TY - JOUR
T1 - An engineered CD81-based combinatorial library for selecting recombinant binders to cell surface proteins
T2 - Laminin binding CD81 enhances cellular uptake of extracellular vesicles
AU - Vogt, Stefan
AU - Bobbili, Madhusudhan Reddy
AU - Stadlmayr, Gerhard
AU - Stadlbauer, Katharina
AU - Kjems, Jørgen
AU - Rüker, Florian
AU - Grillari, Johannes
AU - Wozniak-Knopp, Gordana
PY - 2021/9
Y1 - 2021/9
N2 - The research of extracellular vesicles (EVs) has boomed in the last decade, with the promise of them functioning as target-directed drug delivery vehicles, able to modulate proliferation, migration, differentiation, and other properties of the recipient cell that are vital for health of the host organism. To enhance the ability of their targeted delivery, we employed an intrinsically overrepresented protein, CD81, to serve for recognition of the desired target antigen. Yeast libraries displaying mutant variants of the large extracellular loop of CD81 have been selected for binders to human placental laminin as an example target. Their specific interaction with laminin was confirmed in a mammalian display system. Derived sequences were reformatted to full-length CD81 and expressed in EVs produced by HeLa cells. These EVs were examined for the presence of the recombinant protein and were shown to exhibit an enhanced uptake into laminin-secreting mammalian cell lines. For the best candidate, the specificity of antigen interaction was demonstrated with a competition experiment. To our knowledge, this is the first example of harnessing an EV membrane protein as mediator of de novo target antigen recognition via in vitro molecular evolution, opening horizons to a broad range of applications in various therapeutic settings.
AB - The research of extracellular vesicles (EVs) has boomed in the last decade, with the promise of them functioning as target-directed drug delivery vehicles, able to modulate proliferation, migration, differentiation, and other properties of the recipient cell that are vital for health of the host organism. To enhance the ability of their targeted delivery, we employed an intrinsically overrepresented protein, CD81, to serve for recognition of the desired target antigen. Yeast libraries displaying mutant variants of the large extracellular loop of CD81 have been selected for binders to human placental laminin as an example target. Their specific interaction with laminin was confirmed in a mammalian display system. Derived sequences were reformatted to full-length CD81 and expressed in EVs produced by HeLa cells. These EVs were examined for the presence of the recombinant protein and were shown to exhibit an enhanced uptake into laminin-secreting mammalian cell lines. For the best candidate, the specificity of antigen interaction was demonstrated with a competition experiment. To our knowledge, this is the first example of harnessing an EV membrane protein as mediator of de novo target antigen recognition via in vitro molecular evolution, opening horizons to a broad range of applications in various therapeutic settings.
KW - antigen-binding CD81
KW - directed evolution
KW - extracellular vesicle uptake
KW - targeted extracellular vesicles
KW - yeast display
UR - http://www.scopus.com/inward/record.url?scp=85115138745&partnerID=8YFLogxK
U2 - 10.1002/jev2.12139
DO - 10.1002/jev2.12139
M3 - Journal article
C2 - 34514736
AN - SCOPUS:85115138745
SN - 2001-3078
VL - 10
JO - Journal of Extracellular Vesicles
JF - Journal of Extracellular Vesicles
IS - 11
M1 - e12139
ER -