The use of recombinant adeno-associated virus for skeletal gene therapy.
Dai Juan,Rabie A Bakr M
Orthodontics & craniofacial research
OBJECTIVES:To provide a comprehensive literature review describing recent developments of the recombinant adeno-associated virus (rAAV) vector and exploring the therapeutic application of rAAV for bone defects, cartilage lesions and rheumatoid arthritis. DESIGN:Narrative review. RESULT:The review outlines the serotypes and genome of AAV, integration and life cycle of the rAAV vectors, the immune response and regulating system for AAV gene therapy. Furthermore, the advancements of rAAV gene therapy for bone growth together with cartilage repair are summarized. CONCLUSION:Recombinant adeno-associated virus vector is perceived to be one of the most promising vector systems for bone and cartilage gene therapy approaches and further investigations need to be carried out for craniofacial research.
10.1111/j.1601-6343.2007.00381.x
Modulation of the Wnt pathway through inhibition of CLK2 and DYRK1A by lorecivivint as a novel, potentially disease-modifying approach for knee osteoarthritis treatment.
Osteoarthritis and cartilage
OBJECTIVES:Wnt pathway upregulation contributes to knee osteoarthritis (OA) through osteoblast differentiation, increased catabolic enzymes, and inflammation. The small-molecule Wnt pathway inhibitor, lorecivivint (SM04690), which previously demonstrated chondrogenesis and cartilage protection in an animal OA model, was evaluated to elucidate its mechanism of action. DESIGN:Biochemical assays measured kinase activity. Western blots measured protein phosphorylation in human mesenchymal stem cells (hMSCs), chondrocytes, and synovial fibroblasts. siRNA knockdown effects in hMSCs and BEAS-2B cells on Wnt pathway, chondrogenic genes, and LPS-induced inflammatory cytokines was measured by qPCR. In vivo anti-inflammation, pain, and function were evaluated following single intra-articular (IA) lorecivivint or vehicle injection in the monosodium iodoacetate (MIA)-induced rat OA model. RESULTS:Lorecivivint inhibited intranuclear kinases CDC-like kinase 2 (CLK2) and dual-specificity tyrosine phosphorylation-regulated kinase 1A (DYRK1A). Lorecivivint inhibited CLK2-mediated phosphorylation of serine/arginine-rich (SR) splicing factors and DYRK1A-mediated phosphorylation of SIRT1 and FOXO1. siRNA knockdowns identified a role for CLK2 and DYRK1A in Wnt pathway modulation without affecting β-catenin with CLK2 inhibition inducing early chondrogenesis and DYRK1A inhibition enhancing mature chondrocyte function. NF-κB and STAT3 inhibition by lorecivivint reduced inflammation. DYRK1A knockdown was sufficient for anti-inflammatory effects, while combined DYRK1A/CLK2 knockdown enhanced this effect. In the MIA model, lorecivivint inhibited production of inflammatory cytokines and cartilage degradative enzymes, resulting in increased joint cartilage, decreased pain, and improved weight-bearing function. CONCLUSIONS:Lorecivivint inhibition of CLK2 and DYRK1A suggested a novel mechanism for Wnt pathway inhibition, enhancing chondrogenesis, chondrocyte function, and anti-inflammation. Lorecivivint shows potential to modify structure and improve symptoms of knee OA.
10.1016/j.joca.2019.05.006
A novel Wnt pathway inhibitor, SM04690, for the treatment of moderate to severe osteoarthritis of the knee: results of a 24-week, randomized, controlled, phase 1 study.
Yazici Y,McAlindon T E,Fleischmann R,Gibofsky A,Lane N E,Kivitz A J,Skrepnik N,Armas E,Swearingen C J,DiFrancesco A,Tambiah J R S,Hood J,Hochberg M C
Osteoarthritis and cartilage
OBJECTIVE:To assess the safety, pharmacokinetics, and exploratory efficacy of SM04690, a novel Wnt pathway inhibitor, as a potential disease modifying treatment for knee osteoarthritis (OA). DESIGN:Subjects with Kellgren-Lawrence grade 2-3 knee OA were randomized in successive dose-escalation cohorts to receive a knee intra-articular (IA) injection with 0.03, 0.07, or 0.23 mg SM04690, or placebo (PBO) (4:1 ratio). Safety, pharmacokinetics, efficacy (WOMAC Total/Function/Pain, Pain VAS, Physician Global Assessment [MDGA], and OMERACT-OARSI Response), OA-related biomarker (P1NP, ß-CTX, and cartilage oligomeric matrix protein [COMP]), and radiographic/imaging data were collected at baseline and during 24-week follow-up. RESULTS:61 subjects (SM04690 n = 50; PBO n = 11) enrolled. Two dose limiting toxicities (DLTs), increased pain following injection and paroxysmal tachycardia (also the single serious AE), were reported in the 0.07 mg cohort. A total of 72 AEs were reported; Sixteen (occurring in eight subjects) were considered related to study medication. There were three discontinuations; one due to an AE (0.03 mg cohort). Bone marrow edema (BME) remained constant for most subjects. No doses were excluded from further study due to DLT criteria. Plasma levels of SM04690 were below the limit of detection at all time points. At Week 24, improvements from baseline were seen in all cohorts for the exploratory measures WOMAC Total, WOMAC Function, WOMAC Pain, MDGA, Pain VAS, and OMERACT-OARSI response. Joint space width (JSW) improvement was observed in the 0.07 mg cohort (P = 0.02 vs PBO). CONCLUSION:SM04690 appeared safe and well tolerated, with no evidence of systemic exposure. Exploratory efficacy analyses suggested positive trends for measurements of OA pain, function and disease-modifying osteoarthritis drug (DMOAD) properties. CLINICALTRIALS. GOV REGISTRATION:NCT02095548.
10.1016/j.joca.2017.07.006
A small-molecule inhibitor of the Wnt pathway (SM04690) as a potential disease modifying agent for the treatment of osteoarthritis of the knee.
Deshmukh V,Hu H,Barroga C,Bossard C,Kc S,Dellamary L,Stewart J,Chiu K,Ibanez M,Pedraza M,Seo T,Do L,Cho S,Cahiwat J,Tam B,Tambiah J R S,Hood J,Lane N E,Yazici Y
Osteoarthritis and cartilage
OBJECTIVES:Osteoarthritis (OA) is a degenerative disease characterized by loss of cartilage and increased subchondral bone within synovial joints. Wnt signaling affects the pathogenesis of OA as this pathway modulates both the differentiation of osteoblasts and chondrocytes, and production of catabolic proteases. A novel small-molecule Wnt pathway inhibitor, SM04690, was evaluated in a series of in vitro and in vivo animal studies to determine its effects on chondrogenesis, cartilage protection and synovial-lined joint pathology. DESIGN:A high-throughput screen was performed using a cell-based reporter assay for Wnt pathway activity to develop a small molecule designated SM04690. Its properties were evaluated in bone-marrow-derived human mesenchymal stem cells (hMSCs) to assess chondrocyte differentiation and effects on cartilage catabolism by immunocytochemistry and gene expression, and glycosaminoglycan breakdown. In vivo effects of SM04690 on Wnt signaling, cartilage regeneration and protection were measured using biochemical and histopathological techniques in a rodent acute cruciate ligament tear and partial medial meniscectomy (ACLT + pMMx) OA model. RESULTS:SM04690 induced hMSC differentiation into mature, functional chondrocytes and decreased cartilage catabolic marker levels compared to vehicle. A single SM04690 intra-articular (IA) injection was efficacious in a rodent OA model, with increased cartilage thickness, evidence for cartilage regeneration, and protection from cartilage catabolism observed, resulting in significantly improved Osteoarthritis Research Society International (OARSI) histology scores and biomarkers, compared to vehicle. CONCLUSIONS:SM04690 induced chondrogenesis and appeared to inhibit joint destruction in a rat OA model, and is a candidate for a potential disease modifying therapy for OA.
10.1016/j.joca.2017.08.015
Angiopoietin-like 3-derivative LNA043 for cartilage regeneration in osteoarthritis: a randomized phase 1 trial.
Nature medicine
Osteoarthritis (OA) is a common, debilitating, chronic disease with no disease-modifying drug approved to date. We discovered LNA043-a derivative of angiopoietin-like 3 (ANGPTL3)-as a potent chondrogenesis inducer using a phenotypic screen with human mesenchymal stem cells. We show that LNA043 promotes chondrogenesis and cartilage matrix synthesis in vitro and regenerates hyaline articular cartilage in preclinical OA and cartilage injury models in vivo. LNA043 exerts at least part of these effects through binding to the fibronectin receptor, integrin αβ on mesenchymal stem cells and chondrocytes. In a first-in-human (phase 1), randomized, double-blinded, placebo-controlled, single ascending dose, single-center trial ( NCT02491281 ; sponsored by Novartis Pharmaceuticals), 28 patients with knee OA were injected intra-articularly with LNA043 or placebo (3:1 ratio) either 2 h, 7 d or 21 d before total knee replacement. LNA043 met its primary safety endpoint and showed short serum pharmacokinetics, cartilage penetration and a lack of immunogenicity (secondary endpoints). Post-hoc transcriptomics profiling of cartilage revealed that a single LNA043 injection reverses the OA transcriptome signature over at least 21 d, inducing the expression of hyaline cartilage matrix components and anabolic signaling pathways, while suppressing mediators of OA progression. LNA043 is a novel disease-modifying OA drug candidate that is currently in a phase 2b trial ( NCT04864392 ) in patients with knee OA.
10.1038/s41591-022-02059-9
Repair of bone defects in vivo using tissue engineered hypertrophic cartilage grafts produced from nasal chondrocytes.
Bardsley Katie,Kwarciak Agnieska,Freeman Christine,Brook Ian,Hatton Paul,Crawford Aileen
Biomaterials
The regeneration of large bone defects remains clinically challenging. The aim of our study was to use a rat model to use nasal chondrocytes to engineer a hypertrophic cartilage tissue which could be remodelled into bone in vivo by endochondral ossification. Primary adult rat nasal chondrocytes were isolated from the nasal septum, the cell numbers expanded in monolayer culture and the cells cultured in vitro on polyglycolic acid scaffolds in chondrogenic medium for culture periods of 5-10 weeks. Hypertrophic differentiation was assessed by determining the temporal expression of key marker genes and proteins involved in hypertrophic cartilage formation. The temporal changes in the genes measured reflected the temporal changes observed in the growth plate. Collagen II gene expression increased 6 fold by day 7 and was then significantly downregulated from day 14 onwards. Conversely, collagen X gene expression was detectable by day 14 and increased 100-fold by day 35. The temporal increase in collagen X expression was mirrored by increases in alkaline phosphatase gene expression which also was detectable by day 14 with a 30-fold increase in gene expression by day 35. Histological and immunohistochemical analysis of the engineered constructs showed increased chondrocyte cell volume (31-45 μm), deposition of collagen X in the extracellular matrix and expression of alkaline phosphatase activity. However, no cartilage mineralisation was observed in in vitro culture of up to 10 weeks. On subcutaneous implantation of the hypertrophic engineered constructs, the grafts became vascularised, cartilage mineralisation occurred and loss of the proteoglycan in the matrix was observed. Implantation of the hypertrophic engineered constructs into a rat cranial defect resulted in angiogenesis, mineralisation and remodelling of the cartilage tissue into bone. Micro-CT analysis indicated that defects which received the engineered hypertrophic constructs showed 38.48% in bone volume compared to 7.01% in the control defects. Development of tissue engineered hypertrophic cartilage to use as a bone graft substitute is an exciting development in regenerative medicine. This is a proof of principal study demonstrating the potential of nasal chondrocytes to engineer hypertrophic cartilage which will remodel into bone on in vivo transplantation. This approach to making engineered hypertrophic cartilage grafts could form the basis of a new potential future clinical treatment for maxillofacial reconstruction.
10.1016/j.biomaterials.2016.10.014
Gene therapy approaches to regenerating the musculoskeletal system.
Evans Christopher H,Huard Johnny
Nature reviews. Rheumatology
Injuries to the musculoskeletal system are common, debilitating and expensive. In many cases, healing is imperfect, which leads to chronic impairment. Gene transfer might improve repair and regeneration at sites of injury by enabling the local, sustained and potentially regulated expression of therapeutic gene products; such products include morphogens, growth factors and anti-inflammatory agents. Proteins produced endogenously as a result of gene transfer are nascent molecules that have undergone post-translational modification. In addition, gene transfer offers particular advantages for the delivery of products with an intracellular site of action, such as transcription factors and noncoding RNAs, and proteins that need to be inserted into a cell compartment, such as a membrane. Transgenes can be delivered by viral or nonviral vectors via in vivo or ex vivo protocols using progenitor or differentiated cells. The first gene transfer clinical trials for osteoarthritis and cartilage repair have already been completed. Various bone-healing protocols are at an advanced stage of development, including studies with large animals that could lead to human trials. Other applications in the repair and regeneration of skeletal muscle, intervertebral disc, meniscus, ligament and tendon are in preclinical development. In addition to scientific, medical and safety considerations, clinical translation is constrained by social, financial and logistical issues.
10.1038/nrrheum.2015.28
Regeneration of articular cartilage of the knee.
Rodriguez-Merchan E Carlos
Rheumatology international
Cartilage therapy for focal articular lesions of the knee has been implemented for more than a decade, and it is becoming increasingly available. What do we know on the healing response of cartilage lesions? What do we know on the treatment of focal cartilage lesions of the knee and the prognostic factors involved? PubMed articles related to articular cartilage regeneration of the knee in clinical studies were searched from January 2006 to November 2012, using the following key words: articular cartilage, regeneration, clinical studies, and knee. A total of 44 reports were found. They showed the following possibilities for the treatment of focal lesions of the articular cartilage of the knee: cartilage regeneration and repair including cartilage reparation with gene-activated matrices, autologous chondrocyte implantation (ACI) and matrix-induced ACI (MACI), microfracture, osteochondral autograft transfer (mosaicplasty), biological approaches (scaffolds, mesenchymal stem cells-MSCs, platelet-rich plasma, growing factors-GF, bone morphogenetic proteins-BMPs, magnetically labeled synovium-derived cells-M-SDCs, and elastic-like polypeptide gels), osteotomies, stem-cell-coated titanium implants, and chondroprotection with pulsed electromagnetic fields. Untreated cartilage lesions on the femoral condyles had a superior healing response compared to those on the tibial plateaus, and in the patellofemoral joint. Clinical outcome regarding the treatment of medial defects is better than that of the lateral defects. Improvement from baseline was better for patients < or = 30 years compared with patients > or = 30 years. ACI, MACI, and mosaicplasty have shown similar results. The results of comparative clinical studies using ACI have shown some superiority over conventional microfracturing in medium or large defects and in long-term durability. Some biological methods such as scaffolds, MSCs, GF, M-SDCs, BMPs, and elastic-like polypeptide gels still need more research.
10.1007/s00296-012-2601-3
Cell-based articular cartilage repair: the link between development and regeneration.
Caldwell K L,Wang J
Osteoarthritis and cartilage
Clinical efforts to repair damaged articular cartilage (AC) currently face major obstacles due to limited intrinsic repair capacity of the tissue and unsuccessful biological interventions. This highlights a need for better therapeutic strategies. This review summarizes the recent advances in the field of cell-based AC repair. In both animals and humans, AC defects that penetrate into the subchondral bone marrow are mainly filled with fibrocartilaginous tissue through the differentiation of bone marrow mesenchymal stem cells (MSCs), followed by degeneration of repaired cartilage and osteoarthritis (OA). Cell therapy and tissue engineering techniques using culture-expanded chondrocytes, bone marrow MSCs, or pluripotent stem cells with chondroinductive growth factors may generate cartilaginous tissue in AC defects but do not form hyaline cartilage-based articular surface because repair cells often lose chondrogenic activity or result in chondrocyte hypertrophy. The new evidence that AC and synovium develop from the same pool of precursors with similar gene profiles and that synovium-derived chondrocytes have stable chondrogenic activity has promoted use of synovium as a new cell source for AC repair. The recent finding that NFAT1 and NFAT2 transcription factors (TFs) inhibit chondrocyte hypertrophy and maintain metabolic balance in AC is a significant advance in the field of AC repair. The use of synovial MSCs and discovery of upstream transcriptional regulators that help maintain the AC phenotype have opened new avenues to improve the outcome of AC regeneration.
10.1016/j.joca.2014.11.004
Gene- and stem cell-based therapeutics for cartilage regeneration and repair.
Tang Ying,Wang Bing
Stem cell research & therapy
Cell-based regeneration of damaged or diseased articular cartilage still faces significant clinical challenge due to inadequate environmental regulation of stem cell proliferation and chondrogenic differentiation. The role of insulin-like growth factor in critical steps of human bone marrow-derived mesenchymal stem cell chondrogenesis has potential in optimizing the therapeutic use of mesenchymal stem cells in cartilage disorders. In addition to the previously described benefits of recombinant adeno-associated viral vector for in vivo gene therapy, demonstrated by Frisch and colleagues, such vector is also a safe and efficient delivery system for the genetic modification of human bone marrow-derived mesenchymal stem cells via ex vivo insulin-like growth factor 1 gene transfer, so that implanted mesenchymal stem cells continuously release a therapeutic level of insulin-like growth factor 1 to achieve sustained mesenchymal stem cell chondrogenesis for cartilage regeneration.
10.1186/s13287-015-0058-5
Gene therapy for repair and regeneration of bone and cartilage.
Grol Matthew W,Lee Brendan H
Current opinion in pharmacology
Gene therapy refers to the use of viral and non-viral vectors to deliver nucleic acids to tissues of interest using direct (in vivo) or transduced cell-mediated (ex vivo) approaches. Over the past few decades, strategies have been adopted to express therapeutic transgenes at sites of injury to promote or facilitate repair of bone and cartilage. Targets of interest have typically included secreted proteins such as growth factors and anti-inflammatory mediators; however, work has also begun to focus intracellularly on signaling components, transcription factors and small, regulatory nucleic acids such as microRNAs (miRNAs). In recent years, a number of single therapeutic gene approaches (termed 'monotherapies') have proven effective in preclinical models of disease, and several are being evaluated in clinical trials. In particular, an ex vivo TGF-β1 gene therapy was approved in Korea in 2017 for treatment of moderate-to-severe osteoarthritis (OA). The ability to utilize viral vectors for context-specific and combinatorial gene therapy is also being investigated, and these strategies are likely to be important in more robustly addressing the complexities of tissue repair and regeneration in skeletal disease. In this review, we provide an overview of viral gene therapies being developed for treatment of bone and cartilage pathologies, with an emphasis on emerging combinatorial strategies as well as those targeting intracellular mediators such as miRNAs.
10.1016/j.coph.2018.03.005