Engineered B cells expressing an anti-HIV antibody enable memory retention, isotype switching and clonal expansion

Nahmad, Alessio D.; Raviv, Yuval; Horovitz-Fried, Miriam; Sofer, Ilan; Akriv, Tal; Nataf, Daniel; Dotan, Iris; Carmi, Yaron; Burstein, David; Wine, Yariv; Benhar, Itai; Barzel, Adi

doi:10.1038/s41467-020-19649-1

Cited by 60 publications

(59 citation statements)

References 43 publications

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“…Such single chain coding can further allow the expression of bi-specific bNAbs, which may be required to provide long-term protection from HIV resurgence 1 . Safety may be further improved by using more specific nucleases 22,23 and by having the bNAb gene preceded by a splice acceptor rather than by a promoter, to reduce expression from off-target integration 7,12 . Both safety and efficacy may benefit from embedding B cell specific targeting moieties in the AAV vector 24 or in a non-viral alternative 25 .…”

Section: Resultsmentioning

confidence: 99%

In vivoengineered B cells retain memory and secrete high titers of anti-HIV antibodies in mice

Nahmad

Zelikson

et al. 2021

Preprint

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As a potential single-shot HIV therapy, transplanted engineered B cells allow robust secretion of broadly neutralizing antibodies (bNAbs). However, ex vivo engineering of autologous B cells is expensive and requires specialized facilities, while allogeneic B cell therapy necessitates MHC compatibility. Here, we develop in vivo B cell engineering, by injecting two adeno associated viral vectors, one coding for saCas9 and another coding for a bNAb. Following immunizations, we demonstrate memory retention and bNAb secretion at neutralizing titers. We observed minimal CRISPR/Cas9 off-target cleavage, using unbiased CHANGE-Seq analysis, while on-target cleavage at undesired tissues is reduced by expressing saCas9 from a B cell specific promoter. In vivo B cell engineering is thus a safe, potent and scalable method for expressing desired antibodies against HIV and beyond.

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Section: Resultsmentioning

confidence: 99%

In vivoengineered B cells retain memory and secrete high titers of anti-HIV antibodies in mice

Nahmad

Zelikson

et al. 2021

Preprint

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“…AAV vectors are invaluable reagents for site-specific genome editing of human hematopoietic cells, with AAV6 serotypes in particular being widely used to deliver homology donors to HSPCs, 2, 4, 11, 12 T cells, 13–15 and B cells. 16, 17, 19 The in vitro tropism of AAV6 for human hematopoietic cells, 2, 13, 16 as well its weak induction of innate immune pathways that could trigger harmful biological consequences in engineered cells, 41, 42 may contribute to its success. In addition, the vector’s ability to transduce both dividing and quiescent cells 43 may also be beneficial, particularly if the genome is used as a homology template only after second-strand synthesis as some have suggested, 7 though the G2/S phase restriction of cellular factors required for HDR 25 may limit the advantages conferred by this attribute.…”

Section: Discussionmentioning

confidence: 99%

Optimization of AAV6 transduction enhances site-specific genome editing of primary human lymphocytes

Rogers

Huang

Clark

et al. 2021

Preprint

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Adeno-associated virus serotype 6 (AAV6) is a valuable reagent for genome editing of hematopoietic cells due to its ability to serve as a homology donor template. However, a comprehensive study of AAV6 transduction of hematopoietic cells in culture, with the goal of maximizing ex vivo genome editing, has not been reported. Here, we evaluated how the presence of serum, culture volume, transduction time, and electroporation parameters could influence AAV6 transduction. Based on these results, we identified an optimized protocol for genome editing of human lymphocytes based on a short, highly concentrated AAV6 transduction in the absence of serum, followed by electroporation with a targeted nuclease. In human CD4+ T cells and B cells, this protocol improved editing rates up to 7-fold and 21-fold respectively, when compared to standard AAV6 transduction protocols described in the literature. As a result, editing frequencies could be maintained using 50-100-fold less AAV6, which also reduced cellular toxicity. Our results highlight the important contribution of cell culture conditions for ex vivo genome editing with AAV6 vectors and provide a blueprint for improving AAV6-mediated homology-directed editing of human T and B cells.

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“…One personalized approach is to collect memory B cells from an individual living with HIV and replace the BCR with a bNAb of choice through CRISPR/Cas9 editing, then infuse the engineered cells back into the individual, mirroring the approach used for CAR-T cell therapy approved for some lymphomas ( 159 , 160 ). This approach has shown success in mouse models performed by different groups introducing bNAb VRC01 ( 161 ) or 3BNC117 ( 162 ) as the BCR, with in vivo transfer resulting in a memory B cell population that retains the ability to class switch, undergo somatic hypermutation, and clonally expand and differentiate into plasmablasts following immunization with HIV Env.…”

Section: Cultivating Bnab Development For Treatment and Preventionmentioning

confidence: 99%

Advancing HIV Broadly Neutralizing Antibodies: From Discovery to the Clinic

Spencer

Shapiro

Haigwood

et al. 2021

Front. Public Health

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Despite substantial progress in confronting the global HIV-1 epidemic since its inception in the 1980s, better approaches for both treatment and prevention will be necessary to end the epidemic and remain a top public health priority. Antiretroviral therapy (ART) has been effective in extending lives, but at a cost of lifelong adherence to treatment. Broadly neutralizing antibodies (bNAbs) are directed to conserved regions of the HIV-1 envelope glycoprotein trimer (Env) and can block infection if present at the time of viral exposure. The therapeutic application of bNAbs holds great promise, and progress is being made toward their development for widespread clinical use. Compared to the current standard of care of small molecule-based ART, bNAbs offer: (1) reduced toxicity; (2) the advantages of extended half-lives that would bypass daily dosing requirements; and (3) the potential to incorporate a wider immune response through Fc signaling. Recent advances in discovery technology can enable system-wide mining of the immunoglobulin repertoire and will continue to accelerate isolation of next generation potent bNAbs. Passive transfer studies in pre-clinical models and clinical trials have demonstrated the utility of bNAbs in blocking or limiting transmission and achieving viral suppression. These studies have helped to define the window of opportunity for optimal intervention to achieve viral clearance, either using bNAbs alone or in combination with ART. None of these advances with bNAbs would be possible without technological advancements and expanding the cohorts of donor participation. Together these elements fueled the remarkable growth in bNAb development. Here, we review the development of bNAbs as therapies for HIV-1, exploring advances in discovery, insights from animal models and early clinical trials, and innovations to optimize their clinical potential through efforts to extend half-life, maximize the contribution of Fc effector functions, preclude escape through multiepitope targeting, and the potential for sustained delivery.

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Engineered B cells expressing an anti-HIV antibody enable memory retention, isotype switching and clonal expansion

Cited by 60 publications

References 43 publications

In vivoengineered B cells retain memory and secrete high titers of anti-HIV antibodies in mice

In vivoengineered B cells retain memory and secrete high titers of anti-HIV antibodies in mice

Optimization of AAV6 transduction enhances site-specific genome editing of primary human lymphocytes

Advancing HIV Broadly Neutralizing Antibodies: From Discovery to the Clinic

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