(6af) Systems Biology Approach to Endocrine Signaling

Advances in understanding the molecular basis of normal physiology and disease states are critical to developing improved diagnostic tests and therapeutic interventions for human health. A new avenue toward this objective has arisen from the advent of biomimetic tissue culture, increasing the relevance of experimental results, and of systems biology, comprehending molecular mechanisms in terms of integrated networks rather than as mere isolated components. Combined, these two technological revolutions offer the promise of more powerful studies of cell behavior in a physiologically appropriate microenvironment that examines the integration of external cues (e.g., extracellular matrix and soluble factors) and intrinsic cues (e.g., gene expression levels).

Developing a 3D Microenvironment for Ovarian Follicle Development

My graduate work, under Dr. Lonnie Shea and Dr. Teresa Woodruff, focused on the importance of matching the microenvironment to the needs of a tissue for appropriate development. We applied this conceptual question to the development of the ovarian follicle, in order to generate an alternative method of producing mature eggs for fertility treatments. In my research, I developed three-dimensional synthetic matrices that promoted ovarian follicle development and produced meiotically competent oocytes for fertilization. In this system, ovarian follicles were isolated from immature mice, encapsulated, and cultured within alginate matrices to maintain cell-cell connections which are critical for growth and maturation of the enclosed oocyte. Through modification of the scaffold material and culture conditions, I examined the regulation of follicle development by the extracellular matrix and follicle stimulating hormone (FSH). These distinct signals integrated to regulate the quality of the oocyte. As an example, earlier stages of follicle development were promoted by the addition of collagen I, while FSH was critical for growth of later stage follicles and oocyte maturation.

Systems Biology Analysis of the Effects of RAS Mutations in Apoptosis Signaling

In my postdoctoral research, with Dr. Douglas Lauffenburger, I have been developing computational models of a ?cue ? signal ? response' form, where the cues to the cell are both external (e.g., cytokine treatments) and internal (e.g., genetic mutations). In particular, in order to examine the role of RAS mutations, which are identified in many human cancers, we are developing a partial least-squares regression model for apoptosis in an isogenic panel of colon carcinoma cells lines with varying RAS mutations. Our hypothesis is that cell fate decisions are regulated through a quantitative interplay between the intracellular signaling pathways downstream of apoptotic stimuli and RAS, and that elucidation of this function will enable us to discern the role of mutant RAS in apoptosis. To develop this model, we are constructing an extensive dataset of apoptotic responses to a combination of extracellular cytokine stimuli and RAS genotype variations, along with the corresponding intracellular signal measurements downstream of RAS and tumor necrosis factor (TNF) receptor. Our results indicate that the K-RAS mutation sensitizes cells to TNF stimulated apoptosis, increasing cell death nearly 33% by 48 hours. Our results indicate that in addition to the phenotypic differences with varying RAS state, there are distinct signaling profiles in response to treatment with TNF. Future work will extend this system biology approach to animal models that conditionally express these mutations in the intestinal epithelium.

Combined Application of these Approaches to Understanding Fundamental Biological Questions of Importance in Women's Health

Integrating my doctoral and postdoctoral research experiences, my research program will investigate fundamental biological questions using a systems biology/engineering approach and disease models of importance in women's health. I will extend a systems biology approach to incorporate endocrine signaling, which provides a master level of biological regulation through organ to organ communication and is critically related to a variety of disease states in women. Here, I outline initial studies which will incorporate both experimental and modeling approaches to improve understanding of:

? The impact of immediate versus delayed signals in response to cellular cues - we will develop a ?cue-signal-response' model of a panel of estrogen mimicking compounds in breast cancer cells, examining the multiple modes of estrogen-induced signaling.

? How cells interpret multiple synergistic and antagonistic cues - we will examine the extensive network of cross-talk between the EGF family receptors and the estrogen receptors.

? How mutations such as those found in cancer rewire the cell network response to physiological inputs - we will examine mutations implicated in the progression of ovarian cancer, and the cellular response to endogenous paracrine and endocrine ligands.