Boris D. Zlatopolskiy

Comparison of [18F]DCFPyL and [68Ga]Ga-PSMA-HBED-CC for PSMA-PET Imaging in Patients with Relapsed Prostate Cancer

PurposeGallium-68 (Ga-68)-labeled tracers for imaging expression of the prostate-specific membrane antigen (PSMA) such as the [68Ga]Ga-PSMA-HBED-CC have already demonstrated high potential for the detection of recurrent prostate cancer. However, compared to Ga-68, a labeling with fluorine-18 (F-18) would offer advantages with respect to availability, production amount, and image resolution. [18F]DCFPyL is a promising F-18-labeled candidate for PSMA-positron emission tomography (PET) imaging that has been recently introduced. In the current study, we aimed to compare [68Ga]Ga-PSMA-HBED-CC and [18F]DCFPyL for clinical use in biochemically relapsed prostate cancer.ProceduresIn 14 selected patients with PSA relapse of prostate cancer, [18F]DCFPyL PET/X-ray computed tomography (CT) was performed in addition to [68Ga]Ga-PSMA-HBED-CC PET/CT. A systematic comparison was carried out between results obtained with both tracers with regard to the number of detected PSMA-positive lesions, the standardized uptake value (SUV)max and the lesion to background ratios.ResultsAll suspicious lesions identified by [68Ga]Ga-PSMA-HBED-CC were also detected with [18F]DCFPyL. In three patients, additional lesions were observed using [18F]DCFPyL PET/CT. The mean SUVmax in the concordant [18F]DCFPyL PSMA-positive lesions was significantly higher as compared to [68Ga]Ga-PSMA-HBED-CC (14.5 vs. 12.2, p = 0.028, n = 15). The mean tumor to background ratios (n = 15) were significantly higher for [18F]DCFPyL compared to [68Ga]Ga-PSMA-HBED-CC using kidney, spleen, or parotid as reference organs (p = 0.006, p = 0.002, p = 0.008), but no significant differences were found using the liver (p = 0.167) or the mediastinum (p = 0.363) as reference organs.Conclusion[18F]DCFPyL PET/CT provided a high image quality and visualized small prostate lesions with excellent sensitivity. [18F]DCFPyL represents a highly promising alternative to [68Ga]Ga-PSMA-HBED-CC for PSMA-PET/CT imaging in relapsed prostate cancer.

PSA-stratified performance of 18F- and 68Ga-labeled tracers in PSMA-PET imaging of patients with biochemical recurrence of prostate cancer.

Several studies outlined the sensitivity of 68Ga-labeled PET tracers against the prostate-specific membrane antigen (PSMA) for localization of relapsed prostate cancer in patients with renewed increase in the prostate-specific antigen (PSA), commonly referred to as biochemical recurrence. Labeling of PSMA tracers with 18F offers numerous advantages, including improved image resolution, longer half-life, and increased production yields. The aim of this study was to assess the PSA-stratified performance of the 18F-labeled PSMA tracer 18F-DCFPyL and the 68Ga-labeled reference 68Ga-PSMA-HBED-CC. Methods: We examined 191 consecutive patients with biochemical recurrence according to standard acquisition protocols using 18F-DCFPyL (n = 62, 269.8 MBq, PET scan at 120 min after injection) or 68Ga-PSMA-HBED-CC (n = 129, 158.9 MBq, 60 min after injection). We determined PSA-stratified sensitivity rates for both tracers and corrected our calculations for Gleason scores using iterative matched-pair analyses. As an orthogonal validation, we directly compared tracer distribution patterns in a separate cohort of 25 patients, sequentially examined with both tracers. Results: After prostatectomy (n = 106), the sensitivity of both tracers was significantly associated with absolute PSA levels (P = 4.3 × 10−3). Sensitivity increased abruptly, when PSA values exceeded 0.5 μg/L (P = 2.4 × 10−5). For a PSA less than 3.5 μg/L, most relapses were diagnosed at a still limited stage (P = 3.4 × 10−6). For a PSA of 0.5–3.5 μg/L, PSA-stratified sensitivity was 88% (15/17) for 18F-DCFPyL and 66% (23/35) for 68Ga-PSMA-HBED-CC. This significant difference was preserved in the Gleason-matched-pair analysis. Outside of this range, sensitivity was comparably low (PSA < 0.5 μg/L) or high (PSA > 3.5 μg/L). After radiotherapy (n = 85), tracer sensitivity was largely PSA-independent. In the 25 patients examined with both tracers, distribution patterns of 18F-DCFPyL and 68Ga-PSMA-HBED-CC were strongly comparable (P = 2.71 × 10−8). However, in 36% of the PSMA-positive patients we detected additional lesions on the 18F-DCFPyL scan (P = 3.7 × 10−2). Conclusion: Our data suggest that 18F-DCFPyL is noninferior to 68Ga-PSMA-HBED-CC, while offering the advantages of 18F labeling. Our results indicate that imaging with 18F-DCFPyL may even exhibit improved sensitivity in localizing relapsed tumors after prostatectomy for moderately increased PSA levels. Although the standard acquisition protocols, used for 18F-DCFPyL and 68Ga-PSMA-HBED-CC in this study, stipulate different activity doses and tracer uptake times after injection, our findings provide a promising rationale for validation of 18F-DCFPyL in future prospective trials.

Neither azeotropic drying, nor base nor other additives: a minimalist approach to 18F-labeling

A novel, efficient, time-saving and reliable radiolabeling procedure via nucleophilic substitution with [(18)F]fluoride is described. Different radiolabeled aliphatic and aromatic compounds were prepared in high radiochemical yields simply by heating of quaternary anilinium, diaryliodonium and triarylsulfonium [(18)F]fluorides in suitable solvents. The latter were obtained via direct elution of (18)F(-) from an anion exchange resin with alcoholic solutions of onium precursors. Neither azeotropic evaporation of water, nor a base, nor any other additives like cryptands or crown ethers were necessary. Due to its simplicity this method should be highly suitable for automated radiosyntheses, especially in microfluidic devices.

Radiosynthesis of a new PSMA targeting ligand ([18F]FPy-DUPA-Pep)

Due to the specificity of expression of PSMA (prostate specific membrane antigen) particularly in prostate cancer cells (e.g. LNCaP), numerous PSMA ligands have been synthesized until now. In the current study, we synthesized DUPA-Pep having 2-[3-(1,3-dicarboxypropyl)ureido]pentanedioic acid (DUPA) linked via 8-aminooctanoic acid to two phenylalanine residues and chose 6-[(18)F]fluoronicotinic acid 2,3,5,6-tetrafluorophenyl ester [(18)F]FPy-TFP as a prosthetic group for coupling. [(18)F]FPy-DUPA-Pep was obtained in a radiochemical yield of 48±0.9% (decay uncorrected) within 50 min with a chemical purity of >98%.

123I-ITdU-mediated nanoirradiation of DNA efficiently induces cell kill in HL60 leukemia cells and in doxorubicin-, beta-, or gamma-radiation-resistant cell lines.

Resistance to radiotherapy or chemotherapy is a common cause of treatment failure in high-risk leukemias. We evaluated whether selective nanoirradiation of DNA with Auger electrons emitted by 5-123I-iodo-4′-thio-2′-deoxyuridine (123I-ITdU) can induce cell kill and break resistance to doxorubicin, β-, and γ-irradiation in leukemia cells. Methods: 4′-thio-2′-deoxyuridine was radiolabeled with 123/131I and purified by high-performance liquid chromatography. Cellular uptake, metabolic stability, DNA incorporation of 123I-ITdU, and the effect of the thymidylate synthase (TS) inhibitor 5-fluoro-2′-deoxyuridine (FdUrd) were determined in HL60 leukemia cells. DNA damage was assessed with the comet assay and quantified by the olive tail moment. Apoptosis induction and irradiation-induced apoptosis inhibition by benzoylcarbonyl-Val-Ala-Asp-fluoromethyl ketone (z-VAD.fmk) were analyzed in leukemia cells using flow cytometry analysis. Results: The radiochemical purity of ITdU was 95%. Specific activities were 900 GBq/μmol for 123I-ITdU and 200 GBq/μmol for 131I-ITdU. An in vitro cell metabolism study of 123I-ITdU with wild-type HL60 cells demonstrated an uptake of 1.5% of the initial activity/106 cells of 123I-ITdU. Ninety percent of absorbed activity from 123I-ITdU in HL60 cells was specifically incorporated into DNA. 123I-ITdU caused extensive DNA damage (olive tail moment > 12) and induced more than 90% apoptosis in wild-type HL60 cells. The broad-spectrum inhibitor of caspases zVAD-fmk reduced 123I-ITdU–induced apoptosis from more than 90% to less than 10%, demonstrating that caspases were central for 123I-ITdU–induced cell death. Inhibition of TS with FdUrd increased DNA uptake of 123I-ITdU 18-fold and the efficiency of cell kill about 20-fold. In addition, 123I-ITdU induced comparable apoptotic cell death (>90%) in sensitive parental leukemia cells and in leukemia cells resistant to β-irradiation, γ-irradiation, or doxorubicin at activities of 1.2, 4.1, 12.4, and 41.3 MBq/mL after 72 h. This finding indicates that 123I-ITdU breaks resistance to β-irradiation, γ-irradiation, and doxorubicin in leukemia cells. Conclusion: 123I-ITdU–mediated nanoirradiation of DNA efficiently induced apoptosis in sensitive and resistant leukemia cells against doxorubicin, β-irradiation, and γ-irradiation and may provide a novel treatment strategy for overcoming resistance to conventional radiotherapy or chemotherapy in leukemia. Cellular uptake and cell kill are highly amplified by inhibiting TS with FdUrd.

A Practical One-Pot Synthesis of Positron Emission Tomography (PET) Tracers via Nickel-Mediated Radiofluorination.

Recently a novel method for the preparation of 18F-labeled arenes via oxidative [18F]fluorination of easily accessible and sufficiently stable nickel complexes with [18F]fluoride under exceptionally mild reaction conditions was published. The suitability of this procedure for the routine preparation of clinically relevant positron emission tomography (PET) tracers, 6-[18F]fluorodopamine (6-[18F]FDA), 6-[18F]fluoro-l-DOPA (6-[18F]FDOPA) and 6-[18F]fluoro-m-tyrosine (6-[18F]FMT), was evaluated. The originally published base-free method was inoperative. However, a “low base” protocol afforded protected radiolabeled intermediates in radiochemical conversions (RCCs) of 5–18 %. The subsequent deprotection step proceeded almost quantitatively (>95 %). The simple one-pot two-step procedure allowed the preparation of clinical doses of 6-[18F]FDA and 6-[18F]FDOPA within 50 min (12 and 7 % radiochemical yield, respectively). In an unilateral rat model of Parkinsons disease, 6-[18F]FDOPA with high specific activity (175 GBq μmol−1) prepared using the described nickel-mediated radiofluorination was compared to 6-[18F]FDOPA with low specific activity (30 MBq μmol−1) produced via conventional electrophilic radiofluorination. Unexpectedly both tracer variants displayed very similar in vivo properties with respect to signal-to-noise ratio and brain distribution, and consequently, the quality of the obtained PET images was almost identical.

Synthesis of 18F-labelled β-lactams by using the Kinugasa reaction

Owing to their broad spectrum of biological activities and low toxicity, β-lactams are attractive lead structures for the design of novel molecular probes. However, the synthesis of positron emission tomography (PET)-isotope-labelled β-lactams has not yet been reported. Herein, we describe the simple preparation of radiofluorinated β-lactams by using the fast Kinugasa reaction between (18)F-labelled nitrone [(18)F]-1 and alkynes of different reactivity. Additionally, (18)F-labelled fused β-lactams were obtained through the reaction of a cyclic nitrone 7 with radiofluorinated alkynes [(18)F]-6 a,b. Radiochemical yields of the Kinugasa reaction products could be significantly increased by the use of different Cu(I) ligands, which additionally allowed a reduction in the amount of precursor and/or reaction time. Model radiofluorinated β-lactam-peptide and protein conjugates ([(18)F]-10 and (18)F-labelled BSA conjugate) were efficiently obtained in high yield under mild conditions (aq. MeCN, ambient temperature) within a short reaction time, demonstrating the suitability of the developed method for radiolabelling of sensitive molecules such as biopolymers.

Alcohol-Enhanced Cu-Mediated Radiofluorination

The potential of many 18 F-labeled (hetero)aromatics for applications in positron emission tomography remains underexplored because convenient procedures for their radiosynthesis are lacking. Consequently, simple methods to prepare radiofluorinated (hetero)arenes are highly sought after. Herein, we report the beneficial effect of primary and secondary alcohols on Cu-mediated 18 F-labeling. This observation contradicts the assumption that such alcohols are inappropriate solvents for aromatic fluorination. Therefore, we developed a protocol for rapid radiolabeling of an extraordinarily broad scope of boronic and stannyl substrates under general reaction conditions. Notably, radiofluorinated indoles, phenols, and anilines were synthesized directly from the corresponding unprotected precursors. Furthermore, the novel method enabled the preparation of radiofluorinated tryptophans, [18 F]F-DPA, [18 F]DAA1106, 6-[18 F]FDA, and 6-[18 F]FDOPA.

C-(4-[18F]fluorophenyl)-N-phenyl nitrone: A novel 18F-labeled building block for metal free [3+2]cycloaddition.

Radiofluorination via [3+2]-nitrone-alkene cycloaddition was studied using the model reaction between (18)F-labeled C-(4-fluorophenyl)-N-phenyl nitrone ([(18)F]1) and substituted maleimides 2a-c. [(18)F]1 was prepared in RCY of 73.6±5.8% and radiochemical purity of >95%. Cycloaddition of [(18)F]1 to 2a in toluene at 80°C and in EtOH at 110°C gave the respective isoxazolidine [(18)F]5a in >80% RCY at 10min reaction time. Reaction between [(18)F]1 and 2b, c also went smoothly to afford the respective cycloaddition products in high radiochemical yields.

Preferential tumor targeting and selective tumor cell cytotoxicity of 5-[131/125I]iodo-4'-thio-2'-deoxyuridine.

Purpose: Auger electron emitting radiopharmaceuticals are attractive for targeted nanoirradiation therapy, provided that DNA of malignant cells is selectively addressed. Here, we examine 5-[123/125/131I]iodo-4′-thio-2′-deoxyuridine (ITdU) for targeting DNA in tumor cells in a HL60 xenograft severe combined immunodeficient mouse model. Experimental Design: Thymidine kinase and phosphorylase assays were done to determine phosphorylation and glycosidic bond cleavage of ITdU, respectively. The biodistribution and DNA incorporation of ITdU were determined in severe combined immunodeficient mice bearing HL60 xenografts receiving pretreatment with 5-fluoro-2′-deoxyuridine (FdUrd). Organ tissues were dissected 0.5, 4, and 24 h after radioinjection and uptake of [131I]ITdU (%ID/g tissue) was determined. Cellular distribution of [125I]ITdU was imaged by microautoradiography. Apoptosis and expression of the proliferation marker Ki-67 were determined by immunohistologic staining using corresponding paraffin tissue sections. Results: ITdU is phosphorylated by thymidine kinase 1 and stable toward thymidylate phosphatase-mediated glycosidic bond cleavage. Thymidylate synthase-mediated deiodination of [123/125/131I]ITdU was inhibited with FdUrd. Pretreatment with FdUrd increased preferentially tumor uptake of ITdU resulting in favorable tumor-to-normal tissue ratios and tumor selectivity. ITdU was exclusively localized within the nucleus and incorporated into DNA. In FdUrd-pretreated animals, we found in more than 90% of tumor cells apoptosis induction 24 h postinjection of ITdU, indicating a highly radiotoxic effect in tumor cells but not in cells of major proliferating tissues. Conclusion: ITdU preferentially targets DNA in proliferating tumor cells and leads to apoptosis provided that the thymidylate synthase is inhibited.