Comparative study on DOTA-derivatized bombesin analog labeled with 90Y and 177Lu: in vitro and in vivo evaluation

Introduction: The aim of the study was to compare in vitro and in vivo a novel DOTA-chelated bombesin (BN) analog of the amino acid sequence, QRLGNQWAVGHLM-CONH2 (BN[2-14]NH2), labeled with 90Y and 177Lu, for its potential use in targeted radiotherapy of tumors expressing gastrin releasing peptide (GRP) receptors. The same amino acid sequence, but with different chelator, referred as BN1.1 (Gly-Gly-Cys-Aca-QRLGNQWAVGHLM-CONH2), has already been studied and reported; however, the DOTA-chelated one, suitable for labeling with M+3 type radiometals, was not yet described. Methods: The conditions for labeling of DOTA-BN[2-14]NH2 with noncarrier added 90Y and with 177Lu [specific activity (SA), 15 Ci/mg Lu] were investigated and optimized to provide 90Y-DOTA-BN[2-14]NH2 and 177Lu-DOTA-BN[2-14]NH2 of high SA. The stability of the radiolabeled compounds in human serum was evaluated over a period of 24 h. The human prostate cancer cell line PC-3, known to express GRP receptors, was used for in vitro evaluation of radiolabeled peptide affinity to GRP receptors and for assessment of cytotoxicity of both nonlabeled and radiolabeled peptide. Biodistribution accompanied by receptor blocking was studied in normal Swiss mice. Results: 90Y-DOTA-BN[2-14]NH2 and 177Lu-DOTA-BN[2-14]NH2 were obtained with radiochemical yield >98% and high SA (67.3 GBq 90Y/μmol and 33.6 GBq 177Lu/μmol, respectively). They were stable when incubated in human serum for up to 24 h. The binding affinities of DOTA-BN[2-14]NH2 and both natY- and natLu-labeled analogs to GRP receptors were high (IC50=1.78, 1.99, and 1.34 nM, respectively), especially for the natLu-DOTA-BN[2-14]NH2 complex. The cytotoxicity study of DOTA-BN[2-14]NH2 to PC-3 cells revealed an IC50=6300 nM after 72 h of exposition, while the labeled derivatives showed no significant cytotoxic effect. The internalization rate to PC-3 cells was more rapid for 177Lu-labeled peptide (84.87%) than for the 90Y-labeled one (80.79%), while the efflux rate was slower for 177Lu-DOTA-BN[2-14]NH2 (46.8% vs. 61.74%). The biodistribution study of both derivatives in normal mice revealed a specific binding to GRP receptor-positive tissues, which could be blocked by coinjection of cold peptide. The effect of receptor blockage in vivo was also more pronounced for the 177Lu-labeled peptide than that for the 90Y-labeled (81% vs. 42%, respectively). Conclusions: Our studies demonstrated that DOTA-BN[2-14]NH2 can be labeled with 90Y (NCA) and 177Lu (CA) with high radiochemical yields. The in vitro and in vivo comparison between 90Y-DOTA-BN[2-14]NH2 and 177Lu-DOTA-BN[2-14]NH2 indicated that the change of radiometal in the complex from Y to Lu influence the binding affinity to the GRP receptors with preference to the 177Lu-labeled derivative.