Treatment of chronically FIV-infected cats with suberoylanilide hydroxamic acid

Abstract

Feline immunodeficiency virus (FIV) is a naturally-occurring, large animal model of lentiviral-induced immunodeficiency syndrome, and has been used as a model of HIV pathogenesis and therapeutic interventions. HIV reservoirs in the form of latent virus remain the primary roadblock to viral eradication and cure, and FIV has been previously established an animal model of lentiviral latency. The goal of this study was to determine whether administration of the histone deacetylase inhibitor (HDACi) suberoylanilide hydroxamic acid (SAHA) to aviremic, chronically FIV-infected cats would induce latent viral reactivation in vivo. A proof-of-concept experiment in a Transwell co-culture system demonstrated the ability of SAHA to reactivate latent virus which was replication competent and able to infect naïve cells. Oral SAHA (250mg/m(2)) was administered with food to four asymptomatic, experimentally FIV-infected cats and one uninfected control cat, and a limited pharmacokinetic and pharmacodynamic analysis was performed. A statistically significant increase in cell-associated FIV RNA was detected in the cat with the greatest serum SAHA exposure, and cell-free viral RNA was detected at one time point in the three cats that achieved the highest levels of SAHA in serum. Interestingly, there was a significant decrease in viral DNA burden at 2h post drug administration in the same three cats. Though the sample size is small and the drug response was modest, this study provides evidence that in vivo treatment of FIV-infected cats with the HDACi SAHA can induce viral transcriptional reactivation, which may be dependent upon the concentration of SAHA achieved in blood. Importantly, alternative putative antilatency therapy drugs, and multimodal drug combinations, could be studied in this in vivo system. The FIV/cat model provides a unique opportunity to test novel therapeutic interventions aimed at eradicating latent virus in vivo.

Keywords: Animal model; Antilatency therapy; FIV; HDAC inhibitor; HIV; SAHA.

Figure 1

Viral DNA load from peripheral blood mononuclear cells (PBMC) in the upper culture well of the Transwell co-culture system at 7 and 14 days post ex vivo culture with PBMC from cat 165 (A) and 187 (B). Cells were cultured in media alone (NoTx), media containing 1 uM SAHA (S), SAHA and allogenic SPF PBMC (S+P), or SAHA, allogeneic PBMC, and a mitogen (5 ug/mL concanavalin A) (S+P+M). FIV gag transcripts were normalized to cellular GAPDH; the average and standard deviation of triplicate measurements are displayed (BLD – below the limit of detection). A schematic of the concept behind the Transwell co-culture system is shown in (C), where naïve SPF PBMC are co-cultured with FIV-infected PBMC separated by a 400 nm permeable membrane. Putative antilatency therapy (ALT) is used to reactivate latent virus in the lower well.

Figure 2

Pharmacokinetics of SAHA in 4 FIV+ cats (165, 184, 186, and 187) and 1 FIV- control cat (185) administered 250 mg/m² oral SAHA with food. The average (dotted black line) represents data excluding cat #165 (see text). Serum samples were analyzed by UPLC coupled to tandem mass spectrometry using deuterated SAHA as an internal standard.

Figure 3

Pharmacodynamics of SAHA in 5 cats administered 250 mg/m² oral SAHA with food. PBMC collected at 0 (pre-dose), 1, and 2 hours after SAHA administration were analyzed by Western blotting for both acetylated and total histone H3. Densitometry ratios of acetylH3/totalH3 were normalized to pre-dose controls (time 0). Values above the dashed black line (1) have a relative increase in histone H3 acetylation.

Figure 4

Cell (PBMC)-associated viral RNA at various time points before (0) and after oral SAHA administration. FIV gag transcripts were normalized to cellular GAPDH; the average and standard deviation of triplicate measurements are shown. A one-way ANOVA followed by Dunnett’s multiple comparison post-test using time 0 (pre-dose) as the control was performed to determine if there was a significant change in the level of vRNA after drug administration. *p < 0.05; **p < 0.01; ns = not significant; BLD = below the limit of detection, 5d = 5 days.

Figure 5

PBMC FIV DNA load at various time points before (0) and after oral SAHA administration to 4 FIV+ cats (165, 184, 186, and 187). Copies of FIV gag were normalized to cellular GAPDH. The average and standard deviation of triplicate measurements for individual animals are shown in (A), and the average and standard deviation of all 4 FIV+ animals are graphed in (B). A one-way ANOVA followed by Dunnett’s multiple comparison post-test using time 0 (pre-dose) as the control was performed to determine if there was a significant change in the level of viral DNA after drug administration; *p < 0.05. DNA from the FIV- control cat (185) was below the limit of detection of FIV gag at all time points (not shown).