Parvovirus Capsid-Antibody Complex Structures Reveal Conservation of Antigenic Epitopes Across the Family

Emmanuel, Shanan N.; Mietzsch, Mario; Tseng, Yu Shan; Smith, J. W. G.; Agbandje‐McKenna, Mavis

doi:10.1089/vim.2020.0022

Cited by 21 publications

(30 citation statements)

References 139 publications

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“…Anti-AAV8 antibodies recognize overlapping capsid antigenic footprints. In order to identify the antigenic regions of AAV8 capsids cryo-electron microscopy (cryo-EM) and image reconstruction was used to determine the three-dimensional (3D) structures of capsid-antibody complexes (18). For this purpose, purified capsids were mixed with purified Fabs at ratios of 1:120 (capsid:antibody) and the complex used to collect cryo-EM micrographs for structure determination.…”

Section: Resultsmentioning

confidence: 99%

“…Overall, these antibodies bound to common antigenic regions found among the parvoviruses. According to the recently proposed classification system of the parvoviral antibodies, HL2372 would be assigned to group I, ADK8/9 to group II, and ADK8, HL2381, and HL2383 to group III (18).…”

Section: Downloaded Frommentioning

confidence: 99%

“…A 2D roadmap can be used to depict receptor and antigenic footprint on October 30, 2020 by guest http://jvi.asm.org/ Downloaded from residues mapped by cryo-EM and localized to the capsid surface. These studies provide the basis for engineering the AAV capsid surface to potentially achieve improved cross-functionality (18).…”

Section: Introductionmentioning

confidence: 98%

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Coevolution of Adeno-associated Virus Capsid Antigenicity and Tropism through a Structure-Guided Approach

Havlik

Simon

Smith

et al. 2020

J Virol

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Adeno-associated viruses (AAV) are composed of non-enveloped, icosahedral protein shells that can be adapted to package and deliver recombinant therapeutic DNA. Approaches to engineer recombinant capsids for gene therapy applications have focused on rational design or library-based approaches that can address one or two desirable attributes; however, there is an unmet need to comprehensively improve AAV vector properties. Such cannot be achieved by utilizing sequence data alone, but requires harnessing the 3D structural properties of AAV capsids. Here, we solve the structures of a natural AAV isolate complexed with antibodies using cryo-electron microscopy and harness this structural information to engineer AAV capsid libraries through saturation mutagenesis of different antigenic footprints. Each surface loop was evolved by infectious cycling in the presence of a helper Adenovirus to yield a new AAV variant that then serves as a template for evolving the next surface loop. This stepwise process yielded a humanized AAV8 capsid (AAVhum.8) displaying non-natural surface loops that displays simultaneously tropism for human hepatocytes, increased gene transfer efficiency and neutralizing antibody evasion. Specifically, AAVhum.8 can better evade neutralizing antisera from multiple species compared to AAV8. Further, AAVhum.8 displays robust transduction in a human liver xenograft mouse model with expanded tropism for both murine and human hepatocytes. This work supports the hypothesis that critical properties such as AAV capsid antibody evasion and tropism can be co-evolved by combining rational design and library-based evolution for clinical gene therapy. Importance Clinical gene therapy with recombinant AAV vectors has largely relied on natural capsid isolates. There is an unmet need to comprehensively improve AAV tissue tropism, transduction efficiency and antibody evasion. Such cannot be achieved by utilizing capsid sequence data alone, but requires harnessing the 3D structural properties of AAV capsids. Here, we combine rational design and library-based evolution to co-evolve multiple, desirable properties onto AAV by harnessing 3D structural information.

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Section: Resultsmentioning

confidence: 99%

Section: Downloaded Frommentioning

confidence: 99%

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Coevolution of Adeno-associated Virus Capsid Antigenicity and Tropism through a Structure-Guided Approach

Havlik

Simon

Smith

et al. 2020

J Virol

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“…While the leakage of ligands during elution from the affinity columns is likely low, neutralization by AVB helps understand the functions of the capsids. In AAV1 and AAV5, both neutralized by AVB, the AVB binding site overlaps with the binding epitope of neutralizing antibodies ADK1b, ADK5a, and ADK5b, 30 respectively. Binding of AVB to the AAV5 capsid likely blocks binding of AAVR to cause neutralization, 35 but not AAV1 since AAVR binds differently in AAV1 capsids.…”

Section: Discussionmentioning

confidence: 99%

“… 16 These VRs result in structural differences between serotypes and contribute to functional phenotypes, including receptor attachment, tissue tropism, transduction efficiency, and antigenic reactivity. 29 , 30 …”

Section: Introductionmentioning

confidence: 99%

Characterization of AAV-Specific Affinity Ligands: Consequences for Vector Purification and Development Strategies

Mietzsch

Smith

et al. 2020

Molecular Therapy - Methods & Clinical Development

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Affinity-based purification of adeno-associated virus (AAV) vectors has replaced density-based methods for vectors used in clinical settings. This method utilizes camelid single-domain antibodies recognizing AAV capsids. These include AVB Sepharose (AVB) and POROS CaptureSelect affinity ligand for AAV8 (CSAL8) and AAV9 (CSAL9). In this study, we utilized cryo-electron microscopy and 3D image reconstruction to map the binding sites of these affinity ligands on the capsids of several AAV serotypes, including AAV1, AAV2, AAV5, AAV8, and AAV9, representing the range of sequence and structure diversity among AAVs. The AAV-ligand complex structures showed that AVB and CSAL9 bound to the 5-fold capsid region, although in different orientations, and CSAL8 bound to the side of the 3-fold protrusion. The AAV contact residues required for ligand binding, and thus AAV purification, and the ability of the ligands to neutralize infection were analyzed. The data show that only a few residues within the epitopes served to block affinity ligand binding. Neutralization was observed for AAV1 and AAV5 with AVB, for AAV1 with CSAL8, and for AAV9 with CSAL9, associated with regions that overlap with epitopes for neutralizing monoclonal antibodies against these capsids. This information is critical and could be generally applicable in the development of novel AAV vectors amenable to affinity column purification.

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Adeno-associated virus-vectored delivery of HIV biologics: the promise of a “single-shot” functional cure for HIV infection

Hahn

Martins²

2023

Journal of Virus Eradication

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Parvovirus Capsid-Antibody Complex Structures Reveal Conservation of Antigenic Epitopes Across the Family

Cited by 21 publications

References 139 publications

Coevolution of Adeno-associated Virus Capsid Antigenicity and Tropism through a Structure-Guided Approach

Coevolution of Adeno-associated Virus Capsid Antigenicity and Tropism through a Structure-Guided Approach

Characterization of AAV-Specific Affinity Ligands: Consequences for Vector Purification and Development Strategies

Adeno-associated virus-vectored delivery of HIV biologics: the promise of a “single-shot” functional cure for HIV infection

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