Rational design and synthesis of 11beta-substituted estradiol hybrid agents for the study and treatment of hormone positive breast cancer and applications in nanomedicine

The relationship between the human estrogen receptor alpha (ERalpha) and breast cancer was first hypothesized by Dr. Jensen in 1971 and demonstrated by Dr. McGuire in 1975. These discoveries laid the groundwork for the development and use of anti-hormone therapy in the treatment of cancer as illustrated two years later when the FDA approved Tamoxifen for the treatment of breast cancer in postmenopausal women. Tamoxifen results in a reduction in the recurrence and death rates of women with estrogen receptor positive (ER+) breast cancer. This is a direct result of Tamoxifen's binding to the ERalpha inducing an antagonist conformation forcing helix 12 to fold in such a way the coactivator binding site is buried in the receptor, thus suppressing transcription. This makes the ERalpha an attractive target for the prevention and treatment of ER+ breast cancer. However, acquired resistance and undesired side effects through agonist activity in endometrial tissue establish a need for new compounds with a higher degree of specificity. Chapter 1 of this thesis presents the rationale for this work and the information that directed the course of research for this project.;Chapter 2 focuses on the rational design of two ERalpha antagonists that are derivatives of estradiol. Several key structural features were identified in steroidal and non-steroidal antagonists that imparted high binding affinity through key receptor interactions. The combination of these structural features determined that substitution at the 11beta--position of the steroid with a specific functional moiety generates ERalpha antagonists via disruption of helix 12 of the receptor. This antagonism is enhanced by the incorporation of a basic functional group, which interacts with aspartic acid 351 of ERalpha. With these design criteria, two steroidal ERalpha antagonists were synthesized possessing strong affinity for the receptor.;Chapter 3 presents the extension of the antiestrogens into the construction of hybrid agents, utilizing the steroid as a targeting agent. For these compounds a modular approach allowing for a diverse combination of compounds is highly desirable. For this reason the antiestrogens were synthesized to utilize Sharpless' "Click Chemistry" for the ligation step via the incorporation of an azide or alkyne. The 11beta position was chosen as the site for functionalization, not only because it generates ERalpha antagonists, but also because it is substituent tolerant. Four hybrid agents were aimed to address hormone therapy resistance by targeting two independent mechanisms that downregulate the ERalpha transcription pathway. Three of these were 11betaE2-GDA hybrids aimed to destabilize the heat shock protein ER complex leading to the complex's degradation. A small molecule tyrosine kinase inhibitor targeted the epidermal growth factor receptor pathway as a way to inhibit ligand independent ERalpha activation. The final hybrid developed was designed to serve as a molecular probe, providing data regarding the dynamic nature of ligands binding to ERalpha. Incorporation of a nitroxide radical allowed for the real time study of the structural adaptations of the receptor using electron spin resonance spectroscopy.;Chapter 4 looks at the translation of the work from the previous Chapters 2 and 3 into the emerging field of nanomedicine utilizing gold nanoparticles as a drug delivery vehicle. This chapter focuses on a model system utilizing a convergent approach toward the functionalization of nanoparticles in a controlled manner. This convergent method was compared to current approaches to nanoparticle construction; "top down" and "bottom up". A bifunctional linker was synthesized providing termini allowing for tight coordination to the gold nanoparticle surface and the incorporation of functional moiety in a chemoselective manner. This convergent approach successfully achieved the monofunctionaliztion of gold nanoparticles with a fluorescent dye which was shown to be viable in vivo.;Chapter 5 summarizes the significance of this research and offers recommendations for future studies based on the research presented in this thesis.