Yttrium-90 and Rhenium-188 Radiopharmaceuticals for Radionuclide Therapy

separate administration of the MAb and radiolabel [9.26, 9.27]. Owing to the fact
that avidin binds to biotin selectively with extremely high affinity (K d = 10 –15 ),
the avidin biotin system was first applied for pretargeting. In the most complex
three step avidin biotin system, the antibodies were coupled with biotin, which
was used as the primary targeting agent (step 1), this was followed by bridging
with avidin (cleaning agent, step 2) and then administration of biotin conjugated
effector with radionuclide (step 3).
Within the IAEA CRP Development of Therapeutic Radiopharmaceuticals
based on 188 Re and 90 Y for Radionuclide Therapy, the antibody biotinylation
was investigated using MAb rituximab (Roche) and sulpho NHS biotin (Pierce
Biotechnology) as the biotinylating agent. Sulpho NHS biotin effectively reacts
at pH7–9 with primary amino groups of proteins forming stable amide bonds
(see Fig. 9.18). The MAb at concentration 2 mg/mL in PBS was reacted with
a 20-fold molar excess of sulpho NHS biotin solution. After 1 h incubation at
room temperature, unreacted and hydrolysed biotinylation reagent was removed
by dialysis with PBS. The effect of MAb biotinylation was evaluated by adding
an aliquot of biotinylated antibody to the mixture of HABA and avidin. The
spectrophotometric method of biotin determination is based on the decrease
of HABA avidin complex absorbance when HABA is replaced by biotin. The
number of biotin molecules conjugated to MAbs using the HABA avidin method
was 3.9.
Among the radionuclides for radioimmunotherapy, 90 Y is of particular
interest owing to its superior properties including pure β emission
(E b max = 2.2 MeV) and 64.1 h half-life. Bifunctional macrocyclic chelating
agents such as DOTA analogues can be complexed by metal ions with high
stability [9.28].
Investigation of biotin labelling with 90 Y was performed using the conjugate
of biotin with DOTA as DOTA biotin sarcosine (Macrocyclics) (see Fig. 9.19).
FIG. 9.18. Biotinylation of proteins using sulpho NHS biotin.
164