Dramatic effect of the tridentate ligand on the stability of 99mTC "3 + 1" oxo complexes bearing arylpiperazine derivatives.

Mixed-ligand model complexes of general formula [(99m)Tc(O)(kappa(3)-PNX)(kappa(1)-SPh))] [X = O (1a), S (2a)] were prepared in a one-step procedure from [(99m)TcO(4)(-)] using stannous chloride as reducing agent. Stability studies and challenge experiments with glutathione showed that complex 2a presented promising features for pursuing animal studies. The activity in the brain (% dose injected/organ) at 5 min (0.14% +/- 0.03) and 120 min (0.11% +/- 0.02) pi encouraged the synthesis of several mixed-ligand "3 + 1" oxo complexes of general formula [M(O)(kappa(3)-PNS)(kappa(1)-SL))] (M = (99m)Tc, 3a-6a, Re, 3-6), in which the tridentate ligand is the heterofunctionalized phosphine 2-(diphenylphosphanyl)-N-(2-thioethyl)benzamide (PNS) and the co-ligands are different arylpiperazine derivatives (HSL1-HSL4). The (99m)Tc complexes have been characterized by comparison of their retention times in the HPLC chromatogram (gamma-detection) with the retention times of the analogous Re complexes (UV detection at 254 nm). The (99m)Tc complexes, obtained with radiochemical purity higher than 95%, after HPLC purification, are stable in saline, 0.01 M PBS (pH 7.4), rat plasma (4 h, 37 degrees C), and glutathione (10 mM solutions, 2h, 37 degrees C). Binding affinity and selectivity for 5-HT(1A) receptors (relative to the 5-HT(2A) receptor) were determined, complex 5 demonstrating the best values (IC(50) for the 5-HT(1A) 2.35 +/- 0.02 nM; competitor 5-HT(2A) 372 +/- 11 nM). Biodistribution and stability studies in mice indicated a preferred hepatobiliary excretion, a high in vivo stability, but a poor brain uptake.

Novel tumortropic ester derivatives of 99mTc(V) mesodimercapto succinic acid with low affinity for bone tissue.

Starting from our previous finding that 99mTc(V) dimercaptosuccinic acid (99mTc(V)-DMSA), a useful agent for the localization of osteosarcoma and bone metastases, loses its bone affinity when one ester group is introduced into the complex we studied the impact of esterification in more detail. This paper reports on the evaluation of various ester complexes of 99mTc(V)-DMSA in rats and tumour-bearing nude mice with regard to their tumour retention and improvement of the tumour to tissue ratios. The distribution patterns of the complexes [99mTcO(DMSA)2]- (A), [99mTcO(DMSA/DMSEt)]- (B) and [99mTcO(DMSEt)2]- (C) are gradually changed with the number of ester groups in the anionic complex. However, the asymmetric diester complex [99mTcO(DMSA/DMSEt2)]- (D) is very slowly cleared, especially from the blood of nude mice. Moreover, this complex differs significantly from the symmetrical complex C in its elimination behaviour from the liver and kidneys. The tumour uptake is maintained with complexes that contain one or two non-hydrolysable ester functions. Preliminary biodistribution studies of the monoethyl and diethyl ester complexes B, C and D in comparison with A in tumour-bearing nude mice showed similar uptake into the human squamous cell carcinoma (FaDu) as well as into the human colonic cell carcinoma (HT29) of nude mice. The low bone accumulation of B, C and D results in excellent tumour-to-bone ratios, e.g., approx. 3:1 for the ester complex B compared to approx. 1:2 for complex A. Differences were observed in the accumulation and elimination behaviour of the complexes A and B in various bone structures of rats. The age-dependent uptake of A and B was compared in long bone (femur) and in cranial bone of rats. The results suggest that 99mTc(V)-DMSA complexes that contain a functional ester, and their 188Re analogues, may be superior to 99mTc(V)/188Re(V)-DMSA in diagnostic and therapeutic nuclear medicine.

Tc and Re chelates of 8alpha-amino-6-methyl-ergoline: synthesis and affinity to the dopamine D2 receptor.

The influence of structural changes at the 8alpha-amino position of 8alpha-amino-6-methyl-ergoline on the lipophilicity and affinity to the D2 receptor was studied. 8alpha-amino-6-methyl-ergoline (1) was converted into the derivatives (2a-f) by mercaptoacetylation of the amino group to make it possible to prepare the rhenium and technetium complexes (3, 4a,b). Binding tests on cloned human dopamine D2 receptors show that the affinities of the coordination compounds (IC50 values between 50 and 240 nM) are less than those of the derivatives 2a-f (IC50=3-50 nM) but more than those of the parent compound 1. Biodistribution studies of the Tc complexes 4a,b performed on Wistar rats show a slow blood clearance with substantial accumulation and retention in the liver and kidneys and low brain uptake.

Synthesis, biological and autoradiographic evaluation of a novel Tc-99m radioligand derived from WAY 100635 with high affinity for the 5-HT(1A) receptor and the alpha1-adrenergic receptor.

This paper reports the synthesis, biological evaluation, in vitro and ex vivo autoradiography of the first Tc-99m ligand with subnanomolar affinity for the 5-HT(1A) receptor and a remarkably high affinity for the alpha1-adrenergic receptor. The neutral "3+1" mixed-ligand complex combines 4-(6-mercaptohexyl)-1-(2-methoxyphenyl)piperazine as monodentate and 3-(N-methyl)azapentane-1,5-dithiol as tridentate unit with oxotechnetium(V). The analogous rhenium complex was synthesized for complete structural characterization and used in receptor binding assays. In competition experiments both complexes display subnanomolar affinity for the 5-HT(1A) receptor (IC(50)0.24 nM for Re, 0.13 nM for Tc) but also very high affinities for the alpha1-adrenergic receptor (IC(50) 0.05 nM for Re, 0.03 nM for Tc). Biodistribution studies show a brain uptake in rat of 0.22% ID five minutes post injection. In vitro autoradiographic studies in rat brain and postmortem human brain indicate accumulation of the Tc-99m complex in brain areas which are rich in 5-HT(1A) receptors or in alpha1-adrenergic receptors. This in vitro enrichment can be blocked respectively by the 5-HT(1A) receptor agonist 8-OH-DPAT or by prazosin hydrochloride, an alpha1-adrenergic receptor antagonist. Ex vivo autoradiographic studies in rats show a slight accumulation of the Tc-99m complex in 5-HT(1A) receptor-rich areas of the brain, which could not be blocked, as well as in regions rich in alpha1-adrenergic receptors, which could be blocked by prazosin hydrochloride.

Synthesis and biological evaluation of technetium(III) mixed-ligand complexes with high affinity for the cerebral 5-HT(1A) receptor and the alpha1-adrenergic receptor.

Tc(III) and Re(III) complexes [M(NS(3))(CNR)] (M = Re, 99mTc, NS(3) = 2,2',2"-nitrilotris(ethanethiol), CNR = functionalized isocyanide bearing a derivative of WAY 100635) have been synthesized and characterized. Re was used as Tc surrogate for chemical characterization and in vitro receptor-binding studies. For two representatives subnanomolar affinities for the 5-HT(1A) as well as for the alpha1-adrenergic receptor were reached. Biodistribution studies in rats of the 99mTc complexes showed brain uptakes between 0.3 and 0.5% ID/organ (5 min p.i.). In vitro autoradiography of one 99mTc representative in sections of post mortem human brain indicate its accumulation in 5-HT(1A) receptor-rich brain regions. However, addition of the specific 5-HT(1A) receptor agonist 8-OH-DPAT as well as the alpha1-adrenoceptor antagonist prazosin could not substantially block this tracer accumulation. A preliminary SPET study in a monkey showed negligible brain uptake.

Mixed-ligand technetium(III) complexes with tetradendate/monodendate NS(3)/isocyanide coordination: a new nonpolar technetium chelate system for the design of neutral and lipophilic complexes stable in vivo.

Starting from the tripodal ligand 2,2',2' '-nitrilotris(ethanethiol) (NS(3)) and isocyanides (CNR) as co-ligands, neutral mixed-ligand technetium(III) complexes of the general formulation [Tc(NS(3))(CNR)] have been synthesized and characterized. The (99)Tc complexes can be( )()obtained by a two-step reduction/substitution procedure starting from [TcO(4)](-) via the phosphine-containing precursor complex [Tc(NS(3))(PMe(2)Ph)]. As shown by X-ray structural analyses, the complexes adopt a nearly ideal trigonal-bipyramidal geometry with the trigonal plane formed by the three thiolate sulfurs of the tripodal ligand. The central nitrogen atom of the chelate ligand and the monodendate isocyanides occupy the apical positions. The no-carrier-added preparation of the corresponding (99m)Tc complexes was performed by a one-step procedure starting from (99m)[TcO(4)](-) with stannous chloride as reducing agent. Biodistribution studies in the rat demonstrated for the nonpolar, lipophilic compounds a significant initial brain uptake. In vitro challenge experiments with glutathione clearly indicated that no transchelation reaction occurs. Furthermore, there were no indications for reoxidation of Tc(III) to Tc(V) species or pertechnetate. We propose this type of complexes as a useful tool in the design of lipophilic (99m)Tc or (186)Re/(188)Re radiopharmaceuticals.

Ligand-exchange reaction of labile "3 + 1"99mTc(V) complexes with SH group-containing proteins.

The reactivity of labile 3 + 1 mixed-ligand 99mTc complexes of the type [99mTcO(SES)(RS)] with SES being a tridentate dithiol ligand and glutathione or dimethylcysteamine as monodentate ligands RSH towards proteins was investigated in vitro and in vivo. It was found that the complexes undergo reversible transchelation reactions with SH group-containing components of blood such as albumin or haemoglobin. High labelling yields were obtained when 3 + 1 complexes with the tridentate SSS ligand were used. The biodistribution of blood proteins labelled by ligand-exchange reaction with the [99TcO(SSS)] or [99mTcO(SNMeS)] core was studied and compared with the in vivo distribution of the labile 3 + 1 complexes containing glutathione as monodentate ligand.

Permeation studies in vitro and in vivo of potential radiopharmaceuticals with affinity to neuro receptors.

PURPOSE: To check the influence of structural characteristics on their permeation through the blood-brain barrier (BBB), a set of radioactive [99mTc]chelates bearing amine groups was synthesized and tested in vitro as well as in vivo. METHODS: Compounds with different log P and pKa values were obtained by complex forming reactions of [99mTc]pertechnetate with varying substituents. Transport was studied in rats and mice, as well as in an ECV304 cell culture model. RESULTS: In vitro higher permeation was found for compounds with electron attracting substituents in beta-position to the amine group (pKa values 7.4 to 8.3) than for those with more basic amine groups (pKa values > 8.9) even for similar log DH 7.4. In vivo brain uptake between 0.8 and 4.8% of the injected dose (ID) per organ was found for the former, whereas <0.4% ID were present for the latter. CONCLUSIONS: Three structurally diverse classes of [99mTc]chelates showed distinct patterns with regard to brain uptake in vivo and BBB permeability in vitro which could not be predicted by their lipophilicity alone. The close correlation between the data from rats and mice and those obtained with cell cultures render the ECV304 cells an attractive model for the screening of new compounds.

Radiochemical synthesis and tissue distribution of Tc-99m-labeled 7alpha-substituted estradiol complexes.

The diagnosis and staging of breast cancer could be improved by the development of radiopharmaceutical imaging agents that provide a noninvasive determination of the estrogen receptor (ER) status of tumor cells. Agents labeled with (99m)Tc would be especially valuable in this regard. In attempting to achieve this goal, we synthesized four (99m)Tc-labeled 7alpha-substituted estradiol complexes. One complex utilizes the "3+1" mixed ligand design to introduce the Tc metal, whereas the other three took advantage of the cyclopentadienyltricarbonylmetal (CpTM) design. The Tc moieties were attached to the 7alpha position of estradiol with a hexyl tether, a monoether tether, or a polyether tether. The corresponding rhenium compounds have binding affinities for the ER of 20-45% compared with estradiol. Radiochemical yields of the (99m)Tc-labeled compounds ranged from approximately 15% for the CpT-Tc complexes to 95% for the 3 + 1 inorganic complex. Tissue distribution studies in immature female rats showed low nonreceptor-mediated uptake in the target organs and high uptake in nontarget organs such as the liver and fat. These complexes represent the first time that estradiol has been labeled at the 7alpha position with (99m)Tc and provide a further refinement of our understanding of ligand structure-binding affinity correlations for the ER.

Chemical and biological characterization of technetium(I) and Rhenium(I) tricarbonyl complexes with dithioether ligands serving as linkers for coupling the Tc(CO)(3) and Re(CO)(3) moieties to biologically active molecules.

The organometallic precursor (NEt(4))(2)[ReBr(3)(CO)(3)] was reacted with bidendate dithioethers (L) of the general formula H(3)C-S-CH(2)CH(2)-S-R (R = -CH(2)CH(2)COOH, CH(2)-C&tbd1;CH) and R'-S-CH(2)CH(2)-S-R' (R' = CH(3)CH(2)-, CH(3)CH(2)-OH, and CH(2)COOH) in methanol to form stable rhenium(I) tricarbonyl complexes of the general composition [ReBr(CO)(3)L]. Under these conditions, the functional groups do not participate in the coordination. As a prototypic representative of this type of Re compounds, the propargylic group bearing complex [ReBr(CO(3))(H(3)C-S-CH(2)CH(2)-S-CH(2)C&tbd1;CH)] Re2 was studied by X-ray diffraction analysis. Its molecular structure exhibits a slightly distorted octahedron with facial coordination of the carbonyl ligands. The potentially tetradentate ligand HO-CH(2)CH(2)-S-CH(2)CH(2)-S-CH(2)CH(2)-OH was reacted with the trinitrato precursor [Re(NO(3))(3)(CO)(3)](2-) to yield a cationic complex [Re(CO)(3)(HO-CH(2)CH(2)-S-CH(2)CH(2)-S-CH(2)CH(2)-OH)]NO(3) Re8 which shows the coordination of one hydroxy group. Re8 has been characterized by correct elemental analysis, infrared spectroscopy, capillary electrophoresis, and X-ray diffraction analysis. Ligand exchange reaction of the carboxylic group bearing ligands H(3)C-S-CH(2)CH(2)-S-CH(2)CH(2)-COOH and HOOC-CH(2)-S-CH(2)CH(2)-S-CH(2)-COOH with (NEt(4))(2)[ReBr(3)(CO)(3)] in water and with equimolar amounts of NaOH led to complexes in which the bromide is replaced by the carboxylic group. The X-ray structure analysis of the complex [Re(CO)(3)(OOC-CH(2)-S-CH(2)CH(2)-S-CH(2)-COOH)] Re6 shows the second carboxylic group noncoordinated offering an ideal site for functionalization or coupling a biomolecule. The no-carrier-added preparation of the analogous (99m)Tc(I) carbonyl thioether complexes could be performed using the precursor fac-[(99m)Tc(H(2)O)(3)(CO)(3)](+), with yields up to 90%. The behavior of the chlorine containing (99m)Tc complex [(99m)TcCl(CO)(3)(CH(3)CH(2)-S-CH(2)CH(2)-S-CH(2)CH(3))] Tc1 in aqueous solution at physiological pH value was investigated. In saline, the chromatographically separated compound was stable for at least 120 min. However, in chloride-free aqueous solution, a water-coordinated cationic species Tc1a of the proposed composition [(99m)Tc(H(2)O)(CO)(3)(CH(3)CH(2)-S-CH(2)CH(2)-S-CH(2)CH(3))](+) occurred. The cationic charge of the conversion product was confirmed by capillary electrophoresis. By the introduction of a carboxylic group into the thioether ligand as a third donor group, the conversion could be suppressed and thus the neutrality of the complex preserved. Biodistribution studies in the rat demonstrated for the neutral complexes [(99m)TcCl(CO)(3)(CH(3)CH(2)-S-CH(2)CH(2)-S-CH(2)CH(3))] Tc1 and [(99m)TcCl(CO)(3)(CH(2)-S-CH(2)CH(2)-S-CH(2)-C&tbd1;CH)] Tc2 a significant initial brain uptake (1.03 +/- 0.25% and 0.78 +/- 0.08% ID/organ at 5 min. p.i.). Challenge experiments with glutathione clearly indicated that no transchelation reaction occurs in vivo.

Synthesis and autoradiographic evaluation of a novel high-affinity Tc-99m ligand for the 5-HT2A receptor.

The synthesis and in vitro autoradiography of a novel Tc-99m ligand with subnanomolar affinity to the 5-HT2A receptor is reported. The complex combines the 4-(4-fluoro)-benzoyl piperidine portion derived from the 5-HT2A receptor antagonist ketanserin with a neutral oxotechnetium(V) chelate in form of a mixed ligand "3 + 1" unit containing the SNS/S donor set. The analogous rhenium compound has been synthesized as a surrogate for the Tc-99m complex for use in receptor binding assays and for complete structural characterization. In competition experiments the Tc-99 complex as well as its Re analogue display subnanomolar affinity toward the 5-HT2A receptor (Ki 0.44 nM for Tc, 0.25 nM for Re). The subnanomolar 5-HT2A receptor binding of the Re complex was confirmed by functional in vitro antagonism of contractile effects evoked by 5-HT in rat arterial tissue. Re 1 inhibited 5-HT-induced, 5-HT2A receptor-mediated contractions of isolated rat tail artery in a competitive fashion and possessed nanomolar affinity (pA2 = 9.08, pA2 representing the negative decadic logarithm of the Re 1/5-HT2A-receptor dissociation constant [mol/L]). Like ketanserin, Re 1 displayed moderate affinity for adrenergic alpha1D (pA2 = 8.23) and histamine H1 receptors (pA2 = 8.00), and was >600-fold up to 10,700-fold less active at several neurotransmitter receptor subtypes. In vitro autoradiographic studies clearly indicate the accumulation of the Tc-99m compound in 5-HT2A-receptor-rich areas of the brain. This enrichment can be blocked by 5-HT2A receptor antagonists such as mianserin and ketanserin and is therefore specific.

Stability versus reactivity of "3+1" mixed-ligand technetium-99m complexes in vitro and in vivo.

"3+1" technetium-99m mixed-ligand complexes as relevant to the development of a third generation of 99mTc radiopharmaceuticals were investigated in vivo and in vitro in the blood of rats. Surprisingly, in whole blood the complexes, which proved to be stable in saline, PBS of pH 7.4 and in plasma, were converted into two radioactive, more hydrophilic metabolites. Small structural differences in the complex molecule have a profound influence on the rate of metabolism of the complexes. To obtain an understanding of this unexpected reactivity, transchelation reactions with glutathione (GSH) were hypothesized and this hypothesis substantiated by challenge experiments carried out with a series of 99mTc and analogous rhenium complexes and GSH. In vitro studies in human plasma, whole blood and erythrocytes also revealed conversion of the complexes, though, at a much slower rate. Structural parameters influencing the stability of the complexes and consequences for the radiopharmaceutical design are discussed.

Structural modification of receptor-binding technetium-99m complexes in order to improve brain uptake.

Low brain uptake is a generally accepted problem in developing technetium-99m brain receptor imaging agents. For a class of potential 5-HT2A receptor-binding agents we tried to improve the original low brain uptake of 0.4% injected dose (ID) in rats 5 min p.i. by modifying the lipophilic properties of the molecules. Because of the presence of a protonable nitrogen, which according to the pKa value leads to ionization of the molecule at blood pH, the pKa value was considered to be the parameter most suitable for adjustment of lipophilicity. Insertion of ether-oxygen in the molecule of five candidates lowers the apparent pKa value from 10.0 to 8.3 and dramatically increases the brain uptake to 1.3% ID at 5 min. The direct relationship between brain uptake and apparent pKa cannot be simply explained by the increase in the pKa-governed proportion of the neutral species.

Technetium(V) and rhenium(V) complexes for 5-HT2A serotonin receptor binding: structure-affinity considerations.

Starting from the lead structure of ketanserin, a prototypic serotonin (5-HT) antagonist, new oxotechnetium(V) and oxorhenium(V) complexes were synthesized that are able to compete with [3H]ketan-serin in receptor-binding assays. To imitate organic 5-HT2 receptor ligands, fragments of ketanserin were combined with chelate moieties. Neutral compounds of the general formula [MOL1L2] (M = Tc, Re; L1 = HS-CH2CH2-S-CH2CH2-SH, N-(2-mercaptophenyl)salicylideneimine, N-(2-mercaptoethyl)-salicylideneimine, 3-(2-([N,N-bis(2-mercapto-S-ethyl)]-amino)ethyl)-2,4-(1H, 3H)-quinazolinedione and L2 = HS-R with R = subst. alkyl) were prepared by common action of a Tc(V) or Re(V) precursor with a mixture of equimolar amounts of a tridentate ligand L1 and a monodentate thiolate L2 bearing fragments of the lead structure. Lipophilic complexes consisting of a small S4 thiolate/thioether chelate unit, protonable nitrogen-containing spacer, and simple benzyl moiety significantly inhibited the specific binding of [3H]ketan-serin with IC50 values between 10 and 50 nM.

Preparation of [99mTc(DMPE)2Cl2]+ for myocardial imaging by using Fe-(DMPE) complexes.

The preparation of [99mTc(1,2-bis(dimethylphosphino)ethane)2Cl2]+ ([99mTc(DMPE)2Cl2]+) for imaging the myocardium is investigated. Starting from Fe(III)- or Fe(II)-DMPE and 99mTcO4- different preparation variants are compared. In these reactions either ascorbic acid or DTPA serves as an agent for complexing iron. A simple procedure using lyophilized initial components is represented. Its application yields [99mTc(DMPE)2Cl2]+ with a radiochemical purity of more than 95%. Organ distribution studies performed in rats emphasize the high myocardial accumulation of this preparation.