Inhibition of bromodomain and extra-terminal (BET) proteins with JQ1 exacerbates acetaminophen-induced hepatotoxicity by altering detoxification pathways and oxidative stress responses

Abstract

Acetaminophen (APAP) is a widely used over-the-counter drug for the treatment of fever and pain. At therapeutic doses, APAP has a relatively safe profile. However, at supratherapeutic doses it can produce liver injury and even fatal hepatotoxicity. Antioxidant genes regulated by the Kelch-like ECH-associated protein 1 (KEAP1)-nuclear factor, erythroid 2-like 2 (NRF2) pathway play a crucial role in hepatoprotection against APAP-induced hepatotoxicity and oxidative stress. Recent studies suggest that bromodomain and extra-terminal motif (BET) proteins, epigenetic readers, act as putative regulators of the KEAP1-NRF2 pathway, but their role in acute drug-induced liver injury (DILI) remains unclear. In this study, we employed complementary in vitro and in vivo approaches utilizing pharmacological inhibition and gene knockdown techniques to examine the role of BET proteins in APAP-induced toxicity. Our findings indicate that APAP treatment significantly alters the gene and protein expression of BET proteins in both mouse liver and the HC-04 cell line. Cytotoxicity analysis using lactate dehydrogenase (LDH) leakage assay revealed that treatment with the small molecule BET inhibitor JQ1 did not alter APAP-induced cytotoxicity. However, siRNA-mediated knockdown of the BET genes Brd3 and Brd4, but not Brd2, reduced APAP-induced cytotoxicity in HC-04 cells. In hepatic gene expression analysis experiments, JQ1 pretreatment in mice activated the Nrf2 pathway and altered antioxidant genes such as Gclc, Gclm, Ho-1, and Txnrd1, suggesting an enhancement of cellular defenses against APAP-induced oxidative stress at 12 h timepoint. However, by 24 h, histopathological findings revealed significant liver necrosis and inflammation in the JQ1-APAP treated group, indicating that while BET inhibition may confer early protection, it may not fully prevent long-term liver injury.