Targeting of the Cholecystokinin-2 Receptor with the Minigastrin Analog Lu-DOTA-PP-F11N: Does the Use of Protease Inhibitors Further Improve In Vivo Distribution?

Patients with metastatic medullary thyroid cancer (MTC) have limited systemic treatment options. The use of radiolabeled gastrin analogs targeting the cholecystokinin-2 receptor (CCK2R) is an attractive approach. However, their therapeutic efficacy is presumably decreased by their enzymatic degradation in vivo. We aimed to investigate whether the chemically stabilized analog Lu-DOTA-PP-F11N (Lu-DOTA-(dGlu)-Ala-Tyr-Gly-Trp-Nle-Asp-Phe-NH) performs better than reference analogs with varying in vivo stability, namely Lu-DOTA-MG11 (Lu-DOTA-dGlu-Ala-Tyr-Gly-Trp-Met-Asp-Phe-NH) and Lu-DOTA-PP-F11 (Lu-DOTA-(dGlu)-Ala-Tyr-Gly-Trp-Met-Asp-Phe-NH), and whether the use of protease inhibitors further improves CCKR2 targeting. First human data on Lu-DOTA-PP-F11N are also reported. In vitro stability of all analogs was assessed against a panel of extra- and intracellular endoproteases, whereas their in vitro evaluation was performed using the human MTC MZ-CRC-1 and the transfected A431-CCK2R(+) cell lines. Biodistribution without and with the protease inhibitors phosphoramidon and thiorphan was assessed 4 h after injection in MZ-CRC-1 and A431-CCK2R(+) dual xenografts. Autoradiography of Lu-DOTA-PP-F11N (without and with phosphoramidon) and NanoSPECT/CT were performed. SPECT/CT images of Lu-DOTA-PP-F11N in a metastatic MTC patient were also acquired. Lu-DOTA-PP-F11N is less of a substrate for neprilysins than the other analogs, whereas intracellular cysteine proteases, such as cathepsin-L, might be involved in the degradation of gastrin analogs. The uptake of all radiotracers was higher in MZ-CRC-1 tumors than in A431-CCK2R(+), apparently because of the higher number of binding sites on MZ-CRC-1 cells. Lu-DOTA-PP-F11N had the same biodistribution as Lu-DOTA-PP-F11; however, uptake in the MZ-CRC-1 tumors was almost double (20.7 ± 1.71 vs. 11.2 ± 2.94 %IA [percentage injected activity]/g, = 0.0002). Coadministration of phosphoramidon or thiorphan increases Lu-DOTA-MG11 uptake significantly in the CCK2R(+) tumors and stomach. Less profound was the effect on Lu-DOTA-PP-F11, whereas no influence or even reduction was observed for Lu-DOTA-PP-F11N (20.7 ± 1.71 vs. 15.6 ± 3.80 [with phosphoramidon] %IA/g, < 0.05 in MZ-CRC-1 tumors). The first clinical data show high Lu-DOTA-PP-F11N accumulation in tumors, stomach, kidneys, and colon. The performance of Lu-DOTA-PP-F11N without protease inhibitors is as good as the performance of Lu-DOTA-MG11 in the presence of inhibitors. The human application of single compounds without unessential additives is preferable. Preliminary clinical data spotlight the stomach as a potential dose-limiting organ besides the kidneys.