ABSTRACT
AR-12 has been evaluated in clinical trials as an anti-cancer agent but also has demonstrated host-directed, broad-spectrum clearance of bacteria. We have previously shown that AR-12 has activity in vitro against Salmonella enterica serovar Typhimurium and Francisella species by inducing autophagy and other host immune pathways. AR-12 treatment of S. Typhimurium-infected mice resulted in a 10-fold reduction in bacterial load in the liver and spleen and an increased survival time. However, AR-12 treatment did not protect mice from death, likely due poor formulation. In the current study, AR-12 was encapsulated in a microparticulate carrier formulated from the novel degradable biopolymer acetalated dextran (Ace-DEX) and subsequently evaluated for its activity in human monocyte-derived macrophages (hMDMs). Our results show that hMDMs efficiently internalized Ace-DEX microparticles (MPs), and that encapsulation significantly reduced host cell cytotoxicity compared to unencapsulated AR-12. Efficient macrophage internalization of AR-12 loaded MPs (AR-12/MPs) was further demonstrated by autophagosome formation that was comparable to free AR-12 and resulted in enhanced clearance of intracellular Salmonella. Taken together, these studies provide support that Ace-DEX encapsulated AR-12 may be a promising new therapeutic agent to control intracellular bacterial pathogens of macrophages by targeting delivery and reducing drug toxicity.
Subject(s)
Anti-Bacterial Agents/administration & dosage , Dextrans/chemistry , Drug Carriers/chemistry , Pyrazoles/administration & dosage , Salmonella typhimurium/drug effects , Sulfonamides/administration & dosage , Acetals/chemistry , Anti-Bacterial Agents/pharmacology , Autophagy/drug effects , Blotting, Western , Cell Survival/drug effects , Cells, Cultured , Drug Compounding , Humans , Macrophages/drug effects , Macrophages/microbiology , Microscopy, Electron, Scanning , Microscopy, Fluorescence , Microtubule-Associated Proteins/metabolism , Pyrazoles/pharmacology , Salmonella Infections/drug therapy , Salmonella Infections/microbiology , Sulfonamides/pharmacology , Surface PropertiesABSTRACT
Electrospun acetalated dextran (Ac-DEX) scaffolds were fabricated to encapsulate resiquimod, an immunomodulatory toll-like-receptor (TLR) agonist. Ac-DEX has been used to fabricate scaffolds for sustained and temporal delivery of therapeutics because it has tunable degradation rates that are dependent on its synthesis reaction time or the molecular weight of dextran. Additionally, as opposed to commonly electrospun polyesters that shift the local pH upon degradation, the degradation products of Ac-DEX are pH-neutral: dextran, an alcohol, and the metabolic byproduct acetone. Formulations of Ac-DEX with two different degradation rates were used in this study. The effects of electrospinning conditions on the scaffold size and morphology were examined as well as fibroblast adhesion as imaged with fluorescence microcopy and scanning electron microscopy. Macrophage (MΦ) viability further indicates that the scaffolds are cytocompatible. Also, the controlled release profiles of resiquimod from loaded scaffolds and nitric oxide (NO) production by MΦ incubated with these scaffolds show the potential for Ac-DEX scaffolds to be used to temporally and efficiently deliver therapeutics. Overall, we present a novel scaffold that can have tunable and unique drug release rates for tissue engineering, drug delivery, immunomodulation, and wound healing applications.
