Gelation chemistries for the encapsulation of nanoparticles in composite gel microparticles for lung imaging and drug delivery

Abstract

The formation of 10-40 μm composite gel microparticles (CGMPs) comprised of ∼100 nm drug containing nanoparticles (NPs) in a poly(ethylene glycol) (PEG) gel matrix is described. The CGMP particles enable targeting to the lung by filtration from the venous circulation. UV radical polymerization and Michael addition polymerization reactions are compared as approaches to form the PEG matrix. A fluorescent dye in the solid core of the NP was used to investigate the effect of reaction chemistry on the integrity of encapsulated species. When formed via UV radical polymerization, the fluorescence signal from the NPs indicated degradation of the encapsulated species by radical attack. The degradation decreased fluorescence by 90% over 15 min of UV exposure. When formed via Michael addition polymerization, the fluorescence was maintained. Emulsion processing using controlled shear stress enabled control of droplet size with narrow polydispersity. To allow for emulsion processing, the gelation rate was delayed by adjusting the solution pH. At a pH = 5.4, the gelation occurred at 3.5 h. The modulus of the gels was tuned over the range of 5 to 50 kPa by changing the polymer concentration between 20 and 70 vol %. NP aggregation during polymerization, driven by depletion forces, was controlled by the reaction kinetics. The ester bonds in the gel network enabled CGMP degradation. The gel modulus decreased by 50% over 27 days, followed by complete gel degradation after 55 days. This permits ultimate clearance of the CGMPs from the lungs. The demonstration of uniform delivery of 15.8 ± 2.6 μm CGMPs to the lungs of mice, with no deposition in other organs, is shown, and indicates the ability to concentrate therapeutics in the lung while avoiding off-target toxic exposure.

Figure 1

Reaction scheme for (a.) radical gelation and (b.) Michael addition gelation. (a.) Aradical (1) attacks the acrylate groups on the PEG triacrylate (2) to form a gel with a polyacrylic acid backbone (3). (b.) The acrylate groups on the PEG triacrylate (2) undergo nucleophilic attack from deprotonated thiols on DL-dithiothreitol (5) to form a sulfur carbon bond (6) and subsequently form a gel (6′). The PEG backbone (4) is denoted as a wavy line.

Figure 2

Fluorescence of EtTP-5 nanoparticles and GFP in sodium acetate buffer or in a solution of 25 vol% PEG in sodium acetate buffer as a function of UV exposure time. (a) EtTP-5 nanoparticles in sodium acetate buffer incubated with no initiator (), IRG (), ACVA (), and AMPA (). (For each point, n=3.) (b) GFP in sodium acetate buffer incubated with no initiator (), IRG (), and ACVA (). (For each point, n=3.) (c) EtTP-5 nanoparticles in a solution of 25 vol% PEG-TA in sodium acetate buffer no initiator (), IRG (), and ACVA (). EtTP-5 nanoparticles in a solution of 25 vol% glycerol ethoxylate in sodium acetate buffer no initiator (). (For each point, n>3.) (d) GFP in a solution of 25 vol% PEG-TA in sodium acetate buffer incubated with no initiator (), IRG (), and ACVA (). (For each point, n>3.)

Figure 3

Images of composite gel microparticles (CGMPs) with 25 vol% PEG triacrylate before and after UV exposure. (a) A droplet containing EtTP-5 nanoparticles (NPs) prior to UV exposure. (b) An CGMP with encapsulated NPs after exposure to 15 minutes of UV using ACVA as the initiator. (c) An CGMP with encapsulated NPs after exposure to 15 minutes of UV using Irgacure as the initiator. (d) An CGMP with encapsulated NPs after exposure to 15 minutes of UV with no initiator. (e) A droplet containing GFP prior to UV exposure. (f) An CGMP with encapsulated GFP after exposure to 15 minutes of UV using ACVA as the initiator. (g) An CGMP with encapsulated GFP after exposure to 15 minutes of UV using Irgacure as the initiator. (h) An CGMP with encapsulated GFP after exposure to 15 minutes of UV with no initiator. (e.-h. To improve contrast for printing purposes, the GFP fluorescence image was enhanced prior to creating the overlayed image by converting all pixels above a desired threshold to red. The unenhanced overlayed images can be found in the supplemental information.)

Figure 4

Control of the gel point and modulus via pH. (a) The gel point of a 40 vol% PEG triacrylate in 30 mM sodium acetate buffer as a function of the solution pH. The longest delay in gelation is observed at a solution pH of 5.40. (For each formulation, n=3.) (b) The modulus of swollen gels made from 40 vol% PEG triacrylate in 30 mM sodium acetate buffer at different pHs. Gels were reacted overnight and then swollen for 24 hours prior to the oscillatory shear measurement. (For each formulation, n=3.)

Figure 5

Gel storage modulus of gels formed via Michael addition under slow gelling conditions in 30 mM sodium acetate buffer solution, pH 4.3, () and under fast gelling conditions in 1 mM triethylamine, pH 11.5, () as a function of PEG triacrylate concentration. (For each formulation, n=1.)

Figure 6

Comparison of storage moduli between swollen gels formed with ACVA, Irgacure and via Michael addition. All gels were formulated with 40 vol% PEG triacrylate either in water for the UV exposed gels or in 30 mM sodium acetate buffer (pH 4.32) for the Michael addition gels. The ACVA and Irgacure samples were exposed to UV for 15 minutes. The Michael addition samples were allowed to react overnight. All samples were swollen in excess DI water for 24 hours prior to the oscillatory shear measurement. (For each formulation, n=3.)

Figure 7

The degradation of a bulk 40 vol% PEG triacrylate gel formed via Michael addition. The modulus drops to 50% of its original value by day 27. (For each point, n=3.)

Figure 8

Images of composite gel microparticles (CGMPs) formed via a Michael addition reaction before and after the reaction. (a) A droplet with EtTP-5 nanoparticles (NPs) prior to a reaction with slow kinetics. (b) A droplet with GFP prior to reaction with slow kinetics. (c) A droplet with NPs prior to a reaction with fast kinetics. (d) An CGMP with NPs post reaction with slow kinetics. Flocculation depletion is observed as indicated by the increase in fluorescence at the surface of the particle. (e) An CGMP with GFP post reaction with slow kinetics. (f) An CGMP with NPs post reaction with fast kinetics. Flocculation depletion is not observed as indicated by the homogenous fluorescence of the particle.

Figure 9

In Vivo Lung Targeting with Nanoparticle Loaded Composite Gel Microparticles (CGMP) (a) An image of the nearly monodispersed CGMPs with embedded fluorescent EtTP-5 nanoparticles dispersed in phosphate buffered saline. (b) A lung cryosection of a CGMP treated mouse. The fluorescent CGMPs (red) are trapped in a lung capillary. (c) The fluorescence images of organs from a control mouse (top) and from a CGMP treated mouse (bottom). Fluorescence is observed in the treated lung, indicating the lodging of fluorescent CGMPs in the lung. No increase in fluorescence was observed between the heart, kidneys, spleen and liver of a control mouse (top) and a CGMP treated mouse (bottom). The livers exhibited auto-fluorescence.