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categories: ["genetics", "bodies"]categories: ["genetics", "bodies"]tags: ["gene therapy", "delivery", "viral vectors", "immunology", "manufacturing", "rare disease"]tags: ["gene therapy", "delivery", "viral vectors", "immunology", "manufacturing", "rare disease"]summary: "Engineered adeno-associated viruses used to carry therapeutic genes into human cells, the workhorse of in vivo gene therapy and the source of most of its limits."summary: "Engineered adeno-associated viruses used to carry therapeutic genes into human cells, the workhorse of in vivo gene therapy and the source of most of its limits."updated: "2026-07-27"updated: "2026-10-29"humanEvidence: "Licensed AAV products treat inherited retinal dystrophy, spinal muscular atrophy and haemophilia in people; the same clinical record shows capsid immunity blocking redosing and fatal liver injury at high systemic doses."humanEvidence: "Licensed AAV products treat inherited retinal dystrophy, spinal muscular atrophy and haemophilia in people; the same clinical record shows capsid immunity blocking redosing and fatal liver injury at high systemic doses."access: "Obtainable only as specific licensed gene therapies given in specialist centres, at one-time list prices in the millions of dollars; patients with pre-existing anti-capsid antibodies are screened out."access: "Obtainable only as specific licensed gene therapies given in specialist centres, at one-time list prices in the millions of dollars; patients with pre-existing anti-capsid antibodies are screened out."reversibility: "context"reversibility: "context"lines 35–41 → 35–4120 unchanged lines not shown
## How it works## How it works Wild-type AAV carries about 4.7 kilobases of single-stranded DNA between two inverted terminal repeats, encoding two gene cassettes — *rep*, for replication, and *cap*, for the capsid. It cannot complete its own life cycle without a helper virus such as adenovirus or herpes simplex, which is why it was first noticed in 1965 as a contaminant in adenovirus preparations rather than as a pathogen in its own right.Wild-type AAV carries about 4.7 kilobases of single-stranded DNA between two inverted terminal repeats, encoding two gene cassettes: *rep*, for replication, and *cap*, for the capsid. It cannot complete its own life cycle without a helper virus such as adenovirus or herpes simplex, which is why it was first noticed in 1965 as a contaminant in adenovirus preparations rather than as a pathogen in its own right. To make a vector, everything between the terminal repeats is deleted and replaced with the therapeutic cassette: a promoter, the gene, a polyadenylation signal. The *rep* and *cap* functions are supplied separately during manufacturing, so the resulting particle can enter a cell and deliver its cargo but cannot replicate. Once inside, the single-stranded genome is converted to double-stranded DNA and circularises into episomes that persist in the nucleus, transcribing for years in non-dividing tissue. A small fraction integrates at semi-random sites; wild-type AAV's preference for the AAVS1 locus depends on Rep protein, which vectors lack.To make a vector, everything between the terminal repeats is deleted and replaced with the therapeutic cassette: a promoter, the gene, a polyadenylation signal. The *rep* and *cap* functions are supplied separately during manufacturing, so the resulting particle can enter a cell and deliver its cargo but cannot replicate. Once inside, the single-stranded genome is converted to double-stranded DNA and circularises into episomes that persist in the nucleus, transcribing for years in non-dividing tissue. A small fraction integrates at semi-random sites; wild-type AAV's preference for the AAVS1 locus depends on Rep protein, which vectors lack. lines 47–53 → 47–535 unchanged lines not shown
The natural AAV serotypes differ chiefly in their capsid surface, and therefore in which tissues they enter. AAV2 was the first characterised and remains common in the eye. AAV8 and AAV5 favour the liver. AAV9 crosses the blood-brain barrier in newborns well enough to underpin an approved systemic therapy for spinal muscular atrophy. AAV1, AAV6, and AAVrh74 are used for muscle.The natural AAV serotypes differ chiefly in their capsid surface, and therefore in which tissues they enter. AAV2 was the first characterised and remains common in the eye. AAV8 and AAV5 favour the liver. AAV9 crosses the blood-brain barrier in newborns well enough to underpin an approved systemic therapy for spinal muscular atrophy. AAV1, AAV6, and AAVrh74 are used for muscle. Capsid engineering aims to do better than nature. Directed evolution, DNA shuffling, and peptide-display libraries have produced capsids with sharply improved central nervous system delivery in mice.[^deverman2016] Machine-learning models fitted to library-screening data now propose viable capsid variants directly, an application of [[ai-protein-design]] to the delivery problem rather than to the therapeutic protein. The best-known cautionary tale is AAV-PHP.B, which crosses the murine blood-brain barrier with startling efficiency and then turned out to depend on a receptor, Ly6a, present in some inbred mouse strains and absent in primates.[^hordeaux2018] Its failure to translate is the standard example of why capsid performance in one species predicts little about another — the same species-gap problem that recurs throughout [[targeted-drug-delivery]].Capsid engineering aims to do better than nature. Directed evolution, DNA shuffling, and peptide-display libraries have produced capsids with sharply improved central nervous system delivery in mice.[^deverman2016] Machine-learning models fitted to library-screening data now propose viable capsid variants directly, an application of [[ai-protein-design]] to the delivery problem rather than to the therapeutic protein. The best-known cautionary tale is AAV-PHP.B, which crosses the murine blood-brain barrier with unusually high efficiency and then turned out to depend on a receptor, Ly6a, present in some inbred mouse strains and absent in primates.[^hordeaux2018] Its failure to translate is the standard example of why capsid performance in one species predicts little about another, the same species-gap problem that recurs throughout [[targeted-drug-delivery]]. > [!caution] Mouse capsids are not human capsids> [!caution] Mouse capsids are not human capsids> Every claim about a new capsid's tropism should be read with the question: in which species, and does the receptor it exploits exist in people? Several widely cited enhanced capsids do not work in non-human primates at all.> Every claim about a new capsid's tropism should be read with the question: in which species, and does the receptor it exploits exist in people? Several widely cited enhanced capsids do not work in non-human primates at all.lines 59–75 → 59–755 unchanged lines not shown
{ "year": "1965", "title": "Discovery", "text": "AAV is identified as a small contaminating particle in adenovirus preparations and shown to require a helper virus." }, { "year": "1965", "title": "Discovery", "text": "AAV is identified as a small contaminating particle in adenovirus preparations and shown to require a helper virus." }, { "year": "1984", "title": "First recombinant vector", "text": "Researchers demonstrate that the viral genes can be removed and replaced with a transgene between the inverted terminal repeats." }, { "year": "1984", "title": "First recombinant vector", "text": "Researchers demonstrate that the viral genes can be removed and replaced with a transgene between the inverted terminal repeats." }, { "year": "2006", "title": "The immune barrier appears", "text": "In a haemophilia B trial, factor IX expression rises then falls as capsid-specific T cells destroy transduced hepatocytes, defining the field's central obstacle." }, { "year": "2006", "title": "The immune barrier appears", "text": "In a haemophilia B trial, factor IX expression rises then falls as capsid-specific T cells destroy transduced hepatocytes, defining the field's central obstacle." }, { "year": "2012–2017", "title": "First approvals", "text": "Glybera is licensed in Europe and later withdrawn; Luxturna is approved in the United States for RPE65 retinal dystrophy." }, { "year": "2012-2017", "title": "First approvals", "text": "Glybera is licensed in Europe and later withdrawn; Luxturna is approved in the United States for RPE65 retinal dystrophy." }, { "year": "2019", "title": "Systemic dosing at scale", "text": "Zolgensma is approved for spinal muscular atrophy, delivering AAV9 intravenously at doses orders of magnitude above earlier local injections." }, { "year": "2019", "title": "Systemic dosing at scale", "text": "Zolgensma is approved for spinal muscular atrophy, delivering AAV9 intravenously at doses orders of magnitude above earlier local injections." }, { "year": "2020–2025", "title": "Dose-limiting toxicity", "text": "Deaths from acute liver failure and thrombotic microangiopathy in high-dose systemic trials, including for X-linked myotubular myopathy and Duchenne muscular dystrophy, force reassessment of dose ceilings." } { "year": "2020-2025", "title": "Dose-limiting toxicity", "text": "Deaths from acute liver failure and thrombotic microangiopathy in high-dose systemic trials, including for X-linked myotubular myopathy and Duchenne muscular dystrophy, force reassessment of dose ceilings." }]]`````` ## Limitations## Limitations **Cargo.** The 4.7 kb ceiling excludes many important genes outright — dystrophin, most [[crispr-cas9]] constructs once guides and regulatory elements are added, and the still larger dCas9 fusions used for [[epigenome-editing]]. Workarounds include micro-genes such as the truncated dystrophins used in Duchenne programmes, dual-vector systems that reconstitute a protein from two halves via split inteins, and pairing AAV with smaller nucleases. Each adds inefficiency. Muscle-directed constructs expressing follistatin, discussed under [[myostatin-inhibition]], are small enough to fit and have been used in both clinical and unsanctioned settings.**Cargo.** The 4.7 kb ceiling excludes many important genes outright: dystrophin, most [[crispr-cas9]] constructs once guides and regulatory elements are added, and the still larger dCas9 fusions used for [[epigenome-editing]]. Workarounds include micro-genes such as the truncated dystrophins used in Duchenne programmes, dual-vector systems that reconstitute a protein from two halves via split inteins, and pairing AAV with smaller nucleases. Each adds inefficiency. Muscle-directed constructs expressing follistatin, discussed under [[myostatin-inhibition]], are small enough to fit and have been used in both clinical and unsanctioned settings. **Pre-existing immunity.** Because AAV circulates naturally, a substantial fraction of adults — depending on serotype and region, roughly a third to more than half — carry neutralising antibodies that inactivate the vector before it reaches its target. Those patients are screened out of trials, which distorts both eligibility and evidence.**Pre-existing immunity.** Because AAV circulates naturally, a substantial fraction of adults, depending on serotype and region roughly a third to more than half, carry neutralising antibodies that inactivate the vector before it reaches its target. Those patients are screened out of trials, which distorts both eligibility and evidence. **Redosing.** A treated patient develops high antibody titres against the capsid, so a second dose of the same serotype is ineffective. Switching serotypes helps only partially because of cross-reactivity. Strategies under investigation include plasmapheresis, IgG-cleaving bacterial proteases, and B-cell-depleting or tolerising regimens; none is routine practice as of 2026.**Redosing.** A treated patient develops high antibody titres against the capsid, so a second dose of the same serotype is ineffective. Switching serotypes helps only partially because of cross-reactivity. Strategies under investigation include plasmapheresis, IgG-cleaving bacterial proteases, and B-cell-depleting or tolerising regimens; none is routine practice as of 2026. lines 100–106 → 100–10624 unchanged lines not shown
## Outlook## Outlook AAV is unlikely to be displaced soon for durable gene addition to non-dividing tissue: nothing else delivers a gene to photoreceptors or motor neurons and keeps it expressing for years. Its share of new programmes is nonetheless shrinking, because [[lipid-nanoparticles]] are better suited to the hit-and-run logic of genome editing, can be redosed, and are cheaper to make. The most consequential open questions are whether capsid engineering can produce a vector that reaches muscle or brain at doses low enough to avoid complement activation, and whether any of the antibody-clearing strategies makes redosing routine. A negative answer to both would leave AAV where it is now — extraordinarily effective for a small number of tissues and diseases, and a poor fit for anything requiring a second dose or a large payload. Delivery, not editing chemistry, has been the binding constraint on genetic medicine for thirty years, and it remains one for both [[casgevy|ex vivo cell products]] and any future application to [[gene-therapy-for-aging]].AAV is unlikely to be displaced soon for durable gene addition to non-dividing tissue: nothing else delivers a gene to photoreceptors or motor neurons and keeps it expressing for years. Its share of new programmes is nonetheless shrinking, because [[lipid-nanoparticles]] are better suited to the hit-and-run logic of genome editing, can be redosed, and are cheaper to make. The most consequential open questions are whether capsid engineering can produce a vector that reaches muscle or brain at doses low enough to avoid complement activation, and whether any of the antibody-clearing strategies makes redosing routine. A negative answer to both would leave AAV where it is now: extraordinarily effective for a small number of tissues and diseases, and a poor fit for anything requiring a second dose or a large payload. Delivery rather than editing chemistry has been the binding constraint on genetic medicine for thirty years, and it remains one for both [[casgevy|ex vivo cell products]] and any future application to [[gene-therapy-for-aging]]. ## See also## See also removed, struck through added, underlinedLine numbers count the serialised markdown of each revision, frontmatter included.
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