Kick, kill, contain – researchers examine three possible steps in a cure for HIV

The 7th International AIDS Society Conference on HIV Pathogenesis, Treatment and Prevention (IAS 2013) in Kuala Lumpur featured a special two-day symposium on progress towards a cure for HIV and also featured a number of presentations on possible approaches towards a cure in the main body of the conference.

Both the symposium and the conference heard about a number of cases of 'functional cure' or 'remission' from HIV replication in people who had started antiretroviral therapy (ART) soon after infection, including a German case that can now be added to the 'Mississippi baby' case and 14 members of the French VISCONTI cohort. These individuals manage to maintain an undetectable viral load after coming off ART and it is becoming clear that there may be other early-treated patients who might be able to come off their medication too.

The majority of people who did not start ART till later, however, are unlikely to achieve control of their HIV without more help. The problem is that unless HIV is suppressed early, it integrates into the genetic material of some of the long-lived central memory cells in the immune system. As long as people are on ART, HIV is prevented from replicating but as soon as it is stopped, these cells – only one in a million of which may contain HIV genes – start producing HIV again, it reappears in the blood, and more cells are infected.

Several different approaches are being tried to halt ongoing infection, but the one that has received the most attention has been the so-called 'kick and kill' approach.

It may be a number of years before [these experiments] are turned into an effective strategy, even if the 'kick and kill' approach turns out to be the right one.

Initially, gene-stimulating drugs are given that 'kick' the normally quiescent central memory cells into becoming activated and producing some HIV. As long as this remains suppressed at controllable levels with ART, the hope is that by becoming activated, the cells turn into 'effector' cells with short lives and the so-called reservoir of long-term infected cells is drained.

It is possible that, if the reservoir is sufficiently emptied, the body's own immune surveillance could keep HIV suppressed in the very few left. However, one study where HIV reappeared in someone with fewer than two-in-a-billion HIV infected reservoir cells casts doubt on this.

There may need to be further stages where drugs are given that actively target and kill off the activated reservoir cells, driving their number down still further, and then an immune therapy might be given that magnifies the body's natural immune response to HIV and contains the activation of the tiny number of HIV-infected cells that remain.

Kick – HDAC and beta-catenin inhibitors

The class of drugs furthest along in investigations into reservoir-cell activators is a group called HDAC inhibitors. Some are already in use as anti-cancer drugs, as they activate genes that can kill cancer cells. Professor Martin Tolstrup of Aarhus University in Denmark summarised his team's recent research into a drug called panobinostat at the cure symposium. (This research was the subject of an article in the UK's Sunday Telegraph which implied in its headline that a cure for HIV might be achieved “within months”. The team issued a correction and the article was subsequently modified.)

An HDAC inhibitor called vorinostat had already been shown to induce a 4.8-fold increase in HIV gene expression in reservoir cells after one dose, Professor Tolstrup said. However, so far no production of the viral proteins which would stimulate an immune reaction to HIV had been seen.

Panobinostat reaches levels ten times higher in the cells than vorinostat and the team gave 15 men with HIV twelve doses of it over eight weeks, three doses a week on a week-on, week-off basis. The men were all in late middle age but varied in their medical history: they had been diagnosed for between 6.5 and 28 years, and had spent between 2.5 and 16 years on ART. One man had started ART immediately on diagnosis, while another had not started until 18 years after testing HIV positive.

The researchers used a test for HIV RNA (gene products) to detect low levels of HIV or HIV proteins in the blood. This is the same technology used in viral load testing but this was sensitive to as little as one copy per millilitre of HIV RNA. They found that, after the first dose, 60% of participants expressed low but detectable levels of HIV RNA in their blood, compared with only 28% before panobinostat, and that only one of the 15 participants showed no detectable HIV RNA throughout the study period.

The team will publish data on HIV RNA and DNA detected within cells soon, and will test groups of reservoir cells to find out how many remain with hidden HIV infections and how many can produce replication-competent HIV. The hope is that, if the panobinostat can drain the reservoir sufficiently, it might be safe to take people off ART for a monitored treatment interruption.

HDAC inhibitors are not the only drugs capable of turning long-lived cells with occult HIV infection into short-lived cells visible to the immune system. Dr Mathias Lichterfeld of Massachusetts General Hospital introduced a class of drugs called beta-catenin inhibitors. Beta-catenin is a body protein that stops stem cells differentiating into memory cells, and drugs that inhibit it might be able to reach a small but important part of the HIV reservoir which appears to be the one most hidden and least susceptible to activation.

A beta-catenin inhibitor turned three-quarters of former stem cells into effector-memory cells (stimulated ones that are actively producing HIV) in test-tube trials, and use in combination with panobinostat appeared to roughly double the cell-stimulation effect of beta-cateninin. When given to cells from two people with no reaction to panobinostat alone, it multiplied the production of HIV RNA within the cell 20-fold, and doubled it in cells from a patient who had a negative response (i.e. whose cells had actually become less activated on panobinostat).

Kill – targeted cell toxins

As mentioned above, death in activated cells and immune surveillance may not be enough to deplete the HIV-infected reservoir sufficiently. We may need additional therapies that actively seek out and destroy the cells activated by HDAC or beta-catenin inhibitors.

Dr Victor Garcia of the University of North Carolina introduced such a cell-killing missile. In this molecule, a so-called broadly neutralising antibody, 3B3, that attaches itself exclusively to HIV surface proteins, is joined on to a toxin, PE38, derived from the Pseudomonas bacterium. The antibody attaches the molecule to activated cells from which HIV is budding and the toxin then enters the cells and kills them.

This molecule was injected into mice that had been genetically altered so they could be infected with human HIV. The mice were infected with HIV and then three weeks later were started on ART. Four weeks after that they were given two weekly doses of 3B3-PE38, a smaller then a larger dose.

Even though ART dramatically suppresses production of HIV within immune cells, some still remains. The ART, as expected, produced a 2.1 log (125-fold) drop in HIV RNA inside cells, but the bacterial toxin produced a further 6.5-fold drop (0.8 logs). More importantly, the absolute number of cells expressing HIV RNA decreased from between 1100 and 20,000 per gram of tissue to between 600 and 3000 per gram, an approximately sixfold drop in the presumed size of the reservoir.

Contain – a natural-killer-cell vaccine

Immune therapies and therapeutic vaccines that contain any onward infection of HIV from activated cells to other ones may also be a crucial part of the 'kick and kill' strategy, both in order to encourage the body to kill or contain the tiny fraction of HIV-infected cells left after reservoir 'draining' and to prevent onward infection of HIV into new cells during the 'kick' phase.

In this study, presented by Uriel Nieves of the Institut Pasteur in France, the immune-system cells called natural killer (NK) cells, which represent the body's first line of defence against viruses, were used. Firstly, immune cells of the type called dendritic cells were incubated in the test tube with a candidate vaccine of the MVA type, in which HIV proteins are wrapped up inside the shell of another, harmless virus called modified vaccinia Ankara.

These dendritic cells were then mixed with NK cells from the same people. The NK cells, now sensitised to HIV but the dentritic cells that "presented" HIV proteins to them, became able to recognise and kill cells expressing HIV proteins. They were then mixed with CD4 cells and other dendritic cells. HIV was then introduced. The presence of the sensitised NK cells reduced the proportion of dendritic cells infected with HIV from 45 to 25% and of CD4 cells from 35 to 20%.

This is only one of a large number of therapeutic vaccines devised, but it stimulates an immune response in a different cell type, NK cells, than has been sensitised to HIV before. The activity of HIV-sensitised NK cells, because their response to HIV is less tuned to specific variants of HIV, may be less prone to HIV-infected cells evading its immune control.

It is one of a whole number of measures in early stages of study designed to drive ongoing HIV infection down to the absolute minimum, and one where it may be, in the long run, possible to take even chronically infected people off ART for long periods without HIV reappearing.

However, as we can see, these experiments are in very early clinical or preclinical stages, and it may be a number of years before they are turned into an effective strategy, even if the 'kick and kill' approach turns out to be the right one.