Vorinostat eliminates multicellular resistance of mesothelioma 3D spheroids via restoration of Noxa expression

Our plain-English summary of Vorinostat eliminates multicellular resistance of mesothelioma 3D spheroids via restoration of Noxa expression, by D. Barbone, P. Cheung, S. Battula, et al (PloS One, 2012).

The study

• Authors: D. Barbone, P. Cheung, S. Battula, S. Busacca, S.G. Gray, D.B. Longley, R. Bueno, D.J. Sugarbaker, D.A. Fennell, V.C. Broaddus
• Journal: PloS One
• Published: 2012;7(12):e52753
• Cited by: 29 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

When grown in 3D cultures as spheroids, mesothelioma cells acquire a multicellular resistance to apoptosis that resembles that of solid tumors. We have previously found that resistance to the proteasome inhibitor bortezomib in 3D can be explained by a lack of upregulation of Noxa, the pro-apoptotic BH3 sensitizer that acts via displacement of the Bak/Bax-activator BH3-only protein, Bim. We hypothesized that the histone deacetylase inhibitor vorinostat might reverse this block to Noxa upregulation in 3D. Indeed, we found that vorinostat effectively restored upregulation of Noxa protein and message and abolished multicellular resistance to bortezomib in the 3D spheroids. The ability of vorinostat to reverse resistance was ablated by knockdown of Noxa or Bim, confirming the essential role of the Noxa/Bim axis in the response to vorinostat. Addition of vorinostat similarly increased the apoptotic response to bortezomib in another 3D model, the tumor fragment spheroid, which is grown from human mesothelioma ex vivo. In addition to its benefit when used with bortezomib, vorinostat also enhanced the response to cisplatin plus pemetrexed, as shown in both 3D models. Our results using clinically relevant 3D models show that the manipulation of the core apoptotic repertoire may improve the chemosensitivity of mesothelioma. Whereas neither vorinostat nor bortezomib alone has been clinically effective in mesothelioma, vorinostat may undermine chemoresistance to bortezomib and to other therapies thereby providing a rationale for combinatorial strategies.

In plain English

This laboratory study tested whether vorinostat could overcome mesothelioma cells' resistance to cell death (apoptosis) when grown as three-dimensional clusters (spheroids). It did not study asbestos exposure or patients. The authors used four mesothelioma cell lines and tumor fragment spheroids grown from six surgically removed mesothelioma tumors. Spheroids from all four lines resisted the drug bortezomib; adding vorinostat restored apoptosis to the same or greater level as in flat cell layers, though vorinostat alone did not induce apoptosis. With the combination, spheroid Noxa message rose as much as in flat layers. Blocking Noxa, or its partner protein Bim, stopped vorinostat from increasing cell death. In tumor fragment spheroids, vorinostat alone, bortezomib alone, and cisplatin plus pemetrexed together each failed to significantly increase apoptosis. Vorinostat combined with bortezomib, or with cisplatin plus pemetrexed, significantly increased apoptosis in samples from each tumor. The authors concluded these studies may provide a basis for combining these drugs in future clinical trials. They saw no resistance to vorinostat over 2 days; another study reported resistance to Noxa manipulation developing over 10 days, a duration they did not 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 experiment (in vitro and ex vivo). Mesothelioma cell lines were grown as flat monolayers and as 3D spheroids, and tumor fragment spheroids were grown from human mesothelioma tissue.
• Who was studied: Four human mesothelioma cell lines (M28, REN, SARC and VAMT). Also tumor fragment spheroids from six mesothelioma tumor samples, obtained by extrapleural pneumonectomy or pleurectomy at Brigham and Women's Hospital, Boston. Three tumors were tested with bortezomib and three with cisplatin plus pemetrexed.
• Exposure: Not an asbestos exposure study. Cells and tissue were treated with bortezomib (25 nM in 24-hour apoptosis assays, 100 nM in short-term experiments), vorinostat, and cisplatin plus pemetrexed, alone or in combination.
• Main results:
  • In all four cell lines grown as 3D spheroids, adding vorinostat to bortezomib abolished the resistance to bortezomib. Apoptosis returned to the same or greater level as in 2D monolayers.
  • Vorinostat given alone did not induce apoptosis in monolayers or spheroids.
  • In M28 and REN cells, Noxa message after bortezomib did not increase significantly in 3D spheroids. With vorinostat plus bortezomib, it increased to the same extent in 3D as in 2D.
  • After vorinostat plus bortezomib in spheroids, Noxa message increased 10 to 20 fold, while Bim and Bmf message increased no more than 2 to 3 fold.
  • In M28 spheroids, the combination increased the amount of Noxa bound to the protein Mcl-1 and reduced the amount of Bim bound to it.
  • When Noxa or Bim was knocked down by siRNA in REN cells, vorinostat failed to increase bortezomib-induced apoptosis in spheroids.
  • In tumor fragment spheroids, vorinostat alone, bortezomib alone, and cisplatin plus pemetrexed together (without vorinostat) each failed to significantly increase apoptosis of mesothelioma cells. Vorinostat added to bortezomib, or to cisplatin plus pemetrexed, significantly increased apoptosis in samples from each tumor.
  • In cell-line spheroids, vorinostat also amplified the apoptotic response to cisplatin plus pemetrexed.
• The authors' conclusion: The authors concluded that vorinostat undermined multicellular resistance in their 3D models by restoring Noxa. They wrote that these studies may provide a basis for using these agents in combination in future clinical trials.
• Limitations the authors state:
  • Another study reported that resistance to manipulation of Noxa can develop over 10 days. The authors did not study their spheroids for that duration, and saw no resistance to vorinostat over 2 days.
  • The authors noted that drug resistance, drug penetrance and toxicity will be important considerations for clinical use of the combination.

Topics

Medical terms in this study, explained in our Lexicon: Mesothelioma, Apoptosis, Cisplatin, Pemetrexed (Alimta).

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