Transduction Efficiency and Immune Response Associated With the Administration of AAV8 Vector Into Dog Skeletal Muscle

Abstract

Recombinant adeno-associated virus (rAAV)-mediated gene transfer is an attractive approach to the treatment of Duchenne muscular dystrophy (DMD). We investigated the muscle transduction profiles and immune responses associated with the administration of rAAV2 and rAAV8 in normal and canine X-linked muscular dystrophy in Japan (CXMD(J)) dogs. rAAV2 or rAAV8 encoding the lacZ gene was injected into the skeletal muscles of normal dogs. Two weeks after the injection, we detected a larger number of beta-galactosidase-positive fibers in rAAV8-transduced canine skeletal muscle than in rAAV2-transduced muscle. Although immunohistochemical analysis using anti-CD4 and anti-CD8 antibodies revealed less T-cell response to rAAV8 than to rAAV2, beta-galactosidase expression in rAAV8-injected muscle lasted for <4 weeks with intramuscular transduction. Canine bone marrow-derived dendritic cells (DCs) were activated by both rAAV2 and rAAV8, implying that innate immunity might be involved in both cases. Intravenous administration of rAAV8-lacZ into the hind limb in normal dogs and rAAV8-microdystrophin into the hind limb in CXMD(J) dogs resulted in improved transgene expression in the skeletal muscles lasting over a period of 8 weeks, but with a declining trend. The limb perfusion transduction protocol with adequate immune modulation would further enhance the rAAV8-mediated transduction strategy and lead to therapeutic benefits in DMD gene therapy.

Efficient transduction of non-human primate motor neurons after intramuscular delivery of recombinant AAV serotype 6

“…Acute immune responses and elimination of transduced myofibers after the delivery of rAAV to skeletal muscle has been observed in canine models of muscular dystrophy. [14][15][16] Likewise, T cell-mediated immune responses have previously been observed after rAAV2/2 delivery to non-human primate muscle. 17 This immune response to rAAV in muscle can be circumvented through longterm immunosuppression in cynomolgus macaques.…”

“…As host immune responses have proven an important scale up issue from rodents to larger animals, [14][15][16] we have determined whether administration of the rAAV2/6 vector mediated cellular toxicity or an immune response in the monkeys. Infiltrating cells were observed in the injected muscle and corresponded to recruitment of both macrophages and T cells (Figure 4a and b).…”

Systemic human Netrin-1 gene delivery by adeno-associated virus type 8 alters leukocyte accumulation and atherogenesis in vivo

“…Animal preparation was carried out as described earlier. 35,36 Briefly, each mouse was anesthetized using 1.5% isoflurane (Isothesia; Abbot Laboratories, Chicago, IL, USA) with oxygen and laid supine on a thermostatically heated platform with all legs taped to electrocardiography electrodes for cardiac function monitoring. Abdominal hair was removed with a shaver and a chemical hair remover (Church & Dwight Co., Inc., NJ, USA) and a pre-warmed US gel (Medline Industries Inc., Mundelein, IL, USA) was spread over the skin as a coupling medium for the transducer.…”

Improvement of cardiac fibrosis in dystrophic mice by rAAV9-mediated microdystrophin transduction

“…12,13 To achieve transduction of an entire extremity, a limb perfusion by retrograde intravenous administration of the rAAV in larger animal models is being developed. 14 For cardiac transduction, we utilized rAAV serotype 9, which can be administered systemically with excellent cardiac tropism. 15 In this study, we used mdx mice which are considered to be a representative animal model of DMD.…”