Cannabinoid action induces autophagy-mediated cell death through stimulation of ER stress in human glioma cells
María Salazar ... Patricia Boya, Guillermo Velasco
María Salazar ... Patricia Boya, Guillermo Velasco
Published May 1, 2009
Citation Information: J Clin Invest. 2009;119(5):1359-1372. doi:10.1172/JCI37948.
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Categories: Research Article Oncology

Cannabinoid action induces autophagy-mediated cell death through stimulation of ER stress in human glioma cells

Abstract

Autophagy can promote cell survival or cell death, but the molecular basis underlying its dual role in cancer remains obscure. Here we demonstrate that Δ9-tetrahydrocannabinol (THC), the main active component of marijuana, induces human glioma cell death through stimulation of autophagy. Our data indicate that THC induced ceramide accumulation and eukaryotic translation initiation factor 2α (eIF2α) phosphorylation and thereby activated an ER stress response that promoted autophagy via tribbles homolog 3–dependent (TRB3-dependent) inhibition of the Akt/mammalian target of rapamycin complex 1 (mTORC1) axis. We also showed that autophagy is upstream of apoptosis in cannabinoid-induced human and mouse cancer cell death and that activation of this pathway was necessary for the antitumor action of cannabinoids in vivo. These findings describe a mechanism by which THC can promote the autophagic death of human and mouse cancer cells and provide evidence that cannabinoid administration may be an effective therapeutic strategy for targeting human cancers.

Authors

María Salazar, Arkaitz Carracedo, Íñigo J. Salanueva, Sonia Hernández-Tiedra, Mar Lorente, Ainara Egia, Patricia Vázquez, Cristina Blázquez, Sofía Torres, Stephane García, Jonathan Nowak, Gian María Fimia, Mauro Piacentini, Francesco Cecconi, Pier Paolo Pandolfi, Luis González-Feria, Juan L. Iovanna, Manuel Guzmán, Patricia Boya, Guillermo Velasco

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Autophagy is upstream of apoptosis in cannabinoid-induced cancer cell de...

Figure 5

Autophagy is upstream of apoptosis in cannabinoid-induced cancer cell death.

(A) Effect of THC and the pan-caspase inhibitor ZVAD (10 μM) on the viability of Atg5+/+ and Atg5–/– MEFs (36 h; percentage of viable cells relative to the corresponding Atg5+/+ vehicle-treated cells, mean ± SD; n = 3). (B) Effect of THC on the apoptosis of Bax/Bak WT and Bax/Bak DKO MEFs as determined by cytofluorometric analysis of Annexin V/propidium iodide (PI) (24 h; mean ± SD; n = 3). The mean ± SD percentage of Annexin V–positive/PI-positive and Annexin V–positive, PI-negative cells is shown in the upper and lower corners, respectively. (C) Effect of THC on eIF2α phosphorylation (3 h; n = 3) and LC3 lipidation (18 h; n = 4) of Bax/Bak WT and DKO MEFs. (D) Left: Effect of THC on autophagy and apoptosis of U87MG cells transfected with siC or siATG1. Green bars, cells with LC3 dots; red bars, active caspase-3–positive cells; white bars, cells with both LC3 dots and active caspase-3 staining. Data correspond to the percentage of cells with LC3 dots (green bars), active caspase-3–positive cells (red bars), and cells with LC3 dots and active caspse-3 staining (white bars) relative to the total number of transfected cells at each time point (mean ± SD; n = 3). Right: Representative photomicrographs (36 h; scale bar: 20 μm). (E and F) Effect of THC on apoptosis (E; 24 h; n = 3) and loss of mitochondrial membrane potential as determined by DiOC6(3) staining (F; 24 h; n = 4) of Atg5+/+ and Atg5–/– MEFs. In E, the mean ± SD percentage of Annexin V–positive/PI-positive and Annexin V–positive, PI-negative cells is shown in the upper and lower corners, respectively. **P < 0.01 compared with THC-treated Atg5+/+ (A, E, and F) and Bax/Bak WT (B) MEFs and from THC-treated, siC-transfected cells (D).