Muscle-Directed Delivery of an AAV1 Vector Leads to Capsid-Specific T Cell Exhaustion in Nonhuman Primates and Humans

Abstract

With the US Food and Drug Administration (FDA) and European Medicines Agency (EMA) approvals for Zolgensma, Luxturna, and Glybera, recombinant adeno-associated viruses (rAAVs) are considered efficient tools for gene transfer. However, studies in animals and humans demonstrate that intramuscular (IM) AAV delivery can trigger immune responses to AAV capsids and/or transgenes. IM delivery of rAAV1 in humans has also been described to induce tolerance to rAAV characterized by the presence of capsid-specific regulatory T cells (Tregs) in periphery. To understand mechanisms responsible for tolerance and parameters involved, we tested 3 muscledirected administration routes in rhesus monkeys: IM delivery, venous limb perfusion, and the intra-arterial push and dwell method. These 3 methods were well tolerated and led to transgene expression. Interestingly, gene transfer in muscle led to Tregs and exhausted T cell infiltrates in situ at both day 21 and day 60 post-injection. In human samples, an in-depth analysis of the functionality of these cells demonstrates that capsid-specific exhausted T cells are detected after at least 5 years post-vector delivery and that the exhaustion can be reversed by blocking the checkpoint pathway. Overall, our study shows that persisting transgene expression after gene transfer in muscle is mediated by Tregs and exhausted T cells.

Intramuscular injection of vectorized-scFvMC1 reduces pathological tau in two different tau transgenic models

“…Depending on the administration route, the dose, and/or the AAV serotype, gene transfer can result in a deleterious immune response [ 89 , 90 ] or tolerance [ 61 , 91 , 92 ]. Two studies have shown that AAV1 IM injection can lead to regulatory T cells (Tregs) infiltration in injected muscles in AATD (Alpha-1 Antitrypsin Deficiency) and LPLD (Lipoprotein Lipase Deficiency) patients [ 61 , 91 ].…”

“…Two studies have shown that AAV1 IM injection can lead to regulatory T cells (Tregs) infiltration in injected muscles in AATD (Alpha-1 Antitrypsin Deficiency) and LPLD (Lipoprotein Lipase Deficiency) patients [ 61 , 91 ]. Gernoux et al [ 92 ] have recently correlated results obtained in monkeys to those obtained with patients and further demonstrated that tolerance and persisting transgene expression after AAV1 gene transfer in muscle is mediated by Tregs and exhausted T cells. These findings support the IM approach proposed in the present study and its translational potential for humans.…”

T Cell-Mediated Immune Responses to AAV and AAV Vectors

“…A substantial portion of the muscle infiltrating lymphocytes were regulatory T cells suggesting that they had suppressed vector-induced effector T cell responses and thereby prevented loss of AAV-transduced muscle cells (87,88). A further follow-up study conducted 5 years after gene transfer detected Tregs and AAV capsid-specific CD8 + T cells within the injected muscle (89).…”

AAV8 locoregional delivery induces long-term expression of an immunogenic transgene in macaques despite persisting local inflammation

“…37,38 Detection of regulatory T cells in the perfused limb A recent study suggested that, in animal models, the characterization of muscle-infiltrated cells following rAAV delivery would be more relevant than the analysis of peripheral immune response to understand the absence of a deleterious T cell response. 39 Therefore, we performed a hematoxylin-phloxin-saffron (HPS) staining on perfused and contralateral muscles sections to analyze the presence of mononuclear cell infiltrates. Muscle-infiltrated cells were detected in all monkeys up to 12 months post-injection in perfused limb (Figure 3A), but not in contralateral limb (Figure S2).…”

“…Two cell populations responsible for the absence of a deleterious immune response have been described in AAV gene transfer: regulatory T cells (CD3+CD4+FoxP3+ cells) and exhausted T cells (CD3+CD8+PD-1+). [39][40][41] In this study, we performed an immunostaining on muscle collected at necropsy to verify the presence of these cells in the injected limb to explain long-term GFP expression. Infiltrated regulatory T cells (CD4+Foxp3+ cells; Figure 4) but no exhausted T cells (CD3+PD1+ cells; Figure S4) were detected in situ after 1 year post-injection in both desmin and CMV groups.…”