Abstract
1106
Objectives: The melanocortin-1 receptor (MC1R) is expressed in cutaneous and metastatic melanomas. As an endogenous ligand, alpha-melanocyte stimulating hormone (αMSH) binds to MC1R with high affinity. We previously developed a 68Ga-labeled αMSH derivative (CCZ01048) that showed excellent melanoma visualization and tumor-to-background contrast using positron emission tomography. In this study, we performed single-photon emission computed tomography (SPECT) and biodistribution studies with 177Lu-labeled CCZ01048 in a preclinical model of melanoma. In addition, to further increase tumor uptake, we introduced a novel albumin binder to the sequence of CCZ01048 to extend its retention time in blood. The resulting albumin-binding αMSH derivative (CCZ01082) was also radiolabeled with 177Lu for evaluation.
Methods: DOTA-Pip-Nle-CycMSHhex (CCZ01048) was produced by standard Fmoc solid phase synthesis. For CCZ01082, Fmoc-Lys(Mtt)-OH, Fmoc-Glu(OtBu)-OH and 4-(p-iodophenyl)butyric acid was sequentially coupled to NH2-Pip-Nle-CycMSHhex. After selective removal of the Mtt protecting group, a DOTA chelator was conjugated to the amino group at Lys side chain. For both tracers, 177Lu labeling was performed in sodium acetate buffer (0.1 M, pH 4.5) at 90°C for 15 min followed by HPLC purification. Receptor binding affinity was measured by competitive binding assays using B16F10 cells with non-radioactive lutetium-complexed peptides. SPECT/CT imaging and biodistribution studies were performed in C57BL/6J mice bearing B16F10 tumors at 1, 4, 24 and 120 h post-injection (p.i.).
Results: Both natLu-CCZ01048 and natLu-CCZ01082 were prepared in high purity (> 98%), and both had excellent binding affinity to MC1R (Ki = 0.21 ± 0.03 and 0.41 ± 0.02 nM respectively, n =3). 177Lu-labeled peptides were prepared with high radiochemical purity (> 95%) and specific activity ranging from 85.1 to 177.6 MBq/nmol. SPECT imaging and biodistribution studies showed high tumor uptake of 16.28 ± 3.75 and 16.71 ± 3.86 percent injected dose per gram of tissue (%ID/g) for 177Lu-CCZ01048, and 13.32 ± 3.34 and 20.82 ± 5.37 %ID/g for 177Lu-CCZ01082 at 1 h and 4 h p.i. (n ≥ 5), respectively. Minimal background activity, e.g. blood (0.50 ± 0.02 %ID/g) and muscle (0.13 ± 0.02 %ID/g), was observed for 177Lu-CCZ01048 at 1 h p.i., besides the kidneys (5.36 ± 0.32 %ID/g) and thyroid (2.57 ± 0.62 %ID/g). 177Lu-CCZ01048 cleared rapidly through the kidneys. In contrast, with the addition of an albumin binder in CCZ01082, the radioligand retained in circulation longer as blood uptake was 8.32 ± 2.74 and 2.82 ± 1.17 %ID/g at 1 and 4 h p.i., respectively, and decreased to 0.07 ± 0.00 %ID/g at 24 h p.i. The longer blood retention for 177Lu-CCZ01082 resulted in higher tumor uptake of 7.59 ± 1.70 and 0.79 ± 0.39 %ID/g at 24 and 120 h (n ≥ 5), respectively, compared to the tumor uptake of 177Lu-CCZ01048 at 4.39 ± 1.36 and 0.41 ± 0.21 %ID/g at 24 and 120 h, respectively. Conclusion: Excellent tumor uptake was achieved for both 177Lu-labeled CCZ01048 and CCZ01082 at early time points (1 and 4 h p.i.) in a preclinical model of melanoma. The introduction of a novel albumin binder led to longer radioactivity retention in blood, and resulted in higher tumor uptake at later time points (24 and 120 h p.i.).