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IMPLEMENTATION OF 3D CELLULAR MODELS FOR DRUG AND TOXIN TESTING IN HEALTHY AND CANCEROUS TISSUE

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abstract
Background: In the development of pharmaceutical drugs, it is imperative that microphysiological systems represent their parent tissue as accurately as possible. It is essential that any preclinical model is human, able to recapitulate the organization, cell type, and proper cell to cell interactions. Human 2D cell line models and animal models often fail to meet these criteria, and these failures have cost thousands of lives and billions of dollars. Methods: To add ress these shortcomings in the form of a potential screening tool, we employed bioengineered 3D liv er and cardiac organoids biofabricated from multiple physiologically relevant human cell groups to represent healthy tissue , and tumor organoids derived from colorectal cancer cells from established cell lines to screen multiple materials for toxic effect, including environmental toxins, FDA recalled drugs , and chemotherapeutics. and charted the response of the organoids to these compounds. With liver and card iac organoids, t ests were performed that measured ATP activity, LIVE/DEAD viability and cytotoxicity staining, and cardiac organoid beating activity to demonstrate a comprehensive picture of cellular death under toxin exposure . With tumor organoids, MTS an d additional IHC staining was performed on to evaluate drug efficacy and potential mechanisms of action. Tests were performed in both 2D and 3D systems to compare sensitivity to toxin exposure in organoids versus more traditional 2D cultures. Results: All compounds tested induced toxicity in the organoids. Both ATP and LIVE/DEAD results showed toxicity in cardiac and liver organoids with most systems showing 3D systems having more sensitivity than 2D , with organ specific toxicity displayed in recalled drugs . Additionally, cardiac beat rates were affected at drug and toxin concentrations much lower than those shown to induce toxicity as per ATP or LIVE/DEAD assays. Chemotherapeutic testing in 3D colo rectal cancer organoids displayed EGFR pathway mutation targ eting drug s are given an advantage over the 2D cultures; the reverse was true for most non - EGFR targeting drugs. Additionall y, IHC staining showed upregulation of EGFR and its downstream components in 3D over 2D. Overall, t hese results suggest that the 3D organoids have significant utility to be deployed 8 in additional toxicity screening applications, and future development of treatments targeting certain mutations present in cancer systems with the capacity to involve human derived primary tumors . Conclus ion: Taken together, these results show the potential application of human - based 3D organoids to be applied in a variety of toxicity screening applica tions, such as drug development, environmental toxin detection , and as a diagnostic tool for the treatment of various cancers .
subject
contributor
Forsythe, Steven (author)
Skardal, Aleksander (committee chair)
Soker, Shay (committee member)
Murphy, Sean (committee member)
date
2018-05-24T08:36:15Z (accessioned)
2020-05-23T08:30:17Z (available)
2018 (issued)
degree
Biomedical Science – MS (discipline)
embargo
2020-05-23 (terms)
identifier
http://hdl.handle.net/10339/90744 (uri)
language
en (iso)
publisher
Wake Forest University
title
IMPLEMENTATION OF 3D CELLULAR MODELS FOR DRUG AND TOXIN TESTING IN HEALTHY AND CANCEROUS TISSUE
type
Thesis

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