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Imaging the Spatial Distribution of Efavirenz in Intact HIV Tissue Reservoirs
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Reported by Jules Levin
CROI 2015 Feb 23-26, Seattle, WA
Elias P. Rosen1, Corbin G. Thompson2, Mark T. Bokhart1, Craig Sykes2, Yuri Fedoriw2, Paul Luciw3, David C. Muddiman1, and Angela DM Kashuba2
1North Carolina State University, Raleigh, NC, USA, 2University of North Carolina at Chapel Hill, Chapel Hill, NC, USA 3University of California , Davis, CA, USA
program abstract-
Background: Methods to accurately evaluate ARV biodistribution within tissues are needed to design effective HIV therapy and eradication strategies. Here, we characterize the spatial distribution of efavirenz (EFV) within suspected reservoir tissues of a primate model using a novel approach to mass spectrometry imaging (MSI).
Methods: Reservoir tissues (GALT, lymph nodes, brain, testes) were removed at necropsy from an uninfected rhesus macaque dosed orally to steady-state with EFV. 10 μm cryosections of snap frozen tissue were discretized into 10-4 mm3 voxels, resolving 100 μm features, and analyzed using an infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI) source coupled to a Thermo Q-Exactive mass spectrometer. Response was calibrated by EFV standards on blank tissue, with a limit of detection of 180 attomole/voxel (57 fg/mm3 tissue). The results were visualized using custom analysis software. Serial sections of tissue were utilized to validate MSI results by LC-MS, and stained to correlate observed EFV response with tissue morphology (H&E) and immunohistochemistry (CD3 staining).
Results: The presence of EFV was confirmed in all reservoir tissues by MSI, with varying total EFV penetration observed between tissue types. Mapping of EFV response indicated heterogeneous drug exposure. EFV concentration was substantially increased within the mucosa and lamina propria of the colorectal epithelium, specifically corresponding to high density of CD3+ T cells. No such mucosal enhancement was observed in the ileum. Lymph nodes showed focally increased signal in association with some, but not all, primary follicles. Within the brain, grey matter had enhanced EFV exposure relative to white matter. EFV concentration was lowest (167 pg/g tissue) in the basal ganglia, increasing to approximately two-fold in most other tissues (cerebrum, lymph nodes, spleen, testes, and most GALT), and highest in rectal tissue (3.6 fold).
Conclusions: This study is the first to map the biodistribution of an ARV in viral reservoir tissues. Differences in mucosal enhancement in the gut suggest potential differences in biologic transporter activity. Heterogeneous lymph node distribution may indicate insufficient exposure at important sites of viral replication. By differentiating and quantifying drug exposure between cell types within tissue, IR-MALDESI MSI offers a new capability to evaluate drug efficacy and will help inform the selection of optimal interventions to target active viral reservoirs.
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