(221e) Injectable, Biodegradable Pnipam-Magnetite Nanoparticle Hydrogels

Injectable,
biodegradable PNIPAM-magnetite nanoparticle hydrogels

Scott
Campbell and Todd Hoare

Department of Chemical Engineering, McMaster
University, Hamilton, ON, Canada

INTRODUCTION: The
development of versatile hydrogel networks that elicit some change in response
to one or more external stimuli could have a diverse range of potential
applications. For example, composites with heat induction capabilities could be
used for externally-activated but locally-induced hyperthermia treatment (i.e.
exposing cancerous regions to elevated temperatures (~43°C) that will lyse the
more temperature sensitive cancer tissues while being safe for normal tissues¹)
and/or as scaffolds for the delivery of therapeutic agents. Alternatively, thermosensitive
drug delivering agents, such as micelles or microgels, could be incorporated
into these scaffolds to allow for externally controlled drug release.²
Such treatments may be combined into a single device to further increase the
efficacy of treatment. Consequently, stategies involving heat inducting,  thermosensitive composite materials utilizing
magnetite nanoparticles in alternating magentic fields (AMF) have been widely
pursued in recent years.

While many polymer-magnetite composite
materials have been fabricated before, few are able to be directly injected to
the desired site of interest with the hydrolytically-degradable crosslinks that
are formed immediately after injection. The development of injectable materials
that can provide similar release profiles to currently reported,
macroscale-systems^3,4 would be highly beneficial to expand the
potential applications and patient convenience of such devices. The further ability
of these composite materials to degrade allows for the clearance of their
comprising compontents from the body over time.  Additionally, while most currently reported
systems physically entrap the magnetite nanoparticles in a hydrogel matrix, by
functionalizing the surface of the nanoparticles so that they also participate
in the gelation process, the rheological properties of the final composites may
be significantly altered, leading to exceptionally strong or elastomer-like hydrogels
with unique properties and other potential applications (e.g. embolic or
structural biomaterials). In response, this work describes the fabrication of
novel thermosensitive, injectable, covalently-cross-linked, stimuli-responsive
hydrogels in which one of the polymers used to form the hydrogel is physically
adsorbed to magnetite nanoparticles.  The
hydrogel composites are characterized for their rheological and degradation
properties, biocompatibility, and drug delivery properties.

EXPERIMENTAL: Magnetic, thermosensitive, in situ-injectable hydrogels were
prepared by mixing hydrazide-functionalized poly(N-isopropylacrylamide) (PNIPAM)
coated magnetite nanoparticles, in which the PNIPAM component is physically
adsorbed to the magnetite nanoparticles, and aldehyde-functionalized dextran. The
PNIPAM-based polymer is a copolymer of acrylic acid (20 wt.%) and NIPAM that is
functionalized with hydrazide groups via the carbodiimide-mediated conjugation
of adipic acid dihydrazide (ADH).⁵ Adsoprtion of this polymer to the
surface of iron oxide nanoparticles is performed directly after their
synthesis. Here, in a similar procedure to that used by Hoare et al. (2009), FeCl₃
and FeCl₂ in a 2:1 molar ratio are coprecipated with ammonium hydroxide
to form magnetite nanoparticles that are then heated to 70°C in the presence of
hydrazide-functionalized PNIPAM to peptize the polymer to the surface of the
nanopatrticles.³ Oxidizing dextran with sodium periodate is used to
synthesize the aldehyde-functionalized dextran.⁷ The hydrogels are
manufactured by placing PBS-based solutions of the hydrazide-functionalized NIPAM
coated magnetite nanoparticles and aldehyde-functionalized dextran in opposite
compartments of a double barrel syringe (with the desired drug if it is
intended for drug delivery purposes). The hydrogels are formed when the
solutions are co-extruded upon injection into pre-fabricated molds and the
hydrazide and aldehyde groups of either polymer precursor condense to form
hydrazone crosslinks (Figure 1).

Figure 1:
Hydrogel fabrication process. The double-barrel; syringe has two compartments:
one with hydrazide functionalized NIPAM and the other with aldehyde
functionalized dextran in PBS solutions. Upon injection, these two solutions
mix and hydrazone bonds form in prefabricated moulds.

The hydrazone crosslinks that form will
hydrolytically degrade over the course of several months under physiological
conditions.⁵ The PNIPAM component of the hydrogels is
thermosensitive such that the hydrogels reversibly decrease in size when the
temperature of their environment exceeds their lower critical solution
temperature (LCST).⁷ The magnetite nanoparticles can generate heat
when placed in an AMF via hysteresis losses.¹ When the
thermosensitive hydrogel and magnetite nanoparticles are combined in the same
nanocomposite, heating of the nanoparticles induces a phase transition in the
NIPAM component, which will cause the hydrogel to deswell and release a burst
of pharmaceutical agents. Site-specificity is achieved by injecting the
composite at the desired site, where it quickly gels in vivo using the rapid hydrazide-aldehyde chemistry. The
thermosensitive nature of the hydrogel can be adjusted, along with numerous
other parameters (for example, the degree of functionalization and the concentration
of hydrogel precursor polymers, the phase transition temperature of the polymer
precursors via copolymerization, etc.) to attempt to control the rate of drug
release and the on-demand control of drug release under the presence of an AMF.

RESULTS: Stable, injectable, mechanically robust hydrogels
were successfully formed with 2-10 wt% aldehyde-functionalized dextran content.
The dried gels range from 70-75% iron oxide based by mass, relating to hydrated
gels that have 10 - 22 wt.% iron oxide content. The hydrogels themselves have
shear storage moduli orders of magnitude greater, and elasticities
significantly greater, than conventional hydrogel materials, behaving on a bulk
scale more like a conventional elastomer than a hydrogel (Figure 2).

Figure 2: Storage
(G') and loss (G") moduli of composites fabricated with varying dextran
polymer (Dex B) contents (6 wt.% and 8 wt.%) in the
hydrogel precursor solution.

Composites have
been designed that are also able to be quickly broken apart under the application
of an AMF, likely due to their lower crosslink density than conventional gels
and the thermal motion of the  magnetite
nanoparticles under an AMF. These hydrogels could be used in alternate
applications, such as a blood stopper in surgeries that can be broken apart
after the operation is completed and as a quick supply iron source for the body.
Drug-loaded hydrogels that are more stable under the presence of an AMF have
also shown high-low release characteristics upon applications of short 5-10
minute pulses of an AMF (Figure 3). Increases in drug
release when the AMF is applied may be partially attributed to the induction-enhanced
degradation of the hydrogel itself. Additionally, the performance of these composites in cell viability
assays will be discussed.

Figure 3: Rate of
bupivacaine drug release under pulsed AMF conditions. The red line represents
release without the application of an AMF, while the green lines indicate the
enhanced rate of release after 5 minutes of applied AMF.

CONCLUSIONS: The
combination of thermosensitive polymers with magnetite nanoparticles is a
powerful tool in the fabrication of composites for therapeutic purposes. The
injectable and externally-triggerable drug delivery composite material described
herein offers significant advantages over many current therapies in terms of
facilitating triggered changes in the composite properties using a
patient-friendly and non-invasive triggering technology.

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ACKNOWLEDGEMENTS:
This research is funded by the Natural Sciences and Engineering Research
Council of Canada (NSERC) and the J.P. Bickell Foundation (Medical Research
Grant).