Stratford EW, Bostad M, Castro R, Skarpen E, Berg K, Hogset A, Myklebost O, Selbo PK (2013) Photochemical internalization of CD133-targeting immunotoxins efficiently depletes sarcoma cells with stem-like properties and reduces tumorigenicity. Biochim Biophys Acta 1830(8):4235-4243. doi: 10.1016/j.bbagen.2013.04.033
Summary: In this work the authors used photochemical internalization (PCI) to facilitate local cytosolic toxin release from various biotinylated-anti-CD133 antibodies coupled with Streptavidin-ZAP (Cat. #IT-27) in SW872 cells. This technique demonstrates potential for non-invasive sarcoma therapy.
How can I target B-cells? Which secondary conjugate should I use?
ATS makes secondary conjugates that use your primary antibody to target cells for elimination. Just mix your primary antibody with a secondary antibody conjugated to the ribosome-inactivating protein, Saporin, to screen your antibodies. The cells targeted by your primary antibody are eliminated.
Recognizes whole IgG
Hum-ZAP (Cat. #IT-22) is made with a bivalent secondary antibody that recognizes whole IgG. B-Cells have endogenous IgGs.
Fab-ZAP (Cat. #IT-51) is made with a monovalent secondary antibody that recognizes whole IgG. B-Cells have endogenous IgGs.
Recognizes Fc region
FabFc-ZAP (Cat. #IT-65) is made with a monovalent secondary antibody that recognizes ONLY the FC portion of IgG.
Daniotti JL (2013) Featured Article: Antibodies to glycosphingolipids: An attractive tool for targeted delivery of cytotoxic agents to tumor cells. Targeting Trends 14(2)
Hernandez L, Flenker K, Hernandez F, Klingelhutz A, McNamara J, Giangrande P (2013) Methods for evaluating cell-specific, cell-internalizing RNA aptamers. Pharmaceuticals (Basel) 6:295-319. doi: 10.3390/ph6030295
Objective: Isolate aptamers that internalize upon binding to their cognate receptor on the cell surface
Summary: Among the methods used to characterize aptamers that internalize is a way to monitor for cytoplasmic delivery using the ribosome inactivating protein-based (RNA-RIP) assay. Biotin-labeled A9g was conjugated to streptavidin-modified saporin (streptavidin-ZAP). First, it was verified that conjugation of biotinylated aptamer to Streptavidin-ZAP (A9g-SAP) did not affect the inhibitory effect of the aptamer. Next, the effect was examined of A9g-SAP on PC3(PSMA+) and PC3(PSMA-) cells. Cells were treated with varying amounts of aptamer-saporin conjugate for 72 h at 37°C and then an assay was performed to determine potential cytotoxicity of the conjugate. Results confirm that A9g is internalized preferentially into target cells and that A9g is efficiently accessing the cytoplasm of target cells possibly through a mechanism of endosomal escape, resulting in inhibition of protein synthesis and ultimate cell-death. FGF-SAP was used as a control.
Rust S, Guillard S, Sachsenmeier K, Hay C, Davidson M, Karlsson A, Karlsson R, Brand E, Lowne D, Elvin J, Flynn M, Kurosawa G, Hollingsworth R, Jermutus L, Minter R (2013) Combining phenotypic and proteomic approaches to identify membrane targets in a ‘triple negative’ breast cancer cell type. Mol Cancer 12:11. doi: 10.1186/1476-4598-12-11
Summary: The authors investigated a phenotypic antibody screening technique, in which antibodies are selected by function rather than target specificity. One facet of the screening procedure for hybridomas generated using a cancer cell line as antigen was the use of Mab-ZAP (Cat. #IT-04) to assess cell binding and internalization.
Ren C, Luan L, Wui-Man Lau B, Huang X, Yang J, Zhou Y, Wu X, Gao J, Pickard GE, So KF, Pu M (2013) Direct retino-raphe projection alters serotonergic tone and affective behavior. Neuropsychopharmacology 38(7):1163-1175. doi: 10.1038/npp.2013.35
Summary: Although recent work has shown that some intrinsically photosensitive retinal ganglion cells (ipRGCs) are responsible for processing nonimage-forming visual functions, it is unclear whether the ipRGCs or conventional RGCs modulate affective behavior. The authors injected 2 μg of melanopsin-SAP (Cat. #IT-44) into each eye of gerbils, or biotinylated CTB monoclonal antibody coupled to Streptavidin-ZAP (Cat. #IT-27). The data suggest that retino-raphe signals modulate dorsal raphe nucleus serotonergic tone and affective behavior.
Torres Demichelis V, Vilcaes AA, Iglesias-Bartolome R, Ruggiero FM, Daniotti JL (2013) Targeted delivery of immunotoxin by antibody to ganglioside GD3: A novel drug delivery route for tumor cells. PLoS One 8(1):e55304. doi: 10.1371/journal.pone.0055304
Summary: The authors used the mouse monoclonal antibody R24 against ganglioside G3 with Mab-ZAP (Cat. #IT-04) to test the viability of ganglioside G3 as a cancer therapy target. Varying concentrations of R24 were used on various cell lines with either 0.95 nM or 9.5 nM Mab-ZAP depending on the cell line.
Tuscano JM, Kato J, Pearson D, Xiong C, Newell L, Ma Y, Gandara DR, O’Donnell RT (2012) CD22 antigen is broadly expressed on lung cancer cells and is a target for antibody-based therapy. Cancer Res 72(21):5556-5565. doi: 10.1158/0008-5472.CAN-12-0173
Summary: The median overall survival of patients with advanced, unresectable, non-small cell lung cancer is 9-12 mos. A potential therapeutic target is CD22, a protein expressed on lung cancer cells. The authors examined the use of the monoclonal antibody HB22.7 as an antitumor agent. To assess internalization of the antibody, it was first incubated with 10 μg/ml Mab-ZAP (Cat. #IT-04) then applied to two different cancer cell lines in culture. Analysis of cell viability demonstrated that CD22 internalized when bound by the antibody-toxin complex, suggesting that targeting CD22 has therapeutic potential.
Daniels-Wells TR, Helguera G, Rodriguez JA, Leoh LS, Erb MA, Diamante G, Casero D, Pellegrini M, Martinez-Maza O, Penichet ML (2013) Insights into the mechanism of cell death induced by saporin delivered into cancer cells by an antibody fusion protein targeting the transferrin receptor 1. Toxicol In Vitro 27(1):220-231. doi: 10.1016/j.tiv.2012.10.006
Summary: The antibody-avidin fusion protein ch128.1Av has been shown to target the human transferrin receptor 1 (TfR1) and kill malignant B cells by blocking the use of iron. Combination of this construct with a mono-biotinylated saporin custom conjugate produces an iron-independent toxicity to TfR1-expressing cells, even those that are resistant to ch128.1Av alone. The saporin-containing conjugate induces a transcriptional response consistent with oxidative stress and DNA damage. The data also show that the saporin conjugate is not toxic to human hematopoeietic stem cells.
Usage: An antibody-avidin fusion protein (ch128.1Av) was mixed with MonoBiotin-ZAP to make an immunotoxin that targets the human transferrin receptor 1 (TfR1).
Berstad MB, Weyergang A, Berg K (2012) Photochemical internalization (PCI) of HER2-targeted toxins: Synergy is dependent on the treatment sequence. Biochim Biophys Acta 1820(12):1849-1858. doi: 10.1016/j.bbagen.2012.08.027
Summary: A majority of patients develop acquired resistance to trastuzumab, the monoclonal antibody recognizing HER2, coupled to a toxin as a breast cancer therapeutic. One of the modes of resistance is that the therapeutic molecule is trapped inside an endocytic vesicle. PCI is a technique that facilitates cytosolic release of molecules in vesicles. The authors investigated the potency of biotinylated trastuzumab combined with streptavidin-ZAP (Cat. #IT-27) on several cell lines.
