Comparative bioactivation of the novel anti-tuberculosis agent PA-824 in Mycobacteria and a subcellular fraction of human liver.
OBJECTIVE
PA-824 is a 2-nitroimidazooxazine prodrug currently in Phase II clinical trial for tuberculosis therapy. It is bioactivated by a deazaflavin (F(420) )-dependent nitroreductase (Ddn) isolated from Mycobacterium tuberculosis to form a des-nitro metabolite. This releases toxic reactive nitrogen species which may be responsible for its anti-mycobacterial activity. There are no published reports of mammalian enzymes bioactivating this prodrug. We have investigated the metabolism of PA-824 following incubation with a subcellular fraction of human liver, in comparison with purified Ddn, M. tuberculosis and Mycobacterium smegmatis.


METHODS
PA-824 (250 µM) was incubated with the 9000 × g supernatant (S9) of human liver homogenates, purified Ddn, M. tuberculosis and M. smegmatis for metabolite identification by liquid chromatography mass spectrometry analysis.


RESULTS
PA-824 was metabolized to seven products by Ddn and M. tuberculosis, with the major metabolite being the des-nitro product. Six of these products, but not the des-nitro metabolite, were also detected in M. smegmatis. In contrast, only four of these metabolites were observed in human liver S9; M3, a reduction product previously proposed as an intermediate in the Ddn-catalyzed des-nitrification and radiolytic reduction of PA-824; two unidentified metabolites, M1 and M4, which were products of M3; and a haem-catalyzed product of imidazole ring hydration (M2).


CONCLUSIONS
PA-824 was metabolized by des-nitrification in Ddn and M. tuberculosis, but this does not occur in human liver S9 and M. smegmatis. Thus, PA-824 was selectively bioactivated in M. tuberculosis and there was no evidence for 'cross-activation' by human enzymes.
Department of Molecular Medicine and Pathology, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand.
M
M
Dogra
B D
BD
Palmer
G
G
Bashiri
M D
MD
Tingle
S S
SS
Shinde
R F
RF
Anderson
R
R
O'Toole
E N
EN
Baker
W A
WA
Denny
N A
NA
Helsby
D000995:Antitubercular Agents / Q000493:pharmacokinetics* / Q000494:pharmacology; D001483:Base Sequence; D001711:Biotransformation; D017931:DNA Primers; D006801:Humans; D008099:Liver / Q000378:metabolism*; D013058:Mass Spectrometry; D020102:Mycobacterium smegmatis / Q000187:drug effects* / Q000378:metabolism; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000378:metabolism; D009593:Nitroimidazoles / Q000493:pharmacokinetics* / Q000494:pharmacology; D013347:Subcellular Fractions / Q000378:metabolism*
D000995:Antitubercular Agents; D017931:DNA Primers; D009593:Nitroimidazoles; C410767:pretomanid