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Title: A Potent d-Protein Antagonist of VEGF-A is Nonimmunogenic, Metabolically Stable, and Longer-Circulating in Vivo

Abstract

Polypeptides composed entirely of d-amino acids and the achiral amino acid glycine (d-proteins) inherently have in vivo properties that are proposed to be near-optimal for a large molecule therapeutic agent. Specifically, d-proteins are resistant to degradation by proteases and are anticipated to be nonimmunogenic. Furthermore, d-proteins are manufactured chemically and can be engineered to have other desirable properties, such as improved stability, affinity, and pharmacokinetics. Thus, a well-designed d-protein therapeutic would likely have significant advantages over l-protein drugs. Toward the goal of developing d-protein therapeutics, we previously generated RFX001.D, a d-protein antagonist of natural vascular endothelial growth factor A (VEGF-A) that inhibited binding to its receptor. However, RFX001.D is unstable at physiological temperatures (Tm = 33 °C). Here, we describe RFX037.D, a variant of RFX001.D with extreme thermal stability (Tm > 95 °C), high affinity for VEGF-A (Kd = 6 nM), and improved receptor blocking. Comparison of the two enantiomeric forms of RFX037 revealed that the d-protein is more stable in mouse, monkey, and human plasma and has a longer half-life in vivo in mice. Significantly, RFX037.D was nonimmunogenic in mice, whereas the l-enantiomer generated a strong immune response. These results confirm the potential utility of synthetic d-proteins as alternativesmore » to therapeutic antibodies.« less

Authors:
 [1];  [2];  [2];  [3];  [4];  [5];  [5];  [1];  [6];  [2];  [1]
  1. Banting and Best Department of Medical Research and Department of Molecular Genetics, the Donnelly Centre, University of Toronto, Toronto, Ontario, Canada M5S 3E1
  2. Department of Chemistry, Department of Biochemistry and Molecular Biology, Institute for Biophysical Dynamics, University of Chicago, Chicago, Illinois 60637, United States
  3. Pharmacokinetics &, Drug Metabolism, Amgen, Inc., Thousand Oaks, California 91320, United States
  4. Therapeutic Discovery, Amgen, Inc., Thousand Oaks, California 91320, United States
  5. Antibody Solutions, Sunnyvale, California 94089, United States
  6. Reflexion Pharmaceuticals, Inc., San Francisco, California 94104, United States
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES)
OSTI Identifier:
1236725
Alternate Identifier(s):
OSTI ID: 1249238
Grant/Contract Number:  
AC02-06CH11357
Resource Type:
Published Article
Journal Name:
ACS Chemical Biology
Additional Journal Information:
Journal Name: ACS Chemical Biology Journal Volume: 11 Journal Issue: 4; Journal ID: ISSN 1554-8929
Publisher:
American Chemical Society (ACS)
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; immunology; peptides and proteins; rodent models; plasma; stability

Citation Formats

Uppalapati, Maruti, Lee, Dong Jun, Mandal, Kalyaneswar, Li, Hongyan, Miranda, Les P., Lowitz, Joshua, Kenney, John, Adams, Jarrett J., Ault-Riché, Dana, Kent, Stephen B. H., and Sidhu, Sachdev S. A Potent d-Protein Antagonist of VEGF-A is Nonimmunogenic, Metabolically Stable, and Longer-Circulating in Vivo. United States: N. p., 2016. Web. https://doi.org/10.1021/acschembio.5b01006.
Uppalapati, Maruti, Lee, Dong Jun, Mandal, Kalyaneswar, Li, Hongyan, Miranda, Les P., Lowitz, Joshua, Kenney, John, Adams, Jarrett J., Ault-Riché, Dana, Kent, Stephen B. H., & Sidhu, Sachdev S. A Potent d-Protein Antagonist of VEGF-A is Nonimmunogenic, Metabolically Stable, and Longer-Circulating in Vivo. United States. https://doi.org/10.1021/acschembio.5b01006
Uppalapati, Maruti, Lee, Dong Jun, Mandal, Kalyaneswar, Li, Hongyan, Miranda, Les P., Lowitz, Joshua, Kenney, John, Adams, Jarrett J., Ault-Riché, Dana, Kent, Stephen B. H., and Sidhu, Sachdev S. Wed . "A Potent d-Protein Antagonist of VEGF-A is Nonimmunogenic, Metabolically Stable, and Longer-Circulating in Vivo". United States. https://doi.org/10.1021/acschembio.5b01006.
@article{osti_1236725,
title = {A Potent d-Protein Antagonist of VEGF-A is Nonimmunogenic, Metabolically Stable, and Longer-Circulating in Vivo},
author = {Uppalapati, Maruti and Lee, Dong Jun and Mandal, Kalyaneswar and Li, Hongyan and Miranda, Les P. and Lowitz, Joshua and Kenney, John and Adams, Jarrett J. and Ault-Riché, Dana and Kent, Stephen B. H. and Sidhu, Sachdev S.},
abstractNote = {Polypeptides composed entirely of d-amino acids and the achiral amino acid glycine (d-proteins) inherently have in vivo properties that are proposed to be near-optimal for a large molecule therapeutic agent. Specifically, d-proteins are resistant to degradation by proteases and are anticipated to be nonimmunogenic. Furthermore, d-proteins are manufactured chemically and can be engineered to have other desirable properties, such as improved stability, affinity, and pharmacokinetics. Thus, a well-designed d-protein therapeutic would likely have significant advantages over l-protein drugs. Toward the goal of developing d-protein therapeutics, we previously generated RFX001.D, a d-protein antagonist of natural vascular endothelial growth factor A (VEGF-A) that inhibited binding to its receptor. However, RFX001.D is unstable at physiological temperatures (Tm = 33 °C). Here, we describe RFX037.D, a variant of RFX001.D with extreme thermal stability (Tm > 95 °C), high affinity for VEGF-A (Kd = 6 nM), and improved receptor blocking. Comparison of the two enantiomeric forms of RFX037 revealed that the d-protein is more stable in mouse, monkey, and human plasma and has a longer half-life in vivo in mice. Significantly, RFX037.D was nonimmunogenic in mice, whereas the l-enantiomer generated a strong immune response. These results confirm the potential utility of synthetic d-proteins as alternatives to therapeutic antibodies.},
doi = {10.1021/acschembio.5b01006},
journal = {ACS Chemical Biology},
number = 4,
volume = 11,
place = {United States},
year = {2016},
month = {2}
}

Journal Article:
Free Publicly Available Full Text
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https://doi.org/10.1021/acschembio.5b01006

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