Abstract
Biologic tumor necrosis factor inhibitors (TNFIs) include TNF decoy receptors (TNFRs). TNFα plays a pathologic role in both acute and chronic brain disease. However, biologic TNFIs cannot be developed as brain therapeutics because these large molecule drugs do not cross the blood-brain barrier (BBB). To enable penetration of the brain via receptor-mediated transport, the human TNFR type II was re-engineered as an IgG fusion protein, where the IgG part is a chimeric monoclonal antibody (MAb) against the mouse transferrin receptor (TfR), and this fusion protein is designated cTfRMAb-TNFR. The cTfRMAb part of the fusion protein acts as a molecular Trojan horse to ferry the TNFR across the BBB via transport on the endogenous BBB TfR. cTfRMAb-TNFR was expressed by stably transfected Chinese hamster ovary cells and purified by affinity chromatography to homogeneity on electrophoretic gels. The fusion protein reacted with antibodies to both mouse IgG and the human TNFR and bound TNFα with high affinity (Kd = 96 ± 34 pM). cTfRMAb-TNFR was rapidly transported into mouse brain in vivo after intravenous administration, and the brain uptake of the fusion protein was 2.8 ± 0.5% of injected dose per gram of brain, which is >45-fold higher than the brain uptake of an IgG that does not recognize the mouse TfR. This new IgG-TNFR fusion protein can be tested in mouse models of brain diseases in which TNFα plays a pathologic role.
Footnotes
- Received August 19, 2010.
- Accepted September 30, 2010.
This work was supported by the National Institutes of Health National Institute on Aging [Grant R01-AG032244].
Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
doi:10.1124/dmd.110.036012.
ABBREVIATIONS:
- TNFα
- tumor necrosis factor alpha
- PD
- Parkinson's disease
- AD
- Alzheimer's disease
- TNFI
- TNF inhibitor
- TNFR-II ECD
- extracellular domain of the type II TNF receptor
- CNS
- central nervous system
- BBB
- blood-brain barrier
- MTH
- molecular Trojan horse
- MAb
- monoclonal antibody
- HIR
- human insulin receptor
- IV
- intravenous
- TfR
- transferrin receptor
- CHO
- Chinese hamster ovary
- TV
- tandem vector
- LC
- light chain
- AA
- amino acid
- HC
- heavy chain
- ELISA
- enzyme-linked immunosorbent assay
- PAGE
- polyacrylamide gel electrophoresis
- TBS
- Tris-buffered saline
- PBS
- phosphate-buffered saline
- TCA
- trichloroacetic acid
- IP
- intraperitoneal
- ID
- injection dose
- VD
- volume of distribution
- DPM
- disintegrations per minute
- AUC
- area under the plasma concentration curve
- PS
- permeability-surface area
- C
- constant
- BAA06141
- mouse immunoglobulin gamma-1 light chain
- BAC44885
- mouse immunoglobulin gamma-1 heavy chain.
- Copyright © 2011 by The American Society for Pharmacology and Experimental Therapeutics
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