The effect of a monoclonal antibody coupled to ricin A chain-derived peptides on endothelial cells in vitro: insights into toxin-mediated vascular damage.
Immunotoxins (ITs) containing plant or bacterial toxins have a dose-limiting toxicity of vascular leak syndrome (VLS) in humans. The active A chain of ricin toxin (RTA), other toxins, ribosome-inactivating proteins, and the VLS-inducing cytokine IL-2 contain the conserved sequence motif (x)D(y) where x = L, I, G, or V and y = V, L, or S. RTA-derived LDV-containing peptides attached to a monoclonal antibody, RFB4, induce endothelial cell (EC) damage in vitro and vascular leak in two animal models in vivo. We have now investigated the mechanism(s) by which this occurs and have found that (1) the exposed D75 in the LDV sequence in RTA and the C-terminal flanking threonine play critical roles in the ability of RFB4-conjugated RTA peptide to bind to and damage ECs and (2) the LDV sequence in RTA induces early manifestations of apoptosis in HUVECs by activating caspase-3. These data suggest that RTA-mediated inhibition of protein synthesis (due to its active site) and apoptosis (due to LDV) may be mediated by different portions of the RTA molecule. These results suggest that ITs prepared with RTA mutants containing alterations in LDVT may kill tumor cells in vivo in the absence of EC-mediated VLS.
Baluna
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R
The Cancer Immunology Center, The Immunology Graduate Program, The Cancer Education Summer Program, 5323 Harry Hines Boulevard, Dallas, Texas 75235-8756, USA.
Coleman
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Jones
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Ghetie
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Vitetta
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ES
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology*; D015703:Antigens, CD / Q000276:immunology*; D000944:Antigens, Differentiation, B-Lymphocyte / Q000276:immunology*; D017209:Apoptosis; D001224:Aspartic Acid / Q000378:metabolism; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D015815:Cell Adhesion Molecules; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects*; D004789:Enzyme Activation; D006801:Humans; D000922:Immunotoxins / Q000494:pharmacology*; D037102:Lectins; D051379:Mice; D008969:Molecular Sequence Data; D010455:Peptides / Q000378:metabolism*; D012276:Ricin / Q000494:pharmacology*; D051918:Sialic Acid Binding Ig-like Lectin 2; D013912:Threonine; D014471:Umbilical Veins / Q000166:cytology
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D000944:Antigens, Differentiation, B-Lymphocyte; C497867:CD22 protein, human; C497868:Cd22 protein, mouse; D015815:Cell Adhesion Molecules; D000922:Immunotoxins; D037102:Lectins; D010455:Peptides; D051918:Sialic Acid Binding Ig-like Lectin 2; D013912:Threonine; D001224:Aspartic Acid; D012276:Ricin; C505407:CASP3 protein, human; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases