Tumor cells do not constitutively exhibit invasive activity, but rather, can be transiently induced to adhere and form lesions. We report here that the expression of seprase, a dominant EDTA-resistant gelatinase in malignant tumors, is dependent on tumor cell exposure to type I collagen gel (TICg). The induced seprase expression of ovarian tumor cells influences their collagen contraction and invasion capability. Importantly, tumor cells with reduced seprase expression, due to manipulation by RNA interference, showed a reduction of TICg contraction in the gel contractility assay, inhibition of tumor cell invasion through TICg as shown by a transwell migration assay and inhibition of peritoneal membrane tumor lesion in a mouse model. In addition, mAb C27, an antibody against beta1 integrin, which blocks cellular avidity to TICg, can induce seprase RNA expression and promote the invasive phenotype and metastatic potential of ovarian tumor cells. Thus, collagenous matrices in the tumor cell niche induce the expression of seprase and initiate tumor invasion and metastatic cascades.