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Development of Coagulation Factor XII Antibodies for Inhibiting Vascular Device-Related Thrombosis

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Abstract

Introduction

Vascular devices such as stents, hemodialyzers, and membrane oxygenators can activate blood coagulation and often require the use of systemic anticoagulants to selectively prevent intravascular thrombotic/embolic events or extracorporeal device failure. Coagulation factor (F)XII of the contact activation system has been shown to play an important role in initiating vascular device surface-initiated thrombus formation. As FXII is dispensable for hemostasis, targeting the contact activation system holds promise as a significantly safer strategy than traditional antithrombotics for preventing vascular device-associated thrombosis.

Objective

Generate and characterize anti-FXII monoclonal antibodies that inhibit FXII activation or activity.

Methods

Monoclonal antibodies against FXII were generated in FXII-deficient mice and evaluated for their binding and anticoagulant properties in purified and plasma systems, in whole blood flow-based assays, and in an in vivo non-human primate model of vascular device-initiated thrombus formation.

Results

A FXII antibody screen identified over 400 candidates, which were evaluated in binding studies and clotting assays. One non-inhibitor and six inhibitor antibodies were selected for characterization in functional assays. The most potent inhibitory antibody, 1B2, was found to prolong clotting times, inhibit fibrin generation on collagen under shear, and inhibit platelet deposition and fibrin formation in an extracorporeal membrane oxygenator deployed in a non-human primate.

Conclusion

Selective contact activation inhibitors hold potential as useful tools for research applications as well as safe and effective inhibitors of vascular device-related thrombosis.

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Abbreviations

ACT:

Activated clotting time

aPTT:

Activated partial thromboplastin time

BK:

Bradykinin

BT:

Bleeding time

CAS:

Contact activation system

CBC:

Complete blood count

cDNA:

Complementary DNA

CFT:

Clot formation time

CT:

Clotting time

ECMO:

Extracorporeal membrane oxygenation

ECOS:

Extracorporeal organ support

F:

Factor

HGFA:

Hepatocyte growth factor activator

HK:

High molecular weight kininogen

IgG:

Immunoglobulin G

IP:

Intraperitoneal

mAb:

Monoclonal antibody

MCF:

Maximum clot firmness

NATEM:

Non-activated thromboelastometry

NHP:

Non-human primate

PCR:

Polymerase chain reaction

PK:

Prekallikrein

polyP:

Polyphosphate

PPP:

Platelet-poor plasma

PT:

Prothrombin time

TAT:

Thrombin-antithrombin

VAD:

Ventricular assist device

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Acknowledgments

We thank H. Lakshmanan, M. Carris, and J. Pang for technical assistance and valuable discussions. We also thank Dr. Sally Nofs (Nashville Zoo at Grassmere, Nashville, TN) for mammalian plasma samples. This study was supported in part by the National Heart, Lung, and Blood Institute Grants: HL126235 to E.I. Tucker and M. Wallisch, R35HL140025 to D. Gailani, and HL144113 to O.J.T. McCarty and M.T. Hinds; and the Oregon National Primate Research Center NIH Grant Award P51OD011092.

Conflict of interest

M. Wallisch, C.U. Lorentz, E.I. Tucker, and A. Gruber are employees of Aronora, Inc., and they as well as Oregon Health & Science University may have a financial interest in the results of this study. J.J. Shatzel reports receiving consulting fees from Aronora, Inc. T.C.L. Kohs, J. Johnson, C. Puy, S.R. Olson, D. Gailani, M. T. Hinds, and O.J.T. McCarty state that they have no conflicts of interest.

Research Involving Human Rights

All human subjects research was carried out in accordance with institutional guidelines approved by the Oregon Health & Science University Institutional Review Board.

Research Involving Animal Rights

All animal studies were approved by the Oregon Health & Science University Institutional Animal Care and Use Committee. All institutional and national guidelines for the care and use of laboratory animals were followed and approved by our institutional committee.

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Authors and Affiliations

  1. Department of Biomedical Engineering, Oregon Health & Science University, Portland, OR, 97239, USA

    T. C. L. Kohs, C. U. Lorentz, J. Johnson, C. Puy, S. R. Olson, J. J. Shatzel, M. T. Hinds, E. I. Tucker, A. Gruber, O. J. T. McCarty & M. Wallisch

  2. Aronora Inc., Portland, OR, USA

    C. U. Lorentz, E. I. Tucker, A. Gruber & M. Wallisch

  3. Division of Hematology& Medical Oncology, Department of Medicine, Oregon Health & Science University, Portland, OR, USA

    S. R. Olson, J. J. Shatzel, A. Gruber & O. J. T. McCarty

  4. Department of Pathology, Microbiology, and Immunology, Vanderbilt University School of Medicine, Nashville, TN, USA

    D. Gailani

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  1. T. C. L. Kohs
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  2. C. U. Lorentz
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  3. J. Johnson
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  11. O. J. T. McCarty
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Contributions

MW, AG, EIT were responsible for the project concept and design. MW, JJ, and MTH executed and led the primate studies. CUL, CP, TK, and DG contributed in vitro data acquisition and analysis. MW performed data analysis and statistics for in vivo primate data. TK, MW, OJTM, and AG drafted the manuscript. CUL, JJS, SO, DG, and EIT critically reviewed and revised the manuscript.

Corresponding author

Correspondence to T. C. L. Kohs.

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Associate Editor Michael R. King oversaw the review of this article.

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Kohs, T.C.L., Lorentz, C.U., Johnson, J. et al. Development of Coagulation Factor XII Antibodies for Inhibiting Vascular Device-Related Thrombosis. Cel. Mol. Bioeng. 14, 161–175 (2021). https://doi.org/10.1007/s12195-020-00657-6

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  • DOI: https://doi.org/10.1007/s12195-020-00657-6

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