Preparation of 177Lu-PSMA-617 in Hospital Radiopharmacy: Convenient Formulation of a Clinical Dose Using a Single-Vial Freeze-Dried PSMA-617 Kit Developed In-House.

OBJECTIVE: In the recent time, endoradionuclide therapy for metastatic castration-resistant prostate carcinoma employing 177Lu-PSMA-617 has yielded encouraging results and several clinical trials with the agent are currently ongoing. Routine preparation of 177Lu-PSMA-617 patient doses can be made simpler and convenient, if the ingredients essential for radiolabeling are made available in a ready-to-use lyophilized form. METHODS: PSMA-617 freeze-dried kit was formulated and used for the preparation of 177Lu-PSMA-617 clinical dose with high radiochemical purity using low/medium specific activity 177Lu. Detailed radiochemical studies were performed to determine the maximum activity and volume of 177LuCl3, which can be added in the kit for the formulation of 177Lu-PSMA-617. Studies were also performed to determine the shelf life of the kit to ensure its long-term usage. Studies were performed in buffer as well as human serum medium to determine the stability of the 177Lu-PSMA-617 complex after storing in respective media up to 7 days postpreparation. About ten patient doses of 177Lu-PSMA-617 were administered, and posttherapy scans were acquired. RESULTS: The formulated freeze-dried kit of PSMA-617 could be radiolabeled with an average percentage radiochemical purity > 98.53 ± 0.38. The freeze-dried kit was found suitable for tolerating up to 0.5 mL of 177LuCl3 (in 0.01 N HCl) and specific activity of 555 MBq/µg (15 mCi/µg) for the preparation of the patient dose of 177Lu-PSMA-617. The 177Lu-PSMA-617 complex prepared using the freeze-dried kit of PSMA-617 was observed to maintain % radiochemical purity (RCP) of 96.74 ± 0.87 and 94.81 ± 2.66, respectively, even after storing up to 7 days in buffer and human serum, respectively. 177Lu-PSMA-617 prepared using the in-house formulated freeze-dried kit of PSMA-617 exhibited accumulation in metastatic lesions picked up in a pretherapy PET scan. Reduction in number as well as size of lesions was observed in posttherapy scans acquired after two months of administering the first therapeutic dose of 177Lu-PSMA-617. CONCLUSIONS: The freeze-dried kit of PSMA-617 could be used for the preparation of 177Lu-PSMA-617 with high radiochemical purity (>98%) in a reproducible manner. 177Lu-PSMA-617 prepared using the developed kit was successfully evaluated in patients suffering from metastatic prostate cancer.

Automated cassette-based production of high specific activity [203/212Pb]peptide-based theranostic radiopharmaceuticals for image-guided radionuclide therapy for cancer.

A method for preparation of Pb-212 and Pb-203 labeled chelator-modified peptide-based radiopharmaceuticals for cancer imaging and radionuclide therapy has been developed and adapted for automated clinical production. Pre-concentration and isolation of radioactive Pb2+ from interfering metals in dilute hydrochloric acid was optimized using a commercially-available Pb-specific chromatography resin packed in disposable plastic columns. The pre-concentrated radioactive Pb2+ is eluted in NaOAc buffer directly to the reaction vessel containing chelator-modified peptides. Radiolabeling was found to proceed efficiently at 85°C (45min; pH 5.5). The specific activity of radiolabeled conjugates was optimized by separation of radiolabeled conjugates from unlabeled peptide via HPLC. Preservation of bioactivity was confirmed by in vivo biodistribution of Pb-203 and Pb-212 labeled peptides in melanoma-tumor-bearing mice. The approach has been found to be robustly adaptable to automation and a cassette-based fluid-handling system (Modular Lab Pharm Tracer) has been customized for clinical radiopharmaceutical production. Our findings demonstrate that the Pb-203/Pb-212 combination is a promising elementally-matched radionuclide pair for image-guided radionuclide therapy for melanoma, neuroendocrine tumors, and potentially other cancers.

Preparation of 68Ga-labelled DOTA-peptides using a manual labelling approach for small-animal PET imaging.

(68)Ga-DOTA-peptides are a promising PET radiotracers used in the detection of different tumours types due to their ability for binding specifically receptors overexpressed in these. Furthermore, (68)Ga can be produced by a (68)Ge/(68)Ga generator on site which is a very good alternative to cyclotron-based PET isotopes. Here, we describe a manual labelling approach for the synthesis of (68)Ga-labelled DOTA-peptides based on concentration and purification of the commercial (68)Ga/(68)Ga generator eluate using an anion exchange-cartridge. (68)Ga-DOTA-TATE was used to image a pheochromocytoma xenograft mouse model by a microPET/CT scanner. The method described provides satisfactory results, allowing the subsequent (68)Ga use to label DOTA-peptides. The simplicity of the method along with its implementation reduced cost, makes it useful in preclinical PET studies.

Development of a freeze-dried kit formulation for the preparation of 99mTc-NTP 15-5, a radiotracer for scintigraphic imaging of proteoglycans.

The cartilage-targeting strategy is based on the strong affinity of quaternary ammonium (QA) functions for cartilage proteoglycans. We use a bifunctional agent containing QA moiety and a polyazamacrocycle structure able to complex technetium-99m. (99m)Tc-NTP 15-5 was selected for its high stability and its high affinity for proteoglycans in vivo. Labeling conditions of NTP 15-5 were optimized, and a lyophilized kit was developed for radiolabeling of (99m)Tc-NTP 15-5 (radiochemical yields 94.6±1.8%). (99m)Tc-NTP 15-5 was stable and resulted in favorable biological evaluations.

A novel array of chemiluminescence sensors for sensitive, rapid and high-throughput detection of explosive triacetone triperoxide at the scene.

Triacetone triperoxide (TATP) is relatively easy to make and has been used in various terrorist acts. Early but easy detection of TATP is highly desired. We designed a new type sensor array for H2O2. The unique CL sensor array was based on CeO2 nanoparticles' membranes, which have an excellent catalytic effect on the luminol-H2O2 CL reaction in alkaline medium. It exhibits a linear range for the detection of H2O2 from 1.0×10(-8) to 5.0×10(-5)M (R(2)=0.9991) with a 1s response time. The detection limit is 1.0×10(-9)M. Notably, the present approach allows the design of CL sensor array assays in a more simple, time-saving, long-lifetime, high-throughput, and economical approach when compared with conventional CL sensor. It is conceptually different from conventional CL sensor assays. The novel sensor array has been successfully applied for the detection of TATP at the scene.

Determination of polyether ionophores in urban sewage sludge by pressurised liquid extraction and liquid chromatography-tandem mass spectrometry: study of different clean-up strategies.

A method for the determination of five polyether ionophores in urban sewage sludge has been developed. The extraction of compounds was performed by pressurised liquid extraction using acetone, while florisil was used for in-cell clean-up to minimise the matrix effect in the sludge extracts. An amide polar-embedded reversed-phase column was used for the chromatographic separation with a rapid-resolution liquid chromatograph coupled to a tandem mass spectrometer. Moreover, several clean-up strategies such as in-cell and on-cell clean-up and solid-phase extraction clean-up, among others, were tested and their results are discussed in the present paper. Recoveries (10 and 250 µg/kg in dry weight (d.w.), n=6) were close to 90%, repeatability and reproducibility (%RSD, 10 and 250 µg/kg (d.w.), n=6) were less than 10% and 12%, respectively. Limits of detection (LODs) and limits of quantification (LOQs) ranged between 0.5 and 1 µg/kg (d.w.) and between 1 and 5 µg/kg (d.w.), respectively. The method was applied to samples collected in five sewage treatment plants in Catalonia. Monensin and narasin were determined in some sludge samples at concentrations from

Fast detection of triacetone triperoxide (TATP) from headspace using planar solid-phase microextraction (PSPME) coupled to an IMS detector.

Triacetone triperoxide (TATP) is a high explosive synthesized from easily available reactants making it accessible for illicit uses. In this study, fast detection of TATP is achieved using a novel planar solid-phase microextraction (PSPME) as a preconcentration and sampling device for headspace analysis offering improved sensitivity and reduced sampling time over the conventional fiber-based solid-phase microextraction (SPME) when followed by ion mobility spectrometer (IMS) detection. Quantitation and comparison of the retention capabilities of PSPME as compared to the commercially available SPME were determined using TATP standards and analyzed using gas chromatography-mass spectrometry for SPME analysis and a commercial IMS with no instrumental modification for PSPME. Static and dynamic headspace extractions were used and compared for PSPME extractions, in which low milligram quantities of TATP were detected within 30 s of static mode sampling and less than 5 s in the dynamic mode sampling for PSPME-IMS.

A novel phytotoxic nonenolide from Phomopsis sp. HCCB03520.

A novel phytotoxic nonenolide, (6S,7R,9R)-6,7-dihydroxy-9-propylnon-4-eno-9-lactone (1), was isolated from solid cultures of the endophytic fungus Phomopsis sp. HCCB03520, together with three known compounds, cytochalasin H (2), cytochalasin N (3), and epoxycytochalasin H (4). The structures of these compounds were elucidated through spectroscopic analysis, and the absolute configurations were determined by CD spectroscopy. Phytotoxic activities of compounds 1-4 were also investigated. Compound 1 showed phytotoxic activity on germination and radicle growth of Medicago sativa, Trifolium hybridum, and Buchloe dactyloides.

Molluscicidal metabolites from an assemblage of Palmyra Atoll cyanobacteria.

Molluscicides can play an important role in the control of schistosomiasis because snails of the genus Biomphalaria act as intermediate hosts for the parasite. Schistosomiasis is one of 13 neglected tropical diseases with high morbidity and mortality that collectively affect one billion of the world's poorest population, mainly in developing countries. Thiopalmyrone (1) and palmyrrolinone (2), metabolites isolated from extracts of a Palmyra Atoll environmental assemblage of two cyanobacteria, cf. Oscillatoria and Hormoscilla spp., represent new and potent molluscicidal chemotypes against Biomphalaria glabrata (LC50=8.3 and 6.0 µM, respectively). A slight enhancement in molluscicidal effect (LC50=5.0 µM) was observed when these two natural products were utilized as an equimolar binary mixture.

1,5-diazacyclohenicosane, a new cytotoxic metabolite from the marine sponge Mycale sp.

A new cyclic diamine, 1,5-diazacyclohenicosane (1), was isolated from samples of the marine sponge Mycale sp. collected at Lamu Island (Kenya). Its structure was determined by a combination of spectroscopic techniques, including (+)-HRESIMS and 1D and 2D NMR spectroscopy. The compound displayed cytotoxicity at the muM level against three human tumor cell lines.

Stagonolides G-I and modiolide A, nonenolides produced by Stagonospora cirsii, a potential mycoherbicide for Cirsium arvense.

Stagonospora cirsii Davis, a fungal pathogen isolated from Cirsium arvense (commonly called Canada thistle) and proposed as a potential mycoherbicide of this perennial noxious weed, produces phytotoxic metabolites in liquid and solid cultures. Stagonolide, the main phytotoxic metabolite, and five new related nonenolides, named stagonolides B-F, were isolated from the fungus. When grown on solid culture, nonenolide yields increased. A further four nonenolides were isolated and characterized by spectroscopy. Three were new compounds and named stagonolides G-I, and the fourth was identified as modiolide A, previously isolated from Paraphaeosphaeria sp., a fungus separated from the horse mussel. Leaf disk-puncture assays at 1 mg/mL of stagonolides H-I and modiolide A were phytotoxic to C. arvense. Only stagonolide H inhibited chicory seedling root growth. The most potent toxin, stagonolide H, indicated selectivity when tested on leaves of eight different plants: Canada thistle was most sensitive to the compound.

Alpha-glucosidase inhibitor from Kothala-himbutu (Salacia reticulata WIGHT).

A polyhydroxylated cyclic 13-membered sulfoxide (1) was isolated from an aqueous extract of Kothala-himbutu ( Salacia reticulata WIGHT). The structure of compound 1 was elucidated by 1D and 2D NMR and APCI-MS methods. The alpha-glucosidase inhibitory activity of compound 1 (IC 50: maltase, 0.227 microM; sucrase, 0.186 microM; isomaltase, 0.099 microM) was much greater than the inhibitory activity of salacinol and kotalanol, which were previously isolated from Kothala-himbutu.

Stagonolides B-F, nonenolides produced by Stagonospora cirsii, a potential mycoherbicide of Cirsium arvense.

Stagonospora cirsii, a fungal pathogen isolated from Cirsium arvense and proposed as a potential mycoherbicide of this perennial noxious weed, produces phytotoxic metabolites in liquid and solid cultures. Recently, the main metabolite, stagonolide (1), with interesting phytotoxic properties, was isolated from a liquid culture and characterized as a new nonenolide. In the present work this same fungus, grown in solid culture, exhibited an increased capacity to produce nonenolides. Five new nonenolides, named stagonolides B-F (2-6), were isolated and characterized using spectroscopic methods. When tested by a leaf disk puncture assay at a concentration of 1 mg/mL, these compounds showed no toxicity to C. arvense and Sonchus arvensis, whereas stagonolide (1) was highly toxic. Stagonolide (1) and stagonolide C (3) were weakly toxic to Colpoda steinii, a protozoan, when tested at 0.05 mg/mL, with the other stagonolides nontoxic. A number of structure-activity relationship observations were made.

[64Cu-NOTA-8-Aoc-BBN(7-14)NH2] targeting vector for positron-emission tomography imaging of gastrin-releasing peptide receptor-expressing tissues.

Radiolabeled peptides hold promise as diagnostic/therapeutic targeting vectors for specific human cancers. We report the design and development of a targeting vector, [(64)Cu-NOTA-8-Aoc-BBN(7-14)NH(2)] (NOTA = 1,4,7-triazacyclononane-1,4,7-triacetic acid, 8-Aoc = 8-aminooctanoic acid, and BBN = bombesin), having very high selectivity and affinity for the gastrin-releasing peptide receptor (GRPr). GRPrs are expressed on a variety of human cancers, including breast, lung, pancreatic, and prostate, making this a viable approach toward site-directed localization or therapy of these human diseases. In this study, [NOTA-X-BBN(7-14)NH(2)] conjugates were synthesized, where X = a specific pharmacokinetic modifier. The IC(50) of [NOTA-8-Aoc-BBN(7-14)NH(2)] was determined by a competitive displacement cell-binding assay in PC-3 human prostate cancer cells using (125)I-[Tyr(4)]-BBN as the displacement ligand. An IC(50) of 3.1 +/- 0.5 nM was obtained, demonstrating high binding affinity of [NOTA-8-Aoc-BBN] for the GRPr. [(64)Cu-NOTA-X-BBN] conjugates were prepared by the reaction of (64)CuCl(2) with peptides in buffered aqueous solution. In vivo studies of [(64)Cu-NOTA-8-Aoc-BBN(7-14)NH(2)] in tumor-bearing PC-3 mouse models indicated very high affinity of conjugate for the GRPr. Uptake of conjugate in tumor was 3.58 +/- 0.70% injected dose (ID) per g at 1 h postintravenous injection (p.i.). Minimal accumulation of radioactivity in liver tissue (1.58 +/- 0.40% ID per g, 1 h p.i.) is indicative of rapid renal-urinary excretion and suggests very high in vivo kinetic stability of [(64)Cu-NOTA-8-Aoc-BBN(7-14)NH(2)] with little or no in vivo dissociation of (64)Cu(2+) from the NOTA chelator. Kidney accumulation at 1 h p.i. was 3.79 +/- 1.09% ID per g. Molecular imaging studies in GRPr-expressing tumor models produced high-contrast, high-quality micro-positron-emission tomography images.

Trigoxazonane, a monosubstituted trioxazonane from Trigonella foenum-graecum root exudate, inhibits Orobanche crenata seed germination.

Orobanche crenata is a major threat to grain legume production. Fenugreek (Trigonella foenum-graecum) is an annual legume that has been shown to effectively reduce O. crenata infection when intercropped with grain legumes. In this paper, we point that this can be attributed to allelopathy, through inhibition of the germination of O. crenata by fenugreek root exudates. The main inhibitory metabolite was isolated and characterized. Allelopathy was demonstrated in different bioassays, by inhibition of O. crenata seeds germination both by growing fenugreek and pea plants together (intercropped), and by application of fenugreek root exudates. Fenugreek root exudates were extracted with organic solvent and fractionated giving several fractions, two of which showed moderate (27%) and strong (54%) inhibition of O. crenata seed germination, respectively. The most active metabolite is a new monosubstituted trioxazonane, characterized by spectroscopic methods as the 2-butyl-[1,4,7,2]trioxazonane and named trigoxazonane.

Remediation of RDX- and HMX-contaminated groundwater using organic mulch permeable reactive barriers.

Organic mulch is a complex organic material that is typically populated with its own consortium of microorganisms. The organisms in mulch breakdown complex organics to soluble carbon, which can then be used by these and other microorganisms as an electron donor for treating RDX and HMX via reductive pathways. A bench-scale treatability study with organic mulch was conducted for the treatment of RDX- and HMX-contaminated groundwater obtained from a plume at the Pueblo Chemical Depot (PCD) in Pueblo, Colorado. The site-specific cleanup criteria of 0.55 ppb RDX and 602 ppb HMX were used as the logical goals of the study. Column flow-through tests were run to steady-state at the average site seepage velocity, using a 70%:30% (vol.:vol.) mulch:pea gravel packing to approach the formation's permeability. Significant results included: (1) Complete removal of 90 ppb influent RDX and 8 ppb influent HMX in steady-state mulch column effluent; (2) pseudo-first-order steady-state kinetic rate constant, k, of 0.20 to 0.27 h(-1) based on RDX data, using triplicate parallel column runs; (3) accumulation of reduced RDX intermediates in the steady-state column effluent at less than 2% of the influent RDX mass; (4) no binding of RDX to the column fill material; and (5) no leaching of RDX, HMX or reduction intermediates from the column fill material. The results of the bench-scale study will be used to design and implement a pilot-scale organic mulch/pea gravel permeable reactive barrier (PRB) at the site.

DNA-templated organic synthesis and selection of a library of macrocycles.

The translation of nucleic acid libraries into corresponding synthetic compounds would enable selection and amplification principles to be applied to man-made molecules. We used multistep DNA-templated organic synthesis to translate libraries of DNA sequences, each containing three "codons," into libraries of sequence-programmed synthetic small-molecule macrocycles. The resulting DNA-macrocycle conjugates were subjected to in vitro selections for protein affinity. The identity of a single macrocycle possessing known target protein affinity was inferred through the sequence of the amplified DNA template surviving the selection. This work represents the translation, selection, and amplification of libraries of nucleic acids encoding synthetic small molecules rather than biological macromolecules.

Remediating munitions-contaminated soil with zerovalent iron and cationic surfactants.

Soils contaminated from military operations often contain mixtures of HMX (octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine), RDX (hexahydro-1,3,5-trinitro-1,3,5-triazine), and TNT (2,4,6-trinitrotoluene) rather than a single explosive. Differences among explosives in solubility and reactivity make developing a single remediation treatment difficult. When Fe(0) was used to treat a munitions-contaminated soil, we observed high rates of destruction for RDX and TNT (98%) but not HMX. Our objective was to determine if HMX destruction by Fe(0) could be enhanced by increasing HMX solubility by physical (temperature) or chemical (surfactants) means. To determine electron acceptor preference, we treated RDX and HMX with Fe(0) in homogeneous solutions and binary mixtures. Increasing aqueous temperature (20 to 55 degrees C) increased HMX solubility (2 to 22 mg L(-1)) but did not increase destruction by Fe(0) in a contaminated soil slurry that also contained RDX and TNT. Batch experiments using equal molar concentrations of RDX and HMX demonstrated that RDX was preferentially reduced over HMX by Fe(0). By testing various surfactants, we found that the cationic surfactants (HDTMA [hexadecyltrimethylammonium bromide], didecyl, and didodecyl) were most effective in increasing HMX concentration in solution. Didecyl and HDTMA were also found to be highly effective in facilitating the transformation of HMX by Fe(0). Using HDTMA or didecyl solutions (3%, w/v) containing solid-phase HMX, we observed that 100% of the added HMX was transformed by Fe(0) in the didecyl matrix and 60% in the HDTMA matrix. These results indicate that cationic surfactants can increase HMX solubility and facilitate Fe(0)-mediated transformation kinetics but HMX destruction rates will be slowed when RDX is present.

68Ga-labelled DOTA-derivatised peptide ligands.

68Ge/68Ga generators provide cyclotron-independent access to positron emission tomography (PET) radiopharmaceuticals. We describe a system which allows the safe and efficient handling of 68Ge/68Ga generator eluates for labelling of DOTA-derivatised peptide ligands. The system comprises concentration and purification of the 68Ga eluate as well as labelling and purification steps for peptides, and can be used with different 68Ge/68Ga generator types. The suitability and efficiency were tested with two different DOTA-derivatised somatostatin derivatives and a DOTA-derivatised bombesin derivative. Amounts of 10-20 nmol of the peptides were sufficient and resulted in labelling yields of 50% for all peptides. The built-in safety precautions have proven to be appropriate in allowing use of the method for routine clinical applications. The system was set up and operated in a "hot lab" by personnel with no previous experience in the preparation of PET radiopharmaceuticals.

Heteroaromatic monothiocarboxylic acids from Pseudomonas spp.

Pyridine derivatives substituted with monothiocarboxylic acid groups are the unique metabolites of certain Pseudomonas species. Pyridine-2,6-di-(monothiocarboxylic acid) 1a was found during a screening program for antibiotically active bacterial metabolites due to its ability to complex Fe3+. The structure of this complex, its redox behavior and the biogenesis of the ligand molecule were studied in detail. This lead to the discovery of a new class of natural products, viz. acylsulfenic acid derivatives. Interest in la was revived shortly when complexes with other metals were studied as models for sulfur-containing enzymes. It could also be shown that a quinoline monothiocarboxylic acid derivative acted as an alternative siderophore for Pseudomonas fluorescens. But a real renaissance was observed only when the role of la in the degradation of CCl4 by Pseudomonas stutzeri became evident.