In vivo binding and retention of CD4-specific DARPin 57.2 in macaques.
BACKGROUND
The recently described Designed Ankyrin Repeat Protein (DARPin) technology can produce highly selective ligands to a variety of biological targets at a low production cost.


RESULTS
To investigate the in vivo use of DARPins for future application to novel anti-HIV strategies, we identified potent CD4-specific DARPins that recognize rhesus CD4 and followed the fate of intravenously injected CD4-specific DARPin 57.2 in rhesus macaques. The human CD4-specific DARPin 57.2 bound macaque CD4(+) cells and exhibited potent inhibitory activity against SIV infection in vitro. DARPin 57.2 or the control E3_5 DARPin was injected into rhesus macaques and the fate of cell-free and cell-bound CD4-specific DARPin was evaluated. DARPin-bound CD4(+) cells were detected in the peripheral blood as early as 30 minutes after the injection, decreasing within 6 hours and being almost undetectable within 24 hours. The amount of DARPin bound was dependent on the amount of DARPin injected. CD4-specific DARPin was also detected on CD4(+) cells in the lymph nodes within 30 minutes, which persisted with similar kinetics to blood. More extensive analysis using blood revealed that DARPin 57.2 bound to all CD4(+) cell types (T cells, monocytes, dendritic cells) in vivo and in vitro with the amount of binding directly proportional to the amount of CD4 on the cell surface. Cell-free DARPins were also detected in the plasma, but were rapidly cleared from circulation.


CONCLUSIONS
We demonstrated that the CD4-specific DARPin can rapidly and selectively bind its target cells in vivo, warranting further studies on possible clinical use of the DARPin technology.
Pugach
Pavel
P
Center for Biomedical Research, Population Council, New York, New York, United States of America.
Krarup
Anders
A
Gettie
Agegnehu
A
Kuroda
Marcelo
M
Blanchard
James
J
Piatak
Michael
M
Lifson
Jeffrey D
JD
Trkola
Alexandra
A
Robbiani
Melissa
M
D000818:Animals; D017089:Ankyrin Repeat; D015704:CD4 Antigens / Q000378:metabolism*; D005260:Female; D006678:HIV / Q000187:drug effects; D006801:Humans; D007275:Injections, Intravenous; D007700:Kinetics; D008253:Macaca mulatta; D011485:Protein Binding; D015202:Protein Engineering / Q000379:methods*; D011506:Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism* / Q000493:pharmacokinetics; D013379:Substrate Specificity
D015704:CD4 Antigens; D011506:Proteins
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