RNA Aptamer Binds Heparin-Binding Epidermal Growth Factor-Like Growth Factor with High Affinity and Specificity and Neutralizes Its Activity
Background: Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is a member of the EGF family and is involved in various diseases including cancers. Aptamers are synthetic oligonucleotides (RNA or DNA) that fold into unique three-dimensional structures and specifically bind to their...
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Format: | Article |
Language: | English |
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Taiwan Society of Geriatric Emergency and Critical Medicine (TSGECM)
2017-09-01
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Series: | International Journal of Gerontology |
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Online Access: | http://www.sciencedirect.com/science/article/pii/S1873959817300145 |
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record_format |
Article |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Masaki Yamato Takashi Matsuzaki Ryo Araki Shota Tsuchida Keiji Okuda Hai Ying Fu Shoji Sanada Hiroshi Asanuma Yoshihiro Asano Masanori Asakura Hiroomi Torii Kentaro Noi Hirotsugu Ogi Ryo Iwamoto Eisuke Mekada Seiji Takashima Masafumi Kitakaze Yasushi Sakata Tetsuo Minamino |
spellingShingle |
Masaki Yamato Takashi Matsuzaki Ryo Araki Shota Tsuchida Keiji Okuda Hai Ying Fu Shoji Sanada Hiroshi Asanuma Yoshihiro Asano Masanori Asakura Hiroomi Torii Kentaro Noi Hirotsugu Ogi Ryo Iwamoto Eisuke Mekada Seiji Takashima Masafumi Kitakaze Yasushi Sakata Tetsuo Minamino RNA Aptamer Binds Heparin-Binding Epidermal Growth Factor-Like Growth Factor with High Affinity and Specificity and Neutralizes Its Activity International Journal of Gerontology aptamer heparin-binding epidermal growth factor-like growth factor |
author_facet |
Masaki Yamato Takashi Matsuzaki Ryo Araki Shota Tsuchida Keiji Okuda Hai Ying Fu Shoji Sanada Hiroshi Asanuma Yoshihiro Asano Masanori Asakura Hiroomi Torii Kentaro Noi Hirotsugu Ogi Ryo Iwamoto Eisuke Mekada Seiji Takashima Masafumi Kitakaze Yasushi Sakata Tetsuo Minamino |
author_sort |
Masaki Yamato |
title |
RNA Aptamer Binds Heparin-Binding Epidermal Growth Factor-Like Growth Factor with High Affinity and Specificity and Neutralizes Its Activity |
title_short |
RNA Aptamer Binds Heparin-Binding Epidermal Growth Factor-Like Growth Factor with High Affinity and Specificity and Neutralizes Its Activity |
title_full |
RNA Aptamer Binds Heparin-Binding Epidermal Growth Factor-Like Growth Factor with High Affinity and Specificity and Neutralizes Its Activity |
title_fullStr |
RNA Aptamer Binds Heparin-Binding Epidermal Growth Factor-Like Growth Factor with High Affinity and Specificity and Neutralizes Its Activity |
title_full_unstemmed |
RNA Aptamer Binds Heparin-Binding Epidermal Growth Factor-Like Growth Factor with High Affinity and Specificity and Neutralizes Its Activity |
title_sort |
rna aptamer binds heparin-binding epidermal growth factor-like growth factor with high affinity and specificity and neutralizes its activity |
publisher |
Taiwan Society of Geriatric Emergency and Critical Medicine (TSGECM) |
series |
International Journal of Gerontology |
issn |
1873-9598 |
publishDate |
2017-09-01 |
description |
Background: Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is a member of the EGF family and is involved in various diseases including cancers. Aptamers are synthetic oligonucleotides (RNA or DNA) that fold into unique three-dimensional structures and specifically bind to their targets with high affinity. We aimed to generate an aptamer with high affinity and specificity for HB-EGF.
Methods: Recombinant human HB-EGF (rhHB-EGF), comprised of the extracellular EGF-like and heparin-binding domains of HB-EGF, was used as the target. The aptamer against HB-EGF (the anti-HB-EGF aptamer) was obtained by systematic evolution of ligands by exponential enrichment (SELEX).
Results: After the 10th round of SELEX, aptamers were reverse-transcribed and PCR-amplified. Within obtained forty-six clones, twenty-three were identical (the anti-HB-EGF aptamer). The analysis using wireless-electrodeless quartz crystal microbalance revealed that the anti-HB-EGF aptamer had high affinity for rhHB-EGF (KD value: 12.2 ± 1.1 nmol/L). The dot-blot analysis revealed that the anti-HB-EGF aptamer specifically bound to rhHB-EGF. The analysis using confocal microscopy indicated that the anti-HB-EGF aptamer also bound to membrane-bound HB-EGF. Western-blot assay indicated that the anti-HB-EGF aptamer inhibited the phosphorylation of rhHB-EGF-mediated EGF receptor (EGFR).
Conclusion: We identified a novel RNA aptamer that bound with high affinity and specificity to rhHB-EGF and potently inhibited the rhHB-EGF-mediated phosphorylation of EGFR. The anti-HB-EGF aptamer may be a promising therapeutic agent for specifically neutralizing HB-EGF signaling. |
topic |
aptamer heparin-binding epidermal growth factor-like growth factor |
url |
http://www.sciencedirect.com/science/article/pii/S1873959817300145 |
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doaj-c70a3e26851f4d079843bf28b2606e382020-11-24T22:12:27ZengTaiwan Society of Geriatric Emergency and Critical Medicine (TSGECM)International Journal of Gerontology1873-95982017-09-0111319119610.1016/j.ijge.2017.03.006RNA Aptamer Binds Heparin-Binding Epidermal Growth Factor-Like Growth Factor with High Affinity and Specificity and Neutralizes Its ActivityMasaki Yamato0Takashi Matsuzaki1Ryo Araki2Shota Tsuchida3Keiji Okuda4Hai Ying Fu5Shoji Sanada6Hiroshi Asanuma7Yoshihiro Asano8Masanori Asakura9Hiroomi Torii10Kentaro Noi11Hirotsugu Ogi12Ryo Iwamoto13Eisuke Mekada14Seiji Takashima15Masafumi Kitakaze16Yasushi Sakata17Tetsuo Minamino18Department of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of Internal Medicine, Meiji University of Integrative Medicine, Kyoto, JapanDepartment of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Osaka, JapanCardiovascular Division, Department of Internal Medicine, Hyogo College of Medicine, Hyogo, JapanGraduate School of Engineering Science, Osaka University, Osaka, JapanDepartment of Molecular Cell Biology, Institute of Molecular Embryology and Genetics, Kumamoto University, Kumamoto, JapanGraduate School of Engineering Science, Osaka University, Osaka, JapanDepartment of Cell Biology, Research Institute for Microbial Diseases, Osaka University, Osaka, JapanDepartment of Cell Biology, Research Institute for Microbial Diseases, Osaka University, Osaka, JapanDepartment of Medical Biochemistry, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of Clinical Research and Development, National Cerebral and Cardiovascular Center, Osaka, JapanDepartment of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of Cardiorenal and Cerebrovascular Medicine, Faculty of Medicine, Kagawa University, Kagawa, JapanBackground: Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is a member of the EGF family and is involved in various diseases including cancers. Aptamers are synthetic oligonucleotides (RNA or DNA) that fold into unique three-dimensional structures and specifically bind to their targets with high affinity. We aimed to generate an aptamer with high affinity and specificity for HB-EGF. Methods: Recombinant human HB-EGF (rhHB-EGF), comprised of the extracellular EGF-like and heparin-binding domains of HB-EGF, was used as the target. The aptamer against HB-EGF (the anti-HB-EGF aptamer) was obtained by systematic evolution of ligands by exponential enrichment (SELEX). Results: After the 10th round of SELEX, aptamers were reverse-transcribed and PCR-amplified. Within obtained forty-six clones, twenty-three were identical (the anti-HB-EGF aptamer). The analysis using wireless-electrodeless quartz crystal microbalance revealed that the anti-HB-EGF aptamer had high affinity for rhHB-EGF (KD value: 12.2 ± 1.1 nmol/L). The dot-blot analysis revealed that the anti-HB-EGF aptamer specifically bound to rhHB-EGF. The analysis using confocal microscopy indicated that the anti-HB-EGF aptamer also bound to membrane-bound HB-EGF. Western-blot assay indicated that the anti-HB-EGF aptamer inhibited the phosphorylation of rhHB-EGF-mediated EGF receptor (EGFR). Conclusion: We identified a novel RNA aptamer that bound with high affinity and specificity to rhHB-EGF and potently inhibited the rhHB-EGF-mediated phosphorylation of EGFR. The anti-HB-EGF aptamer may be a promising therapeutic agent for specifically neutralizing HB-EGF signaling.http://www.sciencedirect.com/science/article/pii/S1873959817300145aptamerheparin-binding epidermal growth factor-like growth factor |