Cell-based tissue-engineered reconstruction of large osteochondral defects in the minipig knee.

State of the art The availability of regenerative solutions for deep cartilage defects (osteochondral defects) in human joints is known as an unmet medical need. A long-term follow-up study from Mayoclinic recently demonstrated a significant higher risk of early osteoarthritis for knees with osteochondritis dissecans regardless if treatment was conservative or operative (1,2). The surgical treatment of these large osteochondral defects in osteochondritis dissecans or after trauma, is based on different techniques that all have certain advantages and disadvantages: - osteochondral autograft (3): 1 or more osteochondral plugs are harvested from a non-loaded area in the knee. In this setting the body’s own tissue (autologous) tissue is used, that moreover consist of normal articular cartilage with an intact ‘tidemark’ transition zone between cartilage and bone. As the amount of plugs that can be harvested is limited, this technique is only suited for small sized osteochondral defects (max 4 cm2). - osteochondral allograft (4): there is no limitation on the defect size. For extremely large defects even massive allografts can be used to replace an entire condyle or trochlea. On the contrary since allogenous tissue is used, this treatment strategy has a risk on poor graft incorporation and subsidence of the construct. Furthermore concerns on higher infection rates remain. The availability of fresh allografts is another major problem, causing a rather limited use in Europe (5). - Sandwich technique (6): the aim of this technique is to reconstruct both phases of the osteochondral defect seperately by means of impacted (autologous) bone grafts in the bottom of the defect, which are subsequently covered with either autologous chondrocytes (arthroscopically harvested in a previous surgical session and next cultured and grown in a lab), either minced cartilage fragments. This second phase can be covered with a collagen membrane or the cells are seeded on a scaffold. Autologous chondrocty implantation is an expensive technique, needs 2 sessions of surgery and has limited availability. In Belgium reimbursement for this technique was terminated in 2016 (7). The minced cartilage technique can again only be applied in smaller defects. - Autologous matrix induced chondrogenenis (AMIC) (8) is an alternative technique using only the impaction of bone grafts in the defect and subsequent covering with a collagen membrane. This strategy relies on the recruitment of mesenchymal stem cells (MSC) that promote the formation of fibrocartilage. This fibrocartilage does not express the same properties as hyaline articular cartilage and often leads to genesis of intralesional osteophytes and early degeneration. - Osteochondral scaffolds (9,10): These cellfree biomaterials likewise employ recruitment of MSCs and corresponding generation of fibrocartilage. Such scaffolds are once again expensive and questions remain about their efficacy. The formerly described techniques result in fair clinical results (lower osteoarthritis rates compared with untreated groups), but the enumerated shortcomings imply that the treatment of large osteochondral defects remains suboptimal in clinical practice resulting in the use of prosthetic solutions in rather young patients. This exposes the patient to particular risks of prosthetic surgery including loosening, stiffness, infection and chronic pain. From Orthopride, the Belgian knee prosthesis Registry (11), it appears among others that the mean age for revision knee arthroplasty is lower than the mean age for primary knee arthroplasties, signifying a considerable amount of young patients with a knee prosthesis who are dissatisfied. As such prevention of premature prosthetic surgery through a delay of the arthritic process remains an important goal for knee surgeons. As a result of the shortcomings of the currently available techniques, an important and promising role is anticipated in this process for ‘Tissue Engineering’ (TE) techniques (12). Aim(s) of the project Development of an autologous biological osteochondral implant using tissue engineering in a large animal model with ultimately translation to clinical application on human osteochondral defects. The resulting implant ideally includes the following properties: - Autologous tissue without donor site morbidity - Regeneration of both chondral and osseous phases + formation of a tidemark - Possibility to develop patient and defect specific implant - Stable fixation and structural support to facilitate early rehabilitation - Simple and single stage surgical procedures Research plan and methodology of the project Supported by Prometheus, the musculoskeletal Tissue engineering division of KU Leuven, a new osteochondral implant will be developed based on TE. This research project receives ‘competitive funding’ from Regmed XB en JOINTPROMISE (Horizon 2020). Recent insights from this research group concerning the development and growing of the chondrocyte phase in a small animal model enabled further progression to apply the technique in larger animal models (13,14). In this research project, this TE technique will be used in Aachener Minipigs (15) as in this animal model important similarities exist with humane subchondral bone. First, a critical size defect (a defect that cannot heal spontaneously) will be created in the minipig trochlea. Different strategies to recreate the subchondral bone will be evaluated in combination with the tissue-engineered autologous chondrocyte phase units, called micromasses. Once a ‘proof of concept’ is achieved, this technique will be applied on a condylar model in the minipig knee/stifle joint. When this condylar model leads to a ‘proof of concept’, the technique can finally be translated to human clinical application. During the different phases of the project, different aspects that can influence this translation will be critically evaluated and corrected if necessary (transport, handling, instruments, …). Workplan and timeline Work package 0 6-12 months: Review of the currently available literature (systematic review) on the outcomes of surgical treatment of deep osteochondral defects. Retrospective study on the results of surgical treatment of osteochondral defects of the knee in UZ Leuven. Retrospective study on a prospective cohort of patients treated with a focal metal implant (Episealer) for a large osteochondral defect. These studies have an important role in supporting the statement of an unmet medical need for deep and large osteochondral defects. Work package 1 6-12 months: Optimalisation of the animal model and trochlear defect model: preliminary data were collected in a previous stage. Evaluation of radiological, macroscopical and histological results of empty defects, osteochondral autografts and defects treated with a 2nd generation implant (copios or ceramics + micromass) . This includes short (3 months) and longer (12 months) term results. Work package 2 12-18 months: Evaluation of radiological, macroscopical and histological results of a 3th generation fully cell-based implant. Preliminary data collection is in process. Work package 3 24-36 months: Upscaling of the defect to medial femoral condyle and increase size (8mm to 12-15mm). Work package 4 36-48 months: Clinical translation - optimalisation of transport and logistical points of attention - optimalisation of surgical technique pivotal elements - preparation GMP (good manufacturing process) in collaboration with Anicells. References 1. 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