Functional inhibition of heat shock protein 70 by VER-155008 suppresses pleural mesothelioma cell proliferation via an autophagy mechanism

Our plain-English summary of Functional inhibition of heat shock protein 70 by VER-155008 suppresses pleural mesothelioma cell proliferation via an autophagy mechanism, by K. Sakai, M. Inoue, S. Mikami, et al (Thoracic Cancer, 2021).

The study

• Authors: K. Sakai, M. Inoue, S. Mikami, H. Nishimura, Y. Kuwabara, A. Kojima, M. Toda, Y. Ogawa-Kobayashi, S. Kikuchi, Y. Hirata, Y. Mikami-Saito, H. Kyoyama, G. Moriyama, M. Shiibashi, M. Seike, A. Gemma, K. Uematsu
• Journal: Thoracic Cancer
• Published: 2021;12(4):491-503
• Cited by: 20 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Background: Pleural mesothelioma, a devastating asbestos-associated malignancy, urgently requires a novel effective therapy. Heat shock protein 70 (HSP70), which is synthesized in the cell response to protein damage, is expected to be a new target for antitumor treatment. In addition to its well-known protein refolding function, HSP70 regulates cell proliferation through different pathways, including PI3K/AKT/mTOR, and autophagy in malignant cells. In this study, we attempted to clarify the effects of VER-155008, an HSP70 inhibitor, on pleural mesothelioma.

Methods: Human pleural mesothelioma cell lines 211H, H2452 and H28 were cultured with VER-155008, and protein expression, cell proliferation, colony formation, cell cycle, synergistic effect with cisplatin, and autophagy induction were analyzed.

Results: In mesothelioma cell lines, VER-155008 (5.0 μM or more) inhibited cell growth and colony formation, accompanied by G1 cell cycle arrest. According to western blot analysis, VER-155008 reduced p-AKT expression. However, VER-155008 failed to show a synergistic effect with cisplatin on cell growth. Mesothelioma cells transfected with the novel plasmid pMRX-IP-GFP-LC3-RFP-LC3ΔG, which was developed for the quantitative and statistical estimation of macroautophagy, showed enhanced macroautophagy upon treatment with VER-155008 and gefitinib which is an EGFR-tyrosine kinase inhibitor. In addition, fetal bovine serum deprivation induced macroautophagy was further enhanced by VER-155008.

Conclusions: On the basis of these results, functional HSP70 inhibition by VER-155008 suppressed cell growth in pleural mesothelioma cells, accompanied by enhanced macroautophagy. HSP70 inhibition is thus expected to become a new strategy for treating mesothelioma.

Key Points: Significant findings of the study In pleural mesothelioma cells, inhibition of HSP70 function by VER-155008 suppressed cell proliferation accompanied by induction of autophagy which was synergistically enhanced under the starvation condition, whereas gefitinib, an EGFR-TKI, did not show the same synergistic effect in autophagy. What this study adds The inhibition of HSP70 induced autophagy and suppressed cell proliferation in mesothelioma cells.

© 2020 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.

In plain English

This was a laboratory study of three human pleural mesothelioma cell lines (211H, H2452 and H28). It did not involve patients and did not study asbestos exposure. The authors treated the cells with VER-155008, a drug that blocks the function of heat shock protein 70 (HSP70). They used it alone or with the chemotherapy drug cisplatin or the drug gefitinib. VER-155008 at 5.0 μM or more reduced cell viability in all three lines, and at 20.0 μM it inhibited colony formation. It increased the share of cells in the G1 phase of the cell cycle in 211H and H28 cells but not in H2452 cells. It also lowered phosphorylated AKT, a growth-signaling protein. Adding cisplatin did not enhance the growth suppression. Adding gefitinib did not enhance it in 211H and H2452 cells. In a fluorescent marker test, VER-155008 increased macroautophagy, a process in which cells break down their own contents, and the effect was stronger when cells were deprived of serum. The authors concluded that HSP70 inhibition suppressed mesothelioma cell line growth. They said it is still unclear whether this autophagy protects or harms the cells.

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) experiment on human pleural mesothelioma cell lines
• Who was studied: Three human pleural mesothelioma cell lines: 211H, H2452 and H28 (no human subjects)
• Exposure: No asbestos exposure was studied. The cells were treated with the HSP70 inhibitor VER-155008 (0.5 to 40.0 μM), alone or combined with cisplatin (0.5 to 5.0 μM) or gefitinib (0.5 to 5.0 μM), and with or without fetal bovine serum (FBS) deprivation.
• Main results:
  • VER-155008 reduced cell viability. 211H viability was decreased by 5.0 μM after 24 hours. H2452 and H28 viability was suppressed by 5.0 μM after 48 hours. All lines were suppressed by 10.0 to 40.0 μM. IC50 values at 72 hours were 2.2 μM (211H), 1.5 μM (H2452) and 3.1 μM (H28).
  • At 20.0 μM, VER-155008 inhibited colony formation in all three cell lines.
  • After 48 hours with 20.0 μM VER-155008, the G1 phase cell population rose in 211H cells (82.7 ± 0.9% vs 69.9 ± 3.5% with DMSO control, P = 0.003871). It also rose in H28 cells (85.2 ± 3.3% vs 77.0 ± 2.3%, P = 0.008483).
  • In H2452 cells the G1 phase population was not altered (70.0 ± 1.2% with VER-155008 vs 71.3 ± 4.6% with DMSO).
  • In all three cell lines, phosphorylated AKT decreased after VER-155008 while total AKT was unchanged.
  • Combining VER-155008 with cisplatin did not produce the expected additive effect on growth suppression.
  • Gefitinib did not adequately suppress growth. Adding VER-155008 did not enhance growth inhibition in 211H and H2452 cells, while H28 cells tended toward synergistic suppression.
  • In the fluorescent plasmid system, VER-155008 induced macroautophagy in serum-supplemented conditions, and the effect was significantly enhanced under serum deprivation.
  • Under serum deprivation, gefitinib did not significantly change the macroautophagy measure compared with control (P = 0.0581 for 211H, 0.248053 for H2452 and 0.0604 for H28).
  • Compared with gefitinib alone, the combination of VER-155008 and gefitinib significantly lowered the GFP-to-RFP signal ratio (indicating more macroautophagy).
• The authors' conclusion: The authors concluded that inhibiting HSP70 with VER-155008 disrupts the PI3K/AKT/mTOR pathway and suppresses mesothelioma cell line proliferation by inducing G1 cell cycle arrest.
• Limitations the authors state:
  • The authors could not determine whether the induced macroautophagy protects the cells or leads to their death.
  • The unchanged LAMP-2A result (a marker of chaperone-mediated autophagy) may reflect limited sensitivity of the detection methods.
  • The authors stated that conventional western blot analysis is inadequate to measure autophagy progression.

Topics

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

Medical terms in this study, explained in our Lexicon: Pleural Mesothelioma, Synergistic Effect, Cisplatin, Mesothelioma, EGFR, Tyrosine Kinase Inhibitor.

Related studies

• Combined inhibition of MET and EGFR suppresses proliferation of malignant mesothelioma cells — K. Kawaguchi, H. Murakami, et al. (2009)
• LIM-domain protein AJUBA suppresses malignant mesothelioma cell proliferation via Hippo signaling cascade — I. Tanaka, H. Osada, et al. (2015)
• The flavonoid resveratrol suppresses growth of human malignant pleural mesothelioma cells through direct inhibition of specificity protein 1 — K.A. Lee, Y.J. Lee, et al. (2012)
• Inhibition of autophagy sensitizes malignant pleural mesothelioma cells to dual PI3K/mTOR inhibitors — N. Echeverry, G. Ziltener, et al. (2015)
• Functional Genomic Screen in Mesothelioma Reveals that Loss of Function of BRCA1-Associated Protein 1 Induces Chemoresistance to Ribonucleotide Reductase Inhibition — A. Okonska, S. Bühler, et al. (2020)
• Effect of syndecan-1 overexpression on mesenchymal tumour cell proliferation with focus on different functional domains — F. Zong, E. Fthenou, et al. (2010)

Browse more research in Thoracic Cancer, or return to medical research.