HIF2 ODD-luciferase reporter: the most sensitive assay for HIF prolyl hydroxylase inhibitors

Journal Article · Mon Jan 15 04:00:00 EST 2018 · Russian Chemical Bulletin

DOI:https://doi.org/10.1007/S11172-018-2051-5· OSTI ID:22863395

Smirnova, N. A.; Osipyants, A. I.; Khristichenko, A. Yu.; Hushpulian, D. M.; Nikulin, S. V. ^[1]; Chubar, T. A. ^[2]; Zakhariants, A. A. ^[3]; Tishkov, V. I. ^[2]; Gazaryan, I. G.

---

1. D. Rogachev Federal Scientific and Clinical Center of Pediatric Hematology, Oncology, and Immunology (Russian Federation)
2. M. V. Lomonosov Moscow State University, Department of Chemical Enzymology, Faculty of Chemistry (Russian Federation)
3. Innovation and High Technologies MSU Ltd (Russian Federation)

Reporters expressing fusion proteins of HIF2 and HIF3 C-terminal oxygen degradable domain (ODD) with the firefly luciferase, HIF2 ODD-luc and HIF3 ODD-luc, were constructed and briefly characterized. Stable neuroblastoma cell lines expressing either reporter were generated, and their response to the known HIF prolyl hydroxylase inhibitors: dimethyloxalylglycine, ciclopirox, and adaptaquin, was studied and compared with the HIF1 ODD-luc reporter. The HIF2 ODD-luc reporter exhibited the highest sensitivity: its response in absolute luminescence value was almost an order of magnitude higher than that of the HIF1 ODD-luc reporter. The new reporter can be used for a fine discrimination of enzyme inhibitors stabilizing HIF2, and further structural optimization of adaptaquin discovered earlier by using the HIF1 ODD-luc reporter. The higher sensitivity of HIF2 ODD-luc reporter could be most likely explained by the lower affinity of the endogenous enzyme for this HIF isoform in comparison with the two others, which also resulted in the increased efficiency of inhibitors under the reporter assay conditions.

• OSTI ID: 22863395
• Journal Information: Russian Chemical Bulletin, Journal Name: Russian Chemical Bulletin Journal Issue: 1 Vol. 67; ISSN RCBUEY; ISSN 1066-5285
• Country of Publication: United States
• Language: English

Related Subjects

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY
AFFINITY
ENZYME INHIBITORS
HYDROXYLASES
LUCIFERASE
LUMINESCENCE
OPTIMIZATION
OXYGEN
POLYMERASE CHAIN REACTION