Efavirenz and Efavirenz-like Compounds Activate Human, Murine, and Macaque Hepatic IRE1α-XBP1

Abstract

Efavirenz (EFV), a widely used antiretroviral drug, is associated with idiosyncratic hepatotoxicity and dyslipidemia. Here we demonstrate that EFV stimulates the activation in primary hepatocytes of key cell stress regulators: inositol-requiring 1a (IRE1a) and X-box binding protein 1 (XBP1). Following EFV exposure, XBP1 splicing (indicating activation) was increased 35.7-fold in primary human hepatocytes. In parallel, XBP1 splicing and IRE1a phosphorylation (p-IRE1a, active IRE1a) were elevated 36.4-fold and 4.9-fold, respectively, in primary mouse hepatocytes. Of note, with EFV treatment, 47.2% of mouse hepatocytes were apoptotic; which was decreased to 23.9% in the presence of STF 083010, an inhibitor of XBP1 splicing. Experiments performed using pregnane X receptor (PXR)-null mouse hepatocytes revealed that EFV-mediated XBP1 splicing and hepatocyte death were not dependent on PXR, which is a nuclear receptor transcription factor that plays a crucial role in the cellular response to xenobiotics. Interestingly, incubation with the primary metabolite of EFV, 8-hydroxyefavirenz (8-OHEFV), only resulted in 10.3-and 2.9-fold increased XBP1 splicing in human and mouse hepatocytes and no change in levels of p-IRE1a in mouse hepatocytes. To further probe the structure-activity relationship of IRE1a-XBP1 activation by EFV, 16 EFV analogs were employed. Of these, an analog in which the EFV alkyne is replaced with an alkene and an analog in which the oxazinone oxygen is replaced by a carbon stimulated XBP1 splicing in human, mouse, and macaque hepatocytes. These data demonstrate that EFV and compounds sharing the EFV scaffold can activate IRE1a-XBP1 across human, mouse, and macaque species.

Twelfth-Position Deuteration of Nevirapine Reduces 12-Hydroxy-Nevirapine Formation and Nevirapine-Induced Hepatocyte Death

“…Primary hepatocytes were stained and imaged, and those images were processed as previously described. 56 …”

NNRTI and Liver Damage: Evidence of Their Association and the Mechanisms Involved

“…JNKs are a family of serine/threonine kinases that are important regulators of cellular responses to stress, including cell death. There is also evidence that points to the capacity of EFV to induce ER stress and unfolded protein response (UPR) in human hepatic cells [ 98 , 99 ]. It activates key ER stress/UPR controllers, inositol-requiring 1α (IRE1α) and X-box binding protein 1 (XBP1) through increased formation of ROS, and this effect is thought to contribute to EFV-mediated hepatocyte death [ 99 ].…”

“…There is also evidence that points to the capacity of EFV to induce ER stress and unfolded protein response (UPR) in human hepatic cells [ 98 , 99 ]. It activates key ER stress/UPR controllers, inositol-requiring 1α (IRE1α) and X-box binding protein 1 (XBP1) through increased formation of ROS, and this effect is thought to contribute to EFV-mediated hepatocyte death [ 99 ]. Our research shows that, concomitantly with mitochondrial dysfunction and ER stress, EFV-exposed cells display altered autophagy [ 100 ].…”

Apoptosis of Hepatocytes: Relevance for HIV-Infected Patients under Treatment

“…JNKs are a family of serine/threonine kinases that are important regulators of cellular responses to stress, including modulation of cell death. A recent study by the same group demonstrated that EFV and several EFV-like compounds activate key cell stress controllers such as inositol-requiring 1α (IRE1α) and X-box binding protein 1 (XBP1), a regulator of the mammalian UPR [ 89 ]. The molecular mechanism by which EFV activated IRE1α and, subsequently, IRE1α-catalyzed splicing of XBP1 mRNA involved increased formation of ROS, an event that can lead to activation of the ER stress response, including the IRE1α-XBP1 signaling axis, whose activation was demonstrated in humans, mice, and macaques.…”

“…The molecular mechanism by which EFV activated IRE1α and, subsequently, IRE1α-catalyzed splicing of XBP1 mRNA involved increased formation of ROS, an event that can lead to activation of the ER stress response, including the IRE1α-XBP1 signaling axis, whose activation was demonstrated in humans, mice, and macaques. Thus, activation of the IRE1α-XBP1 axis is thought to contribute to EFV-mediated hepatocyte death [ 89 ] ( Figure 2 ). In another investigation, this NNRTI was used as a model of drug-induced inhibition of complex I in primary mouse hepatocytes, and the authors concluded that this effect was the cause of cytotoxicity, as inhibition of complex I resulted in increased oxidative stress via the formation of peroxynitrite, which can trigger mitochondria-mediated cell death [ 90 ].…”