2263: Nanotherapeutic Approaches for the Treatment of Post-traumatic Osteoarthritis

Luke Springer¹, Kelsey Collins², Huimin Yan², Ying Hu³, Farshid Guilak², Samuel Wickline⁴, Hua Pan² and Christine Pham², ¹Washington University School of Medicine, St. Louis, MO, ²Washington University School of Medicine, Saint Louis, MO, ³Washington University, Saint Louis, MO, ⁴University of South Florida, Tampa, FL

Background/Purpose: Osteoarthritis (OA) represents the most common form of arthritis and is a major cause of morbidity in the aging population. Post-traumatic OA (PTOA) is a form of OA that develops after a joint injury and in which the nature and time of trauma is known. Studies have documented a robust inflammatory response in the immediate aftermath of joint injury that sets in motion a sequence of events, resulting in eventual PTOA. This inflammatory response likely contributes to chondrocyte death, impaired cartilage repair and PTOA. We have previously shown that intra-articular (IA) delivery of a nanoparticle (NP) comprising a peptide, p5RHH, complexed to siRNA suppressed NF-kB p65 expression and significantly mitigated early cartilage changes in a non-invasive joint injury model (Yan et al. Proc Natl Acad Sci USA. 2016 113:E6199-E6208). We hypothesize that suppression of the inflammatory response combined with overexpression of an anabolic factor to maintain cartilage homeostasis will mitigate pain and structural damage in a mouse model of PTOA.

Methods: We employed the amphipathic peptide p5RHH to form nanocomplexes, which have been proven safe and stable for IA delivery of both siRNA and mRNA. Mice were challenged with unilateral destabilization of the medial meniscus (DMM) to provide proof-of-concept that IA delivery of siRNA to knockdown NF-kB p65 (anti-inflammatory) combined with simultaneous delivery of mRNA to overexpress WNT16 (anabolic) will have additive effects compared to treatment with individual agent. Comparisons between groups (≥ 3) were performed by one-way ANOVA followed by Bonferroni's post-test. P < 0.05 was considered significant.

Results: Preliminary data show that p5RHH-p65 siRNA nanotherapy mitigated early DMM changes at 4 weeks, as evidenced by a lower Modified Mankin score (figure 1). We next challenged groups of mice with DMM and treated them with different NP combinations. IA treatment with a combination of p65 siRNA and WNT16 mRNA significantly reduced pressure-pain hyperalgesia in the DMM limb of both male and female mice when measured at 12 weeks post-destabilization (figure 2).

Conclusion: IA drug delivery for OA treatment hold key benefits due to increased local drug availability and reduced systemic bystander effects. However, these benefits are limited by rapid drug clearance from the joint space. We show that p5RHH-based delivery of anti-inflammatory and anabolic factors mitigates hyperalgesia, which has been shown to correlate with structural joint damage. This nanotherapeutic approach promises to overcome IA clearance deficits to halt cartilage loss and maintain cartilage homeostasis, translating to better long-term outcomes in PTOA treatment.
Figure 1. Twelve weeks old male mice were subjected to DMM surgery on the left knee. NPs were administered IA (0.1 ug of scrambled siRNA or NF-kB p65 siRNA) in a volume of 20 uL, 24 hours after surgery and then weekly thereafter (4 doses total). Mice were sacrificed at 4 weeks and both knees were harvested and analyzed histologically. A representative image of Safranin-O/Fast green stained knee joint section is shown for each treatment group. The right knee served as control. Treatment with NF-kB p65 siRNA NP mitigated cartilage fibrillation/clefts (arrow) and loss of Safranin-O staining (*) in the DMM knee. Scale bar = 100 m. *P < 0.05 by Student’s t test.

Figure 2. Twelve weeks old mice were subjected to DMM surgery on the left knee. NPs were administered IA with the indicated siRNA and mRNA combination (0.1 ug each) in a volume of 20 uL, 24 hours after surgery and then on weeks 1, 2, 4, 6, 8, 10. Mice were subjected to SMALGO at week 12 post-DMM. The pressure-pain hyperalgesia threshold (g) was obtained in DMM (red triangles) knee and contralateral non-surgery knee (blue circle). *P < 0.05, **P < 0.01, ****P < 0.0001 by one-way ANOVA and Student’s t test.
Disclosures: L. Springer, None; K. Collins, None; H. Yan, None; Y. Hu, None; F. Guilak, Cytex Therapeutics Inc.; S. Wickline, Altamira Therapeutics; H. Pan, Altamira Therapeutics; C. Pham, None.