The N-glycoform of sRAGE is the key determinant for its therapeutic efficacy to attenuate injury-elicited arterial inflammation and neointimal growth

Hyun Jin Tae; Ji Min Kim; Sungha Park; Noboru Tomiya; Geng Li; Wen Wei; Natalia Petrashevskaya; Ismayil Ahmet; John Pang; Stefanie Cruschwitz; Rebecca A. Riebe; Yinghua Zhang; Christopher H. Morrell; David Browe; Yuan Chuan Lee; Rui Ping Xiao; Mark I. Talan; Edward G. Lakatta; Li Lin

doi:10.1007/s00109-013-1091-4

The N-glycoform of sRAGE is the key determinant for its therapeutic efficacy to attenuate injury-elicited arterial inflammation and neointimal growth

Hyun Jin Tae, Ji Min Kim, Sungha Park, Noboru Tomiya, Geng Li, Wen Wei, Natalia Petrashevskaya, Ismayil Ahmet, John Pang, Stefanie Cruschwitz, Rebecca A. Riebe, Yinghua Zhang, Christopher H. Morrell, David Browe, Yuan Chuan Lee, Rui Ping Xiao, Mark I. Talan, Edward G. Lakatta, Li Lin

Department of Internal Medicine

Research output: Contribution to journal › Article › peer-review

19 Citations (Scopus)

Abstract

Signaling of the receptor for advanced glycation end products (RAGE) has been implicated in the development of injury-elicited vascular complications. Soluble RAGE (sRAGE) acts as a decoy of RAGE and has been used to treat pathological vascular conditions in animal models. However, previous studies used a high dose of sRAGE produced in insect Sf9 cells (sRAGE^Sf9)and multiple injections to achieve the therapeutic outcome. Here, we explore whether modulation of sRAGE N-glycoform impacts its bioactivity and augments its therapeutic efficacy. We first profiled carbohydrate components of sRAGE produced in Chinese hamster Ovary cells (sRAGE^CHO) to show that a majority of its N-glycans belong to sialylated complex types that are not shared by sRAGE^Sf9. In cell-based NF-κB activation and vascular smooth muscle cell (VSMC) migration assays, sRAGE^CHO exhibited a significantly higher bioactivity relative to sRAGE^Sf9 to inhibit RAGE alarmin ligand-induced NF-κB activation and VSMC migration. We next studied whether this N-glycoform-associated bioactivity of sRAGE^CHO is translated to higher in vivo therapeutic efficacy in a rat carotid artery balloon injury model. Consistent with the observed higher bioactivity in cell assays, sRAGE^CHO significantly reduced injury-induced neointimal growth and the expression of inflammatory markers in injured vasculature. Specifically, a single dose of 3 ng/g of sRAGE^CHO reduced neointimal hyperplasia by over 70 %, whereas the same dose of sRAGE^Sf9 showed no effect. The administered sRAGE^CHO is rapidly and specifically recruited to the injured arterial locus, suggesting that early intervention of arterial injury with sRAGE^CHO may offset an inflammatory circuit and reduce the ensuing tissue remodeling. Our findings showed that the N-glycoform of sRAGE is the key determinant underlying its bioactivity and thus is an important glycobioengineering target to develop a highly potent therapeutic sRAGE for future clinical applications. Key message: The specific N-glycoform modification is the key underlying sRAGE bioactivity Markedly reduced sRAGE dose to attenuate neointimal hyperplasia and inflammation Provide a molecular target for glycobioengineering of sRAGE as a therapeutic protein Blocking RAGE alarmin ligands during acute injury phase offsets neointimal growth

Original language	English
Pages (from-to)	1369-1381
Number of pages	13
Journal	Journal of Molecular Medicine
Volume	91
Issue number	12
DOIs	https://doi.org/10.1007/s00109-013-1091-4
Publication status	Published - 2013 Dec

Bibliographical note

Funding Information:
Acknowledgments We thank Robert Monticone for rat VSMCs and advice on cell migration assays. We also thank reviewers of this manuscript for their inputs that improve our work. The work was supported by the intramural research program of the NIH, National Institute on Aging (LL, MIT, RPX, and EGL), and the Korea Research Foundation grant KRF-2009-013-E00008 (SP). WW was supported in part by the Oak Ridge Institute for Science and Education’s Research Associates Program at NIH; JP was a recipient of the 2011 Johns Hopkins “Excellent in Medical Student Research Award”; and RAR and DB were supported by intramural research training awards from NIH.

All Science Journal Classification (ASJC) codes

Molecular Medicine
Drug Discovery
Genetics(clinical)

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10.1007/s00109-013-1091-4

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Tae, H. J., Kim, J. M., Park, S., Tomiya, N., Li, G., Wei, W., Petrashevskaya, N., Ahmet, I., Pang, J., Cruschwitz, S., Riebe, R. A., Zhang, Y., Morrell, C. H., Browe, D., Lee, Y. C., Xiao, R. P., Talan, M. I., Lakatta, E. G., & Lin, L. (2013). The N-glycoform of sRAGE is the key determinant for its therapeutic efficacy to attenuate injury-elicited arterial inflammation and neointimal growth. Journal of Molecular Medicine, 91(12), 1369-1381. https://doi.org/10.1007/s00109-013-1091-4

@article{67f833972ec04e09953fbd99347b7470,

title = "The N-glycoform of sRAGE is the key determinant for its therapeutic efficacy to attenuate injury-elicited arterial inflammation and neointimal growth",

abstract = "Signaling of the receptor for advanced glycation end products (RAGE) has been implicated in the development of injury-elicited vascular complications. Soluble RAGE (sRAGE) acts as a decoy of RAGE and has been used to treat pathological vascular conditions in animal models. However, previous studies used a high dose of sRAGE produced in insect Sf9 cells (sRAGESf9)and multiple injections to achieve the therapeutic outcome. Here, we explore whether modulation of sRAGE N-glycoform impacts its bioactivity and augments its therapeutic efficacy. We first profiled carbohydrate components of sRAGE produced in Chinese hamster Ovary cells (sRAGECHO) to show that a majority of its N-glycans belong to sialylated complex types that are not shared by sRAGESf9. In cell-based NF-κB activation and vascular smooth muscle cell (VSMC) migration assays, sRAGECHO exhibited a significantly higher bioactivity relative to sRAGESf9 to inhibit RAGE alarmin ligand-induced NF-κB activation and VSMC migration. We next studied whether this N-glycoform-associated bioactivity of sRAGECHO is translated to higher in vivo therapeutic efficacy in a rat carotid artery balloon injury model. Consistent with the observed higher bioactivity in cell assays, sRAGECHO significantly reduced injury-induced neointimal growth and the expression of inflammatory markers in injured vasculature. Specifically, a single dose of 3 ng/g of sRAGECHO reduced neointimal hyperplasia by over 70 %, whereas the same dose of sRAGESf9 showed no effect. The administered sRAGECHO is rapidly and specifically recruited to the injured arterial locus, suggesting that early intervention of arterial injury with sRAGECHO may offset an inflammatory circuit and reduce the ensuing tissue remodeling. Our findings showed that the N-glycoform of sRAGE is the key determinant underlying its bioactivity and thus is an important glycobioengineering target to develop a highly potent therapeutic sRAGE for future clinical applications. Key message: The specific N-glycoform modification is the key underlying sRAGE bioactivity Markedly reduced sRAGE dose to attenuate neointimal hyperplasia and inflammation Provide a molecular target for glycobioengineering of sRAGE as a therapeutic protein Blocking RAGE alarmin ligands during acute injury phase offsets neointimal growth",

author = "Tae, {Hyun Jin} and Kim, {Ji Min} and Sungha Park and Noboru Tomiya and Geng Li and Wen Wei and Natalia Petrashevskaya and Ismayil Ahmet and John Pang and Stefanie Cruschwitz and Riebe, {Rebecca A.} and Yinghua Zhang and Morrell, {Christopher H.} and David Browe and Lee, {Yuan Chuan} and Xiao, {Rui Ping} and Talan, {Mark I.} and Lakatta, {Edward G.} and Li Lin",

note = "Funding Information: Acknowledgments We thank Robert Monticone for rat VSMCs and advice on cell migration assays. We also thank reviewers of this manuscript for their inputs that improve our work. The work was supported by the intramural research program of the NIH, National Institute on Aging (LL, MIT, RPX, and EGL), and the Korea Research Foundation grant KRF-2009-013-E00008 (SP). WW was supported in part by the Oak Ridge Institute for Science and Education{\textquoteright}s Research Associates Program at NIH; JP was a recipient of the 2011 Johns Hopkins “Excellent in Medical Student Research Award”; and RAR and DB were supported by intramural research training awards from NIH.",

year = "2013",

month = dec,

doi = "10.1007/s00109-013-1091-4",

language = "English",

volume = "91",

pages = "1369--1381",

journal = "Journal of Molecular Medicine",

issn = "0946-2716",

publisher = "Springer Verlag",

number = "12",

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Tae, HJ, Kim, JM, Park, S, Tomiya, N, Li, G, Wei, W, Petrashevskaya, N, Ahmet, I, Pang, J, Cruschwitz, S, Riebe, RA, Zhang, Y, Morrell, CH, Browe, D, Lee, YC, Xiao, RP, Talan, MI, Lakatta, EG & Lin, L 2013, 'The N-glycoform of sRAGE is the key determinant for its therapeutic efficacy to attenuate injury-elicited arterial inflammation and neointimal growth', Journal of Molecular Medicine, vol. 91, no. 12, pp. 1369-1381. https://doi.org/10.1007/s00109-013-1091-4

TY - JOUR

T1 - The N-glycoform of sRAGE is the key determinant for its therapeutic efficacy to attenuate injury-elicited arterial inflammation and neointimal growth

AU - Tae, Hyun Jin

AU - Kim, Ji Min

AU - Park, Sungha

AU - Tomiya, Noboru

AU - Li, Geng

AU - Wei, Wen

AU - Petrashevskaya, Natalia

AU - Ahmet, Ismayil

AU - Pang, John

AU - Cruschwitz, Stefanie

AU - Riebe, Rebecca A.

AU - Zhang, Yinghua

AU - Morrell, Christopher H.

AU - Browe, David

AU - Lee, Yuan Chuan

AU - Xiao, Rui Ping

AU - Talan, Mark I.

AU - Lakatta, Edward G.

AU - Lin, Li

N1 - Funding Information: Acknowledgments We thank Robert Monticone for rat VSMCs and advice on cell migration assays. We also thank reviewers of this manuscript for their inputs that improve our work. The work was supported by the intramural research program of the NIH, National Institute on Aging (LL, MIT, RPX, and EGL), and the Korea Research Foundation grant KRF-2009-013-E00008 (SP). WW was supported in part by the Oak Ridge Institute for Science and Education’s Research Associates Program at NIH; JP was a recipient of the 2011 Johns Hopkins “Excellent in Medical Student Research Award”; and RAR and DB were supported by intramural research training awards from NIH.

PY - 2013/12

Y1 - 2013/12

N2 - Signaling of the receptor for advanced glycation end products (RAGE) has been implicated in the development of injury-elicited vascular complications. Soluble RAGE (sRAGE) acts as a decoy of RAGE and has been used to treat pathological vascular conditions in animal models. However, previous studies used a high dose of sRAGE produced in insect Sf9 cells (sRAGESf9)and multiple injections to achieve the therapeutic outcome. Here, we explore whether modulation of sRAGE N-glycoform impacts its bioactivity and augments its therapeutic efficacy. We first profiled carbohydrate components of sRAGE produced in Chinese hamster Ovary cells (sRAGECHO) to show that a majority of its N-glycans belong to sialylated complex types that are not shared by sRAGESf9. In cell-based NF-κB activation and vascular smooth muscle cell (VSMC) migration assays, sRAGECHO exhibited a significantly higher bioactivity relative to sRAGESf9 to inhibit RAGE alarmin ligand-induced NF-κB activation and VSMC migration. We next studied whether this N-glycoform-associated bioactivity of sRAGECHO is translated to higher in vivo therapeutic efficacy in a rat carotid artery balloon injury model. Consistent with the observed higher bioactivity in cell assays, sRAGECHO significantly reduced injury-induced neointimal growth and the expression of inflammatory markers in injured vasculature. Specifically, a single dose of 3 ng/g of sRAGECHO reduced neointimal hyperplasia by over 70 %, whereas the same dose of sRAGESf9 showed no effect. The administered sRAGECHO is rapidly and specifically recruited to the injured arterial locus, suggesting that early intervention of arterial injury with sRAGECHO may offset an inflammatory circuit and reduce the ensuing tissue remodeling. Our findings showed that the N-glycoform of sRAGE is the key determinant underlying its bioactivity and thus is an important glycobioengineering target to develop a highly potent therapeutic sRAGE for future clinical applications. Key message: The specific N-glycoform modification is the key underlying sRAGE bioactivity Markedly reduced sRAGE dose to attenuate neointimal hyperplasia and inflammation Provide a molecular target for glycobioengineering of sRAGE as a therapeutic protein Blocking RAGE alarmin ligands during acute injury phase offsets neointimal growth

AB - Signaling of the receptor for advanced glycation end products (RAGE) has been implicated in the development of injury-elicited vascular complications. Soluble RAGE (sRAGE) acts as a decoy of RAGE and has been used to treat pathological vascular conditions in animal models. However, previous studies used a high dose of sRAGE produced in insect Sf9 cells (sRAGESf9)and multiple injections to achieve the therapeutic outcome. Here, we explore whether modulation of sRAGE N-glycoform impacts its bioactivity and augments its therapeutic efficacy. We first profiled carbohydrate components of sRAGE produced in Chinese hamster Ovary cells (sRAGECHO) to show that a majority of its N-glycans belong to sialylated complex types that are not shared by sRAGESf9. In cell-based NF-κB activation and vascular smooth muscle cell (VSMC) migration assays, sRAGECHO exhibited a significantly higher bioactivity relative to sRAGESf9 to inhibit RAGE alarmin ligand-induced NF-κB activation and VSMC migration. We next studied whether this N-glycoform-associated bioactivity of sRAGECHO is translated to higher in vivo therapeutic efficacy in a rat carotid artery balloon injury model. Consistent with the observed higher bioactivity in cell assays, sRAGECHO significantly reduced injury-induced neointimal growth and the expression of inflammatory markers in injured vasculature. Specifically, a single dose of 3 ng/g of sRAGECHO reduced neointimal hyperplasia by over 70 %, whereas the same dose of sRAGESf9 showed no effect. The administered sRAGECHO is rapidly and specifically recruited to the injured arterial locus, suggesting that early intervention of arterial injury with sRAGECHO may offset an inflammatory circuit and reduce the ensuing tissue remodeling. Our findings showed that the N-glycoform of sRAGE is the key determinant underlying its bioactivity and thus is an important glycobioengineering target to develop a highly potent therapeutic sRAGE for future clinical applications. Key message: The specific N-glycoform modification is the key underlying sRAGE bioactivity Markedly reduced sRAGE dose to attenuate neointimal hyperplasia and inflammation Provide a molecular target for glycobioengineering of sRAGE as a therapeutic protein Blocking RAGE alarmin ligands during acute injury phase offsets neointimal growth

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The N-glycoform of sRAGE is the key determinant for its therapeutic efficacy to attenuate injury-elicited arterial inflammation and neointimal growth

Abstract

Bibliographical note

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