Calretinin is essential for mesothelioma cell growth/survival in vitro: a potential new target for malignant mesothelioma therapy?

Our plain-English summary of Calretinin is essential for mesothelioma cell growth/survival in vitro: a potential new target for malignant mesothelioma therapy?, by W. Blum, B. Schwaller (International Journal of Cancer, 2013).

The study

• Authors: W. Blum, B. Schwaller
• Journal: International Journal of Cancer
• Published: 2013;133(9):2077-88
• Cited by: 45 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

Malignant mesothelioma (MM) are highly aggressive asbestos-related neoplasms, which show strong chemotherapy resistance, and there is no effective cure for MM so far. Calretinin (CR) is widely used as a diagnostic marker for epithelioid and mixed (biphasic) mesothelioma; however, little is known about CR's putative functions in tumorigenesis. CR protects against asbestos-induced acute cytotoxicity mediated by the AKT/PI3K pathway, and furthermore, SV40 early region genes are able to upregulate CR in mesothelial cells. However, the precise role of CR in mesothelioma is still unknown. Downregulation of CR via lentiviral-mediated short-hairpin RNA significantly decreased the viability and proliferation of mesothelioma cells in vitro. The effect was strong in epithelioid-dominated cell lines (ZL55 and MSTO-211H). A weaker and delayed effect was observed in mesothelioma cells with prevalent sarcomatoid morphology (SPC111, SPC212 and ZL34). The specificity of the effect was confirmed by stable enhanced green fluorescent protein-CR expression in mesothelioma cell lines and subsequent downregulation. Depletion of CR led these cancer cell lines to enter apoptosis within 72 hr postinfection via strong activation of the intrinsic caspase 9-dependent pathway. Downregulation of CR in immortalized mesothelial cells LP9/TERT-1 strongly blocked proliferation and caused a G1 block without decreasing viability or activating apoptosis pathways. These results demonstrate that downregulation of CR had a strong effect on the viability of MM cells and that CR is essential for cells derived from MM. The authors anticipate these findings to reveal CR as a highly interesting new putative therapeutic target for mesothelioma treatment of especially the epithelioid, as well as of the mixed and sarcomatoid type.

© 2013 UICC.

In plain English

This was a laboratory study of cells grown in culture. It did not study people or asbestos exposure. The authors used a lentivirus carrying short-hairpin RNA to reduce calretinin, a protein used as a diagnostic marker for mesothelioma, in human mesothelioma cell lines and in an immortalized mesothelial cell line, LP9/TERT-1. MSTO-211H (mixed type) and ZL55 (epithelioid) cells had high calretinin. In these cells a viability test signal fell to about 30% and about 60% of control-treated levels by 144 hours, and the cells showed signs of apoptosis (programmed cell death) through the caspase 9 pathway. Cell lines with mainly sarcomatoid features (SPC111, SPC212, ZL34) were barely affected at 120 hours and showed a decrease of about 30% at 138 hours. LP9/TERT-1 cells stopped dividing, with 90% held in the G1/G0 phase compared with 45% of control cells in G1, without signs of cell death. A mouse fibroblast line was not affected. The authors concluded that calretinin is essential for mesothelioma-derived cells in vitro and suggested it as a possible treatment target.

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 cultured cell lines, using lentivirus-delivered short-hairpin RNA to reduce calretinin
• Who was studied: Human mesothelioma cell lines (MSTO-211H, ZL5, ZL55, SPC111, SPC212, ZL34), the immortalized human mesothelial cell line LP9/TERT-1 (a model of reactive mesothelial cells), and the mouse fibroblast line L929 as a negative control; work at the University of Fribourg, Switzerland; dates of the experiments not stated
• Exposure: No asbestos exposure was studied. Cells were treated with lentivirus carrying shRNA against CALB2, the calretinin gene, compared with a control GFP shRNA, at a multiplicity of infection of 10 (5 in some assays).
• Main results:
  • At 144 hours, the MTT viability signal of calretinin-shRNA-treated cells, relative to GFP shRNA-treated cells, fell to about 30% in MSTO-211H cells and to about 60% in ZL55 cells
  • At 120 hours, the MTT signal of LP9/TERT-1 immortalized mesothelial cells decreased by 80%
  • At 120 hours, biphasic (SPC111, SPC212) and sarcomatoid (ZL34) mesothelioma cells were barely affected. At 138 hours their MTT signal decreased by about 30%.
  • L929 fibroblast cells, used as a negative control, were not affected by the calretinin shRNA
  • In MSTO-211H cells at 72 hours, the percentage of undamaged ('healthy') cells dropped from 74.4% to 48.8% after calretinin shRNA treatment
  • Caspase 9 (intrinsic apoptosis pathway) was activated in MSTO-211H and ZL55 cells. Caspase 8 activity was unaffected, which the authors took to suggest selective activation of the intrinsic pathway.
  • In LP9/TERT-1 cells, 45% of control cells were in G1. After calretinin reduction, 90% were in G1/G0, a growth arrest without loss of viability or cell death.
  • Specificity check in MSTO-211H cells expressing a green-fluorescent calretinin fusion protein: GFP shRNA reduced the fluorescence to less than 20% of control without decreasing proliferation. Calretinin shRNA reduced both fluorescence and proliferation.
• The authors' conclusion: The authors concluded that reducing calretinin strongly affected the viability of mesothelioma cells and that calretinin is essential for mesothelioma-derived cells. They suggested it as a possible new therapeutic target.
• Limitations the authors state:
  • The authors noted high variability in the percentage of apoptotic cells, which they attributed to small differences in experimental conditions around the time of peak apoptosis

Topics

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

Medical terms in this study, explained in our Lexicon: Calretinin, Mesothelioma, Malignant Mesothelioma, Neoplasm, Chemotherapy, Epithelioid, Biphasic Mesothelioma, Tumorigenesis.

Related studies

• SV40-induced expression of calretinin protects mesothelial cells from asbestos cytotoxicity and may be a key factor contributing to mesothelioma pathogenesis — T. Henzi, W.V. Blum, et al. (2009)
• Overexpression or absence of calretinin in mouse primary mesothelial cells inversely affects proliferation and cell migration — W. Blum, L. Pecze, et al. (2015)
• Expression of calretinin by breast carcinoma and the potential for misdiagnosis of mesothelioma — G. Powell, H. Roche, et al. (2011)
• Calretinin. A selective marker of normal and neoplastic mesothelial cells in serous effusions — M.C. Barberis, M. Faleri, et al. (1997)
• Comparison of antibodies to HBME-1 and calretinin for the detection of mesothelial cells in effusion cytology — P.A. Fetsch, A. Simsir, et al. (2001)
• ERK2 is essential for the growth of human epithelioid malignant mesotheliomas — A. Shukla, J.M. Hillegass, et al. (2011)

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