Kar Lai Poon*, XingangWang*, Serene G. P. Lee†, Ashley S. Ng*, Wei Huang Goh*, Zhonghua Zhao*, Muthafar Al-Haddawi*, Haishan Wang*, Sinnakaruppan Mathavan†, Philip W. Ingham*,‡, Claudia McGinnis*,ll,1 and Tom J. Carney*,‡,1,2
* Institute of Molecular and Cell Biology (IMCB), A*STAR (Agency for Science, Technology and Research), 61 Biopolis Drive, 138673 Singapore;
† Genome Institute of Singapore, A*STAR (Agency for Science, Technology
and Research), 60 Biopolis Street, 138672 Singapore;
‡ Lee Kong Chian School of Medicine, Nanyang Technological University, 50 Nanyang Avenue, 639798 Singapore;
ll Roche Pharmaceutical Research & Early Development (pRED), Pharmaceutical Sciences, Roche Innovation Center Basel, Basel, Switzerland
1 Joint senior authors.
2 To whom correspondence should be addressed. E-mail: firstname.lastname@example.org.
Published in Toxicological Sciences on 7 March 2017.
Please see here for Editors’ Highlights.
Organ toxicity, particularly liver toxicity, remains one of the major reasons for the termination of drug candidates in the development pipeline as well as withdrawal or restrictions of marketed drugs. A screening-amenable alternative in vivo model such as zebrafish would, therefore, find immediate application in the early prediction of unacceptable organ toxicity. To identify highly upregulated genes as biomarkers of toxic responses in the zebrafish model, a set of well-characterized reference drugs that cause drug-induced liver injury (DILI) in the clinic were applied to zebrafish larvae and adults. Transcriptome microarray analysis was performed on whole larvae or dissected adult livers. Integration of data sets from different drug treatments at different stages identified common upregulated detoxification pathways. Within these were candidate biomarkers which recurred in multiple treatments. We prioritized 4 highly upregulated genes encoding enzymes acting in distinct phases of the drug metabolism pathway. Through promoter isolation and fosmid recombineering, eGFP reporter transgenic zebrafish lines were generated and evaluated for their response to DILI drugs. Three of the 4 generated reporter lines showed a dose and time-dependent induction in endodermal organs to reference drugs and an expanded drug set. In conclusion, through integrated transcriptomics and transgenic approaches, we have developed parallel independent zebrafish in vivo screening platforms able to predict organ toxicities of preclinical drugs.
Reporter transgenic lines responding to DILI drug induction in vivo. Fluorescent images of the intestine region of 5 dpf larvae carrying Tg(cyp2k18:egfp) (A–H), TgFOSMID(gsr:egfp) (I–P) or the TgFOSMID(gstt1b:egfp) (Q–Y) transgenes. The larvae were either untreated (A, I, Q) or challenged for 24h with the DILI compounds: amiodarone (B, J, R), diclofenac (C, K, S), flutamide (D, L, T), Rosiglitazone (E, M, U), Troglitazone (F, N, V), Acetaminophen (APAP; G, O, W) and ROXXXXX53 (H, P, Y). The treatment dosage used for each compound is the respective predetermined LC50 concentration and is given on the left axis for each drug.
For more information on Philip INGHAM’s and Thomas CARNEY’s labs, please click here.