Consultant: Darrell J. Irvine and Patrick S. Doyle.
Department: Massachusetts Institute of Technology. Dept. of Chemical Engineering.
Writer: Massachusetts Institute of Technology
Date Issued: 2008
Therapeutics for example proteins, DNA, or siRNA, are only able to exert their function within the cell cytosol or nucleus. However, many of them are cell membrane impermeable molecules that may simply be adopted by cells via endocytosis or phagocytosis. Such drug molecules therefore are limited in endolysosomes, where reduced pH and degradative enzymes may destroy them without therapeutic gain. Efficient escape of drug molecules towards the cytosol before destruction in endolysosomes is really a major challenge for intracellular drug delivery. To deal with this problem, we developed a pH-sensitive core-covering nanoparticle to segregate the functions from the particle into an endosome-disrupting pH-responsive core that will absorb protons at endolysosomal pH, along with a covering whose composition might be tuned to facilitate particle targeting, cell binding, and drug binding. Two-stage surfactant-free emulsion polymerization of two-diethylamino ethyl methacrylate (DEAEMA) (core) and a pair of-amino ethyl methacrylate (AEMA) (covering) in the existence of a crosslinker was utilized for that synthesis of monodisperse core-covering hydrogel nanoparticles of 200 nm across. The protonation of tertiary amine groups around the polyDEAEMA core on moving from extracellular to endolysosomal pH led to reversible swelling from the nanoparticles having a 2.8-fold diameter change. Using pH-sensitivity of those nanoparticles, efficient cytosolic delivery of calcein (with
95% efficiency) was achieved by disrupting endolysosomes via proton sponge effect. The main amine wealthy covering was discovered to facilitate cell and drug binding, and provided minimal cytotoxicity by sequestering the proton sponge component from the direct interactions with cells. These particles shown a helpful way to deliver therapeutic molecules towards the cytosol of cells of great interest efficiently.(cont.) The applying nanoparticles demonstrated significant improvement in delivering one antigen vaccine protein ovalbumin (OVA) to primary dendritic cells for T cell activation, and promising knockdown of mRNA by delivering siRNA to epithelial cells for gene silencing. To increase this method to some fully biodegradable system, nanoparticles having a cleavable crosslinker bis (acryloyl) cystamine (BAC) were synthesized. Preliminary explorations of the approach demonstrated that such particles can degrade in the existence of glutathione in vitro, a reducing peptide present at mM concentrations within the cytosol of mammalian cells. This design may potentially function as a drug releasing mechanism to improve delivery efficiency.
Thesis (Ph. D.)–Massachusetts Institute of Technology, Dept. of Chemical Engineering, 2008.This electronic version was posted through the student author. The certified thesis will come in the Institute Archives and Special Collections.Vita.Includes bibliographical references (p. 193-208).
Keywords: Chemical Engineering.