Intracellular metabolism of favipiravir (T-705) in uninfected and influenza A (H5N1) virus-infected cells.
OBJECTIVE
To determine the metabolism of favipiravir (T-705, 6-fluoro-3-hydroxy-2-pyrazinecarboxamide) to its ribosylated, triphosphorylated form (T-705 RTP) in uninfected and influenza A/Duck/MN/1525/81 (H5N1) virus-infected cells. Effects of treatment on intracellular guanosine triphosphate (GTP) pools and influenza virus-inhibitory activity were also assessed.


METHODS
A strong anion exchange HPLC separation method with UV detection was used to quantify T-705 RTP and GTP levels in Madin-Darby canine kidney cells. Antiviral activity was determined by virus yield reduction assay.


RESULTS
Accumulation of T-705 RTP in uninfected cells increased linearly from 3 to 320 pmol/10(6) cells in cells exposed to 1-1000 microM extracellular T-705 for 24 h, approaching maximum levels by 9 h. Virus infection did not result in greater T-705 RTP accumulation compared with uninfected cells. Catabolism of T-705 RTP occurred after removal of T-705 from the extracellular medium, with a half-life of decay of 5.6 +/- 0.6 h. Based upon these results, short-term incubation of T-705 with H5N1 virus-infected cells was predicted to provide an antiviral benefit. Indeed, 4-8 h 10-100 microM T-705 treatment of cells resulted in virus yield reductions, but less than continuous exposure. A 100-fold higher extracellular concentration of T-705 was required to inhibit intracellular GTP levels compared with ribavirin, which helps explain ribavirin's greater toxicity.


CONCLUSIONS
The favourable intracellular metabolic properties of T-705 combined with its reduced cell-inhibitory properties make this compound an attractive candidate for treating human influenza virus infections.
Smee
Donald F
DF
Department of Animal, Dairy and Veterinary Sciences, Institute for Antiviral Research, Utah State University, Logan, UT, USA. don.smee@usu.edu
Hurst
Brett L
BL
Egawa
Hiroyuki
H
Takahashi
Kazumi
K
Kadota
Takumi
T
Furuta
Yousuke
Y
D000577:Amides / Q000378:metabolism* / Q000494:pharmacology*; D000818:Animals; D000998:Antiviral Agents / Q000378:metabolism* / Q000494:pharmacology*; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D003600:Cytosol / Q000737:chemistry; D004285:Dogs; D006160:Guanosine Triphosphate / Q000032:analysis; D053124:Influenza A Virus, H5N1 Subtype / Q000187:drug effects* / Q000254:growth & development*; D011719:Pyrazines / Q000378:metabolism* / Q000494:pharmacology*; D013056:Spectrophotometry, Ultraviolet / Q000379:methods
D000577:Amides; D000998:Antiviral Agents; D011719:Pyrazines; D006160:Guanosine Triphosphate; C462182:favipiravir