Novel Aptamers for the Reactivation of Latent HIV.

Abstract

A "Shock and Kill" strategy has been proposed to eradicate the HIV latent viral reservoir. Effective Latency Reversal Agents (LRA) are a key requirement for this strategy. The search for LRAs with a novel mechanism of action is ongoing. This is the first study to propose aptamers for the reactivation of HIV. The purpose of this study was to identify an aptamer that potentially reactivates HIV via the NF-κβ pathway, specifically by binding to IkB and releasing NF-κβ. Aptamer selection was performed at Aptus Biotech (www.aptusbiotech.es), using ikB human recombinant protein with His tag bound to Ni-NTA agarose resin using the SELEX procedure. Activation of NF-κβ was measured by SEAP Assay. HIV reactivation was measured in JLat cells using a BD FACS-Canto™ II flow cytometer. All flow cytometry data were analyzed using Kaluza analyzing software. Clones that had equivalent or greater activation than the positive control in the SEAP assay were regarded as potential reactivators of the NF-κβ pathway and were sequenced. The three ikb clones namely R6-1F, R6-2F, and R6-3F were found to potentially activate the NF-κβ pathway. Toxicity was determined by exposing lymphocytes to serial dilutions of the aptamers; the highest concentration of the aptamers that did not decrease viability by > 20% was used for the reactivation experiments. The three novel aptamers R6-1F, R6-2F, and R6-3F resulted in 4,07%, 6,72% and 3,42% HIV reactivation, respectively, while the untreated control showed minimal (<0.18%) fluorescence detection. This study demonstrated the reactivation of latent HIV by aptamers that act via the NF-κβ pathway. Although the effect was modest and unlikely to be of clinical benefit, future studies are warranted to explore ways of enhancing reactivation.