For this UG3-UH3 transition we plan to maintain our original milestones as originally proposed. These proposed milestones will aim to develop next-generation technologies that will be used to create a transformative model of polycystic ovary syndrome as well as novel screening tool for identifying chemicals and compounds for endocrine disruption. The first of these milestones will focus on the development and integration of downstream biosensors using molecularly imprinted polymers. Previously, hormone production in our model systems was characterized using ELISA techniques. While we are currently transitioning to the use of mass-spectroscopy to improve the accuracy of our quantitative measurements, we are also looking forward to improving the throughput of these measurements. Currently these measurements are performed on media aliquots collected from culture systems on a daily basis resulting in 14 measurements across a 14-day culture timeline. However, through the integration of in-line molecularly imprinted polymers to detect specific molecules, specifically estradiol, progesterone, and testosterone, could provide near real-time concentration measurements via electrochemical sensors. The desired detection range for these compounds is 10-20,000 pg/mL with a frequency of twice daily at a minimum, thus doubling our quantitative capacity. To further increase the quantitative output of our model systems, for our second milestone we will establish fallopian tube epithelial (FTE) and endometrial cell lines with hormone responsive biosensors. These cell lines will respond in near real-time to exogenous estradiol, progesterone and testosterone within a range of 10-100 pg/mL via incorporated reporter constructs with different colors/wavelengths specific to the estrogen receptor (ER), progesterone receptor (PR) and androgen receptor (AR). Following validation of the BLAC TIE system via the UG3 milestones, we will deploy the BLAC TIE PCOS model to screen drug compounds from our pharmaceutical partner, AstraZeneca, for our third UH3 milestone. WE plan to screen 10 different drug compounds currently drugs in development for the treatment of symptoms associated with PCOS across 8 different doses for a period of 14 days using our validated PCOS culture profile. As the ultimate goal of this project and the development of the BLAC TIE system is to use the ex vivo multi‐organ disease model of PCOS to test new therapies and investigate the biological mechanism of existing drugs, this milestone will serve as the cornerstone of our current work. By working together with our industry partner, we can validate that the newly developed system provides relevant data for drug discovery and screening to identify new treatments for diseases such as PCOS. Finally, in order to demonstrate the broad functionality of the BLAC- IE system, for our final milestone we will Use BLAC TIE PCOS model to screen chemicals from the EPA Endocrine Disruption Screening Program. Endocrine disrupting chemicals are believed to be contributors to the development of PCOS. Therefore, we will test the functionality of our integrated multi‐tissue system as new tool for studying how EDC’s may contribute to PCOS pathogenesis. This milestone will include as screen of 10% of the 1,800 chemicals of interest with data from the EDSP21 Dashboard to determine any potential contribution to PCOS pathophysiology. The ability to screen this large number of compounds on a complex multi-tissue in vitro model illustrates the important technological development that is being performed by our group. The
|Effective start/end date||9/15/19 → 8/31/23|
- National Institute of Environmental Health Sciences (3UH3ES029073-05S1)
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