Measurements of Tumor Cell Autophagy Predict Invasiveness, Resistance to Chemotherapy, and Survival in Melanoma

Abstract

Purpose: Autophagy consists of lysosome-dependent degradation of cytoplasmic contents sequestered by autophagic vesicles (AV). The role of autophagy in determining tumor aggressiveness and response to therapy in melanoma was investigated in this study.Experimental Design: Autophagy was measured in tumor biopsies obtained from metastatic melanoma patients enrolled on a phase II trial of temozolomide and sorafenib and correlated to clinical outcome. These results were compared with autophagy measurements in aggressive and indolent melanoma cells grown in two-and three-dimensional (3D) culture and as xenograft tumors. The effects of autophagy inhibition with either hydroxychloroquine or inducible shRNA (short hairpin RNA) against the autophagy gene ATG5 were assessed in three-dimensional spheroids.Results: Patients whose tumors had a high autophagic index were less likely to respond to treatment and had a shorter survival compared with those with a low autophagic index. Differences in autophagy were less evident in aggressive and indolent melanoma cells grown in monolayer culture. In contrast, autophagy was increased in aggressive compared with indolent melanoma xenograft tumors. This difference was recapitulated when aggressive and indolent melanoma cells were grown as spheroids. Autophagy inhibition with either hydroxychloroquine or inducible shRNA against ATG5 resulted in cell death in aggressive melanoma spheroids, and significantly augmented temozolomide-induced cell death.Conclusions: Autophagy is a potential prognostic factor and therapeutic target in melanoma. Three dimensional culture mimics the tumor microenvironment better than monolayer culture and is an appropriate model for studying therapeutic combinations involving autophagy modulators. Autophagy inhibition should be tested clinically in patients with melanoma. Clin Cancer Res; 17(10); 3478-89. Ó2011 AACR.

Autophagy inhibitor Lys05 has single-agent antitumor activity and reproduces the phenotype of a genetic autophagy deficiency

“…Cell lines C8161 and PC-9 were maintained in DMEM, LN229, and 1205Lu in RPMI-1640, and both supplemented with 10% (vol/vol) FBS (Sigma), in an atmosphere of 5% CO 2 at 37°C. LN229 GFP-LC3 was generated as previously described (22).…”

“…Except for certain models of pancreatic cancer (21), in many animal tumor models, where high levels of autophagy are likely present in untreated tumors (4,22), treatment with single-agent HCQ does not impair tumor growth (23,24). To determine if a more potent autophagy inhibitor such as Lys05 could significantly impair tumor growth as a single agent, 1205Lu xenografts were generated in the flanks of nude mice.…”

Oncogenic BRAF induces chronic ER stress condition resulting in increased basal autophagy and apoptotic resistance of cutaneous melanoma

“…15,16 Dysregulation of autophagy has accordingly been postulated as a secondary event contributing to melanoma progression and, importantly, also has a key role in chemoresistance. [18][19][20] Autophagy is the principal catabolic process for the bulk degradation and recycling of aged/damaged cellular components, organelles and proteins through the formation of a double-membraned cytosolic vesicle able to wrap targeted material. The subsequent fusion with lysosomes and degradation of cargo provide nutrients in times of environmental stress, such as nutrient deprivation or hypoxia.…”

The Quinone Methide Aurin Is a Heat Shock Response Inducer That Causes Proteotoxic Stress and Noxa-dependent Apoptosis in Malignant Melanoma Cells

“…Dysregulation of proteotoxic stress has been observed in human melanoma tissue contributing to the notorious chemoresistance of metastatic melanoma cells. Cumulative evidence suggests the involvement of autophagic dysregulation in melanomagenesis and the emerging role of autophagy as a prognostic factor and therapeutic target in melanoma has been substantiated recently (21)(22)(23)(24)(25)(26). Moreover, pathological altera-* This work was supported, in whole or in part, by National Institutes of Health Grants R01CA122484, R03CA167580, R21CA166926, ES007091, ES006694, and Arizona Cancer Center Support Grant CA023074.…”