Reactive stimuli-responsive poly(vinyl lactam) copolymers for design of nanogels based protein and drug carriers
- Reaktive stimulationsresponsive Poly(vinyllactam)-Copolymere zur Ausbildung nanogel-basierter Protein- und Wirkstoffträger
Peng, Huan; Pich, Andrij (Thesis advisor); Schwaneberg, Ulrich (Thesis advisor)
Dissertation / PhD Thesis
Dissertation, RWTH Aachen University, 2016
This thesis deals with synthesis, characterization and applications of (biohybrid) nanogels based on (water-soluble) poly(N-vinyl lactams) copolymers functionalized with reactive side groups. RAFT (Reversible Addition-Fragmentation chain Transfer) copolymerizations of methacrylic acid N-hydroxysuccinimide ester and cyclic N-vinylamides derivatives (N-vinylpyrrolidone, N-vinylpiperidone and N-vinylcaprolactam) were successfully performed with methyl 2-(ethoxycarbonothioylthio)-propanoate as chain transfer agent (CTA). Biohybrid nanogels were constructed via chemical crosslinking of water soluble reactive copolymers poly(N-vinylpyrrolidone-co-N-methacryloxysuccinimide) with proteins such as enhanced green fluorescent protein (EGFP) and cellulase via water-in-oil emulsion. Biofluorescent nanogels crosslinked with varying amounts of EGFP were obtained. The biocatalytic activity of cellulase-conjugated nanogels (CNG) can be tunable via crosslinking density. Circular dichroism (CD) spectra and fluorescence resonance energy transfer (FRET) based study disclosed the mechanism how the crosslinking density influence upon the catalytic activity of the biohybrid colloids. The biohybrid nanogels displayed enhanced stability compared with free cellulase after storing at room temperature for three weeks. Thermo and redox dual sensitive nanogels based on N-vinylcaprolactam (VCL) and N-succinimidyl methacrylate (Suma) crosslinked with diallyl disulfide were synthesized via a facile and straightforward method, with tunable size from 200 nm to 1.1 μm. Doxorubicin (DOX) was covalently bound to the nanogel network via coupling with succinimide groups at mild conditions to obtain prodrug nanogels. The drug delivery system exhibited a sustainable delivery profile in the intracellular release study and high antitumor activity.A series of novel pyridyldisulfide (PDS)-functionalized poly(N-vinyl lactams) statistical copolymers were developed via RAFT polymerization. The PDS-functionalized reactive polymers were amenable to a variety of thiol-containing molecules under mild conditions. The contact angle measurements results and fluorescence microscopy study indicated that the reactive films based on the PDS-functionalized copolymers allowed facile, direct, and environmental-friendly surface engineering in aqueous solution, which can control cell attachment and growth on the surface to meet demands of a particular application.Redox-responsive polymeric nanogels were synthesized via in situ crosslinking of PDS-functionalized poly(N-vinylpyrrolidone). Cellulase as a representative protein was encapsulated in the nanogels (PNG1 and PNG2). The enzymatic activity was greatly deactivated when encapsulated in the nanogels while rapidly recovered in 10 mM DTT solution. The FRET based experiments were performed and revealed that the recovered enzymatic activity was mainly ascribed to reduction inducing degradation of the disulfide crosslinking network, ultimately causing protein release. A series of well-defined primary amine-functionalized statistical copolymers based on cyclic N-vinyl lactam monomers and N-vinylformamide (VFA) were successfully synthesized via RAFT polymerization. Water soluble primary amine-based reactive copolymers were obtained by controlled hydrolysis of the VFA components in alkaline condition. Functional nanogels were prepared in W/O emulsions utilizing the coupling between the reactive amine side-groups of polymer chains and diacrylate functionalities of poly(ethylene glycol) crosslinker via Michael addition reaction.Biodegradable nanogels based on primary amine-functionalized poly(N-vinylpyrrolidone) copolymers were prepared in water-in-oil emulsion via Michael addition reaction with a redox-responsive cross-linker. DOX-loaded nanogels demonstrated decreased drug release rate with increased crosslinking density in presence of 10 mM GSH at 37℃. The in vitro release studies revealed that maximum 85% DOX was released in 24 h under a reductive environment whereas only maximum 13% DOX was released in 24 h under nonreductive conditions. Intracellular drug release experiment indicated that the DOX delivery rate is tunable via varying crosslinking densities of the nanogels. Cell viability assay demonstrated that the free nanogels were biocompatible in wide concentrations while the DOX-loaded nanogels displayed medium antitumor activity.