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Antibodies Prevent HIV Transmission in Mice
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Download the PDF here
Download the PDF here
This new study was just published......whether antibodies can be harnessed to prevent HIV transmission has been a debated & controversial issue but in light of this there have been several studies listed below including this latest in Nature medicine which comes along following their publication in 2011 Nature Medicine, pdf below.
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"Taken together, our results suggest that, by providing broadly neutralizing antibodies through VIP, it is possible to protect humanized mice against infection by strains of HIV similar to those that are responsible for human transmission......Despite these limitations, it seems reasonable to examine whether a sufficiently high circulating concentration of broadly neutralizing antibody might substantially reduce the probability of sexual transmission of HIV between humans."
Vectored immunoprophylaxis protects humanized mice from mucosal HIV transmission
Nature Medicine 09 February 2014
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http://jaxmice.jax.org/news/2012/HIV_VIP.html......the Balazs-Baltimore team developed a novel vectored immunoprophylactic (VIP) that protects mice from HIV infection. This VIP consists of a non-heparin-binding capsid and genes that encode naturally occurring, full-length, human HIV-neutralizing antibodies. When injected intramuscularly in mice, it confers long-lasting protection against even very high-dose HIV infections. The team believes that a similar VIP approach would confer broad, powerful HIV protection to humans. The approach may also be very effective in protecting against other infectious diseases and in therapies that require continuous production of monoclonal antibodies.
HIV Prevention: Nobel Laureate David Baltimore Talks VIP, New Developments (VIDEO)- http://www.huffingtonpost.com/2013/01/14/hiv-prevention-david-baltimore_n_2435387.html
Why is HIV so resistant to HIV....http://www.youtube.com/watch?v=77hSLuvIIjw
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New Studies, Nature Oct 2013 [HIV: Antibodies advance the search for a cureá Louis J. Picker & Steven G. Deeks] - HIV Antibodies Advance the search for a cure/Supercharged antibodies fight HIV-related virus in monkeys.....http://www.natap.org/2013/HIV/103113_01.htm
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Nature Medicine 15, 901 - 906 (2009)
Published online: 17 May 2009 | doi:10.1038/nm.1967
Vector-mediated gene transfer engenders long-lived neutralizing activity and protection against SIV infection in monkeys - pdf attached
Philip R Johnson1, Bruce C Schnepp1, Jianchao Zhang2, Mary J Connell1, Sean M Greene1, Eloisa Yuste3, Ronald C Desrosiers3 & K Reed Clark2
Abstract
The key to an effective HIV vaccine is development of an immunogen that elicits persisting antibodies with broad neutralizing activity against field strains of the virus. Unfortunately, very little progress has been made in finding or designing such immunogens. Using the simian immunodeficiency virus (SIV) model, we have taken a markedly different approach: delivery to muscle of an adeno-associated virus gene transfer vector expressing antibodies or antibody-like immunoadhesins having predetermined SIV specificity. With this approach, SIV-specific molecules are endogenously synthesized in myofibers and passively distributed to the circulatory system. Using such an approach in monkeys, we have now generated long-lasting neutralizing activity in serum and have observed complete protection against intravenous challenge with virulent SIV. In essence, this strategy bypasses the adaptive immune system and holds considerable promise as a unique approach to an effective HIV vaccine.
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Engineered AAV vector minimizes in vivo targeting of transduced hepatocytes by capsid-specific CD8+ T cells....
(2011, Nature Medicine) Antibody-based protection against HIV infection by vectored immunoprophylaxis - pdf attached
Alejandro B. Balazs1, Joyce Chen1, Christin M. Hong1, Dinesh S. Rao2, Lili Yang1 & David Baltimore1
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"Taken together, our results suggest that, by providing broadly neutralizing antibodies through VIP, it is possible to protect humanized mice against infection by strains of HIV similar to those that are responsible for human transmission......Despite these limitations, it seems reasonable to examine whether a sufficiently high circulating concentration of broadly neutralizing antibody might substantially reduce the probability of sexual transmission of HIV between humans."
Vectored immunoprophylaxis protects humanized mice from mucosal HIV transmission - pdf attached
Nature Medicine 09 February 2014
Alejandro B Balazs1,4, Yong Ouyang1, Christin M Hong1,4, Joyce Chen1, Steven M Nguyen1, Dinesh S Rao2,
Dong Sung An3 & David Baltimore1
1Division of Biology, California Institute of Technology, Pasadena, California, USA.
2Department of Pathology and Laboratory Medicine, David Geffen School of Medicine, University of California at Los Angeles (UCLA), Los Angeles, California, USA.
3Department of Translational Sciences, School of Nursing, UCLA AIDS Institute, UCLA, Los Angeles, California, USA.
4Present address: Ragon Institute of MGH, MIT and Harvard, Cambridge, Massachusetts, USA.
The vast majority of new HIV infections result from relatively inefficient transmission1, 2 of the virus across mucosal surfaces during sexual intercourse3. A consequence of this inefficiency is that small numbers of transmitted founder viruses initiate most heterosexual infections4. This natural bottleneck to transmission has stimulated efforts to develop interventions that are aimed at blocking this step of the infection process5. Despite the promise of this strategy, clinical trials of preexposure prophylaxis have had limited degrees of success in humans, in part because of lack of adherence to the recommended preexposure treatment regimens6, 7. In contrast, a number of existing vaccines elicit systemic immunity that protects against mucosal infections, such as the vaccines for influenza8 and human papilloma virus9. We recently demonstrated the ability of vectored immunoprophylaxis (VIP) to prevent intravenous transmission of HIV in humanized mice using broadly neutralizing antibodies10. Here we demonstrate that VIP is capable of protecting humanized mice from intravenous as well as vaginal challenge with diverse HIV strains despite repeated exposures. Moreover, animals receiving VIP that expresses a modified VRC07 antibody were completely resistant to repetitive intravaginal challenge by a heterosexually transmitted founder HIV strain11, suggesting that VIP may be effective in preventing vaginal transmission of HIV between humans.
The description of numerous broadly neutralizing HIV antibodies has invigorated strategies aimed at eliciting similar antibodies in naive patients12. In addition, it has been proposed that such antibodies could prevent the transmission of HIV if administered to patients by passive transfer. We and others have described the use of non-integrating adeno-associated virus (AAV) vectors to deliver the genes encoding such antibodies or immunoadhesins to muscle tissues, resulting in their long-term, systemic production. This has led to several demonstrations of the effectiveness of such a strategy to prevent intravenous transmission of simian immunodeficiency virus13 or laboratory strains of HIV10. However, it remains to be shown whether such an approach can be effective against HIV transmission between humans, which typically occurs across mucosal surfaces by HIV strains that are distinct from the laboratory strains currently used to test interventions.
To explore this issue, we determined the ability of VIP to prevent intravenous transmission of both JR-CSF, a CCR5-tropic primary isolate14, and REJO.c, a CCR5-tropic transmitted molecular founder strain11. We administered AAV vectors intramuscularly to establish groups of mice expressing high levels of human b12 IgG, VRC01 IgG or luciferase as a negative control (Fig. 1a). After administration and engraftment of human peripheral blood mononuclear cells (PBMCs), we challenged humanized mice intravenously with either JR-CSF or REJO.c (Fig. 1b). With either virus challenge we observed robust depletion of CD4+ T cells in animals expressing luciferase. Similarly, all b12-expressing mice challenged with REJO.c exhibited CD4+ T cell depletion, which is consistent with the previously observed resistance of this strain to b12 in vitro (Supplementary Fig. 1). Moreover, only three of the eight animals expressing b12 exhibited CD4+ T cell protection after JR-CSF challenge (Fig. 1b). To determine whether viral escape was responsible for the loss of CD4+ T cells in the remaining animals, we sequenced the viral envelope from mice exhibiting CD4+ T cell depletion and compared these sequences to the known wild-type sequence of JR-CSF (Fig. 1c).
Notably, envelope sequences obtained from mice expressing the b12 antibody exhibited many of the same common mutations that were present in luciferase-expressing control animals, but the sequences from the b12-expressing mice also contained additional unique mutations at JR-CSF Env residues V372 or M373 (numbered relative to the HXB2 reference strain), both of which have been implicated previously in escape from b12 neutralization15 (Fig. 1d). To determine whether these mutations were responsible for the in vivo escape of JR-CSF from b12, we engineered each individual mutation into otherwise wild-type JR-CSF and tested the sensitivity of the resulting viral stocks to either b12 or VRC01 in vitro (Fig. 1e). We found that either mutation enabled nearly complete resistance to b12, but neither mutation had an effect on VRC01 neutralization. This was true despite both antibodies targeting the CD4 binding site of the viral envelope and likely results from their distinct modes of CD4 binding site recognition16 (Supplementary Fig. 2). When we challenged humanized mice expressing VRC01 with JR-CSF or REJO.c, all animals except a single REJO.c-challenged mouse showed at least partial protection of CD4+ T cells (Fig. 1b). The single VRC01-expressing mouse that lost all CD4+ T cells exhibited no detectable viral load at any time point tested, and efforts to amplify envelope sequences from either plasma RNA or genomic DNA from this mouse were unsuccessful (data not shown). Consequently, we suspect that this mouse lost CD4+ T cells for reasons unrelated to HIV challenge. Together these results demonstrate that mice can be protected against CCR5-tropic HIV strains by VRC01 but that the b12 monoclonal antibody, which provided robust protection against the CXCR4-tropic NL4-3 strain10, is easily escaped by the CCR5-tropic JR-CSF strain.
Although these results show that VRC01 is capable of preventing the intravenous transmission of diverse strains, the predominant route of HIV infection worldwide is through heterosexual contact3. The bone marrow-liver-thymus (BLT) humanized mouse model, in contrast to nondiabetic severe combined immunodeficient γc (NOD-SCID-γc) humanized mice engrafted with PBMCs (huPBMC-NSG mice), exhibits extensive engraftment of human immune cells into mucosal tissues, which allows for HIV transmission to occur across mucosal surfaces17. To create a model of heterosexual transmission that better reflects the stochastic nature of human transmission, we modified the established high-dose, single vaginal challenge model in BLT humanized mice18 to implement a repetitive, non-abrasive, low-dose viral challenge similar to those used in non-human primates19. After administration of VIP encoding VRC01 to BLT mice, we observed production of human IgG specific for HIV gp120 at over 100 μg ml-1 in serum, whereas a luciferase-encoding vector produced no specific antibody (Fig. 2a). To determine the concentration of antibody reaching the challenge site, we analyzed vaginal wash fluid by ELISA and detected nearly 100 ng ml-1 of VRC01 4 weeks after AAV injection and 1 day before the first challenge (Fig. 2b). We believe this value to be a minimum estimate of the concentration because of the uncertainty of the dilution resulting from washing a small volume of vaginal mucus. We challenged mice weekly by intravaginal administration of JR-CSF and collected blood samples to monitor CD4+ T cell depletion and viral load. We observed limited but detectable depletion of CD4+ T cells in mice expressing luciferase but a steady or rising number of CD4+ T cells in mice expressing VRC01 (Fig. 2c). At the conclusion of the study, we subjected spleens from both groups to immunohistochemistry and observed a substantial number of p24-expressing cells in the spleens of luciferase-expressing control mice, which were largely absent from mice expressing VRC01 (Fig. 2d and Supplementary Fig. 3). Although we observed very limited depletion of CD4+ T cells in the spleen by FACS, we found a significant depletion of CD4+ T cells in the vaginal lamina propria of luciferase-expressing mice as compared to VRC01-expressing mice (Fig. 2e). Serum samples collected at terminal time points demonstrated that all luciferase-expressing mice were infected, whereas five of eight VRC01-expressing mice exhibited no detectable virus using an ultrasensitive clinical viral load assay (Fig. 2f). To determine the time course of infection, we subjected longitudinal serum samples to a viral load assay with reduced sensitivity and observed infection of control mice with a mean of 4.25 ± 1.32 (95% confidence interval (CI)) challenges (Fig. 2g). In contrast, this assay indicated that only two VRC01-expressing mice became infected over the duration of the experiment, and this infection occurred only after 13 or 15 exposures, indicating that VIP expressing VRC01 substantially reduced the risk of infection (Fig. 2h).
As the available repertoire of broadly neutralizing antibodies against HIV has expanded substantially since our original study20, 21, 22, we set out to determine the minimum protective dose of recently isolated antibodies to ascertain their in vivo potency. We gave mice decreasing doses of AAV vectors encoding each antibody or a luciferase-encoding AAV as a control to establish groups of animals expressing a range of antibody concentrations (Supplementary Fig. 4). After administration and engraftment of human PBMCs, we challenged mice intravenously with 10 ng p24 of NL4-3 and monitored them weekly for CD4+ T cell decline. We observed protection of humanized mice with a number of antibodies at concentrations as low as 350 ng ml-1 (Table 1 and Supplementary Fig. 5). Factoring together the activity we observed and the published breadth of each antibody, we selected the recently described VRC07 antibody containing a G54W mutation23, 24 for further study.
The bottleneck that occurs during mucosal transmission of HIV between humans appears to result in the selection of strains with unique properties that may enhance infectivity25. To determine the potential for VRC07 G54W to prevent mucosal transmission in BLT mice, we conducted a second repetitive challenge study with the REJO.c transmitted molecular founder strain of HIV11. VRC07 G54W was expressed at concentrations similar to those of VRC01, achieving a concentration of nearly 100 μg ml-1 in the serum within 4 weeks of intramuscular injection of the vector (Fig. 3a). Notably, we observed levels of VRC07 G54W in the vaginal wash fluid that approached 1 μg ml-1 at 4 weeks after AAV injection and 1 day before the first challenge (Fig. 3b). After the initiation of weekly intravaginal challenges with REJO.c, we observed that peripheral blood CD4+ T cells were relatively unperturbed in mice expressing luciferase but showed a gradual rise in mice expressing VRC07 G54W (Fig. 3c). At the conclusion of the experiment, we analyzed vaginal tissues by immunohistochemistry and observed vaginal lamina propria lymphocytes displaying HIV p24 antigen only in mice expressing luciferase, suggesting that a local infection was taking place at the site of challenge (Fig. 3d). FACS analysis of splenic tissue demonstrated a modest but statistically significant reduction in the level of CD4+ T cells (Fig. 3e). We observed significant differences among CD4+ T cell populations within gut intraepithelial and lamina propria lymphocytes, as well as in vaginal lamina propria lymphocytes, suggesting that VRC07 G54W was able to protect CD4+ T cells in mucosal tissues, which are typically depleted during the initial phase of HIV infection in patients26 (Fig. 3e and Supplementary Fig. 6). In agreement with this observation, the ultrasensitive clinical viral load assay detected infection in nearly all luciferase-expressing control animals, whereas none of the mice producing VRC07 G54W exhibited detectable virus in plasma despite 21 consecutive weekly challenges with REJO.c (Fig. 3f). To determine the number of vaginal exposures necessary for infection in this model, we quantified the viral load in longitudinal plasma samples using the less sensitive method described above (Fig. 3g). Luciferase-expressing control mice became infected by REJO.c with a mean of 7.45 ± 3.31 (95% CI) challenges. However, in one mouse, just two challenges were sufficient to initiate an infection, whereas in another animal, infection required 18 challenges. The two control animals that failed to become infected exhibited declining health during the course of the experiment and were euthanized after 8 or 14 challenges. Remarkably, none of the mice expressing VRC07 G54W exhibited a sustained viral load above 1,000 copies ml-1 throughout the course of the experiment despite 21 consecutive exposures (Fig. 3h).
Taken together, our results suggest that, by providing broadly neutralizing antibodies through VIP, it is possible to protect humanized mice against infection by strains of HIV similar to those that are responsible for human transmission. Notably, we detected viral escape from antibody neutralization despite our use of virus produced from the transfection of molecular clones that would not be expected to result in a diverse virus stock containing many preexisting mutations. We hypothesize that limited viral replication may occur in vivo-perhaps locally in the mucosa-despite the presence of neutralizing antibodies, which might allow for selection of resistant strains. Whereas we observed rapid CD4+ T cell decline in huPBMC-NSG control mice challenged with HIV, the kinetics of CD4+ T cell depletion after infection appeared to be substantially slower in BLT control mice. We hypothesize that this difference may be a result of lower levels of xenogenic activation of CD4+ T cells that develop in the BLT model and the regeneration of T cells from engrafted stem cells. Despite the IgG1 isotype expressed by VIP, we found that these antibodies reached the vaginal mucosa. It is unclear whether this mucosal antibody alone was sufficient to prevent transmission or whether protection also required the high concentrations of circulating antibody that were present in our mice. Our results demonstrate that repetitive challenge with a transmitted founder strain can be used to mimic the inefficient nature of vaginal HIV transmission in humans and highlight the utility of such a model as a relatively low-cost approach toward testing new prophylactic interventions. However, it is important to note that in addition to the substantial anatomical differences between mice and humans, the existing BLT humanized mouse model does not entirely recapitulate a functional human immune system. Despite these limitations, it seems reasonable to examine whether a sufficiently high circulating concentration of broadly neutralizing antibody might substantially reduce the probability of sexual transmission of HIV between humans.
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