Modulation of Calretinin Expression in Human Mesothelioma Cells Reveals the Implication of the FAK and Wnt Signaling Pathways in Conferring Chemoresistance towards Cisplatin

Our plain-English summary of Modulation of Calretinin Expression in Human Mesothelioma Cells Reveals the Implication of the FAK and Wnt Signaling Pathways in Conferring Chemoresistance towards Cisplatin, by J. Wörthmüller, V. Salicio, A. Oberson, et al (International Journal of Molecular Sciences, 2019).

The study

• Authors: J. Wörthmüller, V. Salicio, A. Oberson, W. Blum, B. Schwaller
• Journal: International Journal of Molecular Sciences
• Published: 2019;20(21)
• Cited by: 11 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Malignant mesothelioma (MM) is an aggressive asbestos-linked neoplasm, characterized by dysregulation of signaling pathways. Due to intrinsic or acquired chemoresistance, MM treatment options remain limited. Calretinin is a Ca2+-binding protein expressed during MM tumorigenesis that activates the FAK signaling pathway, promoting invasion and epithelial-to-mesenchymal transition. Constitutive calretinin downregulation decreases MM cells' growth and survival, and impairs tumor formation in vivo. In order to evaluate early molecular events occurring during calretinin downregulation, we generated a tightly controlled IPTG-inducible expression system to modulate calretinin levels in vitro. Calretinin downregulation significantly reduced viability and proliferation of MM cells, attenuated FAK signaling and reduced the invasive phenotype of surviving cells. Importantly, surviving cells showed a higher resistance to cisplatin due to increased Wnt signaling. This resistance was abrogated by the Wnt signaling pathway inhibitor 3289-8625. In various MM cell lines and regardless of calretinin expression levels, blocking of FAK signaling activated the Wnt signaling pathway and vice versa. Thus, blocking both pathways had the strongest impact on MM cell proliferation and survival. Chemoresistance mechanisms in MM cells have resulted in a failure of single-agent therapies. Targeting of multiple components of key signaling pathways, including Wnt signaling, might be the future method-of-choice to treat MM.

In plain English

This laboratory study used human malignant mesothelioma cell lines. It did not study patients or asbestos exposure. The authors built a system that lowers calretinin, a protein made by many mesothelioma cells, when the chemical IPTG is added. In MSTO-211H cells, calretinin fell by around 90 to 95% from day 5. After nine days, an MTT assay of cell growth and viability showed a decrease of approximately 60% in MSTO-211H cells; in ZL55 cells the MTT signal fell by 30%. Surviving MSTO-211H cells migrated more slowly, but they were more resistant to the chemotherapy drug cisplatin at all concentrations tested. Compared with control cells, these cells had higher mRNA levels of Wnt pathway genes, including WNT5A and WNT5B (up three- and six-fold). With 2.5 micromolar cisplatin, the Wnt inhibitor 3289-8625 completely removed their extra resistance; this did not happen in ZL55 cells. In five mesothelioma cell lines, blocking both the FAK and Wnt pathways lowered MTT signals most. The authors concluded that targeting calretinin together with pathways such as Wnt may be considered as a potential new treatment strategy.

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: in vitro laboratory study on human mesothelioma cell lines, using an IPTG-switchable (inducible) shRNA system to lower calretinin levels
• Who was studied: Human malignant mesothelioma cell lines: mainly MSTO-211H, also ZL55, H226, SPC212 and ZL34. The comparison cell lines were Met-5a (SV40-immortalized mesothelial cells) and LP9/TERT-1 (a model for reactive mesothelial cells). No patients were studied.
• Exposure: No asbestos exposure was studied. The cells were exposed to: calretinin downregulation with IPTG (1 mM); calretinin overexpression; cisplatin at 0.625 to 5 micromolar; the FAK inhibitor VS-6063 at 2.5 to 10 micromolar; and the Wnt inhibitor 3289-8625 at 100 micromolar.
• Main results:
  • In MSTO-211H cells, calretinin levels were decreased by around 90 to 95% from day 5 of IPTG treatment.
  • At day 9 of IPTG treatment, MTT assays in MSTO-211H cells with lowered calretinin showed a decrease of approximately 60% in proliferation rate and number of viable cells compared with untreated cells (p less than 0.0001).
  • In ZL55 cells with lowered calretinin, the effect was smaller: a 30% drop in the MTT signal.
  • MSTO-211H cells with lowered calretinin closed a scratch wound after 12-13 h, compared with an average of 6 h for control cells.
  • MSTO-211H cells overexpressing calretinin were more viable than control cells only at the two lower cisplatin concentrations (0.625 and 1.25 micromolar).
  • Surviving MSTO-211H cells with lowered calretinin were more resistant to cisplatin than control cells at all concentrations tested.
  • In MSTO-211H cells with lowered calretinin, WNT5A and WNT5B transcript (mRNA) levels were upregulated three- and six-fold, respectively, compared with control cells.
  • The Wnt inhibitor 3289-8625 was tested by giving it for at least 24 h before exposing the cells to cisplatin at 2.5 micromolar.
  • Under these conditions, in MSTO-211H cells with lowered calretinin, the inhibitor 3289-8625 (which blocks the protein DVL2) completely removed the extra cisplatin resistance.
  • In ZL55 cells, the 3289-8625 inhibitor did not affect the added cisplatin resistance.
  • In all five mesothelioma cell lines, blocking both the FAK and Wnt pathways gave the largest decrease in MTT signals (a combined measure of cell proliferation and viability).
  • In the non-transformed mesothelial lines (Met-5a and LP9/TERT-1), all treatments lowered the MTT signal by about 20%, and the cisplatin plus 3289-8625 combination lowered it by 40%.
• The authors' conclusion: The authors concluded that co-targeting calretinin together with other key signaling pathways such as Wnt may be considered as a potential new strategy to improve mesothelioma treatment.

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: Calretinin, Mesothelioma, Cisplatin, Malignant Mesothelioma, Neoplasm, Tumorigenesis.

Related studies

• Calretinin Functions in Malignant Mesothelioma Cells Cannot Be Replaced by the Closely Related Ca2+-Binding Proteins Calbindin-D28k and Parvalbumin — J. Wörthmüller, A. Oberson, et al. (2018)
• Interaction and cross-resistance of cisplatin and pemetrexed in malignant pleural mesothelioma cell lines — M. Kitazono-Saitoh, Y. Takiguchi, et al. (2012)
• Expression profile and function of Wnt signaling mechanisms in malignant mesothelioma cells — S.A. Fox, A.K. Richards, et al. (2013)
• Putative cancer stem cells in malignant pleural mesothelioma show resistance to cisplatin and pemetrexed — L. Cortes-Dericks, G.L. Carboni, et al. (2010)
• Overexpression or absence of calretinin in mouse primary mesothelial cells inversely affects proliferation and cell migration — W. Blum, L. Pecze, et al. (2015)
• Cell-growth and migration inhibition of human mesothelioma cells induced by 3-O-methylfunicone from Penicillium pinophilum and cisplatin — E. Buommino, A. De Filippis, et al. (2012)

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