scholarly journals Membrane Env liposomes facilitate immunization with multivalent full-length HIV spikes

2021 ◽  
Author(s):  
Daniel P. Leaman ◽  
Armando Stano ◽  
Yajing Chen ◽  
Lei Zhang ◽  
Michael B. Zwick
Keyword(s):  
Neutralizing Antibodies ◽  
Vaccine Development ◽  
Serum Antibody ◽  
Glycosylation Site ◽  
Broadly Neutralizing Antibodies ◽  
Vaccine Platform ◽  
Antibody Recognition ◽  
Correct Orientation ◽  
Cd4 Binding Site ◽  
Antibody Specificities

A major goal of HIV vaccine design is to elicit broadly neutralizing antibodies (bnAbs). Such bnAbs target HIV’s trimeric, membrane embedded envelope glycoprotein spikes, (m)Env. Soluble (s)Env trimers have been used as vaccines, but engineering sEnvs for stability, multivalency and desired antigenicity is problematic, and deletes key neutralizing epitopes on glycoprotein (gp)41 while creating neoepitopes that elicit unwanted antibodies. Meanwhile, multivalent mEnv vaccines are challenging to develop due to trimer instability and low mEnv copy number amid other extraneous proteins on virus-like particles. Here, we describe a multivalent mEnv vaccine platform that does not require protein engineering or extraneous proteins. MEnv trimers were fixed, purified and combined with naked liposomes in mild detergent. On removal of detergent, mEnv spikes were observed embedded in liposome particles (mean diameter 133 nm) in correct orientation. These particles were recognized by HIV bnAbs and not non-nAbs and are designated mEnv liposomes (MELs). Following a sequential immunization scheme in rabbits, MELs elicited antibodies that neutralized tier 2 HIV isolates. Analysis of serum antibody specificities, including those to epitopes involving a missing conserved N-glycosylation site at position 197 near the CD4 binding site on two of the immunogens, provide clues on how nAb responses may be improved with modified immunogens. In sum, MELs are a biochemically defined platform that enable rational immunization strategies to elicit HIV bnAbs using multimerized mEnv. Importance A vaccine that induced broadly neutralizing antibodies against HIV would likely end the AIDS pandemic. Such antibodies target membrane embedded envelope glycoprotein spikes (m)Env that HIV uses to enter cells. Due to HIV Env’s low expression and instability, soluble stabilized Env trimers have been used as vaccine candidates, but these have an altered base that disrupts targets of HIV broadly neutralizing antibodies that bind near the membrane and are not available for all HIV isolates. Here, we describe membrane Env liposomes (MELs) that display a multivalent array of stable mEnvs on liposome particles. MELs showed the expected antibody recognition properties including targeting parts of mEnv missing on soluble Envs. Immunization with MELs elicited antibodies that neutralized diverse HIV isolates. The MEL platform facilitates vaccine development with potentially any HIV Env at high valency, and a similar approach may be useful for eliciting antibodies to membrane embedded targets of therapeutic interest.

scholarly journals Membrane Env liposomes for immunization with HIV spikes

2020 ◽  
Author(s):  
Daniel P. Leaman ◽  
Armando Stano ◽  
Yajing Chen ◽  
Lei Zhang ◽  
Michael B. Zwick
Keyword(s):  
Envelope Glycoprotein ◽  
Neutralizing Antibodies ◽  
Vaccine Development ◽  
Vaccine Design ◽  
Glycosylation Site ◽  
Hiv Vaccine ◽  
Tier 2 ◽  
Broadly Neutralizing Antibodies ◽  
Vaccine Platform ◽  
Cd4 Binding Site

AbstractA key goal in HIV vaccine design remains to elicit broadly neutralizing antibodies (bnAbs) against the membrane-embedded envelope glycoprotein spike (mEnv). However, mEnv has lagged behind engineered soluble Envs in vaccine development due to low expression yields and the presence of extraneous proteins on particles. Here, we describe a mEnv vaccine platform that requires no extra proteins or protein engineering. MEnv trimers were fixed, purified and combined with liposomes in mild detergent. On removal of detergent, mEnvs were observed embedded in particles, designated mEnv liposomes (MELs), which were recognized by HIV bnAbs but not non-nAbs. Following sequential immunization in rabbits, MEL antisera neutralized select tier 2 HIV isolates. Variations between the Env immunogens, including a missing N-glycosylation site at position 197 near the CD4 binding site, provide insights into the specificities elicited and possible ways to improve immunogens. MELs can facilitate vaccine design to elicit HIV bnAbs using biochemically defined and multimerized mEnv.

scholarly journals Griffithsin Retains Anti-HIV-1 Potency with Changes in gp120 Glycosylation and Complements Broadly Neutralizing Antibodies PGT121 and PGT126

2019 ◽  
Vol 64 (1) ◽  
Author(s):  
Kathryn Fischer ◽  
Kimberly Nguyen ◽  
Patricia J. LiWang
Keyword(s):  
Neutralizing Antibodies ◽  
Glycosylation Site ◽  
Significant Loss ◽  
Broadly Neutralizing Antibodies ◽  
Clade B ◽  
Key Factor ◽  
Clade C ◽  
Cd4 Binding Site ◽  
Gp120 Glycosylation ◽  
Hiv 1

ABSTRACT Griffithsin (Grft) is an antiviral lectin that has been shown to potently inhibit HIV-1 by binding high-mannose N-linked glycosylation sites on HIV-1 gp120. A key factor for Grft potency is glycosylation at N295 of gp120, which is directly adjacent to N332, a target glycan for an entire class of broadly neutralizing antibodies (bNAbs). Here, we unify previous work on the importance of other glycans to Grft potency against HIV-1 and Grft’s role in mediating the conformational change of gp120 by mutating nearly every glycosylation site in gp120. In addition to a significant loss of Grft activity by the removal of glycosylation at N295, glycan absence at N332 or N448 was found to have moderate effects on Grft potency. Interestingly, in the absence of N295, Grft effectiveness could be improved by a mutation that results in the glycan at N448 shifting to N446, indicating that the importance of individual glycans may be related to their effect on glycosylation density. Grft’s ability to alter the structure of gp120, exposing the CD4 binding site, correlated with the presence of glycosylation at N295 only in clade B strains, not clade C strains. We further demonstrate that Grft can rescue the activity of the bNAbs PGT121 and PGT126 in the event of a loss or a shift of glycosylation at N332, where the bNAbs suffer a drastic loss of potency. Despite targeting the same region, Grft in combination with PGT121 and PGT126 produced additive effects. This indicates that Grft could be an important combinational therapeutic.

scholarly journals Impact of glycan positioning on HIV-1 Env glycan shield density, function, and antibody recognition

2020 ◽  
Author(s):  
Qing Wei ◽  
Audra A. Hargett ◽  
Barbora Knoppova ◽  
Alexandra Duverger ◽  
Reda Rawi ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Glycosylation Site ◽  
Cell Entry ◽  
Virus Infectivity ◽  
Ripple Effect ◽  
Dynamic Simulations ◽  
Antibody Recognition ◽  
Cd4 Binding Site ◽  
Env Protein ◽  
Hiv 1

AbstractN-glycans, which represent >50% mass of the HIV-1 envelope (Env) trimer, play important roles for virus-cell entry and immune evasion. How each glycan unit interacts to shape the Env protein-sugar complex and affects Env function is not well understood. Here, high-resolution glycomics analysis of two Env variants from the same donor, with differing functional characteristics and N-glycosylation-site composition, revealed that changes to key N-glycosylation-site not only affected the Env structure at distant locations, but also had a ripple effect on Env-wide glycan processing, virus infectivity, and antibody recognition and virus neutralization. Specifically, the N262 glycan, although not located in the CD4-binding site, controlled Env binding to the CD4 receptor, affected the recognition of Env by several glycan-dependent broadly neutralizing antibodies, and altered heterogeneity of glycosylation at several sites, with N156, N160, and N448 displaying limited glycan processing. Molecular dynamic simulations visualized how specific oligosaccharide positions can move to compensate for loss of a glycan. This study demonstrates how changes in individual glycan units can alter molecular dynamics and processing of the Env-glycan shield and, consequently, Env function.

scholarly journals Multiple Antibody Lineages in One Donor Target the Glycan-V3 Supersite of the HIV-1 Envelope Glycoprotein and Display a Preference for Quaternary Binding

2016 ◽  
Vol 90 (23) ◽  
pp. 10574-10586 ◽  
Author(s):  
Nancy S. Longo ◽  
Matthew S. Sutton ◽  
Andrea R. Shiakolas ◽  
Javier Guenaga ◽  
Marissa C. Jarosinski ◽  
...  
Keyword(s):  
Envelope Glycoprotein ◽  
Neutralizing Antibodies ◽  
Vaccine Development ◽  
Culture Method ◽  
High Serum ◽  
Broadly Neutralizing Antibodies ◽  
Monomeric Gp120 ◽  
Antibody Specificities ◽  
V3 Region ◽  
Hiv 1

ABSTRACT One of the goals of HIV-1 vaccine development is the elicitation of neutralizing antibodies against vulnerable regions on the envelope glycoprotein (Env) viral spike. Broadly neutralizing antibodies targeting the Env glycan-V3 region (also called the N332 glycan supersite) have been described previously, with several single lineages each derived from different individual donors. We used a high-throughput B-cell culture method to isolate neutralizing antibodies from an HIV-1-infected donor with high serum neutralization breadth. Clonal relatives from three distinct antibody lineages were isolated. Each of these antibody lineages displayed modest breadth and potency but shared several characteristics with the well-characterized glycan-V3 antibodies, including dependence on glycans N332 and N301, VH4 family gene utilization, a heavy chain complementarity-determining region 2 (CDRH2) insertion, and a longer-than-average CDRH3. In contrast to previously described glycan-V3 antibodies, these antibodies preferentially recognized the native Env trimer compared to monomeric gp120. These data indicate the diversity of antibody specificities that target the glycan-V3 site. The quaternary binding preference of these antibodies suggests that that their elicitation likely requires the presentation of a native-like trimeric Env immunogen. IMPORTANCE Broadly neutralizing antibodies targeting the HIV-1 glycan-V3 region with single lineages from individual donors have been described previously. Here we describe three lineages from a single donor, each of which targets glycan-V3. Unlike previously described glycan-V3 antibodies, these mature antibodies bind preferentially to the native Env trimer and weakly to the gp120 monomer. These data extend our knowledge of the immune response recognition of the N332 supersite region and suggest that the mode of epitope recognition is more complex than previously anticipated.

scholarly journals Engineering HIV envelope protein to activate germline B cell receptors of broadly neutralizing anti-CD4 binding site antibodies

2013 ◽  
Vol 210 (4) ◽  
pp. 655-663 ◽  
Author(s):  
Andrew T. McGuire ◽  
Sam Hoot ◽  
Anita M. Dreyer ◽  
Adriana Lippy ◽  
Andrew Stuart ◽  
...  
Keyword(s):  
Binding Site ◽  
Neutralizing Antibodies ◽  
Variable Region ◽  
Neutralizing Antibody ◽  
Glycosylation Site ◽  
Affinity Maturation ◽  
Broadly Neutralizing Antibodies ◽  
Cd4 Binding Site ◽  
Broadly Neutralizing Antibody ◽  
Hiv Envelope

Broadly neutralizing antibodies (bnAbs) against HIV are believed to be a critical component of the protective responses elicited by an effective HIV vaccine. Neutralizing antibodies against the evolutionarily conserved CD4-binding site (CD4-BS) on the HIV envelope glycoprotein (Env) are capable of inhibiting infection of diverse HIV strains, and have been isolated from HIV-infected individuals. Despite the presence of anti–CD4-BS broadly neutralizing antibody (bnAb) epitopes on recombinant Env, Env immunization has so far failed to elicit such antibodies. Here, we show that Env immunogens fail to engage the germline-reverted forms of known bnAbs that target the CD4-BS. However, we found that the elimination of a conserved glycosylation site located in Loop D and two glycosylation sites located in variable region 5 of Env allows Env-binding to, and activation of, B cells expressing the germline-reverted BCRs of two potent broadly neutralizing antibodies, VRC01 and NIH45-46. Our results offer a possible explanation as to why Env immunogens have been ineffective in stimulating the production of such bNAbs. Importantly, they provide key information as to how such immunogens can be engineered to initiate the process of antibody-affinity maturation against one of the most conserved Env regions.

scholarly journals Structural basis for germline antibody recognition of HIV-1 immunogens

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Louise Scharf ◽  
Anthony P West ◽  
Stuart A Sievers ◽  
Courtney Chen ◽  
Siduo Jiang ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Affinity Maturation ◽  
Structural Basis ◽  
Binding Modes ◽  
Broadly Neutralizing Antibodies ◽  
Antibody Recognition ◽  
Immunogen Design ◽  
Cd4 Binding Site ◽  
Modified Forms ◽  
Hiv 1

Efforts to elicit broadly neutralizing antibodies (bNAbs) against HIV-1 require understanding germline bNAb recognition of HIV-1 envelope glycoprotein (Env). The VRC01-class bNAb family derived from the VH1-2*02 germline allele arose in multiple HIV-1–infected donors, yet targets the CD4-binding site on Env with common interactions. Modified forms of the 426c Env that activate germline-reverted B cell receptors are candidate immunogens for eliciting VRC01-class bNAbs. We present structures of germline-reverted VRC01-class bNAbs alone and complexed with 426c-based gp120 immunogens. Germline bNAb–426c gp120 complexes showed preservation of VRC01-class signature residues and gp120 contacts, but detectably different binding modes compared to mature bNAb-gp120 complexes. Unlike typical antibody-antigen interactions, VRC01–class germline antibodies exhibited preformed antigen-binding conformations for recognizing immunogens. Affinity maturation introduced substitutions increasing induced-fit recognition and electropositivity, potentially to accommodate negatively-charged complex-type N-glycans on gp120. These results provide general principles relevant to the unusual evolution of VRC01–class bNAbs and guidelines for structure-based immunogen design.

scholarly journals Functional Stability of HIV-1 Envelope Trimer Affects Accessibility to Broadly Neutralizing Antibodies at Its Apex

2017 ◽  
Vol 91 (24) ◽  
Author(s):  
Syna Kuriakose Gift ◽  
Daniel P. Leaman ◽  
Lei Zhang ◽  
Arthur S. Kim ◽  
Michael B. Zwick
Keyword(s):  
Neutralizing Antibodies ◽  
Vaccine Development ◽  
Inverse Correlation ◽  
Functional Stability ◽  
Broadly Neutralizing Antibodies ◽  
Antibody Recognition ◽  
Partial Neutralization ◽  
The Stability ◽  
Hiv 1 ◽  
Trimeric Envelope

ABSTRACT The trimeric envelope glycoprotein spike (Env) of HIV-1 is the target of vaccine development to elicit broadly neutralizing antibodies (bnAbs). Env trimer instability and heterogeneity in principle make subunit interfaces inconsistent targets for the immune response. Here, we investigate how functional stability of Env relates to neutralization sensitivity to V2 bnAbs and V3 crown antibodies that engage subunit interfaces upon binding to unliganded Env. Env heterogeneity was inferred when antibodies neutralized a mutant Env with a plateau of less than 100% neutralization. A statistically significant correlation was found between the stability of mutant Envs and the MPN of V2 bnAb, PG9, as well as an inverse correlation between stability of Env and neutralization by V3 crown antibody, 447-52D. A number of Env-stabilizing mutations and V2 bnAb-enhancing mutations were identified in Env, but they did not always overlap, indicating distinct requirements of functional stabilization versus antibody recognition. Blocking complex glycosylation of Env affected V2 bnAb recognition, as previously described, but also notably increased functional stability of Env. This study shows how instability and heterogeneity affect antibody sensitivity of HIV-1 Env, which is relevant to vaccine design involving its dynamic apex. IMPORTANCE The Env trimer is the only viral protein on the surface of HIV-1 and is the target of neutralizing antibodies that reduce viral infectivity. Quaternary epitopes at the apex of the spike are recognized by some of the most potent and broadly neutralizing antibodies to date. Being that their glycan-protein hybrid epitopes are at subunit interfaces, the resulting heterogeneity can lead to partial neutralization. Here, we screened for mutations in Env that allowed for complete neutralization by the bnAbs. We found that when mutations outside V2 increased V2 bnAb recognition, they often also increased Env stability-of-function and decreased binding by narrowly neutralizing antibodies to the V3 crown. Three mutations together increased neutralization by V2 bnAb and eliminated binding by V3 crown antibodies. These results may aid the design of immunogens that elicit antibodies to the trimer apex.

scholarly journals Frequent development of broadly neutralizing antibodies in early life in a large cohort of HIV-infected children

2021 ◽  
Author(s):  
Genevieve G. Fouda ◽  
Amanda Lucier ◽  
Youyi Fong ◽  
Shuk Hang Li ◽  
Maria Dennis ◽  
...  
Keyword(s):  
Early Life ◽  
Neutralizing Antibodies ◽  
Igg Subclass ◽  
Tier 2 ◽  
Broadly Neutralizing Antibodies ◽  
Significant Difference ◽  
Cd4 Binding Site ◽  
Binding Antibody ◽  
Antibody Specificities ◽  
Living With Hiv

Recent studies conducted in small cohorts of children have indicated that broadly neutralizing antibodies (bnAbs) may develop earlier after HIV infection compared to adults. To define the frequency and kinetics of bnAb responses in a larger pediatric cohort, we evaluated plasma from 212 ART-naïve, children living with HIV aged 1 to 3 years. Neutralization breadth and potency was assessed using a panel of 10 tier-2 viruses and compared to those of adults with chronic HIV. Further, the magnitude, epitope specificity and IgG subclass distribution of Env-specific antibodies were also assessed using a binding antibody multiplex assay. We found that 1-year-old children demonstrated neutralization breadth comparable to that of chronically-infected adults, and breadth continued to increase with age such that the pediatric cohort overall exhibited significantly greater neutralization breadth than adults (p= 0.014). Similarly, binding antibody responses increased with age, and the levels in 2 to 3 year-old children were comparable to those of adults. Overall, there was no significant difference in antibody specificities or IgG subclass distribution between the pediatric and adult cohorts. Interestingly, the neutralization activity was mapped to a single epitope (CD4 binding site, V2 or V3 glycans) in only 5 of 38 pediatric broadly neutralizing samples, suggesting a polyclonal neutralization response may develop in most children. These results contribute to a growing body of evidence suggesting that the early life immune system may present advantages for the development of an effective HIV vaccine.

scholarly journals Broadly neutralizing antibodies and vaccine design against HIV-1 infection

2019 ◽  
Vol 14 (1) ◽  
pp. 30-42 ◽  
Author(s):  
Qian Wang ◽  
Linqi Zhang
Keyword(s):  
Neutralizing Antibodies ◽  
Vaccine Development ◽  
Vaccine Design ◽  
Therapeutic Interventions ◽  
Type I ◽  
Broadly Neutralizing Antibodies ◽  
Dynamic Studies ◽  
Immunodeficiency Virus ◽  
Hiv 1 ◽  
Remarkable Progress

AbstractRemarkable progress has been achieved for prophylactic and therapeutic interventions against human immunodeficiency virus type I (HIV-1) through antiretroviral therapy. However, vaccine development has remained challenging. Recent discoveries in broadly neutralizing monoclonal antibodies (bNAbs) has led to the development of multiple novel vaccine approaches for inducing bNAbs-like antibody response. Structural and dynamic studies revealed several vulnerable sites and states of the HIV-1 envelop glycoprotein (Env) during infection. Our review aims to highlight these discoveries and rejuvenate our endeavor in HIV-1 vaccine design and development.

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