Women, Men and Microbicides

We've heard a lot about HIV entry inhibitors in 2003. The new injectable fusion blocker Fuzeon (T-20, enfuvirtide) is the first member of this new class of HIV drugs to hit the market, and several oral drugs intended to block HIV entry into cells at other steps are just starting clinical trials. As a class, the entry inhibitors are designed to keep HIV from merging with a new cell in someone who is already infected. But before HIV can put down roots and start infecting those billions of immune cells, at least one viral particle must make its way past the body's skin or mucosa -- barriers that are supposed to keep the outside out, and the inside in. This premiere moment is called transmission, and new understanding about how the virus first enters the body is stimulating progress in the fields of therapies, vaccines, and topical microbicides.

Direct blood-to-blood transmission of HIV seems straightforward. People infected through a transfusion, blood products or shared injection equipment probably received a significant dose of the virus in multiple forms that efficiently found its way into their immune system. Immediate post-exposure prophylaxis with antiretroviral (ARV) drugs has a good record for preventing blood-borne infections from needle-stick accidents and may prevent some sexually transmitted infections as well. Another main route of exposure is through mother-to-child transmission during pregnancy or birth. It was eventually discovered that most newborns could be spared from infection by reducing the viral load of the mother with antiretroviral drugs before labor, or in some cases, by choosing C-section over vaginal delivery.

But sexual transmission of HIV is the leading source of infection, and although infections between men having sex were most common at the beginning of the epidemic, nowadays infections passed between men and women are pushing the scale of the world AIDS crisis into uncharted territory. With an estimated 14,000 new infections occurring every day, and as many as 42 million infected worldwide, preventing new infections between men and women is an unmet emergency. The most available lines of defense are education, behavior change and condom use. Simply not having sex or assiduously using a condom every time can do wonders for reducing the infection rate. But like all miracle cures, education and condom use has proved too good to be true. Most new infections occur among young people -- a group powerfully motivated to have sex -- and among people with little control over the material conditions of their lives, such as men and women in resource poor areas who lack the power, education and opportunities to protect themselves.

Beyond education and condoms, public health policy has pushed for development of a vaccine. In a few historical cases, vaccines have been able to effectively protect mass populations from communicable pathogens at a rock bottom, one-shot and on-with-your-life cost that sounds like salvation. As HIV spreads unrestrained in some regions; when as many as 40 percent of a nation's citizens are in danger of becoming infected, even a partially effective vaccine that can't reliably protect any particular individual would have a huge impact on the social catastrophe of AIDS.

But a vaccine for HIV has proved difficult to solve. In every year since the mid-1980s, someone has estimated that a vaccine will appear in the next 5 to 10 years; wise scientists now refuse to guess when one will come. Because HIV infects and influences the very immune system called upon to fight it, vaccine research remains vexed by unanswered questions of basic science. Meanwhile others have been looking for simpler solutions. Over a dozen years ago, seeing the rising worldwide death toll prefigured in epidemiology and social reality, some public health thinkers (such as Zena Stein) began to propose novel ways to augment conventional, barrier-based prevention methods. They first recognized that male condoms would never be a viable option for every woman because, among many reasons, the technology depended upon male participation to be effective. In too many cases and cultures, men simply refuse to accept condoms, and women become infected, powerless to object. With the chances of infection during one episode of heterosexual sex put at 1 in 200 or less, even a partially protective method could lower that risk and start saving lives.

Enter the Microbicide

One clever proposal for keeping HIV out of the body is to apply a liquid or gel substance before sex that could block infection by physical, chemical or medicinal means. Known generically as a microbicide, the idea was derived from products to prevent pregnancy that were already on the market. (One contraceptive product, N-9 or nonoxynol-9, was initially thought to have anti-HIV properties until it was discovered that N-9 actually increased susceptibility to HIV infection by breaking down the body's natural cellular barriers to microbes.) Despite the exciting potential for an inexpensive method that can be used without the participation of a partner, microbicide research has met with some of the same problems slowing vaccine development. And microbicides have posed a new set of challenges altogether, ranging from applicator design to placebo validation. Simply coming up with a gel to stop viruses from sticking to their targets has not proved as elementary as many thought.

A highly effective microbicide would likely protect against HIV transmission in multiple ways and might even act against several other kinds of microbial invaders. Begin with the gels and foams. These are based on a medium that would be inserted, squirted or secreted into a vagina or rectum and may or may not carry other active ingredients. But before a product can be found effective, it must first be found safe, tolerable and acceptable. For example, if the product is a gel, the physical properties must be right -- not too sticky, too runny, or prone to dry out. These are qualities that matter to the user, and different users are likely to have different needs and preferences. Then there are the chemical properties to consider. Some forms of gel might be designed to simply keep HIV from ever coming into contact with the mucus membranes that line our vulnerable body cavities. Or it may contain chemicals to help maintain the naturally protective acidic environment of the vagina. Another gel might be optimized to carry an active ingredient that disrupts HIV's lipid membrane. Still another type might be best for delivering an antibody, a vaccine or a drug to the mucosal tissue so it can penetrate into the submucosal layers and become active. A good gel must avoid harming the mucosal barrier cells or any submucosal cells that become exposed through tiny breaks and tears in the vagina, cervix or rectum. It must not cause irritation if it is left on overnight, especially to unsuspecting penises. Furthermore, the gel must not set off any local or general immune response in the vast majority of people who use it; the last thing you want to do is attract immune cells -- the primary target of HIV -- to the scene. All of these stringent requirements mean that any gel expected to perform in a mass-produced microbicide -- not to mention the active ingredients it carries -- must be thoroughly tested to prove it is safe.

Although there is a long list of potential microbicide candidates, only a handful of products have advanced far enough through the clinical trials process for them to realistically become available within the next five years. And lack of investment by industry and government is holding back more rapid development. The candidates furthest along the pipeline work in non-specific ways; most by impeding viral access to vulnerable mucosal tissues.

How HIV Gets Through Mucosal Tissue
How HIV Gets Through Mucosal Tissue

Our Mucosa -- Basic Biology

The biology of the mucosal tissues that line our body parts susceptible to infection determines how likely and how preventable sexual transmission of HIV will be. These tissues are composed of various types of epithelial cells arranged in either layers or in columns. The vagina, outer cervix, anus and foreskin (of an uncircumcised penis) are covered by overlapping layers of epithelium in a fish scale-like structure called pluristratified mucosa. The upper cervix and rectum are lined with a single layer of columnar shaped epithelium called monostratified mucosa (see illustration). There is also a brief transition zone that bridges these tissue types in both the vagina and the rectum. Another theoretically infectable region of mucosa is in the mouth and throat, although saliva seems to provide a natural microbicidal action and infection of these tissues, while it does occur, is relatively rare.

In healthy, intact pluristratified mucosa, as in the vagina, one mechanism for infection is thought to use a type of immune cell called a dendritic cell, which moves through the submucosal layers of tissue and sends its dendrites (octopus-like arms) into the stratified epithelium to scan for foreign pathogens. Dendritic cells (DC) carry a cell-surface receptor called DC-SIGN that is capable of binding to HIV's gp-120 spike. In CD4 T-cells, the predominant target for HIV, attachment of one of the viral envelope spikes to a CD4 receptor begins the process of fusion and infection. But in dendritic cells (mature cells at least; immature DCs may be infectable), attachment to HIV causes the virus particle to be taken inside a bubble-like vesicle within the cell where it stays while the DC continues its immune patrol of the mucosa. After the dendritic cell finally leaves the mucosa and makes its way back to the immune system's regional headquarters in a nearby lymph node (a process that may take several days), it puts the HIV virion on display for other immune cells to inspect. When a CD4 T-cell comes along that recognizes the virus as an outsider, contact is made. Unfortunately, HIV uses this very act of self-defense to enter the CD4 cell and hijack it into making new viral copies. As new HIV virions start to bud off from the infected cell, they quickly board other CD4 cells in the lymphatic neighborhood and the primary infection is launched. Within days, millions of immune cells are infected and distributing the virus throughout the body.

How the Leading Microbicides Work
How the Leading Microbicides Work

In healthy, intact monstratified mucosa, the process is thought to be a little different. Although it's not exactly clear how, it seems that a receptor on the columnar cells with properties similar to DC-SIGN attaches to HIV and causes it to be internalized into the cell where it is passed through to the other side in a process called transcytosis. Once deposited on the submucosal underside of the epithelial barrier, the virus may come into contact with a patrolling CD4 cell or other immune sentry cell and infect it directly. As with dendritic cells, the CD4 cell likely carries the virus back to a lymph node where the infection is amplified.

But mucosal tissue is rarely completely healthy and intact, and variations on these paths to infection may be common. Microscopic nicks and tears in the mucosal barrier due to physical abrasion may allow direct contact between the outer environment and the submucosal immune cells to occur. The monostratified cells of the rectum are particularly vulnerable to physical damage during sex. Vaginal infections such as chlamydia or herpes may disrupt the protective mucosa and enhance HIV transmission by attracting target immune cells to the inflamed region. There are other wrinkles and exceptions to these basic modes of sexual transmission. For instance, not all virus is free floating; HIV may also be transmitted via a virus-laden cell in the semen that crosses the epithelial barrier like a Trojan Horse. But once HIV has made it into the mucosa, the barrier-based methods have failed. The next challenge for microbicide research is to find ways to stop an infection in its earliest stages.

Products at the Head of the Pipeline

Product NamePipeline StatusSponsorHow It Works
CarraguardPhase II safety studies in progress. Phase III to begin in 2004 .Population CouncilForms protective coating
PRO2000Phase IIB planned.Indevus PharmaceuticalsForms protective coating
BuffergelPhase IIB planned.ReProtect LLCMaintains normal vaginal pH
Savvy (C-21G)Phase I/II completed.Biosyn, Inc.Disrupts viral membranes
Emmelle (dextrin-2-sulfate)Phase II safety studies in progress.Multiple SponsorsForms protective coating
Ushercell (cellulose sulfate)Phase II in progress.Multiple SponsorsForms protective coating

The Bottleneck in Microbicides Development

In the absence of major pharmaceutical industry participation, a number of universities and small, independent biopharmaceutical firms have taken the lead on microbicide research. But in order to fund their research, these entities require public grants and -- to the extent they can raise it -- venture capital. The result: chronic underfunding and a clogged research pipeline.

In 2002, a major economic analysis of the field concluded that if a single pharmaceutical company were managing all microbicide research leads, that company would have to invest $775 million over five years to ensure the production of at least one safe, effective product. The Rockefeller analysis was a "bare bones" scenario that only considered the costs directly related to product development, omitting other necessities like basic research, discovery of new leads and work to assure that the products will be acceptable and accessible to users.

The report also showed that if current funding levels continue, the amount spent on microbicide research and development (R&D) worldwide between 2001 and 2005 would total about $230 million. This leaves a $545 million shortfall at minimum between current funding levels and the expected cost of getting one successful microbicide on the market. Even the recent generosity of the Bill & Melinda Gates Foundation doesn't cover that gap.

Over 60 potential microbicides have been identified to date, yet most are stuck in the preclinical phase because funding to move them into human trials isn't available. The few candidate products of proven safety have not moved forward in 2003 as planned because their sponsors are unable to support the cost of large Phase III effectiveness trials.

Microbicide R&D cannot advance efficiently without substantially increased governmental and foundation funding. At present, the U.S. National Institutes of Health (NIH) invests only about 2% of its AIDS-related research budget in microbicide R&D.

Anna Forbes -- Global Campaign for Microbicides Advocacy

New Research

Certain areas of advanced microbicide research overlap with work going on in the HIV vaccine field seeking to target the virus with exquisite selectivity. In the early pipeline are proposals ranging from small molecule inhibitors to mucosally-directed vaccines. At the AIDS Vaccine Conference held in New York City in September 2003, some new ideas emerged about how HIV enters the body that may help focus the development of a topical (applied to the skin) method for preventing sexual transmission using antibodies to neutralize infectious HIV.

One new observation has to do with the specific kind and quality of HIV that can be transmitted sexually. It's long been recognized that only viruses that use the CD4 cell's CCR5 receptor during entry seem to be present in newly infected people. It was thought that the barrier epithelium or dendritic cells filtered out the CXCR4-using HIV at the point of transmission. This less-common form of HIV sometimes appears later in the disease and is associated with rapidly progressing AIDS.

New work by Eric Hunter and colleagues from the University of Alabama, Birmingham (UAB) now suggest that the kind of virus most likely to be transmitted may also be especially vulnerable to immune attack -- that is if the immune system has been prepared to recognize it. Their work drew upon a highly productive research project conducted by UAB in collaboration with researchers in Zambia. The study has followed a cohort of sero-discordant (one positive, one negative) couples in Lusaka for over eight years. Couples in the study are counseled about safe sex and provided with condoms. About 8 percent of the partners become infected each year; yet because counseling and condoms are effective, this is a reduction from an expected infection rate of 20 percent per year.

The researchers obtain blood samples from participants throughout the study. If a partner becomes HIV-positive, the pair's samples are analyzed to determine the genetic sequence of the gp120 viral envelope protein responsible for attachment and entry. Hunter presented 8 cases of transmission, with 4 from male to female and 4 from female to male. In every case, the received virus was of the CCR5-using type. Unexpectedly, the genetic analysis showed that a particular envelope region on the received virus was unusually compact, lacking several features characteristic of a typical virus as found in established infections. Furthermore, this envelope region of the received virus did not resemble that on the virus obtained from the donor's blood. What was particularly exciting about this finding was that the received virus was unusually susceptible to neutralization by several specific antibodies, while the virus obtained from the donor's blood was protected against these antibodies. This may mean that a virus that is especially suited for sexual transmission may also be especially vulnerable to antibody neutralization. If so, then there is a possibility that, one day, these antibodies might be elicited in the mucosa by a vaccine or perhaps delivered by a microbicide to attack newly transmitted virus.

While the UAB study did not find a virus in the blood of the donor that matched the virus that was transmitted, they were not able to look at virus that may have been contained within the genital tracts. It is possible that a protected compartment allows the transmission-specialized virus to exist without competition from the more accessible viral strain coursing through the blood.

Although some vaccines under investigation seek to benefit chronically infected individuals by stimulating cellular immunity, the findings about a vulnerability in transmitted virus mainly applies to the uninfected, since HIV seems to begin mutating a protective carbohydrate (glycan) cover for its vulnerable spots soon after a primary infection has taken hold. However, there has been some evidence that these vulnerable epitopes may eventually reappear on gp120 as an infection matures; once the evolving shield of protective glycans begins to let down its guard against those early, presumably long gone, antibodies.

Although some vaccines under investigation seek to benefit chronically infected individuals by stimulating cellular immunity, the findings about a vulnerability in transmitted virus mainly applies to the uninfected, since HIV seems to begin mutating a protective carbohydrate (glycan) cover for its vulnerable spots soon after a primary infection has taken hold. However, there has been some evidence that these vulnerable epitopes may eventually reappear on gp120 as an infection matures; once the evolving shield of protective glycans begins to let down its guard against those early, presumably long gone, antibodies.

With so much still to be learned about the basic science of HIV infection, and so much intractable about the social reality that allows HIV to flourish, the impact of either an effective vaccine or microbicide remains many years away. Yet workers in the field remain hopeful -- and with good reason. The energy and commitment evidenced by microbicide and vaccine advocates, and the increasing elucidation of the underlying science, argues that an eventual breakthrough is inevitable.