| Abstract: |
Multistep targeting can improve the therapeutic index of antibody-based targeting, particularly relevant to pediatric tumors where acute toxicity and late effects of treatment are major concerns. Neuroblastoma is uniquely suited for such investigations because of its abundance of surface ganglioside GD2. Methods: 5F11scFv (scFv = single-chain variable fragment) was constructed from the variable regions of the heavy (VH) and κ-light (VL) chain complementary DNA (cDNA) of anti-GD2 IgM hybridoma 5F11 and ligated to full-length streptavidin cDNA for expression in Escherichia coli. Purified 5F11-scFv-streptavidin (5F11-scFv-SA) was a homotetramer and showed comparable avidity to 5F11 IgM and a 30-fold improvement over monomeric scFv. Biodistribution of 5F11-scFv-SA was studied in nude mice xenografted with neuroblastoma LAN-1. Twenty-four hours after intravenous injection of 300-900 μg 5F11-scFv-SA, 150-450 μg of a thiogalactoside-containing clearing agent, (Gal-NAc)16-α-S-C5H10-NH-LC-N-Me- biotin (molecular weight, 8,652), were administered intravenously, followed by ∼2.5 μg (1.85-3.7 MBq) 111In-1,4,7,10-tetraazacyclododecane-1, 4,7,10-tetraacetic acid-biotin (111In-DOTA-biotin) intravenously 4 h later and clocked as time 0. Results: Tumor uptake (percentage of injected dose per gram [%ID/g]) at 2 h was 7 %ID/g and decayed with a half-life of 72 h, whereas blood %ID/g rapidly decreased to <1/500 of that of tumor after the first 24 h. The tumor-to-nontumor (T/NT) ratio at 72 h was high (median, 106; range, 3.4 [kidney] to 1,660 [blood]). When the area under the radioactivity curve was computed, the T/NT organ ratio was favorable (4.8 for kidney and 162 for blood). When human and murine tumors were surveyed, the T/NT ratio of 111In-DOTA-biotin uptake correlated with their levels of GD2 expression as assayed by flow cytometry. Biotinylated polypeptides (bovine serum albumin and vasointestinal peptides) achieved selective tumor targeting when the multistep strategy was applied. Conclusion: Improvement in the T/NT ratio using pretargeting strategy may increase the efficacy and safety of scFv-based approaches in cancer therapy. Additionally, since biotinylated polypeptides can be rendered tumor selective, a large repertoire of agents can potentially be explored. |
| Keywords: |
controlled study; protein expression; nonhuman; protein analysis; mouse; animal; metabolism; mouse mutant; animals; mice; in vivo study; cancer cell culture; in vitro study; mice, scid; tumor cells, cultured; mice, inbred c57bl; c57bl mouse; indium radioisotopes; cell culture; nude mouse; mice, nude; neuroblastoma; ganglioside gd2; dosimetry; protein purification; escherichia coli; radioactivity; neoplasms, experimental; experimental neoplasm; high performance liquid chromatography; 1,4,7,10 tetraazacyclododecane 1,4,7,10 tetraacetic acid; single chain fragment variable antibody; streptavidin; scintigraphy; complementary dna; biodistribution; gangliosides; ganglioside; indium; humans; human; priority journal; article; multistep targeting; scfv-streptavidin; single-chain variable fragment; sialogangliosides
|
