Design, Synthesis, and Biological Evaluation of the First Radio-Metalated Neurotensin Analogue Targeting Neurotensin Receptor 2.

Neurotensin receptor 2 (NTS 2 ) is a well-known mediator of central opioid-independent analgesia. Seminal studies have highlighted NTS 2 overexpression in a variety of tumors including prostate cancer, pancreas adenocarcinoma, and breast cancer. Herein, we describe the first radiometalated neurotensin analogue targeting NTS 2 . JMV 7488 (DOTA-(βAla) 2 -Lys-Lys-Pro-(D)Trp-Ile-TMSAla-OH) was prepared using solid-phase peptide synthesis, then purified, radiolabeled with 68 Ga and 111 In, and investigated in vitro on HT-29 cells and MCF-7 cells, respectively, and in vivo on HT-29 xenografts. [ 68 Ga]Ga-JMV 7488 and [ 111 In]In-JMV 7488 were quite hydrophilic (logD 7.4 = -3.1 ± 0.2 and -2.7 ± 0.2, respectively, p < 0.0001). Saturation binding studies showed good affinity toward NTS 2 ( K D = 38 ± 17 nM for [ 68 Ga]Ga-JMV 7488 on HT-29 and 36 ± 10 nM on MCF-7 cells; K D = 36 ± 4 nM for [ 111 In]In-JMV 7488 on HT-29 and 46 ± 1 nM on MCF-7 cells) and good selectivity (no NTS 1 binding up to 500 nM). On cell-based evaluation, [ 68 Ga]Ga-JMV 7488 and [ 111 In]In-JMV 7488 showed high and fast NTS 2 -mediated internalization of 24 ± 5 and 25 ± 11% at 1 h for [ 111 In]In-JMV 7488, respectively, along with low NTS 2 -membrane binding (<8%). Efflux was as high as 66 ± 9% at 45 min for [ 68 Ga]Ga-JMV 7488 on HT-29 and increased for [ 111 In]In-JMV 7488 up to 73 ± 16% on HT-29 and 78 ± 9% on MCF-7 cells at 2 h. Maximum intracellular calcium mobilization of JMV 7488 was 91 ± 11% to that of levocabastine, a known NTS 2 agonist on HT-29 cells demonstrating the agonist behavior of JMV 7488. In nude mice bearing HT-29 xenograft, [ 68 Ga]Ga-JMV 7488 showed a moderate but promising significant tumor uptake in biodistribution studies that competes well with other nonmetalated radiotracers targeting NTS 2 . Significant uptake was also depicted in lungs. Interestingly, mice prostate also demonstrated [ 68 Ga]Ga-JMV 7488 uptake although the mechanism was not NTS 2 -mediated.