Prostate cancer diagnosis: Radiochemical characterization of 68Ga-PSMA

Objectives To optimize the radiolabelling of Glu-NH-CO-NH-Lys-(Ahx)-HBED-CC with 68Ga (68Ga-PSMA) for prostate cancer diagnosis. The optimization included: 1) Evaluation of the ratio activity/precursor. 2) Examination of the stability at room temperature (RT) under different incubation conditions. 3) Development of a chromatographic method to resolve 68Ga-PSMA diastereomers from impurities improving the radiochemical purity (RP) determination. 4) Synthesis under GMP and quality control to achieve maximal specific activity (SA) before the use in patients.

Methods The labelling of Glu-NH-CO-NH-Lys(Ahx)-HBED-CC with 68Ga eluted from a 68Ge/68Ga generator (360MBq in 1000µL HCl 0.05M) was optimized varying the amount of the precursor from 0.04 to 1.25µg (0.04-1.32nmol). The reaction pH 4.0 was adjusted with 250µL NaOAc 0.25M. Incubation was done at RT or 100ºC. At different incubation times (1 to 5min), the reaction was stopped by addition of an excess of gallium nitrate in order to study kinetics and stability. RP was controlled by RP-HPLC, testing different solvent gradients and flow. The presence of 68Ga-colloid was analysed by ITLC-SG in MeOH:NH4OAc 1M. The scaling up production of 68Ga-PSMA for patients was done with a microfluidics system inside a shielded laminar flow hood Class A. Post labelling purification was done by SPE-C18.

Results 68Ga was totally bound to the precursor from amounts of 0.32nmol, giving a maximum SA of 1171MBq/nmol. There was evidence that the 68Ga incorporation to the precursor formed two diastereomers (one more lipophilic and the other more hydrophilic), which were isolated with the solvent gradient optimized from the 15 systems tested. A conversion of the more lipophilic diastereomer into the more hydrophilic was observed. The incidence in the diagnosis is still uncertain. The rate of conversion was positively correlated with the temperature and time of incubation as well as with the time elapsed after labelling, so the optimal reaction conditions were achieved at 100ºC for at least 5min. For patient administration, 1131±29MBq of 68Ga and 2,64nmol of precursor were used. A global yield of 90±1% (ndc), a RP of 97±1% and a SA of 418±15MBq/nmol were obtained.

Conclusions Two diastereomers were isolated from the radiochemical impurities and from free 68Ga with the optimized HPLC system. More research is required to determine the molecular characteristics of each diastereomer and their incidence in the diagnosis. The high specific activities required for the clinical application of this radiopharmaceutical demands careful control of the ratio activity/precursor, the time of incubation and the temperature. 68Ga-PSMA with high RP and SA is used clinically at our Centre with excellent results.