Targeting NETs using dual-active DNase1 variants
BackgroundNeutrophil Extracellular Traps (NETs) are key mediators of immunothrombotic mechanisms and defective clearance of NETs from the circulation underlies an array of thrombotic, inflammatory, infectious, and autoimmune diseases. Efficient NET degradation depends on the combined activity of two...
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Frontiers Media S.A.
2023-05-01
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| Series: | Frontiers in Immunology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fimmu.2023.1181761/full |
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| author | Hanna Englert Josephine Göbel Danika Khong Maryam Omidi Nina Wolska Sandra Konrath Maike Frye Reiner K. Mailer Manu Beerens Julian C. Gerwers Roger J. S. Preston Jacob Odeberg Jacob Odeberg Lynn M. Butler Lynn M. Butler Lynn M. Butler Lynn M. Butler Coen Maas Evi X. Stavrou Evi X. Stavrou Tobias A. Fuchs Tobias A. Fuchs Thomas Renné Thomas Renné Thomas Renné |
| author_facet | Hanna Englert Josephine Göbel Danika Khong Maryam Omidi Nina Wolska Sandra Konrath Maike Frye Reiner K. Mailer Manu Beerens Julian C. Gerwers Roger J. S. Preston Jacob Odeberg Jacob Odeberg Lynn M. Butler Lynn M. Butler Lynn M. Butler Lynn M. Butler Coen Maas Evi X. Stavrou Evi X. Stavrou Tobias A. Fuchs Tobias A. Fuchs Thomas Renné Thomas Renné Thomas Renné |
| author_sort | Hanna Englert |
| collection | DOAJ |
| description | BackgroundNeutrophil Extracellular Traps (NETs) are key mediators of immunothrombotic mechanisms and defective clearance of NETs from the circulation underlies an array of thrombotic, inflammatory, infectious, and autoimmune diseases. Efficient NET degradation depends on the combined activity of two distinct DNases, DNase1 and DNase1-like 3 (DNase1L3) that preferentially digest double-stranded DNA (dsDNA) and chromatin, respectively.MethodsHere, we engineered a dual-active DNase with combined DNase1 and DNase1L3 activities and characterized the enzyme for its NET degrading potential in vitro. Furthermore, we produced a mouse model with transgenic expression of the dual-active DNase and analyzed body fluids of these animals for DNase1 and DNase 1L3 activities. We systematically substituted 20 amino acid stretches in DNase1 that were not conserved among DNase1 and DNase1L3 with homologous DNase1L3 sequences.ResultsWe found that the ability of DNase1L3 to degrade chromatin is embedded into three discrete areas of the enzyme's core body, not the C-terminal domain as suggested by the state-of-the-art. Further, combined transfer of the aforementioned areas of DNase1L3 to DNase1 generated a dual-active DNase1 enzyme with additional chromatin degrading activity. The dual-active DNase1 mutant was superior to native DNase1 and DNase1L3 in degrading dsDNA and chromatin, respectively. Transgenic expression of the dual-active DNase1 mutant in hepatocytes of mice lacking endogenous DNases revealed that the engineered enzyme was stable in the circulation, released into serum and filtered to the bile but not into the urine.ConclusionTherefore, the dual-active DNase1 mutant is a promising tool for neutralization of DNA and NETs with potential therapeutic applications for interference with thromboinflammatory disease states. |
| first_indexed | 2024-03-13T10:01:27Z |
| format | Article |
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| institution | Directory Open Access Journal |
| issn | 1664-3224 |
| language | English |
| last_indexed | 2024-03-13T10:01:27Z |
| publishDate | 2023-05-01 |
| publisher | Frontiers Media S.A. |
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| series | Frontiers in Immunology |
| spelling | doaj.art-9aed362b20df48998e4383fc4e6df5562023-05-23T05:07:27ZengFrontiers Media S.A.Frontiers in Immunology1664-32242023-05-011410.3389/fimmu.2023.11817611181761Targeting NETs using dual-active DNase1 variantsHanna Englert0Josephine Göbel1Danika Khong2Maryam Omidi3Nina Wolska4Sandra Konrath5Maike Frye6Reiner K. Mailer7Manu Beerens8Julian C. Gerwers9Roger J. S. Preston10Jacob Odeberg11Jacob Odeberg12Lynn M. Butler13Lynn M. Butler14Lynn M. Butler15Lynn M. Butler16Coen Maas17Evi X. Stavrou18Evi X. Stavrou19Tobias A. Fuchs20Tobias A. Fuchs21Thomas Renné22Thomas Renné23Thomas Renné24Institute of Clinical Chemistry and Laboratory Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, GermanyInstitute of Clinical Chemistry and Laboratory Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, GermanyInstitute of Clinical Chemistry and Laboratory Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, GermanyInstitute of Clinical Chemistry and Laboratory Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, GermanyInstitute of Clinical Chemistry and Laboratory Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, GermanyInstitute of Clinical Chemistry and Laboratory Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, GermanyInstitute of Clinical Chemistry and Laboratory Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, GermanyInstitute of Clinical Chemistry and Laboratory Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, GermanyInstitute of Clinical Chemistry and Laboratory Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, GermanyInstitute of Clinical Chemistry and Laboratory Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, GermanyIrish Centre for Vascular Biology, School of Pharmacy and Biomolecular Sciences, Royal College of Surgeons in Ireland, Dublin, IrelandDepartment of Clinical Medicine, The Arctic University of Norway, Tromsø, NorwayScience for Life Laboratory, Department of Protein Science, CBH, KTH Royal Institute of Technology, Stockholm, SwedenInstitute of Clinical Chemistry and Laboratory Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, GermanyDepartment of Clinical Medicine, The Arctic University of Norway, Tromsø, NorwayScience for Life Laboratory, Department of Protein Science, CBH, KTH Royal Institute of Technology, Stockholm, SwedenDepartment of Molecular Medicine and Surgery, Karolinska Institute, Stockholm, SwedenDepartment of Clinical Chemistry and Haematology, University Medical Center Utrecht, Utrecht University, Utrecht, NetherlandsMedicine Service, Section of Hematology-Oncology, Louis Stokes Veterans Administration Medical Center, Cleveland, OH, United StatesDepartment of Medicine, Hematology and Oncology Division, Case Western Reserve University School of Medicine, Cleveland, OH, United StatesInstitute of Clinical Chemistry and Laboratory Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, GermanyNeutrolis, Inc., Cambridge, MA, United StatesInstitute of Clinical Chemistry and Laboratory Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, GermanyIrish Centre for Vascular Biology, School of Pharmacy and Biomolecular Sciences, Royal College of Surgeons in Ireland, Dublin, Ireland0Center for Thrombosis and Hemostasis (CTH), Johannes Gutenberg University Medical Center, Mainz, GermanyBackgroundNeutrophil Extracellular Traps (NETs) are key mediators of immunothrombotic mechanisms and defective clearance of NETs from the circulation underlies an array of thrombotic, inflammatory, infectious, and autoimmune diseases. Efficient NET degradation depends on the combined activity of two distinct DNases, DNase1 and DNase1-like 3 (DNase1L3) that preferentially digest double-stranded DNA (dsDNA) and chromatin, respectively.MethodsHere, we engineered a dual-active DNase with combined DNase1 and DNase1L3 activities and characterized the enzyme for its NET degrading potential in vitro. Furthermore, we produced a mouse model with transgenic expression of the dual-active DNase and analyzed body fluids of these animals for DNase1 and DNase 1L3 activities. We systematically substituted 20 amino acid stretches in DNase1 that were not conserved among DNase1 and DNase1L3 with homologous DNase1L3 sequences.ResultsWe found that the ability of DNase1L3 to degrade chromatin is embedded into three discrete areas of the enzyme's core body, not the C-terminal domain as suggested by the state-of-the-art. Further, combined transfer of the aforementioned areas of DNase1L3 to DNase1 generated a dual-active DNase1 enzyme with additional chromatin degrading activity. The dual-active DNase1 mutant was superior to native DNase1 and DNase1L3 in degrading dsDNA and chromatin, respectively. Transgenic expression of the dual-active DNase1 mutant in hepatocytes of mice lacking endogenous DNases revealed that the engineered enzyme was stable in the circulation, released into serum and filtered to the bile but not into the urine.ConclusionTherefore, the dual-active DNase1 mutant is a promising tool for neutralization of DNA and NETs with potential therapeutic applications for interference with thromboinflammatory disease states.https://www.frontiersin.org/articles/10.3389/fimmu.2023.1181761/fullneutrophil extracellular traps (NETs)NETosisNET degradationDNase1DNase1-like 3thromboinflammation |
| spellingShingle | Hanna Englert Josephine Göbel Danika Khong Maryam Omidi Nina Wolska Sandra Konrath Maike Frye Reiner K. Mailer Manu Beerens Julian C. Gerwers Roger J. S. Preston Jacob Odeberg Jacob Odeberg Lynn M. Butler Lynn M. Butler Lynn M. Butler Lynn M. Butler Coen Maas Evi X. Stavrou Evi X. Stavrou Tobias A. Fuchs Tobias A. Fuchs Thomas Renné Thomas Renné Thomas Renné Targeting NETs using dual-active DNase1 variants Frontiers in Immunology neutrophil extracellular traps (NETs) NETosis NET degradation DNase1 DNase1-like 3 thromboinflammation |
| title | Targeting NETs using dual-active DNase1 variants |
| title_full | Targeting NETs using dual-active DNase1 variants |
| title_fullStr | Targeting NETs using dual-active DNase1 variants |
| title_full_unstemmed | Targeting NETs using dual-active DNase1 variants |
| title_short | Targeting NETs using dual-active DNase1 variants |
| title_sort | targeting nets using dual active dnase1 variants |
| topic | neutrophil extracellular traps (NETs) NETosis NET degradation DNase1 DNase1-like 3 thromboinflammation |
| url | https://www.frontiersin.org/articles/10.3389/fimmu.2023.1181761/full |
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