Submitted April 4, 2007
Accepted January 10, 2008
Human antibody RNase fusion protein targeting CD30+ lymphomas
Christian Menzel, Thomas Schirrmann, Zoltan Konthur, Thomas Jostock, and Stefan Dubel*
Department of Biotechnology, Technical University of Braunschweig, Institute of Biochemistry and Biotechnology, Braunschweig, Germany
Department of Vertebrate Genomics, Max Planck Institute of Molecular Genetics, Berlin, Germany
* Corresponding author; email: s.duebel{at}tu-bs.de.
"Targeted RNases" (TR) are immunoenzymes with ribonucleases as cytotoxic effector domains which are less immunogenic as plant or bacterial toxin components of classical immunotoxins.
In this study, we show the generation and production of the first entirely human TR (huTR) directed against CD30+ lymphomas. The scFv-Fc-RNase construct was produced in HEK293T cells, yielding of up to 4 mg/L soluble protein after purification by proteinA affinity chromatography. Size exclusion chromatography revealed a homodimer of the predicted molecular mass. Surface plasmon resonance analysis revealed an affinity to CD30 of KD < 1 nM for both the scFv-Fc and the scFv-Fc-RNase protein. Internalization of the scFv-Fc-RNase protein by CD30+ Karpas-299 cell was demonstrated by confocal microscopy. Proliferation of CD30+ lymphoma cell line Karpas-299 was strongly inhibited by CD30 specific huTR protein (IC50 = 3.3 nM).
The huTR is a promising candidate for the immunotherapy of CD30+ lymphomas, because of its expected low immunogenicity, good production yields and potent effector function upon target cell binding and internalization. Its modular design is set to target other internalizing tumor antigens using different antibody domains.
