Radiolabeling and biological evaluation of DOTA-Ph-Al derivative conjugated to anti-EGFR antibody ior egf/r3 for targeted tumor imaging and therapy

Abstract

An appropriate bifunctional chelating agent namely DOTA-Ph-Al was developed for the conjugation with biological vectors (anti EGFr antibody). We hereby report the synthesis of p-bromoacetamidobenzyl derivative of DOTA and its conjugation to monoclonal antibody anti-EGFR ior egf/r3. Immunoconjugate was prepared by conjugation of p- bromoacetamidobenzyl derivative of DOTA with ior egf/r3. Modified antibody was purified on size exclusion chromatography. DOTA-Ph-Al-ior egf/r3 exhibited quantitative 99mTc labeling (>96%) with specific activity 10-20 mCi/mg of protein and 90Y- labeling with specific activity 2-5 mCi/mg. Immunoreactivity was determined by flow cytometry. Receptor ligand assay on murine cell line EAT and human tumor cell line U- 87MG showed Kd =2.87 nM and 4.86 nM respectively. The stability in serum indicated that 99mTc remained bound to antibodies up to 24h and 98% 90Y was associated with the mAb for 5 days. Biodistribution characteristics of Ab-conjugate radiolabeled to 99mTc and 90Y radionuclide was examined in BALB/c mice grafted with EAT and athymic mice with U-87MG cell line demonstrated high tumor uptake with 5.5 ± 1.3 and 7.85 ± 1.2 %ID/g at 4 and 24 h for 99mTc- DOTA-Ph-Al-ior egf/r3 in EAT tumors after post injection respectively. Maximal radiotracer uptake peaked 17.6 ± 2.5 %ID/g in EAT tumor and 12.89 ± 0.66 % ID/g in U-87MG tumor at 48h for 90Y. The drug excreted through renal routes as the activity in the kidneys was 13.42 ± 0.33 %ID/g at 1h and 4.51± 1.2 %ID/g at 4h for 99mTc- DOTA-Ph-Al-ior egf/r3.