Improving the Pharmacodynamics and In Vivo Activity of ENPP1‐Fc Through Protein and Glycosylation Engineering
Enzyme replacement with ectonucleotide pyrophosphatase phospodiesterase‐1 (ENPP1) eliminates mortality in a murine model of the lethal calcification disorder generalized arterial calcification of infancy. We used protein engineering, glycan optimization, and a novel biomanufacturing platform to enha...
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doaj-eb3329ba4a5240e181f00b874aa1535e2021-02-11T20:06:36ZengWileyClinical and Translational Science1752-80541752-80622021-01-0114136237210.1111/cts.12887Improving the Pharmacodynamics and In Vivo Activity of ENPP1‐Fc Through Protein and Glycosylation EngineeringPaul R. Stabach0Kristin Zimmerman1Aaron Adame2Dillon Kavanagh3Christopher T. Saeui4Christian Agatemor5Shawn Gray6Wenxiang Cao7Enrique M. De La Cruz8Kevin J. Yarema9Demetrios T. Braddock10Department of Pathology Yale University School of Medicine New Haven Connecticut USADepartment of Pathology Yale University School of Medicine New Haven Connecticut USADepartment of Pathology Yale University School of Medicine New Haven Connecticut USADepartment of Pathology Yale University School of Medicine New Haven Connecticut USADepartment of Biomedical Engineering Translational Tissue Engineering Center The Johns Hopkins University Baltimore Maryland USADepartment of Biomedical Engineering Translational Tissue Engineering Center The Johns Hopkins University Baltimore Maryland USADepartment of Molecular Biophysics and Biochemistry Yale University New Haven Connecticut USADepartment of Molecular Biophysics and Biochemistry Yale University New Haven Connecticut USADepartment of Molecular Biophysics and Biochemistry Yale University New Haven Connecticut USADepartment of Biomedical Engineering Translational Tissue Engineering Center The Johns Hopkins University Baltimore Maryland USADepartment of Pathology Yale University School of Medicine New Haven Connecticut USAEnzyme replacement with ectonucleotide pyrophosphatase phospodiesterase‐1 (ENPP1) eliminates mortality in a murine model of the lethal calcification disorder generalized arterial calcification of infancy. We used protein engineering, glycan optimization, and a novel biomanufacturing platform to enhance potency by using a three‐prong strategy. First, we added new N‐glycans to ENPP1; second, we optimized pH‐dependent cellular recycling by protein engineering of the Fc neonatal receptor; finally, we used a two‐step process to improve sialylation by first producing ENPP1‐Fc in cells stably transfected with human α‐2,6‐sialyltransferase (ST6) and further enhanced terminal sialylation by supplementing production with 1,3,4‐O‐Bu3ManNAc. These steps sequentially increased the half‐life of the parent compound in rodents from 37 hours to ~ 67 hours with an added N‐glycan, to ~ 96 hours with optimized pH‐dependent Fc recycling, to ~ 204 hours when the therapeutic was produced in ST6‐overexpressing cells with 1,3,4‐O‐Bu3ManNAc supplementation. The alterations were demonstrated to increase drug potency by maintaining efficacious levels of plasma phosphoanhydride pyrophosphate in ENPP1‐deficient mice when the optimized biologic was administered at a 10‐fold lower mass dose less frequently than the parent compound—once every 10 days vs. 3 times a week. We believe these improvements represent a general strategy to rationally optimize protein therapeutics.https://doi.org/10.1111/cts.12887 |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Paul R. Stabach Kristin Zimmerman Aaron Adame Dillon Kavanagh Christopher T. Saeui Christian Agatemor Shawn Gray Wenxiang Cao Enrique M. De La Cruz Kevin J. Yarema Demetrios T. Braddock |
spellingShingle |
Paul R. Stabach Kristin Zimmerman Aaron Adame Dillon Kavanagh Christopher T. Saeui Christian Agatemor Shawn Gray Wenxiang Cao Enrique M. De La Cruz Kevin J. Yarema Demetrios T. Braddock Improving the Pharmacodynamics and In Vivo Activity of ENPP1‐Fc Through Protein and Glycosylation Engineering Clinical and Translational Science |
author_facet |
Paul R. Stabach Kristin Zimmerman Aaron Adame Dillon Kavanagh Christopher T. Saeui Christian Agatemor Shawn Gray Wenxiang Cao Enrique M. De La Cruz Kevin J. Yarema Demetrios T. Braddock |
author_sort |
Paul R. Stabach |
title |
Improving the Pharmacodynamics and In Vivo Activity of ENPP1‐Fc Through Protein and Glycosylation Engineering |
title_short |
Improving the Pharmacodynamics and In Vivo Activity of ENPP1‐Fc Through Protein and Glycosylation Engineering |
title_full |
Improving the Pharmacodynamics and In Vivo Activity of ENPP1‐Fc Through Protein and Glycosylation Engineering |
title_fullStr |
Improving the Pharmacodynamics and In Vivo Activity of ENPP1‐Fc Through Protein and Glycosylation Engineering |
title_full_unstemmed |
Improving the Pharmacodynamics and In Vivo Activity of ENPP1‐Fc Through Protein and Glycosylation Engineering |
title_sort |
improving the pharmacodynamics and in vivo activity of enpp1‐fc through protein and glycosylation engineering |
publisher |
Wiley |
series |
Clinical and Translational Science |
issn |
1752-8054 1752-8062 |
publishDate |
2021-01-01 |
description |
Enzyme replacement with ectonucleotide pyrophosphatase phospodiesterase‐1 (ENPP1) eliminates mortality in a murine model of the lethal calcification disorder generalized arterial calcification of infancy. We used protein engineering, glycan optimization, and a novel biomanufacturing platform to enhance potency by using a three‐prong strategy. First, we added new N‐glycans to ENPP1; second, we optimized pH‐dependent cellular recycling by protein engineering of the Fc neonatal receptor; finally, we used a two‐step process to improve sialylation by first producing ENPP1‐Fc in cells stably transfected with human α‐2,6‐sialyltransferase (ST6) and further enhanced terminal sialylation by supplementing production with 1,3,4‐O‐Bu3ManNAc. These steps sequentially increased the half‐life of the parent compound in rodents from 37 hours to ~ 67 hours with an added N‐glycan, to ~ 96 hours with optimized pH‐dependent Fc recycling, to ~ 204 hours when the therapeutic was produced in ST6‐overexpressing cells with 1,3,4‐O‐Bu3ManNAc supplementation. The alterations were demonstrated to increase drug potency by maintaining efficacious levels of plasma phosphoanhydride pyrophosphate in ENPP1‐deficient mice when the optimized biologic was administered at a 10‐fold lower mass dose less frequently than the parent compound—once every 10 days vs. 3 times a week. We believe these improvements represent a general strategy to rationally optimize protein therapeutics. |
url |
https://doi.org/10.1111/cts.12887 |
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