Date of Award

2011-01-01

Degree Name

Doctor of Philosophy

Department

Biological Sciences

Advisor(s)

Marc Cox

Abstract

Steroid hormone receptors require the ordered assembly of various chaperone and cochaperone proteins in order to reach a functional state. The final stage in the receptor maturation process requires the formation of a mutimeric complex consisting of Hsp90 dimer, p23, and one of several large immunophilins. Studies conducted previously demonstrated that the large immunophilin FKBP52 acts to potentiate glucocorticoid, androgen, and progesterone receptor signaling pathways. The aim of these studies was to identify and characterize FKBP52-specific inhibitors that would not only serve as tools for the pharmacological analysis of FKBP52-receptor interactions, but may also lead to novel drugs with significant therapeutic potential. A modified receptor-mediated reporter assay in yeast used to screen a natural compound library for FKBP52 inhibitors led to the identification of multiple hits. Structure activity relationship studies using the yeast-based assays led to the characterization of fifteen additional inhibitors, some of which display dramatically increased potency. Surface plasmon resonance studies confirmed that these inhibitors disrupt FKBP52 function through interaction with the AR hormone binding domain. In addition, scintillation proximity binding and fluorescence polarization assays demonstrated that the compounds bind to a previously unrecognized regulatory surface domain on the AR LBD termed BF3. Interestingly, mutations within the BF3 surface lead to increased dependence on FKBP52 for normal function. ELISA and Western immunoblotting analysis in prostate cancer cells demonstrated that the compounds inhibit Prostate-Specific Antigen and FKBP51 expression. In addition, androgen-dependent cell proliferation was prevented after treatment with the inhibitory molecules. In summary we have identified molecules that inhibit FKBP52 regulation of AR function and represent an exciting new approach for the treatment of prostate cancer. In addition, these studies provide new insight into FKBP52-AR structural and functional interactions.

Language

en

Provenance

Received from ProQuest

File Size

127 pages

File Format

application/pdf

Rights Holder

Johanny Tonos De Leon

Download

Share

COinS