Prevention of radiolysis of monoclonal antibody during labeling.
Monoclonal antibody may undergo loss of immunoreactivity due to radiation damage when labeled with large amounts of 131I or 90Y for therapy. Our aim was to develop a method to protect an antibody during the labeling procedure.


METHODS
As a model we used T101, a murine monoclonal antibody directed against CD5 antigen. Iodine-125-T101 (100 micrograms, 1 ml) was mixed with 90Y-DTPA (0.64 MBq to 165.9 MBq) for 24 hr in order to deliver doses of 5 Gy to 1280 Gy to the solution. In separate experiments, 125I-T101 solutions were irradiated with 60Co external beam delivering radiation doses of 40 Gy to 1280 Gy. The effect of radiation on T101 immunoreactivity was tested by using the CCRF-CEM cell line, and the bound T101 radioactivity was determined. In a final experiment, we directly labeled a DTPA conjugated T101 using 561 MBq of 90Y under conditions delivering approximately 640 Gy to the solution. Previously used radioprotectants including human serum albumin, cysteamine and glycerol were evaluated. We focused on ascorbic acid because it is an FDA approved drug that does not interfere with the radiolabeling process.


RESULTS
The immunoreactivity of 125I-T101 was approximately 83%, but at 640 Gy the immunoreactivity decrease to 7%. In contrast, in the presence of radioprotectants this decrease could be abrogated. External irradiation also showed a dose dependent decrease in immunoreactivity to as low as 0.3% at 1280 Gy. Adding ascorbic acid (5.5 mg/ml) to the solutions prior to the irradiation largely abrogated this decrease. The immunoreactivity of T101 labeled with 90Y without protectant showed 46% immunoreactivity whereas, in presence of ascorbic acid (11 mg/ml) full retention of immunoreactivity was observed.


CONCLUSIONS
Various radioprotectants can successfully prevent the loss of immunoreactivity or breakdown as a result of radiolysis. Ascorbic acid is an effective radioprotectant that can be used to prevent loss of antibody immunoreactivity during the labeling process.
Chakrabarti
M C
MC
Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, Bethesda, Maryland 20892-1180, USA.
Le
N
N
Paik
C H
CH
De Graff
W G
WG
Carrasquillo
J A
JA
D000818:Animals; D000911:Antibodies, Monoclonal / Q000528:radiation effects*; D001205:Ascorbic Acid / Q000494:pharmacology*; D003037:Cobalt Radioisotopes; D003543:Cysteamine / Q000494:pharmacology; D005990:Glycerol / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D007457:Iodine Radioisotopes / Q000627:therapeutic use*; D007553:Isotope Labeling; D051379:Mice; D008807:Mice, Inbred BALB C; D011829:Radiation Dosage; D011837:Radiation-Protective Agents / Q000494:pharmacology*; D016499:Radioimmunotherapy; D012709:Serum Albumin / Q000494:pharmacology; D015021:Yttrium Radioisotopes / Q000627:therapeutic use*
D000911:Antibodies, Monoclonal; D003037:Cobalt Radioisotopes; D007457:Iodine Radioisotopes; D011837:Radiation-Protective Agents; D012709:Serum Albumin; D015021:Yttrium Radioisotopes; D003543:Cysteamine; D005990:Glycerol; D001205:Ascorbic Acid