Three-dimensional testicular cell co-culture model for reproductive toxicity screening

Summary Reproductive and developmental disorders caused by drugs or environmental chemicals are a prominent health issue worldwideCurrent toxicity testing methods set by regulatory agencies predominately rely on animal testingThe cost and time associated with animal testing for risk assessment seriously limit efforts for extensive toxicity screeningThese data gaps necessitate the implementation of a new vision for toxicity testing as reflected in the National Research Council s reportToxicity Testing in thest Centuryand the restructuring of risk assessment proceduresToxicity testing in thest century must move beyond classical methods of risk assessment by incorporating modern molecular knowledgesuch as adverse outcome pathwaysAOPswith advanced in vitro modelsCurrentlyin vitro reproductive toxicity testing models are actively being developedUnder the support of RESgrantReproTOX established a Mini Testis model from testicular cell lines and found this Mini Testis model could identify reproductive toxicantsBoth morphological and cell specific biomarkers assessment demonstrated that this co culture model created an in vitro like nicheformed a three dimensional cytoskeleton bundle structure and supported germ cells within theD environmentWe initially validated this model by discriminating reproductive toxicants amongcompounds and observed a strong correlation between the in vitro ICand in vitro rLOAELthe lowest observed adverse effect levelfor these selected compoundsThe calculation of concordancesensitivityand specificity further supported the reliability of this modelOur results from the phase I study suggested that our in vitro MiniTestis model might be a valuable screening tool for reproductive toxicant assessment and to prioritize chemicals for further testingIn this phase II proposalwe will further examine and optimize the protocol for the in vitro Mini Testis modelWe will test the Mini Testis model with an expanding list ofreference testing compoundsand validate blindly our predictive model of pathway based high content and high throughput platformThe Specific Aims areto complete the development of high content assays based on the adverse outcome pathways associated with the reproductive toxicity in an in vitro Mini testis modelto apply the multi parametric high throughput and high content analysisHT HCAto examine the sensitivity and specificity through comparison between the ICobtained from the in vitro model and in vivo reproductive lowest observed adverse effect levelrLOAELto validate the in vitro Mini testis modelto conduct the intraand inter laboratory reproducibility study to validate the in vitro Mini testis modelwe will test the variability within test and the reproducibility of the test within and among laboratoriesAltogetherthis proposed Phase II will focus on the continued development of thisD Mini Testis screening platform toward a validated quantitative pathway based HT HCA screen for reproductive toxicityCommercialization of this in vitro platform and its ability to screen up to a thousand compounds via focusing on multiple endpoints associated with adverse reproductive effects caused by agents from a variety of chemical classesThis MiniTestis model based platform has the potential to offer predictive reproductive and developmental toxicity without the use of animals in a high throughput formatand will be a critical part of an integrated testing strategy for Reproductive and developmental toxicity Project Narrative Reproductive and developmental disorders caused by drugs or environmental chemicals are a prominent health issue worldwideCurrent toxicity testing methods set by regulatory agencies predominately rely on animal testing and the cost and time associated with animal testing for risk assessment severely limit efforts for extensive toxicity screeningThis proposal to establish in vitroD Mini testis model has the potential to offer predictive toxicity screening without the use of animals in a high throughput formatand will be a critical part of an integrated testing strategy for Reproductive and developmental toxicity testing