OU‐MRU Vol.72 November 2019

Okayama University Medical Research Updates （OU‐MRU）

2019.11 Vol.72

Source: Okayama University (JAPAN), Public Relations Division
For immediate release: 4 November 2019
Okayama University research: Synthetic compound provides fast screening for potential drugs(Okayama, 4 November) A simple assay may benefit drug discovery for treating diabetes,
Parkinson’s, and Alzheimers disease, as well as studies of functional food and endocrine
disruptor report researchers at Okayama University in the Journal of Medicinal Chemistry.
The assay hinges on a synthetic compound that allows faster screening with fewer
hardware resource requirements than existing methods.
Retinoid X receptors (RXRs) are a type of nuclear receptor ‐ proteins that regulate anorganism’s development, homeostasis and metabolism. They usually operate asheterodimers alongside other proteins and receptors, so the ligands targeting them are key
to controlling their activity. RXR activators have attracted particular interest recently becauseof their potential to treat diabetes, Alzheimers and Parkinsons disease. They are also
associated with functional foods and processes by which environmental pollutants damage
health. However, methods for screening compounds for their potential RXR targeting ligand
activity have so far proved slow and awkward. Associate Professor KAKUTA Hiroki at
University of Okayama Graduate School of Medicine and Shogo Nakano at the University of
Shizuoka in Japan, and their colleagues have now demonstrated an assay based on a synthetic
compound CU‐6PMN ‐ referred to as 10 ‐ that can screen for RXR targeting ligand activity inhours instead of days with no complex equipment or radioactive isotopes needed.
The researchers based the chemical structure of synthetic compound 10, on the RXR activator
CD3254, referred to as compound 9. “Because 9 has a cinnamic acid structure, we anticipatedthat this structure could be developed toward an umbelliferone structure,” they explain in
their report of the work. The significance of umbelliferone is its fluorescence. Not only is thefluorescence of umbelliferone relatively easy to detect ‐ widely available filter sets can detectit ‐ but the compound can also be modified so that its fluorescence intensity increases inaqueous environments. This means that if a compositely binding RXR ligand displaces thereceptor bound compound, the fluorophore will be exposed to a more aqueous environment,
its fluorescence will increase, and the activity of the ligand can be detected.
With compound 10 the researchers showed they could detect RXR targeting ligand activity injust a few hours with standard fluorescence microplate readers and no need for complicatedprocesses. In their report they conclude, “We believe it will be useful not only for identifying
RXR binders in drug discovery studies but also for studies of functional foods and endocrine
disruptors, though it should be noted that fluorescence‐based assays often suffer frominterference when used to screen natural products.”

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OU‐MRU Vol.72 November 2019

Background
Nuclear receptor activity mechanisms
Nuclear receptors are found in cells. They sense the presence of small molecules such as
steroid and thyroid hormones and regulate the expression of genes to control bodily
processes. The human body has 48 types of nuclear receptor. The RXR activator bexarotene
is already used clinically to treat cutaneous T cell lymphoma, and recent studies haverecommended its potential for treating diabetes, Alzheimer’s disease, and Parkinson’sdisease. Polyunsaturated fatty acids including docosahexaenoic acid (DHA) are naturally
occurring RXR targeting ligands and are linked to improved memory and metabolic syndrome.
Conversely the impact of environmental polluters on nuclear receptors can inhibit theirinteraction with hormones and disrupt the endocrine system.
Current assay techniques
To test for the presence and activity of substances, researchers in medicine, pharmacologyand environmental and molecular biology use assays. Previous assays for RXR targeting ligand
activity have also used fluorescence, but they have had drawbacks. Those based on time‐
resolved fluorescence resonance energy transfer, require a special reader plate, while others
have been based on quenching the autofluorescence of the molecule tryptophan, which hasa weak autofluorescence signal at a wavelength that standard readers cannot detect.
Alternative assays have used reporter genes and take 3‐4 days to detect ligand activity, or
they have used radioisotope labelled ligands, where the issues that surround use ofradioactive isotopes further complicate an already complex procedure.

CaptionThe fluorescence from compound 10 increases when an RXR targeting ligand displaces it,
making it a useful indicator for ligand activity in assays.

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1-1-1 Tsushima-naka, Kita-ku, Okayama 700-8530 Japan© OKAYAMA University

OU‐MRU Vol.72 November 2019

Reference
Shoya Yamada, Mayu Kawasaki, Michiko Fujihara, Masaki Watanabe, Yuta Takamura, MahoTakioku, Hiromi Nishioka, Yasuo Takeuchi, Makoto Makishima, Tomoharu Motoyama, Sohei
Ito, Hiroaki Tokiwa, Shogo Nakano, Hiroki Kakuta Competitive binding assay with anumbelliferone‐based fluorescent rexinoid for retinoid X receptor ligand screening. Journal ofMedicinal Chemistry 62, 8809−8818 4 September 2019.
DOI: 10.1021/acs.jmedchem.9b00995
https://pubs.acs.org/doi/abs/10.1021/acs.jmedchem.9b00995

Correspondence toAssociate Professor KAKUTA Hiroki, Ph.D.
Division of Pharmaceutical Sciences,
Graduate School of Medicine, Dentistry and PharmaceuticalSciences, Okayama University, 1‐1‐1, Tsushima‐naka, Kita‐ku,Okayama 700‐8530, JapanE‐mail: kakuta‐h@okayama‐u.ac.jp
Associate Professor
http://www.cc.okayama‐u.ac.jp/~kakuta‐h/kakuta_lab_english/index.html
KAKUTA Hiroki

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OU‐MRU Vol.72 November 2019

◆About Okayama UniversityOkayama University is one of the largest comprehensive universities in Japan withroots going back to the Medical Training Place sponsored by the Lord of Okayama andestablished in 1870. Now with 1,300 faculty and 13,000 students, the University offers
courses in specialties ranging from medicine and pharmacy to humanities and physicalsciences.
Okayama University is located in the heart of Japan approximately 3 hours west ofTokyo by Shinkansen.
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