The development of
HIV protease inhibitors (PIs) has been one of the most significant advances of the past decade in controlling
HIV infection. Unfortunately, the benefits of HIV PIs are compromised by serious side effects. One of the most frequent and deleterious side effects of HIV PIs is severe gastrointestinal (GI) disorders including mucosal erosions, epithelial barrier dysfunction, and leak-flux
diarrhea, which occurs in 16-62% of patients on HIV PIs. Although the underlying mechanisms behind HIV PI-associated serious adverse side effects remain to be identified, our recent studies have shown that activation of endoplasmic reticulum (ER) stress response plays a critical role in HIV PI-induced GI complications. The objective of this study was to develop a novel self-microemulsifying drug delivery system (SMEDDS) using various
antioxidants as
surfactants and cosurfactants to reduce the GI side effects of the most commonly used HIV PI,
ritonavir. The
biological activities of this SMSDDS of
ritonavir were compared with that of
Norvir, which is currently used in the clinic. Rat normal intestinal epithelial cells (IEC-6) and mouse Raw 264.7 macrophages were used to examine the effect of new SMEDDS of
ritonavir on activation of ER stress and oxidative stress. Sprague-Dawley rats and C57/BL6 mice were used for pharmacokinetic studies and in vivo studies. The intracellular and plasma
drug concentrations were determined by HPLC analysis. Activation of ER stress was detected by Western blot analysis and secreted
alkaline phosphatase (SEAP) reporter assay.
Reactive oxygen species (ROS) was measured using dichlorodihydrofluorescein diacetate as a probe. Cell viability was determined by Roche's cell proliferation
reagent WST-1.
Protein levels of inflammatory
cytokines (TNF-alpha and IL-6) were determined by
enzyme-linked
immunosorbent assays (ELISA). The intestinal permeability was assessed by
luminal enteral administration of
fluorescein isothiocyanate conjugated
dextran (
FITC-dextran, 4 kDa). The pathologic changes in intestine were determined by histological examination. The results indicated that incorporation of
antioxidants in this new SMEDDS not only significantly reduced
ritonavir-induced ER stress activation, ROS production and apoptosis in intestinal epithelial cells and macrophages, but also improved the solubility, stability and bioavailability of
ritonavir, and significantly reduced
ritonavir-induced disruption of intestinal barrier function in vivo. In conclusion, this new SMEDDS of
ritonavir has less GI side effects compared to
Norvir. This new SMEDDS can be used for other HIV PIs and any insoluble
antiviral drug with serious GI side effects.