Abstract
RNAi-based bone anabolic gene therapy has demonstrated initial success, but many practical challenges are still unmet. Here, we demonstrate that a recombinant adeno-associated virus 9 (rAAV9) is highly effective for transducing osteoblast lineage cells in the bone. The adaptor protein Schnurri-3 (SHN3) is a promising therapeutic target for osteoporosis, as deletion of shn3 prevents bone loss in osteoporotic mice and short-term inhibition of shn3 in adult mice increases bone mass. Accordingly, systemic and direct joint administration of an rAAV9 vector carrying an artificial-microRNA that targets shn3 (rAAV9-amiR-shn3) in mice markedly enhanced bone formation via augmented osteoblast activity. Additionally, systemic delivery of rAAV9-amiR-shn3 in osteoporotic mice counteracted bone loss and enhanced bone mechanical properties. Finally, we rationally designed a capsid that exhibits improved specificity to bone by grafting the bone-targeting peptide motif (AspSerSer)6 onto the AAV9-VP2 capsid protein. Collectively, our results identify a bone-targeting rAAV-mediated gene therapy for osteoporosis.
Highlights
RNAi-based bone anabolic gene therapy has demonstrated initial success, but many practical challenges are still unmet
An self-complementary AAVs (scAAVs) vector construct expressing the enhanced green fluorescent protein (Egfp) reporter gene was packaged into 14 conventional associated virus (AAV) capsids (AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV6.2, AAV7, AAV8, AAV9, AAVrh.[8], AAVrh.[10], AAVrh.[39], and AAVrh.43)[25] and incubated with mouse calvarial osteoblasts (COBs), bone marrow-derived osteoclast precursors (BM-OCPs), and chondrocyte progenitor cells (ATDC5s) at three different MOIs
Cortical bone at the femoral diaphysis showed no significant change in thickness (Supplementary Fig. 7b). These results demonstrate that systemically delivered recombinant adeno-associated virus 9 (rAAV9)-Cre in Shn3fl/fl mice targets osteoblast lineage cells and mediates shn[3] deletion to increase bone mass
Summary
RNAi-based bone anabolic gene therapy has demonstrated initial success, but many practical challenges are still unmet. Developed agents including the antisclerostin antibody[6] and a small molecule inhibitor of Cathepsin K7 can increase bone mass and reduce fracture risk in osteoporosis. These drugs show adverse cardiovascular events in clinical trials[8]. SHN3 deficiency is not associated with any observable phenotypes in nonskeletal tissues[10,12] These properties together make SHN3 inhibition an attractive approach to promote bone formation to treat osteoporosis. Replacement of the Rep and Cap genes with a transgene of interest produces a replication-defective recombinant AAV (rAAV) genome that can transduce target tissues as a potent vector[15,16]. AAV-based gene therapies for bone and joint disorders are limited[22]
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