THROMBIN INCREASES THE METASTATIC POTENTIAL OF TUMOR-CELLS

Initial arrest of tumor cells in the microvasculature and their attach ment to the endothelium and subendothelial matrix (SEM) are essential prerequisites for metastasis to occur. Factors mediating these interac tions are viewed as important determinants of the tumor-cell metastati c phenotype. In this work we have studied the effects of thrombin, its analogs and its precursors on the adhesive properties and metastatic potential of tumor cells. We show that alpha-thrombin, the native form of the key coagulation enzyme, is capable of enhancing tumor-cell adh esion to both the endothelium and SEM components represented by fibron ectin. Subclotting, physiological concentrations of alpha-thrombin pro duced a 2- to 5-fold increase in tumor-cell adhesion. A bell-shaped do se-response curve was observed, with maximal effect at 0.1 U/ml. Maxim um effect occurred when cells were exposed to the agonist for 15 min a nd exposure for up to 4 hr resulted in enhanced tumor-cell adhesion. P rolonged incubation with thrombin resulted in a decline in the thrombi n-enhanced adhesion which reached unstimulated control levels by 24 hr . Thrombin precursors and active-site-inhibited thrombin analogs only had minimal adhesion-enhancing activity; nitro- and exosite-alpha-thro mbin, which retain a functional active site, mimicked, although to a l esser degree, the action of alpha-thrombin. Tumor-cell incubation with thrombin resulted in an upregulated cell-surface expression of the al pha(IIbk)beta3 integrin, a receptor mediating interactions between tum or cells and endothelial cells, and between tumor cells and SEM. Antib odies against alpha(IIb)beta3 integrin effectively inhibited thrombin- enhanced tumor-cell adhesion. Thrombin effects on tumor cells involved the PKC signal transduction pathway as thrombin-enhanced adhesion was inhibited by pre-incubation with PKC inhibitors and a transient PKC t ranslocation from cytosol to membrane was observed following thrombin challenge. In vivo, thrombin-treated tumor cells demonstrated a 2-fold increase in their lung-colonizing ability. In contrast to the adhesio n results, the metastasis-enhancing effects of alpha-thrombin were mim icked by a thrombin precursor (prothrombin) and thrombin analogs. (C) 1993 Wiley-Liss, Inc.