Quantitative characterisation of extracellular vesicles designed to decoy or compete with SARS-CoV-2 reveals differential mode of action across variants of concern and highlights the diversity of Omicron
Abstract Background The converging biology between enveloped viruses and extracellular vesicles (EVs) has raised interest in the application of engineered EVs as antiviral therapeutics. Following the recent COVID-19 pandemic, EVs engineered with either the ACE2-receptor or Spike-protein have been pr...
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BMC
2025-07-01
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| Series: | Cell Communication and Signaling |
| Online Access: | https://doi.org/10.1186/s12964-025-02223-x |
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| author | Melanie Schürz Isabel Pagani Eva Klinglmayr Heloisa Melo Benirschke Martin Mayora Neto Luis J. V. Galietta Arianna Venturini Nicoletta Pedemonte Valeria Capurro Sandra Laner-Plamberger Christoph Grabmer Essi Emminger Martin Wolf Marianne Steiner Cyrus Kohlmetz Niklas Mayr Liliia Paniushkina Katharina Schallmoser Dirk Strunk Hans Brandstetter Martin Hintersteiner Nigel Temperton Elisa Vicenzi Nicole Meisner-Kober |
| author_facet | Melanie Schürz Isabel Pagani Eva Klinglmayr Heloisa Melo Benirschke Martin Mayora Neto Luis J. V. Galietta Arianna Venturini Nicoletta Pedemonte Valeria Capurro Sandra Laner-Plamberger Christoph Grabmer Essi Emminger Martin Wolf Marianne Steiner Cyrus Kohlmetz Niklas Mayr Liliia Paniushkina Katharina Schallmoser Dirk Strunk Hans Brandstetter Martin Hintersteiner Nigel Temperton Elisa Vicenzi Nicole Meisner-Kober |
| author_sort | Melanie Schürz |
| collection | DOAJ |
| description | Abstract Background The converging biology between enveloped viruses and extracellular vesicles (EVs) has raised interest in the application of engineered EVs as antiviral therapeutics. Following the recent COVID-19 pandemic, EVs engineered with either the ACE2-receptor or Spike-protein have been proposed as strategy to either decoy SARS-CoV-2, or to compete with its cell entry. For generic use as a platform for future pandemic preparedness, a systematic and quantitative comparison of both strategies is required to assess their limitations and benefits across different variants of concern. Methods Here we generated EVs decorated with either the ACE2-receptor or the Spike-protein of (Wuhan)-SARS-CoV-2 and used single vesicle imaging for in-depth quantitative characterisation. These vesicles were then systematically tested for anti-viral activity across SARS-CoV-2 variants of concern using both, pseudotype and live virus cellular infection models including primary human bronchial and nasal explants. Results Spike-protein EVs or ACE2-EVs recovered from transiently transfected HEK293T cells comprised only a small fraction of the EV secretome (5% or 20%, respectively) and were primarily derived from the plasma membrane rather than multivesicular bodies. Redirecting intracellular trafficking of the Spike protein by mutating its transmembrane or subcellular localisation domains did not increase the yields of Spike-EVs. Both types of vesicles inhibited SARS-CoV-2 (D614G) in a dose dependent manner with kinetics and immunohistochemistry consistent with an inhibition at the initial cell entry stage. ACE2-EVs were more potent than Spike-EVs and at least 500–1000 times more potent than soluble antibodies in a pseudotype model. Surprisingly, ACE2-EVs switched from an inhibitory to an enhancer activity for the Omicron BA.1 variant whereas Spike-EVs retained their activity across all variants of concern. Conclusions While our data show that both types of engineered EVs potently inhibit SARS-CoV, the decoy versus competition strategy may result in diverging outcomes when considering viral evolution into new variants of concern. While Spike-EVs retain their competition for receptor binding even against higher affinity viral Spike mutations, the formation of complexes between ACE2-EVs and the virus may not only result in inhibition by decoy. As EVs are actively internalised by cells themselves, they may shuttle the virus into cells, resulting in a productive alternative cell entry route for variants such as Omicron, that diverge from strict plasma membrane protease cleavage to the use of endosomal proteases for release of their genome. |
| format | Article |
| id | doaj-art-322ae71a051a4c7983dab96c8b742e6f |
| institution | Kabale University |
| issn | 1478-811X |
| language | English |
| publishDate | 2025-07-01 |
| publisher | BMC |
| record_format | Article |
| series | Cell Communication and Signaling |
| spelling | doaj-art-322ae71a051a4c7983dab96c8b742e6f2025-08-20T04:03:07ZengBMCCell Communication and Signaling1478-811X2025-07-0123113310.1186/s12964-025-02223-xQuantitative characterisation of extracellular vesicles designed to decoy or compete with SARS-CoV-2 reveals differential mode of action across variants of concern and highlights the diversity of OmicronMelanie Schürz0Isabel Pagani1Eva Klinglmayr2Heloisa Melo Benirschke3Martin Mayora Neto4Luis J. V. Galietta5Arianna Venturini6Nicoletta Pedemonte7Valeria Capurro8Sandra Laner-Plamberger9Christoph Grabmer10Essi Emminger11Martin Wolf12Marianne Steiner13Cyrus Kohlmetz14Niklas Mayr15Liliia Paniushkina16Katharina Schallmoser17Dirk Strunk18Hans Brandstetter19Martin Hintersteiner20Nigel Temperton21Elisa Vicenzi22Nicole Meisner-Kober23Chemical Biology and Biological Therapeutics, Department of Biosciences and Medical Biology, Paris Lodron University SalzburgViral Pathogenesis and Biosafety Unit, Division of Immunology, Transplantation, and Infectious Diseases, IRCCS San Raffaele Scientific InstituteChemical Biology and Biological Therapeutics, Department of Biosciences and Medical Biology, Paris Lodron University SalzburgChemical Biology and Biological Therapeutics, Department of Biosciences and Medical Biology, Paris Lodron University SalzburgViral Pseudotype Unit, Medway School of Pharmacy, University of KentDepartment of Translational Medical Sciences, Telethon Institute of Genetics and Medicine, University of Napoli“Federico II,”NaplesDepartment of Translational Medical Sciences, Telethon Institute of Genetics and Medicine, University of Napoli“Federico II,”NaplesUOC Genetica Medica, IRCCS Istituto Giannina GasliniUOC Genetica Medica, IRCCS Istituto Giannina GasliniDepartment for Transfusion Medicine, University Hospital of SalzburgDepartment for Transfusion Medicine, University Hospital of SalzburgCell Therapy Institute, Paracelsus Medical UniversityCell Therapy Institute, Paracelsus Medical UniversityChemical Biology and Biological Therapeutics, Department of Biosciences and Medical Biology, Paris Lodron University SalzburgChemical Biology and Biological Therapeutics, Department of Biosciences and Medical Biology, Paris Lodron University SalzburgChemical Biology and Biological Therapeutics, Department of Biosciences and Medical Biology, Paris Lodron University SalzburgCell Therapy Institute, Paracelsus Medical UniversityDepartment for Transfusion Medicine, University Hospital of SalzburgCell Therapy Institute, Paracelsus Medical UniversityChemical Biology and Biological Therapeutics, Department of Biosciences and Medical Biology, Paris Lodron University SalzburgEvoBiotiX SAViral Pseudotype Unit, Medway School of Pharmacy, University of KentViral Pathogenesis and Biosafety Unit, Division of Immunology, Transplantation, and Infectious Diseases, IRCCS San Raffaele Scientific InstituteChemical Biology and Biological Therapeutics, Department of Biosciences and Medical Biology, Paris Lodron University SalzburgAbstract Background The converging biology between enveloped viruses and extracellular vesicles (EVs) has raised interest in the application of engineered EVs as antiviral therapeutics. Following the recent COVID-19 pandemic, EVs engineered with either the ACE2-receptor or Spike-protein have been proposed as strategy to either decoy SARS-CoV-2, or to compete with its cell entry. For generic use as a platform for future pandemic preparedness, a systematic and quantitative comparison of both strategies is required to assess their limitations and benefits across different variants of concern. Methods Here we generated EVs decorated with either the ACE2-receptor or the Spike-protein of (Wuhan)-SARS-CoV-2 and used single vesicle imaging for in-depth quantitative characterisation. These vesicles were then systematically tested for anti-viral activity across SARS-CoV-2 variants of concern using both, pseudotype and live virus cellular infection models including primary human bronchial and nasal explants. Results Spike-protein EVs or ACE2-EVs recovered from transiently transfected HEK293T cells comprised only a small fraction of the EV secretome (5% or 20%, respectively) and were primarily derived from the plasma membrane rather than multivesicular bodies. Redirecting intracellular trafficking of the Spike protein by mutating its transmembrane or subcellular localisation domains did not increase the yields of Spike-EVs. Both types of vesicles inhibited SARS-CoV-2 (D614G) in a dose dependent manner with kinetics and immunohistochemistry consistent with an inhibition at the initial cell entry stage. ACE2-EVs were more potent than Spike-EVs and at least 500–1000 times more potent than soluble antibodies in a pseudotype model. Surprisingly, ACE2-EVs switched from an inhibitory to an enhancer activity for the Omicron BA.1 variant whereas Spike-EVs retained their activity across all variants of concern. Conclusions While our data show that both types of engineered EVs potently inhibit SARS-CoV, the decoy versus competition strategy may result in diverging outcomes when considering viral evolution into new variants of concern. While Spike-EVs retain their competition for receptor binding even against higher affinity viral Spike mutations, the formation of complexes between ACE2-EVs and the virus may not only result in inhibition by decoy. As EVs are actively internalised by cells themselves, they may shuttle the virus into cells, resulting in a productive alternative cell entry route for variants such as Omicron, that diverge from strict plasma membrane protease cleavage to the use of endosomal proteases for release of their genome.https://doi.org/10.1186/s12964-025-02223-x |
| spellingShingle | Melanie Schürz Isabel Pagani Eva Klinglmayr Heloisa Melo Benirschke Martin Mayora Neto Luis J. V. Galietta Arianna Venturini Nicoletta Pedemonte Valeria Capurro Sandra Laner-Plamberger Christoph Grabmer Essi Emminger Martin Wolf Marianne Steiner Cyrus Kohlmetz Niklas Mayr Liliia Paniushkina Katharina Schallmoser Dirk Strunk Hans Brandstetter Martin Hintersteiner Nigel Temperton Elisa Vicenzi Nicole Meisner-Kober Quantitative characterisation of extracellular vesicles designed to decoy or compete with SARS-CoV-2 reveals differential mode of action across variants of concern and highlights the diversity of Omicron Cell Communication and Signaling |
| title | Quantitative characterisation of extracellular vesicles designed to decoy or compete with SARS-CoV-2 reveals differential mode of action across variants of concern and highlights the diversity of Omicron |
| title_full | Quantitative characterisation of extracellular vesicles designed to decoy or compete with SARS-CoV-2 reveals differential mode of action across variants of concern and highlights the diversity of Omicron |
| title_fullStr | Quantitative characterisation of extracellular vesicles designed to decoy or compete with SARS-CoV-2 reveals differential mode of action across variants of concern and highlights the diversity of Omicron |
| title_full_unstemmed | Quantitative characterisation of extracellular vesicles designed to decoy or compete with SARS-CoV-2 reveals differential mode of action across variants of concern and highlights the diversity of Omicron |
| title_short | Quantitative characterisation of extracellular vesicles designed to decoy or compete with SARS-CoV-2 reveals differential mode of action across variants of concern and highlights the diversity of Omicron |
| title_sort | quantitative characterisation of extracellular vesicles designed to decoy or compete with sars cov 2 reveals differential mode of action across variants of concern and highlights the diversity of omicron |
| url | https://doi.org/10.1186/s12964-025-02223-x |
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