Polymer-based amphiphilic systems that self-assemble into micelles are widely studied, promising molecular designs for the delivery of hydrophobic drug molecules, which are otherwise difficult to deliver due to their poor water solubility. Incorporating stimuli-sensitive character into these polymeric assemblies has elevated the usefulness of these molecular systems in drug delivery applications due to their ability to unload non-covalently encapsulated guest molecules in response to specific stimuli. Among these nano-sized polymeric micelles, dendrimer-based micellar assemblies have received particular attention due to the fact that dendrimers are well-defined-monodispersed molecular architectures. The monodisperse nature of dendrimers provides a unique advantage in studying the structure-property relationship of amphiphilic supramolecular assemblies and stimuli-sensitive disassemblies. In this dissertation, we incorporate stimuli-sensitive characteristics into facially amphiphilic dendrimers designed and synthesized by our group. Our design principle renders these dendrimers responsive to different stimuli such as proteins, redox potential and light. We first study the self-assembly and encapsulation properties of these stimuli-responsive dendrimers in aqueous media. Next, it is demonstrated that these dendritic micellar assemblies disassemble in response to (i) an external stimulus such as light, ( ii) protein-ligand interactions, and (iii) a combination of an enzymatic reaction and redox potential. In the third molecular design, we show that combination of two stimuli enhances the release kinetics of guest molecules as compared to the independent effect of each stimulus. We also demonstrate that disassembly of these dendritic supramolecular assemblies takes place with a concomitant release of hydrophobic guest molecules trapped within the assembly.

Currently, most of the commercially available therapeutics are all targeting cell surface receptors which constitutes only a small portion of the targets found in the cells. Therefore, reaching intracellular targets would provide many new opportunities to treat various diseases. However, intracellular delivery of therapeutic molecules has always been a challenge due to the poor permeability of cell membrane to large, negatively charged macromolecules and their restricted biodistribution. In the past decades, cell penetrating peptides (CPPs), also known as protein transduction domains (PTDs), are shown to improve the intracellular delivery of bioactive molecules and among the PTDs, arginine-rich peptides are highlighted as the most effective subclass. In the light of this information, using the power of polymer chemistry, protein transduction domain mimics (PTDMs) based on ring opening metathesis polymerization (ROMP) of functionalized oxanorbornene derivatives are aimed to be designed. This thesis demonstrates that these PTDMs can adopt cell penetrating activity and show superior properties compared to peptide analogues (i.e. nonaarginine, R9, Pep-1). The structure-activity relationship is studied by guanidinium functionalized monomers. The impact of number of guanidiniums, density of guanidiniums, molecular length and hydrophobicity on cellular internalization is investigated. Further, the siRNA delivery ability of designed PTDMs is also studied. Efficient downregulation of NOTCH1 protein using PTDM-based non-covalent siRNA delivery system in T cell lines and primary blood cells is demonstrated. Two different structures of PTDMs are studied to understand the structural requirements for an efficient carrier. Apart from in vitro testing of PTDM/siRNA complexes, their size and surface charge are also characterized. Further, PTDM-based siRNA delivery system is used to study the function of NOTCH1 in in vitro in primary human blood cells and as well as in humanized mouse model of graft vs host disease as an in vivo environment. In addition to siRNA delivery, novel protein transporter PTDMs which are inspired by primary amphipathic peptides is introduced. The effects of different functional groups and different block lengths on protein delivery efficiency are studied. Successful delivery of functional proteins is demonstrated using Cre Recombinase and Runx1.d190.

The potential applications afforded by elastin-based materials are primarily a product of their stimulus-responsive nature at both the microscopic and macroscopic level. The details of this report are intended as an investigation of stimulus response at or near condition boundaries. The first description of elastin-like peptides, or ELPs, presented here is considered to be largely relevant to the advent of protein engineering. Guest residue modulation offers a potential control of both temperature stimulation and chemical functionality. The reported influence of ELPs with Tryptophan (Trp), Tyrosine (Tyr), or Histamine (His) at lengths defined by n≤6 were studied in a context relevant to protein engineering. Primary results indicate that recombinantly expressed ELPs can exert a distortion of local secondary structure; further, under the constraints of system design, the hydrophobicity of guest residue and number of pentapeptide repeats did not exert a significant influence in determination of the temperature value associated with conformation change, Tt. The second description of elastin-like proteins, or ELPs, is of their use as a molecular probe. Ionic liquids are a novel class of solvents for which the potential application is still being realized. Generally, the literature reports of macroscopic phase behavior describe the solvency of macromolecules by ionic liquids and potential cosolvent mixtures, to an extent. There remains, however, a need to expand on the relationship between micro- and macroscopic transitions of elastin-like molecules. In this work, the phase behavior of an ELP having a particular defined length (n=128), concentration (C=20μM), and sequence (X=V) was considered in the context of four IL/buffer mixtures. Primary results support a description of the variation in phase behavior with IL/buffer composition that can largely be defined according to the physical properties of the complicated IL anion relationship between the physical properties of the anion and the solvent strength of the ionic liquid. Ultimately, this report is intended to provide a novel description of elastin-based materials, such that the information will be useful to the development of two highly relevant technologies: protein engineering and green solvents.

Very selective and highly sensitive ways to detect peptides or proteins of interest remain important goals in proteomics applications. This dissertation focuses on the use of amphiphilic polymeric materials that self-assemble as reverse micelles in apolar solvents as part of a liquid-liquid extraction methodology to selectively extract and concentrate peptides from aqueous solutions. After extraction the polymer-peptide mixtures are amenable to direct analysis by mass spectrometry (MS) using matrix-assisted laser desorption/ionization (MALDI). The polymeric materials that self-assemble in apolar solvents have charged interiors that allow oppositely-charged peptides to readily migrate into the aggregate's core, while similarly-charged peptides do not. This charge complementarity can be controlled by varying aqueous phase pH such that peptides having a narrow range of a pI values can be selectively isolated. Insights into the extraction mechanism and how these materials can potentially open the way for multidimensional separations were also studied. Furthermore, amino acid-specific covalent labels are introduced inside the reverse micelles to selectively react with the extracted peptides having the amino acid of interest. This reaction leads to a mass signature that can be used to identify peptides with amino acids of interest. In addition, bulk ampholytic gels and small commercial surfactants were investigated as extraction materials. Overall, our results reveal that the amphiphilic polymeric reverse micelles are far superior for selectively and efficiently extracting peptides from complex mixtures.