9th International Workshop on Adverse Drug Reactions and Lipodystrophy in HIV


19-21 July 2007, Sydney, Australia

ACUTE INHIBITION OF MITOCHONDRIAL RESPIRATION BY EFAVIRENZ IN HEPATIC CELLS: A NEW MECHANISM OF DAMAGE FOLLOWING BIOENERGETIC STRESS

Antiviral Therapy 2007; 12(Suppl. 2):L25 (abstract no. P-05)

A Blas-García, M Rocha, F Baixauli, A Alvarez, N Martínez-Martín, VM Víctor and JV Esplugues
Department of Pharmacology, Faculty of Medicine, University of Valencia, Valencia, Spain

BACKGROUND: Liver toxicity is the most relevant adverse effect of non-nucleoside reverse transcriptase inhibitors (NNRTIs) although there is no clear explanation of the mechanisms responsible. The present study evaluates in vitro the acute effects of efavirenz on mitochondrial function and its implication in the energetic metabolism of the hepatic cell.

METHODS: Non-HIV-infected Hep3B cells, cultured in minimum essential medium, were placed in gas-tight chambers and O2 consumption was measured with a Clark-type O2 electrode. Following incubation with efavirenz, for 1 and 4 h respectively, intracellular ATP was measured by fluorescence (ATP Bioluminiscence Assay Kit HSII, Roche), whereas mitochondrial cytochrome c release was measured in the cytosolic fraction and analysed by Western blotting. Ritonavir or lamivudine were used as comparison.

RESULTS: The presence of efavirenz induces significant and dose-dependent (one-way ANOVA followed by Student’s t-test) inhibition of mitochondrial O2 consumption, which was noticeable immediately after the addition of the drug. This inhibition of mitochondrial function resulted in a similar dose-dependent and significant reduction of intracellular ATP following 1 h incubation with efavirenz. Both these effects were also present with lamivudine but not with ritonavir. Incubation with efavirenz (4 h) resulted in a dose-dependent increase in cytosolic cytochrome c.




Table 1. (Abstract P-05)
Treatment, µM % of reduction in O2 consumption versus control (n=3) ATP, nmol/mg protein (n=6)
Control – 16.97 ±2.38
Efavirenz
  10 26.57 ±5.73‡ 13.68 ±2.89
  25 50.60 ±8.09‡ 9.42 ±2.04*
  50 54.67 ±1.35‡ 4.82 ±0.96†
Lamivudine
  25 44.03 ±11.18† 6.77 ±2.13†
Ritonavir
  25 9.40 ±3.27 –
*P<0.05, †P<0.01, ‡P<0.001

DISCUSSION: These preliminary results suggest that clinically used concentrations of efavirenz acutely reduce mitochondrial function in hepatic cells. This is followed by a significant diminution in intracellular ATP and probably leads to metabolic stress, which is accompanied by damage in the mitochondrial membranes as suggested by the release of mitochondrial cytochrome c. These mechanisms could be involved in the toxic effects of efavirenz in the liver.

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2007-07-24
P-05

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