Scientists at the Scripps Research Institute in California have discovered a new type of drug that may permanently inhibit HIV from becoming reactivated in the cells that are chronically infected with it. The new drug, didehydro-cortistatin A (dCA), appears to permanently change the cellular environment so that even after it is removed, the cells of the latently infected reservoir cells remain 100 times less likely to ‘wake up’ and start producing new viruses.
This drug fills a gap in the anti-HIV arsenal that researchers have been in search of for a long time: it is an effective, apparently non-toxic and potent tat inhibitor.
The HIV protein tat is one of the first proteins that HIV induces infected cells to make when it infects them. It does a number of different things and contributes directly to HIV's cell-killing activity and to the immune over-stimulation seen in untreated HIV infection. But its most important job is to amplify HIV replication. Without tat, HIV-infected cells only produce new viruses very sluggishly: tat tremendously accelerates the rate of DNA transcription in the cell and ensures that a productive infection is initiated and maintained.
The reappearance of HIV when antiretroviral therapy (ART) is stopped in people with a viral load which is undetectable by even the most sensitive tests is one of the frustrations of researchers working towards an HIV cure, and tat is part of what enables that reappearance. Even when the changes happen in a cell that cause it to become quiescent and enter the long-lived reservoir of permanently infected but non-productive cells, tat ensures that they will reactivate when ART is stopped. Its effects seem permanent; experiments suggest that tat has epigenetic effects, meaning that it permanently changes the structure of chromatin, the protein 'scaffolding' for DNA, in particular by removing or repositioning chromatin components that physically prevent gene transcription.
The tat protein is a unique product of HIV, has no human cellular counterpart, is a toxin in itself and appears to be an obligatory component of clinically significant HIV infection; for all these reasons a viable tat inhibitor has been a long-sought goal of HIV researchers. An Italian team is researching a therapeutic vaccine containing small doses of tat but, despite lengthy work which documents some promising immune changes in volunteers, it has not produced a vaccine with the efficacy needed to take into fully fledged clinical trials. dCA is a small molecule and could possibly be dosed orally.
In the Scripps research, some of which was presented at this year’s Conference on Retroviruses and Opportunistic Infections (CROI) in February, the scientists extracted HIV-infected CD4 reservoir cells from nine people with HIV who had been on ART for at least three years. They specifically extracted cells that produced no complete HIV viral particles spontaneously but which readily produced HIV when given immune stimulants. The cells were maintained on ART in the lab dish. Even on ART, infected reservoir cells continue to produce low levels of HIV proteins and have occasional bursts of production of complete viruses; it is these 'blips' that replenish the reservoir. However, when these cells were given dCA, it reduced the amount of p24, a viral core protein indicative of HIV replication, by a mean of 93%. Even this average figure is misleading as it includes one set of cells where p24 production was only inhibited by 55%; in five out of nine patients’ cells, p24 production was reduced by over 99%.
In a second experiment, HIV-infected CD4 reservoir cells from two patients were treated in the lab dish with either ART alone or ART plus dCA. When ART was stopped and production of new HIV particles was measured six days later, the dCA-treated cells produced only 7% as much p24 as the non-treated cells. When ART was not stopped, but instead cells were given prostratin, an activator of gene expression, in the cells that had been on ART alone, seven times as much virus was produced by the cells as the cells in which ART was stopped. However, in cells stimulated with prostratin that had been on dCA also until six days previously, there was no sign of viral production at all.
In another experiment, the researchers treated two different lab-grown cell lines with dCA. In one line, a poor virus producer, residual low-level p24 production declined to undetectability an average of 82 days after they were started on ART plus dCA; in contrast, p24 production remained unchanged, at 1000 times the undetectability level, in cells on ART alone. In a line more strongly producing HIV, near-undetectability was achieved after 225 days of treatment.
In the first cell line, when dCA was withdrawn 24 days after starting it, residual HIV p24 production started rebounding at day 66 and was back to previous levels around 100 days after first starting dCA. In contrast, when the researchers waited for 100 days before stopping dCA treatment, residual p24 replication continued to remain undetectable for the five-month duration of the experiment.
In a final experiment, the researchers tried to reverse the effect of dCA by culturing reservoir cells with an immune stimulant plus extra tat protein that had been produced elsewhere. This ‘exogenous’ tat increased residual p24 production in ART-treated reservoir cells by a factor of 800, showing the effect tat has on viral production. In cells also treated with dCA, adding in exogenous tat produced a burst of p24 where it had previously been undetectable, showing that large amounts of tat can overwhelm dCA. However, even with this extra tat, p24 production started declining ten days later and was back to undetectable again ten days after that.
These last two experiments offer the strongest evidence that dCA treatment may at least have very long-lasting effects or even permanent ones.
What are the implications? If animal and human experiments replicate the results of these lab-dish ones, then for the first time we may have a drug that is capable of steadily and progressively reducing the amount of HIV expressed by the previously untouchable long-lived reservoir cells. It may even render them permanently incapable of producing any more HIV, "preventing viral rebound and maintaining a permanent state of latency," to quote the researchers.
Another cure strategy, "kick and kill", has tried to eliminate the long-lived viral reservoir by using drugs called HDAC inhibitors to induce the reservoir cells to come out of hiding, on the assumption that the immune system will then "see" them and deal with them. However, while the stimulation has woken up reservoir cells, the immune system did not react sufficiently to the woken-up cells to reduce the size of the reservoir, possibly because HDAC inhibitors have additional, unforeseen immune-suppressant effects.
dCA would work in exactly the opposite way: it may possibly be able to maintain the reservoir cells in a lifelong sleep. It is not a drug that initially could be used to induce a state where patients could be taken off ART and maintain an undetectable viral load. However, if given in conjunction with ART or started at the point HIV becomes undetectable in the blood, it could induce a further stage of viral decay such that eventually there was no residual HIV replication left, and possibly even no cells capable of producing HIV. This could happen either because the effects of dCA really are permanent or, even if they are not, because there would be none of the low-level viral replication that is the mechanism that replenishes the reservoir, so it would dwindle slowly through natural cell death. Once there are either no latently infected cells left, or no infected cell capable of producing new viruses, then we would have an HIV cure. For the time being, these possibilities remain speculative, and much further research will be needed to understand the potential uses of dCA.
Mousseau G et al. The tat inhibitor didehydro-cortistatin A prevents HIV-1 reactivation from latency. mBio Journal 6(4):e00465-15. doi:10.1128/mBio.00465-15. July 2015.
