Yui Kuroda1, Miki Nonaka2, Yuji Kamikubo3, Haruo Ogawa4, Takashi Murayama3, Nagomi Kurebayashi3, Hakushun Sakairi3, Kanako Miyano5, Akane Komatsu1, Tetsushi Dodo6, Kyoko Nakano-Ito7, Keisuke Yamaguchi8, Takashi Sakurai3, Masako Iseki9, Masakazu Hayashida9, Yasuhito Uezono10. 1. Department of Anesthesiology and Pain Medicine, Juntendo University Graduate School of Medicine, Tokyo, Japan; Department of Pain Control Research, The Jikei University School of Medicine, Tokyo, Japan. 2. Department of Pain Control Research, The Jikei University School of Medicine, Tokyo, Japan. 3. Department of Cellular and Molecular Pharmacology, Juntendo University Graduate School of Medicine, Tokyo, Japan. 4. Institute of Molecular and Cellular Biosciences, The University of Tokyo, Tokyo, Japan. 5. Department of Pain Control Research, The Jikei University School of Medicine, Tokyo, Japan; Division of Cancer Pathophysiology, National Cancer Center Research Institute, Tokyo, Japan. 6. Strategy Planning & Operations, Medicine Development Center, Eisai Co., Ltd., Ibaraki, Japan. 7. Global Drug Safety, Medicine Development Center, Eisai Co., Ltd., Ibaraki, Japan. 8. Department of Pain Medicine, Juntendo University Graduate School of Medicine, Tokyo, Japan. 9. Department of Anesthesiology and Pain Medicine, Juntendo University Graduate School of Medicine, Tokyo, Japan; Department of Pain Medicine, Juntendo University Graduate School of Medicine, Tokyo, Japan. 10. Department of Pain Control Research, The Jikei University School of Medicine, Tokyo, Japan; Department of Pain Medicine, Juntendo University Graduate School of Medicine, Tokyo, Japan; Supportive and Palliative Care Research Support Office, National Center Hospital East, Chiba, Japan; Project for Supportive Care Research, Exploratory Oncology Research and Clinical Trial Center, National Cancer Center, Tokyo, Japan. Electronic address: yuezono@jikei.ac.jp.
Abstract
BACKGROUND: The misuse of opioids has led to an epidemic in recent times. The endothelin A receptor (ETAR) has recently attracted attention as a novel therapeutic target to enhance opioid analgesia. We hypothesized that endothelin A receptors may affect pain mechanisms by heterodimerization with μ opioid receptors. We examined the mechanisms of ETAR-mediated pain and the potential therapeutic effects of an ETAR antagonist, Compound-E, as an agent for analgesia. METHODS: Real-time in vitro effect of Compound-E on morphine response was assessed in HEK293 cells expressing both endothelin A and μ opioid receptors through CellKey™ and cADDis cAMP assays. Endothelin A/μ opioid receptor dimerization was assessed by immunoprecipitation and live cell imaging. The in vivo effect of Compound-E was evaluated using a morphine analgesia mouse model that observed escape response behavior, body temperature, and locomotor activity. RESULTS: In CellKey™ and cAMP assays, pretreatment of cells with endothelin-1 attenuated morphine-induced responses. These responses were improved by Compound-E, but not by BQ-123 nor by bosentan, an ETAR and endothelin B receptor antagonist. Dimerization of ETARs and μ opioid receptors was confirmed by Western blot and total internal reflection fluorescence microscopy in live cells. In vivo, Compound-E potentiated and prolonged the analgesic effects of morphine, enhanced hypothermia, and increased locomotor activity compared to morphine alone. CONCLUSION: The results suggest that attenuation by endothelin-1 of morphine analgesia may be caused by dimerization of Endothelin A/μ opioid receptors. The novel ETAR antagonist Compound-E could be an effective adjunct to reduce opioid use.
BACKGROUND: The misuse of opioids has led to an epidemic in recent times. The endothelin A receptor (ETAR) has recently attracted attention as a novel therapeutic target to enhance opioid analgesia. We hypothesized that endothelin A receptors may affect pain mechanisms by heterodimerization with μ opioid receptors. We examined the mechanisms of ETAR-mediated pain and the potential therapeutic effects of an ETAR antagonist, Compound-E, as an agent for analgesia. METHODS: Real-time in vitro effect of Compound-E on morphine response was assessed in HEK293 cells expressing both endothelin A and μ opioid receptors through CellKey™ and cADDis cAMP assays. Endothelin A/μ opioid receptor dimerization was assessed by immunoprecipitation and live cell imaging. The in vivo effect of Compound-E was evaluated using a morphine analgesia mouse model that observed escape response behavior, body temperature, and locomotor activity. RESULTS: In CellKey™ and cAMP assays, pretreatment of cells with endothelin-1 attenuated morphine-induced responses. These responses were improved by Compound-E, but not by BQ-123 nor by bosentan, an ETAR and endothelin B receptor antagonist. Dimerization of ETARs and μ opioid receptors was confirmed by Western blot and total internal reflection fluorescence microscopy in live cells. In vivo, Compound-E potentiated and prolonged the analgesic effects of morphine, enhanced hypothermia, and increased locomotor activity compared to morphine alone. CONCLUSION: The results suggest that attenuation by endothelin-1 of morphine analgesia may be caused by dimerization of Endothelin A/μ opioid receptors. The novel ETAR antagonist Compound-E could be an effective adjunct to reduce opioid use.
Authors: David Speck; Gunnar Kleinau; Michal Szczepek; Dennis Kwiatkowski; Rusan Catar; Aurélie Philippe; Patrick Scheerer Journal: Front Endocrinol (Lausanne) Date: 2022-04-19 Impact factor: 6.055
Authors: Sam R J Hoare; Paul H Tewson; Shivani Sachdev; Mark Connor; Thomas E Hughes; Anne Marie Quinn Journal: Front Cell Neurosci Date: 2022-01-17 Impact factor: 5.505