On the TRAIL to Overcome BRAF-Inhibitor Resistance Peter Geserick, Meenhard Herlyn, Martin Leverkus  Journal of Investigative Dermatology  Volume 134, Issue 2, Pages 315-318 (February 2014) DOI: 10.1038/jid.2013.348 Copyright © 2014 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 The role of RAF-inhibitor-induced signaling for the crosstalk of extrinsic and intrinsic cell death signaling pathway in melanomas. The influence of the pan-RAF inhibitor (L-779450) and BRAF-specific inhibitor (vemurafenib) for extrinsic and intrinsic cell death pathways is marked in blue color. Consequences of this treatment are marked in red color. Treatment with the pan-RAF-inhibitor L-779450 inhibits both wild-type and mutant BRAF, whereas vemurafenib suppresses only BRAFV600E. Treatment with RAF inhibitors results in (a) the upregulation of LC3 (autophagy), and (b) inhibition of extracellular signal–regulated kinase (ERK) phosphorylation and resulting upregulation of Bim without direct cell death induction (“priming” of melanoma cells to death). Activation of tumor necrosis factor–related apoptosis-inducing ligand (TRAIL) death receptors results in procaspase-8 cleavage, subsequent Bid cleavage, and further activation of Bim. This is sufficient to ultimately induce cell death by the inhibition of antiapoptotic Bcl-2 proteins, and subsequent Bax, but not Bak activation. This is sufficient for MOMP, loss of mitochondrial membrane potential (Ψ), and release of cytochrome c and apoptosis-inducing factor. In turn, caspase-9 activation in the apoptosome complex as well as release of smac that suppresses X-linked inhibitor of apoptosis protein leads to melanoma cell death. Either TRAIL-mediated activation of caspase-8 or mitochondria-facilitated caspase-9 activation (by Bim-facilitated MOMP) leads to caspase-3 activation and apoptosis execution. The role of the caspase-8 inhibitor cellular FLICE-like inhibitory protein and the impact of BRAF for the regulation of autophagy needs to be substantiated by further experiments but represents an intriguing twist of BRAF-inhibitor resistance in melanoma. However, acute interference with RAF signaling seems to modulate the intrinsic signaling pathway directly, opening a “window of opportunity” to overcome resistance mechanisms by combining BRAF inhibitors with death receptor ligands, autophagy inhibitors, or other compounds that attack the mitochondria and therefore overcome the “primed to death” status of melanoma cells and effectively kill them. MAPK, mitogen-activated protein kinase; MEK, MAPK/ERK kinase. ?, indicates currently unknown interactions suggested by Berger et al., (2014). Journal of Investigative Dermatology 2014 134, 315-318DOI: (10.1038/jid.2013.348) Copyright © 2014 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 The TRAIL to overcome BRAF-inhibitor resistance: BRAF inhibitor (vemurafenib), pan-RAF inhibitor (L-779450), and MEK inhibitor carry a high clinically measurable potential for melanoma therapy. However, all inhibitors induce only low levels of cell death and rather prime the melanoma cells to cell death. RAF inhibitors suppress cell proliferation substantially but cannot trigger melanoma cell death sufficiently. Without further death stimuli, long-term stimulation with RAF inhibitors therefore promotes development of secondary resistance by alternative activation of other MAPK signaling pathways such as ARAF, NRAS, and CRAF. Furthermore, RAF-resistant melanomas circumvent the cell death priming event by the induction of autophagy. Both reactivation of alternative MAPKs as well as activation of the cellular recycling process of autophagy during RAF-inhibitor treatment can lead to increased melanoma remission. We speculate that RAF and MEK inhibitors as cell death priming compounds that are rapidly followed by direct cell death stimuli such as TRAIL or BH3-mimetics may prove to be highly beneficial for melanomas. Melanoma cells that survive such combination therapies may benefit from combination treatment with the autophagy-inhibitor hydroxychloroquine or with pan-RAF inhibitors that target all MAPK. The overall goal of such therapeutic intervention is to overcome secondary resistance by strong and rapid elimination of all tumor cells without targeting primary cells, facilitated by the use of tumor-specific targeting of mutant BRAF. AIF, apoptosis-inducing factor; cFLIP, cellular FLICE-like inhibitory protein; DISC, death-inducing signaling complex; MAPK, mitogen-activated protein kinase; MEK, MAPK/ERK kinase; RIP1, receptor-interacting protein 1; TRAIL, tumor necrosis factor–related apoptosis-inducing ligand; XIAP, X-linked inhibitor of apoptosis protein. Journal of Investigative Dermatology 2014 134, 315-318DOI: (10.1038/jid.2013.348) Copyright © 2014 The Society for Investigative Dermatology, Inc Terms and Conditions

Journal of Investigative Dermatology 2014 134, 315-318DOI: (10 Journal of Investigative Dermatology 2014 134, 315-318DOI: (10.1038/jid.2013.348) Copyright © 2014 The Society for Investigative Dermatology, Inc Terms and Conditions