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Matrix-anchored Nanoparticles for MMP13 RNA Interference Block Post-Traumatic Osteoarthritis

ABSTRACT: Targeting MMP13 directly by RNA interference (RNAi) silenced MMP13 in a murine post-traumatic osteoarthritis (PTOA) and was found to have broad effects on overall joint health, including both the articular cartilage and surrounding hard and soft tissues. To further characterize the global impacts of targeted MMP13 inhibition, the mechanical loading mouse model was applied with samples were harvested at 4 weeks to capture an intermediate stage of disease. The nanoString nCounter Inflammation panel was used to broadly quantify gene expression of the knee joint samples (articular cartilage, meniscal, and synovial tissue). Abstract: Osteoarthritis (OA) is a debilitating and prevalent chronic disease, but there are no approved disease modifying OA drugs (DMOADs), only pharmaceuticals for pain management. OA progression, particularly for post-traumatic osteoarthritis (PTOA), is associated with inflammation and enzymatic degradation of the extracellular matrix. In particular, matrix metalloproteinase 13 (MMP13) breaks down collagen type 2 (CII), a key structural component of cartilage extracellular matrix, and consequently, matrix degradation fragments perpetuate inflammation and a degenerative cycle that leads to progressive joint pathology. Here, we tested extracellular matrix-binding MMP13 RNA interference (RNAi) nanoparticles (NPs) as a DMOAD. The retention approach pursued here deviates from the convention of targeting specific cell types (e.g., through cell surface receptors) and instead leverages a monoclonal antibody (mAbCII) that targets extracellular CII that becomes uniquely accessible in OA-damaged cartilage. CII monoclonal antibody-functionalized nanoparticles carrying small interfering ribonucleic acid (siRNA) (mAbCII-siNPs) create an in situ NP depot for retention and potent activity within OA joints. The mAbCII-siNPs loaded with MMP13 siRNA (mAbCII-siNP/siMMP13) potently suppressed MMP13 expression (95% silencing) in TNFα-stimulated chondrocytes in vitro and had higher binding to trypsin-damaged porcine cartilage than control NPs. In an acute mechanical injury mouse model of PTOA, mAbCII-siNP/siMMP13 achieved 80% reduction in MMP13 expression (p = 0.00231), whereas control siNP/siMMP13 achieved only 55% silencing. In a more severe, longer-term PTOA model, weekly mAbCII-siNP/siMMP13 treatment provided significant protection of cartilage integrity (0.45+/-.3 vs 1.6+/-.5 on the OARSI scale; p=0.0013) and overall joint structure (1.3+/-.6 vs 2.8+/-.2 on the Degenerative Joint Disease scale; p=0.0418), including reduction of osteophyte formation (siMMP13 vs siNEG: 0.088+/-0.074 vs 0.47+/-0.107 mm femoral osteophyte outgrowth; p<0.0001). Multiplexed gene expression analysis of 254 inflammation-related genes showed that MMP13 inhibition suppressed clusters of genes associated with tissue restructuring, angiogenesis, innate immune response, and proteolysis. Finally, in benchmarking studies, intra-articular mAbCII-siNPs better reduced OA disease progression relative to either one time or weekly treatment with the clinical gold standard steroid methylprednisolone. Here, we introduce the concept of anchoring to unique local extracellular matrix signatures to sustain retention and increase delivery efficacy of biologics with intracellular activity and also validates the promise of MMP13 RNAi as a DMOAD in a clinically-relevant therapeutic context.

ORGANISM(S): Mus musculus

PROVIDER: GSE171031 | GEO | 2021/04/17

REPOSITORIES: GEO

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Project description:As the most common degenerative joint disease, osteoarthritis (OA) contributes significantly to pain and disability during aging. Several genes of interest involved in articular cartilage damage in OA have been identified. However, the direct causes of OA are poorly understood. Evaluating the public human RNA-seq dataset showed that Cbfβ, (subunit of a heterodimeric Cbfβ/Runx1,Runx2, or Runx3 complex) expression is decreased in the cartilage of patients with OA. Here, we found that the chondrocyte-specific deletion of Cbfβ in tamoxifen-induced Cbfβf/fCol2α1-CreERT mice caused a spontaneous OA phenotype, worn articular cartilage, increased inflammation, and osteophytes. RNA-sequencing analysis showed that Cbfβ deficiency in articular cartilage resulted in reduced cartilage regeneration, increased canonical Wnt signaling and inflammatory response, and decreased Hippo/YAP signaling and TGF-β signaling. Immunostaining and western blot validated these RNA-seq analysis results. ACLT surgery-induced OA decreased Cbfβ and Yap expression and increased active β-catenin expression in articular cartilage, while local AAV-mediated Cbfβ overexpression promoted Yap expression and diminished active β-catenin expression in OA lesions. Remarkably, AAV-mediated Cbfβ overexpression in knee joints of mice with OA showed the significant protective effect of Cbfβ on articular cartilage in the ACLT OA mouse model. Overall, this study, using loss-of-function and gain-of-function approaches, uncovered that low expression of Cbfβ may be the cause of OA. Moreover, Local admission of Cbfβ may rescue and protect OA through decreasing Wnt/β-catenin signaling, and increasing Hippo/Yap signaling and TGFβ/Smad2/3 signaling in OA articular cartilage, indicating that local Cbfβ overexpression could be an effective strategy for treatment of OA. Using unbiased genome-wide RNA-seq data from Cbfβf/f;Col2α1-Cre hip joint articular cartilage and Cbfβf/f;Aggrecan-cre knee joint articular cartilage and their controls, we examined Cbfβ-mediated transcriptional targets for articular cartilage regeneration in OA.

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Matrix-anchored Nanoparticles for MMP13 RNA Interference Block Post-Traumatic Osteoarthritis

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