Endothelial cells internalize monoclonal antibody to angiotensin
-converting enzyme.
Muzykantov, Vladimir R., Elena N. Atochina, Alice Kuo#, Elliott S.
Barnathan, Kathy Notarfrancesco, Henry Shuman, Chandra Dodia, and
Aron B. Fisher.
Institute for Environmental Medicine and Department of Medicine,
School of Medicine, University of Pennsylvania
APStracts 2:0225L, 1995.
We investigated the fate of Mab 9B9, a monoclonal antibody to
angiotensin-converting enzyme (ACE), which binds to endothelium both
in vitro and in vivo. Using cultured human umbilical vein endothelial
cells (HUVEC) and isolated perfused rat lungs (IPL), we demonstrated
specific and saturable binding of 125I-Mab 9B9 at 4oC (Kd = 20-50 nM,
Bmax = (1.5-3.0)x105 sites/cell). When 125I-Mab 9B9 was bound to
HUVEC at 37oC, only 40% of cell-associated radioactivity was acid
elutable, suggesting antibody internalization. This was confirmed by
finding that: 1) the amount of Mab 9B9 uptake at 37oC was higher than
at 4oC both in HUVEC and IPL; 2) binding of 125I-streptavidin with
HUVEC and IPL pre-treated with biotinylated Mab 9B9 (b-Mab 9B9) was
diminished in a temperature- and time-dependent fashion at 37oC; and,
3) b-Mab 9B9 bound to HUVEC at 37oC was found intracellularly by
ultrastructural analysis using streptavidin-gold. Intracellular 125I
-Mab 9B9 was found in microsomal fractions of lung homogenate from IPL
and after iv injections in rats. Degradation of internalized Mab 9B9
was minimal, since >90% of cell-associated 125Iodine label
remained precipitable by trichloracetic acid in HUVEC, IPL and in
vivo. Radioautography of SDS-PAGE of lung homogenates made as long as
several days after iv injections of 125I-Mab 9B9 in rats,
demonstrated a predominant band above 140 kD. These data indicate
that endothelial cells either in vitro or in vivo internalize ACE
ligand, Mab 9B9, without significant intra cellular degradation.
Therefore, Mab 9B9 may be useful for selective intracellular delivery
of drugs to the pulmonary vascular endothelium after systemic
administration.
Received 4 August 1995; accepted in final form 30 November 1995.
APS Manuscript Number L244-5.
Article publication pending Am. J. Physiol. (Lung Cell. Mol.
Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 23 December 95