Preparation and characterization of [99TcO] apcitide: a technetium labeled peptide

[99mTcO] apcitide (99mTcO(P246)), the technetium complex of the 13 amino acid, apcitide, cyclo-(D-Tyr-Apc-Gly-Asp-Cys)-Gly-Gly-Cys(Acm)-Gly-Cys(Acm)-Gly-Gly-Cys-NH2, where Apc is L-[S-(3-aminopropyl)]cysteine (an arginine mimetic) and Acm is the acetamidomethyl protecting group, has high affinity and selectivity for the GPIIb/IIIa receptor that is expressed on the membrane surface of activated platelets and plays an integral role in platelet aggregation and thrombus formation. Bibapcitide, a 26 amino acid, bis-succinimidomethyl ether-linked dimer of the peptide apcitide has been formulated as a single-vial, lyophilized kit having the trade name AcuTect. When sterile, nonpyrogenic sodium pertechnetate (99mTcO4-) in 0.9% sodium chloride is added to the AcuTect radiopharmaceutical kit and the resulting kit is heated, [99mTcO] apcitide forms. This is the first radiopharmaceutical to target acute deep vein thrombosis (DVT) in the lower extremities. We report here the preparation, purification, and isolation of the 99Tc complex of apcitide and its characterization to determine the mode of binding of Tc to apcitide. [99TcO] apcitide was prepared, on the macroscopic level, by reaction of [99TcOCl4]- with apcitide, purified by preparative HPLC and isolated as a trifluoroacetate salt. [99TcO] apcitide can also be formed from the reaction of bibapcitide and 99TcO4- in the presence of Sn(II) and glucoheptonate at 80 degrees C, conditions that mimic the radiopharmaceutical kit preparation. FTIR data show a Tc=O stretch at 961.2 cm(-1), in the range observed for anionic [TcVO]3+ amide thiolate complexes. The mass spectral data is in agreement with the formula, [C51H73O20N17S5Tc]-, consistent with retention of Acm groups and the Tc binding in the Gly11-Gly12-Cys13 region of the peptide. Despite significant spectral overlap due to numerous similar amino acids, all protons of apcitide and [99TcO] apcitide were unambiguously assigned. The observation of two nonequivalent Acm groups and the observation of only 10 NH-CH cross-peaks in the TOCSY and COSY spectra of [99TcO] apcitide (NH-CH cross-peaks were absent for Gly11-Gly12-Cys13), compared to all 13 cross-peaks found in apcitide, provided compelling evidence to support the 99Tc binding to the terminal Gly11-Gly12-Cys13 region of apcitide.     

Novel 99mTc labeled σ receptor ligand as a potential tumor imaging agent

A novel 99mTc labeled complex, [N-[2-((2-oxo-2-(4-(3-phenylpropyl)piperazin-1-yl)ethyl) (2-mercaptoethyl)amino)acetyl]-2-aminoethanethiolato]Technetium(V) oxide (PPPE-MAMA′-99mTcO) ([99mTc]-2) has been designed and prepared based on the integrated approach. The corresponding rhenium complex (PPPE-MAMA′-ReO)(Re-2) has been prepared and characterized. In vitro competition binding assays show moderate affinity of Re-2 towards σ1 and σ2 receptors with Ki values of 8.67 ± 0.07 and 5.71 ± 1.88 μmol, respectively. Planar images obtained at 0.5 h, 4 h, 20 h after I.v. Injection indicate the accumulation of [99mTc]-2 in MCF-7 human breast tumor bearing mice at 20 h. Furthermore, the accumulation of [99mTc]-2 has been inhibited at 20 h after co-injection of [99mTc]-2 plus haloperidol (1 mg/kg). Biodistribution studies of [99mTc]-2 display an in vivo tumor uptake of 0.14% ± 0.01% ID/g at 24 h post I.v. Injection with a tumor/muscle ratio of 6.02 ± 0.87. The above results suggest that [99mTc]-2, derived from a previously published lead compound, retains certain tumor uptake and affinity for σ receptors. [99mTc]-2 may be used as a basis for further structural modifications to develop tumor imaging agents with high affinity for σ receptors.     

Bridging the gap between in vitro and in vivo imaging: isostructural Re and 99mTc complexes for correlating fluorescence and radioimaging studies

A bifunctional ligand that is capable of forming Re and 99mTc complexes as complementary fluorescent and radioactive probes was developed. The tridentate bis(quinoline) amine ligand, which is referred to as the SAACQ system, was prepared in a single step from Fmoc protected lysine in high yield. Reaction of the SAACQ ligand with [Re(CO)3Br3]2- resulted in the formation of the SAACQ-(Re(CO)3)+complex which exhibits favorable fluorescence properties including a long lifetime and a large Stoke's shift. Because the SAACQ ligand is derived from an amino acid, it can readily be linked to or incorporated within peptides as a means of targeting the probe to specific receptors. To demonstrate this feature, the SAACQ ligand and the SAACQ-Re complex were incorporated into fMLFG, a peptide that binds to the formyl peptide receptor (FPR). Uptake of the fMLF[(SAACQ-Re(CO)3)+]G conjugate into human leukocytes in vitro was visualized by fluorescence microscopy, and the observed distribution of the peptide was similar to that of a well-established fluorescent FPR probe. The corresponding Tc complex, fMLF[(SAACQ-99mTc(CO)3)+]G, was prepared in excellent yield from [99mTc(CO)3(OH2)3]+, which affords the opportunity to correlate the results of the microscopy experiments with in vivo radioimaging studies because the probes are isostructural.     

Synthesis and Characterization of Oxotechnetium(V) Mixed-Ligand Complexes Containing a Tridentate N-Substituted Bis(2-mercaptoethyl)amine and a Monodentate Thiol

A series of 22 mixed-ligand complexes of the general formula TcOL(1)L(2), where L(1)H(2) are N-substituted bis(2-mercaptoethyl)amine ligands, [SN(R)S], and L(2)H are monodentate thiols as coligand, is reported. The complexes were prepared by the ligand exchange method using Tc-gluconate as precursor and equimolar quantities of the two ligands. In all cases the syn stereoisomer was formed in high yield and isolated as a crystalline product. In four cases HPLC analysis demonstrated the presence of the anti stereoisomer in the reaction mixture. Although the yield was less than 1%, one anti isomer, 4a, was successfully isolated as brown crystals. The isolated complexes were characterized by spectroscopic methods and elemental analysis. The formation of the two diastereomers, syn and anti, was expected due to the configuration of the nitrogen substituent (R) with respect to the central TcO core. The X-ray crystallography showed that the coordination geometry of the syn isomers 9, 11, and 18 is trigonal bipyramidal while for the anti isomer 4a it is distorted square pyramidal. This is the first documentation of syn/anti isomerism in N-substituted TcO[SN(R)S][S] mixed-ligand complexes.     

Influence of different 99mTc cores on the physicochemical and biodistribution behaviours of 99mTc-labelled complexes of pamidronate dithiocarbamate

Journal of Radioanalytical and Nuclear Chemistry - This study sought to evaluate the impact of 99mTcO and 99mTc≡N cores on the physicochemical and biodistribution properties of 99mTc-labelled...     

Aqueous fluoride and the preparation of [99mTc(CO)3(OH2)3]+ and 99mTc-carborane complexes

A method for the preparation of eta5-metallocarborane complexes of technetium-99m in water was developed. The key to the procedure is the use of aqueous sodium or potassium fluoride, which prevents premature degradation of the Tc(I) starting material used to prepare the carborane complexes. Solid-phase extraction was used to purify Tc-metallocarboranes derived from both ortho and meta isomers, which were isolated in good to excellent yields in high radiochemical purities. In conjunction with these studies, a series of fluoride-based "kits" were developed to produce the key precursor [99mTc(CO)3(H2O)3]+ in the absence of any other stabilizing ligand. Using this approach, [99mTc(CO)3(H2O)3]+ could be prepared directly from 99mTcO4- under a range of pH values, including neutral pH, which affords the opportunity to develop one-pot labeling procedures for base-sensitive targeting vectors.     

^9^9^mTcO^3^+和^9^9^mTcN^2^+的二胺基二硫醇配合 物在脑中滞留机制的量子 化学研究

N,N'-二(2-巯乙基)-乙二胺(DTEN)作为二胺基二硫醇类配体的模型化合物,CH~3SH作为还原型谷胱甘肽(GSH)的模拟物,用从头算分子轨道理论研究了这两种配合物与CH~3S^-的反应产物。结果表明,CH~3S^-配位到TcO-DTEN的Tc原子上,而将其转化为不能穿越血脑屏障的[TcO-DTEN-SCH~3]^-离子。与此相反,CH~3S^-不能与TcN-DTEN中的Tc配位,后者仍保持可以穿越血脑屏障的中性分子状态。以此解释了^9^9^mTcO^3^+的二胺基二硫醇配合物比相应的^9^9^mTcN^2^+配合物在脑中有较高的滞留。还用从头算分子轨道理论研究了胺基配体与TcO^3^+及TcN^2^+配位时从胺基N上脱去质子的规律。     

Purity control of the radiopharmaceutical thiamine pyrophosphate-Sn-99mTc

A new purity control method was developed in order to assure the radiochemical purity of thiamine pyrophosphate-Sn-^99mTc. The method described consists in ascending cromatography using Whatman N^o 1 for support and 5% sodium citrate as solvent. It is capable of detecting free^99mTcO ₄ ^– and colloid formation, the most frequent impurities found in^99mTc radiopharmaceuticals.     

Determination of residual activities in 99mTc generators and 81mKr generators

The residual activities present in spent 99Mo-99mTc generators and 81Rb-81mKr generators were determined from a view point of the disposal of radiopharmaceuticals. The results showed that seven kinds of residual nuclides such as 95Zr-95Nb, 103Ru, 90Sr, etc. were found in 99mTc generators with maximum level of 740 Bq/3.7 GBq (0.02 muCi/100 mCi) at one year after the reference date. In 81mKr generators one year old, six nuclides such as 57Co, 83Rb, 84Rb and others remained and these activities were less than 3.7 kBq/370 MBq (0.1 muCi/10 mCi). On the basis of these data, the disposal of 99mTc generators and 81mKr generators is discussed.     

Synthesis of chiral vicinal diamines by highly diastereoselective three-component phenolic Mannich reaction: temperature-dependent stereodivergency

[reaction: see text] A diastereoselective three-component synthesis of chiral o-1,2-diaminoalkyl phenols from an electron-rich phenol, an amine, and a chiral alpha-N,N-dibenzylamino aldehyde is developed. The diastereoselectivity of this phenolic Mannich reaction is temperature-dependent,and either anti or syn diastereomer can be prepared by controlling the reaction conditions. Low reaction temperature (-20 degrees C) favors the formation of anti adduct 1, whereas higher temperature (60 degrees C) under otherwise identical conditions produces mainly the syn isomer.     

Rhenium analogues of promising renal imaging agents with a [99mTc(CO)3]+ core bound to cysteine-derived dipeptides, including lanthionine

The coordination chemistry of lanthionine (LANH2) and cystathionine (CSTH2) dipeptides, which respectively consist of two cysteines and one cysteine and one homocysteine linked by a thioether bridge, is almost unstudied. Recently for fac-[99mTc(CO)3(LAN)]- isomers, the first small 99mTc(CO)3 agents evaluated in humans were found to give excellent renal images and to have a high specificity for renal excretion. Herein we report the synthesis and characterization of Re complexes useful for interpreting the nature of tracer 99mTc radiopharmaceuticals. Treatment of [Re(CO)3(H2O)3]OTf with commercially available LANH2 (a mixture of meso (d,l) and chiral (dd,ll) isomers) gave three HPLC peaks, 1A, 1B, and 1C, but treatment with CSTH2 (l,l isomer) gave one major product, Re(CO)3(CSTH) (2). Crystalline Re(CO)3(LANH) products were best obtained with synthetic LANH2, richer in meso or chiral isomers. X-ray crystallography showed that these dipeptides coordinate as tridentate N2S-bound ligands with two dangling carboxyls. The LANH ligand is meso in 1A and 1C and chiral in 1B. While 1A (kinetically favored) is stable at ambient temperature for days, it converted into 1C (thermodynamically favored) at 100 degrees C; after 6 h, equilibrium was reached at a 1A:1C ratio of 1:2 at pH 8. The structures provide a rationale for this behavior and for the fact that 1A and 1C have simple NMR spectra. This simplicity results from fluxional interchange between an enantiomer with both chelate rings having the same delta pucker and an enantiomer with both chelate rings having the same lambda pucker. Agents with the [99mTc(CO)3]+ core and N2S ligands show promise of becoming an important class of 99mTc radiopharmaceuticals. The chemistry of Re analogues with these ligands, such as the LAN2- complexes reported here, provides a useful background for designing new small agents and also tagged large agents because two uncoordinated carboxyl groups are available for conjugation with biological molecules such as proteins.     

Using Tc-Re chemical analogy to synthesize and identify new99mTc complexes

The strong chemical resemblance between Tc and Re is applied to design and evaluate experiments with^99mTc complexes. A combination of spectrophotometric and electrophoretic techniques allows to propose the formula [TcO₂/amine/₂]⁺ for compounds prepared by reduction of^99mTcO ₄ ^– with Zn /solid phase/ in presence of several /bidentate/ amines.     

Synthesis and biological evaluation of novel 99mTcN-labeled bisnitroimidazole complexes containing monoamine-monoamide dithiol as potential tumor hypoxia markers

Tumor hypoxia can decrease the efficacy of clinical therapy due to resistance toward radiation damage and chemotherapy, thus detection of tumor hypoxia by radiolabeled hypoxia markers is important for the control of tumor. Radiopharmaceuticals with two bioreductive groups, such as propylene amine oxime-bisnitroimidazole or monoamine-monoamide dithiol (MAMA) -bisnitroimidazole, have potential to improve hypoxia selectivity. In order to obtain radiopharmaceuticals with better features, we synthesized two novel [^99mTcN]²⁺ complexes with bisnitroimidazole moieties and MAMA ligand for targeting tumor hypoxia. Their physicochemical characters and biodistribution were also investigated. Both the [^99mTcN]²⁺ complexes show good stability and hydrophilicity. They show faster clearance from blood and soft tissues, better tumor retention and favorable tumor-to-tissue ratios compared with a control complex without nitroimidazole group. In addition, both of them show more favorable biodistribution patterns than the corresponding [^99mTcO]³⁺ complexes. These results indicate that the ^99mTcN-labeled MAMA-bisnitroimidazole complexes would have potential to image tumor hypoxia in vivo.     

Ether and crown ether-containing cationic 99mTc complexes useful as radiopharmaceuticals for heart imaging

While radiopharmaceutical research has been focused on the development of target-specific radiotracers for early detection and radiotherapy of cancers in the last decade, there is a limited effort on new cationic 99mTc radiotracers for heart imaging. This review will summarize some of the most recent developments in ether- and crown ether-containing cationic 99mTc radiotracers that have a fast liver clearance with a heart/liver ratio substantially better than that of 99mTc-Sestamibi and 99mTc-Tetrofosmin, the two commercial 99mTc radiopharmaceuticals currently available for myocardial perfusion imaging. Fast liver clearance might shorten the duration of imaging protocols (<30 min post-injection), and allow for early acquisition of heart images of high quality. Improvement of heart/liver ratio may permit better detection of the presence and extent of coronary artery disease. Identification of such a new radiotracer that allows for the improved non-invasive delineation of myocardial perfusion would be of considerable benefit in treatment of patients with suspected coronary artery disease.     

Convenient route leading to neutral fac-M(CO)3(NNO) complexes (M = Re, 99mTc) coupled to amine pharmacophores

The synthesis and characterization of three neutral tricarbonyl fac-M(CO)3(NNO) (M = Re, (99m)Tc) complexes based on the picolylamine N,N-diacetic acid (PADA) ligand is reported. One of the two carboxylate groups of the PADA ligand is efficiently and conveniently derivatized with an amine nucleophile through the use of the PADA anhydride. In this work, aniline, benzylamine and pyrrolidine were used as model amine nucleophiles. The rhenium complexes were synthesized using the [NEt4]2[Re(CO)3Br3] precursor and fully characterized by elemental analysis, spectroscopic methods, and X-ray crystallography. The analogous technetium-99m complexes were also prepared quantitatively using the [(99m)Tc(CO)3(H2O)3](+) precursor. The reaction scheme presented for the synthesis of the fac-M(CO)3(NNO) (M = Re, (99m)Tc) complexes can be applied to the development of target-specific radiopharmaceuticals because, in principle, any bioactive pharmacophore bearing an amine group can be used in the place of the model amine nucleophiles.     

Investigation of isomer formation upon coordination of bifunctional histidine analogues with 99mTc/Re(CO)3

Histidine is a convenient tridentate chelator used in the synthesis of technetium-99m radiopharmaceuticals, as it can be pendantly attached to a biomolecule for molecular imaging applications. Once coordinated, it forms a neutral complex that is capable of forming diastereomers at the alpha amine of the histidine. This is demonstrated through the synthesis and characterization of four different histidine chelators; three small molecule chelators, which consist of a benzylated histidine at the alpha amine, and one modified dipeptide, containing a phenylalanine derivative at the C-terminus and a histidine at the N-terminus. Upon rhenium coordination, two products are observed, each having the desired exact mass of the metal-containing species. The two products have been characterized through LC-MS, (1)H, gCOSY, NOESY and ROESY NMR experiments, and the relative stereochemistry determined. The implications of diastereomer formation when using this chelation system for creating molecular imaging agents is also discussed.     

一种新的潜在心肌显像剂[99mTc(CO)3(TBI)3]+的制备及其生物分布研究

以[99mTcO4]-为起始物,在0.1MPa下制备了中间体[99mTc(CO)3(H2O)1]+,通过配体交换反应,叔丁基异腈(TBI)配体取代该配合物中的3个水分子,制得一种标记率大于90%的[99mTc(CO)1(TBI)1]+配合物.该配合物在室温下放置6h以上,标记率无明显变化.在正常昆明小鼠体内的生物分布实验结果表明,[99mTc(CO)3(TBI)3]+具有较高的心肌摄取,且滞留也相当好,在静脉注射5min和60min后时的心肌摄取值分别为(19.07±0.81)%(ID/g)和(18.24±2.41)%(ID/g).该配合物的非靶本底摄取较低,注射60min后的心/肝、心/肺和心/血摄取比值分别为1.02,5.83和23.69,有望发展为一种新的心肌显像剂.     

Derivatization of glucose and 2-deoxyglucose for transition metal complexation: substitution reactions with organometallic 99mTc and Re precursors and fundamental NMR investigations

Synthetic strategies for the bifunctionalization of glucose and 2-deoxyglucose at position C-1 for transition metal coordination are reported. In particular organometallic technetium and rhenium complexes for potential use in diagnostic nuclear medicine were synthesized and investigated. Specifically, a common iminodiacetic acid (IDA) moiety was O-glycosidically connected through an ethylene spacer group to produce the pure alpha- (in case of 2-deoxyglucose) and beta-anomer (in case of glucose). Reaction of the sugar derivatives with the organometallic precursor [M(H2O)3(CO)3]+ (M = 99mTc, Re) produced single products in high yield, which are water-soluble and water-stable. The displacement of the three water molecules of the metal precursor and thus the tridentate coordination of the metal-tricarbonyl core exclusively via the amine and the two carboxylic acid functionalities of the IDA chelate was verified by means of 1D and 2D 1H NMR spectroscopy, mass spectrometry, and IR spectroscopy. The radioactive-labeled products (99mTc) proved their excellent stability in vitro in physiological phosphate buffer (pH = 7.4) and human plasma over a period of 24 h at 37 degrees C.     

Conjugation of a novel histidine derivative to biomolecules and labelling with [99mTc(OH2)3(CO)3]+

The new histidine derivative 3-[1-[3-(9H-fluoren-9-ylmethoxycarbonylamino)-propyl]-1H-imidazol-4-yl]-2-(3-trimethylsilanyl-ethylcarboxyamino)-propionic acid methyl ester (7) has been prepared via alkylation of the histidine urea derivative (7S)-5,6,7,8-tetrahydro-7-(methoxycarbonyl)-5-oxoimidazo-[1,5-c]-pyrimidine (2) with Fmoc-protected 3-iodopropyl-amine, followed by ring opening with 2-trimethylsilylethanol. After Fmoc cleavage by HNEt2, the histidine amine derivative was coupled to biotin, to the pentapeptide leucine-enkephalin and to Vitamin B12-b-acid by amide formation, employing TBTU as the coupling reagent. In order to make the histidine accessible for labelling, the teoc protecting group was removed by either NBu4F (for the biotin conjugate) or by TFA (for the enkephalin and B12 conjugates). Reaction of a 10(-4) M solution of the bioconjugates with [99mTc(H2O)3(CO)3]+ at 50 degrees C for 30 min led to the formation of one single new peak in the HPLC radiochromatogram in each case, confirming quantitative labelling of the respective biomolecules. To assess the nature of the labelled compounds, the rhenium analogues with Re(CO)3 were also synthesised and similar retention times confirmed the identity with the 99mTc labelled conjugates.     

Synthesis and biological evaluation of novel 99mTc-oxo and 99mTc-nitrido complexes with phenylalanine dithiocarbamate for tumor imaging

In this study, phenylalanine dithiocarbamate (PHEDTC) ligand was successfully synthesized and then radiolabeled with [^99mTcO]³⁺ core and [^99mTc≡N]²⁺ core to produce ^99mTcO–PHEDTC and ^99mTcN–PHEDTC, respectively. Both complexes were prepared with high radiochemical purity and had good stability. The partition coefficient results showed they were hydrophilic, while ^99mTcN–PHEDTC was more hydrophilic than ^99mTcO–PHEDTC. The biodistribution study in mice bearing S 180 tumor showed that ^99mTcO–PHEDTC and ^99mTcN–PHEDTC had high tumor uptake at 2 h post-injection, 1.91 and 1.21, respectively. The good uptake and retention in tumor together with favorable tumor-to-muscle ratios make them promising candidates for further evaluation as potential tumor imaging agents.