title: IMP-GMP specific cytosolic 5'-nucleotidase regulates nucleotide pool and prodrug metabolism.
abstract: BACKGROUND
  Type II cytosolic 5'-nucleotidase (cN-II) catalyzes the hydrolysis of purine and, to some extent, of pyrimidine monophosphates. Recently, a number of papers demonstrated the involvement of cN-II in the mechanisms of resistance to antitumor drugs such as cytarabine, gemcitabine and fludarabine. Furthermore, cN-II is involved in drug resistance in patients affected by hematological malignancies influencing the clinical outcome. Although the implication of cN-II expression and/or activity appears to be correlated with drug resistance and poor prognosis, the molecular mechanism by which cN-II mediates drug resistance is still unknown.
  
  
  METHODS
  HEK 293 cells carrying an expression vector coding for cN-II linked to green fluorescent protein (GFP) and a control vector without cN-II were utilized. A highly sensitive capillary electrophoresis method was applied for nucleotide pool determination and cytotoxicity exerted by drugs was determined with 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay.
  
  
  RESULTS
  Over-expression of cN-II causes a drop of nucleoside triphosphate concentration and a general disturbance of nucleotide pool. Over-expressing cells were resistant to fludarabine, gemcitabine and cytarabine independently of cN-II ability to hydrolyze their monophosphates.
  
  
  CONCLUSIONS
  An increase of cN-II expression is sufficient to cause both a general disturbance of nucleotide pool and an increase of half maximal inhibitory concentration (IC50) of the drugs. Since the monophosphates of cytarabine and gemcitabine are not substrates of cN-II, the protection observed cannot be directly ascribed to drug inactivation.
  
  
  CONCLUSIONS
  Our results indicate that cN-II exerts a relevant role in nucleotide and drug metabolism through not only enzyme activity but also a mechanism involving a protein-protein interaction, thus playing a general regulatory role in cell survival.
  Resistance to fludarabine, gemcitabine and cytarabine can be determined by an increase of cN-II both through dephosphorylation of active drugs and perturbation of nucleotide pool.
authors: Dipartimento di Biologia, Unità di Biochimica, Università di Pisa, Via San Zeno 51, 56127, Pisa, Italy.
authors: Federico
authors: F
authors: Cividini
authors: Dipartimento di Biologia, Unità di Biochimica, Università di Pisa, Via San Zeno 51, 56127, Pisa, Italy; Dipartimento di Chimica e Farmacia, Università di Sassari, Via Muroni 23A, 07100, Sassari, Italy.
authors: Daniela Nicole
authors: DN
authors: Filoni
authors: Dipartimento di Biologia, Unità di Biochimica, Università di Pisa, Via San Zeno 51, 56127, Pisa, Italy.
authors: Rossana
authors: R
authors: Pesi
authors: Dipartimento di Chimica e Farmacia, Università di Sassari, Via Muroni 23A, 07100, Sassari, Italy. Electronic address: enomis@uniss.it.
authors: Simone
authors: S
authors: Allegrini
authors: Dipartimento di Biologia, Unità di Biochimica, Università di Pisa, Via San Zeno 51, 56127, Pisa, Italy.
authors: Marcella
authors: M
authors: Camici
authors: Dipartimento di Biologia, Unità di Biochimica, Università di Pisa, Via San Zeno 51, 56127, Pisa, Italy.
authors: Maria Grazia
authors: MG
authors: Tozzi
mesh_terms: D015720:5'-Nucleotidase / Q000235:genetics / Q000378:metabolism*; D000970:Antineoplastic Agents / Q000378:metabolism* / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D003561:Cytarabine / Q000378:metabolism / Q000494:pharmacology; D003841:Deoxycytidine / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004351:Drug Resistance / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006157:Guanosine Monophosphate / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D015151:Immunoblotting; D007291:Inosine Monophosphate / Q000378:metabolism; D009711:Nucleotides / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011355:Prodrugs / Q000378:metabolism* / Q000494:pharmacology; D013379:Substrate Specificity; D014740:Vidarabine / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D000093542:Gemcitabine
chemical_list: D000970:Antineoplastic Agents; D009711:Nucleotides; D011355:Prodrugs; D003561:Cytarabine; D003841:Deoxycytidine; D007291:Inosine Monophosphate; D049452:Green Fluorescent Proteins; D006157:Guanosine Monophosphate; D015720:5'-Nucleotidase; D014740:Vidarabine; C024352:fludarabine; D000093542:Gemcitabine
keywords: Chemotherapy; Cytarabine; Fludarabine; Gemcitabine; Resistance; cN-II