Abstract P6-14-08: Lucanthone Inhibits Autophagy and Promotes Cathepsin D-Mediated Apoptosis in Breast Cancer Cells
Authors: Carew JS , Espitia CM , Esquivel II JA , Mahalingam D , Kelly KR , Reddy G , Giles FJ , Nawrocki ST. 
Published: April 2011

Abstract
Autophagy is an evolutionarily conserved degradation pathway that eliminates certain proteins, defective organelles, and protein aggregates in order to maintain protein homeostasis and prevent abnormal cells from replicating. However, the autophagy machinery may also be utilized to generate ATP from organelles and other cellular components to promote survival under metabolic/hypoxic stress conditions. Since nutrient and oxygen deprivation are hallmark characteristics of the tumor microenvironment, disrupting autophagic degradation may be a promising approach to selectively kill cancer cells. Chloroquine and its derivatives are the only clinically relevant autophagic inhibitors that are currently being evaluated for their potential applications in cancer therapy. However, these drugs are known to cause ocular toxicity, especially irreversible retinopathy, and this underscores the need for additional inhibitors of autophagy with more favorable therapeutic indices. Lucanthone is an anti-schistome agent that is being investigated for cancer therapy as a sensitizer to chemotherapy and radiation. Here we report a novel mechanism of action for lucanthone that is characterized by the disruption of lysosomal function, inhibition of autophagy, and induction of apoptosis. Exposure to lucanthone resulted in the processing and recruitment of microtubule-associated protein 1 light chain 3 (LC3) to autophagosomes, but induced lysosomal membrane permeabilization and impaired autophagic degradation as revealed by electron microscopy and the accumulation of p62/SQSTM1. Importantly, lucanthone displayed more potent activity against a panel of breast cancer cell lines compared to chloroquine. Cathepsin D is a lysosomal protease that functions to promote apoptosis following its release into the cytosol upon lyososomal disruption. In order to elucidate the mechanism by which lucanthone induces apoptosis in cancer cells, we investigated the potential effects of lucanthone on cathepsin D expression and subcellular localization. As expected based on its effects on the integrity of lysosomes, lucanthone treatment stimulated a highly significant increase in cytosolic cathepsin D expression, which was associated with apoptosis induction. Accordingly, cathepsin D-targeted siRNA blocked the ability of lucanthone to stimulate its expression and significantly reduced lucanthone-mediated apoptosis. Subsequent experiments revealed that the anticancer activity of lucanthone occurred through a mechanism independent of the functional status of the p53 tumor suppressor. Considering that p53 is functionally inactivated in the majority of cancers, the ability of lucanthone to retain anticancer activity under p53 deficient conditions could have important clinical implications. Collectively, this study establishes a novel mechanism of action for lucanthone as an autophagy inhibitor and provides a rationale for its use in combination with anticancer agents that induce this important survival pathway.
Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P6-14-08.