Gastrin-releasing peptide receptor as theranostic target in estrogen-receptor positive breast cancer: A preclinical study of the theranostic pair [⁵⁵Co]Co- and [¹⁷⁷Lu]Lu-DOTA-RM26

Background: Patients with advanced metastatic estrogen receptor-positive breast cancer often develop resistance to standard treatments, leading to uncontrolled progression. Thus, innovative therapies are urgently needed. The gastrin-releasing peptide receptor (GRPR) is overexpressed in various cancers, including breast cancer, making it an interesting theranostic target. RM26, a GRPR-targeting antagonist, has demonstrated promising in vivo kinetics in prostate cancer models. This study evaluated the theranostic capabilities of [⁵⁵Co]Co-/[¹⁷⁷Lu]Lu-DOTA-RM26 in vitro in estrogen receptor-positive breast cancer cells and assessed the diagnostic potential of [⁵⁵Co]Co-DOTA-RM26 in vivo in a breast cancer mouse model.

Methods: We analyzed the binding specificity of [⁵⁷Co]Co-/[¹⁷⁷Lu]Lu-DOTA-RM26 in T47D breast cancer cells, using [⁵⁷Co]Co-DOTA-RM26 as a surrogate for [⁵⁵Co]Co-DOTA-RM26. The therapeutic efficacy of increasing [¹⁷⁷Lu]Lu-DOTA-RM26 concentrations was determined via viability assay in vitro. Ex vivo biodistribution of [⁵⁷Co]Co-DOTA-RM26 (17.2 ± 2.7 kBq, 33 ± 5.2 pmol/mouse) was investigated in 12 mice (n= 4/group) with orthotopic breast cancer tumors. The mice were sacrificed at 4 and 24 h post-injection (pi), including a blocking group (20 nmol of unlabeled [Tyr4]-Bombesin) at 4 h pi. For imaging, two tumor-bearing mice underwent [⁵⁵Co]Co-DOTA-RM26 PET/CT, 4 and 24 h pi (2.8 ± 0.2 MBq, 167.5 ± 0.5 pmol/mouse), with or without GRPR blocking.

Results: In vitro studies revealed high, specific binding of [⁵⁷Co]Co-DOTA-RM26 (43 ± 1 % of total added activity per 10⁶ cells (%IA/10⁶)) and [¹⁷⁷Lu]Lu-DOTA-RM26 (37 ± 4 %IA/10⁶). The activity was predominantly localized at the cell surface: 71 ± 3 % and 80 ± 6 % for [⁵⁷Co]Co-DOTA-RM26 and [¹⁷⁷Lu]Lu-DOTA-RM26, respectively. [¹⁷⁷Lu]Lu-DOTA-RM26 significantly reduced cell viability at all activity concentrations >0.625 MBq/mL (p < 0.0001), with cell viability below 1 % at concentrations ≥5 MBq/mL. Biodistribution data (n = 12) indicated a high, specific tumor uptake of [⁵⁷Co]Co-DOTA-RM26, surpassing all other tissues significantly at both time points, 3.7 ± 0.6 % of the injected activity per gram (%IA/g) 4 h pi and 0.98 ± 0.05 %IA/g 24 h pi. The kidneys showed the second-highest uptake (2.0 ± 0.1 %IA/g 4 h pi), followed by the pancreas (1.4 ± 0.4 %IA/g 4 h pi). PET/CT imaging with [⁵⁵Co]Co-DOTA-RM26 supported the biodistribution data and, distinctly visualized the tumor 24 h pi and showed an improved tumor-to-background compared to the earlier time points. Effective GRPR blocking significantly reduced tumor uptake in the PET images 24 h pi.

Conclusion: These findings suggest that the theranostic pair [⁵⁵Co]Co-/[¹⁷⁷Lu]Lu-DOTA-RM26 holds significant promise as a theranostic agent for estrogen receptor-positive breast cancer.