Cabanas - A Supramolecular Host-Guest Carrier System for Growth Factors Employing VHH Fragments

Jordi Cabanas-Danés,† Emilie Dooms Rodrigues, Ellie Landman, Jasper van Weerd,†, Clemens van Blitterswijk, Theo Verrips, Jurriaan Huskens, Marcel Karperien and Pascal Jonkheijm

JACS 2014


A supramolecular strategy is presented for the assembly of growth factors employing His6-tagged singledomain antibodies (VHH). A combination of orthogonal supramolecular interactions of β-cyclodextrin (βCD)−adamantyl
(Ad) host−guest and N-nitrilotriacetic acid (NTA)− histidine (His) interactions was employed to generate reversible and homogeneous layers of growth factors. A single-domain antibody VHH fragment was identified to bind to the human bone morphogenetic protein-6 (hBMP6) growth factor and could be recombinantly expressed in E. coli. The VHH fragment was equipped with a C-terminal hexahistidine (His6) tether to facilitate the assembly on βCD surfaces using a linker that contains an Ad group to bind to the βCD receptors and an NTA moiety to interact with the His6-tag upon cocomplexation of Ni2+ ions. After exploring the thermodynamic and kinetic stability of the VHH assemblies on βCD surfaces using a variety of experimental techniques including microcontact printing (μCP), surface plasmon resonance (SPR), microscale thermophoresis (MST), and theoretical models for determining the thermodynamic behavior of the system, hBMP6 was assembled onto the VHH-functionalized surfaces. After analyzing the immobilized hBMP6 using immunostaining, the biological activity of hBMP6 was demonstrated in cell differentiation experiments. Early osteogenic differentiation was analyzed in terms of alkaline phosphatase (ALP) activity of KS483-4C3 mouse progenitor cells, and the results indicated that the reversibly immobilized growth factors were functionally delivered to the cells. In conclusion, the supramolecular strategy used here offers the necessary affinity, reversibility, and temporal control to promote biological function of the growth factors that were delivered by this strategy.