Couple-close construction of polycyclic rings from diradicals.

Heteroarenes are ubiquitous motifs in bioactive molecules, conferring favourable physical properties when compared to their arene counterparts1-3. In particular, semisaturated heteroarenes possess attractive solubility properties and a higher fraction of sp3 carbons, which can improve binding affinity and specificity. However, these desirable structures remain rare owing to limitations in current synthetic methods4-6. Indeed, semisaturated heterocycles are laboriously prepared by means of non-modular fit-for-purpose syntheses, which decrease throughput, limit chemical diversity and preclude their inclusion in many hit-to-lead campaigns7-10. Herein, we describe a more intuitive and modular couple-close approach to build semisaturated ring systems from dual radical precursors. This platform merges metallaphotoredox C(sp2)-C(sp3) cross-coupling with intramolecular Minisci-type radical cyclization to fuse abundant heteroaryl halides with simple bifunctional feedstocks, which serve as the diradical synthons, to rapidly assemble a variety of spirocyclic, bridged and substituted saturated ring types that would be extremely difficult to make by conventional methods. The broad availability of the requisite feedstock materials allows sampling of regions of underexplored chemical space. Reagent-controlled radical generation leads to a highly regioselective and stereospecific annulation that can be used for the late-stage functionalization of pharmaceutical scaffolds, replacing lengthy de novo syntheses.

Catalytic Activation of Bioorthogonal Chemistry with Light (CABL) Enables Rapid, Spatiotemporally Controlled Labeling and No-Wash, Subcellular 3D-Patterning in Live Cells Using Long Wavelength Light.

Described is the spatiotemporally controlled labeling and patterning of biomolecules in live cells through the catalytic activation of bioorthogonal chemistry with light, referred to as "CABL". Here, an unreactive dihydrotetrazine (DHTz) is photocatalytically oxidized in the intracellular environment by ambient O2 to produce a tetrazine that immediately reacts with a trans-cyclooctene (TCO) dienophile. 6-(2-Pyridyl)dihydrotetrazine-3-carboxamides were developed as stable, cell permeable DHTz reagents that upon oxidation produce the most reactive tetrazines ever used in live cells with Diels-Alder kinetics exceeding k2 of 106 M-1 s-1. CABL photocatalysts are based on fluorescein or silarhodamine dyes with activation at 470 or 660 nm. Strategies for limiting extracellular production of singlet oxygen are described that increase the cytocompatibility of photocatalysis. The HaloTag self-labeling platform was used to introduce DHTz tags to proteins localized in the nucleus, mitochondria, actin, or cytoplasm, and high-yielding subcellular activation and labeling with a TCO-fluorophore were demonstrated. CABL is light-dose dependent, and two-photon excitation promotes CABL at the suborganelle level to selectively pattern live cells under no-wash conditions. CABL was also applied to spatially resolved live-cell labeling of an endogenous protein target by using TIRF microscopy to selectively activate intracellular monoacylglycerol lipase tagged with DHTz-labeled small molecule covalent inhibitor. Beyond spatiotemporally controlled labeling, CABL also improves the efficiency of "ordinary" tetrazine ligations by rescuing the reactivity of commonly used 3-aryl-6-methyltetrazine reporters that become partially reduced to DHTzs inside cells. The spatiotemporal control and fast rates of photoactivation and labeling of CABL should enable a range of biomolecular labeling applications in living systems.

Tunable and Practical Homogeneous Organic Reductants for Cross-Electrophile Coupling.

The syntheses of four new tunable homogeneous organic reductants based on a tetraaminoethylene scaffold are reported. The new reductants have enhanced air stability compared to current homogeneous reductants for metal-mediated reductive transformations, such as cross-electrophile coupling (XEC), and are solids at room temperature. In particular, the weakest reductant is indefinitely stable in air and has a reduction potential of -0.85 V versus ferrocene, which is significantly milder than conventional reductants used in XEC. All of the new reductants can facilitate C(sp2)-C(sp3) Ni-catalyzed XEC reactions and are compatible with complex substrates that are relevant to medicinal chemistry. The reductants span a range of nearly 0.5 V in reduction potential, which allows for control over the rate of electron transfer events in XEC. Specifically, we report a new strategy for controlled alkyl radical generation in Ni-catalyzed C(sp2)-C(sp3) XEC. The key to our approach is to tune the rate of alkyl radical generation from Katritzky salts, which liberate alkyl radicals upon single electron reduction, by varying the redox potentials of the reductant and Katritzky salt utilized in catalysis. Using our method, we perform XEC reactions between benzylic Katritzky salts and aryl halides. The method tolerates a variety of functional groups, some of which are particularly challenging for most XEC transformations. Overall, we expect that our new reductants will both replace conventional homogeneous reductants in current reductive transformations due to their stability and relatively facile synthesis and lead to the development of novel synthetic methods due to their tunability.

Chemiluminescent probe for the detection of inverse electron demand Diels-Alder reaction between tetrazine and trans-Cyclooctene.

A chemiluminescent probe has been developed, consisting of phenoxy-dioxetane moiety covalently attached to trans-cyclooctene. The inverse electron demand Diels-Alder reaction with tetrazine produces a cycloaddition product which undergoes a series of spontaneous rearrangements resulting in emission of green light. The chemiluminescent probe can be applied to study bioconjugation chemistry with tetrazine-modified biomaterials, which have recently been shown to have great potential for anticancer drug delivery. This work describes in vitro studies, including NMR and spectroscopic investigation of chemiluminescence, which will pave way for future in vivo bioconjugation experiments.

Comparison of phosphonate, hydroxypropyl and carboxylate pendants in Fe(III) macrocyclic complexes as MRI contrast agents.

Fe(III) macrocyclic complexes containing a macrocycle and three pendant groups including phosphonate (NOTP =1,4,7-triazacyclononane-1,4,7-triyl-tris(methylenephosphonic acid), carboxylate (NOTA = 1,4,7 - triazacyclononane - N,N',N″ - triacetate) or hydroxypropyl (NOHP =(2S,2'S,2"S)-1,1',1″-(1,4,7-triazonane-1,4,7-triyl)tris(propan-2-ol)) were studied in order to compare the effect of these donor groups on solution chemistry and water proton relaxivity. All three complexes, Fe(NOTP), Fe(NOHP) and Fe(NOTA), display a large degree of kinetic inertness to dissociation in the presence of phosphate and carbonate, under acidic conditions of 100 mM HCl or 1 M HCl or to trans-metalation with Zn(II). The r1 proton relaxivity of the complexes at 1.4 T, 33 °C is compared over the pH range of 1 to 10. At pH 7.4, 33 °C, 1.4 T, Fe(NOHP) has the largest relaxivity (1.5 mM-1 s-1), Fe(NOTP) is second at 1.0 mM-1 s-1, whereas Fe(NOTA) is the lowest at 0.61 mM-1 s-1. Fe(NOTP), Fe(NOHP) and Fe(NOTA) all show an increase in relaxivity at very acidic pH values (< 3) that is consistent with an acid-catalyzed process. Variable temperature 17O NMR studies at near neutral pH are consistent with the absence of an inner-sphere water molecule for Fe(NOTP) and Fe(NOHP), supporting second-sphere or outer-sphere water contributions to proton relaxation. Fe(NOTP) shows contrast enhancement in T1 weighted MRI studies in mice and clears through a renal pathway.

Synthesis, Radiolabeling, and Preliminary in vivo Evaluation of [68Ga] IPCAT-NOTA as an Imaging Agent for Dopamine Transporter.

INTRODUCTION: Novel radiotracer development for imaging dopamine transporters is a subject of interest because although [99mTc]TRODAT-1, [123I]ß-CIT, and [123I]FP-CIT are commercially available; 99Mo/99mTc generator is in short supply and 123I production is highly dependent on compact cyclotron. Therefore, we designed a novel positron emission tomography (PET) tracer based on a tropane derivative through C-2 modification to conjugate NOTA for chelating 68Ga, a radioisotope derived from a 68Ge/68Ga generator. METHODS: IPCAT-NOTA 22 was synthesized and labeled with [68Ga]GaCl4 - at room temperature. Biological studies on serum stability, LogP, and in vitro autoradiography (binding assay and competitive assay) were performed. Furthermore, ex vivo autoradiography, biodistribution, and dynamic PET imaging studies were performed in Sprague Dawley rats. RESULTS: [68Ga]IPCAT-NOTA 24 obtained had a radiochemical yield of ≥90% and a specific activity of 4.25 MBq/nmol. [68Ga]IPCAT-NOTA 24 of 85% radiochemical purity (RCP%) was stable at 37°C for up to 60 minutes in serum with a lipophilicity of 0.88. The specific binding ratio (SBR%) reached 15.8 ± 6.7 at 60 minutes, and the 85% specific uptake could be blocked through co-injection at 100- and 1000-fold of the cold precursor in in vitro binding studies. Tissue regional distribution studies in rats with [68Ga]IPCAT-NOTA 24 showed striatal uptake (0.02% at 5 minutes and 0.007% at 60 minutes) with SBR% of 6%, 25%, and 62% at 5-15, 30-40, and 60-70 minutes, respectively, in NanoPET studies. The RCP% of [68Ga]IPCAT-NOTA 24 at 30 minutes in vivo remained 67.65%. CONCLUSION: Data described here provide new information on the design of PET probe of conjugate/pendent approach for DAT imaging. Another chelator or another direct method of intracranial injection must be used to prove the relation between [68Ga]IPCAT-NOTA 24 uptake and transporter localization.

Seeking Illumination: The Path to Chemiluminescent 1,2-Dioxetanes for Quantitative Measurements and In Vivo Imaging.

Chemiluminescence is a fascinating phenomenon that evolved in nature and has been harnessed by chemists in diverse ways to improve life. This Account tells the story of our research group's efforts to formulate and manifest spiroadamantane 1,2-dioxetanes with triggerable chemiluminescence for imaging and monitoring important reactive analytes in living cells, animals, and human clinical samples. Analytes like reactive sulfur, oxygen and nitrogen species, as well as pH and hypoxia can be indicators of cellular function or dysfunction and are often implicated in the causes and effects of disease. We begin with a foundation in binding-based and activity-based fluorescence imaging that has provided transformative tools for understanding biological systems. The intense light sources required for fluorescence excitation, however, introduce autofluorescence and light scattering that reduces sensitivity and complicates in vivo imaging. Our work and the work of our collaborators were the first to demonstrate that spiroadamantane 1,2-dioxetanes had sufficient brightness and biological compatibility for in vivo imaging of enzyme activity and reactive analytes like hydrogen sulfide (H2S) inside of living mice. This launched an era of renewed interest in 1,2-dioxetanes that has resulted in a plethora of new chemiluminescence imaging agents developed by groups around the world. Our own research group focused its efforts on reactive sulfur, oxygen, and nitrogen species, pH, and hypoxia, resulting in a large family of bright chemiluminescent 1,2-dioxetanes validated for cell monitoring and in vivo imaging. These chemiluminescent probes feature low background and high sensitivity that have been proven quite useful for studying signaling, for example, the generation of peroxynitrite (ONOO-) in cellular models of immune function and phagocytosis. This high sensitivity has also enabled real-time quantitative reporting of oxygen-dependent enzyme activity and hypoxia in living cells and tumor xenograft models. We reported some of the first ratiometric chemiluminescent 1,2-dioxetane systems for imaging pH and have introduced a powerful kinetics-based approach for quantification of reactive species like azanone (nitroxyl, HNO) and enzyme activity in living cells. These tools have been applied to untangle complex signaling pathways of peroxynitrite production in radiation therapy and as substrates in a split esterase system to provide an enzyme/substrate pair to rival luciferase/luciferin. Furthermore, we have pushed chemiluminescence toward commercialization and clinical translation by demonstrating the ability to monitor airway hydrogen peroxide in the exhaled breath of asthma patients using transiently produced chemiluminescent 1,2-dioxetanedione intermediates. This body of work shows the powerful possibilities that can emerge when working at the interface of light and chemistry, and we hope that it will inspire future scientists to seek out ever brighter and more illuminating ideas.

Bright chemiluminescent dioxetane probes for the detection of gaseous transmitter H2S.

Hydrogen sulfide (H2S), the third gaseous transmitter after CO and NO, is a double-edged sword in the human body. A specific concentration of H2S can attenuate myocardial ischemia-reperfusion injury by preserving mitochondrial function, in contrast, cause illness, including inflammation and stroke. There are already some probes for the real-time monitoring of the level of H2S in the biological environment. However, they have some disadvantages, such as phototoxicity, low sensitivity, and low quantum yield. In this research, by linking 4-dinitrophenyl-ether (DNP), a specific recognition group for H2S, with a chemiluminophore 1,2-dioxetane, we designed and synthesized the probe SCL-1. To tackle the barrier that the traditional chemiluminescent group has a short emission wavelength and is not easy to penetrate deep tissues, an acrylonitrile electron-withdrawing substituent was installed to the ortho-position of the 1,2-dioxanol hydroxy group. According to the same design strategy as SCL-1, the probe SCL-2 was designed with the modified chemiluminescent group. Studies have shown that SCL-2 with electron-withdrawing acrylonitrile has higher luminescence quantum yield and high sensitivity than SCL-1, realizing real-time detection of H2S in vitro and in vivo. The LOD of SCL-2 was 0.185 µM, which was the best among the currently available luminescent probes for detecting H2S. We envisage that SCL-2 may be a practical toolbox for studying the biological functions of H2S and H2S-related diseases.

Overview of Syntheses and Molecular-Design Strategies for Tetrazine-Based Fluorogenic Probes.

Various bioorthogonal chemistries have been used for fluorescent imaging owing to the advantageous reactions they employ. Recent advances in bioorthogonal chemistry have revolutionized labeling strategies for fluorescence imaging, with inverse electron demand Diels-Alder (iEDDA) reactions in particular attracting recent attention owing to their fast kinetics and excellent specificity. One of the most interesting features of the iEDDA labeling strategy is that tetrazine-functionalized dyes are known to act as fluorogenic probes. In this review, we will focus on the synthesis, molecular-design strategies, and bioimaging applications of tetrazine-functionalized fluorogenic probes. Traditional Pinner reaction and "Pinner-like" reactions for tetrazine synthesis are discussed here, as well as metal-catalyzed C-C bond formations with convenient tetrazine intermediates and the fabrication of tetrazine-conjugated fluorophores. In addition, four different quenching mechanisms for tetrazine-modified fluorophores are presented.

64Cu/177Lu-DOTA-diZD, a Small-Molecule-Based Theranostic Pair for Triple-Negative Breast Cancer.

Despite advances in targeted therapies, the prognosis for patients with triple-negative breast cancer (TNBC) is poor because there are few actionable molecular targets. The dependence of solid tumor growth on angiogenesis prompted our development of angiogenic-receptor-targeted radionuclide therapy (TRT) to treat TNBC by targeted delivery of therapeutic doses of ionizing radiation to tumors. A high-affinity vascular endothelial growth factor receptor (VEGFR)-targeted agent, diZD, was synthesized and labeled with 177Lu and 64Cu by 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) chelator giving the TRT agent, 177Lu-DOTA-diZD, and PET imaging agent, 64Cu-DOTA-diZD. We showed that "64Cu/177Lu"-DOTA-diZD radiotracers are a promising theranostic pair for TNBC. 4T1-bearing mice treated with 177Lu-DOTA-diZD-based TRT survived with a median of 28 days, which was significantly longer than that of control mice as 18 days. Anti-PD1 immunotherapy resulted in a shorter median survival of 16 days. This work presents for the first time that small-molecule VEGFR-oriented TRT is a promising therapeutic option to treat "immunogenic cold" TNBC.

Stabilization Strategies for Linear Minigastrin Analogues: Further Improvements via the Inclusion of Proline into the Peptide Sequence.

Minigastrin (MG) analogues, known for their high potential to target cholecystokinin-2 receptor (CCK2R) expressing tumors, have limited clinical applicability due to low enzymatic stability. By introducing site-specific substitutions within the C-terminal receptor-binding sequence, reduced metabolization and improved tumor targeting can be achieved. In this work, the influence of additional modification within the N-terminal sequence has been explored. Three novel 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA)-conjugated CCK2R ligands with proline substitution at different positions were synthesized. Substitution did not affect CCK2R affinity, and the conjugates labeled with indium-111 and lutetium-177 showed a high enzymatic stability in different incubation media as well as in vivo (57-79% intact radiopeptide in blood of BALB/c mice at 1 h p.i.) combined with enhanced tumor uptake (29-46% IA/g at 4 h in xenografted BALB/c nude mice). The inclusion of Pro contributes significantly to the development of CCK2R ligands with optimal targeting properties for application in targeted radiotherapy.

Facile preparation of a novel Ga-67-labeled NODAGA-conjugated lactam-cyclized alpha-MSH peptide at room temperature for melanoma targeting.

In this study, the melanoma targeting property of 67Ga-NODAGA-GGNle-CycMSHhex {1,4,7-triazacyclononane,1-gluteric acid-4,7-acetic acid-GlyGlyNle-c[Asp-His-D-Phe-Arg-Trp-Lys]-CONH2} was determined on B16/F10 melanoma-bearing C57 mice to demonstrate the feasibility of NODAGA as a radiometal chelator for facile room temperature radiolabeling of NODAGA-GGNle-CycMSHhex. The IC50 value of NODAGA-GGNle-CycMSHhex was 0.87 ± 0.12 nM on B16/F10 melanoma cells. 67Ga-NODAGA-GGNle-CycMSHhex was readily prepared at room temperature with greater than 98% radiolabeling yield and displayed MC1R-specific binding on B16/F10 melanoma cells. The B16/F10 melanoma uptake of 67Ga-NODAGA-GGNle-CycMSHhex was 10.31 ± 0.78, 14.96 ± 1.34, 13.7 ± 3.33 and 10.4 ± 2.2% ID/g at 0.5, 2, 4 and 24 h post-injection, respectively. Approximately 85% of the injected dose was cleared out the body via urinary system at 2 h post-injection. 67Ga-NODAGA-GGNle-CycMSHhex showed high tumor/blood, tumor/muscle and tumor/skin uptake ratios after 2 h post-injection. Overall, 67Ga-NODAGA-GGNle-CycMSHhex could be easily prepared at room temperature and exhibited favorable melanoma targeting property, suggesting the potential use of NODAGA as a radiometal chelator for facile room temperature radiolabeling of α-MSH peptides.

Combinatorial discovery of thermoresponsive cycloammonium ionic liquids.

This work demonstrated, for the first time, the combinatorial discovery and rational identification of small-molecule cycloammonium-based thermoresponsive ionic liquids that exhibit LCST phase transition and carry attractive Tc values in water.

A Toxic RNA Templates the Synthesis of Its Own Fluorogenic Inhibitor by Using a Bio-orthogonal Tetrazine Ligation in Cells and Tissues.

Expanded RNA repeats cause more than 30 incurable diseases. One approach to mitigate their toxicity is by using small molecules that assemble into potent, oligomeric species upon binding to the disease-causing RNA in cells. Herein, we show that the expanded repeat [r(CUG)exp] that causes myotonic dystrophy type 1 (DM1) catalyzes the in situ synthesis of its own inhibitor using an RNA-templated tetrazine ligation in DM1 patient-derived cells. The compound synthesized on-site improved DM1-associated defects at picomolar concentrations, enhancing potency by 10 000-fold, compared to its parent compounds that cannot undergo oligomerization. A fluorogenic reaction is also described where r(CUG)exp templates the synthesis of its own imaging probe to enable visualization of the repeat in its native context in live cells and muscle tissue.

In Vivo Evaluation of Indium-111-Labeled 800CW as a Necrosis-Avid Contrast Agent.

PURPOSE: Current clinical measurements for tumor treatment efficiency rely often on changes in tumor volume measured as shrinkage by CT or MRI, which become apparent after multiple lines of treatment and pose a physical and psychological burden on the patient. Detection of therapy-induced cell death in the tumor can be a fast measure for treatment efficiency. However, there are no reliable clinical tools for detection of tumor necrosis. Previously, we studied the necrosis avidity of cyanine-based fluorescent dyes, which suffered long circulation times before tumor necrosis could be imaged due to low hydrophilicity. We now present the application of radiolabeled 800CW, a commercially available cyanine with high hydrophilicity, to image tumor necrosis in a mouse model. PROCEDURES: We conjugated 800CW to DOTA via a PEG linker, for labeling with single-photon emission-computed tomography isotope indium-111, yielding [111In]In-DOTA-PEG4-800CW. We then investigated specific [111In]In-DOTA-PEG4-800CW uptake by dead cells in vitro, using both fluorescence and radioactivity as detection modalities. Finally, we investigated [111In]In-DOTA-PEG4-800CW uptake into necrotic tumor regions of a 4T1 breast tumor model in mice. RESULTS: We successfully prepared a precursor and developed a reliable procedure for labeling 800CW with indium-111. We detected specific [111In]In-DOTA-PEG4-800CW uptake by dead cells, using both fluorescence and radioactivity. Albeit with a tumor uptake of only 0.37%ID/g at 6 h post injection, we were able to image tumor necrosis with a tumor to background ratio of 7:4. Fluorescence and radioactivity in cryosections from the dissected tumors were colocalized with tumor necrosis, confirmed by TUNEL staining. CONCLUSIONS: [111In]In-DOTA-PEG4-800CW can be used to image tumor necrosis in vitro and in vivo. Further research will elucidate the application of [111In]In-DOTA-PEG4-800CW or other radiolabeled hydrophilic cyanines for the detection of necrosis caused by chemotherapy or other anti-cancer therapies. This can provide valuable prognostic information in treatment of solid tumors.

Synthesis and Therapeutic Applications of Iminosugars in Cystic Fibrosis.

Iminosugars are sugar analogues endowed with a high pharmacological potential. The wide range of biological activities exhibited by these glycomimetics associated with their excellent drug profile make them attractive therapeutic candidates for several medical interventions. The ability of iminosugars to act as inhibitors or enhancers of carbohydrate-processing enzymes suggests their potential use as therapeutics for the treatment of cystic fibrosis (CF). Herein we review the most relevant advances in the field, paying attention to both the chemical synthesis of the iminosugars and their biological evaluations, resulting from in vitro and in vivo assays. Starting from the example of the marketed drug NBDNJ (N-butyl deoxynojirimycin), a variety of iminosugars have exhibited the capacity to rescue the trafficking of F508del-CFTR (deletion of F508 residue in the CF transmembrane conductance regulator), either alone or in combination with other correctors. Interesting results have also been obtained when iminosugars were considered as anti-inflammatory agents in CF lung disease. The data herein reported demonstrate that iminosugars hold considerable potential to be applied for both therapeutic purposes.

An efficient synthetic approach toward a sporadic heterocyclic scaffold: 1,3-Oxathiol-2-ylidenes; alkaline phosphatase inhibition and molecular docking studies.

We developed a simple and robust method for synthesis of 1,3-oxathiol-2-ylidene benzamides (4a-m) a sporadic class of heterocycles, by reacting freshly prepared aroyl isothiocyanates, with ethyl 2-chloroacetoacetate in presence of N-methylimidazole in dry acetonitrile. The synthesized compounds were explored for their inhibition against alkaline phosphatases and HeLa cancer cell lines. The results suggest that almost all the compounds possess good % inhibition against both enzymes, with compound 4m showing dual inhibition while 4g and 4i as potent and selective inhibitors of TNAP and c-IAP respectively. Structure activity relationship for the active members of series has been carried out based on molecular docking studies. The result of SAR shows the involvement of active inhibitors in H-bonding at various sites with different amino acid residues in addition to secondary metal ion interactions with Zn ions inside the active pocket of the enzyme. The π-π interactions between the 1,3-oxathiole ring and imidazole ring of His321 and His 317 further defines the dual mode of inhibition by compound 4m. These compounds also possess inhibition potential against cervical cell lines in the range of 2.42-69.03% with the maximum inhibition shown by the unsubstituted member 4a compared to the reference drug cisplatin.

Structure-Activity Relationship of Antischistosomal Ozonide Carboxylic Acids.

Semisynthetic artemisinins and other bioactive peroxides are best known for their powerful antimalarial activities, and they also show substantial activity against schistosomes-another hemoglobin-degrading pathogen. Building on this discovery, we now describe the initial structure-activity relationship (SAR) of antischistosomal ozonide carboxylic acids OZ418 (2) and OZ165 (3). Irrespective of lipophilicity, these ozonide weak acids have relatively low aqueous solubilities and high protein binding values. Ozonides with para-substituted carboxymethoxy and N-benzylglycine substituents had high antischistosomal efficacies. It was possible to increase solubility, decrease protein binding, and maintain the high antischistosomal activity in mice infected with juvenile and adult Schistosoma mansoni by incorporating a weak base functional group in these compounds. In some cases, adding polar functional groups and heteroatoms to the spiroadamantane substructure increased the solubility and metabolic stability, but in all cases decreased the antischistosomal activity.

Automated radiosynthesis and preclinical evaluation of Al[18F]F-NOTA-P-GnRH for PET imaging of GnRH receptor-positive tumors.

INTRODUCTION: Gonadotropin releasing hormone (GnRH) receptor is overexpressed in many human tumors. Previously we developed a 18F-labelled GnRH peptide. Although the GnRH-targeted PET probe can be clearly visualized by microPET imaging in a PC-3 xenograft model, clinical applications of the probe have been limited by complex labeling procedures, poor radiochemical yield, and unwanted accumulation in GnRH receptor negative tissues. In this study, we have designed a new 18F-labelled GnRH peptide that is more amenable to clinical development. METHODS: GnRH peptide analogues NOTA-P-GnRH was synthesized and automated radiolabeled with 18F using a Al[18F]F complex on a modified PET-MF-2V-IT-I synthesis module. The GnRH receptor affinities of AlF-NOTA-P-GnRH and NOTA-P-GnRH were determined by in vitro competitive binding assay. For in vitro characterization determination of stability and partition coefficients were carried out, respectively. Dynamic microPET and biodistribution studies of Al[18F]F-NOTA-P-GnRH were evaluated in xenograft tumor mouse models. RESULTS: The total radiochemical synthesis and purification of Al[18F]F-NOTA-P-GnRH was completed within 35 min with a decay-corrected yield of 35 ± 10%. The logP value of Al[18F]F-NOTA-P-GnRH was -2.74 ± 0.04 and the tracer was stable in phosphate-buffered saline, and bovine and human serum. The IC50 values of AlF-NOTA-P-GnRH and NOTA-P-GnRH were 116 nM and 56.2 nM, respectively. Dynamic PET imaging together with ex vivo biodistribution analyses revealed that Al[18F]F-NOTA-P-GnRH was clearly delineated in both PC-3 and MDA-MB-231 xenografted tumors. CONCLUSION: Al[18F]F-NOTA-P-GnRH can be efficiently produced on a commercially available automated synthesis module and has potential for use in clinical diagnosis of GnRH receptor-positive tumors. ADVANCES IN KNOWLEDGE: Our studies developed the automated radiosynthesis of a new 18F-labelled GnRH tracer and preclinical evaluation for future clinical application. IMPLICATIONS FOR PATIENT CARE: Quantitative and noninvasive imaging of GnRH expression would provide information for diagnosis and treatment of cancer patients.

Synthesis, Labeling and Preclinical Evaluation of a Squaric Acid Containing PSMA Inhibitor Labeled with 68 Ga: A Comparison with PSMA-11 and PSMA-617.

The L-lysine urea-L-glutamate (KuE) represents a key motif in recent diagnostic and therapeutic radiopharmaceuticals targeting the prostate specific membrane antigen (PSMA). Using a squaric acid moiety for coupling of KuE with a radioactive label, the squaric acid as a linker in the PSMA ligand seems to mimic the aromatic structure of the naphthylalanine unit on PSMA-617. In this work, we investigate the influence of squaric acid moiety on the biological activity of the compound carrying a KuE motif and three typical chelates. The derivatives TRAM.SA.KuE, DOTAGA.SA.KuE and NODAGA.SA.KuE were all synthesized in straightforward organic reactions and purified by HPLC afterward. Different amounts of tracer were labeled at different temperatures with 68 Ga. PET examinations were performed on NMRInu/nu nude mice with an LNCaP tumor on the right hind leg including ex vivo investigations of the organs. For comparison, 68 Ga-derivatives of PSMA-11 and PSMA-617, the derivatives most commonly used in clinics, were investigated in the same animal model.