Research Seminar Abstract
The insulin receptor (IR) and insulin-like growth factor receptor (IGF1R) system is implicated in the pathogenesis of breast cancer. As members of the tyrosine kinase receptor family, these receptors are able to dimerize in different combinations forming IR-IR and IGF1R-IGF1R homodimers and IR-IGF1R heterodimers. Overexpression of one and/or both receptors on the cell surface represents a clinical risk factor for the disease. However, it seems apparent that the risk is not solely attributable to the absolute amounts of receptor expressed on the cell surface, but also to their relative amounts and distribution between the different dimeric forms. Despite great homology in the structures of and signal transduction pathways induced by these receptors, it is theorized that each species might generate differential downstream effects on cell growth, division, and apoptosis in response to binding of hormone. Thus the ability to quantitate these different species and their hormone binding affinities is of paramount importance. To date, we know of no such technique able to accomplish such a feat. We propose fluorescence correlation spectroscopy, a technique with good spatial resolution, a large temporal range, short acquisition times, and robust photon statistics, as a method to examine membrane-localized fluorescence in situ. Herein this technique is applied to determine hormone-binding affinities and estimate receptor dimer surface densities through probe-labeled hormone-saturation experiments. We also propose a new and improved method implementing our 4-detector fluorescence correlation apparatus for determining receptor dimer surface densities by direct labeling of receptors via transient transfection using cDNA vectors. Ultimately, the information gained from these experiments, in tandem with endocrinological studies of hormone regulation and studies aiming to elucidate intracellular signaling pathways, will shed light on the role of this system in the pathophysiology of breast cancer and inform future therapeutic approaches.