Spatial transcriptomics and single-nucleus RNA sequencing reveal rAAV2- and rAAV9-specific transduction signatures in the mouse liver.

DOI
Publisher DOI
PubMed ID
41813829
Abstract
The liver is a primary target for recombinant adeno-associated viral (rAAV) vectors, yet the influence of serotype, sex, and liver zonation on transduction and transcriptomic changes remains incompletely understood. This proof-of-concept study employs spatial transcriptomics alongside single-nucleus RNA sequencing to map the spatial distribution and impacts of rAAV2- and rAAV9-CMV-EGFP vectors in male and female mouse livers. Spatial transcriptomics provided precise transgene mapping and highlighted that CMV-EGFP rAAV vectors deregulate hepatocellular lipid metabolism, the circadian clock, and the immune/stress response with sex specific differences. Lipid metabolism genes (Elovl3, Chka, Irs2, Ppard), were consistently deregulated across all zones of the liver lobule in male and female rAAV2- and rAAV9-CMV-EGFP-treated mice, while Srebf1, Tlcd4, Cpt2, and Acot1 exhibited sex-specific patterns. Circadian clock modulators (Dbp, Tef, Arntl, Nfil3, Nr1d1/Nr1d2) were altered independently of zonation. The study found sex-specific downregulation of immune and stress-response genes and pathways, including Gadd45g and hypoxia pathways. TGF-β and EGFR pathways were upregulated sex-independently. Spatial transcriptomics further enabled examination of transgene and rAAV entry factor co-expression, identifying known and novel factors like Rpsa, Dpp4, Sdc1, and solute carrier proteins, highlighting its role in supporting targeted screening. Our findings demonstrate spatial transcriptomics as a powerful tool in gene therapy research and reveal novel rAAV vector effects on liver biology.
Date Issued
2026-05
Publication Type
Article
Subject(s)
Language(s)
en
Author(s)
Köchl, Fabian
Kumpesa, Nadine
Prasad, Megana
Bochner, Filip
Hernández-Obiols, Mar
Otteneder, Michael B
Stalder, Lucas
Shaffo, Frances
Nowrouzi, Ali
Markusic, David
Haegel, Hélène
Xicluna, Rebecca
Sultan, Marc
Jacobsen, Björn
Institute of Animal Pathology, Laboratory Cancer Therapy Escape I
Valdeolivas, Alberto
Schwalie, Petra C
Additional Credits
Institute of Animal Pathology, Laboratory Cancer Therapy Escape I
Journal
Gene Therapy
Publisher
Springer Nature [academic journals on nature.com]
ISSN
1476-5462
0969-7128
Access(Rights)
open.access
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