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By identifying a protein that restricts the release of HIV-1 virus from human cells, scientists believe they may be closer to identifying new approaches to treatment. The research is published in the advance online edition of Nature Medicine.

Scientists have known that most human cells contain a factor that regulates the release of virus particles, but until now they have been uncertain about the factor’s identity. Now a research team from Emory University School of Medicine, Vanderbilt University School of Medicine, and Mayo Medical School has identified CAML (calcium-modulating cyclophilin ligand) as the cellular protein that inhibits the release of HIV particles.

CAML works by inhibiting a very late step in the virus lifecycle, leading to the retention of HIV particles on the membrane of the cell. The virus has developed a means of counteracting CAML, through the action of the viral Vpu protein. When Vpu is absent, HIV particles don’t detach from the plasma membrane and instead accumulate by a protein tether at the cell surface.

When the research team depleted CAML in human cells in the laboratory, they found that Vpu was no longer required for the efficient exit of HIV-1 particles from the cell. When they expressed CAML in cell types that normally allow particles to exit freely, the particles remained attached to the cell surface.

“This research is important because it identifies CAML as an innate defense mechanism against HIV,” says senior author Paul Spearman, professor of pediatrics (infectious diseases) at Emory University School of Medicine. “We are continuing to work on the mechanism that Vpu uses to counteract CAML and on defining exactly how CAML leads to virus particle retention on the infected cell membrane. We hope this will lead us to new treatments.”

Nature Medicine

Identification of calcium-modulating cyclophilin ligand as a human host restriction to HIV-1 release overcome by Vpu

Vasundhara Varthakavi, Ellen Heimann-Nichols, Rita M Smith, Yuehui Sun, Richard J Bram, Showkat Ali, Jeremy Rose, Lingmei Ding & Paul Spearman

Published online: 25 May 2008; | doi:10.1038/nm1778

Researchers have developed what they believe is the first new mechanism in nearly 20 years for inhibiting a common target used to treat all HIV patients, which could eventually lead to a new class of AIDS drugs.

Image of the molecule researchers discovered that keeps the flaps from closing on the HIV-1 protease. It inhibits the flaps from closing, and prevents the protease from assembling an active virus. (Credit: Kelly Damm)

Researchers at the University of Michigan used computer models to develop the inhibiting compound, and then confirmed in the lab that the compound does indeed inhibit HIV protease, which is an established target for AIDS treatment. The protease is necessary to replicate the virus, says Heather Carlson, U-M professor of medicinal chemistry and principal investigator of the study.

Carlson stresses this is a preliminary step, but still significant.

“It’s very easy to make an inhibitor, (but) it’s very hard to make a drug,” said Carlson, who also has an appointment in chemistry. “This compound is too weak to work in the human body. The key is to find more compounds that will work by the same mechanism.”

What’s so exciting is how differently that mechanism works from the current drugs used to keep the HIV from maturing and replicating, she says. Current drugs called protease inhibitors work by debilitating the HIV-1 protease. This does the same, but in a different way, Carlson says.

A protease is an enzyme that clips apart proteins, and in the case of HIV drugs, when the HIV-1 protease is inhibited it cannot process the proteins required to assemble an active virus. In existing treatments, a larger molecule binds to the center of the protease, freezing it closed.

The new mechanism targets a different area of the HIV-1 protease, called the flap recognition pocket, and actually holds the protease open. Scientists knew the flaps opened and closed, but didn’t know how to target that as a mechanism, Carlson says.

Carlson’s group discovered that this flap, when held open by a very small molecule—half the size of the ones used in current drug treatments—also inhibits the protease.

In addition to a new class of drugs, the compound is key because smaller molecules have better drug-like properties and are absorbed much more easily.

“This new class of smaller molecules could have better drug properties (and) could get around current side effects,” Carlson said. “HIV dosing regimes are really difficult. You have to take medicine several times in the day. Maybe you wouldn’t have to do that with these smaller molecules because they would be absorbed differently.”

Kelly Damm, a former student and now at Johnson & Johnson, initially had the idea to target the flaps in this new way, Carlson says.

“In a way, this works like a door jam. If you looked only at the door when it’s shut, you’d not know you could put a jam in it,” she said. “We saw a spot where we could block the closing event, but because everyone else was working with the closed form, they couldn’t see it.”

Kelly L. Damm, Peter M. U. Ung, Jerome J. Quintero, Jason E. Gestwicki, Heather A. Carlson.

A poke in the eye: Inhibiting HIV-1 protease through its flap-recognition pocket.

Biopolymers. Volume 89, Issue 8 , Pages 643 - doi: 652.10.1002/bip.20993

Abstract
A novel mechanism of inhibiting HIV-1 protease (HIVp) is presented. Using computational solvent mapping to identify complementary interactions and the Multiple Protein Structure method to incorporate protein flexibility, we generated a receptor-based pharmacophore model of the flexible flap region of the semiopen, apo state of HIVp. Complementary interactions were consistently observed at the base of the flap, only within a cleft with a specific structural role. In the closed, bound state of HIVp, each flap tip docks against the opposite monomer, occupying this cleft. This flap-recognition site is filled by the protein and cannot be identified using traditional approaches based on bound, closed structures. Virtual screening and dynamics simulations show how small molecules can be identified to complement this cleft. Subsequent experimental testing confirms inhibitory activity of this new class of inhibitor. This may be the first new inhibitor class for HIVp since dimerization inhibitors were introduced 17 years ago. © 2008 Wiley Periodicals, Inc. Biopolymers 89: 643-652, 2008.

Australian researchers have unveiled a new immunotherapy technique to help prevent the progression from HIV infection to AIDS. Details of the simple, cost-effective technique are published May 2nd in the open-access journal PLoS Pathogens.

There is an overwhelming need for effective immunotherapies for HIV, as current therapies are expensive, impractical, and often highly toxic. The authors, led by Professor Stephen Kent, propose a technique named OPAL therapy–Overlapping Peptide-pulsed Autologous CeLls–a reinfusion of fresh blood cells incubating with overlapping SIV peptides. The OPAL technique was successfully tested in animal trials for stimulation of immunity, control of virus levels, and prevention of AIDS.

Vaccination diminished the levels of virus 10-fold lower than in controls, and was shown to be durable for over one year past initial vaccination. Therefore, viral replication was shown to be prolonged and more manageable, resulting in fewer deaths from AIDS.

The study is the result of collaboration among researchers from the University of Melbourne, the National Serology Reference Laboratory, and the University of New South Wales. The researchers plan to conduct future OPAL-therapy clinical trials in HIV-infected humans.

PLoS Pathog 4(5): e1000055. doi:10.1371/journal.ppat.1000055

Any hunter will tell you that when your quarry goes into hiding, you have to flush it out to get a good shot at it. Such is the case with HIV, the virus that causes AIDS.

Though antiretroviral “cocktails” can target an active infection, they cannot get at the virus when it retreats inside the host’s T cells, where it may lie dormant for decades, waiting for an opportunity to burst forth in a fresh round of infection. What HIV hunters need is a good bird dog.

Now, Stanford chemist Paul Wender and his coworkers have found a way to synthesize better bird dogs, agents that can be tailored to flush HIV out into the open where the immune system and antiretroviral therapies can destroy it. Wender is senior author of a paper about the research in the May 2 issue of Science.

“We’re not sure how far this will go, but certainly, from a theoretical point of view, it has promise of taking therapy to the next level—that is, addressing issues related to eradication of the disease, of the virus, at least,” said Wender, the Francis W. Bergstrom Professor.

Wender and his co-workers Jung-Min Kee and Jeff Warrington have developed a way to synthesize prostratin and DPP, two compounds that occur naturally in plants, in the laboratory. Prostratin, found in the Mamala plant (Homalanthus nutans) that grows in the Samoan rainforest, has shown promise in previous studies as an activator of dormant HIV. DPP, a molecular relative of prostratin found in resin spurge (Euphorbia resinifera), which grows in arid regions, also has shown potential.

Research has been hampered, though, because the compounds are difficult to obtain, particularly in the quantities needed for practical lab work on their mode of action and therapeutic potential. The yield from both plants is low and highly variable; the availability of the plants is limited; and isolating the compound is difficult. Heavy harvesting of the wild plants, especially in Samoa, also could cause ecological damage.

But synthetic prostratin and DPP, which now can be readily made in the lab, changes that equation.

“We have now minimized, if not eliminated, the issue of availability of prostratin and DPP,” Wender said. “But equally, if not more importantly, we have opened access to other compounds that might be similar in structure but superior in function.”

Previous work done in mice by researchers at the University of California-Los Angeles indicates that prostratin, used in combination with interleukin-7, an immune system stimulator made in bone marrow, managed to flush out and eliminate approximately 80 percent of the dormant virus. But with HIV, 80-percent efficiency is not enough. Anything less than 100 percent means the virus is still lurking in the T-cells and will spring back to action as soon as an opportunity presents itself.

“Nature has produced these compounds for various reasons in the plants from which they’re derived, but certainly not to treat human maladies,” Wender said. “They’re not optimized for human therapy.”

But with synthetic prostratin and DPP available, researchers can take the basic compounds and tinker with the structure and related function. “We could find out how to improve them by reverse engineering: figuring out what is important and what isn’t important,” Wender said. “We could begin to design and synthesize molecules that would never be found in nature but might actually be therapeutically more beneficial than what has been found thus far.”

In the Science paper, Wender and his team detail how both compounds can be synthesized, but also show the initial phase of designing and making new derivative compounds.

Although prostratin has long been used by traditional Samoan healers without their patients experiencing acute side effects, it is possible that undesirable effects could show up in an immune-impaired patient taking prostratin or DPP. But Wender noted that engineering the compounds in a lab would permit scientists to circumvent these problems. “Usually these kinds of side effects are a result of a drug hitting multiple targets. So it hits one target, which is beneficial, but then it hits some other target, too,” he said. “But by modifying the structures, you could select for the beneficial activity over the non-beneficial activity.”

“It’s a little bit like draw poker,” Wender said. “The important point is that we’re not forced to use the hand we get. We’ll get a hand and we’ll return a few cards if we don’t like it, because we can keep on tuning this until we get it right, so that a royal flush, hopefully, can be realized.”

Wender’s team developed their method of synthesizing prostratin and DPP by using a renewable resource, croton oil, made from the seeds of a small tree (Croton tiglium) cultivated in Asia. They derived phorbol from the croton oil and then converted it into the structure of prostratin.

The conversion process required some engineering finesse; they had to overcome a hurdle when, by removing an oxygen atom, they triggered a series of anticipated but seemingly undesired changes.

“To the credit of my coworkers, Jung-Min Kee and Jeff Warrington, they employed a strategy that sometimes is missed,” Wender said. “Rather than fighting the flow, they went with it.” They found a way to redirect the chemical complications into a solution to the problem that proved even better than the route they had initially sought to follow.

“Eventually they produced a shorter, more economical way of connecting our starting material, phorbol, to our target, prostratin,” Wender said. The process Kee and Warrington came up with requires only five steps, which is of tremendous importance in making it economically feasible. As Wender pointed out, “steps cost money and human time.”

Wender emphasized that the work of his team is the most recent chapter in efforts of a truly global community, starting with the Samoan healers, who willingly shared their knowledge with Paul Cox, an ethnobotanist who saw them prescribing a tea made from Mamala bark for patients with hepatitis-like symptoms. Cox, in turn, sent samples to the National Institutes of Health, in hopes that the bark might have antiviral properties useful in fighting some cancers. Researchers at NIH then analyzed the bark and isolated prostratin.

Prostratin belongs to a class of compounds called tiglianes, many of which promote tumor growth, so it had no initially perceived use in fighting cancer. But NIH researchers found that prostratin was not a tumor promoter and checked to see if perhaps it could help combat HIV, which is when its remarkable ability to flush out the dormant virus was discovered. Significantly, prostratin has also been found to block uptake of the purged virus, offering yet another potentially therapeutic benefit.

“The whole effort is a testimonial to a global community working to deal with what I think is a global, and top priority, problem,” Wender said.

The research was funded by the National Institutes of Health. At the time of the study, Kee was a doctoral candidate in chemistry and Warrington was a postdoctoral scholar at Stanford. Kee is now a postdoctoral scholar at Rockefeller University, and Warrington is working in the biotech industry.

cience 2 May 2008: Vol. 320. no. 5876, pp. 649 - 652 DOI: 10.1126/science.1154690
Reports
Practical Synthesis of Prostratin, DPP, and Their Analogs, Adjuvant Leads Against Latent HIV
Paul A. Wender,* Jung-Min Kee, Jeffrey M. Warrington

Department of Chemistry and Department of Chemical and Systems Biology, Stanford University, 337 Campus Drive, Stanford, CA 94305, USA.

A research group supported by the National Institutes of Health (NIH) has uncovered a new route for attacking the human immunodeficiency virus (HIV) that may offer a way to circumvent problems with drug resistance. In findings published April 28 in the online edition of the Proceedings of the National Academy of Sciences, the researchers report that they have blocked HIV infection in the test tube by inactivating a human protein expressed in key immune cells.

Most of the drugs now used to fight HIV, which is the retrovirus that causes acquired immune deficiency syndrome (AIDS), target the virus’s own proteins. However, because HIV has a high rate of genetic mutation, those viral targets change quickly and lead to the emergence of drug-resistant viral strains. Doctors have tried to outmaneuver the rapidly mutating virus by prescribing multi-drug regimens or switching drugs. But such strategies can increase the risk of toxic side effects, be difficult for patients to follow and are not always successful. Recently, interest has grown in attacking HIV on a new front by developing drugs that target proteins of human cells, which are far less prone to mutations than are viral proteins.

In the new study, Pamela Schwartzberg, M.D., Ph.D., a senior investigator at the National Human Genome Research Institute (NHGRI), part of NIH; Andrew J. Henderson, Ph.D., of Boston University; and their colleagues found that when they interfered with a human protein called interleukin-2-inducible T cell kinase (ITK) they inhibited HIV infection of key human immune cells, called T cells. ITK is a signaling protein that activates T cells as part of the body’s healthy immune response.

“This new insight represents an important contribution to HIV research,” said NHGRI Scientific Director Eric D. Green, M.D., Ph.D. “Finding a cellular target that can be inhibited so as to block HIV validates a novel concept and is an exciting model for deriving potential new HIV therapies.”

When HIV enters the body, it infects T cells and takes over the activities of these white blood cells so that the virus can replicate. Eventually, HIV infection compromises the entire immune system and causes AIDS. The new work shows that without active ITK protein, HIV cannot effectively take advantage of many signaling pathways within T cells, which in turn slows or blocks the spread of the virus.

“We were pleased and excited to realize the outcome of our approach,” Dr. Schwartzberg said. “Suppression of the ITK protein caused many of the pathways that HIV uses to be less active, thereby inhibiting or slowing HIV replication.”

In their laboratory experiments, the researchers used a chemical inhibitor and a type of genetic inhibitor, called RNA interference, to inactivate ITK in human T cells. Then, the T cells were exposed to HIV, and the researchers studied the effects of ITK inactivation upon various stages of HIV’s infection and replication cycle. Suppression of ITK reduced HIV’s ability to enter T cells and have its genetic material transcribed into new virus particles. However, ITK suppression did not interfere significantly with T cells’ normal ability to survive, and mice deficient in ITK were able to ward off other types of viral infection, although antiviral responses were delayed.

“ITK turns out to be a great target to examine,” said Dr. Schwartzberg, noting that researchers had been concerned that blocking other human proteins involved in HIV replication might kill or otherwise impair the normal functions of T cells.

According to Dr. Schwartzberg, ITK already is being investigated as a therapeutic target for asthma and other diseases that affect immune response. In people with asthma, ITK is required to activate T cells, triggering lung inflammation and production of excess mucus.

“There are several companies who have published research about ITK inhibitors as part of their target program,” Schwartzberg said. “We hope that others will extend our findings and that ITK inhibitors will be pursued as HIV therapies.”

Published online on April 28, 2008
Proc. Natl. Acad. Sci. USA, 10.1073/pnas.0709659105
OPEN ACCESS ARTICLE

T cell signaling | transcription | tyrosine kinase | viral entry | kinase inhibitors

Scientists in the Laboratoire Hépatite C of the Institut de Biologie de Lille in collaboration with INSERM Unit 602 and a laboratory at Stanford University have provided evidence of a protein, called EWI-2wint, that inhibits the hepatitis C virus at an early stage of its infective cycle. This research suggests possible new perspectives for the development of therapies to block the virus before it enters a cell.

The EWI-2wint protein is not present in hepatocytes (liver cells). When it comes into contact with the hepatocyte, the hepatitis C virus can thus bind to the CD81 protein, which will allow it to enter the cell and pursue its infective cycle. In other types of cells in the body, the EWI-2wint protein is present and interacts with CD81, thus preventing the hepatitis C virus from entering these cells. (Credit: Copyright CNRS 2008 Sophana Ung)

Hepatitis C is a major public health problem that affects some 130 million people throughout the world. In France , where there are about 5000 new cases each year, it is estimated that half a million people could be affected by this disease. The causal agent is the hepatitis C virus (HCV) which targets cells in the liver called hepatocytes. HCV infection is usually chronic (60% to 80% of cases) and in the long term can lead to the development of cirrhosis and liver cancer.

Unlike the hepatitis A and B viruses, there is no vaccine to combat this virus. Furthermore, the treatments employed are only of limited efficacy (the failure rate reaches around 40%), and they involve considerable side effects. It is therefore crucial to develop new antiviral compounds to control this infection.

HCV uses at least three receptors to enter and infect a hepatocyte. One of these receptors is the CD81 protein, which has the particular characteristic of associating with numerous other proteins. It was by studying these CD81 partner proteins that the researchers identified the EWI-2wint protein, which prevents the recognition of CD81 by the hepatitis C virus and inhibits it at a very early stage in its infective cycle. This protein is present in other types of cells, which could explain why they are not infected by HCV. Discovery of the role of EWI-2wint in hepatocytes has demonstrated the complexity of the mechanisms of entry of HCV into its target cells, and opens the way to new therapeutic approaches.

PLoS ONE. 2008 Apr 2;3(4):e1866.

The CD81 partner EWI-2wint inhibits hepatitis C virus entry.

Rocha-Perugini V, Montpellier C, Delgrange D, Wychowski C, Helle F, Pillez A, Drobecq H, Le Naour F, Charrin S, Levy S, Rubinstein E, Dubuisson J, Cocquerel L.

Institut de Biologie de Lille (UMR8161), CNRS, Universités de Lille I et Lille II, Institut Pasteur de Lille, Lille, France.

Two to three percent of the world’s population is chronically infected with hepatitis C virus (HCV) and thus at risk of developing liver cancer. Although precise mechanisms regulating HCV entry into hepatic cells are still unknown, several cell surface proteins have been identified as entry factors for this virus. Among these molecules, the tetraspanin CD81 is essential for HCV entry. Here, we have identified a partner of CD81, EWI-2wint, which is expressed in several cell lines but not in hepatocytes. Ectopic expression of EWI-2wint in a hepatoma cell line susceptible to HCV infection blocked viral entry by inhibiting the interaction between the HCV envelope glycoproteins and CD81. This finding suggests that, in addition to the presence of specific entry factors in the hepatocytes, the lack of a specific inhibitor can contribute to the hepatotropism of HCV. This is the first example of a pathogen gaining entry into host cells that lack a specific inhibitory factor.

Researchers studying deer mice have discovered evidence to support what mothers everywhere have long suspected: the immune system needs food to function properly. In a new study Lynn Martin and coauthors find that reduced food intake leads to a decline in immune function in their subjects. The findings could have profound implications for human health.

Immune System Overview

Why immune activity is variable in many wild animals is a question that has long puzzled researchers. “Animals live different lifestyles, so they may use different types of defenses against infection depending on the situation. Perhaps this is why immune defenses vary seasonally in most species; some may be too expensive to use all the time,” Martin said, referring to previous work on Peromyscus and other small mammals and birds.

While it is known that the immune system expends energy when it gears up to fight a virus or an infection–a fever, for example–the researchers found that restricting their subjects’ diet by 30% significantly decreased the amount of available B cells, which produce antibodies and maintain immune memory. Without these cells, the immune system must relearn how to fight a threat if it reappears.

Research on the relationship between food and the immune system could have profound implications for humans. Martin and fellow researchers cite previous studies that have found that infections are “more frequent and tend to be chronic in malnourished children.” Vaccines, in order to work effectively, must provoke B cells to produce sufficient antibodies for immune memory.

Previous studies have found that vaccines such as those for measles have a significantly lower rate of efficacy among the malnourished. “A 30% restriction in food intake doesn’t affect body mass and only minimally reduces activity in deer mice, but it eliminates the long-term immune protection provided by antibodies. One wonders whether similar moderate food restriction has comparable immune effects in humans,” Martin asked. Although other variables may be at work, the authors propose that for both wild animals and humans, food availability impinges on immunity and future research should determine what specific components of a diet (calories, protein, micronutrients) are responsible.

Physiological and Biochemical Zoology. Volume 81, Issue 3, Page 366–372, May 2008

Food Restriction Compromises Immune Memory in Deer Mice (Peromyscus maniculatus) by Reducing Spleen‐Derived Antibody‐Producing B Cell Numbers

Lynn B. Martin II, Kristen J. Navara, Michael T. Bailey, Chelsea R. Hutch, Nicole D. Powell, John F. Sheridan, and Randy J. Nelson

Abstract

Immune activity is variable in many wild animals, despite presumed strong selection against immune incompetence. Much variation may be due to changes in prevalence and abundance of pathogens (and/or their vectors) in time and space, but the costs of immune defenses themselves may also be important. Induction of immune activity often increases energy and protein expenditure, sometimes to the point of compromising fitness. Whether immune defenses are expensive to maintain once they are generated, however, is less well appreciated. If so, organisms would face persistent challenges of allocating resources between immunity and other expensive physiological processes, which would mandate trade-offs. Mild food restriction (70% ad lib. diet) reduces secondary antibody responses in deer mice (Peromyscus maniculatus), functionally representing a cost of immune memory. In this study, we asked whether compromised immune memory was mediated by a decrease in size of the B cell population responsible for producing antibodies (i.e., spleen-derived B lymphocytes producing immunoglobulin G [IgG]). Two weeks of food restriction reduced total splenocytes, total splenic B lymphocytes (B220+ cells), and splenic B lymphocytes producing IgG (B220+/IgG+ cells) but did not affect body mass or two circulating antibody subclasses (IgG1 and IgG2a) in deer mice. These results further indicate that maintenance of immune memory is expensive and may be subject to trade-offs with other physiological processes.

  HIV-positive women in the United States face strikingly high levels of stigma, according to survey results released today by amfAR, The Foundation for AIDS Research.

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The results of the survey reveal pervasive negative views of HIV-positive women and a high level of discomfort in interacting with them. Many of the responses display a lack of knowledge of how HIV is transmitted and misplaced fear of contracting the virus that indicate a pressing need to scale up prevention education efforts.

Sixty-eight percent of respondents indicated that they would be somewhat or not at all comfortable with an HIV-positive woman as their dentist; 59 percent said they would be somewhat or not at all comfortable with an HIV-positive woman serving as their childcare provider; and 57 percent said they would be somewhat or not at all comfortable having a female physician who is HIV-positive. One in five respondents would be somewhat or not at all comfortable having a close friend who is HIV positive.

Only 14 percent of respondents felt that HIV-positive women should have children. Currently medication exists to prevent mother-to-child transmission of HIV.

The survey also demonstrates significant differences in how Blacks, Hispanics and Caucasians perceive HIV/AIDS and the risk of acquiring it. Of those who know someone with HIV or AIDS, Blacks (34 percent) and Hispanics (32 percent) are much more likely to have a family member with HIV/AIDS than Caucasians (13 percent).

Insights were also gained into public attitudes about HIV testing. Nearly 40 percent were sure they had not been tested for HIV. A majority (80 percent) of these respondents indicated that they did not need a test either because they “knew” they did not have HIV or because they didn’t think they needed to be tested.

However, respondents overwhelmingly supported expanded HIV testing and 65 percent support making HIV testing part of standard routine healthcare. This acceptance may be partially linked to the belief that HIV testing occurs more frequently than it does, with 67 percent mistakenly assuming that they are automatically screened for HIV when they are tested for other sexually transmitted infections. Fifty percent believed that women are automatically tested during prenatal exams.

The survey draws much needed attention to the plight of women living with HIV/AIDS. Forty-six percent of people with HIV/AIDS worldwide – about 15.4 million – are women and girls. In the United States, women account for 27 percent of new AIDS diagnoses, up from only eight percent in 1985. Both domestically and internationally, women continue to face widespread social and gender inequalities that can make it difficult for them to reduce their risk of HIV infection. In addition, women are biologically more susceptible to HIV infection than men.

“In the minds of many people, AIDS in the United States is no longer a crisis,” said Rear Admiral Susan J. Blumenthal, M.D., M.P.A., amfAR’s senior policy and medical advisor and former deputy assistant secretary for women’s health in the U.S. Department of Health and Human Services. “Complacency has obscured the changing face of the epidemic and the dramatic rise in HIV infections in women over the past 25 years. These results should serve as a wake-up call for action across all sectors of society. We need to intensify efforts for science-based education and policy to shatter the stigma that has surrounded this disease for all too long.”

The online survey, conducted by Harris Interactive for amfAR, questioned nearly 5,000 respondents ages 18-44 and covered HIV risk and responsibility, impact of gender-based violence, and women’s access to healthcare and health information, as well as attitudes towards HIV-positive women. The survey was made possible by grants from Broadway Cares /Equity Fights AIDS and the M•A•C AIDS Fund.

The March 31 briefing will feature presentations by international AIDS activist Marvelyn Brown; Regan Hofmann, Editor of POZ Magazine; Helen-Maria Lekas, Ph.D. , Assistant Professor of Clinical Sociomedical Sciences at Columbia University’s Mailman School of Public Health; and Laura C. Nyblade, Ph.D. , Senior Social Scientist, HIV and Stigma, at the International Center for Research on Women. The briefing will be moderated by Dr. Blumenthal.

“Many women erroneously believe that they are not at risk for HIV,” said Hofmann, who is HIV positive. “This is why we are seeing the rate of new infections for women rise significantly in America. While many women accept that they could potentially become pregnant from even just one act of unprotected sex, they feel that they would have to do something ‘extraordinary’ to contract HIV, like be excessively promiscuous or be involved with people the likes of whom they don’t think they would encounter in their everyday world.”

Brown said, “I take seven pills daily that make me sick to my stomach. I experience nausea, diarrhea, vomiting and the worst of all mood swings. But yet it is still not the worst part of having HIV. It is the stigma.”

The amfAR survey results will be presented on March 31at the National Press Club in Washington, D.C. :

FOR IMMEDIATE RELEASE Mon 31-Mar-2008 CONTACT: Rear Admiral Susan Blumenthal, M.D., M.P.A., Senior Policy and Medical Advisor, amfAR - (202) 331-8600 / susan.blumenthal@amfar.org; Donald Kaplan, Director of Program Communications, amfAR - (212) 806-1602 / donald.kaplan@amfar.org Stigma Clings Stubbornly to Women Living With HIV/AIDS amfAR, The Foundation for AIDS Research, to release results of a national survey of public perceptions of women living with HIV/AIDS Washington, D.C. (March 31, 200 – HIV-positive women in the United States face strikingly high levels of stigma, according to survey results released today by amfAR, The Foundation for AIDS Research. The amfAR survey results will be presented at a press conference on Monday, March 31, at 9 a.m. at the National Press Club’s 13th Floor First Amendment Lounge, 529 14th St. N.W., in Washington, D.C. The press conference will be followed by a briefing in which panelists will offer personal and professional perspectives on HIV-related stigma among women. The results of the survey reveal pervasive negative views of HIV-positive women and a high level of discomfort in interacting with them. Many of the responses display a lack of knowledge of how HIV is transmitted and misplaced fear of contracting the virus that indicate a pressing need to scale up prevention education efforts. Sixty-eight percent of respondents indicated that they would be somewhat or not at all comfortable with an HIV-positive woman as their dentist; 59 percent said they would be somewhat or not at all comfortable with an HIV-positive woman serving as their childcare provider; and 57 percent said they would be somewhat or not at all comfortable having a female physician who is HIV-positive. One in five respondents would be somewhat or not at all comfortable having a close friend who is HIV positive. Only 14 percent of respondents felt that HIV-positive women should have children. Currently medication exists to prevent mother-to-child transmission of HIV. The survey also demonstrates significant differences in how Blacks, Hispanics and Caucasians perceive HIV/AIDS and the risk of acquiring it. Of those who know someone with HIV or AIDS, Blacks (34 percent) and Hispanics (32 percent) are much more likely to have a family member with HIV/AIDS than Caucasians (13 percent). Insights were also gained into public attitudes about HIV testing. Nearly 40 percent were sure they had not been tested for HIV. A majority (80 percent) of these respondents indicated that they did not need a test either because they “knew” they did not have HIV or because they didn’t think they needed to be tested. However, respondents overwhelmingly supported expanded HIV testing and 65 percent support making HIV testing part of standard routine healthcare. This acceptance may be partially linked to the belief that HIV testing occurs more frequently than it does, with 67 percent mistakenly assuming that they are automatically screened for HIV when they are tested for other sexually transmitted infections. Fifty percent believed that women are automatically tested during prenatal exams. The survey draws much needed attention to the plight of women living with HIV/AIDS. Forty-six percent of people with HIV/AIDS worldwide – about 15.4 million – are women and girls. In the United States, women account for 27 percent of new AIDS diagnoses, up from only eight percent in 1985. Both domestically and internationally, women continue to face widespread social and gender inequalities that can make it difficult for them to reduce their risk of HIV infection. In addition, women are biologically more susceptible to HIV infection than men. “In the minds of many people, AIDS in the United States is no longer a crisis,” said Rear Admiral Susan J. Blumenthal, M.D., M.P.A., amfAR’s senior policy and medical advisor and former deputy assistant secretary for women’s health in the U.S. Department of Health and Human Services. “Complacency has obscured the changing face of the epidemic and the dramatic rise in HIV infections in women over the past 25 years. These results should serve as a wake-up call for action across all sectors of society. We need to intensify efforts for science-based education and policy to shatter the stigma that has surrounded this disease for all too long.” The online survey, conducted by Harris Interactive for amfAR, questioned nearly 5,000 respondents ages 18-44 and covered HIV risk and responsibility, impact of gender-based violence, and women’s access to healthcare and health information, as well as attitudes towards HIV-positive women. The survey was made possible by grants from Broadway Cares /Equity Fights AIDS and the M•A•C AIDS Fund. The March 31 briefing will feature presentations by international AIDS activist Marvelyn Brown; Regan Hofmann, Editor of POZ Magazine; Helen-Maria Lekas, Ph.D. , Assistant Professor of Clinical Sociomedical Sciences at Columbia University’s Mailman School of Public Health; and Laura C. Nyblade, Ph.D. , Senior Social Scientist, HIV and Stigma, at the International Center for Research on Women. The briefing will be moderated by Dr. Blumenthal. “Many women erroneously believe that they are not at risk for HIV,” said Hofmann, who is HIV positive. “This is why we are seeing the rate of new infections for women rise significantly in America. While many women accept that they could potentially become pregnant from even just one act of unprotected sex, they feel that they would have to do something ‘extraordinary’ to contract HIV, like be excessively promiscuous or be involved with people the likes of whom they don’t think they would encounter in their everyday world.” Brown said, “I take seven pills daily that make me sick to my stomach. I experience nausea, diarrhea, vomiting and the worst of all mood swings. But yet it is still not the worst part of having HIV. It is the stigma.” About amfAR amfAR, The Foundation for AIDS Research, is one of the world’s leading nonprofit organizations dedicated to the support of AIDS research, HIV prevention, treatment education, and the advocacy of sound AIDS-related public policy. Since 1985, amfAR has invested nearly $275 million in its programs and has awarded grants to more than 2,000 research teams worldwide. For more information about amfAR, visit www.amfar.org. About Harris Interactive Harris Interactive is a global leader in custom market research. With a long and rich history in multimodal research that is powered by our science and technology, we assist clients in achieving business results. Harris Interactive serves clients globally through our North American, European and Asian offices and a network of independent market research firms. For more information, please visit www.harrisinteractive.com. Methodology This survey was conducted online within the United States by Harris Interactive on behalf of amfAR between March 22 and April 17, 2007, among 4831 people ages 18 to 44 who were willing to disclose their race. Sample included an oversample of Blacks and Hispanics. No estimates of theoretical sampling error can be calculated; a full methodology is available. Downloadable materials # Agenda (PDF) # Speaker biographies (PDF) # Dr. Blumenthal’s PowerPoint presentation (PDF) # Women and HIV/AIDS Fact Sheet (PDF) spacer spacer spacer Glossary spacer Women

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One Out Of Four HIV Patients Believe Their Doctors Stigmatize Them

Stigma

Impaired brain function is a prominent and still unsolved problem in AIDS. Shortly after an individual becomes infected with HIV, the virus can invade the brain and persist in this organ for life. Many HIV-infected individuals experience disturbances in memory functions and movement, which can progress to serious dementia. How the virus causes brain disease is still unclear.

HI-Virus leaving a cell. (Credit: NIH)

Dr. Ruth Brack-Werner and her team at the Institute of Virology of the German Research Center for Environmental Health previously demonstrated that HIV invades not only brain macrophages but also astrocytes. Astrocytes are the most abundant cells in the brain. They perform many important activities which support functions of nerve cells and protect them from harmful agents. HIV-infected astrocytes normally restrain the virus and prevent its production. However, various factors can cause astrocytes to lose control over the virus, allowing the virus to replicate and to reach the brain. There HIV can infect other brain cells as well as immune cells that patrol the brain and may carry the virus outside the brain.

Thus astrocytes form a reservoir for HIV in infected individuals and represent a serious obstacle to elimination of the virus from infected individuals. Whether this also applies to other types of brain cells was unclear until now. In a study recently published in AIDS, Dr. Brack Werner, together with Ina Rothenaigner and colleagues present data indicating that neural progenitor cells can also form HIV reservoirs in the brain. Neural progenitor cells are capable of developing into different types of brain cells and have an enormous potential for repair processes in the brain.

Dr. Brack-Werner’s team used a multi-potent neural progenitor cell line, which can be grown and developed to different types of brain cells in the laboratory, for their studies. After exposing these neural progenitor cells to HIV, they examined the cultures for signs of virus infection for 115 days. HIV was found to persist in these cultures during the entire observation period.

The cultures released infectious HIV particles for over 60 days and contained information for production of HIV regulatory proteins- Tat, Rev and Nef- for even longer. Dr. Brack-Werner and her team also examined neural progenitor cell populations cells with persisting HIV for differences from uninfected cells. They found that HIV persistence had an influence on the expression of selected genes and on cell morphology, but did not prevent their development to astrocytes. Thus HIV persistence has the potential to change neural progenitor cells.

Dr. Brack-Werner’s summarizes, “Our study indicates that neural progenitor cells are potential reservoirs for HIV and that HIV persistence has the potential to change the biology of these cells.” In future studies the researchers are planning to investigate the influence of HIV infection on important functions of neural progenitor cells. These include migration to diseased regions of the brain and development of different types of brain cells. Subsequently they will investigate how HIV changes neural progenitor cells and, importantly, how to protect neural progenitor cells from harmful effects of the virus in HIV infected individuals.

AIDS. 2007 Nov 12;21(17):2271-81.
Long-term HIV-1 infection of neural progenitor populations.
Rothenaigner I, Kramer S, Ziegler M, Wolff H, Kleinschmidt A, Brack-Werner R.

GSF–National Research Center for Environment and Health, Institute of Molecular Virology, Ingolstaedter Landstrasse 1, 85764 Neuherberg, Germany.

BACKGROUND: HIV can reside in the brain for many years. While astrocytes are known to tolerate long-term HIV infection, the potential of other neural cell types to harbour HIV is unclear. OBJECTIVE: To investigate whether HIV can persist in neural progenitor cell populations. DESIGN: A multipotent human neural stem cell line (HNSC.100) was used to compare HIV infection in neural progenitor and astrocyte cell populations. METHODS: Expression of cellular genes/proteins was analysed by real-time reverse transcriptase PCR, Western blot, immunocytochemistry and flow cytometry. Morphological properties of cells were measured by quantitative fluorescent image analysis. Virus release by cells exposed to HIV-1IIIB was monitored by enzyme-linked immunosorbent assay for Gag. Proviral copy numbers were determined by real-time PCR and early HIV transcripts by reverse transcriptase PCR. Rev activity was determined with a fluorescent-based reporter assay. RESULTS: Progenitor populations differed from astrocyte populations by showing much lower glial fibrillary acidic protein (GFAP) production, higher cell-surface expression of the CXCR4 chemokine receptor, higher Rev activity and distinct cell morphologies. HIV-exposed progenitor cultures released moderate amounts of virus for over 2 months and continued to display cell-associated HIV markers (proviral DNA, early HIV transcripts) during the entire observation period (115 days). Differentiation of HIV-infected progenitor cells to astrocytes was associated with transient activation of virus production. Long-term HIV infection of progenitor populations led to upregulation of GFAP and changes in cell morphology. CONCLUSION: These studies suggest that neural progenitor populations can contribute to the reservoir for HIV in the brain and undergo changes as a consequence of HIV persistence.

A Canada-U.S. research team has solved a major genetic mystery: How a protein in some people’s DNA guards them against killer immune diseases such as HIV. In an advance online edition of Nature Medicine, the scientists explain how the protein, FOX03a, shields against viral attacks and how the discovery will help in the development of a HIV vaccine.

Computer model of AIDS virus (HIV). (Credit: Produced by Richard Feldmann; courtesy of NIH/National Institute of Allergy and Infectious Diseases)

“HIV infection is characterised by the slow demise of T-cells, in particular central memory cells, which can mediate lifelong protection against viruses,” said lead researcher Rafick-Pierre Sékaly, a Université de Montréal professor and a researcher at the Centre Hospitalier de l’Université de Montréal and the French Institut national de la santé et de la recherche médicale (Inserm).

“Our group has found how the key protein, FOX03a, is vital to the survival of central memory cells that are defective in HIV-infected individuals even if they are treated,” added Dr. Sékaly, who produced his study with CHUM and Inserm colleagues including Elias El Haddad and Julien van Grevenynghe. Collaborators also included Jean-Pierre Routy, a McGill University Health Centre researcher and professor at McGill University and Robert S. Balderas, Vice-President of Research and Development at BD Biosciences of San Diego, CA.

Public support for the research came through Genome Canada and Génome Québec, among others, while private contributions came via a segment of BD (Becton, Dickinson and Company). “Public-private collaborations such as this play an important role in advancing medical research,” Robert S. Balderas. “BD Biosciences was pleased to provide powerful research instruments, reagents and technical expertise to support this breakthrough research.”

The breakthrough emerged by studying three groups of men: One HIV-negative sample, a second HIV-positive group whose infection was successfully controlled through tritherapy and a third group whose HIV did not show any symptoms. Called elite controllers, this third group fended off infection without treatment because their immune system, which would normally be attacked by HIV, maintained its resilient immune memory through the regulation of the FOX03a protein.

“Given their perfect resistance to HIV infection, elite controllers represent the ideal study group to examine how proteins are responsible for the maintenance of an immune system with good anti-viral memory,” said Dr. Haddad. “This is the first study to examine, in people rather than animals, what shields the body’s immune system from infection and to pinpoint the fundamental role of FOX03a in defending the body.”

Beyond HIV treatment, Dr. Sékaly said his team’s discovery offers promise for other immune diseases. “The discovery of FOX03a will enable scientists to develop appropriate therapies for other viral diseases that weaken the immune system,” he said, citing cancer, rheumatoid arthritis, hepatitis C, as well as organ or bone marrow transplant rejection.

Paul L’Archevêque, president and CEO of Génome Québec, lauded Dr. Sékaly’s team for their breakthrough and the people who volunteered for the study. “This discovery, the first such study in humans, is a major step forward in the understanding of how our immune system responds to life-threatening infections such as HIV. This advance stems directly from research co-financed by Génome Québec, which demonstrates the impact that genomic research can have in improving healthcare.”

This research was made possible by public and private institutions across Canada, the United States and France: the Université de Montréal, CHUM, Inserm, MUHC, Genome Canada, Génome Québec, Fonds de la recherche en santé du Québec, Canadian Institutes of Health Research, National Institutes of Health and BD Biosciences.

Nature Medicine Published online: 2 March 2008 | doi:10.1038/nm1728
Transcription factor FOXO3a controls the persistence of memory CD4⁺ T cells during HIV infection

Julien van Grevenynghe, Francesco A Procopio, Zhong He, Nicolas Chomont, Catherine Riou, Yuwei Zhang, Sylvain Gimmig, Genevieve Boucher, Peter Wilkinson, Yu Shi, Bader Yassine-Diab, Elias A Said, Lydie Trautmann, Mohamed El Far, Robert S Balderas, Mohamed-Rachid Boulassel, Jean-Pierre Routy, Elias K Haddad & Rafick-Pierre Sekaly

The persistence of central memory CD4⁺ T cells (T_CM cells) is a major correlate of immunological protection in HIV/AIDS, as the rate of T_CM cell decline predicts HIV disease progression. In this study, we show that T_CM cells and effector memory CD4⁺ T cells (T_EM cells) from HIV⁺ elite controller (EC) subjects are less susceptible to Fas-mediated apoptosis and persist longer after multiple rounds of T cell receptor triggering when compared to T_CM and T_EM cells from aviremic successfully treated (ST) subjects or from HIV^- donors. We show that persistence of T_CM cells from EC subjects is a direct consequence of inactivation of the FOXO3a pathway. Silencing the transcriptionally active form of FOXO3a by small interfering RNA or by introducing a FOXO3a dominant-negative form (FOXO3a Nt) extended the long-term survival of T_CM cells from ST subjects to a length of time similar to that of T_CM cells from EC subjects. The crucial role of FOXO3a in the survival of memory cells will help shed light on the underlying immunological mechanisms that control viral replication in EC subjects.