BIOTIN REAGENTS FOR ANTIBODY PRETARGETING - SYNTHESIS, RADIOIODINATION, AND IN-VITRO EVALUATION OF WATER-SOLUBLE, BIOTINIDASE RESISTANT BIOTIN DERIVATIVES

As part of our development of antibody pretargeting for cancer therapy , an investigation has been conducted to examine the stability of wate r solubilized, radioiodinated biotin derivatives toward biotinidase de gradation in mouse and human serum. Eight new biotin derivatives were synthesized to conduct the study. The biotin derivatives synthesized c ontained (I) the biotin moiety, (2) a water solubilizing linker moiety , (3) p-iodobenzoate or p-tri-n-butylstannylbenzoate moieties, and (4) in some of the compounds, N-methyl or alpha-methyl containing moietie s were added to block biotinidase activity. The linker moiety, 4,7,10- trioxa-1,13-tridecanediamine, 5, was included in the biotin derivative s to improve their water solubility, and it also functioned as a 17 An gstrom spacer between the biotin and the benzoyl moieties. Four of the new biotin derivatives (12, 14, 22, and 23) contained a p-tri-n-butyl stannylbenzoyl moiety as precursors which could be radioiodinated in t he last synthetic step. The other four biotin derivatives (13, 15, 24, and 25) contained p-iodobenzoyl moieties and were used as HPLC refere nce standards, Initial studies involved radioiodination of 12 to yield [I-125]13. Radioiodinated 13, which did not contain a moiety far bloc king biotinidase activity, was found to be rapidly degraded in both mo use and human serum at 37 degrees C. Derivatives which were designed t o be stable to biotinidase incorporated N-methyl and alpha-methyl moie ties adjacent to the biotin carboxylate group. In one set of biotin de rivatives (14 and 15), the N-methyl moiety was obtained by incorporati ng N,N-dimethyl-4,7,10-trioxa-1,13-tridecanediamine, 9, as a Linker in the place of 5. In the second set of biotin derivatives (22 and 24), the N-methyl moiety was introduced by incorporating a sarcosine (N-met hylglycine) moiety between biotin and 5. The radioiodinated N-methyl c ontaining biotin derivatives [I-125]15 and [I-125]24 were found to be very stable to biotinidase degradation. An alpha-methyl group was obta ined in a pair of biotin derivatives (23 and 25) by incorporating a 8- aminobutyric acid moiety between biotin and 5. The radioiodinated alph a-methyl containing derivative, [I-125]25, was found to have an interm ediate stability with regards to biotinidase degradation.