Glycosaminoglycan-based hydrogels with tunable sulfation and degradation for anti-inflammatory small molecule delivery

Abstract

Event Abstract Back to Event Glycosaminoglycan-based hydrogels with tunable sulfation and degradation for anti-inflammatory small molecule delivery Liane Tellier1*, Yifeng Peng1* and Johnna S. Temenoff1, 2* 1 Georgia Institute of Technology and Emory University, Coulter Department of Biomedical Engineering, United States 2 Georgia Institute of Technology, Parker H. Petit Institute for Bioengineering and Bioscience, United States Introduction: Crystal violet (CV) is a small molecule shown to have anti-bacterial, anti-fungal & more recently, anti-inflammatory properties in part by acting as a Nox & angiopoietin-2 (ang-2) inhibitor[1]. Though treatment for many inflammatory conditions would ideally be delivered with zero order release kinetics, this remains particularly challenging for small molecule drugs such as CV. Therefore, to tune CV release kinetics we have developed a CV-loaded glycosaminoglycan (GAG)-based hydrogel system with heparin (a highly sulfated GAG) of varying sulfation levels (affects CV affinity) & with varying amounts of degradable crosslinker (affects hydrogel degradation). It is hypothesized that more sulfated heparin hydrogels will release bioactive CV over a longer period than more desulfated heparin hydrogels & that varying hydrogel degradation will allow us to further tune the rate of CV release. Methods: Hydrogels were prepared via free radical polymerization with 90 wt% water, 9 wt% PEG diacrylate (PEGDA), 1 wt% heparin, & 0, 30, or 42 mol% hydrolytically degradable crosslinker (DTT) for non-, slow-, & fast-degrading hydrogels, respectively. N-desulfated (Hep-N), 6O,N-desulfated (Hep-N,-6O) & fully desulfated (Hep-) heparin were prepared & functionalized with methacrylamide groups. CV was preloaded (20mg) into the hydrogel precursor prior to crosslinking. To quantify CV release, hydrogel supernatant was removed & the amount of CV was measured with a plate reader (absorption at 590nm) every 2 days. CV bioactivity was assessed by incubating hydrogel supernatant diluted to ~20μM with bEnd.3 cells & quantifying bEnd.3 ang-2 gene expression via PCR. Results: CV release kinetics were dependent on heparin sulfation level, whereby more sulfated heparin hydrogels (Hep&Hep-N) exhibited a linear release profile, whereas more desulfated hydrogels (Hep-N,-6O&Hep-) released significantly more CV at Day 1, followed by more plateaued release over 9-15 days (Fig1). Moreover, slow-degrading hydrogels required at least 2 additional days to release an equivalent amount of CV as fast-degrading hydrogels with the same heparin derivative. Finally, after incubating hydrogel releasate with bEnd.3 cells, bEnd.3 ang-2 gene expression was significantly lower than the no treatment control & was not significantly different from the 20μM soluble CV, indicating that CV maintained its bioactivity after release from hydrogels (Fig2). Conclusion: We have fabricated GAG-based hydrogels with heparin derivatives of varying sulfation levels (alters CV release kinetics) & with varying concentrations of DTT (alters hydrogel degradation & CV release rate). In this way, we have developed a highly tailorable hydrogel platform & have achieved near zero-order release of bioactive CV in particular formulations. Thus, in the future, this system can be utilized in vivo with the unique advantage of releasing small molecule anti-inflammatories such as CV in a sustained manner over a period of weeks. We would like to acknowledge NIH R01 AR063692.