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Stamatatos Progress Report Abstracts continued

 

Submitted January 25, 2011 (Interim Report)

Our Consortium aims to develop immunogens that would elicit broad and potent anti-HIV-1 neutralizing antibody responses. We are employing two distinct, but complementary, approaches: (a) We engineer immunogens derived from the natural target of anti-HIV NAbs, the HIV Envelope glycoprotein (Env), which contains multiple neutralization epitopes; and (b) we computationally design small non-HIV proteins that express specific, known, HIV-1 neutralization epitopes.

Regarding the first approach: We established methodologies to express and purify trimeric heterotrimeric gp140 proteins composed of clade A and B protomers and performed rabbit immunizations. Our data indicate that these heterotrimeric proteins are eliciting broader neutralizing antibody responses than the corresponding homotrimeric gp140 proteins.

Regarding the second approach: We recently reported that our computationally designed immunogens (CDI) elicit antibody responses to one of the most conserved, but poorly immunogenic neutralization epitopes on HIV-1, the 4E10 epitope (recognized by the broadly neutralizing MAb 4E10).  The antibodies elicited by our CDI fail to neutralize HIV-1 despite the fact that they recognize the 4E10 epitope in a manner which is very similar to that of MAb 4E10. Our findings suggest that neutralization of HIV through the 4E10 epitope depends not only on the proper antibody recognition of the epitope but has additional requirements.

 
Submitted September 1, 2010

Our Consortium is working on developing a vaccine that would elicit neutralizing antibodies against HIV. During this past year, we generated novel information on the structure of the viral envelope glycoprotein, the sole target of anti-HIV neutralizing antibodies, by crystallizing and solving the first structure of a clade C envelope glycoprotein. HIV clade C infections predominate worldwide. This new structural information is relevant to immunogen-design efforts. So far, only the structures of clade B-derived HIV-1 envelopes were available. Our group recently completed an extensive analysis of the immunogenic properties of computationally-designed scaffolds (small non-HIV proteins that are designed to express a single known conserved HIV neutralization epitope). Our major focus for the past three years has been on the ‘4e10 epitope’, which is a highly conserved HIV neutralization epitope and is the target of the broadly neutralizing MAb 4E10. The 4e10 epitope adopts a helical conformation and is partially embedded in the viral lipid membrane. Our recent studies indicate that 4e10-like antibodies are routinely elicited by our scaffolds and that these antibodies have a similar helical binding dependency as MAb 4E10. However, despite similar binding properties with MAb 4E10, these 4e10-like antibodies do not neutralize HIV. We are currently examining the reason(s) for this lack of neutralizing activity. We also made progress in engineering novel soluble HIV envelope proteins composed of clade B and A envelope protomers. These constructs (termed as ‘heterotrimers’) elicit more potent neutralizing antibody responses that the corresponding ‘homotrimers’ (which are commonly used in the HIV vaccine field). We are now following on this promising result and are optimizing our heterotrimeric immunogens for further preclinical evaluation. 

 
Submitted January 28, 2010 (Interim Report)

Our Consortium aims to develop immunogens that would elicit broad and potent anti-HIV-1 neutralizing antibody responses. We are employing two distinct, but complementary, approaches: (a) We engineer immunogens derived from the natural target of anti-HIV NAbs, the HIV Envelope glycoprotein (Env), which contains multiple neutralization epitopes; and (b) we computationally design small non-HIV proteins that express specific, known, HIV-1 neutralization epitopes.

Regarding the first approach: We developed methodologies that allow us perform rapid comparative immunogenicity analysis of diverse Envs, to identify the most promising constructs, and we are evaluating novel adjuvant formulations to identify those that optimize Env immunogenicity.

Regarding the second approach: We recently reported that our computationally designed immunogens (CDI) elicit antibody responses to one of the most conserved, but poorly immunogenic neutralization epitopes on HIV-1, the 4E10 epitope (recognized by the broadly neutralizing MAb 4E10). The antibodies elicited by our CDI fail to neutralize HIV-1 despite the fact that they recognize the 4E10 epitope in a manner which is very similar to that of MAb 4E10. Our findings suggest that neutralization of HIV through the 4E10 epitope depends not only on the proper antibody recognition of the epitope but has additional requirements.

Despite efforts by different groups over many years, the structure of the functional, trimeric, HIV Env (the target of NAbs) remains unsolved. Only structures of the extracellular domain of the HIV-1 Env (the gp120 subunit) derived from clade B isolates have been reported. This seriously limits our ‘structure-based’ immunogen design efforts. Overall we aim at solving the structure of the trimeric form of Env, but we are interested in solving the structure of Env (trimeric or monomeric) derived from non-clade B isolates, since such new information will reveal similarities and differences in the structures of Env from diverse clades . Recently, our group solved the crystal structure of a truncated clade C Env. Clade C infections are predominate world wide.

 
Submitted September 15, 2009

Our Consortium is working on developing a vaccine that would elicit antibodies capable of preventing infection by HIV. Such antibodies are called neutralizing antibodies (NAbs). Our group takes advantage of the latest advances made in various areas of research, including immunology, virology, molecular biology and computational-based design methodologies, to develop novel strategies to design an anti-HIV vaccine. Investigators with diverse but complementary backgrounds participate in our Consortium. Our Consortium has made significant progress on multiple fronts and here we highlight two of them: (a) We have recently obtained and analyzed the first crystal structure of a clade C viral envelope glycoprotein. HIV clade C infections are predominant worldwide and this new information may be important in understanding the prevalence of clade C HIV viruses and potentially allow for more effective interventions, including vaccines. (b) Using computational approaches our group designed candidate vaccines to elicit NAbs against a conserved domain of HIV. We recently obtained very promising results that our computationally-designed vaccines are capable of eliciting antibodies with the appropriate binding specificities. We are improving our design approaches and our immunization methodologies to enhance the neutralizing potential of the elicited antibodies.

 
Submitted February 1, 2009 (Interim Report)

Our Consortium is working on developing a vaccine that would elicit antibodies capable of preventing infection by HIV. Such antibodies are called, neutralizing antibodies (NAbs). Our group takes advantage of the latest advances made in various areas of research, including immunology, virology, molecular biology and computational-based design methodologies, to develop novel strategies to design an anti-HIV vaccine. Investigators with diverse but complementary background participate in our Consortium.

We made significant progress in the past 6 months. We optimized our vaccine-production capabilities and we are now able to express numerous, structurally-complex molecules that are suitable for immunization testing. Tens of vaccine candidates were tested in animals. We are in the process of optimizing selected immunogens and immunization protocols.

We completed a study during which we investigated the prevalence and epitope-specificities of broad neutralizing antibodies during natural HIV-1-infection. Our results indicate that ~20% of anti-retroviral naïve HIV+ patients develop cross-reactive neutralizing antibodies. Analysis of the epitope-specificity of these antibodies, revealed the location of some of these epitopes. These studies can guide our vaccine-design efforts.

 
Submitted September 2, 2008

Our Consortium is working on developing a vaccine that would elicit antibodies capable of preventing infection by HIV. Such antibodies are called, neutralizing antibodies (NAbs). Our group takes advantage of the latest advances made in various areas of research, including immunology, virology, molecular biology and computational-based design methodologies, to develop novel strategies to design an anti-HIV vaccine. Investigators with diverse but complementary background participate in our Consortium. Some of our colleagues have not been involved in HIV research in the past, but their expertise allows us to attack this public health problem from new angles. During the first two years we developed the methodologies and techniques that are necessary for the accomplishment of our goals, and we initiated many experiments and made strong progress in all our objectives. We are continuously optimizing our vaccine candidates, based on the results we obtained and analyzed during the first two years, and we are evaluating them for the elicitation of cross-reactive NAbs according to our initial plan.

 
Submitted February 1, 2008 (Interim Report)

Our Consortium is working on developing a vaccine that would elicit antibodies capable of preventing infection by HIV. Such antibodies are called, neutralizing antibodies (NAbs). Our group takes advantage of the latest advances made in various areas of research, including immunology, virology, molecular biology and computational-based design methodologies, to develop novel strategies to design an anti-HIV vaccine. Investigators with diverse but complementary background participate in our Consortium.

In the past 6 months we engineered several novel vaccine candidates, some of which were solely designed using computers. Several of these novel vaccines have been tested in animals. Although, so far, we have not identified a vaccine candidate that reproducibly elicits the desired neutralizing antibody response to HIV, the results we obtained from these pilot immunizations (performed in rabbits and guinea pigs) have provided us with valuable information which we are currently utilizing to optimize the design of our vaccine candidates. This iterative approach is essential in our efforts to define antigenic and biophysical properties of candidate HIV vaccines that correlate with their ability to elicit protective antibody responses.

In the past six months we optimized the HIV Env expression and purification protocols and we are now routinely producing mg quantities of soluble trimeric HIV Env gp140 from clade A and B viruses. These proteins are used for both crystallization and immunization studies. We have recently developed a protocol that allows us to produce and purify stable clade B/A hetero-oligomeric Envs. These constructs will be characterized antigenically and then tested as vaccine candidates.

Finally, in the past 6 months we continued our efforts to isolate cross-neutralizing anti-HIV antibodies for rhesus macaques that were experimentally infected with the R5-tropic SHIVSF162P4. We have isolated several antibodies that although they display limited cross-neutralizing activities, they recognize unique epitopes on HIV. Specifically, many of these antibodies do not bind to the recombinant Env (trimeric gp140 or monomeric gp120) but bind to virion-associated Env. These antibodies will be used as tools to design Env immunogens with an overall structure most closely related to the virion associated Env structure. Such optimized immunogens may be more effective in eliciting cross-neutralizing antibodies than the current immunogens. We are in the process of further characterizing the epitopes these MAbs recognize and are actively pursuing the isolation of rhesus antibodies that display broader anti-HIV cross-neutralizing activities.

 
Submitted September 4, 2007

Our Consortium is working on developing a vaccine that would elicit antibodies capable of preventing infection by HIV. Such antibodies are called, neutralizing antibodies (NAbs). Our group takes advantage of the latest advances made in various areas of research, including immunology, virology, molecular biology and computational-based design methodologies, to develop novel strategies to design an anti-HIV vaccine. Investigators with diverse but complementary background participate in our Consortium. Some of our colleagues have not been involved in HIV research in the past, but their expertise allows us to attack this public health problem from new angles. In the past year we spend a lot of time in developing the methodologies and techniques that are necessary for the accomplishment of our goals. We initiated many experiments and made strong progress in all our objectives. For example, we engineered several novel vaccine candidates, some of which were solely designed using computers are tools. These novel vaccines are currently tested in animals. We also isolated novel anti-HIV NAbs and we are now defining their biological activities. Despite the fact that we encountered several unexpected hurdles, mostly due to the novelty of our approaches, overall we are very pleased with our efforts in Year 1 and are looking forward to a successful Year 2.

 
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