RT Journal Article SR Electronic T1 Enhancing Tumor Uptake for 177Lu-Labeled MC1R Targeting Radioligands by Combining N-Methylations with an Albumin Binder JF Journal of Nuclear Medicine JO J Nucl Med FD Society of Nuclear Medicine SP 378 OP 378 VO 61 IS supplement 1 A1 Zhang, Chengcheng A1 Zhang, Zhengxing A1 Zeisler, Jutta A1 Colpo, Nadine A1 Lin, Kuo-Shyan A1 Benard, Francois YR 2020 UL http://jnm.snmjournals.org/content/61/supplement_1/378.abstract AB 378Objectives: Radiotracers derived from Melanotan II (MTII), a melanocortin-1 receptor (MC1R) targeting peptide, have been proposed for melanoma imaging and radionuclide therapy. We previously reported CCZ01048, an MTII derivative which showed excellent imaging properties with 68Ga at early time points. However, when radiolabeled with 177Lu, moderate tumor uptake was observed at longer time points, due to the sub-optimal in vivo stability. The aim of this study was to evaluate a panel of novel N-methylated MTIIderivatives for their binding affinity and stability, and characterize the top candidate, conjugated with an albumin binder, for tumor uptake in a preclinical model of melanoma. Methods: The peptides were prepared by Fmoc solid phase synthesis. N-methylation was performed under Mitsunobu conditions. The DOTA-containing peptides were synthesized with one to four N-methylations on the His, Arg, Trp and Lys residues of CCZ01048. For the derivatives conjugated with an albumin binder, Fmoc-Lys(Mtt)-OH, Fmoc-Glu(OtBu)-OH and 4-(p-iodophenyl)butyric acid were used before coupling DOTA chelator. Receptor binding affinity was measured using in vitro competition binding assays on B16F10 cells. The peptides were radiolabeled with 68Ga, and their in vivo stability was performed via radio-HPLC using urine samples at 1 h post-injection (p.i.) from C57BL/6J mice. The top candidate was conjugated with the albumin binder and radiolabeled with 177Lu. SPECT/CT imaging and biodistribution studies were performed using 177Lu-labeled peptides in C57BL/6J mice bearing B16F10 tumors. Results: A panel of six novel MTII derivatives were synthesized with one to four N-methylations, which had a positive impact on their stability, with average in vivo urine stability ranging from 41.4% to ≥ 98% (n ≥ 3), compared to [68Ga]Ga-CCZ01048 at 34%. The N-methylations had an unfavorable impact on binding affinity, with average inhibition constants ranging from 1.5 to 35.9 nM (n ≥ 3), compared to 0.31 nM for natGa-CCZ01048. The best candidate was CCZ01106, which had improved stability (82.5%) while maintaining excellent binding affinity (1.5 nM). An albumin binder was conjugated to CCZ01106 to afford CCZ01118. 177Lu-labeled CCZ01118 was evaluated in B16F10 tumor-bearing C57BL/6J mice at 1, 4, 24 and 120 h p.i., and the tumor uptake was 24.44 ± 1.76, 29.70 ± 5.23, 18.35 ± 1.35, and 2.63 ± 0.21 percent injected dose per gram of tissue (%ID/g), respectively (n = 5). This was a significant improvement over 177Lu-labeled CCZ01048 with respective tumor uptake values of 16.28 ± 3.75, 16.71 ± 3.86, 4.39 ± 1.36, and 0.41 ± 0.21%ID/g at the same time points[FB1] . SPECT imaging also showed consistent findings as much higher radioactivity accumulation was visualized in tumors. Conclusion: With the addition of N-methylations and an albumin binder, we designed CCZ01118, which showed greatly enhanced tumor uptake with 177Lu in SPECT imaging and biodistribution studies. CCZ01118 is therefore a promising compound for radionuclide therapy of melanoma.