(51a) Oral Delivery of Therapeutic Proteins Exhibiting High Isoelectric Points in pH-Responsive Hydrogels

Koetting, M. C., University of Texas at Austin
Peppas, N. A., University of Texas at Austin

Oral Delivery of Therapeutic Proteins
Exhibiting High Isoelectric Points in pH-Responsive Hydrogels

Michael C. Koetting and
Nicholas A. Peppas, The University of Texas at Austin, Department of Chemical
Engineering, Austin, TX

Introduction: Therapeutic
delivery of proteins via intravenous injection is an invasive and often painful
method which has been shown to cause low patient compliance with
doctor-recommended treatment.  Delivery via the oral route is strongly
preferred to avoid the pain and embarrassment of injection and therefore
increase patient compliance and improve quality of life.  Environmentally-responsive
hydrogel systems have shown great promise as drug delivery vehicles to overcome
the obstacles of oral delivery.  Our group has previously developed copolymer
hydrogel delivery vehicles?notably poly(methacrylic acid) grafted with
poly(ethylene glycol), or P(MAA-g-EG)?for the oral delivery of insulin. 
These hydrogels protect the protein through the gastrointestinal tract and release
the protein in the small intestine by swelling at the higher pH found in the

            Our current
work is focused on creating a pH-responsive copolymeric hydrogel delivery platform
for high isoelectric point-exhibiting proteins, specifically salmon calcitonin
which is used to treat osteoporosis and Paget's disease.  Calcitonin displays
very different properties than insulin, primarily a high isoelectric point of
8.86 compared to 5.39 for insulin.  Since roughly half of the human proteome
consists of proteins with similarly high isoelectric points, the ability to
orally deliver these many compounds will be a significant accomplishment toward
making protein therapy cost effective, minimally invasive, and clinically
desirable.  In this work, we investigate potential hydrogel delivery systems to
develop an effective oral delivery platform for achieving high bioavailability
with salmon calcitonin.

Methods and Results:
pH-Responsive hydrogels were synthesized by UV-initiated free radical
polymerization in 0.7 mm thick films.  The hydrogels were composed of either
methacrylic acid (MAA) or itaconic acid (IA) copolymerized with either N-vinyl
pyrrolidone (NVP) or poly(ethylene glycol) methyl ether methacrylate (PEGMMA). 
The hydrogels' swelling characteristics were determined in equilibrium and
dynamic swelling studies, with all gels demonstrating a collapsed network at
low pH (pH 2.0) and a significantly swelled network at neutral pH (pH 7.4) that
is suitable for protection of encapsulated protein through the acidic,
proteolytic stomach environment and for diffusive release in the neutral
environment of the small intestine.  Protein delivery capabilities were tested in
by loading 90-150 μm microparticles (from crushing/sieving of dried
films) with salmon calcitonin by imbibition in solutions of varying ionic
strength.  Calcitonin was then released in a pH 7.4 solution of 0.150 M PBS. 
Loading and release levels were determined by HPLC or MicroBCA assay as
appropriate.  Hydrogels displayed significantly improved delivery capability
when loaded in low ionic strength solutions, achieving up to 15.8 times as much
calcitonin released per unit of hydrogel compared to when loaded in standard
0.150 M PBS.  Additionally, hydrogels based on methacrylic acid, specifically
poly(methacrylic acid-co-N-vinyl pyrrolidone) or P(MAA-co-NVP),
achieved improved delivery levels compared to gels based on itaconic acid.  Together,
the platform achieves delivery levels that would suggest its suitability as a
therapeutic oral delivery platform for salmon calcitonin.

Conclusions:  pH-Responsive
hydrogels were synthesized and screened as candidates for oral delivery of the
therapeutic protein salmon calcitonin, as well as similar high isoelectric
point-exhibiting proteins.  Loading and release studies demonstrated that
hydrogels based on methacrylic acid as the pH-responsive moiety offer greater
delivery capabilities than itaconic acid-based hydrogels.  Importantly, it was
also shown that the ionic strength of the loading solution strongly affects
both loading and release levels, with low ionic strength loading solutions
offering significantly improved delivery capabilities in vitro
Therefore, we have shown improved delivery capability achieving levels
sufficient for clinical therapy. 

Acknowledgment:     This
work was supported by grant 1R01-EB000246-19 from the National Institutes of