In-vivo gene therapy: AAV and lentiviral vectors

01Overview and value chain

Markers: [EC: Reg. (EC) No 1394/2007 ATMP & FDA OTP framework | OECD: Biopharma | Regulator: FDA (USA), EMA (EU), NMPA (China)]

In-vivo gene therapy delivers a functional therapeutic gene directly into a patient’s cells using a engineered viral vector, most often an adeno-associated virus (AAV) and, for ex-vivo-modified autologous products, a lentiviral vector. Since Luxturna became the first FDA-approved direct in-vivo gene therapy in December 2017, eight one-time AAV products have reached the US or EU market, including Zolgensma for spinal muscular atrophy at roughly 2.1 million US dollars per dose and Hemgenix for haemophilia B at about 3.5 million dollars — the world’s most expensive medicine at launch. More than 3000 children have received Zolgensma worldwide, and AAV manufacturing titers in suspension HEK293 cultures now routinely exceed 1e14 viral genomes per batch. Yet the field is also honest about its limits: BioMarin voluntarily withdrew Roctavian from the market in 2024 after poor commercial uptake despite durable five-year Phase 3 efficacy, a signal that clinical success and market viability are not the same.

The key directions of in-vivo gene therapy are:

1. AAV serotype matching (AAV Serotype Matching): tissue-tropic capsids route the payload to the right organ — AAV2 for the retina (Luxturna), AAV9 across the blood-brain barrier (Zolgensma), AAV5 to hepatocytes (Hemgenix, Roctavian), AAVrh74 to skeletal and cardiac muscle (Elevidys).
2. Liver-directed haemophilia platforms (Liver-Directed Haemophilia): a single infusion of an AAV5 or AAVrh74 vector encoding factor IX or VIII achieves multi-year clotting-factor expression, replacing lifelong prophylactic infusions (Hemgenix, Beqvez), though Roctavian’s withdrawal tempers the commercial narrative.
3. Lentiviral ex-vivo platforms (Lentiviral Ex-Vivo): gammaretroviral and lentiviral vectors modify a patient’s own haematopoietic stem cells outside the body before re-infusion, the backbone of Bluebird’s Skysona and Zynteglo and Orchard’s Libmeldy for rare neurological and blood disorders.
4. Manufacturing scale-up and capsid yield (Manufacturing Scale-Up): the move from adherent to suspension HEK293 transient transfection, with triple-plasmid systems and affinity chromatography, drives the titer and cost reductions needed to treat millions rather than thousands.

Sectoral value chain

[capsid + payload design] ──> [plasmid & cell bank] ──> [transfection / production] ──> [harvest + lysis]
                                             │
                                  (AAV serotype selection)
                                             │
                                             ▼
[patient dosing] <─── [fill-finish + QC] <─────┘

Value chain levels

Cross-cutting technologies of the sector:

• Transient triple-plasmid transfection (Transient Transfection): rep/cap, helper and transgene plasmids co-transfected into HEK293 suspension cells for helper-virus-free AAV production.
• Capsid engineering (Capsid Engineering): directed evolution and rational design of capsids that evade pre-existing neutralising antibodies and target novel tissues.
• Tissue-specific regulatory cassettes (Tissue-Specific Cassettes): promoters such as LP1 or MHCK7 restrict transgene expression to hepatocytes or muscle, raising safety and expression.

02US

The United States hosts the largest concentration of approved in-vivo gene therapy sponsors and the FDA’s Office of Therapeutic Products (OTP), which has cleared the majority of the world’s commercial AAV treatments.

capsid platforms, DMD leadership, haemophilia launches

• Spark Therapeutics (Luxturna): the 2017 RPE65 retinal therapy continues to lead inherited-retinal gene therapy with growing market penetration as the field’s first direct in-vivo product.
• Sarepta Therapeutics (Elevidys): the AAVrh74 micro-dystrophin vector for Duchenne muscular dystrophy, accelerated in 2023 and expanded to ambulatory and non-ambulatory boys, is now the largest US in-vivo gene-therapy franchise by revenue, reported in the Q1 2026 results.
• Pfizer (Beqvez): FDA- and Health Canada-approved fidanacogene elaparvovec for haemophilia B in 2024, adding a second one-time AAV haemophilia option alongside a candid commercial caution from the Roctavian withdrawal.

03CN

China’s in-vivo gene therapy is moving from CAR-T-adjacent lentiviral work into AAV-directed platforms, with NMPA pathways opening for domestic capsid engineering and base-editing programmes.

AAV base editing, lentiviral heritage, NMPA pathways

• YolTech Pharmaceuticals (尧唐生物): a Shanghai in-vivo gene-editing company founded in 2021, building AAV-delivered base-editor platforms for cardiovascular and metabolic targets, backed by a more than 300 million yuan B round led by the AstraZeneca-CICC healthcare fund.
• Lentiviral heritage: China’s earlier approved gene therapies lean on ex-vivo lentiviral CAR-T platforms, providing the GMP and regulatory experience now flowing into AAV in-vivo candidates.
• NMPA acceleration: the Centre for Drug Evaluation is opening priority review channels for AAV treatments for rare diseases, tracking the FDA OTP model.

04EU

Europe authored the field’s regulatory backbone through the ATMP Regulation (EC) No 1394/2007 and hosts both the haemophilia-B originator and the CNS gene- therapy franchise.

SMA franchise, haemophilia origin, ATMP regulation

• Novartis (Zolgensma): the AAV9 onasemnogene abeparvovec for spinal muscular atrophy, with full-year 2025 and Q2 2026 sales reaffirming its place as the world’s most widely administered one-time in-vivo gene therapy, delivered across Swiss-based global manufacturing.
• uniQure (Hemgenix): the Dutch originator of the AAV5 etranacogene dezaparvovec, whose five-year durability data published in the New England Journal of Medicine underpins the CSL-marketed haemophilia B therapy and the first patients treated commercially.
• ATMP framework: EMA’s Committee for Advanced Therapies (CAT) provides the centralized authorisation route that licensed Zolgensma and Hemgenix across the bloc, predating several US approvals.

05Leading companies and research institutes

06Tech stack and innovations

The in-vivo gene therapy stack rests on vector engineering, scalable production and immune management, each hardened by a decade of post-Luxturna learning.

1. AAV capsid selection (AAV Capsid Selection):
   • serotype-tissue matching dictates efficacy: AAV2 for retina, AAV9 for CNS and systemic paediatric delivery, AAV5 and AAVrh74 for liver and muscle.
   • case: Zolgensma’s AAV9 crosses the blood-brain barrier to reach spinal motor neurons in SMA type 1.
2. Suspension transient transfection (Suspension Transient Transfection):
   • triple-plasmid (rep/cap + helper + transgene) co-transfection in serum-free suspension HEK293 replaces adherent production for scale.
   • affinity chromatography (e.g. AVB Sepharose) plus anion-exchange polishing lifts batch titers above 1e14 viral genomes.
3. Immune and expression engineering (Immune & Expression Engineering):
   • engineered and selected capsids evade pre-existing neutralising antibodies that would otherwise exclude up to 60 percent of adults from systemic AAV dosing.
   • tissue-specific promoters (LP1, MHCK7) confine transgene expression to hepatocytes or muscle, lifting both safety and expression durability.

07Value chains and production pipelines

Industrial pipeline of an AAV drug product (FDA OTP / EU ATMP GMP)

┌───────────────────────────┐      ┌───────────────────────────┐
│ 1. Capsid + cassette design│ ───> │ 2. Plasmid & cell bank    │
└───────────────────────────┘      └───────────────────────────┘
                                                  │
                                                  ▼
┌───────────────────────────┐      ┌───────────────────────────┐
│ 4. Purification           │ <─── │ 3. Transfection + harvest │
└───────────────────────────┘      └───────────────────────────┘
              │
              ▼
┌───────────────────────────┐      ┌───────────────────────────┐
│ 5. Fill-finish + QC       │ ───> │ 6. Clinical dosing        │
└───────────────────────────┘      └───────────────────────────┘

Stage 1: Capsid and cassette design

A serotype is matched to the target tissue and a transgene cassette with a tissue-specific promoter is assembled in silico, then validated in rodent and large-animal models.

Stage 2: Plasmid and cell bank

GMP-grade rep/cap, helper and transgene plasmids are amplified in bacterial fermentation and a qualified HEK293 master cell bank is established and characterised for adventitious agents.

Stage 3: Transfection and harvest

Triple-plasmid transfection of suspension HEK293 cells in stirred-tank bioreactors produces crude AAV, which is harvested by chemical lysis and nuclease digestion to release intracellular capsids.

Stage 4: Purification

Affinity chromatography captures AAV capsids, followed by anion-exchange polishing and diafiltration to remove empty capsids, host-cell protein and DNA, reaching purity above 95 percent full capsids in modern processes.

Stage 5: Fill-finish and QC

The purified vector is formulated, sterile-filtered, filled into cryo-vials and released against potency, identity, residual host-cell and empty-capsid specifications under FDA OTP and EU ATMP GMP.

Stage 6: Clinical dosing

A one-time intravenous, subretinal or intrathecal infusion delivers the vector; patients are monitored for neutralising-antibody status and transgene expression, with multi-year durability now demonstrated for Luxturna, Zolgensma and Hemgenix.