title: Fludarabine resistance mediated by aminoglycoside-3'-phosphotransferase-IIa and the structurally related eukaryotic cAMP-dependent protein kinase.
abstract: While working with G418-resistant stably transfected cells, we realized the neomycin resistance (NeoR) gene, which encodes the aminoglycoside-3'-phosphotransferase-IIa [APH(3')-IIa], also confers resistance to the nucleoside analog fludarabine. Fludarabine is a cytostatic drug widely used in the treatment of hematologic and solid tumors, as well as in the conditioning of patients before transplantation of hematopoietic progenitors. We present evidence that NeoR-transfected cells do not incorporate fludarabine, thus avoiding DNA damage caused by the drug, evidenced by a lack of FANCD2 monoubiquitination and impaired apoptosis. A screening of other nucleoside analogs revealed that APH(3')-IIa only protects against ATP purine analogs. Moreover, APH(3')-IIa ATPase activity is inhibited by fludarabine monophosphate, suggesting that APH(3')-IIa blocks fludarabine incorporation into DNA by dephosphorylating its active fludarabine triphosphate form. Furthermore, overexpression of the catalytic subunit of the eukaryotic kinase PKA, which is structurally related to APHs, also provides resistance to fludarabine, anticipating its putative utility as a response marker to the drug. Our results preclude the use of Neo marker plasmids in the study of purine analogs and unveils a new resistance mechanism against these chemotherapeuticals.-Sánchez-Carrera, D., Bravo-Navas, S., Cabezón, E., Arechaga, I., Cabezas, M., Yáñez, L., Pipaón, C. Fludarabine resistance mediated by aminoglycoside-3'-phosphotransferase-IIa and the structurally related eukaryotic cAMP-dependent protein kinase.
authors.affiliation: Laboratorio de Hematología Molecular, Servicio de Hematología y Hemoterapia, Hospital Universitario Marqués de Valdecilla-Instituto de Investigación Marqués de Valdecilla (IDIVAL), Santander, Spain.
authors.forename: Dámaso
authors.initials: D
authors.lastname: Sánchez-Carrera
authors.affiliation: Laboratorio de Hematología Molecular, Servicio de Hematología y Hemoterapia, Hospital Universitario Marqués de Valdecilla-Instituto de Investigación Marqués de Valdecilla (IDIVAL), Santander, Spain.
authors.forename: Sara
authors.initials: S
authors.lastname: Bravo-Navas
authors.affiliation: Departamento de Biología Molecular and Instituto de Biomedicina y Biotecnología de Cantabria (IBBTEC), Universidad de Cantabria-Consejo Superior de Investigaciones Cientificas (CSIC), Santander, Spain.
authors.forename: Elena
authors.initials: E
authors.lastname: Cabezón
authors.affiliation: Departamento de Biología Molecular and Instituto de Biomedicina y Biotecnología de Cantabria (IBBTEC), Universidad de Cantabria-Consejo Superior de Investigaciones Cientificas (CSIC), Santander, Spain.
authors.forename: Ignacio
authors.initials: I
authors.lastname: Arechaga
authors.affiliation: Departamento de Biología Molecular and Instituto de Biomedicina y Biotecnología de Cantabria (IBBTEC), Universidad de Cantabria-Consejo Superior de Investigaciones Cientificas (CSIC), Santander, Spain.
authors.forename: Matilde
authors.initials: M
authors.lastname: Cabezas
authors.affiliation: Laboratorio de Hematología Molecular, Servicio de Hematología y Hemoterapia, Hospital Universitario Marqués de Valdecilla-Instituto de Investigación Marqués de Valdecilla (IDIVAL), Santander, Spain.
authors.forename: Lucrecia
authors.initials: L
authors.lastname: Yáñez
authors.affiliation: Laboratorio de Hematología Molecular, Servicio de Hematología y Hemoterapia, Hospital Universitario Marqués de Valdecilla-Instituto de Investigación Marqués de Valdecilla (IDIVAL), Santander, Spain; cpipaon@humv.es.
authors.forename: Carlos
authors.initials: C
authors.lastname: Pipaón
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D001665:Binding Sites; D002461:Cell Line, Transformed; D003001:Cloning, Molecular; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism*; D019008:Drug Resistance, Neoplasm / Q000502:physiology*; D005347:Fibroblasts; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D006801:Humans; D019868:Kanamycin Kinase / Q000235:genetics / Q000378:metabolism*; D015394:Molecular Structure; D013329:Structure-Activity Relationship; D014740:Vidarabine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000494:pharmacology
chemical_list: D000970:Antineoplastic Agents; D019868:Kanamycin Kinase; D017868:Cyclic AMP-Dependent Protein Kinases; D014740:Vidarabine; C024352:fludarabine
keywords: DNA damage; FANC proteins; protein kinase A; purine analogues