The Bcl-2 repertoire of mesothelioma spheroids underlies acquired apoptotic multicellular resistance

Our plain-English summary of The Bcl-2 repertoire of mesothelioma spheroids underlies acquired apoptotic multicellular resistance, by D. Barbone, J.A. Ryan, N. Kolhatkar, et al (Cell Death & Disease, 2011).

The study

• Authors: D. Barbone, J.A. Ryan, N. Kolhatkar, A.D. Chacko, D.M. Jablons, D.J. Sugarbaker, R. Bueno, A.G. Letai, L.M. Coussens, D.A. Fennell, V.C. Broaddus
• Journal: Cell Death & Disease
• Published: 2011;2(6):e174
• Cited by: 44 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Three-dimensional (3D) cultures are a valuable platform to study acquired multicellular apoptotic resistance of cancer. We used spheroids of cell lines and actual tumor to study resistance to the proteasome inhibitor bortezomib in mesothelioma, a highly chemoresistant tumor. Spheroids from mesothelioma cell lines acquired resistance to bortezomib by failing to upregulate Noxa, a pro-apoptotic sensitizer BH3-only protein that acts by displacing Bim, a pro-apoptotic Bax/Bak-activator protein. Surprisingly, despite their resistance, spheroids also upregulated Bim and thereby acquired sensitivity to ABT-737, an inhibitor of anti-apoptotic Bcl-2 molecules. Analysis using BH3 profiling confirmed that spheroids acquired a dependence on anti-apoptotic Bcl-2 proteins and were 'primed for death'. We then studied spheroids grown from actual mesothelioma. ABT-737 was active in spheroids grown from those tumors (5/7, ∼70%) with elevated levels of Bim. Using immunocytochemistry of tissue microarrays of 48 mesotheliomas, we found that most (33, 69%) expressed elevated Bim. In conclusion, mesothelioma cells in 3D alter the expression of Bcl-2 molecules, thereby acquiring both apoptotic resistance and sensitivity to Bcl-2 blockade. Mesothelioma tumors ex vivo also show sensitivity to Bcl-2 blockade that may depend on Bim, which is frequently elevated in mesothelioma. Therefore, mesothelioma, a highly resistant tumor, may have an intrinsic sensitivity to Bcl-2 blockade that can be exploited therapeutically.

In plain English

This laboratory study examined why mesothelioma cells grown as three-dimensional clusters (spheroids) resist the drug bortezomib. It did not study asbestos exposure. The authors used four mesothelioma cell lines, spheroids grown from seven surgically removed tumors, and stained tissue from 48 tumors. In flat layers of cell lines M28 and REN, bortezomib's 48-hour IC50, a potency measure based here on dying-cell counts, was about 18 nM and 9 nM. Spheroids of both lines had an IC50 near 100 nM. In spheroids, bortezomib failed to raise a protein called Noxa, but spheroids held more of the cell-death protein Bim. ABT-737, a drug that blocks Bcl-2 survival proteins, reversed this resistance. Given alone, it triggered cell death in spheroids but not in flat layers. Normal mesothelial cells, tested only as flat layers, did not undergo cell death after bortezomib, ABT-737 or both. ABT-737 increased bortezomib-induced cell death in spheroids from five of the seven tumors, which had high Bim. Of the 48 tumors, 33 (69%) had Bim above normal tissue. The authors called combining such blockers with chemotherapy a promising strategy and said further studies are needed.

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 and ex vivo) study using cell-line spheroids, spheroids grown from fragments of human tumors, and immunohistochemistry of tissue microarrays
• Who was studied: Four human mesothelioma cell lines: M28 and REN (epithelioid) and SARC and VAMT (sarcomatous). Also tumor fragment spheroids from seven mesothelioma tumors removed by extrapleural pneumonectomy or pleurectomy at Brigham and Women's Hospital, Boston. A tissue microarray of 48 mesothelioma tumors and 5 normal pleural samples from Brigham and Women's Hospital and UCSF Medical Center. Normal mesothelial cells were taken from benign human ascites.
• Exposure: Laboratory treatment with the proteasome inhibitor bortezomib, with or without ABT-737 (an inhibitor of Bcl-2/XL/w) and a Noxa peptide (R8-NoxaBH3). The study did not examine asbestos or any fiber exposure.
• Main results:
  • In cell lines grown as flat layers (monolayers), the IC50 at 48 hours was approximately 18 nM for M28 and 9 nM for REN. Spheroids of both lines showed resistance, with an approximate IC50 of 100 nM. Apoptosis was measured by counting Hoechst-stained cells.
  • In spheroids, bortezomib did not increase Noxa, whereas it did in monolayers. Spheroids expressed more Bim than monolayers.
  • In M28 and REN cells, ABT-737 given alone induced apoptosis in spheroids but not in monolayers, and it eliminated the spheroids' acquired resistance to bortezomib.
  • Normal mesothelial cells from benign ascites, studied only as monolayers, did not undergo apoptosis after bortezomib, ABT-737 alone, or the combination.
  • Spheroids of all four cell lines had more Bim than monolayers: a 1.9-fold increase in M28, and 4.5-, 3.2- and 4.0-fold increases in REN, SARC and VAMT.
  • In spheroids grown from five of the seven human tumors, ABT-737 increased bortezomib-induced apoptosis. These responsive tumors had high Bim. The two unresponsive tumors had Bim at low or undetectable levels.
  • Of 48 mesothelioma tumors, 33 (69%) expressed Bim above normal tissue: 20 of 22 epithelioid, 12 of 21 mixed and 1 of 5 sarcomatous.
• The authors' conclusion: The authors concluded that resistance to bortezomib in 3D models is mediated by changes in Bcl-2 family proteins. They described blocking these anti-apoptotic proteins alongside chemotherapy as a promising strategy for mesothelioma, and suggested Bim as a possible marker of response to ABT-737.
• Limitations the authors state:
  • The authors stated that further studies are needed to confirm their predictions about ABT-737 as a therapy and Bim as a biomarker.
  • The authors stated that the reason Bim increases in 3D spheroids is not yet known.
  • The authors noted that the protein Mcl-1 is a potential barrier to using ABT-737 clinically.

Topics

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

Medical terms in this study, explained in our Lexicon: Mesothelioma.

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