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ABSTRACT:Many drug interactions with drugs used for the therapy of human immunodeficiency virus (HIV) occur at the level of different cytochrome P450 isozymes. Increasing evidence suggests that antiretrovirals may also modify activity and expression of active drug transport systems. Such interactions may alter drug absorption, elimination, and also drug distribution and reach clinical importance if thereby access to the target site is affected. Beyond P-glycoprotein, the family of multidrug resistance-related proteins (MRP/ABCC) substantially contributes to the elimination of numerous drugs and their metabolites. Because the interaction of MRPs with non-HIV protease inhibitor antiretrovirals has not been studied thoroughly, we investigated whether important non-nucleoside reverse transcriptase inhibitors (NNRTI) (delavirdine, efavirenz, and nevirapine), nucleoside reverse transcriptase inhibitors (NRTI) (abacavir, emtricitabine, and lamivudine), and tenofovir as a nonnucleotide reverse transcriptase inhibitor can interact with MRP1, MRP2, and MRP3 in vitro. Inhibition of these ABC transporters was quantified by confocal laser-scanning microscopy using the 5-chloromethylfluorescein diacetate assay. With the exception of abacavir, which had no effect on MRP3, all the test compounds increased intracellular 5-chloromethylfluorescein fluorescence in a concentration-dependent manner, and this effect was observed in all the overexpressing cell lines but not in the parental cell line, indicating inhibition of MRP1, MRP2, and MRP3. In conclusion, the present study provides the first evidence for a significant and concentration-dependent inhibition of MRPs by NNRTI, NRTI, and tenofovir, which was most pronounced for delavirdine, efavirenz, and emtricitabine, suggesting that this might contribute to some of the known drug interactions impairing HIV therapy and also to the superior effectiveness of combination pharmacotherapy.Infections with the human immunodeficiency virus (HIV) are typically treated with drug combinations consisting of at least three different antiretroviral drugs. Essential components of this highly active antiretroviral therapy (HAART) are the HIV protease inhibitors (HPI), the non-nucleoside and nucleoside reverse transcriptase inhibitors (NNRTI and NRTI), and the nucleotide reverse transcriptase inhibitor tenofovir. Whereas this combination therapy substantially improves the clinical prognosis for patients infected with HIV, it concurrently increases the risk for drug-drug interactions (Piscitelli and Gallicano, 2001;de Maat et al., 2003).Many drug interactions with antiretrovirals, but by far not all and particularly not those with NRTI, occur at the level of different cytochrome P450 isozymes (Dasgupta and Okhuysen, 2001;Piscitelli and Gallicano, 2001;de Maat et al., 2003). Indeed, increasing evidence suggests that antiretrovirals may also modify activity and expression of active drug transport systems. Such interactions may determine drug absorption, elimination, and also drug distribution and reac...

The drug transporter P-glycoprotein (ABCB1) plays an important role in drug distribution and elimination, and when overexpressed it may confer multidrug resistance (MDR). P-glycoprotein is localized in the plasma membrane, especially within rafts and caveolae, characterized as detergent-resistant membranes (DRMs). This study investigated the effect of cholesterol depletion and repletion as well as saturation on subcellular localization and function of P-glycoprotein to determine the effect of DRM localization on P-glycoprotein-mediated drug efflux. In L-MDR1 overexpressing human P-glycoprotein, cholesterol depletion removed P-glycoprotein from the raft membranes into non-DRM fractions, whereas repletion fully reconstituted raft localization. P-glycoprotein function was assessed by realtime monitoring with confocal laser scanning microscopy using BODIPY-verapamil as substrate. Cholesterol depletion reduced P-glycoprotein function in L-MDR1 cells resulting in intracellular substrate accumulation (159% Ϯ 43, p Ͻ 0.001; control ϭ 100%). Cholesterol repletion reduced intracellular substrate fluorescence (120% Ϯ 36, p Ͻ 0.001) and restored the transporter activity. Addition of surplus cholesterol (saturation) even enhanced drug efflux in L-MDR1 cells, leading to reduced intracellular accumulation of BODIPY-verapamil (69% Ϯ 10, p Ͻ 0.001). Transport of BODIPY-verapamil in cells not expressing human P-glycoprotein (LLC-PK1) was not susceptible to cholesterol alterations. These results demonstrate that cholesterol alterations influence P-glycoprotein localization and function, which might contribute to the large interindividual variability of P-glycoprotein activity known from in vivo studies.

Numerous studies have been published on single aspects of pulmonary adenocarcinoma (ADC). To comprehensively link clinically relevant ADC characteristics, we evaluated established morphological, diagnostic and predictive biomarkers in 425 resected ADCs.Morphology was reclassified. Cytokeratin-7, thyroid transcription factor (TTF)1, napsin A, thymidylate synthase and excision repair cross-complementing rodent repair deficiency complementation group-1 expression, anaplastic lymphoma kinase rearrangements as well as epidermal growth factor receptor (EGFR), V-Ki-ras2 Kirsten rat sarcoma viral oncogene homologue (KRAS) and v-Raf murine sarcoma viral oncogene homologue B1 (BRAF) mutations were analysed. All characteristics were correlated with clinical and survival parameters.Morphological ADC subtypes were significantly associated with smoking history and distinct patterns of diagnostic biomarkers. KRAS mutations were prevalent in male smokers, while EGFR mutations were associated with female sex, nonsmoking and lepidic as well as micropapillary growth patterns. TTF1 expression (hazard ratio (HR) for overall survival 0.61, p50.021) and BRAF mutations (HR for disease-free survival 2.0, p50.046) were found to be morphology-and stage-independent predictors of survival in multivariate analysis. Adjuvant radio-/chemotherapy, in some instances, strongly impacted on the prognostic effect of both diagnostic and predictive biomarkers.Our data draw a comprehensive picture of the prevalence and interplay of established histological and molecular ADC characteristics. These data will help to develop time-and cost-effective diagnostic and treatment algorithms for ADC. @ERSpublications Morphological, diagnostic and predictive biomarkers, and clinical characteristics of pulmonary adenocarcinomas