Inhibition of autophagy sensitizes malignant pleural mesothelioma cells to dual PI3K/mTOR inhibitors

Our plain-English summary of Inhibition of autophagy sensitizes malignant pleural mesothelioma cells to dual PI3K/mTOR inhibitors, by N. Echeverry, G. Ziltener, D. Barbone, et al (Cell Death & Disease, 2015).

The study

• Authors: N. Echeverry, G. Ziltener, D. Barbone, W. Weder, R.A. Stahel, V.C. Broaddus, E. Felley-Bosco
• Journal: Cell Death & Disease
• Published: 2015;6(5):e1757
• Cited by: 44 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Malignant pleural mesothelioma (MPM) originates in most of the cases from chronic inflammation of the mesothelium due to exposure to asbestos fibers. Given the limited effect of chemotherapy, a big effort is being made to find new treatment options. The PI3K/mTOR pathway was reported to be upregulated in MPM. We tested the cell growth inhibition properties of two dual PI3K/mTOR inhibitors NVP-BEZ235 and GDC-0980 on 19 MPM cell lines. We could identify resistant and sensitive lines; however, there was no correlation to the downregulation of PI3K/mTOR activity markers. As a result of mTOR inhibition, both drugs efficiently induced long-term autophagy but not cell death. Autophagy blockade by chloroquine in combination with the dual PI3K/mTOR inhibitors significantly induced caspase-independent cell death involving RIP1 in the sensitive cell line SPC212. Cell death in the resistant cell line Mero-82 was less pronounced, and it was not induced via RIP1-dependent mechanism, suggesting the involvement of RIP1 downstream effectors. Cell death induction was confirmed in 3D systems. Based on these results, we identify autophagy as one of the main mechanisms of cell death resistance against dual PI3K/mTOR inhibitors in MPM. As PI3K/mTOR inhibitors are under investigation in clinical trials, these results may help interpreting their outcome and suggest ways for intervention.

In plain English

This was a laboratory study of mesothelioma cancer cells grown in dishes. No patients were studied, and asbestos exposure was not examined. The authors tested two experimental drugs, NVP-BEZ235 and GDC-0980, on 19 malignant pleural mesothelioma cell lines. Both drugs block a growth signaling pathway called PI3K/mTOR. Sensitivity varied between lines (about 26-fold for NVP-BEZ235, up to 8-fold for GDC-0980). It did not match how strongly the drugs suppressed pathway markers. The drugs slowed growth and increased autophagy, a self-recycling process that helps cells survive stress. No cell death was seen at 72 hours. At 96 hours, the drugs alone caused some cell death, but death was significantly higher only with added chloroquine, which blocks autophagy. This added effect was larger in the sensitive line SPC212 than in the resistant line Mero-82. A blocker of the protein RIP1 significantly reduced this death in SPC212 but did not block it in Mero-82. Similar results were seen in three-dimensional cell clusters. The authors concluded that autophagy is one way mesothelioma cells resist these drugs, and that combining them with chloroquine deserves further study.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory (in vitro) study of human mesothelioma cell lines, in standard 2D culture and in 3D spheroids
• Who was studied: 19 malignant pleural mesothelioma cell lines, with detailed work on 6 lines and mainly on SPC212 (sensitive) and Mero-82 (resistant). A non-transformed mesothelial cell line (SDM104) was used for comparison. The work was done by laboratories in Zürich and San Francisco. No human subjects were studied.
• Exposure: Two dual PI3K/mTOR inhibitor drugs, NVP-BEZ235 (200 nM in functional assays) and GDC-0980 (500 nM), given alone or combined with the autophagy blocker chloroquine (20 μM). Asbestos exposure was not studied; asbestos is mentioned only as background on the cause of the disease.
• Main results:
  • Across the 19 cell lines, the drug concentration needed to inhibit growth by half differed about 26-fold between the most sensitive and most resistant lines for NVP-BEZ235, and by at most 8-fold for GDC-0980.
  • In six representative sensitive and resistant lines, the drugs reduced mTORC1 pathway markers to a similar degree, so sensitivity did not correspond to pathway suppression.
  • In SPC212 and Mero-82 after 72 hours of treatment, the drugs caused growth arrest, which was more pronounced in sensitive lines, but no cell death was observed.
  • Both drugs induced sustained autophagy in SPC212 and Mero-82 after 72 hours.
  • Almost all mesothelioma cell lines had high baseline autophagy compared with the non-transformed mesothelial line SDM104.
  • Silencing the autophagy gene Atg5 significantly impaired growth of both SPC212 and Mero-82 cells.
  • Cell death with the combination of chloroquine and the drugs was detected only after 96 hours.
  • In the 96-hour Annexin V/PI assay, both drugs alone induced some cell death in SPC212 and Mero-82 cells.
  • Cell death was significantly higher only when chloroquine was added to the drugs. The added effect was bigger in sensitive SPC212 than in resistant Mero-82.
  • A broad caspase inhibitor (Z-VAD-FMK) did not prevent the cell death.
  • The RIP1 inhibitor Necrostatin-1 significantly blocked the combination-induced cell death in SPC212 but not in Mero-82.
  • In 3D spheroids of both cell lines, chloroquine plus the drugs had a significant additive effect on lowering ATP, a measure of cell viability.
  • Inhibiting autophagy did not make the normal SDM104 mesothelial cells more sensitive to the drugs.
  • Adding the MEK inhibitor U0126 to GDC-0980 and chloroquine did not induce cell death.
• The authors' conclusion: The authors concluded that PI3K/mTOR inhibitors increase autophagy levels in MPM cells. They concluded that combining these drugs with chloroquine should be further explored, because blocking autophagy blocks this resistance mechanism.
• Limitations the authors state:
  • Because few lines expressed proven wild-type NF2, the authors could not draw conclusions about NF2 status and drug sensitivity.
  • Atg5 silencing itself impaired cell growth, so the authors could not test the drugs in those cells.
  • The mechanisms behind the cells' dependence on autophagy were not studied.

Topics

The National Library of Medicine indexes this study under subjects our site covers: Lung cancer, Mesothelioma.

Medical terms in this study, explained in our Lexicon: Pleural Mesothelioma, Chronic Inflammation, Mesothelium, Chemotherapy, Clinical Trial.

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