Biomimetic Molecules Lower Catabolic Expression and Prevent Chondroitin Sulfate Degradation in an Osteoarthritic ex Vivo Model

Abstract

Aggrecan, the major proteoglycan in cartilage, serves to protect cartilage tissue from damage and degradation during the progression of osteoarthritis (OA). In cartilage extracellular matrix (ECM) aggrecan exists in an aggregate composed of several aggrecan molecules that bind to a single filament of hyaluronan. Each molecule of aggrecan is composed of a protein core and glycosaminoglycan sides chains, the latter of which provides cartilage with the ability to retain water and resist compressive loads. During the progression of OA, loss of aggrecan is considered to occur first, after which other cartilage matrix components become extremely susceptible to degradation. Proteolytic cleavage of the protein core of aggrecan by enzymes such as aggrecanases, prevent its binding to HA and lower cartilage mechanical strength. Here we present the use of HA-binding or collagen type II-binding molecules that functionally mimic aggrecan but lack known cleavage sites, protecting the molecule from proteolytic degradation. These molecules synthesized with chondroitin sulfate backbones conjugated to hyaluronan-or collagen type IIbinding peptides, are capable of diffusing through a cartilage explant and adhering to the ECM of this tissue. The objective of this study was to test the functional efficacy of these molecules in an ex vivo osteoarthritic model to discern the optimal molecule for further studies. Different variations of chondroitin sulfate conjugated to the binding peptides were diffused through aggrecan depleted explants and assessed for their ability to enhance compressive stiffness, prevent CS degradation, and modulate catabolic (MMP-13 and ADAMTS-5) and anabolic (aggrecan and collagen type II) gene expression. A pilot in vivo study assessed the ability to retain the molecule within the joint space of an osteoarthritic guinea pig model. The results indicate chondroitin sulfate conjugated to hyaluronan-binding peptides is able to significantly restore equilibrium modulus and prevent CS degradation. All molecules demonstrated the ability to lower catabolic gene expression in aggrecan depleted explants. In order to enhance biosynthesis and regeneration, the molecules need to be coupled with an external stimulant such as a growth factor. The chondroitin sulfate molecule synthesized with HA-binding peptides demonstrated adherence to

Biomimetic proteoglycans can molecularly engineer early osteoarthritic cartilage in vivo

“…A number of studies have investigated the diffusion of molecules into the articular cartilage ECM as treatments or drug delivery vehicles . Work done by other groups demonstrated diffusion of a positively charged molecule (66 kDa avidin ex vivo and in vivo) a large neutral molecule (500 kDa dextran ex vivo), and antibodies (150 kDa goat‐anti‐mouse antibodies ex vivo) into articular cartilage.…”

Glycosaminoglycan and Proteoglycan Biotherapeutics in Articular Cartilage Protection and Repair Strategies: Novel Approaches to Visco‐supplementation in Orthobiologics

“…This peptidoglycan aggrecan mimetic (mAGC) containing CS chains with attached HABPs replicated the HA-binding properties of native aggrecan ensuring that GAG localization was maintained within the scaffold. [204] mAGC had superior water regain properties to that of native aggrecan and mAGC constructs had superior compressive strength (78% increase), The catabolism of mAGC constructs by MMP-13 and ADAMTS-5 [206,207] was also reduced compared to in animal models of OA. AFM demonstrated that the attached peptide chains in mAGC assumed bottle-brush-type orientations on the CS-core structure similar to the CS chain arrangements in native aggrecan core protein.…”

“…[205] Furthermore, localization of mAGC in the scaffold also stimulated synthesis of type II collagen. [206,208] A peptidoglycan lubricin-mimetic, mLUB15 has also been prepared by similar methodology to that of mAGC using HA binding peptide (GAHWQFNALTVRGGGC) and a type II collagen binding peptide (WYRGRL) attached to the CS backbone [209] (Figure 11b). [204] mAGC had superior water regain properties to that of native aggrecan and mAGC constructs had superior compressive strength (78% increase), The catabolism of mAGC constructs by MMP-13 and ADAMTS-5 [206,207] was also reduced compared to in animal models of OA.…”

“…AFM demonstrated that the attached peptide chains in mAGC assumed bottle-brush-type orientations on the CS-core structure similar to the CS chain arrangements in native aggrecan core protein. [204] mAGC had superior water regain properties to that of native aggrecan and mAGC constructs had superior compressive strength (78% increase), The catabolism of mAGC constructs by MMP-13 and ADAMTS-5 [206,207] was also reduced compared to in animal models of OA. [206,208] A peptidoglycan lubricin-mimetic, mLUB15 has also been prepared by similar methodology to that of mAGC using HA binding peptide (GAHWQFNALTVRGGGC) and a type II collagen binding peptide (WYRGRL) attached to the CS backbone [209] (Figure 11b).…”

Biomimetic proteoglycans diffuse throughout articular cartilage and localize within the pericellular matrix

“…We (Phillips et al, 2018;Prudnikova et al, 2017;Prudnikova et al, 2018;Sarkar, 2011;Sarkar et al, 2017;Sarkar, Lightfoot-Vidal, Schauer, Vresilovic, & Marcolongo, 2012) and others (Bernhard & Panitch, 2012;Place, Kelly, & Kipper, 2014;Sharma, Panitch, & Neu, 2013;Sharma, Vazquez-Portalatin, Calve, & Panitch, 2016) have been investigating mimics to proteoglycans. We designed, synthesized and characterized biomimetic proteoglycans (BPGs) that mimic the molecular architecture and physical properties of natural proteoglycans.…”