Table 2.

Investigative strategies to mitigate the effect of pre-existing anti-AAV antibodies in patients

Potential approach: rationale	Considerations and examples
GTx-related approaches
Direct delivery to target organ: minimize exposure to NAbs	•Minimizes systemic exposure to the vector43,47,115,116,118
	•This option is not suitable for all target tissues, and some administration routes may elicit strong T cell responses against the transgene115
	•Example: Subretinal delivery of AAV2-hRPE65v2 in five patients with Leber congenital amaurosis type 2 resulted in improvement in visual acuity sustained for 3 years115
	•Example: Saline flushing of the liver followed by injection into the portal vein allowed efficient human FIX transduction by AAV8 in non-human primates with NAb titer ≤1:56118
Administer high dose: overcome NAbs	•May be effective in the presence of low-titer (1:5–1:17) NAbs93,96
	•The higher vector load could trigger an anti-capsid T cell response96,136, which is a potential safety risk, particularly in the setting of pre-existing humoral response
	•Example: Tenfold increase in vector dose partially overcame low-level pre-existing antibodies to AAV in mice models137
Administer empty capsids: adsorb anti-AAV antibodies	•Advantages include the lack of pharmacological intervention and the ease of manufacturing empty capsids (as a by-product of AAV vector production)116
	•May lead to greater viral load, potentially triggering an anti-capsid T cell response102, particularly in the setting of pre-existing humoral response
	•Example: Adding empty capsids dose-dependently to adsorb anti-AAV antibodies, successfully facilitating transduction in murine and non-human primate models – even at high titers. Moreover, mutated capsids were reported to adsorb antibodies without entering the target cell117
Modify capsid/switch AAV serotype or engineering/cloaking of AAV: reduce capsid susceptibility to NAbs	•Coupled immunomodulation via cloaking is technically feasible, although such a strategy remains challenging in terms of engineering and manufacturing the capsid115,119
	•Engineering capsid could deleteriously modify tissue tropism and increase cross-reactivity among AAV serotypes47,116,138
	•Example: Engineered AAV vectors elicited reduced immune responses and enhanced gene expression across different tissues, including liver, muscle and retina in clinically relevant mouse and pig models119
Pre-treatment approaches
Immunosuppressive drugs: prevent/eradicate humoral immune response to AAV	•Several B and T cell depletion therapies are available120,139
	•Potential risks are associated with incomplete eradication of pre-existing high-titer NAbs with systemic immunosuppression, such as rituximab116,140
	•Example: Rituximab reduced anti-AAV NAb titers in rheumatoid arthritis patients with titers ≤1:1000, although only a minority dropped to <1:516
	•Example: In a patient with Pompe disease, treatment with rituximab and sirolimus mitigated an immune response to an AAV1-GAA vector, which may allow for repeat administration in the future121
Plasmapheresis: selective depletion of anti-AAV IgGs	•Has been shown to reduce antibody titers64,122
	•Potential risks are associated with incomplete eradication of pre-existing high-titer NAbs123
	•Example: Frequent sessions of plasmapheresis resulted in reduction of NAbs specific for AAV1, 2, 6, and 8 to undetectable levels or titers <1:5 in seropositive patients (mainly when initial titers were ≤1:20)123
IgG cleaving enzymes from certain Streptococcus species: inhibiting an IgG immune response	•IdeS (from Streptococcus pyogenes) treatment has thus far been shown to have a favorable safety profile in solid organ transplant recipients141
	•Has no impact on IgA, IgM, IgD, and IgE antibodies142
	•Potential alternatives include IdeZ from Streptococcus equi subsp. zooepidemicus125
	•Example: IdeS treatment before rAAV vector infusion resulted in enhanced liver transduction in non-human primates, even in the setting of vector re-administration, and reduced anti-AAV antibody levels from human plasma samples in vitro, including plasma from prospective GTx trial participants undergoing GTx for Crigler–Najjar syndrome124
Anti-FcRn antibodies: reduce IgG levels	•FcRn1 helps maintain circulating IgG levels, and its inhibition can potentially reduce NAb titers by ∼80% for 60 days126
	•Has no impact on IgA, IgM, IgD, and IgE antibodies143
	•Example: Anti-FcRn antibodies lowered and maintained the reduction of total IgG and IgG subclasses following multiple subcutaneous injections in healthy adults127

AAV, adeno-associated virus; FcRn, neonatal crystallizable fragment receptor; GAA, acid α-glucosidase; GTx, gene therapy; IdeS, IgG-degrading enzyme of Streptococcus pyogenes; IgA, immunoglobulin A; IgD, immunoglobulin D; IgE, immunoglobulin E; IgG, immunoglobulin G; IgM, immunoglobulin M; NAb, neutralizing antibody.