A hyaluronic acid-based hydrogel enabling CD44-mediated chondrocyte binding and gapmer oligonucleotide release for modulation of gene expression in osteoarthritis

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Abstract

Hyaluronic acid (HA) is an attractive biomaterial for osteoarthritis (OA) treatment due to inherent functional and compatibility properties as an endogenous knee joint component. In this work, we describe a HA-based hydrogel with the dual functionality of increased CD44-dependent chondrocyte binding and controlled release of gapmer antisense oligonucleotides for unassisted cellular entry and subsequent gene silencing activity. A Schiff base-mediated gelation method was used to produce a panel of hydrogels varying in the aldehyde-modified HA (900 kDa) to chitosan ratios (3:7, 5:5 and 7:3) for identifying designs displaying optimal engagement of OA patient-derived CD44-expressing chondrocytes. Correlation was found between cell binding and CD44 expression, with maximal binding exhibited at a HA/chitosan ratio of 7:3, that was 181% higher than CD44-negative MCF-7 cell control cells. Transfection agent-free uptake into OA chondrocytes of fluorescent 13-mer DNA oligonucleotides with a flanked locked nucleic acid (LNA) gapmer design, in contrast to naked siRNA, was demonstrated by confocal and flow cytometric analysis. A sustained and complete release over 5 days was found with the 7:3 hydrogel, in contrast, the 5:5 and 3:7 hydrogel released 60% and 43% of loaded gapmers, respectively over the same period. A COX-2-specific gapmer designed with maximal chondrocyte gene silencing (˜ 70% silencing efficiency at 500 nM compared with a mismatch gapmer sequence) resulted in effective COX-2 silencing over 14 days in hydrogels seeded with OA chondrocytes, with significant difference exhibited between day 3 and 10. This work introduces a novel HA-based CD44-mediated cellular binding and gapmer controlled release platform to modulate cellular gene expression.

Graphical abstract

Bind and repair chondrocytes within HA-based hydrogel.

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