Immune transgene-dependent myocarditis in macaques after systemic administration of adeno-associated virus expressing human acid alpha-glucosidase

Immune responses to human non-self transgenes can present challenges in preclinical studies of adeno-associated virus (AAV) gene therapy candidates in nonhuman primates. Although anti-transgene immune responses are usually mild and non-adverse, they can confound pharmacological readouts and complicate translation of results between species. We developed a gene therapy candidate for Pompe disease consisting of AAVhu68, a clade F AAV closely related to AAV9, that expresses an engineered human acid-alpha glucosidase (hGAA) tagged with an insulin-like growth factor 2 variant (vIGF2) peptide for enhanced cell uptake. Rhesus macaques were administered an intravenous dose of 1x10(13) genome copies (GC)/kg, 5x10(13) GC/kg, or 1 x 10(14) GC/kg of AAVhu68.vIGF2.hGAA. Some unusually severe adaptive immune responses to hGAA presented, albeit with a high degree of variability between animals. Anti-hGAA responses ranged from absent to severe cytotoxic T-cell-mediated myocarditis with elevated troponin I levels. Cardiac toxicity was not dose dependent and affected five out of eleven animals. Upon further investigation, we identified an association between toxicity and a major histocompatibility complex class I haplotype (Mamu-A002.01) in three of these animals. An immunodominant peptide located in the C-terminal region of hGAA was subsequently identified via enzyme-linked immunospot epitope mapping. Another notable observation in this preclinical safety study cohort pertained to the achievement of robust and safe gene transfer upon intravenous administration of 5x10(13) GC/kg in one animal with a low pre-existing neutralizing anti-capsid antibodies titer (1:20). Collectively, these findings may have significant implications for gene therapy inclusion criteria.

Automated summary

Immune responses to human non-self transgenes in adeno-associated virus (AAV) gene therapy candidates in nonhuman primates can be challenging. A gene therapy candidate for Pompe disease consisting of AAVhu68 was developed, and some severe adaptive immune responses to the therapy were observed, with variability between animals. Cardiac toxicity was found to be associated with a specific haplotype, and an immunodominant peptide was identified as a potential cause. Additionally, robust and safe gene transfer was achieved in an animal with low pre-existing neutralizing antibodies.