Nadine Colpo

18F-AmBF3-MJ9: A novel radiofluorinated bombesin derivative for prostate cancer imaging

A novel radiofluorinated derivative of bombesin, (18)F-AmBF3-MJ9, was synthesized and evaluated for its potential to image prostate cancer by targeting the gastrin releasing peptide receptor (GRPR). AmBF3-MJ9 was prepared from an ammoniomethyl-trifluoroborate (AmBF3) conjugated alkyne 2 and azidoacetyl-MJ9 via a copper-catalyzed click reaction, and had good binding affinity for GRPR (Ki=0.5±0.1nM). The (18)F-labeling was performed via a facile one-step (18)F-(19)F isotope exchange reaction, and (18)F-AmBF3-MJ9 was obtained in 23±5% (n=3) radiochemical yield in 25min with >99% radiochemical purity and 100±32GBq/μmol specific activity. (18)F-AmBF3-MJ9 was stable in mouse plasma, and was partially (22-30%) internalized after binding to GRPR. Positron emission tomography (PET) imaging and biodistribution studies in mice showed fast renal excretion and good uptake of (18)F-AmBF3-MJ9 by GRPR-expressing pancreas and PC-3 prostate cancer xenografts. Tumor uptake was 1.37±0.25%ID/g at 1h, and 2.20±0.13%ID/g at 2h post-injection (p.i.) with low background uptake and excellent tumor visualization (tumor-to-muscle ratios of 75.4±5.5). These data suggest that (18)F-AmBF3-MJ9 is a promising PET tracer for imaging GRPR-expressing prostate cancers.

18F-Trifluoroborate Derivatives of [Des-Arg10]Kallidin for Imaging Bradykinin B1 Receptor Expression with Positron Emission Tomography

Bradykinin B1 receptor (B1R) is involved in pain and inflammation pathways and is upregulated in inflamed tissues and cancer. Due to its minimal expression in healthy tissues, B1R is an attractive target for the development of therapeutic agents to treat inflammation, chronic pain, and cancer. The goal of this study is to synthesize and compare two (18)F-labeled peptides derived from potent B1R antagonists B9858 and B9958 for imaging B1R expression with positron emission tomography (PET). Azidoacetyl-B9858 2 and azidoacetyl-B9958 3 were synthesized by a solid-phase approach and subsequently clicked to ammoniomethyl-trifluoroborate (AmBF3)-conjugated alkyne 1 to obtain AmBF3-B9858 and AmBF3-B9958, respectively. AmBF3-B9858 and AmBF3-B9958 bound B1R with high affinity, with Ki values at 0.09 ± 0.08 and 0.46 ± 0.03 nM, respectively, as measured by in vitro competition binding assays. (18)F labeling was performed via an (18)F-(19)F isotope exchange reaction. The radiofluorinated tracers were obtained within a synthesis time of 30 min and with 23-32% non-decay-corrected radiochemical yield, >99% radiochemical purity, and 43-87 GBq/μmol specific activity at the end of the synthesis. PET imaging and biodistribution studies were carried out in mice bearing both B1R-positive (B1R(+)) HEK293T::hB1R and B1R-negative (B1R(-)) HEK293T tumors. Both tracers cleared rapidly from most organs/tissues, mainly through the renal pathway. High uptake in B1R(+) tumors ((18)F-AmBF3-B9858: 3.94 ± 1.24% ID/g, tumor-to-muscle ratio 21.3 ± 4.33; (18)F-AmBF3-B9958: 4.20 ± 0.98% ID/g, tumor-to-muscle ratio 48.6 ± 10.7) was observed at 1 h postinjection. These results indicate that (18)F-AmBF3-B9858 and (18)F-AmBF3-B9958 are promising agents for the in vivo imaging of B1R expression with PET.

Cross-Linked Polyethylene Glycol Beads to Separate 99mTc-Pertechnetate from Low-Specific-Activity Molybdenum

We report a kit-based approach for the purification of sodium pertechnetate (99mTcO4−) from solutions with high MoO42− content. Methods: Cross-linked polyethylene glycol resins (ChemMatrix) were used to separate 99mTc and molybdenum in 4N NaOH. The resins were loaded at various flow rates and eluted with water to release 99mTc. The 99mTc solution was passed through a cation exchange resin and an alumina cartridge, followed by saline elution. This process was tested with cyclotron-produced 99mTc using an automated system and disposable kits. Results: Optimal results were obtained by loading 500 mg of resin at flow rates of up to 3.1 mL/min, with quantitative extraction of 99mTc from the molybdate solution and complete release of 99mTc after elution with water. The automated system was highly efficient at isolating Na99mTcO4 within minutes, with a recovery rate of 92.7% ± 1.1% (mean ± SD) using cyclotron-produced 99mTc. Conclusion: ChemMatrix resins were highly effective at separating 99mTcO4− from molybdate solutions.

Design, synthesis and evaluation of 18F-labeled cationic carbonic anhydrase IX inhibitors for PET imaging

Abstract Carbonic anhydrase IX (CA-IX) is a marker for tumor hypoxia, and its expression is negatively correlated with patient survival. CA-IX represents a potential target for eliminating hypoxic cancers. We synthesized fluorinated cationic sulfonamide inhibitors 1–3 designed to target CA-IX. The binding affinity for CA-IX ranged from 0.22 to 0.96 μM. We evaluated compound 2 as a diagnostic PET imaging agent. Compound 2 was radiolabeled with 18F in 10 ± 4% decay-corrected radiochemical yield with 85.1 ± 70.3 GBq/μmol specific activity and >98% radiochemical purity. 18F-labeled 2 was stable in mouse plasma at 37 °C after 1 h incubation. PET/CT imaging was conducted at 1 h post-injection in a human colorectal cancer xenograft model. 18F-labeled 2 cleared through hepatobiliary and renal pathways. Tumor uptake was approximately 0.41 ± 0.06% ID/g, with a tumor-to-muscle ratio of 1.99 ± 0.25. Subsequently, tumor xenografts were visualized with moderate contrast. This study demonstrates the use of a cationic motif for conferring isoform selectively for CA-IX imaging agents.

Imaging Bradykinin B1 Receptor with 68Ga-Labeled [des-Arg10]Kallidin Derivatives: Effect of the Linker on Biodistribution and Tumor Uptake

Bradykinin B1 receptor (B1R) that is overexpressed in cancers but minimally expressed in normal healthy tissues represents an attractive biomarker for the development of cancer imaging agents. The goal of this study was to evaluate the effect of different linkers on the pharmacokinetics and tumor uptake of a B1R-targeting radio-peptide sequence, 68Ga-DOTA-linker-Lys-Arg-Pro-Hyp-Gly-Cha-Ser-Pro-Leu. Four peptides, SH01078, P03034, P04115, and P04168, with 6-aminohexanoic acid, 9-amino-4,7-dioxanonanoic acid, Gly-Gly, and 4-amino-(1-carboxymethyl)piperidine, respectively, as the linker were synthesized and evaluated. In vitro competition binding assays showed that the Ki values of SH01078, P03034, P04115, and P04168 were 27.8±4.9, 16.0±1.9, 11.4±2.5, and 3.6±0.2 nM, respectively. Imaging and biodistribution studies were performed in mice bearing both B1R-positive HEK293T::hB1R and B1R-negative HEK293T tumors. All tracers showed mainly renal excretion with excellent tumor visualization and minimal background activity except for kidneys and bladder. The average uptake of 68Ga-labeled SH01078, P03034, and P04115 in HEK293T::hB1R tumor was similar (1.96-2.17%ID/g) at 1 h postinjection. 68Ga-P04168 generated higher HEK293T::hB1R tumor uptake (4.15±1.13%ID/g) and lower background activity, leading to a >2-fold improvement in HEK293T::hB1R tumor-to-background (HEK293T tumor, blood, muscle, and liver) contrasts over those of 68Ga-labeled SH01078, P03034, and P04115. Our results indicate that the choice of linker affects binding affinity, pharmacokinetics, and tumor targeting. The use of the cationic 4-amino-(1-carboxymethyl)piperidine linker improved tumor visualization, and the resulting 68Ga-P04168 might be promising for clinical application for imaging B1R-expressing tumors with positron emission tomography.

One-step synthesis of 4-[18F]fluorobenzyltriphenylphosphonium cation for imaging with positron emission tomography

4-[(18) F]Fluorobenzyltriphenylphosphonium cation ((18) F-FBnTP) is a promising negative membrane potential targeting positron emission tomography tracer. However, the reported multistep radiolabeling approach for the synthesis of (18) F-FBnTP poses a challenge for routine clinical applications. In this study, we demonstrated that (18) F-FBnTP can be prepared in good conversion yields (~60%, nondecay corrected) in just one step via a copper-mediated (18) F-fluorination reaction using a pinacolyl arylboronate precursor. In addition, our data suggest that (18) F-labeled (phosphonium) cations can be efficiently prepared via a copper-mediated (18) F-fluoronation by using triflate as the counterion.

Comparative Studies of Three 68Ga-Labeled [Des-Arg10]Kallidin Derivatives for Imaging Bradykinin B1 Receptor Expression with PET

Bradykinin B1 receptor (B1R) is a G-protein–coupled receptor that is overexpressed in a variety of cancers. B1R is not expressed in healthy tissues, making it an attractive cancer imaging marker. Previously, we reported selective uptake of 68Ga-P03034 (68Ga-DOTA-dPEG2-Lys-Arg-Pro-Hyp-Gly-Cha-Ser-Pro-Leu) in B1R-positive (B1R+) HEK293T::hB1R tumor xenografts in mice. In this study, we compare 68Ga-P03034 with 68Ga-labeled P04158 (68Ga-DOTA-dPEG2-Lys-Lys-Arg-Pro-Hyp-Gly-Igl-Ser-D-Igl-Oic) and Z02090 (68Ga-DOTA-dPEG2-Lys-Lys-Arg-Pro-Hyp-Gly-Cpg-Ser-D-Tic-Cpg) derived from 2 potent B1R antagonists, B9858 and B9958, respectively, for imaging B1R expression with PET. Methods: Peptide sequences were assembled on solid-phase. Cold standards were prepared by incubating DOTA-conjugated peptides with GaCl3. Binding affinity was measured via competition binding assays using hB1R-expressing Chinese hamster ovary-K1 cell membranes. 68Ga labeling was performed in N-(2-hydroxyethyl)piperazine-N′-(2-ethanesulfonic acid) buffer with microwave heating and purified by high-performance liquid chromatography. Imaging/biodistribution studies were performed in mice bearing wild-type HEK293T (B1R−) and B1R+ HEK293T::hB1R tumors. Results: P03034, P04158, and Z02090 bound B1R with high affinity, with Ki values at 16.0 ± 2.9, 1.5 ± 1.9, and 1.1 ± 0.8 nM, respectively. 68Ga-labeled P03034, P04159, and Z02090 were obtained in greater than 50% decay-corrected radiochemical yields with more than 99% radiochemical purity. Biodistribution studies showed that all three 68Ga-labeled tracers cleared rapidly from the blood and normal tissues, with excretion mainly via the renal pathway. At 1 h after injection, only the kidneys, bladders, and B1R+ HEK293T::hB1R tumors were clearly visualized in PET images. Uptake values of 68Ga-labeled P03034, P04158, and Z02090 in B1R+ tumors were 2.17 ± 0.49, 19.6 ± 4.50, and 14.4 ± 1.63 percentage injected dose per gram, respectively. Uptake ratios of B1R+ to B1R− tumor, blood, and muscle were 6.23 ± 1.69, 5.72 ± 2.20, and 25.5 ± 13.1 for 68Ga-P03034; 34.5 ± 10.5, 19.2 ± 8.21, and 66.1 ± 17.0 for 68Ga-P04158; and 29.3 ± 9.68, 29.9 ± 5.58, and 124 ± 28.1 for 68Ga-Z02090, respectively. Conclusion: All three 68Ga-labeled B1R-targeting peptides generated specific and high-contrasted images of B1R+ tumors xenografted in mice. With significantly higher tumor uptake and target-to-nontarget ratios, 68Ga-labeled P04158 and Z02090 are superior to P03034 for imaging B1R expression with PET.

Preclinical Melanoma Imaging with 68Ga-Labeled α-Melanocyte-Stimulating Hormone Derivatives Using PET

It is estimated that melanoma accounted for 76,380 new cases and 10,130 deaths in the United States in 2016. The melanocortin 1 receptor (MC1R) is highly expressed in the vast majority of melanomas, which makes it an attractive target for molecular imaging and radionuclide therapy. Lactam bridge-cyclized α-melanocyte-stimulating hormone (Ac-Nle4-cyclo[Asp5-His-D-Phe7-Arg-Trp-Lys10]-NH2, or Nle-CycMSHhex) analogues have been successfully developed and studied for MC1R-targeted imaging, predominantly with single-photon emission computed tomography (SPECT). The goal of this study was to design and evaluate novel peptides for melanoma imaging with positron emission tomography (PET). We designed and synthesized three peptides, DOTA-PEG2-Nle-CycMSHhex (CCZ01047), DOTA-4-amino-(1-carboxymethyl) piperidine (Pip)-Nle-CycMSHhex (CCZ01048), and DOTA-Pip-Pip-Nle-CycMSHhex (CCZ01056). All three peptides exhibited high binding affinity to MC1R with sub-nanomolar Ki values, rapid internalization into B16F10 melanoma cells and high in vivo stability with more than 93% remaining intact at 15 min post-injection (p.i.) in blood plasma. All three 68Ga-labeled tracers produced high contrast PET images in C57BL/6J mice bearing B16F10 tumors, and their respective tumor uptakes were 8.0 ± 3.0, 12.3 ± 3.3, and 6.5 ± 1.4 %ID/g at 1 h p.i. Minimal normal organ activity was observed at 1 h p.i., except for kidneys (5.1 ± 1.4, 4.7 ± 0.5, and 6.2 ± 2.0 %ID/g, respectively), and thyroid (4.1 ± 0.6 %ID/g for CCZ01047 and 2.4 ± 0.6 %ID/g for CCZ01048). Due to high accumulation at tumor sites and rapid background clearance of 68Ga-CCZ01048, we further evaluated it at 2 h p.i., and a tumor uptake of 21.9 ± 4.6 %ID/g was observed, with background activity further decreased. Exceptional image contrast was also achieved, i.e. tumor-to-blood, tumor-to-muscle, tumor-to-bone and tumor-to-kidney ratios were 96.4 ± 13.9, 210.9 ± 20.9, 39.6 ± 11.9 and 4.0 ± 0.9, respectively. A blocking study was also performed by co-injection of excess amount of non-radioactive Ga-coupled of CCZ01048, which confirmed that the tumor uptake was MC1R mediated. In conclusion, the introduction of a cationic Pip linker to Nle-CycMSHhex, CCZ01048, not only improved tumor uptake, but also generated high tumor-to-normal tissue contrast with PET imaging in a preclinical melanoma model. Therefore, CCZ01048 is a promising candidate for PET imaging of melanoma, and potentially as a theranostic agent for radionuclide therapy of melanoma when labeled with α or β emitters.

Synthesis and evaluation of 18F-labeled CJ-042794 for imaging prostanoid EP4 receptor expression in cancer with positron emission tomography

The potent and selective prostanoid EP4 receptor antagonist CJ-042794 was radiolabeled with 18F, and evaluated for imaging EP4 receptor expression in cancer with positron emission tomography (PET). The fluorination precursor, arylboronic acid pinacol ester 4, was prepared in 4 steps with 42% overall yield. 18F-CJ-042794 was synthesized via a copper-mediated 18F-fluorination reaction followed by base hydrolysis, and was obtained in 1.5±1.1% (n=2) decay-corrected radiochemical yield. PET/CT imaging and biodistribution studies in mice showed that 18F-CJ-042794 was excreted through both renal and hepatobiliary pathways with significant retention in blood. The EP4-receptor-expressing LNCaP prostate cancer xenografts were clearly visualized in PET images with 1.12±0.08%ID/g (n=5) uptake value and moderate tumour-to-muscle contrast ratio (2.73±0.22) at 1h post-injection. However, the tumour uptake was nonspecific as it could not be blocked by co-injection of cold standard, precluding the application of 18F-CJ-042794 for PET imaging of EP4 receptor expression in cancer.

Synthesis and evaluation of (18)F-labeled 4-nitrobenzyl derivatives for imaging tumor hypoxia with positron emission tomography: Comparison of 2-[(18)F]fluoroethyl carbonate and 2-[(18)F]fluoroethyl carbamate.

Two 4-nitrobenzyl derivatives, 2-fluoroethyl 4-nitrobenzyl carbonate 1 and 4-nitrobenzyl N-2-fluoroethyl carbamate 2, were radiolabeled with (18)F and evaluated for imaging tumor hypoxia with positron emission tomography. Although good tumor uptake was observed for [(18)F]1 and [(18)F]2 (>2.5%ID/g at 3-h post-injection), the tracers cleared slowly from nontarget tissues (>1.5%ID/g) and exhibited extensive defluorination in vivo (>4.0%ID/g for bone). Therefore, [(18)F]1 and [(18)F]2 are not suitable for imaging tumor hypoxia due to suboptimal tumor-to-background contrasts.