Generation of ammodytoxin-anti-cathepsin B immuno-conjugate as a model for delivery of secretory phospholipase A2 into cancer cells.
Ammodytoxin C (AtxC) is a toxic secreted phospholipase A2 (sPLA2) from the venom of Vipera ammodytes snake. To evaluate its potential to kill cancer cells, the toxin was cross-linked to the monoclonal antibody against cathepsin B which endocytoses upon binding to cathepsin B, an antigen overexpressed on the plasma membrane of cancer cells. A photo-reactive derivative of AtxC, possessing the same biological activity as the native toxin, was reacted with antibodies to form a covalent immuno-conjugate. In conditions of the cytosolic redox potential, AtxC was gradually released from the conjugate due to reduction of the disulfide bond in the spacer arm of the cross-linker. The phospholipase activity of the preparation reached maximum in 10min and then decreased gradually. As demonstrated by fluorescence microscopy, the immuno-conjugate targeted Caco-2 colon adenocarcinoma cells but was very slowly internalized, the likely reason of only slight cytotoxicity being observed. Despite the lack of a clear cytotoxic effect of AtxC-antibody conjugate on Caco-2 cells, we demonstrated in this work a new methodology for the targeted delivery of (toxic) sPLA2 into cells, promising in research or therapy.
Premzl
Ales
A
Department of Biochemistry, Molecular and Structural Biology, Jozef Stefan Institute, Jamova Street 39, SI-1000 Ljubljana, Slovenia.
Kovacic
Lidija
L
Halassy
Beata
B
Krizaj
Igor
I
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000378:metabolism; D000970:Antineoplastic Agents / Q000008:administration & dosage* / Q000627:therapeutic use; D018938:Caco-2 Cells; D002401:Cathepsin B / Q000276:immunology*; D016503:Drug Delivery Systems; D054501:Group II Phospholipases A2 / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D018796:Immunoconjugates; D009369:Neoplasms / Q000188:drug therapy; D054467:Phospholipases A2 / Q000378:metabolism*; D014757:Viper Venoms / Q000276:immunology / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D000970:Antineoplastic Agents; D018796:Immunoconjugates; D014757:Viper Venoms; D054501:Group II Phospholipases A2; D054467:Phospholipases A2; C060161:ammodytoxin C; D002401:Cathepsin B