Targeting of the cholecystokinin-2 receptor with the minigastrin analog 177 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 177 Lu-DOTA-PP-F11N (177 Lu-DOTA-(DGlu)6-Ala-Tyr-Gly-Trp-Nle-Asp-Phe-NH2) performs better than reference analogs with varying in vivo stability, namely 177 Lu-DOTA-MG11 (177 Lu-DOTA-DGlu-Ala-Tyr-Gly-Trp-Met-Asp-Phe-NH2) and 177 Lu-DOTA-PP-F11 (177 Lu-DOTA-(DGlu)6-Ala-Tyr-Gly-Trp-Met-Asp-Phe-NH2), and if the use of protease inhibitors further improves CCKR2-targeting. First human data on 177 Lu-DOTA-PP-F11N are also reported. Methods: In vitro stability of all analogs was assessed against a panel of extra- and intra-cellular endoproteases, while 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 (PA) and thiorphan (TO) was assessed 4h post-injection in MZ-CRC-1 and A431-CCK2R(+) dual xenografts. Autoradiography of 177 Lu-DOTA-PP-F11N (without and with PA) and nanoSPECT/CT images were performed. SPECT/CT images of 177 Lu-DOTA-PP-F11N in a metastatic MTC patient were also acquired. Results: natLu-DOTA-PP-F11N is less of a substrate for neprilysines than the other analogs, while intracellular cysteine proteases, like cathepsins-L, might be involved in the degradation of gastrin analogs. The uptake of all radiotracers was higher in MZ-CRC-1 tumors, compared to A431-CCK2R(+), apparently due to the higher number of binding sites on MZ-CRC-1 cells. 177 Lu-DOTA-PP-F11N has the same biodistribution as 177 Lu-DOTA-PP-F11, however, the uptake in the MZ-CRC-1 tumors is almost double (20.7±1.71 vs 11.2±2.94 %IA/g, P = 0.0002). Co-administration of PA or TO increases significantly the 177 Lu-DOTA-MG11 uptake in the CCK2R(+) tumors and stomach. Less profound is the effect on 177 Lu-DOTA-PP-F11, while no influence or even reduction is observed for 177 Lu-DOTA-PP-F11N (20.7±1.71 vs 15.6±3.80 (with PA) %IA/g, p<0.05 in MZ-CRC-1 tumors). First clinical data show high 177 Lu-DOTA-PP-F11N accumulation in the tumors, stomach, kidneys and colon. Conclusion: The performance of 177 Lu-DOTA-PP-F11N without protease inhibitors is as good as the performance of 177 Lu-DOTA-MG11 in the presence of inhibitors. The human application of single compounds without unessential additives is preferable. Preliminary clinical data spotlight stomach as a potential dose-limiting organ beside the kidneys.