Extensive preclinical evaluation of lutetium-177-labeled PSMA-specific tracers for prostate cancer radionuclide therapy

Various radiolabeled prostate-specific membrane antigen (PSMA)–targeting tracers are clinically applied for prostate cancer (PCa) imaging and targeted radionuclide therapy. The PSMA binding affinities, biodistribution, and DNA-damaging capacities of these radiotracers have not yet been compared in detail. A major concern of PSMA-targeting radiotracers is the toxicity in other PSMA-expressing organs, such as the salivary glands, thus demanding careful evaluation of the most optimal and safest radiotracer. In this extensive preclinical study, we evaluated the clinically applied PSMA-targeting small molecule inhibitors DOTA-PSMA-617 (PSMA-617) and DOTAGA-PSMA-I&T (PSMA-I&T) and the PSMA nanobody DOTA-JVZ-007 (JVZ-007) using PSMA-expressing cell lines, a unique set of PCa patient-derived xenografts (PDX) and healthy human tissues. In vitro displacement studies on PSMA-expressing cells and cryosections of a PSMA-positive PDX revealed high and specific binding affinity for all three tracers labeled with lutetium-177 with IC${}_{50}$ values in the nanomolar range. Interestingly, [${}^{177}$Lu]Lu-JVZ-007 could not be displaced by PSMA-617 or PSMA-I&T, suggesting that this tracer targets an alternative binding site. Autoradiography assays on cryosections of human salivary and renal tissues revealed [${}^{177}$Lu]Lu-PSMA-617 to have the lowest binding to these healthy organs compared with [${}^{177}$Lu]Lu-PSMA-I&T. In vivo biodistribution assays confirmed the in vitro results with comparable tumor uptake of [${}^{177}$Lu]Lu-PSMA-617 and [${}^{177}$Lu]Lu-PSMA-I&T at all timepoints, resulting in induction of similar levels of DNA double-strand breaks in the tumors. However, [${}^{177}$Lu]Lu-PSMA-I&T demonstrated approximately 40× higher renal uptake at 4 and 8 h post injection resulting in an unfavorable tumor-to-kidney ratio. [${}^{177}$Lu]Lu-PSMA-617 has the most favorable biodistribution in mice as well as more favorable binding characteristics in vitro in PSMA-positive cells and human kidney and salivary gland specimens compared with [${}^{177}$Lu]Lu-PSMA-I&T and [${}^{177}$Lu]Lu-JVZ-007. Based on our preclinical evaluation, [${}^{177}$Lu]Lu-PSMA-617 is the best performing tracer to be taken further into clinical evaluation for PSMA-targeted radiotherapeutic development although with careful evaluation of the tracer binding to PSMA-expressing organs.