Humans, wildlife, and the environment in general are being exposed to an increasingly complex milieu of synthetic chemicals/materials whose biological effects aren’t fully understood. To keep pace with this challenge, toxicity testing in the 21st century requires in vitro assays that are both high throughput and more predictive of in vivo effects. ACEA’s real time cell analysis platform, xCELLigence, is well suited to meet this challenge. Using impedance-based microsensors that are both noninvasive and label free, the xCELLigence system monitors adherent cell growth, morphology, and attachment quality continuously over the entire experimental period. This approach mimics continuous monitoring of animal responses and provides significantly more information than what is available via traditional end point assays. ACEA is currently engaged in environmental toxicity testing with two partners:
US Environmental Protection Agency ToxCast™ Program
Launched in 2007, the US Environmental Protection Agency’s (EPA) Toxicity Forecaster (ToxCast™) initiative uses cell-based high-throughput chemical screening technologies to assess toxicity while minimizing the need for laboratory research animals. The wealth of information provided by real time cell analysis (RTCA), and the efficiency with which it is collected using automated impedance recording, have made the xCELLigence platform an asset to ToxCast™ since its inception. To date, ACEA has completed phase I cytotoxicity screening of 320 compounds, and nanotoxicity testing of about 60 different nanomaterials. Moreover, the EPA has adopted two endocrine disruptor assays that were developed on the xCELLigence platform. An assay for detecting estrogen/progesterone mimics was used to screen all 320 phase I and 700 phase II chemicals, as well as the e1K collection of putative endocrine disruptors. The xCELLigence platform is also being used to screen 800 compounds for androgen receptor agonist/antagonist activity. In its ability to monitor dynamic response profiles over the whole experiment, and simultaneously probe both cytotoxicity and hormone-induced responses, xCELLigence is unrivaled. The data from these cytotoxicity and estrogen/progesterone mimics screening are available in the ToxCast database (http://www.epa.gov/ncct/toxcast/), and have also been published (Rotroff et al 2013). After compilation and analysis, the androgen modulator detection screening data will also be publicly available.
Human breast cancer cell line T-47D expresses both estrogen and progesterone receptors. T-47D cells were seeded in a 96-well microtiter E-plate, and monitored on the xCELLigence system. After overnight growth, cells were exposed to two different hormones, 17β-estradiol (E2) and progesterone (Pg). As shown by the red line in each figure, E2 and Pg induce distinctive kinetic responses. While the estrogen antagonist ICI182780 (blue lines) specifically abolished the effect of E2, it had no impact on the effect of Pg.
Collaboration with the Alberta Centre for Toxicology (ACFT)
In 2009 ACEA began a partnership with the Alberta Centre for Toxicology (University of Calgary, Canada). With the goal of developing a new human risk assessment model based on in vitro cytotoxicity assays, key human cell lines are being analyzed on the xCELLigence platform for their response to reference chemicals and, eventually, environmental samples. To date, about 800 reference chemicals have been tested, including more than 100 compounds with known in vivo LD 50 values. Owing to the high quality/reproducibility of the data generated on this innovative system, several different models for predicting toxicity potency have already been developed (Pan et al 2013). Because the time-dependent cell response profile generated on the xCELLigence platform is similar for groups of chemicals that display the same mechanism of action (MOA), the data are being used to develop mathematical models for predicting a chemical’s MOA in the absence of additional information (Xi, et al 2014).
- In vitro cytotoxicity assessment based on KC(50) with real-time cell analyzer (RTCA) assay. Pan et al (2013) Comput Biol Chem. 47:113-20.
- Real-time growth kinetics measuring hormone mimicry for ToxCast chemicals in T-47D human ductal carcinoma cells. Rotroff et al (2013) Chem Res Toxicol. 26:1097-107.
- Mode of action classification of chemicals using multi-concentration time-dependent cellular response profiles. Xi et al (2014) Comput Biol Chem. 49:23-35.