Similar Datasets
Project description:Evolving a novel AAV variant for efficient gene delivery to murine T lymphocytes
| PRJNA893602 | ENA
Project description:Precise targeting of large transgenes to T cells using homology-directed repair has been transformative for adoptive cell therapies and T cell biology. Non-toxic delivery of DNA templates via adeno-associated virus (AAV) has greatly improved knock-in efficiencies, but the tropism of current AAV serotypes restricts their use to human T cells employed in immunodeficient mouse models. To enable targeted knock-ins in murine T cells, we evolved Ark313, a synthetic AAV that exhibits high transduction efficiency in murine T cells. We performed a genome-wide knockout screen and identified QA2 as an essential factor for Ark313 infection. We demonstrate that Ark313 can be used for nucleofection-free DNA delivery, CRISPR/Cas9-mediated knockouts, and targeted integration of large transgenes. Ark313 enables pre-clinical modeling of Trac-targeted CAR-T and transgenic TCR-T cells in immunocompetent models. Efficient gene targeting in murine T cells holds great potential for improved cell therapies and opens new avenues in experimental T cell immunology.
2022-12-31 | GSE216819 | GEO
Project description:Autologous chimeric antigen receptor T (CAR-T) cell therapy has exhibited prominent therapeutic effectiveness in hematologic malignancy and autoimmune disease, but the manufacturing complexity and the required lymphodepletion hindered its wide clinical application. Engineering human T cells in vivo holds promise to conquer these limitations but requires effective T cell-targeted CAR delivery with demonstrated safety. Recombinant adeno-associated virus (rAAV) emerged as a promising vehicle in eight FDA-approved gene therapy drugs and shows attractive potential in tropism redirection via capsid engineering. Here, we showcased that T-cell-tropic rAAV variants successfully enabled CAR expression in human T cells and de-targeted from the liver down to 1% of the parental serotypes. In a humanized mouse model, systematic injection of rAAV-CAR turned up to 8.3% of human T cells into CAR-T cells, which successfully eliminated the number of B cells and controlled tumor progression. Moreover, we identified and verified three cell surface receptors corresponding to the superior transduction efficiencies of the rAAV variants to T cells. This study demonstrated the effectiveness and safety of AAV-based in vivo generation of CAR-T cells in a humanized mouse model, which illustrated a route of in vivo CAR-T therapy via a delivery vector proved in clinics. Furthermore, the identified receptors provide mechanistic insights into the translational potential and guidance to receptor-based capsid engineering.
2026-03-05 | GSE276034 | GEO
Project description:An evolved AAV variant enables efficient genetic engineering of murine T cells
| PRJNA895443 | ENA
Project description: Not available
| S-EPMC11297966 | biostudies-literature
Project description:Introduction: Cellular immunity is one of the main factors affecting sustained expression of gene therapies. AAV capsid-specific CD8+ T cells recognize peptides that are loaded on HLA class I molecules by cells that have been infected or transduced with AAVs. Previous studies performed by others have identified several peptides that activate CD8+ T cells, however the entire repertoire of naturally displayed peptides is unknown.
Methods: Using MHC-associated peptide proteomics, we describe the HLA class I immunopeptidomes of AAV2, AAV6 and AAV9 capsids. Monocyte-derived dendritic cells (MDDCs) were isolated from a panel of 13 healthy donors that have diverse alleles representing more than 50% of the US population. MDDCs were transfected with mRNA encoding for the different AAV capsids. 6 hours post transfection, cells were lysed and peptide:HLA complexes were immunoprecipitated. Peptides were eluted and analyzed by mass spectrometry and PEAKS bioinformatic algorithms.
Results: The HLA-I peptides are distributed along the entire capsids but the number of AAV9-derived peptides was significantly higher than for AAV2 or AAV6. We identified a few peptides that had been described before and are immunogenic. We also observed 14 peptides that are highly conserved among the serotypes. Finally, we observed that more than 60% of the HLA-I peptides are contained within HLA-II clusters.
Discussion: This work is the first comprehensive study identifying the naturally displayed HLA class I peptides derived from the capsid of AAVs and expands on previous studies that have identified immunogenic peptides. The results from this study can be used to generate peptide pools to assess immunogenicity risk of gene therapies in the clinic.
2023-03-01 | MSV000091389 | MassIVE
Project description: Not available
| S-EPMC5872824 | biostudies-literature
Project description: Not available
| S-EPMC10540678 | biostudies-literature
Project description: Not available
| S-EPMC7885987 | biostudies-literature
Project description:Recombinant adeno-associated viral vectors (rAAVs) are among the most commonly used vehicles for in vivo based gene therapies. However, it is hard to predict which AAV capsid will provide the most robust expression in human subjects due to the observed discordance in vector-mediated transduction between species. We used a primate specific capsid, AAV-LK03, and demonstrated that the limitation of this capsid towards transduction of mouse cells was unrelated to cell entry and nuclear transport but rather due to depleted histone H3 chemical modifications related to active transcription, namely H3K4me3 and H3K27ac, on the vector DNA itself. A single-amino acid insertion into the AAV-LK03 capsid enabled efficient transduction and the accumulation of active-related epigenetic marks on the vector chromatin in mouse without compromising transduction efficiency in human cells. Our study suggests that the capsid protein itself is involved in driving the epigenetic status of the vector genome, most likely during the process of uncoating. Programming viral chromatin states by capsid design may enable facile DNA transduction between vector and host species and ultimately led to rationale selection of AAV capsids for use in humans.
2023-02-28 | GSE226167 | GEO