| The work presented here represents three separate research projects. Each explores different approaches to the same problem: how can artificial macromolecules be used to target specific intracellular sites in living organisms, and how may that technology be exploited to treat different disease states? Here, the copolymers based on pHPMA [poly N-(2-hydroxypropyl)methacrylamide)] were used to demonstrate cellular and subcellular targeting in a cancer model. Used as carriers for low molecular weight drugs, pHPMA copolymers may possess great potential for increasing the efficacy of drugs, while also decreasing side effects resulting from drug exposure to healthy tissues and cells.;In the first project, antibody Fab' fragments were dimerized using a multifunctionalized PEG (poly(ethylene glycol)) crosslinker to create a targeting module for pHPMA/drug carriers. The linker was semitrifunctional, able to crosslink 2 Fab' antibody fragments, which could then be attached to an HPMA-doxorubicin anticancer drug carrier. Monoclonal antibody Fab' fragments were chosen that target ovarian carcinoma cells. Using this strategy, the conjugates are selectively uptaken by cancer cells, whereby free doxorubicin is released.;In the second project, HPMA copolymers were created with a terminal mitochondriotropic chemical moiety, TPP (triphenylphosphonium). Lipophilic cations are used to traffic low molecular weight drugs to mitochondria. Studies had reported that terminal TPP can also traffic high molecular weight, uncharged, linear macromolecules into mitochondria, and enable delivery to the cytosol via direct transduction through the plasma membrane. Semitelechelic TPP-HPMA copolymers were synthesized to determine if this effect could be applied to HPMA copolymer drug conjugates.;In the third project, the flexibility of HPMA polymerization chemistry was exploited to create a large "library" or array of different HPMA copolymers with a wide range of chemical properties. Copolymers were made incorporating a variety of cationic, anionic, and hydrophobic sidechains. This array of copolymers was incubated and microinjected directly into the cytoplasm of living cells. The resulting uptake and/or intracellular distribution was observed using time-lapse confocal microscopy and flow cytometry. Using HPMA copolymers as a model, this work functioned as a general survey for cellular uptake, and the intracellular distribution/trafficking that can be expected when artificial macromolecules are internalized into living cells. |
