Targeting ER stress–induced autophagy overcomes BRAF inhibitor resistance in melanoma
Xiao-Hong Ma, … , Constantinos Koumenis, Ravi K. Amaravadi
Xiao-Hong Ma, … , Constantinos Koumenis, Ravi K. Amaravadi
Published February 24, 2014
Citation Information: J Clin Invest. 2014;124(3):1406-1417. https://doi.org/10.1172/JCI70454.
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Research Article

Targeting ER stress–induced autophagy overcomes BRAF inhibitor resistance in melanoma

Abstract

Melanomas that result from mutations in the gene encoding BRAF often become resistant to BRAF inhibition (BRAFi), with multiple mechanisms contributing to resistance. While therapy-induced autophagy promotes resistance to a number of therapies, especially those that target PI3K/mTOR signaling, its role as an adaptive resistance mechanism to BRAFi is not well characterized. Using tumor biopsies from BRAFV600E melanoma patients treated either with BRAFi or with combined BRAF and MEK inhibition, we found that BRAFi-resistant tumors had increased levels of autophagy compared with baseline. Patients with higher levels of therapy-induced autophagy had drastically lower response rates to BRAFi and a shorter duration of progression-free survival. In BRAFV600E melanoma cell lines, BRAFi or BRAF/MEK inhibition induced cytoprotective autophagy, and autophagy inhibition enhanced BRAFi-induced cell death. Shortly after BRAF inhibitor treatment in melanoma cell lines, mutant BRAF bound the ER stress gatekeeper GRP78, which rapidly expanded the ER. Disassociation of GRP78 from the PKR-like ER-kinase (PERK) promoted a PERK-dependent ER stress response that subsequently activated cytoprotective autophagy. Combined BRAF and autophagy inhibition promoted tumor regression in BRAFi-resistant xenografts. These data identify a molecular pathway for drug resistance connecting BRAFi, the ER stress response, and autophagy and provide a rationale for combination approaches targeting this resistance pathway.

Authors

Xiao-Hong Ma, Sheng-Fu Piao, Souvik Dey, Quentin Mcafee, Giorgos Karakousis, Jessie Villanueva, Lori S. Hart, Samuel Levi, Janice Hu, Gao Zhang, Rossitza Lazova, Vincent Klump, John M. Pawelek, Xiaowei Xu, Wei Xu, Lynn M. Schuchter, Michael A. Davies, Meenhard Herlyn, Jeffrey Winkler, Constantinos Koumenis, Ravi K. Amaravadi

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Figure 3

Autophagy inhibition augments growth impairment associated with BRAFi.

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Autophagy inhibition augments growth impairment associated with BRAFi. (...
(A) Percent growth inhibition (mean ± SEM) compared with control for 1 μM PLX4720 treatment with or without 10 μM HCQ (left), or for 1 μM PLX4720 treatment of MEL624shNT, MEL624shATG5, A375PshNT, and A375PshATG5 clones (right). (B) 14 day clonogenic growth assays for BRAFi-sensitive A375P and BRAFi-resistant MEL1617R and MEL624. Cells were plated in triplicate and immediately treated with PBS vehicle, 500 nM PLX4720, 5 μM HCQ, or the combination. Colony counts (mean ± SEM) are presented. (C) 72-hour MTT assay was conducted after treating the indicated cell lines with PBS, 1 μM PLX4720, 3 μM Lys05, or the combination. Results are mean ± SEM normalized to control. (D) MEL624 and A375P cells were treated with 30 nM GSK2118436, 20 nM GSK1120212, 10 μM HCQ, and the indicated combinations. 72-hour MTT (mean ± SEM) was performed. *P < 0.05.