Laboratory of Viral Diseases

Bethesda, MD, United States

Laboratory of Viral Diseases

Bethesda, MD, United States

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News Article | November 10, 2016

Silver Spring, Md. - Researchers have found that an investigational treatment combining a therapeutic vaccine and an immune stimulator improves virologic control and delays viral rebound following the discontinuation of antiretroviral therapy (ART) in non-human primates infected with SIV, the simian form of HIV. The proof-of-concept study examined the combined effects of therapeutic vaccination with an adenovirus serotype 26 vector vaccine and an MVA vector vaccine (Ad26/MVA) and TLR-7 agonist stimulation in ART-suppressed, SIV-infected monkeys. Findings were published online today in Nature. The study was a collaboration led by the Beth Israel Deaconess Medical Center (BIDMC) and the U.S. Military HIV Research Program (MHRP) of the Walter Reed Army Institute of Research (WRAIR), and includes scientists from Janssen Vaccines & Prevention B.V., one of the Janssen Pharmaceutical Companies of Johnson & Johnson, and Gilead Sciences, Inc. All rhesus monkeys were started on suppressive ART seven days after infection with SIV. After 24 weeks, groups of animals then either received a placebo treatment, Ad26/MVA, TLR7 agonist or a combination intervention of Ad26/MVA and TLR-7. TLR7 agonist. At 72 weeks, ART was discontinued to test the ability of the investigational therapies to affect continued virological control. "We found the combination of Ad26/MVA vaccination and TLR7 stimulation proved more effective than either component alone," said Col. Nelson Michael, Director of MHRP, who helped design the preclinical study. "This was especially striking for viral load set-point, which impacts future disease." In the combination group, the mean viral load set-point was reduced by 100 fold in all animals. Researchers saw a 2.5-fold delay of viral rebound as compared with the other groups. TLR-7Stimulation of TLR7 alone did not impact viral load or rebound. The vaccine alone reduced viral load set-point by 10 fold and only marginally delayed rebound. Though all monkeys eventually experienced viral rebound following ART interruption, three of the monkeys in the combination intervention group showed effective virologic control to undetectable viral loads following ART discontinuation. "Current antiretroviral drugs, although they're lifesaving, do not cure HIV. They merely hold it in check. We are trying to develop strategies to achieve ART-free, long-term viral suppression," said senior author Dan Barouch, MD, PhD, Director of the Center for Virology and Vaccine Research at BIDMC and Professor of Medicine at Harvard Medical School. "We reasoned that if we can activate the immune cells that might harbor the virus, then the vaccine-induced immune responses might perform better seeking them out and destroying them." A critical barrier to HIV cure is the viral reservoir that remains hidden and infects cells throughout the body, leading to viral rebound in the vast majority of HIV-infected individuals after they discontinue ART. According to Dr. Merlin Robb, Deputy Director for Clinical Research at MHRP, "the combination of Ad26/MVA vaccination and TLR7 stimulation resulted in decreased levels of viral DNA in both lymph nodes and peripheral blood. With further optimization this combination strategy may show promise to achieve a functional cure for HIV." Additionally, cellular immune breadth correlated inversely with set-point viral loads and correlated directly with time to viral rebound. According to Michael, "This gives us an immunologic correlate which can potentially be used to predict responses in humans, but this needs to be confirmed in human clinical studies." Ad26/MVA is a prime boost vaccine regimen. MHRP, in collaboration with Janssen, recently began evaluating this regimen as a therapeutic vaccine in HIV infected adults who initiated ART during acute HIV infection. That study is being conducted at the Thai Red Cross in Bangkok, and the protocol chair is Dr. Jintanat Ananworanich, MHRP's Associate Director for Therapeutics. The Ad26 vaccine was developed in partnership between the National Institute of Allergy and Infectious Diseases (NIAID), part of the National Institutes of Health (NIH), BIDMC and Janssen. MHRP developed the MVA vaccine in collaboration with the Laboratory of Viral Diseases at NIAID/NIH. The TLR7 agonist (GS-986) was developed by Gilead. Funding for the study was provided by the U.S. Army Medical Research and Materiel Command and the Military HIV Research Program, Walter Reed Army Institute of Research through its cooperative agreement with the Henry M. Jackson Foundation (W81XWH-11-2-0174); NIH (AI096040, AI124377, AI126603); the Ragon Institute of MGH, MIT, and Harvard. About the Walter Reed Army Institute of Research Headquartered in Silver Spring, Maryland, the Walter Reed Army Institute of Research (WRAIR) is the oldest and most diverse biomedical research laboratory in the Department of Defense. WRAIR provides unique research capabilities and innovative solutions to a range of force health and readiness challenges currently facing U.S. Service Members, along with threats anticipated during future operations. With comprehensive research units in Africa, Asia, and the Caucasus region, WRAIR is comprised of two Centers of Excellence, the Center for Infectious Disease Research and the Center for Military Psychiatry and Neuroscience.

Dey B.,Laboratory of Viral Diseases | Lagenaur L.A.,U.S. National Institutes of Health | Lagenaur L.A.,Osel, Inc. | Lusso P.,National Institute of Allergy and Infectious Diseases
Current HIV Research | Year: 2013

Although the development of a protective vaccine remains the most effective strategy for the global control of HIV/AIDS, another practical form of medical intervention would be a microbicide capable of preventing HIV-1 transmission at the mucosal level. A broad spectrum of antiviral molecules have demonstrated in vitro efficacy in proof-of-principle studies, and a selected few have already been tested in pre-clinical and clinical microbicide trials. Nevertheless, major hurdles remain to be overcome and there is still much uncertainty about the choice of inhibitors, formulations and administration vehicles for obtaining a safe and effective microbicide. A special category of HIV-1 microbicides are those based on proteins or peptides that interfere with the earliest steps in the viral infectious cycle. Besides a high degree of target specificity and a limited, if any, systemic absorption, protein-based microbicides offer the unique advantage of being suitable to in vivo expression by engineered bacteria or viral vectors, which might ensure prolonged protection without the need for planned, intercourse-coordinated application. In this respect, vaginal or rectal microbiota such as Lactobacillus spp. represent ideal expression systems as they would not only produce the inhibitor of choice at the mucosal surface, but also easily blend within the resident microflora and offer additional valuable homeostatic effects. In this article, we review the current state of the art on protein-based microbicides. © 2013 Bentham Science Publishers.

News Article | April 1, 2016

A team led by Purdue University researchers is the first to determine the structure of the Zika virus, which reveals insights critical to the development of effective antiviral treatments and vaccines. The team also identified regions within the Zika virus structure where it differs from other flaviviruses, the family of viruses to which Zika belongs that includes dengue, West Nile, yellow fever, Japanese encephalitis and tick-borne encephalitic viruses. A paper detailing the findings was published Thursday, March 31, in the journal Science and is available online. Any regions within the virus structure unique to Zika have the potential to explain differences in how a virus is transmitted and how it manifests as a disease, said Richard Kuhn, director of the Purdue Institute for Inflammation, Immunology and Infectious Diseases (PI4D) who led the research team with Michael Rossmann, Purdue's Hanley Distinguished Professor of Biological Sciences. "The structure of the virus provides a map that shows potential regions of the virus that could be targeted by a therapeutic treatment, used to create an effective vaccine or to improve our ability to diagnose and distinguish Zika infection from that of other related viruses," said Kuhn, who also is head of Purdue's Department of Biological Sciences. "Determining the structure greatly advances our understanding of Zika - a virus about which little is known. It illuminates the most promising areas for further testing and research to combat infection." The Zika virus, a mosquito-borne disease, has recently been associated with a birth defect called microcephaly that causes brain damage and an abnormally small head in babies born to mothers infected during pregnancy. It also has been associated with the autoimmune disease Guillain-Barré syndrome, which can lead to temporary paralysis. In the majority of infected individuals symptoms are mild and include fever, skin rashes and flulike illness, according to the World Health Organization. Zika virus transmission has been reported in 33 countries. Of the countries where Zika virus is circulating 12 have reported an increased incidence of Guillain-Barré syndrome, and Brazil and French Polynesia have reported an increase in microcephaly, according to WHO. In February WHO declared the Zika virus to be "a public health emergency of international concern." "This breakthrough illustrates not only the importance of basic research to the betterment of human health, but also its nimbleness in quickly addressing a pressing global concern," said Purdue President Mitch Daniels. "This talented team of researchers solved a very difficult puzzle in a remarkably short period of time, and have provided those working on developing vaccines and treatments to stop this virus a map to guide their way." Rossmann and Kuhn collaborated with Theodore Pierson, chief of the viral pathogenesis section of the Laboratory of Viral Diseases at the National Institutes of Health National Institute of Allergy and Infectious Diseases. Additional research team members include Purdue graduate student Devika Sirohi and postdoctoral research associates Zhenguo Chen, Lei Sun and Thomas Klose. The team's paper marks the first published success of the new Purdue Institute for Inflammation, Immunology and Infectious Diseases in Purdue's Discovery Park. The university's recently announced $250 million investment in the life sciences funded the purchase of advanced equipment that allowed the team to do in a couple of months what otherwise would have taken years, Rossmann said. "We were able to determine through cryo-electron microscopy the virus structure at a resolution that previously would only have been possible through X-ray crystallography," he said. "Since the 1950s X-ray crystallography has been the standard method for determining the structure of viruses, but it requires a relatively large amount of virus, which isn't always available; it can be very difficult to do, especially for viruses like Zika that have a lipid membrane and don't organize accurately in a crystal; and it takes a long time. Now we can do it through electron microscopy and view the virus in a more native state. This was unthinkable only a few years ago." The team studied a strain of Zika virus isolated from a patient infected during the French Polynesia epidemic and determined the structure to 3.8Å. At this near-atomic resolution key features of the virus structure can be seen and groups of atoms that form specific chemical entities, such as those that represent one of 20 naturally occurring amino acids, can be recognized, Rossmann said. The team found the structure to be very similar to that of other flaviviruses with an RNA genome surrounded by a lipid, or fatty, membrane inside an icosahedral protein shell. The strong similarity with other flaviviruses was not surprising and is perhaps reassuring in terms of vaccine development already underway, but the subtle structural differences are possibly key, Sirohi said. "Most viruses don't invade the nervous system or the developing fetus due to blood-brain and placental barriers, but the association with improper brain development in fetuses suggest Zika does," Sirohi said. "It is not clear how Zika gains access to these cells and infects them, but these areas of structural difference may be involved. These unique areas may be crucial and warrant further investigation." The team found that all of the known flavivirus structures differ in the amino acids that surround a glycosylation site in the virus shell. The shell is made up of 180 copies of two different proteins. These, like all proteins, are long chains of amino acids folded into particular structures to create a protein molecule, Rossmann said. The glycosylation site where Zika virus differs from other flaviviruses protrudes from the surface of the virus. A carbohydrate molecule consisting of various sugars is attached to the viral protein surface at this site. In many other viruses it has been shown that as the virus projects a glycosylation site outward, an attachment receptor molecule on the surface of a human cell recognizes the sugars and binds to them, Kuhn said. The virus is like a menacing stranger luring an unsuspecting victim with the offer of sweet candy. The human cell gladly reaches out for the treat and then is caught by the virus, which, once attached, may initiate infection of that cell. The glycosylation site and surrounding residues on Zika virus may also be involved in attachment to human cells, and the differences in the amino acids between different flaviviruses could signify differences in the kinds of molecules to which the virus can attach and the different human cells it can infect, Rossmann said. "If this site functions as it does in dengue and is involved in attachment to human cells, it could be a good spot to target an antiviral compound," Rossmann said. "If this is the case, perhaps an inhibitor could be designed to block this function and keep the virus from attaching to and infecting human cells." The team plans to pursue further testing to evaluate the different regions as targets for treatment and to develop potential therapeutic molecules, Kuhn said. Kuhn and Rossmann have studied flaviviruses, the family of viruses to which Zika belongs, for more than 14 years. They were the first to map the structure of any flavivirus when they determined the dengue virus structure in 2002. In 2003 they were first to determine the structure of West Nile virus and now they are the first to do so with the Zika virus.

Karttunen H.,New York University | Savas J.N.,Scripps Research Institute | McKinney C.,New York University | McKinney C.,Laboratory of Viral Diseases | And 5 more authors.
Molecular Cell | Year: 2014

DNA damage associated with viral DNA synthesis can result in double-strand breaks that threaten genome integrity and must be repaired. Here, we establish that the cellular Fanconi anemia (FA) genomic stability pathway is exploited by herpes simplex virus 1 (HSV-1) to promote viral DNA synthesis and enable its productive growth. Potent FA pathway activation in HSV-1-infected cells resulted in monoubiquitination of FA effector proteins FANCI and FANCD2 (FANCI-D2) and required the viral DNA polymerase. FANCD2 relocalized to viral replication compartments, and FANCI-D2 interacted with a multisubunit complex containing the virus-encoded single-stranded DNA-binding protein ICP8. Significantly, whereas HSV-1 productive growth was impaired in monoubiquitination-defective FA cells, this restriction was partially surmounted by antagonizing the DNA-dependent protein kinase (DNA-PK), a critical enzyme required for nonhomologous end-joining (NHEJ). This identifies the FA-pathway as a cellular factor required for herpesvirus productive growth and suggests that FA-mediated suppression of NHEJ is a fundamental step in the viral life cycle. © 2014 Elsevier Inc.

Das S.R.,Laboratory of Viral Diseases | Puigbo P.,U.S. National Center for Biotechnology Information | Hensley S.E.,Laboratory of Viral Diseases | Hurt D.E.,Computational Biology Bioinformatics and Computational Biosciences Branch BCBB | And 2 more authors.
PLoS Pathogens | Year: 2010

Antigenic drift in the influenza A virus hemagglutinin (HA) is responsible for seasonal reformulation of influenza vaccines. Here, we address an important and largely overlooked issue in antigenic drift: how does the number and location of glycosylation sites affect HA evolution in man? We analyzed the glycosylation status of all full-length H1 subtype HA sequences available in the NCBI influenza database. We devised the "flow index" (FI), a simple algorithm that calculates the tendency for viruses to gain or lose consensus glycosylation sites. The FI predicts the predominance of glycosylation states among existing strains. Our analyses show that while the number of glycosylation sites in the HA globular domain does not influence the overall magnitude of variation in defined antigenic regions, variation focuses on those regions unshielded by glycosylation. This supports the conclusion that glycosylation generally shields HA from antibody-mediated neutralization, and implies that fitness costs in accommodating oligosaccharides limit virus escape via HA hyperglycosylation.

Yewdell J.W.,Laboratory of Viral Diseases
Molecular Immunology | Year: 2013

The field of antigen processing and presentation has taken tremendous strides since the first international workshop in 1995. While much has been learned, much remains to be discovered. Here I discuss the most recent findings regarding the nature of substrates for the MHC class I antigen processing pathways which provide glimpses of the mist shrouded features remaining to be discovered. © 2012.

Yewdell J.W.,Laboratory of Viral Diseases
Current Opinion in Immunology | Year: 2010

CD8+ T cells play important roles in clearing viral infections and eradicating tumors. Designing vaccines that elicit effective CD8+ T cell responses requires a thorough knowledge of the pathways of antigen presentation in vivo. Here, I review recent progress in understanding the activation of naïve CD8+ T cells in vivo, with particular emphasis on cross-priming, the presentation of protein antigens acquired by dendritic cells from their environment. With the rapid advances in this area of research, the dawn of rational vaccine design is at hand. © 2010.

Dolan B.P.,Laboratory of Viral Diseases | Bennink J.R.,Laboratory of Viral Diseases | Yewdell J.W.,Laboratory of Viral Diseases
Cellular and Molecular Life Sciences | Year: 2011

It has been 15 years since we proposed the defective ribosomal product (DRiP) hypothesis to explain the rapid presentation of viral peptides by MHC class I molecules on the surface of infected cells. Here, we review the evidence for the contribution of DRiPs to antigen processing, pointing to the uncertainties regarding the physical nature of DRiPs, and emphasizing recent findings suggesting that peptide generation is a specialized process involving compartmentalized translation. © Springer Basel (outside the USA) 2011.

Yewdell J.W.,Laboratory of Viral Diseases | Ince W.L.,Laboratory of Viral Diseases
Cell Host and Microbe | Year: 2013

In this issue, Nakamura et al. (2013) describe a robust SCID mouse-based method for isolating human monoclonal antibodies of desired specificity from adoptively transferred human B cells. As proof of principle, they isolate human mAbs that could potentially be used to treat or prevent human infection with any influenza A virus strain. © 2013 Elsevier Inc.

Yewdell J.W.,Laboratory of Viral Diseases
Trends in Immunology | Year: 2011

Defective ribosomal products (DRiPs) are a subset of rapidly degraded polypeptides that provide peptide ligands for major histocompatibility complex (MHC) class I molecules. Here, recent progress in understanding DRiP biogenesis is reviewed. These findings place DRiPs at the center of the MHC class I antigen processing pathway, linking immunosurveillance of viruses and tumors to mechanisms of specialized translation and cellular compartmentalization. DRiPs enable the immune system to rapidly detect alterations in cellular gene expression with great sensitivity. © 2011.

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