Technetium-99m labelling of the anti-tumour antibody PR1A3 by photoactivation.
Irradiation of antibody with ultraviolet light leads to reduction of disulphide bonds. Thus irradiation can be used to generate free thiols prior to direct labelling of antibody with technetium-99m, and has a potential advantage over methods using chemical reducing agents such as mercaptoethanol or tin, in that no purification step is needed to remove excess reducing agent. We have used the photoactivation method developed by Sykes et al. to label the anti-tumour antibody PR1A3 with 99mTc. The antibody was irradiated at 300 nm using a Rayonet photochemical reactor with eight RMR3000 lamps. In a typical experiment, the antibody solution was injected into a nitrogen-filled borosilicate glass vial and purged with nitrogen. A degassed solution containing stannous fluoride and methylene diphosphonate was then added to the antibody and the vial was irradiated. Following the irradiation, [99mTc]pertechnetate was injected into the vial and the reaction mixture was incubated for 30 min at room temperature before being analysed by size-exclusion high-pressure liquid chromatography and instant thin-layer chromatography. Labelling yields greater than 95% were obtained using antibody concentrations ranging from 0.5mg/ml to 5mg/ml. Irradiation times as short as 5 min and tin to antibody ratios in the range between 11 and 32 microg tin per mg antibody gave high labelling yields. Labelling yields greater than 95% were obtained after storage of the photoactivated antibody at -70 degrees C for several weeks. The stability of the 99mTc-labelled photoactivated PR1A3 was similar to that of 99mTc-labelled mercaptoethanol-reduced PR1A3. The mean immunoreactive fraction was 77% for the photoactivation-labelled PR1A3, compared to 93% for PR1A3 labelled by mercaptoethanol reduction. Biodistribution studies were carried out using 99mTc-photoactivation-labelled PR1A3 or PR1A3 labelled by mercaptoethanol reduction in Balb/c mice and in nude mice with MKN-45 human tumour xenografts. There was no significant difference in tumour uptake between the mice that received photoactivated PR1A3 and those that received mercaptoethanol-reduced PR1A3. There was also no significant difference in uptake in most organs in Balb/c mice; however, the photoactivated antibody cleared more rapidly from the blood, and whole-body clearance was also faster for the photoactivated PR1A3. In conclusion, the photoactivation technique provides a very convenient "one-pot" method for labelling antibodies with 99mTc.
Stalteri
M A
MA
Imperial Cancer Research Fund, Department of Nuclear Medicine, St. Bartholomew s Hospital, West Smithfield, London EC1A 7BE, UK.
Mather
S J
SJ
D000818:Animals; D000911:Antibodies, Monoclonal; D058846:Antibodies, Monoclonal, Murine-Derived; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D006801:Humans; D007553:Isotope Labeling / Q000379:methods*; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D016719:Radioimmunodetection / Q000379:methods*; D013670:Sodium Pertechnetate Tc 99m; D014018:Tissue Distribution; D014466:Ultraviolet Rays
D000911:Antibodies, Monoclonal; D058846:Antibodies, Monoclonal, Murine-Derived; C511152:PR1A3 monoclonal antibody; D013670:Sodium Pertechnetate Tc 99m