AAV vectors: The Rubik’s cube of human gene therapy

Pupo, Amaury; Fernández, Audry; Low, Siew Hui; François, Achille; Suárez-Amarán, Lester; Samulski, R. Jude

doi:10.1016/j.ymthe.2022.09.015

Cited by 162 publications

(109 citation statements)

References 340 publications

(524 reference statements)

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“…36 The mean CP/mL and VG/mL values shown in Supplementary Table S1 obtained with the Wyatt column were used to assess linearity and accuracy by recovery. A plot of the theoretical CP/mL and VG/mL against the measured values produced strong linearity ( R 2 > 0.999) over two orders of magnitude ( Fig. 2A, B ).…”

Section: Resultsmentioning

confidence: 84%

“…There was a trend with cryo-EM that suggested a bias toward the quantification of empty capsids over full capsids. Nevertheless, the cryo-EM results were linear with respect to the SV-AUC values ( R 2 = 0.952; Fig. 5 ).…”

Section: Resultsmentioning

confidence: 89%

“…These viruses are nonpathogenic, exhibit lower immunogenicity than most viruses, and are capable of transducing a variety of cell types. 1 , 2 In addition, a viral vector can package ∼4.7 kb of DNA, be produced at scale to meet clinical trial demands with evolved capsids and tissue-specific cassette design that allow for targeted delivery and expression. 3–6 The icosahedral AAV capsid is composed of viral proteins (VP) VP1, VP2, and VP3 in an ∼1:1:10 ratio.…”

Section: Introductionmentioning

confidence: 99%

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Application of Size Exclusion Chromatography with Multiangle Light Scattering in the Analytical Development of a Preclinical Stage Gene Therapy Program

et al. 2023

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To provide safe recombinant adeno-associated viruses (rAAV) to patients, scalable manufacturing processes are required. However, these processes may introduce impurities that impact the performance and quality of the final drug product. Empty rAAV capsids are product-related impurities. Regulatory guidance requires that accurate analytical methods be implemented early in product development to measure the level of empty capsids. A process confirmation vector, produced from 200 L production, was used to develop and optimize a size exclusion chromatography with UV and multiangle light scattering (SEC-MALS) method. Vector produced from a 500 L production was used to assess the full-to-empty ratio using the following analytical methods: sedimentation velocity analytical ultracentrifugation (SV-AUC), droplet digital PCR (ddPCR) with capsid enzyme-linked immunosorbent assay (ELISA), bulk absorbance at 260/280 nm, cryogenic electron microscopy, and SEC-MALS. This test article was used for a 30-day, non-Good Laboratory Practices animal study that assessed biodistribution of the product (STRX-330). SEC-MALS outperformed the other methods and correlated well with SV-AUC values of full-to-empty particles. In addition, SEC-MALS agreed with ddPCR and ELISA measurements for vector genomes/mL and capsid particles/mL, respectively. SEC-MALS was linear, accurate, and precise while achieving chromatography quality control (QC) recommendations. Compared to other stability-indicating assays, SEC-MALS performed similarly to ddPCR, capsid ELISA, and infectivity assays in accelerated stress studies. In response to alkaline, but not acidic stress, SEC-MALS indicated distinct changes in the DNA content of the monomer Adeno-associated viruses (AAV) peak for STRX-330, which was supported by ddPCR data. Conversely, acidic treatment resulted in more aggregated vector, but did not impact the DNA content. This work indicates that SEC-MALS is a valuable analytical tool in the analytical development and QC testing of AAV. In addition, this work suggests that SEC-MALS can provide fundamental understanding of AAV in response to environmental stress. This may impact steps of the manufacturing process to minimize conditions that reduce performance.

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Section: Resultsmentioning

confidence: 84%

Section: Resultsmentioning

confidence: 89%

Section: Introductionmentioning

confidence: 99%

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Application of Size Exclusion Chromatography with Multiangle Light Scattering in the Analytical Development of a Preclinical Stage Gene Therapy Program

et al. 2023

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“…Therefore, this targeting strategy is well suited to target cardiomyocytes, a class of cells with poor replication capacity, and have an important role in driving clinical translation of cardiomyocyte regeneration ( 134 ). Furthermore, similar to exosomes and liposomes, AAV itself has few side effects, high targeting capacity, and is malleable and can likewise be engineered to obtain higher or more unique targeting capacity ( 135 ). Thus using AAV for immunomodulation is a way to consider, for example, the aforementioned Ep3 receptor, where the single-stranded sequence of Ep3 is delivered to monocytes via AAV to promote Ep3 overexpression, thus better attenuating inflammation and promoting repair ( Figure 3C ).…”

Section: Targeting Strategies Worth Considering For Novel Immunomodul...mentioning

confidence: 99%

Immune cells drive new immunomodulatory therapies for myocardial infarction: From basic to clinical translation

Nian

Huang

2023

Front. Immunol.

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The high incidence of heart failure secondary to myocardial infarction (MI) has been difficult to effectively address. MI causes strong aseptic inflammation, and infiltration of different immune cells and changes in the local inflammatory microenvironment play a key regulatory role in ventricular remodeling. Therefore, the possibility of improving the prognosis of MI through targeted immunity has been of interest and importance in MI. However, previously developed immune-targeted therapies have not achieved significant success in clinical trials. Here, we propose that the search for therapeutic targets from different immune cells may be more precise and lead to better clinical translation. Specifically, this review summarizes the role and potential therapeutic targets of various immune cells in ventricular remodeling after MI, especially monocytes/macrophages and neutrophils, as a way to demonstrate the importance and potential of immunomodulatory therapies for MI. In addition, we analyze the reasons for the failure of previous immunomodulatory therapies and the issues that need to be addressed, as well as the prospects and targeting strategies of using immune cells to drive novel immunomodulatory therapies, hoping to advance the development of immunomodulatory therapies by providing evidence and new ideas.

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“…AAVs have 12 or more serotypes, with their capsid proteins varying to a certain extent. Each serotype may have a distinct affinity to specific cell types, allowing AAVs to exhibit preferred tissue targeting features in in vivo treatments [ 56 , 57 , 58 ]. With further capsid manipulation, AAVs are capable of crossing the blood–brain barrier [ 59 ].…”

Section: Crispr Function and Cellular Deliverymentioning

confidence: 99%

New Therapeutics for Extracellular Vesicles: Delivering CRISPR for Cancer Treatment

Yan

Liang

2022

IJMS

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Cancers are defined by genetic defects, which underlines the prospect of using gene therapy in patient care. During the past decade, CRISPR technology has rapidly evolved into a powerful gene editing tool with high fidelity and precision. However, one of the impediments slowing down the clinical translation of CRISPR-based gene therapy concerns the lack of ideal delivery vectors. Extracellular vesicles (EVs) are nano-sized membrane sacs naturally released from nearly all types of cells. Although EVs are secreted for bio-information conveyance among cells or tissues, they have been recognized as superior vectors for drug or gene delivery. Recently, emerging evidence has spotlighted EVs in CRISPR delivery towards cancer treatment. In this review, we briefly introduce the biology and function of the CRISPR system and follow this with a summary of current delivery methods for CRISPR applications. We emphasize the recent progress in EV-mediated CRISPR editing for various cancer types and target genes. The reported strategies for constructing EV-CRISPR vectors, as well as their limitations, are discussed in detail. The review aims to throw light on the clinical potential of engineered EVs and encourage the expansion of our available toolkit to defeat cancer.

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AAV vectors: The Rubik’s cube of human gene therapy

Cited by 162 publications

References 340 publications

Application of Size Exclusion Chromatography with Multiangle Light Scattering in the Analytical Development of a Preclinical Stage Gene Therapy Program

Application of Size Exclusion Chromatography with Multiangle Light Scattering in the Analytical Development of a Preclinical Stage Gene Therapy Program

Immune cells drive new immunomodulatory therapies for myocardial infarction: From basic to clinical translation

New Therapeutics for Extracellular Vesicles: Delivering CRISPR for Cancer Treatment

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