Here we investigated the early effect
Here we investigated the early effect of LEDGINs and evaluated their effect on HIV Rose Bengal site distribution. In addition, we monitored the effect of LEDGINs on the establishment of the latent reservoir and investigated whether retargeting of integration could lead to a silent HIV reservoir resistant to reactivation. In a stepwise approach we first demonstrate that LEDGF/p75 depletion results in reduced integration and a quiescent state of residual integrants. Next, we demonstrate that upon treatment with LEDGINs, blocking the LEDGF/p75–IN interaction, the residual proviral integration shifts away from transcription units. LEDGIN treatment also shifts the 3D localization of the integrated provirus towards the inner nucleus. LEDGIN-induced retargeting results in a silent HIV reservoir in cell lines and primary CD4+ cells. This silent reservoir is refractory to reactivation by latency reversing agents (LRAs). Pushing sufficient proviruses into latency is theoretically predicted to drive the basic reproduction number of HIV below 1, resulting in unsustainable infection (Rouzine et al., 2015). Hence, addition of LEDGINs to cART regimens during acute HIV infection may represent a new strategy to achieve a remission of HIV infection in patients.
Materials and Methods
Discussion The moral duty to respond to the call for an HIV cure calls for exploration of experimental and innovative HIV cure strategies. As a complement to current purge-and-kill approaches aimed at forcing HIV out of its hiding places to obtain a sterilizing cure, we here provide experimental evidence for a strategy to push the virus towards transcriptional quiescence by interference with LEDGF/p75, the main determinant of integration site selection. General belief states that latency is an accident rather than a default pathway of an actively replicating cytopathic virus. The fact that LEDGF/p75, the tethering determinant of HIV integration, directs integration preferentially towards actively transcribed regions, ensuring productive infection, is consistent with this notion. Recently, it was proposed that HIV latency is a hardwired, evolutionarily conserved switch increasing the likelihood of successful mucosal transmission during primary infection (Razooky et al., 2015; Rouzine et al., 2015). Whereas HIV Tat is known to be the main viral determinant in controlling the HIV transcriptional state, the contribution of other viral and host determinants to the transcriptional state of the HIV provirus awaits further clarification. Here we investigated the role of LEDGF/p75 in the establishment of HIV latency. We demonstrate that LEDGF/p75 depletion, known to result in retargeting of integration away from the body of actively transcribed genes (Fadel et al., 2014; Schrijvers et al., 2012a; Shun et al., 2007), increases the fraction of quiescently infected cells and simultaneously decreases the reactivation potential of the proviruses. In light of the recently proposed role of LEDGF/p75 (and the Iws1/Spt6 complex) in post-integration HIV transcriptional repression (Gérard et al., 2015), this observed decrease may even be an underestimation of the contribution of the integration environment to the latent phenotype. Taken together, these observations suggest that disruption of the LEDGF/p75–IN interaction not only inhibits integration but could as well affect the establishment of the latent pool and the reactivation from latency.
Acknowledgements We are grateful to J. De Rijck for critical reading and thank P. Van de Velde, B. Vanremoortel and NJ. Van der Veeken for their technical assistance. Viral vector production was performed at the Leuven Viral Vector Core. LEDGINs were synthesized by Cistim/CD3 (courtesy of Dr. A. Marchand). L. Vranckx is a doctoral fellow supported by the Flemish Fund for Scientific Research (FWO; Fonds voor Wetenschappelijk Onderzoek). Research at KU Leuven received financial support from the FWO, the KU Leuven Research Council (OT; OT/13/098), HIV-ERA EURECA (IWT-SBO-EURECA, ZL345530), the KU Leuven IDO program (IDO/12/008) the Belgian IAP Belvir (ZKC4893 - P7/45-P) and the Creative and Novel Ideas in HIV Research Program (CNIHR, P30-AI027763) through a supplement to the University of California at San Francisco (UCSF) Center For AIDS Research funding (P30 AI027763). This latter funding was made possible by collaborative efforts of the Office of AIDS Research, the National Institute of Allergy and Infectious Diseases, and the International AIDS Society.