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A new method is reported for labeling proteins with the positron-emitting nuclide 18F. Initially, 4-[18F]-fluorobenzylamine was prepared in two steps from aqueous [18F]fluoride in high yield. The 18F acylation agent was formed by reaction of this product with disuccinimidyl suberate. Overall yields for the 4-[18F]fluorobenzylamine succinimidyl ester ([18F]SFBS), decay corrected to the end of cyclotron bombardment, were about 30% in a synthesis time of 60 min. After a 15-min reaction, 30-45% (decay corrected) of the [18F]SFBS could be coupled to intact antibodies and their F(ab')2 and Fab fragments. Coupling yields were dependent on protein concentration but not reaction time. HPLC purification of [18F]SFBS was necessary to obtain optimal coupling efficiency and immunoreactivity. The immunoreactivities of 18F-labeled F(ab')2 and Fab fragments of an antimyosin antibody were 89 +/- 5% and 75 +/- 9%, respectively. Biodistribution studies in normal mice demonstrated similar in vivo behavior of 18F-labeled antibody fragments and those labeled with 125I by using N-succinimidyl 3-[125I]iodobenzoate. These results indicate that this method may be useful for labeling monoclonal antibodies and other proteins and peptides with 18F.

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Fab/c fragments, purified from pepsin digest of mouse IgG2b monoclonal antibody (MoAb) 791T/36, consist of one Fab arm and the intact Fc portion. Pharmacokinetic and biodistribution studies in BALB/c mice of radioiodine-labeled 791T/36 MoAb and its Fab/c fragments showed that, due to the presence of the Fc portion, the Fab/c fragment has the same catabolic rate as whole antibody (T1/2 = 64 h). Due to its lower molecular weight (105,000), the Fab/c fragment extravasated more quickly and to a greater extent than whole MoAb in organs in which the vascular endothelium was fenestrated or continuous. In organs in which the vascular endothelium is sinusoidal, such as in liver and spleen, their diffusion capacities were identical. Therefore, Fab/c fragments reconcile advantages of the intact antibody molecule (slow catabolic rate) and Fab or F(ab)2 fragments (increased extravascular diffusion), features required to improve targeting to solid tumors. Data from biodistribution studies in nude mice bearing subcutaneous 791T tumor (antigen positive) and Colo205 tumor (antigen negative) contralaterally showed important differences in the behavior of whole MoAb and Fab/c fragment: (a) Whole MoAb was cleared more rapidly from the body and from the blood than Fab/c fragment; (b) The MoAb was taken up by the spleen (tissue to blood ratio greater than 1 from 12 h after injection over the 3 days of the experiment) and the liver (0.6), whereas Fab/c fragment tissue to blood ratios were only slightly increased (0.34 and 0.35) compared to control nude mice (0.25 and 0.28) for the spleen and liver, respectively, 3 days after injection. Since both MoAb and Fab/c fragment bear the Fc portion, these data suggest that the reputed "nonspecific uptake" of antibodies due to the Fc portion could be an Fc-mediated specific uptake, e.g., uptake of immune complexes; (c) The tumor to blood ratios were 1.7 and 1.2 for MoAb and Fab/c fragment, respectively, from 24 h throughout the experiment, whereas the percentage of injected dose (% of ID) present/g of 791T tumor was at any time greater for Fab/c fragment (8% maximum of ID) than for MoAb (5% of ID). These results were not expected in view of the low immunoreactivity in vitro of Fab/c fragments compared to whole antibody. It is suggested that the distribution and the catabolic rate of whole antibody and its Fab/c fragment at the tumor level are modulated by their respective valency and immunoreactivity for the target cell.

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Hydroxylapatite high performance liquid chromatography was used to prepare two fractions from 125I- Fab 96.5. One fraction (peak 1) had relatively low immunoreactivity (25-38%) and the second fraction (peak 2) had high immunoreactivity (70-81%). Scatchard analysis showed similar affinity constants for the two preparations (2.9 x 10(9) M-1 for peak 1; 3.4 x 10(9) M-1 for peak 2). In biodistribution and imaging studies in athymic mice with human melanoma (FEMX-II) xenografts the high immunoreactivity preparation rapidly cleared from the blood and nontumor organs while retention of radioactivity in the tumor was prolonged. The low immunoreactivity preparation, had slower blood and nontumor organ clearance but faster tumor clearance than the high immunoreactivity fraction. Thus, in these studies highly immunoreactive antibody gave higher tumor to nontumor ratios and enhanced the target to nontarget image contrast.

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Forty seven patients with suspected malignant disease (mainly colorectal cancer) were studied with 111In labelled F(ab')2 fragments of an anti-CEA monoclonal antibody (BW 431/31). The kinetic data revealed a long whole body retention of the label (62% after 4 days) and a rapid blood clearance (77% within 24 h, 89% within 48 h) leading to an early positive tumour contrast 24 h p.i. and optimal scintigrams 48 h p.i. Diagnostic results were promising in local recurrences of colorectal cancer (8/10 positive = 80%) though false positive findings in patients with inflammatory bowel disease occurred probably due to cross-reaction with a human granulocyte antigen. Liver metastases and tumours neighbouring liver and spleen were often missed (10/27 = 37%) because of high nonspecific uptake in these organs. Thus BW 431/31 proved to be a suitable compound for radioimmunodetection, however, further improvements to optimize tumour affinity and specificity of the antibody are mandatory.

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