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Latency Reversing Agents Activate Latent Reservoirs in the Brain of SIV-Infected Macaques
Lucio Gama1, Sarah Price1, Erin Shirk1, Suzanne E. Queen1, Ming Li1, Brandon Bullock1, Stephen Wietgrefe2, Luiz Pianowski3, M. Christine Zink1, Janice Clements1
1 Molecular and Comparative Pathobiology, Johns Hopkins University School of Medicine, Baltimore, MD, United States. 2 Medicine, University of Minnesota, Minneapolis, MN, United States. 3 Bioqual, Valinhos, Sao Paulo, Brazil.
Background: Our group has been testing the PKC activator ingenol-3-hexanoate (Ing-B) as a potential candidate for a “Kick and Kill” HIV eradication strategy. Preliminary data show that Ing-B treatment caused a temporary but significant increase in plasma viral load (VL) of two virally suppressed SIVmac251-infected cART-treated rhesus macaques. After interrupting the drug regimen (cART and Ing-B), VL stayed undetectable for 45 days before rebounding to pre-cART levels. Here we report results from Ing-B treatment in our consistent and accelerated SIV macaque model for HIV/AIDS and HAND.
Methods: Three pigtailed macaques were dual inoculated with SIVΔB670 and SIV/17E-Fr, and treated at 12 days p.i. with CNS-penetrant cART (TNF, ATZ, RTN, L-870812). After 500 days of viral suppression, one animal was kept as control while two macaques received daily oral doses of Ing-B for 40 days. After a 2-week washout, the same animals received a 10-day treatment of Ing-B in combination with vorinostat (4 daily IV doses in 10 days). Animals were kept on cART until necropsy. SIV RNA was quantitated in plasma and CSF by ddPCR. SIV DNA was assessed in tissues by qPCR. In situ hybridization (ISH) for SIV RNA was performed in samples from brain and mesenteric lymph node. Resting CD4+ T cells were collected before and after latency reversing agents (LRA) treatment for quantitative viral outgrowth assay.
Results: LRA induction caused a significant increase of plasma and CSF VL in one of the LRA-treated macaques. CSF viral load was 10x higher than in plasma, and the animal had to be euthanized due to encephalitis-related symptoms. SIV RNA could be detected by ISH in occipital cortex, despite undetectable levels of SIV DNA measured by qPCR. No change was observed in the other LRA-treated macaque. However, the number of SIV-infected resting CD4+ T cells was reduced after LRA-induction in both treated animals when compared to control.
Conclusions: Treatment with LRAs led to a decrease in latent reservoirs in SIV-infected cART-treated macaques. In one animal, treatment activated viral genomes in basal ganglia, leading to CNS disease, indicating that the brain harbors latent virus and should be seriously considered when novel "Kick and Kill” strategies are designed for HIV eradication.