Sustained Interleukin-10 Transgene Expression Following Intra-Articular AAV5-IL-10 Administration to Horses

Abstract

Joint trauma leads to post-traumatic inflammation with upregulation of inflammatory cytokines and degradative enzymes. If severe enough, this response can lead to irreversible post-traumatic osteoarthritis. Interleukin-10 (IL-10), a cytokine with potent anti-inflammatory effects, has been shown to have chondroprotective effects. A gene therapy approach using a vector to overexpress IL-10 in the joint represents a feasible method of delivering sustained high doses of IL-10 to post-traumatic joints. We hypothesized that an AAV5 vector overexpressing IL-10 would result in rapid and sustained IL-10 expression following direct intra-articular injection and that this increase would not be reflected in systemic circulation. In addition, we hypothesized that intra-articular AAV5-IL-10 injection would not induce a local inflammatory response. Twelve horses were assigned to either treatment (AAV5-IL-10-injected) or control (PBS-injected) groups. Middle carpal joints were injected with 10 12 vector genomes/joint or phosphatebuffered saline (PBS) alone (3 mL). Serial synovial fluid samples were analyzed for inflammatory changes, IL-10 concentration, and vector genome copy number. Serum samples were also analyzed for IL-10 concentration and vector genome copy number. Synovial membrane was collected on day 84. Synovial fluid IL-10 was significantly increased within 48 h of AAV5-IL-10 injection and remained increased, compared to PBS-injected joints, until day 84. Serum IL-10 was not different between groups. Vector administration did not cause a significant synovial inflammatory response. Vector genomes were detectable in the plasma, synovial fluid, and synovial membrane of AAV5-IL-10-injected horses only. IL-10 has the potential to modulate the articular inflammatory response, thereby protecting cartilage from degradation and osteoarthritis. This study demonstrates the feasibility and efficiency of intra-articular AAV5-IL-10, and future studies investigating the chondroprotective effects of IL-10 in inflamed joints in vivo are warranted.

Detection of intra‐articular gene therapy in horses using quantitative real time PCR in synovial fluid and plasma

“…Some of the experimental procedures described below can be found in more detail in a published paper by Moss et al 21 …”

“…A rAAV‐IL10 viral vector was generated by the University of North Carolina Vector Core as described previously 21 . Briefly, the vector consisted of an equine IL‐10 transgene, flanked by AAV2 inverted terminal repeats (ITRs) and under control of a cytomegalovirus (CMV) promoter (Figure 1) incorporated into the AAV5 viral capsule.…”

“…There are many serotypes of AAV and appropriate rAAV vectors are chosen for specific applications based on tissue tropism and transduction efficiency 13–16 . In horses, rAAVs have predominantly been used for intra‐articular delivery of anti‐inflammatory or anabolic genes such as IL1‐RA, 14,15,17,18 IGF‐1, 19,20 and most recently, IL‐10 21 …”

Interleukin-17 pathway activation inEquus caballussupporting limb laminitis

“…Inhibiting IL-17 signaling in the horse will not be as simple as applying human therapies since the most successful psoriasis treatments use monoclonal antibodies to block IL- 17 receptor activation [21]; these antibodies may not bind the equine IL-17 receptor, may be antigenic in equines, and are likely cost-prohibitive. Local inhibition of IL-17 and/or mTORC1 would be desirable, and could be possible using an adeno-associated viral vector to deliver genes for inhibitory cytokines to reduce inflammation, as demonstrated recently [83], or other genes able to inhibit IL-17 and/or mTORC1 activation.…”

Microfluidic Quantitative PCR Detection of 12 Transgenes from Horse Plasma for Gene Doping Control

“…Recent advances in veterinary medical technologies have enabled gene therapy in horses, and several clinical studies have been reported [4][5][6]. Gene therapy may be technically categorized into gene transfer, gene silencing and gene editing.…”