Detection
of Drug Bioactivation in Vivo: Mechanism
of Nevirapine–Albumin Conjugate Formation in Patients
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Abstract
The
non-nucleoside reverse transcriptase inhibitor nevirapine (NVP)
is widely used for the treatment of human immunodeficiency virus type
1 (HIV-1), particularly in developing countries. Despite its therapeutic
benefits, NVP has been associated with skin and liver injury in exposed
patients. Although the mechanism of the tissue injury is not yet clear,
it has been suggested that reactive metabolites of NVP may be involved.
The detection of NVP mercapturate in the urine of patients undergoing
standard antiretroviral chemotherapy indicates that NVP undergoes
bioactivation in vivo. However, covalent binding of drug to protein
in patients remains to be determined. In this study, we investigate
the chemical basis of NVP protein adduct formation by using human
serum albumin (HSA) and glutathione <i>S</i>-transferase
pi (GSTP) as model proteins in vitro. In addition, HSA was isolated
from serum samples of HIV-1 patients undergoing NVP therapy to measure
NVP haptenation. Mass spectrometric analysis of 12-sulfoxyl-NVP-treated
HSA revealed that the drug bound selectively to histidine (His146,
His242, and His338) and a cysteine residue (Cys34). The reaction proceeds
most likely by a concerted elimination–addition mechanism.
This pathway was further confirmed by the observation of NVP-modified
Cys47 in GSTP. Importantly, the same adduct (His146) was detected
in HSA isolated from the blood of patients receiving NVP, providing
direct evidence that NVP modifies protein in vivo, via the formation
of a reactive metabolite