title: Targeting glucosylceramide synthase induction of cell surface globotriaosylceramide (Gb3) in acquired cisplatin-resistance of lung cancer and malignant pleural mesothelioma cells.
abstract: BACKGROUND
  Acquired resistance to cisplatin treatment is a caveat when treating patients with non-small cell lung cancer (NSCLC) and malignant pleural mesothelioma (MPM). Ceramide increases in response to chemotherapy, leading to proliferation arrest and apoptosis. However, a tumour stress activation of glucosylceramide synthase (GCS) follows to eliminate ceramide by formation of glycosphingolipids (GSLs) such as globotriaosylceramide (Gb3), the functional receptor of verotoxin-1. Ceramide elimination enhances cell proliferation and apoptosis blockade, thus stimulating tumor progression. GSLs transactivate multidrug resistance 1/P-glycoprotein (MDR1) and multidrug resistance-associated protein 1 (MRP1) expression which further prevents ceramide accumulation and stimulates drug efflux. We investigated the expression of Gb3, MDR1 and MRP1 in NSCLC and MPM cells with acquired cisplatin resistance, and if GCS activity or MDR1 pump inhibitors would reduce their expression and reverse cisplatin-resistance.
  
  
  METHODS
  Cell surface expression of Gb3, MDR1 and MRP1 and intracellular expression of MDR1 and MRP1 was analyzed by flow cytometry and confocal microscopy on P31 MPM and H1299 NSCLC cells and subline cells with acquired cisplatin resistance. The effect of GCS inhibitor PPMP and MDR1 pump inhibitor cyclosporin A for 72h on expression and cisplatin cytotoxicity was tested.
  
  
  RESULTS
  The cisplatin-resistant cells expressed increased cell surface Gb3. Cell surface Gb3 expression of resistant cells was annihilated by PPMP whereas cyclosporin A decreased Gb3 and MDR1 expression in H1299 cells. No decrease of MDR1 by PPMP was noted in using flow cytometry, whereas a decrease of MDR1 in H1299 and H1299res was indicated with confocal microscopy. No certain co-localization of Gb3 and MDR1 was noted. PPMP, but not cyclosporin A, potentiated cisplatin cytotoxicity in all cells.
  
  
  CONCLUSIONS
  Cell surface Gb3 expression is a likely tumour biomarker for acquired cisplatin resistance of NSCLC and MPM cells. Tumour cell resistance to MDR1 inhibitors of cell surface MDR1 and Gb3 could explain the aggressiveness of NSCLC and MPM. Therapy with GCS activity inhibitors or toxin targeting of the Gb3 receptor may substantially reduce acquired cisplatin drug resistance of NSCLC and MPM cells.
authors.affiliation: Department of Medical Biosciences, Umeå University, S-901 85 Umea, Sweden. Electronic address: andreas.tyler@medbio.umu.se.
authors.forename: Andreas
authors.initials: A
authors.lastname: Tyler
authors.affiliation: Department of Odontology, Umeå University, S-901 85 Umea, Sweden.
authors.forename: Anders
authors.initials: A
authors.lastname: Johansson
authors.affiliation: Department of Radiation Sciences, Oncology, S-901 85 Umea, Sweden.
authors.forename: Terese
authors.initials: T
authors.lastname: Karlsson
authors.affiliation: Department of Medical Biosciences, Umeå University, S-901 85 Umea, Sweden.
authors.forename: Shyam Kumar
authors.initials: SK
authors.lastname: Gudey
authors.affiliation: Department of Medical Biosciences, Umeå University, S-901 85 Umea, Sweden.
authors.forename: Thomas
authors.initials: T
authors.lastname: Brännström
authors.affiliation: Department of Medical Biosciences, Umeå University, S-901 85 Umea, Sweden.
authors.forename: Kjell
authors.initials: K
authors.lastname: Grankvist
authors.affiliation: Department of Medical Biosciences, Umeå University, S-901 85 Umea, Sweden.
authors.forename: Parviz
authors.initials: P
authors.lastname: Behnam-Motlagh
mesh_terms: D018435:ATP Binding Cassette Transporter, Subfamily B / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002518:Ceramides / Q000378:metabolism; D016572:Cyclosporine / Q000494:pharmacology*; D018432:Drug Resistance, Multiple / Q000187:drug effects; D019008:Drug Resistance, Neoplasm / Q000187:drug effects*; D005434:Flow Cytometry; D005964:Glucosyltransferases / Q000037:antagonists & inhibitors*; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology; D008175:Lung Neoplasms / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology; D008614:Meperidine / Q000031:analogs & derivatives / Q000494:pharmacology; D008654:Mesothelioma / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology; D000086002:Mesothelioma, Malignant; D018613:Microscopy, Confocal; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000378:metabolism; D014281:Trihexosylceramides / Q000378:metabolism*; D014407:Tumor Cells, Cultured
chemical_list: D018435:ATP Binding Cassette Transporter, Subfamily B; D002518:Ceramides; D016572:Cyclosporine; D005964:Glucosyltransferases; D007166:Immunosuppressive Agents; D008614:Meperidine; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D014281:Trihexosylceramides; C513055:ABCB1 protein, human; C026733:4-propionyloxy-4-phenyl-N-methylpiperidine; C018549:globotriaosylceramide; C023246:ceramide glucosyltransferase; C421109:multidrug resistance-associated protein 1
keywords: Acquired drug resistance; Cisplatin; Cyclosporin A; DL-threo-1-phenyl-2-palmitoylamino-3-morpholino-1-propanol (PPMP); Globotriaosylceramide (Gb3); Glucosylceramide synthase (GCS); Lung cancer; Malignant pleural mesothelioma (MPM); Multidrug resistance 1/P-glycoprotein (MDR1); Multidrug resistance-associated protein 1 (MRP1)