experimental systems and in man. We explored its functional role by overexpression or down-regulation and by directed mutagenesis in transformed and/or metastatic cells in vivo. Results: DNA array, EST, SAGE, RT-PCR and Northern blot analysis of human tumors revealed expression of the TROP2 gene in the vast majority of human cancers. A corresponding overexpression of the Trop-2 protein was demonstrated by a large scale IHC analysis of human tumors (1755 cases). Trop-2 was demonstrated to potently stimulate the growth of tumor cells, and its down-regulation by siRNA inhibited it. Deletion of the cytoplasmic region of Trop-2 abolished its growth stimulatory capacity, as mutagenesis of the S303 PKC phosphorylation site did. Proteomic and phosphoproteomic analysis demonstrated a Trop-2-dependent activation of PKC, FAK and Raf1, modulation of ERK, MEK and p38 MAPK, and upregulation of NF-B. In vivo imaging demonstrated a dynamic colocalization of PKC and Trop-2 in membrane ruffles and podosomes. Dominant negative PKC and PKC siRNAs selectively abolished the Trop-2-induced growth demonstrating that PKC plays a key role in Trop-2 signaling. Strikingly, comparative global gene expression analysis revealed that TROP2 was the only gene up-regulated across different metastatic models, tumor types and animal species. IHC analysis revealed a dramatic up-regulation in metastases from colon, stomach, breast and ovary tumors in man. To assess if Trop-2 plays a causal role in metastatic spreading, the metastatic potential of TROP2-transfected KM12SM colon cancer cells, orthotopically injected in nude mice, was assessed. TROP2-overexpressing cells indeed demonstrated a profoundly increased metastatic potential to the liver. Deletion of the HIKE domain of Trop-2 severely diminished, whereas that of the whole cytoplasmic region vastly increased metastatic diffusion, indicating the existence of metastatic enhancers and silencers in the Trop-2 cytoplasmic tail. Conclusions: Our findings demonstrate the existence of a previously unsuspected, strikingly widespread mechanism of stimulation of tumor growth and of metastatic spreading in man, and candidate Trop-2 for novel diagnostic and therapeutic procedures.