pH-responsive glycopolymeric nanoparticles for targeted drug delivery to macrophages

Introduction:: Macrophages perform an essential role in the development of a host of inflammatory diseases and therefore serve as vital targets to treat these diseases. While therapeutics designed to modulate macrophage activity have been promising, they tend to suffer from a lack of specificity that results in off-target toxicities. Delivery vehicles that can preferentially convey these therapeutics to macrophages are therefore needed to improve the bioavailability and effectiveness of these drugs. Here, we develop macrophage-targeting nanoparticles (NPs) which feature a polymeric mannose corona for high affinity binding and internalization by macrophages via the mannose receptor. The NPs also feature a pH-responsive core capable of spatiotemporally controlled drug release. These unique properties endow the NPs with tremendous potential to actively target macrophages in inflammatory disease.


Materials and Methods:: Diblock copolymers comprising of poly((2-Diisopropylamino)ethyl methacrylate) (PDPA) and poly(methacrylamidomannose) (PMAM) were synthesized through reversible addition fragmentation transfer (RAFT) polymerization. The PDPA block enables hydrophobic drug loading and pH-triggered release while the PMAM block enables NP stability and macrophage targeting. PDPA-b-PMAG and PDPA-b-PEGMA diblock copolymers were also synthesized as control polymers. NPs were fabricated using the nanoprecipitation/dialysis method and their physicochemical properties characterized via DLS. The pH-responsiveness of the NPs was assessed by incubating fluorescent dye-loaded NPs at varying pHs (5.0-7.4) and measuring the loss of fluorescence with time. Macrophage internalization was assessed by incubating the NPs with macrophages for 2h then measuring uptake through flow cytometry. The mechanism of uptake of the PDPA-b-PMAM NPs was determined by performing blocking experiments with D-mannose. Internalization studies in 4T1 breast tumor cells were performed to further probe the specificity of the PDPA-b-PMAM NPs.

Results, Conclusions, and Discussions:: Results: The PDPA-b-PMAM copolymers had a molecular weight of 41kDa (M_n­) and formed NPs ~130nm in size. The NPs showed a burst release of cargo at pHs below the pKa of PDPA (6.3) but remained intact at physiological pH. The PDPA-b-PMAM NPs showed the highest uptake in macrophages with a 1.4-fold and 12.2-fold increase over the PDPA-b-PMAG and PDPA-b-PEGMA NPs respectively. Blocking the mannose receptor led to a 1.4-fold decrease in macrophage internalization of the PDPA-b-PMAM NPs while no significant differences were observed in the internalization of PDPA-b-PMAG and PDPA-b-PEGMA NPs. Furthermore, the PDPA-b-PMAM NPs showed the lowest internalization of all NP formulations in 4T1 breast cancer cells indicating a preference for cells which express the mannose receptor.

Conclusion: Delivery vehicles capable of targeting macrophages in inflammation are a prominent need to improve the efficacy of macrophage-modulating therapies against inflammatory diseases. To address this need, we fabricated PDPA-b-PMAM NPs that show tremendous promise as macrophage-targeting drug delivery vehicles. The NPs showed the ability to release cargo in a pH-dependent manner enabling the spatiotemporal control of drug delivery. The PDPA-b-PMAM NPs also show superior internalization in macrophages compared to PDPA-b-PMAG and PDPA-b-PEGMA NPs. More so, internalization by macrophages was shown to be dependent on the mannose receptor. In addition, the PDPA-b-PMAM NPs show a specificity for cells that express the mannose receptor as they had the lowest internalization in 4T1 breast cancer cells which do not express the mannose receptor.


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