Rilpivirine-associated aggregation-induced emission enables cell-based nanoparticle tracking

Abstract

Antiretroviral therapy (ART) has improved the quality and duration of life for people living with human immunodeficiency virus (HIV) infection. However, limitations in drug efficacy, emergence of viral mutations and the paucity of cell-tissue targeting remain. We posit that to maximize ART potency and therapeutic outcomes newer drug formulations that reach HIV cellular reservoirs need be created. In a step towards achieving this goal we harnessed the aggregation-induced emission (AIE) property of the non-nucleoside reverse transcriptase inhibitor rilpivirine (RPV) and used it as a platform for drug cell and subcellular tracking. RPV nanocrystals were created with endogenous AIE properties enabling the visualization of intracellular particles in cell and tissuebased assays. The intact drug crystals were easily detected in CD4 + T cells and macrophages, the natural viral target cells, by flow cytometry and ultraperformance liquid chromatography tandem mass spectrometry. We conclude that AIE can be harnessed to monitor cell biodistribution of selective antiretroviral drug nanocrystals.

Europium sulfide nanoprobes predict antiretroviral drug delivery into HIV-1 cell and tissue reservoirs

“…MDM displayed maximal uptake between 2-4 hours post treatment, with 28% of the added EuS particles taken up by the cells by 4 hours ( Figure 4 B ). These levels approximate the 10% NRPV uptake reported in MDM previously 24 . Co-treatment with Dynasore significantly reduced EuS uptake in MDM by 40% ( Figure 4 C ).…”

“…Intact NRPV likewise emits fluorescence, albeit at a shorter wavelength (385 nm), resulting from aggregation induced emission 24 . The MRI compatibility of EuS was also determined as a means of quantifying particle accumulation at different tissue depths.…”

Defining the Innate Immune Responses for SARS-CoV-2-Human Macrophage Interactions

“…For ultrastructural analysis, mock and SARS-CoV-2-challenged MDMs sampled at 1, 3, 5 and 14 days post-viral inoculation were washed 2 times with PBS and fixed in a solution of 2% glutaraldehyde and 2% PFA in 0.1 M Sorenson’s phosphate buffer for 24 hours at 4°C which have been washed 3 times with PBS to clear excess fixative solution. TEM analysis was performed as previously described ( 23 ) in samples post-fixed in a 1% aqueous solution of osmium tetroxide for 30 minutes that were dehydrated in 50, 70, 90, 95, and 100% graded ethanol. Spurr’s resin was used as embedding medium after solvent transition with ethanol and Spurr’s resin (50:50 ethanol:resin, followed by twice immersion in 100% Spurr’s resin for 2-3 hours for each solution), and embedded samples were cured at 60 - 65°C for 24 hours.…”

Defining the Immune Responses for SARS-CoV-2-Human Macrophage Interactions

“…For ultrastructural analysis, mock and SARS-CoV-2challenged MDMs sampled at 1, 3, 5 and 14 days post-viral inoculation were washed 2 times with PBS and fixed in a solution of 2% glutaraldehyde and 2% PFA in 0.1 M Sorenson's phosphate buffer for 24 hours at 4°C which have been washed 3 times with PBS to clear excess fixative solution. TEM analysis was performed as previously described (Mukadam et al, 2020) in samples post-fixed in a 1% aqueous solution of osmium tetroxide for 30 minutes that were dehydrated in 50, 70, 90, 95, and 100% graded ethanol. Spurr's resin was used as embedding medium after solvent transition with ethanol and Spurr's resin (50:50 ethanol:resin, followed by twice immersion in 100% Spurr's resin for 2-3 hours for each solution), and embedded samples were cured at 60 -65°C for 24 hours.…”