Long-lasting fibrin matrices ensure stable and functional angiogenesis by highly tunable, sustained delivery of recombinant VEGF164.
Clinical trials of therapeutic angiogenesis by vascular endothelial growth factor (VEGF) gene delivery failed to show efficacy. Major challenges include the need to precisely control in vivo distribution of growth factor dose and duration of expression. Recombinant VEGF protein delivery could overcome these issues, but rapid in vivo clearance prevents the stabilization of induced angiogenesis. Here, we developed an optimized fibrin platform for controlled delivery of recombinant VEGF, to robustly induce normal, stable, and functional angiogenesis. Murine VEGF164 was fused to a sequence derived from α2-plasmin inhibitor (α2-PI1-8) that is a substrate for the coagulation factor fXIIIa, to allow its covalent cross-linking into fibrin hydrogels and release only by enzymatic cleavage. An α2-PI1-8-fused variant of the fibrinolysis inhibitor aprotinin was used to control the hydrogel degradation rate, which determines both the duration and effective dose of factor release. An optimized aprotinin-α2-PI1-8 concentration ensured ideal degradation over 4 wk. Under these conditions, fibrin-α2-PI1-8-VEGF164 allowed exquisitely dose-dependent angiogenesis: concentrations ≥25 μg/mL caused widespread aberrant vascular structures, but a 500-fold concentration range (0.01-5.0 μg/mL) induced exclusively normal, mature, nonleaky, and perfused capillaries, which were stable after 3 mo. Optimized delivery of fibrin-α2-PI1-8-VEGF164 was therapeutically effective both in ischemic hind limb and wound-healing models, significantly improving angiogenesis, tissue perfusion, and healing rate. In conclusion, this optimized platform ensured (i) controlled and highly tunable delivery of VEGF protein in ischemic tissue and (ii) stable and functional angiogenesis without introducing genetic material and with a limited and controllable duration of treatment. These findings suggest a strategy to improve safety and efficacy of therapeutic angiogenesis.
Cell and Gene Therapy, Department of Biomedicine, University of Basel, and Department of Surgery, Basel University Hospital, CH-4031 Basel, Switzerland;
Veronica
V
Sacchi
Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, Austrian Cluster for Tissue Regeneration, Allgemeine Unfallversicherungsanstalt, A-1200 Vienna, Austria;
Rainer
R
Mittermayr
Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, Austrian Cluster for Tissue Regeneration, Allgemeine Unfallversicherungsanstalt, A-1200 Vienna, Austria;
Joachim
J
Hartinger
Institute of Bioengineering, School of Life Sciences and School of Engineering, Ecole Polytechnique Fédérale de Lausanne, CH-1015 Lausanne, Switzerland;
Mikaël M
MM
Martino
Institute of Bioengineering, School of Life Sciences and School of Engineering, Ecole Polytechnique Fédérale de Lausanne, CH-1015 Lausanne, Switzerland;
Kristen M
KM
Lorentz
Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, Austrian Cluster for Tissue Regeneration, Allgemeine Unfallversicherungsanstalt, A-1200 Vienna, Austria;
Susanne
S
Wolbank
Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, Austrian Cluster for Tissue Regeneration, Allgemeine Unfallversicherungsanstalt, A-1200 Vienna, Austria;
Anna
A
Hofmann
Departments of Urology and.
Remo A
RA
Largo
Obstetrics, Zurich University Hospital, CH-8091 Zurich, Switzerland; andUniversity of Louisville School of Dentistry, Louisville, KY 40202.
Jeffrey S
JS
Marschall
Cell and Gene Therapy, Department of Biomedicine, University of Basel, and Department of Surgery, Basel University Hospital, CH-4031 Basel, Switzerland;
Elena
E
Groppa
Cell and Gene Therapy, Department of Biomedicine, University of Basel, and Department of Surgery, Basel University Hospital, CH-4031 Basel, Switzerland;
Roberto
R
Gianni-Barrera
Obstetrics, Zurich University Hospital, CH-8091 Zurich, Switzerland; and.
Martin
M
Ehrbar
Institute of Bioengineering, School of Life Sciences and School of Engineering, Ecole Polytechnique Fédérale de Lausanne, CH-1015 Lausanne, Switzerland;
Jeffrey A
JA
Hubbell
Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, Austrian Cluster for Tissue Regeneration, Allgemeine Unfallversicherungsanstalt, A-1200 Vienna, Austria;
Heinz
H
Redl
Cell and Gene Therapy, Department of Biomedicine, University of Basel, and Department of Surgery, Basel University Hospital, CH-4031 Basel, Switzerland; Andrea.Banfi@usb.ch.
Andrea
A
Banfi
D000818:Animals; D005260:Female; D005337:Fibrin / Q000493:pharmacokinetics*; D005782:Gels / Q000493:pharmacokinetics; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods; D006614:Hindlimb; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D007511:Ischemia / Q000628:therapy*; D051379:Mice; D008815:Mice, Inbred Strains; D016513:Mice, SCID; D018482:Muscle, Skeletal / Q000098:blood supply; D018919:Neovascularization, Physiologic / Q000502:physiology*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism / Q000493:pharmacokinetics; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics* / Q000378:metabolism / Q000493:pharmacokinetics*
D005782:Gels; D011994:Recombinant Proteins; D042461:Vascular Endothelial Growth Factor A; C467480:vascular endothelial growth factor A, mouse; D005337:Fibrin