Antitransferrin receptor immunotoxin inhibits proliferating human retinal pigment epithelial cells
G. J. Jaffe, K. Earnest, S. Fulcher, G. M. Lui and L. L. Houston
Department of Ophthalmology, Duke University Eye Center, Durham, NC 27710.
Cultured human retinal pigment epithelial cells were exposed to an
immunotoxin composed of a monoclonal antibody, 454A12, directed against
transferrin receptors conjugated to a toxin, recombinant ricin A chain.
Exposure of proliferating human retinal pigment epithelial cells to the
immunotoxin (0.1 to 10,000 ng/mL) caused a statistically significant (P
less than .0001) decrease in the number of cells. This inhibitory effect
was induced after an exposure to the immunotoxin as short as 5 minutes and
was maximal after 24 hours of exposure. The diminution in cell number was
dose dependent over the range from 0.1 to 100 ng/mL. Monoclonal antibody
alone, recombinant ricin A chain alone, or an irrelevant immunotoxin,
MOP21C monoclonal antibody-recombinant ricin A, did not diminish the number
of cells. There was a marked decrease in DNA synthesis measured by nuclear
tritiated thymidine incorporation that accompanied the immunotoxin-mediated
decrease in cell number. Viable cells remaining after exposure to the
immunotoxin (0.1 to 10,000 ng/mL) were morphologically abnormal; typically
the cells had elongated spindle-shaped processes and had lost their normal
cuboidal appearance. In contrast, cell number was not decreased in
confluent human retinal pigment epithelial cells after treatment with
maximal doses of immunotoxin. Morphologic changes similar to those seen in
proliferating cells were observed in confluent cells exposed to more than
100 ng/mL of immunotoxin. The effect of the immunotoxin was species
specific because large doses of immunotoxin did not reduce the number of
viable cells in proliferating or confluent pig retinal pigment epithelial
cells or cause observable morphologic changes in this cell type. Our
results indicate that the immunotoxin selectively inhibited proliferating
retinal pigment epithelial cells by receptor-mediated internalization of
the antitransferrin receptor monoclonal antibody-recombinant ricin A chain
conjugate.
