Abstract B150: Cathepsin L targeting abrogates tumor microenvironment enhanced metastatic potential in prostate cancer cells.

Metastatic dissemination of tumor cells remains the primary cause of therapeutic failure and high mortality in men with prostate cancer. The development of novel therapeutic strategies that impair the metastatic process therefore is critical to improving prostate cancer patient survival. Tumor cells rely on proteolytic enzymes for successful establishment of metastases. Cathepsin L (CTSL) is a lysosomal cysteine protease that is up regulated in a wide range of human cancers including prostate cancer. The elevated secretion of CTSL by aggressively metastasizing tumor cells plays a key role in metastatic spread of tumor cells through proteolytic degradation of extracellular matrix and basement membrane components. Thus intervention strategies targeting CTSL may provide a novel anti-metastatic therapeutic approach to improve treatment outcome. This concept is supported by recent studies in our laboratory which showed that treatment with the specific small molecule CTSL inhibitor 3-bromophenyl-3-hydroxyphenyl-ketone thiosemicarbazone (KGP94) significantly impaired the migratory and invasive potential of metastatic prostate cancer cells. Clinical and experimental observations emphasize that pathophysiological tumor microenvironments characterized by hypoxia and acidic pH augment metastatic aggressiveness. Thus, the goal of the present studies was to explore the role of CTSL in tumor microenvironment-triggered elevation of invasiveness and to elucidate whether the administration of KGP94 could inhibit the metastatic phenotype in prostate cancer cells. Exposure to 1% oxygen and pH values of 6.8 and 6.4, enhanced the invasive phenotype of both the human prostate cancer cell line PC-3 and its highly metastatic subline PC-3-ML cells 1.5 to 2 fold. Treatment with 10 and 25μM KGP94 suppressed the heightened invasive potential (33% inhibition at 10μM and 60% inhibition at 25μM in PC-3 ML cells). Interestingly upon KGP94 treatment, invasiveness of these cells reduced below basal invasion levels observed under normoxic conditions. Aberrant microenvironment induced enhancement of PC-3 and PC-3-ML invasive potential was attributable to elevated activity of secreted CTSL. Molecular assessment revealed that the observed increase in CTSL activity was a consequence of exocytosis of lysosomes containing active CTSL into the extracellular milieu. Lysosomal/endosomal marker LAMP-1immunostaining revealed that in contrast to the peri-nuclear localization observed in control cells, lysosomes in cells exposed to low oxygen tension and pH traffic towards the cell periphery. Observations based on immunostaining were confirmed through biochemical quantification of the release of lysosomal enzyme β-Hexosaminidase into the media after hypoxic or acidic exposures. In conclusion, our findings suggest that hypoxia and acidosis triggered elevation of PC-3 and PC-3-ML invasive potential is associated with CTSL activation. KGP94 mediated CTSL inhibition thus effectively impaired the pathophysiological tumor microenvironment induced enhancement of the metastatic phenotype of prostate cancer cells. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr B150.