Harnessing the multifunctionality of lipid-based drug delivery systems for the local treatment of osteoarthritis.

Osteoarthritis (OA) is a widespread joint condition affecting millions globally, presenting a growing socioeconomic burden thus making the development of more effective therapeutic strategies crucial. This review emphasizes recent advancements in lipid-based drug delivery systems (DDSs) for intra-articular administration of OA therapeutics, encompassing non-steroidal anti-inflammatory drugs, corticosteroids, small molecule disease-modifying OA drugs, and RNA therapeutics. Liposomes, lipid nanoparticles, lipidic mesophases, extracellular vesicles and composite systems exhibit enhanced stability, targeted delivery, and extended joint retention, which contribute to improved therapeutic outcomes and minimized systemic drug exposure. Although active targeting strategies hold promise, further research is needed to assess their targeting efficiency in physiologically relevant conditions. Simultaneously, multifunctional DDSs capable of delivering combinations of distinct therapeutic classes offer synergistic effects and superior OA treatment outcomes. The development of such long-acting systems that resist rapid clearance from the joint space is crucial, where particle size and targeting capabilities emerge as vital factors. Additionally, combining cartilage lubrication properties with sustained drug delivery has demonstrated potential in animal models, meriting further investigation in human clinical trials. This review highlights the crucial need for direct, head-to-head comparisons of novel DDSs with standard treatments, particularly within the same drug class. These comparisons are essential in accurately evaluating their effectiveness, safety, and clinical applicability, and are set to significantly shape the future of OA therapy.

Pathophysiology to advanced intra-articular drug delivery strategies: Unravelling rheumatoid arthritis.

Rheumatoid arthritis (RA) is one of the most prevalent life-long autoimmune diseases with an unknown genesis. It primarily causes chronic inflammation, pain, and synovial joint-associated cartilage and bone degradation. Unfortunately, limited information is available regarding the etiology and pathogenesis of this chronic joint disorder. In the last few decades, an improved understanding of RA pathophysiology about key immune cells, antibodies, and cytokines has inspired the development of several anti-rheumatic drugs and biopharmaceuticals to act on RA-affected joints. However, life-long frequent systemic high doses of commercially available drugs are currently a limiting factor in the efficient management of RA. To address this issue, various single and double-barrier intra-articular drug delivery systems (IA-DDSs) such as nanocarriers, microparticles, hydrogels, and particles-hybrid hydrogel composite have been developed which can exclusively target the RA-affected joint cavity and release the precisely controlled therapeutic drug concentration for prolonged time whilst avoiding the systemic toxicity. This review provides a comprehensive overview of the pathogenesis of RA and discusses the rational design and development of biomaterials-based novel IA-DDs, ranging from conventional to advanced systems, for improved treatment of RA. Therefore, this review aims to unravel the pathophysiology of rheumatoid arthritis and explore cutting-edge IA-DD strategies exploiting biomaterials. It offers researchers a consolidated and up-to-date resource platform to analyze existing knowledge, identify research gaps, and contribute to the scientific literature.

Intra-articular delivery of an indoleamine 2,3-dioxygenase galectin-3 fusion protein for osteoarthritis treatment in male Lewis rats.

OBJECTIVE: Osteoarthritis (OA) is driven by low-grade inflammation, and controlling local inflammation may offer symptomatic relief. Here, we developed an indoleamine 2,3-dioxygenase and galectin-3 fusion protein (IDO-Gal3), where IDO increases the production of local anti-inflammatory metabolites and Gal3 binds carbohydrates to extend IDO's joint residence time. In this study, we evaluated IDO-Gal3's ability to alter OA-associated inflammation and pain-related behaviors in a rat model of established knee OA. METHODS: Joint residence was first evaluated with an analog Gal3 fusion protein (NanoLuc™ and Gal3, NL-Gal3) that produces luminescence from furimazine. OA was induced in male Lewis rats via a medial collateral ligament and medial meniscus transection (MCLT + MMT). At 8 weeks, NL or NL-Gal3 were injected intra-articularly (n = 8 per group), and bioluminescence was tracked for 4 weeks. Next, IDO-Gal3s's ability to modulate OA pain and inflammation was assessed. Again, OA was induced via MCLT + MMT in male Lewis rats, with IDO-Gal3 or saline injected into OA-affected knees at 8 weeks post-surgery (n = 7 per group). Gait and tactile sensitivity were then assessed weekly. At 12 weeks, intra-articular levels of IL6, CCL2, and CTXII were assessed. RESULTS: The Gal3 fusion increased joint residence in OA and contralateral knees (p < 0.0001). In OA-affected animals, both saline and IDO-Gal3 improved tactile sensitivity (p = 0.008), but IDO-Gal3 also increased walking velocities (p ≤ 0.033) and improved vertical ground reaction forces (p ≤ 0.04). Finally, IDO-Gal3 decreased intra-articular IL6 levels within the OA-affected joint (p = 0.0025). CONCLUSION: Intra-articular IDO-Gal3 delivery provided long-term modulation of joint inflammation and pain-related behaviors in rats with established OA.

Application of magnetic particle imaging to evaluate nanoparticle fate in rodent joints.

Nanoparticles are a promising approach for improving intra-articular drug delivery and tissue targeting. However, techniques to non-invasively track and quantify their concentration in vivo are limited, resulting in an inadequate understanding of their retention, clearance, and biodistribution in the joint. Currently, fluorescence imaging is often used to track nanoparticle fate in animal models; however, this approach has limitations that impede long-term quantitative assessment of nanoparticles over time. The goal of this work was to evaluate an emerging imaging modality, magnetic particle imaging (MPI), for intra-articular tracking of nanoparticles. MPI provides 3D visualization and depth-independent quantification of superparamagnetic iron oxide nanoparticle (SPION) tracers. Here, we developed and characterized a polymer-based magnetic nanoparticle system incorporated with SPION tracers and cartilage targeting properties. MPI was then used to longitudinally assess nanoparticle fate after intra-articular injection. Magnetic nanoparticles were injected into the joints of healthy mice, and evaluated for nanoparticle retention, biodistribution, and clearance over 6 weeks using MPI. In parallel, the fate of fluorescently tagged nanoparticles was tracked using in vivo fluorescence imaging. The study was concluded at day 42, and MPI and fluorescence imaging demonstrated different profiles in nanoparticle retention and clearance from the joint. MPI signal was persistent over the study duration, suggesting NP retention of at least 42 days, much longer than the 14 days observed based on fluorescence signal. These data suggest that the type of tracer - SPIONs or fluorophores - and modality of imaging can affect interpretation of nanoparticle fate in the joint. Given that understanding particle fate over time is paramount for attaining insights about therapeutic profiles in vivo, our data suggest MPI may yield a quantitative and robust method to non-invasively track nanoparticles following intra-articular injection on an extended timeline.

An intra-articular injectable phospholipids-based gel for the treatment of rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic inflammatory and destructive arthropathy with a high deformity rate. Despite numerous studies and clinical trials, no curative treatment is available for large weight-bearing joints. Intra-articular (IA) injections could deliver high concentrations of drug to the afflicted joint and improve the drug efficacy while reducing systemic toxicity. However, free drugs are rapidly cleared from synovial fluid and do not significantly halt the progression of joint disease. Herein, a phospholipids-based controlled-release gel was prepared for sustained IA delivery of celastrol (CEL) and the therapeutic efficiency was evaluated in a rheumatoid arthritis rabbit model. The CEL-loaded gel (CEL-gel) contained up to 70% phospholipids yet was easy to inject. After injecting into the joint cavity, CEL-gel achieved sol to gel phase transition without special stimuli and gelling agent. In vitro release and in vivo pharmacokinetic studies evidenced the stable and sustained release action of CEL-gel. A single IA injection of CEL-gel could maintain therapeutic efficiency for about 25 d and showed much better anti-arthritic efficacy compared to repeated injections of free drug solution (CEL-sol). Furthermore, the IA injection of CEL-gel greatly reduced the systemic toxicity of CEL. With good biocompatibility and biodegradability, CEL-gel might be a promising IA drug delivery system.

Poly ε-Caprolactone Nanoparticles for Sustained Intra-Articular Immune Modulation in Adjuvant-Induced Arthritis Rodent Model.

Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disorder with synovitis and articular pathology as its primary expositions. Leflunomide (Lfd) is an anti-rheumatic drug that is effective in the treatment of RA, but displays severe side effects upon prolonged systemic administration. Local therapy might represent a promising strategy to treat rheumatoid arthritis without eliciting systemic adverse effects. In this study, leflunomide-loaded poly(ε-caprolactone) nanoparticles (Lfd-NPs) were prepared and assessed as a local drug delivery system capable of alleviating RA-associated inflammation. Lfd-NPs were optimized using the Quality by Design (QbD) approach, applying a 32 full factorial design. In vitro drug release from NPs was examined in simulated synovial fluid. In addition, the in vivo efficacy of Lfd-NPs was evaluated in the Adjuvant Induced Arthritis (AIA) rodent model. Sustained drug release in simulated synovial fluid was observed for up to 168 h. A gradual reduction in paw volume and knee diameter was observed over the course of treatment, indicating the regression of the disease. In addition, significant reductions in serum proinflammatory markers and cytokines, including the C-reactive protein (CRP), rheumatoid factor (RF), TNF-α, IL1-ß, and IL-6, were verified upon treatment with Lfd-NPs, suggesting the modulation of immune responses at the pathological site. Most importantly, no remarkable signs of toxicity were observed in Lfd-NP-treated animals. Collectively, intra-articularly administered Lfd-NPs might represent a potential therapeutic alternative to systemically administered drugs for the treatment of rheumatoid arthritis, without eliciting systemic adverse effects.

The in vitro and in vivo study of novel formulation of andrographolide PLGA nanoparticle embedded into gelatin-based hydrogel to prolong delivery and extend residence time in joint.

Andrographolide (AG), a well-known traditional medicinal plant in Southeast Asia, is widely used for treatment of many chronic diseases. Interestingly, AG has been reported to have inhibitory effects on osteoclast function and anti-inflammatory properties. Because of these therapeutic properties, this study aimed to develop and optimize the formulation of AG using PLGA nanocarriers and gelatin-based hydrogel to prolong the retention time in the joint. We investigated the in vitro release pattern of the AG nanoparticles formulation which prepared by emulsion solvent evaporation method and embedded into gelatin-based hydrogel. The result showed that the AG loaded ester terminated end group PLGA polymer gradually released AG from the PLGA nanoparticles when compared with AG solution. Importantly, the combined use of gelatin-based hydrogel with AG from the PLGA nanoparticles significantly delayed the AG release more than 1 month. Furthermore, we selected the DiR fluorescence dye to represents AG and monitored the retention time by IVIS imaging. The optimal formulation was administered as intra-articular drug delivery systems in in vivo study. The results successfully displayed a long-term sustained release for implantation (≈2 months) and injection (≥2 months) providing a novel strategy for the local management of osteoarthritis disease.

Osteoarthritis In Vitro Models: Applications and Implications in Development of Intra-Articular Drug Delivery Systems.

Osteoarthritis (OA) is a complex multi-target disease with an unmet medical need for the development of therapies that slow and potentially revert disease progression. Intra-articular (IA) delivery has seen a surge in osteoarthritis research in recent years. As local administration of molecules, this represents a way to circumvent systemic drug delivery struggles. When developing intra-articular formulations, the main goals are a sustained and controlled release of therapeutic drug doses, taking into account carrier choice, drug molecule, and articular joint tissue target. Therefore, the selection of models is critical when developing local administration formulation in terms of accurate outcome assessment, target and off-target effects and relevant translation to in vivo. The current review highlights the applications of OA in vitro models in the development of IA formulation by means of exploring their advantages and disadvantages. In vitro models are essential in studies of OA molecular pathways, understanding drug and target interactions, assessing cytotoxicity of carriers and drug molecules, and predicting in vivo behaviors. However, further understanding of molecular and tissue-specific intricacies of cellular models for 2D and 3D needs improvement to accurately portray in vivo conditions.

Reinforced Supramolecular Hydrogels from Attapulgite and Cyclodextrin Pseudopolyrotaxane for Sustained Intra-Articular Drug Delivery.

Injectable hydrogels for nonsteroidal anti-inflammatory drugs' (NSAIDs) delivery to minimize the side effects of NSAIDs and achieve long-term sustained release at the targeted site of synovial joint are attractive for osteoarthritis therapy, but how to improve its mechanical strength remains a challenge. In this work, a kind of 1D natural clay mineral material, attapulgite (ATP), is introduced to a classical cyclodextrin pseudopolyrotaxane (PPR) system to form a reinforced supramolecular hydrogel for sustained release of diclofenac sodium (DS) due to its rigid, rod-like morphology, and unique structure, which has great potential in tissue regeneration, repair, and engineering. Investigation on the interior morphology and rheological property of the obtained hydrogel points out that the ATP distributed in PPR hydrogel plays a role similar to the "reinforcement in concrete" and exhibits a positive effect on improving the mechanical properties of PPR hydrogel by regulating their interior morphology from a randomly distributed style to the well-ordered porous frame structure. The hybrid hydrogels demonstrate good shear-thinning and thixotropic properties, excellent biocompability, and sustained release behavior both in vitro and in vivo. Furthermore, preliminary in vivo treatment in an acute inflammatory rat model reveals that the ATP hybrid hydrogels present sustained anti-inflammatory effect.

Intra-articular delivery of flurbiprofen sustained release thermogel: improved therapeutic outcome of collagenase II-induced rat knee osteoarthritis.

Knee osteoarthritis (OA) is a common degenerative disease. Intra-articular administration of flurbiprofen is frequently employed in clinic to treat OA, while repeated injections are required because of the limited effective duration. To improve therapeutic outcome and prolong the treatment interval, a poly(ε-caprolactone-co-lactide)-b-poly(ethylene glycol)-b-poly(ε-caprolactone-co-lactide) (PCLA-PEG-PCLA) triblock copolymer based flurbiprofen thermosensitive gel for the sustained intra-articular drug delivery was designed in this study. The anti-OA effects of this flurbiprofen thermogel were investigated on collagenase II-induced rat knee OA model by multiple approaches and compared with that of conventional sodium hyaluronate and flurbiprofen injecta. In vitro drug release studies indicated that flurbiprofen was sustained released from the thermosensitive gel for more than three weeks. This sustained drug release system exerted comparable short-term analgesic effects and distinctly improved long-term analgesic efficacy in terms of the increased percentage of the total ipsilateral paw print intensity and the reduced Knee-Bend scores of OA rats. The inflammatory response was attenuated in the samples of flurbiprofen gel treated group by showing decreased IL-1, IL-6, and IL-11 levels in the joint fluid and down-regulated IL-1, IL-6, IL-11, COX-2, TNF-α, and NF-κB/p65 expression in the articular cartilages. The results suggest the suitability of thermosensitive copolymer PCLA-PEG-PCLA for sustained intra-articular effects of flurbiprofen and provide in vivo experimental evidence for potential clinical application of this flurbiprofen delivery system to better management of OA cases.

Sulfasalazine Resolves Joint Stiffness in a Rabbit Model of Arthrofibrosis.

Joint stiffness due to fibrosis/capsule contracture is a seriously disabling complication of articular injury that surgical interventions often fail to completely resolve. Fibrosis/contracture is associated with the abnormal persistence of myofibroblasts, which over-produce and contract collagen matrices. We hypothesized that intra-articular therapy with drugs targeting myofibroblast survival (sulfasalazine), or collagen production (ß-aminopropionitrile and cis-hydroxyproline), would reduce joint stiffness in a rabbit model of fibrosis/contracture. Drugs were encapsulated in poly[lactic-co-glycolic] acid pellets and implanted in joints after fibrosis/contracture induction. Capsule α-smooth muscle actin (α-SMA) expression and intimal thickness were evaluated by immunohistochemistry and histomorphometry, respectively. Joint stiffness was quantified by flexion-extension testing. Drawer tests were employed to determine if the drugs induced cruciate ligament laxity. Joint capsule fibroblasts were tested in vitro for contractile activity and α-SMA expression. Stiffness in immobilized joints treated with blank pellets (control) was significantly higher than in non-immobilized, untreated joints (normal) (p = 0.0008), and higher than in immobilized joints treated with sulfasalazine (p = 0.0065). None of the drugs caused significant cruciate ligament laxity. Intimal thickness was significantly lower than control in the normal and sulfasalazine-treated groups (p = 0.010 and 0.025, respectively). Contractile activity in the cells from controls was significantly increased versus normal (p = 0.001). Sulfasalazine and ß-aminopropionitrile significantly inhibited this effect (p = 0.005 and 0.0006, respectively). α-SMA expression was significantly higher in control versus normal (p = 0.0021) and versus sulfasalazine (p = 0.0007). These findings support the conclusion that sulfasalazine reduced stiffness by clearing myofibroblasts from fibrotic joints. Statement of clinical significance: The results provide proof-of-concept that established joint stiffness can be resolved non-surgically. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:629-638, 2020.

A rabbit model demonstrates the influence of cartilage thickness on intra-articular drug delivery and retention within cartilage.

For evaluation of new approaches to drug delivery into cartilage, the choice of an animal model is critically important. Since cartilage thickness varies with animal size, different levels of drug uptake, transport and retention should be expected. Simple intra-articular injection can require very high drug doses to achieve a concentration gradient high enough for drug diffusion into cartilage. New approaches involve nanoparticle delivery of functionalized drugs directly into cartilage; however, diffusion-binding kinetics proceeds as the square of cartilage thickness. In this study, we demonstrate the necessity of using larger animals for sustained intra-cartilage delivery and retention, exemplified by intra-articular injection of Avidin (drug-carrier) into rabbits and compared to rats in vivo. Penetration and retention of Avidin within cartilage is greatly enhanced by electrostatic interactions. Medial tibial cartilage was the thickest of rabbit cartilages, which generated the longest intra-cartilage half-life of Avidin (τ1/2 = 154 h). In contrast, Avidin half-life in thinner rat cartilage was 5-6 times shorter (τ1/2 ∼ 29 h). While a weak correlation (R(2) = 0.43) was found between Avidin half-lives and rabbit tissue GAG concentrations, this correlation improved dramatically (R(2) = 0.96) when normalized to the square of cartilage thickness, consistent with the importance of cartilage thickness to evaluation of drug delivery and retention.

Electrostatic interactions enable rapid penetration, enhanced uptake and retention of intra-articular injected avidin in rat knee joints.

Intra-articular (i.a.) drug delivery for local treatment of osteoarthritis remains inadequate due to rapid clearance by the vasculature or lymphatics. Local therapy targeting articular cartilage is further complicated by its dense meshwork of collagen and negatively charged proteoglycans, which can prevent even nano-sized solutes from entering. In a previous in vitro study, we showed that Avidin, due to its size (7 nm diameter) and high positive charge (pI 10.5), penetrated the full thickness of bovine cartilage and was retained for 15 days. With the goal of using Avidin as a nano-carrier for cartilage drug delivery, we investigated its transport properties within rat knee joints. Avidin penetrated the full thickness of articular cartilage within 6 h, with a half-life of 29 h, and stayed inside the joint for 7 days after i.a. injection. The highest concentration of Avidin was found in cartilage, the least in patellar tendon and none in the femoral bone; in contrast, negligible Neutravidin (neutral counterpart of Avidin) was present in cartilage after 24 h. A positive correlation between tissue sGAG content and Avidin uptake (R(2) = 0.83) confirmed the effects of electrostatic interactions. Avidin doses up to at least 1 µM did not affect bovine cartilage explant cell viability, matrix catabolism or biosynthesis.

Intra-articular fate of degradable poly(ethyleneglycol)-hydrogel microspheres as carriers for sustained drug delivery.

A novel degradable microsphere (MS) for intra-articular drug delivery, composed of a polyethylene glycol (PEG) core containing degradable regions made of short poly-(lactic-co-glycolic acid) (PLGA) sequences - named PEG-hydrogel MS - was injected into the cavity of sheep shoulder joint, and compared to non-degradable MS devoid of hydrolysable crosslinker in terms of location, degradation and inflammation. One week after intra-articular injection both groups of MS were localized beneath the synovial lining of the synovial fringes located at bottom of the shoulder joint, while a fraction of particles remained in synovial fluid. Histological analyses made one and 4 weeks after intra-articular injection showed cell proliferation around the non-degradable MS entrapped within the synovium. By contrast, degradable PEG-hydrogel MS were surrounded by few cells. The degradation of degradable PEG-hydrogel MS within the synovium was slow and was not fully complete after four weeks. Our findings indicate that the tissue entrapment of MS below the synovial lining was independent of the material degradability, while degradable PEG-hydrogel MS are less inflammatory than the non-degradable one. Degradable PEG-hydrogel MS offer several advantages over the non-degradable MS as carriers for a sustained drug delivery in synovial tissue according to the low intensity of inflammatory reaction triggered in synovium.