Sustained intra-articular delivery of IL-1RA from a thermally-responsive elastin-like polypeptide as a therapy for post-traumatic arthritis

Abstract

Post-traumatic arthritis (PTA) is a rapidly progressive form of arthritis that develops due to joint injury, including articular fracture. Current treatments are limited to surgical restoration and stabilization of the joint; however, evidence suggests that PTA progression is mediated by the upregulation of pro-inflammatory cytokines, such as interleukin-1 (IL-1) or tumor necrosis factor-α (TNF-α). Although these cytokines provide potential therapeutic targets for PTA, intra-articular injections of anti-cytokine therapies have proven difficult due to rapid clearance from the joint space. In this study, we examined the ability of a cross-linked elastin-like polypeptide (xELP) drug depot to provide sustained intra-articular delivery of IL-1 and TNF-α inhibitors as a beneficial therapy. Mice sustained a closed intra-articular tibial plateau fracture; treatment groups received a single intra-articular injection of drug encapsulated in xELP. Arthritic changes were assessed 4 and 8 weeks after fracture. Inhibition of IL-1 significantly reduced the severity of cartilage degeneration and synovitis. Inhibition of TNF-α alone or with IL-1 led to deleterious effects in bone morphology, articular cartilage degeneration, and synovitis. These findings suggest that IL-1 plays a critical role in the pathogenesis of PTA following articular fracture, and sustained intra-articular cytokine inhibition may provide a therapeutic approach for reducing or preventing joint degeneration following trauma.

Fig. 1

Results of in vitro drug release studies. (a) Close-up image of lyophilized xELP constructs. (b) Drug rehydrated xELP constructs at 4 °C (left) and at 37 °C (right). Change in opacity of the xELP at the transition-induced temperature of 37 °C is seen by comparing the left and right pictures. (c) Drug release of IL-lRa from xELP in 10% serum-containing medium over 180 h. Mean ± standard deviation repo1ted (n = 4). (d) Drug release of sTNFRII from xELP in 10 % semm-containing medium over 180 h. Mean ± standard deviation repo1ted (n = 4).

Fig. 2

Results of in vivo drug release studies. (a) Drug release of IL-1Ra from xELP in mouse serum as measured by ELISA. Fx and xELP[PBS] were run as background controls for ELISA. Data are mean of samples in triplicate (n = 6-11 per data point). (b) Drug release of sTNFRII from xELP in mouse semm as measured by ELISA. Fx and xELP[PBS] were mn as background controls for ELISA. Dataare mean of samples in triplicate (n = 6-11 per data point).

Fig. 3

Assessment of articular cartilage degeneration. (a) Modified Mankin total joint score for limbs at 4 weeks. p values denote significance between control and fractured limb using Wilcoxon Matched Pairs test. Median values and 25^th-75^th quartile range reported (n = 6-8 per group). (b) Modified Mankin total joint score for limbs at 8 weeks. p values denote significance between control and fractured limb using Wilcoxon Matched Pairs test. Median values and 25^th-75^th quartile range reported (n = 6-8 per group). (c) Representative histological pictures of articular cartilage on the lateral tibial plateau taken at 40× magnification for each group with Safranin-O-Fast Green staining at 8 weeks. Scale bar indicates 5 μm.

Fig. 4

Assessment of synovial inflammation. (a) Modified Krenn joint score for medial and lateral sides of each limb at 4 weeks. * denotes p < 0.05 between control and fractured limb using Wilcoxon Matched Pairs test. Mean ± standard deviation reported (n = 5-8 per group). (b) Modified Krenn joint score for medial and lateral sides of each limb at 8 weeks. * denotes p < 0.05 between control and fractured limb using Wilcoxon Matched Pairs test. Mean ± standard deviation reported (n = 5-8 per group). (c) Representative histological pictures of synovium (black arrows) in the lateral tibial plateau quadrant of knee joints at 8 weeks using Hematoxylin and Eosin stained sections. Scale bar indicates 10 μm.

Fig. 5

Assessment of bone morphological changes. (a) Representative microCT images of the fractured limb from each group at 8 weeks. Fractures are present in each image in the lateral tibial plateau. Scale bar indicates 1 mm. (b) Representative microCT images of the tibial plateau of a control limb (first picture) and fractured limbs (pictures 2-6) from each group at 8 weeks. Scale bar indicates 1 mm. (c) Bone volume data shown at 4 weeks for each group. Tukey HSD post-hoc test performed; groups that do not share letters are significantly different from each other (p < 0.05). Mean ± standard deviation reported (n = 5-8 per group). (d) Bone volume data shown at 8 weeks for each group. Tukey HSD post-hoc test performed; groups that do not share letters are significantly different from each other (p < 0.05). Mean ± standard deviation reported (n = 5-8 per group). (e) Bone density data shown at 4 weeks for each group. Tukey HSD post-hoc test performed; groups that do not share letters are significantly different from each other (p < 0.05). Mean ± standard deviation reported (n = 5-8 per group). (f) Bone density data shown at 8 weeks for each group. Tukey HSD post-hoc test performed; groups that do not share letters are significantly different from each other (p < 0.05). Mean ± standard deviation reported (n = 5-8 per group).

Fig. 6

Assessment of serum and synovial fluid biomarkers. (a) Levels of cartilage oligomeric matrix protein (COMP) in synovial fluid in the limbs at 4 weeks. Mean ± standard deviation reported (n = 5-8 per group). # denotes p < 0.05 between groups using Kruskal-Wallis ANOVA. * denotes p < 0.05 between fractured and control limb using Wilcoxon Matched Pairs test. (b) Levels of COMP in synovial fluid in the limbs at 8 weeks. Mean ± standard deviation reported (n = 5-8 per group). # denotes p < 0.05 between groups using Kruskal-Wallis ANOVA. * denotes p < 0.05 between fractured and control limb using Wilcoxon Matched Pairs test. (c) Levels of COMP in serum for each group at 4 and 8 weeks. Mean ± standard deviation reported (n = 5-8 per group). # denotes p < 0.05 between groups using Kruskal-Wallis ANOVA. * denotes p < 0.05 between 4 and 8 weeks using Kruskal-Wallis ANOVA test. (d) Levels of matrix metalloproteinase-3 (MMP-3) in serum for each group at 4 and 8 weeks. Mean ± standard deviation reported (n = 5-8 per group). * denotes p < 0.05 between 4 and 8 weeks using Kruskal-Wallis ANOVA. Groups that do not share letters are significantly different (p < 0.05) at 4 weeks by Kruskal-Wallis ANOVA.