AAV Gene Therapy Vectors

Adeno-Associated Virus (AAV) is the leading viral vector for gene therapy. Safe, effective, but with integration and immune response risks.

Vector Design

class AAVVector:
    """
    AAV vector components:
    - ITRs (Inverted Terminal Repeats): Required for packaging
    - Transgene: Your therapeutic gene
    - Promoter: Tissue-specific expression
    - Capsid: Determines which cells infected
    """

    def __init__(self, transgene, promoter, capsid_serotype):
        self.transgene = transgene  # e.g., "Factor IX" for hemophilia
        self.promoter = promoter    # e.g., "liver-specific"
        self.capsid = capsid_serotype  # AAV8 for liver, AAV9 for CNS

    def design_vector(self):
        """
        AAV genome (max 4.7 kb):
        ITR-Promoter-Transgene-PolyA-ITR

        ⚠️ Size limit: Must fit in 4.7kb including promoter + transgene
        """
        return f"ITR-{self.promoter}-{self.transgene}-PolyA-ITR"

Capsid Engineering

# Different serotypes target different tissues
CAPSID_TROPISM = {
    'AAV1': 'Muscle, heart',
    'AAV2': 'Liver, CNS, retina',
    'AAV5': 'CNS, lung',
    'AAV8': 'Liver (best for hepatic gene therapy)',
    'AAV9': 'CNS, heart (crosses blood-brain barrier)',
    'AAVrh10': 'Broad tropism',
}

def select_capsid(target_tissue):
    """Choose optimal AAV serotype for tissue."""
    if target_tissue == 'liver':
        return 'AAV8'  # 10-100x better liver transduction
    elif target_tissue == 'brain':
        return 'AAV9'  # Crosses BBB after systemic injection
    elif target_tissue == 'muscle':
        return 'AAV1'

Production & Titer

class AAVProduction:
    def produce_aav(self, vector_plasmid, helper_plasmids):
        """
        Triple transfection method:
        1. Vector plasmid (your gene)
        2. AAV rep/cap plasmid (packaging genes)
        3. Adenovirus helper plasmid
        """
        # Transfect HEK293 cells
        cells = HEK293T()
        cells.transfect([vector_plasmid, rep_cap, helper])

        # Harvest after 48-72 hours
        lysate = cells.lyse()

        # Purify AAV
        purified_aav = self.purify_by_iodixanol_gradient(lysate)

        # Titer (measure concentration)
        titer = self.measure_genome_copies(purified_aav)
        return purified_aav, titer

    def measure_genome_copies(self, aav_prep):
        """
        qPCR to quantify AAV genomes.
        Typical: 10^13 - 10^14 vg/mL (genome copies per mL)
        """
        return qpcr_quantification(aav_prep)

Risks ⚠️

def assess_aav_risks(vector):
    risks = []

    # 1. Integration (rare but possible)
    if vector.has_homology_to_genome():
        risks.append("Integration risk at AAVS1 locus")

    # 2. Immune response
    if patient.has_anti_aav_antibodies():
        risks.append("Pre-existing immunity may neutralize vector")

    # 3. Genotoxicity
    if vector.transgene_is_oncogene():
        risks.append("Insertional mutagenesis risk")

    # 4. Off-target transduction
    if not vector.is_tissue_specific():
        risks.append("Non-target tissue expression")

    return risks

Dosing: 10^12 - 10^14 vg/kg (patient weight-based)

Related Chronicles: Synthetic Blood Contamination (2032)

FDA Approved: Luxturna (retina), Zolgensma (SMA)