Much Accomplished, Much More to Achieve

Despite tremendous gains in the realms of HIV treatment and prevention, the focus at AIDS 2014 was on what still needs to be done.

By Neil McKellar-Stewart

The 20th International AIDS Conference (AIDS 2014), held July 20-25 in Melbourne, Australia, commenced on an even sadder and more reflective tone than this biannual meeting typically conjures, as the nearly 14,000 delegates from over 200 countries commemorated the tragic death of their colleagues who died on their way tothe conference as passengers aboard the Malaysian airline flight brought down in eastern Ukraine (see Joep Lange's Long Reach and In Brief).

Although this terrible loss cast a shadow over the entirety of the conference, delegates turned their focus to recent progress in the realms of HIV treatment and prevention research, and the implementation of recently proven effective HIV prevention methods, including voluntary medical male circumcision (VMMC) and pre-exposure prophylaxis (PrEP; the use of antiretrovirals to prevent HIV infection). Salim Abdool Karim, director of the Centre for the AIDS Programme of Research in South Africa (CAPRISA), spoke convincingly of the possibility of controlling the HIV pandemic, even in the absence of an effective vaccine or cure, by implementing existing treatment and prevention approaches. He referenced modeling studies that suggest VMMC, earlier initiation of antiretroviral therapy (ART), and PrEP, if implemented in combination and at ambitious coverage levels, could produce a six-fold decline in global HIV incidence by 2025. However, Karim acknowledged that this would not stop HIV transmission completely.

Anthony Fauci, director of the US National Institute of Allergy and Infectious Diseases (NIAID), concluded that a world without AIDS would require both a vaccine and a cure. Although there was little new data on vaccine research to speak of in Melbourne, cure research once again received top billing as the conference was preceded by the Towards an HIV Cure symposium, the fourth such pre-conference meeting on the topic.

One step forward, two steps back?

At the main conference, Jintanat Ananworanich, associate director for therapeutics research at the US Military HIV Research Program, highlighted a multitude of recent cure-related studies (see box, below), including the recent cases in which transient but encouraging remission from HIV infection was achieved: the two Boston patients who received allogeneic stem cell transplantation, and the Mississippi infant who remained HIV free for several years following early initiation of ART (see A Toddler Stole the ShowIAVI Report, Spring 2013). In early July, the National Institutes of Health announced that the Mississippi infant had relapsed with recrudescence of viremia and the re-emergence of HIV antibodies. The child has subsequently re-initiated ART.

Highlights of Recent HIV Cure Studies  

An uptick in the number of cure-related studies has generated a plethora of data that is helping researchers better characterize the viral reservoir and decipher what strategies may help combat it. Below is a list of some recently published studies and their main findings.

  • Sizeable reservoirs of replication-competent HIV provirus, which although non-inducible in vitro, may nevertheless contribute to ongoing viral re-activation (Cell 155, 540, 2013).
  • The majority of HIV proviruses are not activated using HDAC inhibitors (Proc. Natl. Acad. Sci. USA111, 7078, 2014).
  • A therapeutic rhesus cytomegalovirus (RhCMV)/simian immunodeficiency virus (SIV) vector-based candidate in monkeys demonstrated durable control of infection following challenge with the highly pathogenic SIVmac239 virus (Nature 502, 100, 2013).
  • A cohort of patients in the VISCONTI study who initiated ART very early were subsequently able to control HIV replication in the absence of ongoing therapy, and are now termed “post-treatment controllers” (PLoS Pathog. 9, e1003211, 2013).
  • A dramatically reduced integrated HIV DNA reservoir in central memory CD4+ T cells was achievable by initiating ART in very early infection (CROI Abst 47, 2013).
  • Three perinatally HIV-infected infants who received ART from infancy achieved significantly reduced circulating levels of proviral and replication-competent HIV and sustained ongoing decay of their viral reservoirs (J. Infect. Dis. online May 21, 2014 doi: 10.1093/infdis/jiu297).
  • Macaques infected with the hybrid simian/human virus SHIV-SF162P3 that were administered a single broadly neutralizing antibody (PGT121) had a rapid and precipitous decline in plasma HIV RNA to undetectable levels. PGT121 administration resulted in reduced proviral DNA in peripheral blood, gastrointestinal mucosa, and lymph nodes without the development of viral resistance. Of the 18 antibody-treated macaques, three sustained undetectable viremia through necropsy at 100 days (Nature 503, 224, 2013).
  • A study of 12 chronically infected but virologically suppressed volunteers in a gene therapy study suggests infusion of autologous CD4-enriched T cells modified to remove expression of CCR5 by a zinc-finger nuclease is safe (N. Engl. J. Med. 370, 901, 2014). —NMS


These and other studies indicate that the pace of HIV cure research has picked up in the last two years, a trend confirmed by the latest report from the HIV Vaccines and Microbicides Resource Tracking Working Group (see In Brief). They estimate that global funding for cure research increased by 16% from 2012 to 2013, to a total of US$102.7 million. This may underestimate the contribution by industry, as companies with known programs in cure research did not provide data to the working group. Most of the new funding has come from the public sector, with less than $5 million from philanthropies such as Aides Fonds, amfAR, the Campbell Foundation, and Sidaction.

Ananworanich went on to discuss some of the social and ethical considerations around an HIV cure and suggested that society and individuals living with HIV expected that eradication would mean being free of disease, with no long-term adverse consequences of HIV and diminution of stigma and discrimination. She referred to the Australian research that sought to identify the outcome priorities of participants in a small clinical study of the histone deacetylase inhibitor (HDACi), vorinostat (also known as suberanilohydroxamic acid or SAHA). This study found that the four highest rated priorities for participants were: Not passing HIV onto others (47%); being considered uninfected (32%); not getting HIV a second time (32%); and stopping ART (25%). These priorities were congruent with those from a larger European community survey presented at AIDS 2012.

Fauci covered similar terrain in a review of the critical challenges in HIV cure and vaccine research. He noted that a cure generally denotes permanent remission from disease following cessation of therapy, which in the case of infectious diseases typically involves eradication of the microbe, and in cancer means an absence of relapse for life or for a pre-defined period of time determined as a surrogate for cure. After surveying what is known about the HIV reservoir, Fauci discussed what he thinks will be required for sustained virological remission (SVR) from HIV. He suggested that SVR might be most achievable in individuals who receive early ART, are stimulated to induce natural HIV-specific immunity, and receive passive transfer of HIV-specific antibodies and therapeutic vaccination.

Highlights from the Towards an HIV Cure symposium

Like Fauci, Jeff Lifson, director of the AIDS and Cancer Virus Program at the Frederick National Laboratory for Cancer Research and the keynote speaker at this year’s Towards an HIV Cure symposium, classified a functional cure as sustained off-treatment remission, involving reduction of the viral reservoir to levels low enough that with sufficient host control, HIV pathologies and the risk of transmission are reduced or eliminated. Achieving a functional cure is a much different goal than eliminating the virus entirely. And it remains to be seen what lessons can be learned from the recrudescence of virus in the Mississippi child, or whether a functional cure will indeed be possible if a viral reservoir, regardless how small, still exists, according to Lifson.

He reviewed the spectrum of current approaches to curing HIV, including: transcriptional activators to ‘shock’ HIV out of latency; epigenetic modulators (agents which are able to induce changes in the genes controlling behavior of HIV provirus); immune modulators, including interventions targeting immune checkpoint molecules such as PD-1 and its ligands; immune targeting, including broadly neutralizing monoclonal antibodies; and therapeutic vaccination. Lifson then focused on this final intervention and how non-human primate (NHP) models offer considerable advantages in evaluating therapeutic vaccines. However, NHP models, while accurately recapitulating many essential features of HIV infection, are limited in that HIV is not the same as SIV or the hybrid simian/human virus known as SHIV.

The potential role of therapeutic vaccination received a boost last year with the publication of results of the DCV2/MANON07-ORVACS study (Sci. Transl. Med. 5, 166ra2, 2013). In this study, participants received a therapeutic vaccine utilizing autologous monocyte-derived dendritic cells (MD-DCs) pulsed with autologous, heat-inactivated, whole HIV. The vaccine was administered in three doses before cessation of ART. Participants who received the vaccine pulsed with inactivated HIV had a significantly greater reduction in plasma viral load from baseline levels, with a maximum decrease in viral load at 12 weeks that persisted through 48 weeks.

The use of cytomegalovirus (CMV)-vectored vaccines has also shown promise in animal models. In a previous study, a majority of macaques vaccinated with a rhesus CMV/SIV vaccine established immune control within 12 weeks of established SIV infection following intra-rectal, intra-vaginal, or intravenous challenge, and this viral control persisted until necropsy (Nature 502, 100, 2013). Notably, disseminated infection was controlled, not just infection at the portal of entry. SIV viral load decreased over time at all sites, including plasma, gastrointestinal tract mucosa, lymph nodes, spleen, bone marrow, and tonsils. These results suggest a sustained virological remission occurred in these animals. Additionally, the vaccinated animals appeared to be incapable of infecting other animals. Lifson suggested this apparent viral clearance has many implications for the development of CMV-based therapeutic vaccines.

To further evaluate therapeutic vaccine candidates in NHPs, Lifson and colleagues developed an SIVmac239 clone with molecular tags to track individual infection events. They have also observed, in unpublished research, that the timing of ART initiation can profoundly influence the size and timing of reservoir establishment, as well as the variability in virological control. Animals treated from seven days post-infection all achieved undetectable plasma SIV RNA, and had SIV DNA levels that were 45% less than those who initiated ART weeks later (at day 42).

Additionally, in early treated animals, cell-associated viral DNA was 2.5 logs lower in lymph nodes, 1.5 logs lower in peripheral blood mononuclear cells (PBMCs), and 1.8 logs lower in bone marrow. Lifson suggested that viral persistence in T follicular helper (Tfh) cells might be an obstacle to complete viral clearance, but overall, these finding are encouraging and will be tested further in NHPs.

Can bNAbs Block HIV Dissemination in Dendritic Cells?   

At the Towards an HIV Cure symposium, which took place in advance of the AIDS 2014 conference in Melbourne in July, Bin Su from the Université de Strasbourg presented data illustrating the ability of broadly neutralizing antibodies (bNAbs) to inhibit HIV transmission from primary plasmacytoid dendritic cells (pDCs) to CD4+ T cells. pDCs are particularly rich in the submucosal epithelium, where they very effectively transfer HIV to CD4+ T cells. HIV replicates poorly in pDCs because of their expression of the host restriction factor SAMHD1, which blocks HIV replication in DCs, monocytes, and macrophages. Therefore, if bNAbs are able to block both cell-free transfer of HIV and pDC-facilitated transfer to CD4+ T cells, the presence of bNAbs could stop HIV in the submucosal epithelium before it has the opportunity to disseminate further.

To study this interaction, Su and his colleagues developed an HIV transfer assay to mimic early mucosal transmission of HIV infection. In this model in which pDCs and CD4+T cells were co-cultured, pDCs quite freely exhibited increased HIV replication in the presence of autologous CD4+ T cells, as the function of SAMHD1 was overridden. However, in the presence of VCR01 and PGT121, two recently identified bNAbs, transfer of HIV was inhibited in this model. When a 20μg/ml VCR01 solution was added to co-cultured cells two hours post-infection, 80% of HIV transmission from pDCs to CD4+ T cells was blocked, and the same amount of CD4+ T-cell free infection was inhibited. PGT121 even more effectively blocked HIV transfer from pDCs to CD4+ T cells, and also effectively neutralized cell-free infection.

Su observed that both bNAbs also induced pDC maturation and increased interferon-alpha (IFN-α) expression by infected pDCs in the presence of co-cultured CD4+ T cells. He suggested this increased immune surveillance and pDC maturation during pDC/T cell cross talk might promote effective innate immune responses and contribute to viral control. He suggested that a future role for vaccination might be to induce bNAbs directly at mucosal sites to prevent early dissemination of HIV after sexual exposure. —NMS


Eradicating HIV from lymph nodes

It has been known for almost a decade that the germinal center in lymphoid follicles is an anatomical site where HIV replication is active and where substantial reservoirs of inducible HIV are located. It was encouraging therefore to hear the results of research from Richard Koup, deputy director of the Vaccine Research Center at NIAID, on a novel bi-specific antibody (BibNAb), VRC07(fab)-anti-hCD3 that induces lysis of HIV-infected cells in lymph nodes.

This antibody is termed ‘bi-specific’ because it is active against the CD4 binding site of the HIV envelope and also induces redirected lysis of HIV-expressing CD4+ T cells by CD8+ T-cells. Koup’s team was able to demonstrate that administration of VRC07(fab)-anti-hCD3 led to 35-40% lysis of HIV-infected cells in lymph nodes. This antibody-mediated killing of HIV-infected cells was mediated by caspases (intracellular cysteine proteases) associated with increased secretion of granzyme B and perforin, which are the effectors of cell apoptosis.

Rama Amara, associate professor at the Yerkes National Primate Research Center at Emory University, also presented research involving lymph nodes. Amara’s work is focused specifically on the fate of PD-1+ CD4+ T cells in blood, lymph nodes, and rectal tissue in the presence of vaccine-mediated control of SIV-infection in a rhesus macaque model. Memory CD4+ cells expressing high levels of PD-1 (PD-1 high), accumulate preferentially in lymph nodes, where they are found at three-fold higher levels compared to plasma, and at 18-fold higher levels than in rectal tissues. Additionally, they are phenotypically Tfh cells, defined as such by high levels of the chemokine receptor CXCR5.

Amara and colleagues observed that when vaccinated macaques were challenged with SIV, they had much lower SIV viral load set points than unvaccinated animals (generally 3-4 logs lower). These SIV-controllers also had significant lower proportions of PD-1 high CD4+ T cells (levels similar to SIV-uninfected animals) in all three anatomical sites. This lower frequency of PD-1 high CD4+ T cells was strongly associated with plasma viral load. PD-1 high CD4+ T cells supported both SIV replication and production in lymph nodes in non-controllers during chronic infection.

Amara proposed several possible explanations of this reduced expansion of PD-1 high CD4+ T cells. First, higher frequencies of SIV-specific CD8+ T cells are found in lymph nodes of controllers, and there was a strong inverse correlation between SIV-specific CD8+ T cells (especially those expressing granzyme B) and PD-1 high CD4+ T cells. Secondly, there was a higher frequency of functional follicular CD8+ T cells in controllers. There was also an increased co-location of CD8+ T cells with PD-1 high cells within the lymph nodes of controllers. Amara’s team developed an in vitro CD8+ T-cell killing assay that was able to demonstrate that it was indeed SIV-specific CD8+ T cells located in the lymph node that were able to limit the proliferation of PD-1 high Tfh cells.

These data suggest that it may be possible to develop vaccine-based therapies to reduce or eliminate virus-infected CD4+ T cells located in the B-cell zone of lymphoid follicles by stimulating higher frequencies of cytotoxic T lymphocytes (CTLs). Such vaccine-based therapies might be augmented by other interventions to down-regulate PD-1 and its ligands.

Targeting HIV-infected cells as they come out of latency

David Margolis, director of the Program in Translational Clinical Research at the University of North Carolina, presented additional research on HIV reservoirs and how HIV-infected cells might be targeted as they come out of latency. His team is developing model systems to assess viral clearance from reservoirs, focusing in particular on how CTLs and natural killer (NK) cells might to be used to target reactivated cells.

CTLs have been well characterized through the work of oncologists who used expanded autologous CTLs ex vivoto treat viral infections, such as herpes viruses, in cancer patients. Margolis’s team has produced a suite of such expanded CTLs (styled HXTCs), which consist mainly of CD8+ T effector memory cells that elicit responses to gag,pol, and nef cognate peptides. Such HXTCs in a superinfection assay are demonstrated to blunt active viral replication and potently inhibit autologous reservoir virus. Additionally, they have been demonstrated to clear infected cells that have been stimulated from latency by both the global mitogen phytohemagglutinin (PHA), and also the more selective reactivation agent, vorinostat. Margolis’s team has demonstrated that vorinostat does not impair CD8+ or HXTC antiviral activity at physiologically relevant exposures.

Margolis also reviewed the role of NK cells, which are able to access HIV-infected cells that are protected from antiviral activity of T cells. NK cells are crucial innate immune effectors that do not recognize specific antigens or require prior antigen sensitization, and kill cells by insertion of granzyme and perforin, resulting in apoptosis. Their action may be augmented by clinically applicable cytokines such as interleukin (IL)-2 and IL-15. His team used NK cells stimulated with either of these cytokines to inhibit autologous HIV by over 90%. In a latency clearance assay they were able to demonstrate that cytokine-stimulated NK cells were able to clear almost all virus coming out of latency. Margolis said plans are to test this approach clinically within the next six to eight months.

Advances in HIV prevention

Discussion of PrEP is hardly new at international AIDS conferences, but as evidence of its effectiveness in at-risk populations mounts, recommendations for its use are also being strengthened. In the 2012 guidelines issued by the World Health Organization (WHO), PrEP was recommended upon review of evolving evidence, however, the recently updated guidelines now strongly recommend PrEP use based on high quality evidence.

At a media briefing, Chris Beyrer, incoming president of the International AIDS Society, emphasized that the guidelines recommend PrEP as “an additional prevention option for men who want it,” as part of a comprehensive set of interventions. He explained that an individual’s preferred HIV prevention method, like contraceptive options for women, may change over an individual’s lifetime, and PrEP is now another possible option.

Bob Grant, Director of Gladstone Institute of Virology and Immunology at the University of California, San Francisco, presented late-breaking findings from the iPrEx OLE trial, an opt-in, open-label extension Phase of the original iPrEx trial that showed the fixed dose combination antiretroviral Truvada (tenofovir disoproxil fumarate and emtricitabine, or FTC) was an effective PrEP strategy in men who have sex with men and transgender women. These findings were published simultaneously (Lancet Infect. Dis. online Jul 22, 2014 doi: 10.1016/S1473-3099(14)70847-3).

Among study participants who elected to take PrEP, regardless of the frequency and regularity with which they did so, the annual HIV incidence rate was 1.8%, compared to 2.6% for those who opted out of taking the drug. Overall effectiveness of PrEP during this study was about 50%. Among participants who took at least four doses of drug weekly, there were no new HIV infections.

However, as in the original study, poor adherence was strongly associated with incident infections. Only about one-third of OLE participants took drug regularly, with younger people being less likely to have measurable drug levels in blood. However, adherence was better among people who reported more high-risk sex or more sexual partners, indicating that they may have adopted PrEP as a perceived risk-reduction strategy.

Intermittent PrEP

Given the overall poor adherence to PrEP in clinical trials, the Agence nationale de recherches sur le sida et les hépatites virales’ (ANRS) IPERGAY study is evaluating the efficacy of intermittent or “on demand” PrEP in an ongoing, randomized, double-blind, placebo-controlled trial. The trial began enrolling gay and bisexual men in France and Canada in early 2012. Participants are randomly assigned to take two Truvada or placebo pills 24 hours before they expect to have sex, and one pill at both 24 and 48 hours afterwards.

As the trial is still enrolling, data on effectiveness are not yet available, however, Jean-Michel Molina, from Saint-Louis Hospital reported early findings on adherence in Melbourne. The interim analysis included 129 men with an average age of 35 who reported having a median of two instances of sexual intercourse per week (range: 0-31), and a median of 10 partners over the previous two months. About 80% of participants reported that they had used PrEP the last time they had sex. Based on pill counts, they took an average of 15 pills per month, meaning they were on PrEP about half the time. At any clinic visit approximately 86% of participants had detectable levels of tenofovir in blood, and 82% had detectable levels of FTC.

Circumcision continues to deliver

Male circumcision as an HIV prevention strategy continues to produce encouraging results. The WHO and the Joint United Nations Programme on HIV/AIDS recommend VMMC as an additional intervention for prevention of heterosexually acquired HIV, particularly in settings with generalized HIV epidemics. And the most recent data from Uganda show that in the five years since the Rakai trial—one of the first to establish the efficacy of VMMC in preventing HIV infection—was completed, high effectiveness has been maintained among the men who were circumcised, with a 73% protective effect against HIV infection.

Other benefits of circumcision were reported by Kévin Jean of the Institut national de la santé et de la recherche médicale (INSERM), who presented data from the ANRS-12126 study in Orange Farm, South Africa. He reported for the first time that male circumcision not only protected men but also was beneficial for their female partners. VMMC protects against some sexually transmitted infections in both men and women, reducing the risk of herpes simplex virus-2 and human papilloma virus acquisition in men and their female partners. It is also associated with a reduction in the risk of genital ulcer disease and genital cancers in both men and women. However, protection against HIV in women was not observed in epidemiological studies until now. The ANRS showed that HIV incidence among women who only had sex with circumcised men was reduced by 17% in those aged 15-49 years, and 20% in women aged 15-29 years, a notable outcome in a context where HIV prevalence in women aged 15-45 years is around 32%.

In a related presentation, Jillian Pintye from the University of Washington reported on new data from the Partners PrEP Study that shows circumcised men are also less likely to acquire syphilis. She reported that syphilis incidence in participants in this study involving heterosexual, mostly married, serodiscordant couples in Africa was a statistically significant 42% lower in circumcised men. When stratified by HIV serostatus, the researchers found a significant risk reduction of 62% among HIV-infected men, and a similar non-significant trend among HIV-uninfected men. In women, there was a statistically significant 59% risk reduction of syphilis associated with having a circumcised male partner. These results extend even further the health benefits associated with VMMC.

Neil McKellar-Stewart is HIV Health Promotion Officer at ACON (AIDS Council of New South Wales) and an active member of the National Association of People with HIV Australia Treatment Officers Network. He is a community representative on the Australasian Society of HIV Medicine Sub-Committee for Guidance in HIV Management in Australia.