title: Altered pH gradient at the plasma membrane of osteosarcoma cells is a key mechanism of drug resistance.
abstract: Current therapy of osteosarcoma (OS), the most common primary bone malignancy, is based on a combination of surgery and chemotherapy. Multidrug resistance mediated by P-glycoprotein (P-gp) overexpression has been previously associated with treatment failure and progression of OS, although other mechanisms may also play a role. We considered the typical acidic extracellular pH (pHe) of sarcomas, and found that doxorubicin (DXR) cytotoxicity is reduced in P-gp negative OS cells cultured at pHe 6.5 compared to standard 7.4. Short-time (24-48 hours) exposure to low pHe significantly increased the number and acidity of lysosomes, and the combination of DXR with omeprazole, a proton pump inhibitor targeting lysosomal acidity, significantly enhanced DXR cytotoxicity. In OS xenografts, the combination treatment of DXR and omeprazole significantly reduced tumor volume and body weight loss. The impaired toxicity of DXR at low pHe was not associated with increased autophagy or lysosomal acidification, but rather, as shown by SNARF staining, with a reversal of the pH gradient at the plasma membrane (ΔpHcm), eventually leading to a reduced DXR intracellular accumulation. Finally, the reversal of ΔpHcm in OS cells promoted resistance not only to DXR, but also to cisplatin and methotrexate, and, to a lesser extent, to vincristine. Altogether, our findings show that, in OS cells, short-term acidosis induces resistance to different chemotherapeutic drugs by a reversal of ΔpHcm, suggesting that buffer therapies or regimens including proton pump inhibitors in combination to low concentrations of conventional anticancer agents may offer novel solutions to overcome drug resistance.
authors: Orthopaedic Pathophysiology and Regenerative Medicine Unit, Istituto Ortopedico Rizzoli, Bologna, Italy.
authors: Sofia
authors: S
authors: Avnet
authors: Orthopaedic Pathophysiology and Regenerative Medicine Unit, Istituto Ortopedico Rizzoli, Bologna, Italy.
authors: Silvia
authors: S
authors: Lemma
authors: Orthopaedic Pathophysiology and Regenerative Medicine Unit, Istituto Ortopedico Rizzoli, Bologna, Italy.
authors: Margherita
authors: M
authors: Cortini
authors: Department of Oncology-Pathology, Cancer Center Karolinska, Karolinska Institute, Stockholm, Sweden.
authors: Paola
authors: P
authors: Pellegrini
authors: Orthopaedic Pathophysiology and Regenerative Medicine Unit, Istituto Ortopedico Rizzoli, Bologna, Italy.
authors: Francesca
authors: F
authors: Perut
authors: CNR - National Research Council of Italy, Institute of Molecular Genetics, Bologna, Italy.|Laboratory of Musculoskeletal Cell Biology, Istituto Ortopedico Rizzoli, Bologna, Italy.
authors: Nicoletta
authors: N
authors: Zini
authors: Musculoskeletal Oncology Unit, Takai Hospital, Nara, Japan.
authors: Katsuyuki
authors: K
authors: Kusuzaki
authors: Department of Clinical Laboratory Medicine, Shiga University of Medical Science, Otsu, Shiga, Japan.
authors: Tokuhiro
authors: T
authors: Chano
authors: Department of Medical and Surgical Sciences for Children and Adults, University-hospital of Modena e Reggio Emilia, Modena, Italy.
authors: Giulia
authors: G
authors: Grisendi
authors: Department of Medical and Surgical Sciences for Children and Adults, University-hospital of Modena e Reggio Emilia, Modena, Italy.
authors: Massimo
authors: M
authors: Dominici
authors: Department of Oncology-Pathology, Cancer Center Karolinska, Karolinska Institute, Stockholm, Sweden.
authors: Angelo
authors: A
authors: De Milito
authors: Orthopaedic Pathophysiology and Regenerative Medicine Unit, Istituto Ortopedico Rizzoli, Bologna, Italy.|Department of Biomedical and Neuromotor Sciences, University of Bologna, Bologna, Italy.
authors: Nicola
authors: N
authors: Baldini
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D001859:Bone Neoplasms / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D049109:Cell Proliferation / Q000187:drug effects; D004317:Doxorubicin / Q000494:pharmacology; D018432:Drug Resistance, Multiple; D019008:Drug Resistance, Neoplasm; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008297:Male; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D009113:Muramidase / Q000378:metabolism; D012516:Osteosarcoma / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology*; D014407:Tumor Cells, Cultured; D059016:Tumor Microenvironment / Q000187:drug effects*; D023041:Xenograft Model Antitumor Assays
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000970:Antineoplastic Agents; D004317:Doxorubicin; D009113:Muramidase
keywords: doxorubicin; drug resistance; osteosarcoma; plasma membrane pH gradient; tumor microenvironment