Depolarization of mitochondrial membrane potential is the initial event in non-nucleoside reverse transcriptase inhibitor efavirenz induced cytotoxicity

Abstract

Efavirenz is a non-nucleoside reverse transcriptase inhibitor (NNRTI) and an active constituent of the highly active antiretroviral therapy regime. It has significantly contributed in control and management of human immunodeficiency virus propagation. However, EFV administration has also led to severe adverse effects, several reports highlighted the role of EFV in mitochondrial dysfunction and toxicity but the molecular mechanism has been poorly understood. In present study, human hepatoma cells Huh 7.5 were treated with clinically relevant concentrations of EFV and parameters like cytotoxicity, mitochondrial transmembrane potential, mitochondrial morphology, cytochrome c release, mitochondria-mediated apoptosis, mtDNA and mtRNA levels and EFV distribution into mitochondrial compartment were evaluated to understand sequence of events leading to cell death in EFV-treated cells. EFV at its clinically relevant concentration was significantly toxic after 48 and 72 h of treatments. EFV-mediated toxicity is initiated with the permeabilization of mitochondrial outer membrane and change in mitochondrial membrane potential (Δψm) which triggers a series of events like cytochrome c release, alteration in mitochondrial morphology and mitochondria-mediated apoptosis. Total mitochondrial content is reduced after 48 h of EFV treatment at IC concentration which is also reflected in reduced mitochondrial DNA and RNA levels. After detecting EFV in mitochondrial compartment after 12 h of incubation with EFV, we hypothesize that EFV being a lipophilic molecule is internalized into the mitochondrial compartment causing depolarization of Δψm which subsequently leads to a cascade of events causing cell death.

Combination of tenofovir and emtricitabine with efavirenz does not moderate inhibitory effect of efavirenz on mitochondrial function and cholesterol biosynthesis in human T lymphoblastoid cell line

“…Treatment with EFV also resulted in about 50% reduction in mtDNA compared to treatment with DMSO. Although, EFV does not inhibit Pol-γ, we observed mtDNA depletion in CEM cells treated with 1xC max of EFV; this is consistent with previous report using human hepatoma cells Huh 7.5 (27). Thus, EFV has several affections on mitochondria.…”

“…In this study, we sought to investigate whether combining EFV with TDF/FTC will moderate reported EFV-associated mitochondrial dysfunction (8, 18, 25-27) and perturbation of cholesterol biosynthesis (8, 28, 29). Most of the reported EFV-induced toxicities are from in vitro studies where cells were treated with EFV alone.…”

“…Mitochondrial function depends on intact mitochondrial membrane (40). The loss of mitochondrial ΔΨ is believed to be the first and crucial step in mitochondrial dysfunction, which triggers a cascade of events leading ultimately to cell death(27, 41). It is, therefore, not surprising that the effect of EFV on mitochondrial ΔΨ (Figure 2A) paralleled the effect of EFV on apoptosis (Figure 1B).…”

“…The individual differences in EFV plasma concentration may partly explain why certain patients on EFV-based therapy do not experience effects of EFV on mitochondrial function. Ganta et al detected EFV by HPLC in purified mitochondrial lysate from Huh 7.5 and HepaRG cells after 12 h of treatment with 6.25 µM (27). The investigators explained that since EFV is hydrophilic it can pass freely through the outer mitochondrial membrane and initiate depolarization of the inner mitochondrial membrane and subsequent mitochondrial dysfunction (27).…”

“…Ganta et al detected EFV by HPLC in purified mitochondrial lysate from Huh 7.5 and HepaRG cells after 12 h of treatment with 6.25 µM (27). The investigators explained that since EFV is hydrophilic it can pass freely through the outer mitochondrial membrane and initiate depolarization of the inner mitochondrial membrane and subsequent mitochondrial dysfunction (27). This theory may be consistent with the individual differences in the mitochondrial affections of EFV.…”