Fialuridine and its metabolites inhibit DNA polymerase γ at sites of multiple adjacent analog incorporation, decrease mtDNA abundance, and cause mitochondrial structural defects in cultured hepatoblasts Journal Article
| Authors: | Lewis, W.; Levine, E. S.; Griniuviene, B.; Tankersley, K. O.; Colacino, J. M.; Sommadossi, J. P.; Watanabe, K. A.; Perrino, F. W. |
| Article Title: | Fialuridine and its metabolites inhibit DNA polymerase γ at sites of multiple adjacent analog incorporation, decrease mtDNA abundance, and cause mitochondrial structural defects in cultured hepatoblasts |
| Abstract: | The thymidine analog fialuridine [1-(2-deoxy-2-fluoro-β-D-arabinofuranosyl)-5-iodouracil (FIAU)] was toxic in trials for chronic hepatitis B infection. One mechanism postulated that defective mtDNA replication was mediated through inhibition of DNA polymerase-γ (DNA pol-γ) by FIAU triphosphate (FIAUTP) or by triphosphates of FIAU metabolites. Inhibition kinetics and primer-extension analyses determined biochemical mechanisms of FIAU, 1-(2-deoxy-2-fluoro-β-D-arabinofuranosyl)-5-methyluracil (FAU), and 1-(2-deoxy-2-fluoro-β-D-arabinofuranosyl)uracil triphosphate (TP) inhibition of DNA pol-γ. dTMP incorporation by DNA pol-γ was inhibited competitively by FIAUTP, FMAUTP, and FAUTP (Ki = 0.015, 0.03, and 1.0 μM, respectively). By using oligonucleotide template-primers, DNA pol-γ incorporated each analog into DNA opposite a single adenosine efficiently without effects on DNA chain elongation. Incorporation of multiple adjacent analogs at positions of consecutive adenosines dramatically impaired chain elongation by DNA pol-γ. Effects of FIAU, FMAU, and FAU on HepG2 cell mtDNA abundance and ultrastructure were determined. After 14 days, mtDNA decreased by 30% with 20 μM FlAU or 20 μM FMAU and decreased less than 10% with 100 μM FAU. FIAU and FMAU disrupted mitochondria and caused accumulation of intracytoplasmic lipid droplets. Biochemical and cell biological findings suggest that FIAU and its metabolites inhibit mtDNA replication, most likely at positions of adenosine tracts, leading to decreased mtDNA and mitochondrial ultrastructural defects. |
| Keywords: | controlled study; human cell; hepatitis b; dna replication; microscopy, electron; enzyme inhibition; cell line; molecular sequence data; kinetics; drug mechanism; arabinofuranosyluracil; base sequence; inhibition kinetics; binding sites; fialuridine; dna primers; drug metabolite; dna, mitochondrial; mitochondrial dna; mitochondria, liver; antiviral agents; southern blotting; transmission electron microscopy; fat droplet; dna directed dna polymerase gamma; cell ultrastructure; liver mitochondrion; nucleoside analogs; dna polymerase iii; humans; human; priority journal; article; mitochondrial myopathy |
| Journal Title: | Proceedings of the National Academy of Sciences of the United States of America |
| Volume: | 93 |
| Issue: | 8 |
| ISSN: | 0027-8424 |
| Publisher: | National Academy of Sciences |
| Date Published: | 1996-04-16 |
| Start Page: | 3592 |
| End Page: | 3597 |
| Language: | English |
| PUBMED: | 8622980 |
| PROVIDER: | scopus |
| PMCID: | PMC39655 |
| DOI: | 10.1073/pnas.93.8.3592 |
| DOI/URL: | |
| Notes: | Article -- Export Date: 22 November 2017 -- Source: Scopus |
