Sensitization of melanoma cells for death ligand-induced apoptosis by an indirubin derivative--Enhancement of both extrinsic and intrinsic apoptosis pathways.

Until today effective therapies are lacking for metastatic melanoma. The death ligand TRAIL appears as promising in cancer treatment; however, melanoma cells reveal both preexisting and inducible TRAIL resistance. Here, we present evidence that the recently described indirubin derivative 8-Rha-β enhances melanoma cell sensitivity for death ligands and overcomes resistance to TRAIL and CD95 agonists. Indirubin is known from traditional Chinese medicine and is a potent kinase inhibitor. Unraveling of apoptotic signaling pathways revealed that TRAIL resulted in a quick (within 8h) downregulation of both agonistic TRAIL receptors DR4 and DR5, in a kind of negative feed-back loop. Treatment with indirubin, however, mediated upregulation of both receptors, thus compensating this negative feed-back loop by TRAIL. Furthermore, indirubin activated intrinsic apoptosis pathways, seen in loss of mitochondrial membrane potential and release of cytochrome c. The mitochondrial response appeared as related to upregulation of Bax and Bad and to downregulation of Mcl-1. Remarkably, indirubin in combination with TRAIL was also able to overcome apoptosis resistance due to ectopic Bcl-2 overexpression. The tumor suppressor p53 appeared as master regulator of these propapoptotic changes and is the transactivator of proapoptotic proteins which was upregulated by indirubin. Taking into account the physiological role of death ligands in immune surveillance, sensitization of melanoma cells for death ligands may be supportive for an anti-tumor immune response. Furthermore, combinations with kinase inhibitors, such as indirubin 8-Rha-β may help for a breakthrough of TRAIL-mediated strategies in melanoma.