Development of a homotrimeric PSMA radioligand based on the NOTI chelating platform

Abstract

Background: The NOTI chelating scafold can readily be derivatized for bioconjuga‑ tion without impacting its metal complexation/radiolabeling properties making it an attractive building block for the development of multimeric/-valent radiopharma‑ ceuticals. The objective of the study was to further explore the potential of the NOTI chelating platform by preparing and characterizing homotrimeric PSMA radioconju‑ gates in order to identify a suitable candidate for clinical translation. Results: Altogether, three PSMA conjugates based on the NOTI-TVA scafold with dif‑ ferent spacer entities between the chelating unit and the Glu-CO-Lys PSMA bind‑ ing motif were readily prepared by solid phase-peptide chemistry. Cell experiments allowed the identifcation of the homotrimeric conjugate 9 comprising NaI-Amc spacer with high PSMA binding afnity (IC50=5.9 nM) and high PSMA-specifc internalization (17.8±2.5%) compared to the clinically used radiotracer [ 68Ga]Ga-PSMA-11 with a IC50 of 18.5 nM and 5.2±0.2% cell internalization, respectively. All 68Ga-labeled trimeric con‑ jugates showed high metabolic stability in vitro with [ 68Ga]Ga-9 exhibiting high bind‑ ing to human serum proteins (>95%). Small-animal PET imaging revealed a specifc tumor uptake of 16.0±1.3% IA g−1 and a kidney uptake of 67.8±8.4% IA g−1 for [ 68Ga] Ga-9. Clinical PET imaging allowed identifcation of all lesions detected by [ 68Ga]GaPSMA-11 together with a prolonged blood circulation as well as a signifcantly lower kidney and higher liver uptake of [ 68Ga]Ga-9 compared to [ 68Ga]Ga-PSMA-11. Conclusions: Trimerization of the Glu-CO-Lys binding motif for conjugate 9 resulted in a~threefold higher binding afnity and cellular uptake as well as in an altered bio‑ distribution profle compared to the control [ 68Ga]Ga-PSMA-11 due to its intrinsic high binding to serum proteins. To fully elucidate its biodistribution, future studies in combi‑ nation with long-lived radionuclides, such as 64Cu, are warranted. Its prolonged biologi‑ cal half-life and favorable tumor-to-kidney ratio make this homotrimeric conjugate also a potential candidate for future radiotherapeutic applications in combination with therapeutic radionuclides such as 67Cu.