On the basis of the high and consistent expression of prostatespecific membrane antigen (PSMA) in metastatic prostate cancer (PC), the goal of this study was the development, preclinical evaluation, and first proof-of-concept investigation of a PSMA inhibitor for imaging and therapy (PSMA I&T) for 68 Ga-based PET and 177 Lu-based endoradiotherapeutic treatment in patients with metastatic and castration-resistant disease. Methods: PSMA I&T was synthesized in a combined solid phase and solution chemistry strategy. The PSMA affinity of nat Ga-/ nat Lu-PSMA I&T was determined in a competitive binding assay using LNCaP cells. Internalization kinetics of 68 Ga-and 177 Lu-PSMA I&T were investigated using the same cell line, and biodistribution studies were performed in LNCaP tumor-bearing CD-1 nu/nu mice. Initial human PET imaging studies using 68 Ga-PSMA I&T, as well as endoradiotherapeutic treatment of 2 patients with metastatic PC using 177 Lu-PSMA I&T, were performed. Results: PSMA I&T and its cold gallium and lutetium analog revealed nanomolar affinity toward PSMA. The DOTAGA (1,4,7,10-tetraazacyclododecane-1-(glutamic acid)-4,7,10-triacetic acid) conjugate PSMA I&T allowed fast and high-yield labeling with 68 Ga III and 177 Lu III . Uptake of 68 Ga-/ 177 Lu-PSMA I&T in LNCaP tumor cells is highly efficient and PSMA-specific, as demonstrated by competition studies both in vitro and in vivo. Tumor targeting and tracer kinetics in vivo were fast, with the highest uptake in tumor xenografts and kidneys (both PSMA-specific). First-in-human 68 Ga-PSMA I&T PET imaging allowed high-contrast detection of bone lesions, lymph node, and liver metastases. Endoradiotherapy with 177 Lu-PSMA I&T in 2 patients was found to be effective and safe with no detectable side effects. Conclusion: 68 Ga-PSMA I&T shows potential for high-contrast PET imaging of metastatic PC, whereas its 177 Lu-labeled counterpart exhibits suitable targeting and retention characteristics for successful endoradiotherapeutic treatment. Prospective studies on larger cohorts of patients are warranted and planned.

BackgroundDue to its high expression in prostate cancer, PSMA (prostate-specific membrane antigen) represents an ideal target for both diagnostic imaging and endoradiotherapeutic approaches. Based on a previously published highly specific PSMA ligand ([68Ga]DOTA-FFK(Sub-KuE)), we developed a corresponding metabolically stable 1,4,7,10-tetraazacyclododececane,1-(glutaric acid)-4,7,10-triacetic acid (DOTAGA) construct for theranostic treatment of prostate cancer.MethodsAll ligands were synthesized by a combined solid phase and solution phase synthesis strategy. The affinity of the natgallium and lutetium complexes to PSMA and the internalization efficiency of the radiotracers were determined on PSMA-expressing LNCaP cells. The 68Ga- and 177Lu-labelled ligands were further investigated for lipophilicity, binding specificity, metabolic stability, as well as biodistribution and μPET in LNCaP-tumour-bearing mice.ResultsRadiochemical yields for 68Ga (3 nmol, 5.0 M NaCl/2.7 M HEPES (approximately 5/1), pH 3.5 to 4.5, 5 min, 95°C) and 177Lu labelling (0.7 nmol, 0.1 M NH4OAc, pH 5.5, 30 min, 95°C) were almost quantitative, resulting in specific activities of 250 to 300 GBq/μmol for the 68Ga analogues and 38 GBq/μmol for 177Lu complexes. Due to metabolic instability of l-amino acid spacers, d-amino acids were implemented resulting in a metabolically stable DOTAGA ligand. Compared to the DOTA ligand, the DOTAGA derivatives showed higher hydrophilicity (logP = −3.6 ± 0.1 and −3.9 ± 0.1 for 68Ga and 177Lu, respectively) and improved affinity to PSMA resulting in an about twofold increased specific internalization of the 68Ga- and 177Lu-labelled DOTAGA analogue. Especially, [68Ga]DOTAGA-ffk(Sub-KuE) exhibits favourable pharmacokinetics, low unspecific uptake and high tumour accumulation in LNCaP-tumour-bearing mice.ConclusionsThe pair of diagnostic/therapeutic PSMA-ligands [68Ga/177Lu]DOTAGA-ffk(Sub-KuE) possess remarkable potential for the management of prostate cancer.

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Due to its 3 carbonic acid groups being available for bioconjugation, the TRAP chelator (1,4,7-triazacyclononane-1,4,7-tris(methylene(2-carboxyethylphosphinic acid))) is chosen for the synthesis of trimeric bioconjugates for radiolabelling. We optimized a protocol for bio-orthogonal TRAP conjugation via Cu(I)-catalyzed Huisgen-cycloaddition of terminal azides and alkynes (CuAAC), including a detailed investigation of kinetic properties of Cu(II)-TRAP complexes. TRAP building blocks for CuAAC, TRAP(alkyne)3 and TRAP(azide)3 were obtained by amide coupling of propargylamine/3-azidopropyl-1-amine, respectively. For Cu(II) complexes of neat and triply amide-functionalized TRAP, the equilibrium properties as well as pseudo-first-order Cu(II)-transchelation, using 10 to 30 eq. of NOTA and EDTA, were studied by UV-spectrophotometry. Dissociation of any Cu(II)-TRAP species was found to be independent on the nature or excess of a competing chelator, confirming a proton-driven two-step mechanism. The respective thermodynamic stability constants (log K(ML): 19.1 and 17.6) and dissociation rates (k: 38 × 10(-6) and 7 × 10(-6) s(-1), 298 K, pH 4) show that the Cu(II) complex of the TRAP-conjugate possesses lower thermodynamic stability but higher kinetic inertness. At pH 2-3, its demetallation with NOTA was complete within several hours/days at room temperature, respectively, enabling facile Cu(II) removal after click coupling by direct addition of NOTA trihydrochloride to the CuAAC reaction mixture. Notwithstanding this, an extrapolated dissociation half life of >100 h at 37 °C and pH 7 confirms the suitability of TRAP-bioconjugates for application in Cu-64 PET (cf. t(1/2)(Cu-64) = 12.7 h). To showcase advantages of the method, TRAP(DUPA-Pep)3, a trimer of the PSMA inhibitor DUPA-Pep, was synthesized using 1 eq. TRAP(alkyne)3, 3.3 eq. DUPA-Pep-azide, 10 eq. Na ascorbate, and 1.2 eq. Cu(II)-acetate. Its PSMA affinity (IC50), determined by the competition assay on LNCaP cells, was 18-times higher than that of the corresponding DOTAGA monomer (IC50: 2 ± 0.1 vs. 36 ± 4 nM), resulting in markedly improved contrast in Ga-68-PET imaging. In conclusion, the kinetic inertness profile of Cu(II)-TRAP conjugates allows for simple Cu(II) removal after click functionalisation by means of transchelation, but also confirms their suitability for Cu-64-PET as demonstrated previously (Dalton Trans., 2012, 41, 13803).

Prostate-specific membrane antigen (PSMA) is a promising target for diagnosis and treatment of prostate cancer. EuK-Subkff-68Ga-DOTAGA (68Ga-PSMA Imaging & Therapy [PSMA I&T]) is a recently introduced PET tracer for imaging PSMA expression in vivo. Whole-body distribution and radiation dosimetry of this new probe were evaluated. Methods Five patients with a history of prostate cancer were injected intravenously with 91–148 MBq of 68Ga-PSMA I&T (mean ± SD, 128 ± 23 MBq). After an initial series of rapid whole-body scans, 3 static whole-body scans were acquired at 1, 2, and 4 h after tracer injection. Time-dependent changes of the injected activity per organ were determined. Mean organ-absorbed doses and effective doses were calculated using OLINDA/EXM. Results Injection of 150 MBq of 68Ga-PSMA I&T resulted in an effective dose of 3.0 mSv. The kidneys were the critical organ (33 mGy), followed by the urinary bladder wall and spleen (10 mGy each), salivary glands (9 mGy each), and liver (7 mGy). Conclusion 68Ga-PSMA I&T exhibits a favorable dosimetry, delivering organ doses that are comparable to (kidneys) or lower than those delivered by 18F-FDG.

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Zusammenfassung Ziel: In diesem Beitrag wird ein ?berblick ?ber den aktuellen Stand der radiopharmazeutischen Forschung in Bezug auf die Entwicklung von Radiopharmaka f?r die Diagnostik und Therapie des Prostatakarzinoms (PCa) gegeben. Material und Methode: Hierzu wurden die j?ngsten Entwicklungen zusammengestellt und mit besonderem Augenmerk auf die klinische Anwendung am Patienten bewertet und kommentiert. Ergebnisse: Zahlreiche Radiopharmaka, die seit einiger Zeit f?r die klinische Positronenemissionstomografie (PET) und Single-Photonen-Emissionstomografie (SPECT) Diagnostik des PCa eingesetzt werden, adressieren PCa-assoziierte metabolische Prozesse mit einer mehr oder minder ausgepr?gten Spezifit?t. Beispiele f?r diese Gruppe von Radiopharmaka sind [18F]Fluorid, [11C]Acetat, [18F]Fluoroethylcholin ([18F]FEC), [18F]Fluoromethylcholin ([18F]FMC) und [11C]Cholin. Einen neuen Tracer dieses Typs stellt die k?nstliche Aminos?ure anti-1-amino-3-[18F]fluorocyclobutan-1-carbons?ure ([18F]FACBC) dar. Hierneben werden vornehmlich spezifische Rezeptorliganden, wie 16?-[18F]fluoro-5?-dihydrotestosteron ([18F]FDHT), radiomarkierte GRPR-Liganden (Gastrin Releasing Peptide Rezeptor, BB2) und radiomarkierte Antik?rper gegen das Prostata-spezifische Membranantigen (PSMA, NAALADase I oder Glutamat-Carboxypeptidase II) sowie PSMA-spezifische Inhibitoren untersucht. F?r die Therapie des PCa kommen in j?ngster Zeit PSMA-gerichtete Antik?rper und Inhibitoren zum Einsatz, w?hrend f?r die Radionuklidtherapie oss?rer Metastasen in 2014 [223Ra]RaCl2 (Xofigo) die FDA- und EMA-Zulassung erhielt. Schlussfolgerungen: W?hrend einige ?metabolische? Radiopharmaka nur m??igen Erfolg in klinischen Studien zeigten, stehen mit den j?ngst entwickelten PSMA-Inhibitoren, PSMA-bindenden h?hermolekularen Radiopharmaka (Antik?rper und deren Fragmente) und GRPR-Liganden besonders aussichtsreiche neue Tracer-Klassen f?r die Diagnostik und Radionuklidtherapie des PCa zur Verf?gung. Trotz der bisher herausragenden Ergebnisse mit PSMA-Inhibitoren werden gro? angelegte Vergleichsstudien eine vergleichende Bewertung mit GRPR-Liganden erlauben.

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