June 2017
Volume 58, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2017
Whole-organism HTS Identifies Synergistic Drug Combinations Promoting Rod Photoreceptor Survival in a Zebrafish Model of Retinitis Pigmentosa
Author Affiliations & Notes
  • Liyun Zhang
    Wilmer Eye Inst-Smith Bldg Rm 4001, Johns Hopkins School of Medicine, Baltimore, Maryland, United States
  • Conan Chen
    Wilmer Eye Inst-Smith Bldg Rm 4001, Johns Hopkins School of Medicine, Baltimore, Maryland, United States
  • Jeff S Mumm
    Wilmer Eye Inst-Smith Bldg Rm 4001, Johns Hopkins School of Medicine, Baltimore, Maryland, United States
  • Footnotes
    Commercial Relationships   Liyun Zhang, None; Conan Chen, None; Jeff Mumm, None
  • Footnotes
    Support  Foundation Fighting Blindness / Wynn-Gund TRAP Award
Investigative Ophthalmology & Visual Science June 2017, Vol.58, 4539. doi:
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      Liyun Zhang, Conan Chen, Jeff S Mumm; Whole-organism HTS Identifies Synergistic Drug Combinations Promoting Rod Photoreceptor Survival in a Zebrafish Model of Retinitis Pigmentosa. Invest. Ophthalmol. Vis. Sci. 2017;58(8):4539.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Retinitis pigmentosa (RP) is caused by the death of rod photoreceptors. The purpose of this study was to use a whole-organism high-throughput screening (HTS) platform to identify human-approved drugs which promote the survival of rod photoreceptors in an inducible zebrafish model of RP.

Methods : Transgenic zebrafish which enabled prodrug (metronidazole)-induced rod cell apoptosis were used to identify drugs that protect rod cells. Five day old zebrafish were dispensed into individual wells of a 96-well plate containing test compounds from the John Hopkins Drug Library (2934 predominantly human-approved drugs). Each drug was tested across six concentrations (4µM - 125nM) and 16 fish per condition; with controls, more than 500,000 fish were screened. After a 4 hr pre-exposure to test drugs, larvae were exposed to 2.5 mM metronidazole to induce rod cell apoptosis. Surviving rod cell numbers were then quantified by measuring reporter levels with a fluorescent plate reader. Effect sizes were compared across compounds to establish and prioritize “hits”. Top-performing hits underwent a minimum of three biological repeats for validation; confocal microscopy was used to verify effects on rod cell protection. Validated hits were then assessed in combination to test for additive effects on rod cell protection.

Results : Validation efforts focused on 42 top-performing hits. Of these, 8 compounds passed all validation criteria, including visual verification of preserved rod cell numbers, and have moved forward as lead drug candidates. Interestingly, seven separate signaling pathways, based on known molecular mechanism of action (MoA) data, were predicted to function in rod cell neuroprotection. Combinatorial assays demonstrated additive effects for 7 of 28 pairs tested, producing improved rod cell protection relative to individual drug treatments.

Conclusions : Whole-organism HTS drug screening successfully identified several potentially synergistic neuroprotective compounds in an inducible zebrafish model of RP. We conclude that combinatorial therapies targeting complementary neuroprotective signaling pathways may provide an improved strategy for protecting and maintaining function of rod photoreceptors in RP patients.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.

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