title: Hypoxia increases the heterogeneity of melanoma cell populations and affects the response to vemurafenib.
abstract: A hypoxic microenvironment is one of the predominant reasons for incomplete response to melanoma treatment. Vemurafenib, which targets the mutated BRAF-V600 kinase, improves melanoma patient survival, however, resistance invariably develops. The present study evaluated the effect of hypoxia on three BRAF-V600E mutant melanoma cell lines, M14, A375 and 518A2, treated with vemurafenib. Compared with the other two cell lines, hypoxic vemurafenib-treated A375 cells exhibited an enhanced cell proliferation rate and migratory capacity compared with normoxic vemurafenib-treated A375 cells. Immunoblotting analyses revealed that the expression levels of hypoxia inducible factor (HIF)1α and carbonic anhydrase IX were reduced in vemurafenib‑treated M14 and 518A2 cells, however, not in A375 cells. The expression levels of the mitogen‑activated protein kinase, Janus kinase-signal transducer and activator of transcription, and phosphatidylinositol-4,5-bisphosphate 3‑kinase signaling pathway proteins revealed a cell‑type specific response to vemurafenib and hypoxia. Knockdown experiments of HIF1α performed in hypoxic A375 cells decreased the expression of phosphorylated (p‑)protein kinase B, which was restored following vemurafenib treatment, and increased the expression of p‑extracellular‑signal‑regulated kinases. Therefore, three melanoma cell lines responded to vemurafenib under hypoxia in a cell type‑specific manner, suggesting that a subset of cells provides a treatment-resistant pool, from which disease relapse may originate. These data confirmed that vemurafenib may be successful in treating the proliferating cells, whereas the non‑proliferating subpopulation must be addressed by a combination of vemurafenib with other treatment strategies.
authors.affiliation: Department of Pathophysiology and Allergy Research, Center for Pathophysiology, Infectiology and Immunology, Medical University of Vienna, Vienna A‑1090, Austria.
authors.forename: Daniela
authors.initials: D
authors.lastname: Pucciarelli
authors.affiliation: Department of Pathophysiology and Allergy Research, Center for Pathophysiology, Infectiology and Immunology, Medical University of Vienna, Vienna A‑1090, Austria.
authors.forename: Nina
authors.initials: N
authors.lastname: Lengger
authors.affiliation: Institute of Virology, Department of Molecular Medicine, Slovak Academy of Sciences, Bratislava 81101, Slovakia.
authors.forename: Martina
authors.initials: M
authors.lastname: Takáčová
authors.affiliation: Institute of Virology, Department of Molecular Medicine, Slovak Academy of Sciences, Bratislava 81101, Slovakia.
authors.forename: Lucia
authors.initials: L
authors.lastname: Csaderova
authors.affiliation: Institute of Virology, Department of Molecular Medicine, Slovak Academy of Sciences, Bratislava 81101, Slovakia.
authors.forename: Maria
authors.initials: M
authors.lastname: Bartosova
authors.affiliation: Department of Pathophysiology and Allergy Research, Center for Pathophysiology, Infectiology and Immunology, Medical University of Vienna, Vienna A‑1090, Austria.
authors.forename: Heimo
authors.initials: H
authors.lastname: Breiteneder
authors.affiliation: Institute of Virology, Department of Molecular Medicine, Slovak Academy of Sciences, Bratislava 81101, Slovakia.
authors.forename: Silvia
authors.initials: S
authors.lastname: Pastorekova
authors.affiliation: Department of Pathophysiology and Allergy Research, Center for Pathophysiology, Infectiology and Immunology, Medical University of Vienna, Vienna A‑1090, Austria.
authors.forename: Christine
authors.initials: C
authors.lastname: Hafner
mesh_terms: D000951:Antigens, Neoplasm / Q000235:genetics / Q000378:metabolism; D000970:Antineoplastic Agents / Q000494:pharmacology*; D000071231:Carbonic Anhydrase IX / Q000235:genetics / Q000378:metabolism; D015687:Cell Hypoxia; D045744:Cell Line, Tumor; D002465:Cell Movement; D049109:Cell Proliferation; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D007211:Indoles / Q000494:pharmacology*; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D008545:Melanoma / Q000378:metabolism / Q000473:pathology; D010101:Oxygen Consumption; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D013449:Sulfonamides / Q000494:pharmacology*; D000077484:Vemurafenib
chemical_list: D000951:Antigens, Neoplasm; D000970:Antineoplastic Agents; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007211:Indoles; D034741:RNA, Small Interfering; D013449:Sulfonamides; D000077484:Vemurafenib; D019869:Phosphatidylinositol 3-Kinases; D048493:Proto-Oncogene Proteins B-raf; D051057:Proto-Oncogene Proteins c-akt; D048049:Extracellular Signal-Regulated MAP Kinases; D048031:JNK Mitogen-Activated Protein Kinases; C089226:CA9 protein, human; D000071231:Carbonic Anhydrase IX