Purpose

Osteoarthritis (OA) affects hundreds of million people worldwide. With pain management and surgical treatment merely alleviating symptoms, currently there are no disease modifying treatment options. Non-viral gene therapy has great potential to address this issue by delivering oligonucleotides that can halt inflammatory and degenerative processes, and induce regeneration. In this study we used poly(amido)amine-based polymeric nanoparticles (20Medtx) to deliver mRNA to human osteoarthritic chondrocytes.

Methods and Materials

Human OA chondrocytes (3 donors) were seeded into 96 well plates (20.000/cm²). After 24h they were transfected with nanoparticles (NP) (uncoated or PEGylated) loaded with mRNA coding for EGFP at mRNA concentrations of 0.8 to 3.2 µg/ml (weight ratio mRNA:NP= 1:25). 24h after transfection, LDH assay was used to asses toxicity. Transfection analysis was done using confocal imaging. Moreover, human cartilage explants were transfected 48h with PEGylated nanoparticles loaded with cy3 labelled siRNA to asses diffusion into cartilage.

Results

The transfection efficiency showed a dose dependent increase with particle concentration (for uncoated: increase from 9.43 (SD 7.53) – 36.34% (17.4) , Figure 1 A,B ). PEG-coated NPs showed similar transfection efficiency (highest 45.10% (13.83) for 60 μg/ml, comparison to uncoated 26.96% (8.2). Toxicity was lowest for uncoated NPs at 20 µg/ml 2.34 (9.23) and increased with NP concentration up to 45.96% (17.95) for 80 μg/ml of PEGylated nanoparticles. Confocal imaging could show that PEGylated NPs can enter into the cartilage (Figure 2).

Conclusion

We showed a novel way of transfection of primary human chondrocytes with nanoparticles for mRNA therapy in OA. The two different nanoparticle formulations showed similar transfection efficiency and similar toxicity. We could also show that PEGylated particles are able to diffuse into cartilage. Future experiments will focus to transfect chondrocytes in situ. Additionally, the delivery of biologically relevant targets (i.e. mRNA encoding for regenerative factors) will be tested.

Purpose

To investigate the in vitro effects of a single Methylprednisolone (MP) or Triamcinolone (TA) exposure on the viability, mechanics, and biochemical content of native articular cartilage explants and engineered neocartilage.

Methods and Materials

Articular cartilage explants were harvested from the femoral condyle of the bovine stifle, and neocartilage constructs were engineered from bovine stifle chondrocytes using the scaffold-free, self-assembling process. Both explants and neocartilage were exposed to chondrogenic medium containing a clinical dose of MP or TA for one hour, followed by fresh medium wash and exchange. At t=24 hours post-treatment, samples were assessed for viability (live/dead), mechanical properties (creep indentation and Instron tensile testing), and biochemical (collagen and glycosaminoglycan) content.

Results

Single steroid exposure was chondrotoxic in both explants and neocartilage constructs (Figure 1). Mean cell viability was significantly decreased in native explants and neocartilage constructs exposed to MP (35.5% and 30.2%, respectively) compared to control (49.8%, p < 0.001 and 36.8%, p = 0.062, respectively) and TA (45.7%, p = 0.003 and 37.3%, p = 0.04, respectively). No significant differences in collagen and glycosaminoglycan content were found within steroid treatment groups. There was significant weakening of mechanical properties of steroid-treated native explants when compared to control, with decreases in aggregate modulus (646.3 kPa vs 312.8 kPa (MP) and 257.0 kPa (TA), p < 0.001), shear modulus (370.1 kPa vs 191.2 kPa (MP) and 157.4 kPa (TA), p < 0.001), and ultimate tensile strength (UTS) (9.650 MPa vs 5.648 MPa (MP, p = 0.021) and 6.065 MPa (TA), p = 0.0403) (Figure 2). No significant differences in mechanical properties of engineered neocartilage were found among groups.

Conclusion

Single exposure of MP or TA resulted in chondrotoxicity and weakening of mechanical properties of native cartilage explants compared to control. Clinicians should be judicious regarding use of intra-articular steroids in the setting of cartilage repair.

Purpose

Lorecivivint (LOR) is a novel, investigational, intra-articular (IA) CLK2/DYRK1A inhibitor that modulates the Wnt pathway to potentially treat knee osteoarthritis (OA). We analyzed a 5-year, Phase 3, multicenter, observational extension study for safety and efficacy in subjects with moderate to severe knee OA.

Methods and Materials

Pooled data from 6-, 12-, 24-, and 36-month visits for completer subjects (NCT02951026) from Phase 2a (NCT02536833) and 2b (NCT03122860) LOR trials were analyzed. Subjects received a single LOR or control injection at baseline. Safety outcomes included adverse events (AEs) and serious AEs (SAEs). Efficacy outcomes included WOMAC Pain and Function subscores, analyzed by baseline-adjusted ANCOVA, and radiographic medial joint space width (mJSW). A post hoc analysis of 0.07 mg LOR versus control in a subject subgroup (unilateral symptoms, no widespread pain, 18-month radiograph at study termination) was performed.

Results

Overall, 584/703 (83%) subjects completed the study (mean age 60.7 years, mean BMI 29.1 kg/m², female 61%, KL 3 OA 61.2%). In the safety analysis set (LOR, n=495; control, n=208), 169 AEs were reported by 110 (15.6%) subjects. AE rates were similar between groups. Four AEs in 3 (0.6%) subjects were treatment related; none involved subject withdrawal. Sixty-eight SAEs (none treatment related) were reported by 38 (5.4%) subjects, with one death in the control group. LOR (n=59) showed greater improvements from baseline in WOMAC Pain and Function at 6 (Pain: -8.16, 95% CI [-15.60, -0.71], P=0.032; Function: -9.47 [-17.09, -1.84], P=0.015) and 12 (Pain: -8.51 [-15.17, -1.85], P=0.013; Function: -9.62 [-16.83, -2.42], P=0.009) months versus control (n=70) (Figure 1). No meaningful mJSW progression occurred in any group over 18 months. Limitations included post hoc analysis of small completer groups.

Conclusion

In this post hoc analysis, a single 0.07 mg LOR injection appeared well tolerated in knee OA subjects and demonstrated symptom improvements for up to 12 months versus control.

Purpose

To determine the efficacy of a single intra-articular injection of amniotic suspension allograft (ASA) compared to hyaluronic acid (HA) and saline for the treatment of symptomatic knee osteoarthritis (OA).

Methods and Materials

200 subjects at 12 sites were randomized 1:1:1 to saline, HA, or ASA. Patient-reported outcomes (PROs) were collected out to 12 months. Blinded subjects who reported unacceptable pain at 3 months from saline/HA groups were eligible to cross over to the ASA arm and followed for an additional 12 months (n=95). Changes in the Visual Analog Scale (VAS) and the Knee Injury and Osteoarthritis Outcome Score (KOOS) PROs from baseline were used to evaluate efficacy of treatment.

Results

Subjects who received ASA saw a robust improvement for VAS and KOOS PROs compared to saline/HA out to 12 months (Figure 1). OMERACT-OARSI simplified responder rates following ASA treatment was 54.5% at 12 months compared to 39.1% for HA and 44.1% for saline. Subjects who crossed over and were subsequently treated with ASA saw significant improvements in VAS and KOOS PROs (p<0.05 for all) and had a 56.8% responder rate out to 12 months. Interestingly, comparison of the response of subjects who originally received ASA treatment to those who crossed over showed no significant differences in treatment effect (Figure 2).

Conclusion

Conclusions: In this 200-patient study, subjects originally randomized to ASA showed improvements in pain and function out to 12 months, while subjects who crossed over to ASA following failed treatment with saline/HA showed a robust response to ASA treatment out to 12 months. This study provides evidence supporting the use of ASA to treat symptomatic knee OA.

Purpose

The purpose of this study was to construct an algorithm to optimize clinical outcomes prior to definitive surgical management in patients with symptomatic knee OA.

Methods and Materials

A systematic review of randomized controlled trials on IA injections for symptomatic knee OA without severe degenerative changes from the inception of databases to December 18th, 2019 was performed. Interventions were compared via arm-based Bayesian network meta-analysis in a random-effects model and interventions ranked for each domain (ROM, pain, PROs) via surface under the cumulative ranking curves.

Results

A total of 39 studies comprising 2,764 knees (60% females, age 57 ± 4 years, follow-up 10 ± 5 months) were included. There were no significant differences in preoperative PROs between injection groups. Interventions ranked highest for improved ROM included fetal-derived mesenchymal stem cells (MSC.Fetal), adipose-derived mesenchymal stem cells (MSC.Ad), and bone-marrow derived stem cells (MSC.BM). Interventions ranked highest for improved pain included all MSC injections and platelet rich plasma (PRP), with or without hyaluronic acid (HA). These injections ranked equally high within the Western Ontario and McMaster Universities Arthritis Index (WOMAC) and Knee Injury and Osteoarthritis Outcome Score (KOOS) subscales. Corticosteroid and HA injections ranked relatively poorly across all ROM, pain, and PRO domains.

Conclusion

PRP and MSC injections offer excellent clinical improvements in pain, ROM, and PROs, exceeding minimal clinically important difference thresholds. On the other hand, CSI and HA injections, in isolation, demonstrated relatively poor clinical outcomes with often non-statistically significant effects for pain, ROM, and PRO outcomes. Thus, for patients with symptomatic knee OA amenable to treatment by injection, the authors recommend either PRP, with or without HA, or MSC injection.

Purpose

The purpose of this study is to evaluate long term follow-up of single injection of nSTRIDE® APS for the treatment of unilateral knee osteoarthritis (NCT02138890).

Methods and Materials

Forty-six patients underwent a 2:1 randomization process to either one single injection of APS(n=31) or salin (n=15). APS was prepared with the nSTRIDE APS Kit (Zimmer Biomet). Data up to the 3 year follow up were previously published(1). The APS cohort was asked to participate in unblinded 60-month follow-up and the saline patients were offered cross-over after 12 months and were similarly followed. Efficacy endpoints (WOMAC LK 3.1, KOOS, and VAS) were measured as a change from baseline to each time point. Data is presented as mean ± SD. Missing data was calculated as baseline carried forward(BLCF) for subjects that had exited the trial for knee OA pain.

Results

At the final follow-up for the study (60 months), 17 of the original 31 randomized to APS and 6 of the original 15 saline/crossover remained in the study. The Kaplan-Meier curve for subjects exiting for knee-related events demonstrate a 0.64 survivorship probability[95% CI 0.463,0.820](Figure 1). Using the BLCF method, the APS cohort had a mean WOMAC pain percent improvement from baseline of 64.8±29.5% at 1 year(p<0.0001), 50.4±42.1% at 3 years(p<0.0001), and 39.0±41.2% at 5 years(p<0.0001). For the subjects that accepted crossover after 1 year, the WOMAC pain improvement was 45.5±33.3% at time of crossover(p=0.0002), 44.0±37.6% at 3 years(p=0.0008), and 24.6±33.7% at 5 years(p=0.028).

Conclusion

All in all, treatment with a single, intra-articular injection of APS in subjects with knee osteoarthritis has shown long-lasting pain and function relief and indicates of the long-term durability APS for knee osteoarthritis.

[1]Kon et al. AJSM;2020;48(11):2703-2710.