Substitution of L-Trp by α-methyl-L-Trp in 177Lu-RM2 results in 177Lu-AMTG, a high affinity GRPR ligand with improved in vivo stability

Guenther T, Deiser S, Felber V, Beck R and Wester HJ

14.01.2022 [Original Artikel]

Theranostic applications targeting the gastrin-releasing peptide receptor (GRPR) have shown promising results. When compared with other peptide ligands for radioligand therapy, the most often used GRPR ligand, RM2 (DOTA-Pip5-D-Phe6-Gln7-Trp8-Ala9-Val10-Gly11-His12-Sta13-Leu14-NH2), may be clinically impacted by limited metabolic stability. Aim: With the aim to improve the metabolic stability of RM2, we investigated whether the metabolically unstable Gln7-Trp8 bond within the pharmacophore of RM2 can be stabilized via substitution of L-Trp8 by α-methyl-L-tryptophan (α-Me-L-Trp) and whether the corresponding DOTAGA analogue might also be advantageous. A comparative preclinical evaluation of 177Lu-α-Me-L-Trp8-RM2 (177Lu-AMTG) and its DOTAGA counterpart (177Lu-AMTG2) was carried out using 177Lu-RM2 and 177Lu-NeoBOMB1 as reference compounds. Methods: Peptides were synthesized by solid-phase peptide synthesis (SPPS) and labeled with 177Lu. Lipophilicity was determined at pH 7.4 (logD7.4). Receptor-mediated internalization was investigated on PC-3 cells (37 °C, 60 min), whereas GRPR affinity (IC50) was determined on both PC-3 and T-47D cells. Stability towards peptidases was examined in vitro (human plasma, 37 °C, 72 ± 2 h) and in vivo (murine plasma, 30 min post injection (p.i.)). Biodistribution studies were carried out at 24 h p.i. and single photon emission tomography/computed tomography (µSPECT/CT) images in PC-3 tumor-bearing mice at 1, 4, 8, 24 and 28 h p.i. Results: Syntheses via SPPS yielded 9-15% purified labeling precursors. 177Lu-labeling proceeded quantitatively. Compared to 177Lu-RM2, 177Lu-AMTG showed slightly improved GRPR affinity, a similar low internalization rate, slightly increased lipophilicity, and considerably improved stability in vitro and in vivo. In vivo, 177Lu-AMTG exhibited the highest tumor retention (11.45 ± 0.43 %ID/g) and tumor/blood ratio (2702 ± 321) at 24 h p.i. as well as a favorable biodistribution profile. As demonstrated by μSPECT/CT imaging, 177Lu-AMTG also revealed a less rapid clearance from tumor tissue. Compared to 177Lu-AMTG, 177Lu-AMTG2 did not show any further beneficial effects. Conclusion: The results of this study, particularly the superior metabolic stability of 177Lu-AMTG, strongly recommend a clinical evaluation of this novel GRPR-targeted ligand to investigate its potential for radioligand therapy of GRPR-expressing malignancies.