Periodic Reporting for period 1 - NovelKnees (Targeting Subchondral Bone Marrow Lesions, Microdamage and Remodelling using Novel Bisphosphonate-Loaded Delivery Systems to Prevent Post-Traumatic Osteoarthritis after Acute Knee Injury)
Reporting period: 2018-05-01 to 2020-04-30
The overall problem that was to be addressed by 'NovelKnees' was to “to understand why Anterior Cruciate Ligament (ACL) ruptures so frequently lead to PTOA, and determine the role of subchondral bone microdamage and BMLs in that process - and whether this can be harnessed for a new treatment strategy”.
The proposed work is important for society since injury/disease of skeletal joints is a significant, and growing, problem among all age-groups (especially younger cohorts) and musculoskeletal isssues are a leading cause of pain and disability.
Here, Objective 1 set out to determine the relationship between bone microdamage and joint disease that follows ACL rupture. We determined that a close relationship exists between these two things.
Objective 2 was to determine the biological consequences of subchondral microdamage and Bone Marrow Lesions (BMLs) in term of localized pathological bone activity. We determined that localized increases in osteoclast activity occur in response to subchondral microdamage.
Objective 3 was to develop an intra-articular drug-delivery system to target damaged subchondral bone via sustained release of therapeutic microparticles. We developed alginate based microparticles that successfully release bone targeting agents which reduce likelihood of PTOA development in model systems.
This project set out to develop tissue-specific early intervention therapies to prevent PTOA after acute joint injury and to determine whether subchondral microdamage caused by ACL injury, increases remodeling which overlaps with BMLs, and stimulates a positive feedback-loop of catabolic events resulting in PTOA via bone- cartilage crosstalk.
In conclusion, this action demonstrated that sustained release of bisphosphonates in a suitable carrier can cross the osteochondral junction to inhibit bone remodelling and the catabolic cycle in cartilage and thus has potential to prevent PTOA. This goal was achieved through the development of bone targeting treatment system used in a preclinical joint injury system.
The proposed work is important for society since injury/disease of skeletal joints is a significant, and growing, problem among all age-groups (especially younger cohorts) and musculoskeletal isssues are a leading cause of pain and disability.
Here, Objective 1 set out to determine the relationship between bone microdamage and joint disease that follows ACL rupture. We determined that a close relationship exists between these two things.
Objective 2 was to determine the biological consequences of subchondral microdamage and Bone Marrow Lesions (BMLs) in term of localized pathological bone activity. We determined that localized increases in osteoclast activity occur in response to subchondral microdamage.
Objective 3 was to develop an intra-articular drug-delivery system to target damaged subchondral bone via sustained release of therapeutic microparticles. We developed alginate based microparticles that successfully release bone targeting agents which reduce likelihood of PTOA development in model systems.
This project set out to develop tissue-specific early intervention therapies to prevent PTOA after acute joint injury and to determine whether subchondral microdamage caused by ACL injury, increases remodeling which overlaps with BMLs, and stimulates a positive feedback-loop of catabolic events resulting in PTOA via bone- cartilage crosstalk.
In conclusion, this action demonstrated that sustained release of bisphosphonates in a suitable carrier can cross the osteochondral junction to inhibit bone remodelling and the catabolic cycle in cartilage and thus has potential to prevent PTOA. This goal was achieved through the development of bone targeting treatment system used in a preclinical joint injury system.
In workpackage (WP) 1 we used our novel joint injury model to examine knee joints by MRI and histological analyses for microdamage. Our experience with this process allowed us to qualitatively determine when a test has been successful due to the change in load/displacement readout during the test. However, MRI imaging can definitely assess ACL status. We showed, using sagittal MRI images that all injured groups had MRI confirmed ACL ruptures. We then used T2 weighted fat-supressed imaging to determine the extent of subchondral bone damage (specifically in the form of BMLs) caused by injury. The injured knees display increased signal in the subchondral compartments of both femur and tibia bones. We used histology to assess levels of bone microdamage caused by ACL injury. We found significantly increased level of microdamage in the injured knees, as measured by crack density, and that cracks and BMLs were significantly correlated.
The primary aim of WP2 was to build on WP1 by developing our ability to evaluate the level of co-localization of subchondral microdamage with osteoclast activity which occurs after ACL injury. In carrying out this work we determined that a system of microdamage-mediated bone turnover exists in the subchondral compartment, and that these changes do indeed correlate with subsequent cartilage degeneration. Specifically, we tested a potent bisphosphonate (ZOL) at two different time points post- injury. In addition a fluorochrome bone labeling agent (Calcein) was also given on the day of injury and 3 days prior the end of the experiment. Injury resulted in increased uptake of calcein in the subchondral region. Early ZOL administration blocked calcein uptake almost completely, whereas late administration brought uptake back up to control levels. Finally, we assessed the effect of ZOL treatment, at two different time points after injury, on articular cartilage health and thus PTOA development. We showed that in comparison with Vehicle treatment, ZOL treatment prevented proteoglycan loss to a significant, and moderate level, respectively. This suggests an osteoclast-independent mechanism may be responsible for this aspect of the injury/disease paradigm.
The primary aim of WP3 was to build on the advances made in the previous WPs by developing and characterizing bisphosphonate-loaded alginate microparticles for injectable delivery to the knee joint, targeting subchondral bone, after knee injury to prevent PTOA. This enhances efficacy of the drug, reducing the number of administrations of the drug and minimizing off-target effects. Alginate-based microparticles are excellent candidates for IA drug-delivery because of their hydrophilic nature, biocompatibility, and physical architecture. We made poly(lactic-co-glycolic) (PLGA) microparticles to encapsulate ZOL. Their size and release profile was controlled by adjusting spray drying and material parameters. Release of ZOL as a function of time was measured. We also tested whether the ZOL agent could be reliably and reproducibly delivered by intra-articular (IA) injection to the joint. Here we made use of a ZOL analogue which was tagged with a fluorescent moiety in order to trace the location of the drug in the joint after delivery. Specifically, we wished to test whether ZOL would penetrate the osteochondral junction (from the articular side), to reach the subchondral bone compartment beneath the joint and thus have the intended therapeutic effect on local osteoclastic activity.
As an overview, our results show that increased subchondral microdamage in the injured knees, is related to BMLs and localized remodeling. Furthermore inhibition of this process by bisphosphonates in microparticle carriers is a feasible drug delivery approach.These data can be exploited in terms of a novel early treatment after joint injury, early data has been disseminated at research meeting, journal publications and public health seminars.
The primary aim of WP2 was to build on WP1 by developing our ability to evaluate the level of co-localization of subchondral microdamage with osteoclast activity which occurs after ACL injury. In carrying out this work we determined that a system of microdamage-mediated bone turnover exists in the subchondral compartment, and that these changes do indeed correlate with subsequent cartilage degeneration. Specifically, we tested a potent bisphosphonate (ZOL) at two different time points post- injury. In addition a fluorochrome bone labeling agent (Calcein) was also given on the day of injury and 3 days prior the end of the experiment. Injury resulted in increased uptake of calcein in the subchondral region. Early ZOL administration blocked calcein uptake almost completely, whereas late administration brought uptake back up to control levels. Finally, we assessed the effect of ZOL treatment, at two different time points after injury, on articular cartilage health and thus PTOA development. We showed that in comparison with Vehicle treatment, ZOL treatment prevented proteoglycan loss to a significant, and moderate level, respectively. This suggests an osteoclast-independent mechanism may be responsible for this aspect of the injury/disease paradigm.
The primary aim of WP3 was to build on the advances made in the previous WPs by developing and characterizing bisphosphonate-loaded alginate microparticles for injectable delivery to the knee joint, targeting subchondral bone, after knee injury to prevent PTOA. This enhances efficacy of the drug, reducing the number of administrations of the drug and minimizing off-target effects. Alginate-based microparticles are excellent candidates for IA drug-delivery because of their hydrophilic nature, biocompatibility, and physical architecture. We made poly(lactic-co-glycolic) (PLGA) microparticles to encapsulate ZOL. Their size and release profile was controlled by adjusting spray drying and material parameters. Release of ZOL as a function of time was measured. We also tested whether the ZOL agent could be reliably and reproducibly delivered by intra-articular (IA) injection to the joint. Here we made use of a ZOL analogue which was tagged with a fluorescent moiety in order to trace the location of the drug in the joint after delivery. Specifically, we wished to test whether ZOL would penetrate the osteochondral junction (from the articular side), to reach the subchondral bone compartment beneath the joint and thus have the intended therapeutic effect on local osteoclastic activity.
As an overview, our results show that increased subchondral microdamage in the injured knees, is related to BMLs and localized remodeling. Furthermore inhibition of this process by bisphosphonates in microparticle carriers is a feasible drug delivery approach.These data can be exploited in terms of a novel early treatment after joint injury, early data has been disseminated at research meeting, journal publications and public health seminars.
Among my goals for this proposal was to establish a world-class research group in the area of joint injury and PTOA, and also to take new approaches to important problems in musculoskeletal health. Our ideas on the involvement of subchondral bone in PTOA, especially in the early stage have become an important part of the research agenda in recent years. This is evident by the increased number of papers that have been published which consider these factors. It is also evident by the fact that tI have ben invited to give numerous workshops and talks in that last 5 years relating to this topic. This will continue to have impacts within the field in the future, but also to have wider socio-economic impacts as new treatment ideas begin to emerge and focus on these new pathways for treatment which we and others have discovered. This fellowship has allowed me to develop the therapeutic, applied and translational aspects of my research. I have learned new skills in the development of biomaterials and drug-delivery systems. This has been beneficial for European healthcare, but also for me when it comes to reintegrating in the European research community as an expert researcher. In line with the objectives of the MSCA-IF programme I have built significantly on my existing skill-set, the advanced training and support available at RCSI to become a better all round scientist and group leader.