Cisplatin-induced expression of Gb3 enables verotoxin-1 treatment of cisplatin resistance in malignant pleural mesothelioma cells

Our plain-English summary of Cisplatin-induced expression of Gb3 enables verotoxin-1 treatment of cisplatin resistance in malignant pleural mesothelioma cells, by D. Johansson, C. Andersson, J. Moharer, et al (British Journal of Cancer, 2010).

The study

• Authors: D. Johansson, C. Andersson, J. Moharer, A. Johansson, P. Behnam-Motlagh
• Journal: British Journal of Cancer
• Published: 2010;102(2):383-91
• Cited by: 19 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Background: A major problem with cisplatin treatment is the development of acquired-drug resistance of the tumour cells. Verotoxin-1 (VT-1) exerts its cytotoxicity by targeting the membrane glycolipid globotriasosylceramide (Gb3), a molecule associated with drug resistance. Cisplatin- and VT-1-induced apoptosis involves mitogen-activated protein kinase (MAPK) activation, and deactivation of MAPKs is associated with cisplatin resistance. This study aimed to investigate whether a sub-toxic concentration of VT-1 could enhance cisplatin-induced apoptosis and overcome acquired-cisplatin resistance in cultured cancer cell lines.

Method: P31 and H1299 cells with corresponding cisplatin-resistant sub-lines (P31res/H1299res) were incubated with VT-1 and/or cisplatin followed by determination of Gb3 expression, cell viability, apoptosis, and signalling pathways.

Results: Cells from the resistant sub-lines had elevated Gb3 expression compared with the parental cell lines, and cisplatin further increased Gb3 expression, whereas VT-1 reduced the percentage of Gb3-expressing cells. Combination of cisplatin and sub-toxic concentrations of VT-1 led to a super-additive increase of cytotoxicity and TUNEL staining, especially in the cisplatin-resistant sub-lines. Blockade of Gb3 synthesis by a Gb3 synthesis inhibitor not only led to eradicated TUNEL staining of P31 cells, but also sensitised P31res cells to the induction of apoptosis by cisplatin alone. Cisplatin- and VT-1-induced apoptosis involved the MAPK pathways with increased C-Jun N-terminal kinase and MAPK kinase-3 and -6 phosphorylation.

Conclusions: We show the presence of Gb3 in acquired-cisplatin resistance in P31res and H1299res cells. Cisplatin up-regulated Gb3 expression in all cells and thus sensitised the cells to VT-1-induced cytotoxicity. A strong super-additive effect of combined cisplatin and a sub-toxic concentration of VT-1 in cisplatin-resistant malignant pleural mesothelioma cells were observed, indicating a new potential clinical-treatment approach.

In plain English

This laboratory study used cultured human cancer cell lines: P31 (malignant pleural mesothelioma, MPM) and H1299 (non-small cell lung cancer, NSCLC), each with a cisplatin-resistant sub-line (P31res, H1299res). The authors measured Gb3, the membrane receptor for verotoxin-1 (VT-1), plus cell viability, apoptosis (TUNEL), caspase activity and signalling proteins. Gb3-expressing cells were few in P31 (1%) and H1299 (12%) but higher in P31res (27%) and H1299res (29%); cisplatin 5 mg/l for 72 hours raised these to 31% in P31, 54% in P31res, 15% in H1299 and 40% in H1299res, while VT-1 reduced them. Combining cisplatin with a sub-toxic VT-1 concentration significantly increased cytotoxicity, and the authors described a strong super-additive effect in cisplatin-resistant MPM cells; TUNEL staining in P31res rose from 19% with cisplatin alone to 78% with the combination. PPMP, a Gb3 synthesis inhibitor, eradicated DNA fragmentation induced by VT-1 plus cisplatin (the text says in P31 cells, though it also reports P31 TUNEL staining unaffected by PPMP), and seemed to sensitise P31res cells to cisplatin alone. The authors concluded Gb3-targeted therapy is a possible approach warranting further 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: In vitro laboratory experiment using human cancer cell lines and their cisplatin-resistant sub-lines (no patients, no animals)
• Who was studied: Cultured human cells: the malignant pleural mesothelioma line P31 and the non-small cell lung cancer line H1299, each with a cisplatin-resistant sub-line (P31res, H1299res); experiments generally in triplicate (n = 3), carried out at Umea University, Sweden (published 2010)
• Exposure: Not an asbestos exposure study: cells were exposed to cisplatin (0.1-10 mg/l, or 5 mg/l in most assays) and verotoxin-1 (0.1-5.0 mg/l for P31, 0.001-1.0 mg/l for H1299; sub-toxic doses 0.1 mg/l for P31 and 0.001 mg/l for H1299), alone or combined, plus the Gb3 synthesis inhibitor PPMP (2 mmol/l) and the MDR1/PgP inhibitors verapamil and cyclosporin A (10 mmol/l)
• Main results:
  • Baseline Gb3-expressing cells: 1% of P31 and 12% of H1299 cells, rising to 27% in P31res and 29% in H1299res
  • Cisplatin 5 mg/l for 72 h increased the proportion of Gb3-expressing cells to 31% (P31), 54% (P31res), 15% (H1299) and 40% (H1299res)
  • VT-1 lowered the percentage of Gb3-expressing cells: to 1% (P31) and 4% (P31res) with 0.1 mg/l, and to 3% (H1299) and 20% (H1299res) with 0.001 mg/l
  • VT-1 alone (0.1-5 mg/l, 72 h) showed no cytotoxicity to P31 and only modest cytotoxicity to P31res, while both NSCLC sub-types were sensitive at much lower concentrations
  • Adding VT-1 to cisplatin significantly increased cytotoxicity (p less than 0.05), especially in the cisplatin-resistant sub-lines
  • The authors characterised the combined treatment in cisplatin-resistant MPM cells as strongly super-additive
  • TUNEL staining in mesothelioma cells: VT-1 alone 17% in P31res and no increase in P31; cisplatin 5 mg/l gave 78% in P31 but 19% in P31res; the combination gave 78% in P31res, with no further effect beyond cisplatin alone in P31
  • PPMP results as the article reports them, which are internally inconsistent: the results text says PPMP eradicated the DNA fragmentation induced by VT-1 with cisplatin in P31 cells, the same section says P31 Gb3 expression and TUNEL staining were unaffected by PPMP, and the section heading attributes the eradicated VT-1 super-additive effect to cisplatin-resistant MPM cells; PPMP seemed to sensitise P31res cells to cisplatin alone
  • The same subsection states that PPMP left P31 cells unchanged
  • Caspase assays: cisplatin activated caspase-3 and -9 in P31 but not P31res; VT-1 activated caspase-3 in P31res but not P31
  • Western blotting showed JNK and MKK3/6 phosphorylation after cisplatin in P31 but not P31res, while VT-1 caused phosphorylation in P31res but not P31; MKK3/6 phosphorylation increased further with the combination
  • MDR1/PgP inhibitors verapamil and cyclosporin A did not noticeably reduce MDR1/PgP expression in any cell type, whereas PPMP reduced both Gb3 and MDR1/PgP expression in the resistant sub-lines
• The authors' conclusion: The authors concluded that Gb3 is present in acquired cisplatin resistance in these cells, that cisplatin up-regulates Gb3 and thereby sensitises cells to VT-1, and that Gb3-targeted therapy with VT-1 or its sub-units could be a possible approach
• Limitations the authors state:
  • The authors noted that Gb3 expression alone does not explain VT-1 cytotoxicity, since NSCLC cells were highly sensitive despite modest Gb3 expression, and that the relationship needs further investigation
  • The authors stated that the relationship between drug efflux pump expression and Gb3 in these resistant cells requires further study, which was ongoing
  • Several apoptosis-related proteins tested gave no conclusive changes

Topics

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

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

Related studies

• Resistance to caspase-8 and -9 fragments in a malignant pleural mesothelioma cell line with acquired cisplatin-resistance — V. Janson, A. Johansson, et al. (2010)
• Interaction and cross-resistance of cisplatin and pemetrexed in malignant pleural mesothelioma cell lines — M. Kitazono-Saitoh, Y. Takiguchi, et al. (2012)
• Impact of metallothionein-knockdown on cisplatin resistance in malignant pleural mesothelioma — S. Borchert, P.M. Suckrau, et al. (2020)
• Modulation of Calretinin Expression in Human Mesothelioma Cells Reveals the Implication of the FAK and Wnt Signaling Pathways in Conferring Chemoresistance towards Cisplatin — J. Wörthmüller, V. Salicio, et al. (2019)
• Loss of C/EBP-β LIP drives cisplatin resistance in malignant pleural mesothelioma — J. Kopecka, I.C. Salaroglio, et al. (2018)
• Quercetin and Cisplatin combined treatment altered cell cycle and mitogen activated protein kinase expressions in malignant mesotelioma cells — A. Demiroglu-Zergeroglu, E. Ergene, et al. (2016)

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