Novel anti-inflammatory effect of the anti-retroviral drug rilpivirine in the HepG2 human hepatocyte cell line

A. Chapman1, A. A. Shamsabadi2, V. Tilliridou1, H. Bergin2, J. G. Mabley2. 1Brighton & Sussex Medical School, Brighton, UNITED KINGDOM, 2School of Pharmacy & Biomolecular Sciences, University of Brighton, Brighton, UNITED KINGDOM.

Introduction: Rilpivirine, a second-generation non-nucleoside reverse transcriptase inhibitor, is used globally in the treatment of HIV/AIDS. Like many reverse transcriptase inhibitors, rilpivirine is known to induce clinical hepatotoxic effects, though the mechanisms causing this are unknown (1,2). The aim of this study was to investigate the direct effect of rilpivirine on hepatocytes.

Method: The human hepatocyte cell line, HepG2, was exposed to rilpivirine (3, 10 or 30 µM) either alone or in combination with palmitic acid (300 µM) for 24h. Cell viability was determined using the MTT assay and cell death by propidium iodide/Hoechst staining. Inflammation was assessed by interleukin (IL)-8 release using a specific ELISA. Data is expressed as mean±SEM, statistical analysis was carried out using two-way ANOVA with Bonferroni’s correction.

Results: Rilpivirine exposure at 3 µM had no effect on HepG2 cell viability whilst exposure to 10 or 30 µM significantly reduced cell viability from 100±0.8% to 81±1.2% and 74±1.6% respectively (p<0.05 vs. untreated cells, n=3 with 6 replicates per experiment). Rilpivirine exposure at 3 or 10 µM caused an increase in both apoptosis; from 0.9±0.2% to 3.1±0.6% and 4.5±0.5%, and necrosis; from 4.8±0.8% to 16.5±2.2% and 24±3.2% (p<0.05 vs. untreated cells, n=4 with 2 replicates per experiment) respectively. Rilpivirine exposure had a biphasic effect on basal IL-8 secretion with 3 and 10 µM decreasing IL-8 from 2.6±0.2 to 1.8±0.2 and 1.6±0.2 pg/mg protein respectively while 30 µM increased IL-8 to 3.7±0.5 pg/mg protein (p<0.05 vs. untreated cells, n=5 with 3 replicates per experiment). Rilpivirine (3 or 10 µM) had an anti-inflammatory effect against stimulated IL-8 release, palmitic acid at 300 µM increased IL-8 release from 2.4±0.2 to 14.1±1.8 pg/mg protein, reduced to 5.6±0.5 and 4.2±0.3 pg/mg protein in the presence of 3 or 10 µM rilpivirine respectively (p<0.05 vs. palmitic acid treated cells, n=5 with 3 replicates per experiment). Rilpivirine had no effect on palmitic acid-mediated decrease in cell viability or increased apoptosis/necrosis levels.

Conclusions: Rilpivirines direct hepatocyte-damaging effect at high concentrations on cell viability, death and inflammation may account for its clinically observed hepatotoxic effects. However, the novel anti-inflammatory effect observed at lower concentrations of rilpivirine reducing both basal and stimulated IL-8 release from hepatocytes reveals a possible therapeutic action that may be exploitable in HIV infected individuals.

References:

1) Nunez et al. (2001). J Acquir Immune Defic Syndr 27: 426-431

2) Nelson et al. (2012). J Antimicrob Chemother 67: 2020-2028