Ferenc K. Kálmán

Mn(II) complexes of novel hexadentate AAZTA-like chelators: a solution thermodynamics and relaxometric study

Three novel chelators based on the 6-amino-6-methylperhydro-1,4-diazepine scaffold and possessing three pendant N-acetic or N-α-methylacetic acid have been synthesised. The ligands contain six donor atoms for complexation of Mn(II) ions and thus potentially leave an additional site for coordination of a water molecule. The protonation constants of the ligands and the stability constants of their complexes formed with Mn(II) ion were determined by pH-potentiometric titrations in 0.15 M NaCl solution at 25 °C and compared to those of the parent AAZTA ligand (AAZTA = 6-amino-6-methylperhydro-1,4-diazepine tetraacetic acid). In spite of the similar value of the total basicity (Σlog K), the values of the stability constants of the Mn(II)AAZTA-like complexes are more than three orders of magnitude lower than that of MnAAZTA (log K(MnL) = 14.19). A detailed (1)H and (17)O NMR relaxometric study was carried out on the Mn(II) complexes in aqueous solution as a function of pH, temperature and magnetic field strength. The (1)H NMRD profiles of all the complexes show a similar shape, typical of low-molecular weight systems, but amplitudes that markedly differ to indicate a different degree of hydration. A similar behaviour is shown by the (17)O NMR transverse relaxation rates and chemical shift data as a function of temperature. The experimental data can be rationalised by considering the presence in solution of a mixture of two isomeric species differing in coordination number (7 and 6) and in the number (1 and 0) of bound water molecules. Whereas this type of coordination equilibrium has been previously reported for lanthanide(III) complexes, it is observed for the first time on Mn(II) chelates.

Lanthanide(III) complexes of tris(amide) PCTA derivatives as potential bimodal magnetic resonance and optical imaging agents

Lanthanide complexes of two tris(amide) derivatives of PCTA were synthesized and characterized. The relaxometric and luminescence properties of their lanthanide complexes were investigated as bimodal magnetic resonance (MR) and optical imaging agents. Luminescence studies show that one of the Tb(III) complexes dimerizes in solution at low millimolar concentrations, whereas the other may have a higher than expected coordination number in solution. The corresponding Gd(III) complexes display unusually high T(1) relaxivities and enhanced kinetic inertness compared to GdPCTA. These features suggest that these new chelates may be suitable for in vivo applications. The fast water-exchange rates observed for these complexes make them unsuitable as paramagnetic chemical exchange saturation transfer (PARACEST) agents.

Synthesis and Characterization of a Hypoxia‐Sensitive MRI Probe

Tissue hypoxia occurs in pathologic conditions, such as cancer, ischemic heart disease and stroke when oxygen demand is greater than oxygen supply. An imaging method that can differentiate hypoxic versus normoxic tissue could have an immediate impact on therapy choices. In this work, the gadolinium(III) complex of 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) with a 2-nitroimidazole attached to one carboxyl group via an amide linkage was prepared, characterized and tested as a hypoxia-sensitive MRI agent. A control complex, Gd(DO3A-monobutylamide), was also prepared in order to test whether the nitroimidazole side-chain alters either the water proton T(1) relaxivity or the thermodynamic stability of the complex. The stabilities of these complexes were lower than that of Gd(DOTA)(-) as expected for mono-amide derivatives. The water proton T(1) relaxivity (r(1)), bound water residence lifetime (τ(M)) and rotational correlation time (τ(R)) of both complexes was determined by relaxivity measurements, variable temperature (17) O NMR spectroscopy and proton nuclear magnetic relaxation dispersion (NMRD) studies. The resulting parameters (r(1) =6.38 mM(-1)  s(-1) at 20 MHz, τ(M) =0.71 μs, τ(R) =141 ps) determined for the nitroimidazole derivative closely parallel to those of other Gd(DO3A-monoamide) complexes of similar molecular size. In vitro MR imaging experiments with 9L rat glioma cells maintained under nitrogen (hypoxic) versus oxygen (normoxic) gas showed that both agents enter cells but only the nitroimidazole derivative was trapped in cells maintained under N(2) as evidenced by an approximately twofold decrease in T(1) measured for hypoxic cells versus normoxic cells exposed to this agent. These results suggest that the nitroimidazole derivative might serve as a molecular reporter for discriminating hypoxic versus normoxic tissues by MRI.

Picolinate-containing macrocyclic Mn2+ complexes as potential MRI contrast agents.

We report the synthesis of the ligand Hnompa (6-((1,4,7-triazacyclononan-1-yl)methyl)picolinic acid) and a detailed characterization of the Mn(2+) complexes formed by this ligand and the related ligands Hdompa (6-((1,4,7,10-tetraazacyclododecan-1-yl)methyl)picolinic acid) and Htempa (6-((1,4,8,11-tetraazacyclotetradecan-1-yl)methyl)picolinic acid). These ligands form thermodynamically stable complexes in aqueous solution with stability constants of logKMnL = 10.28(1) (nompa), 14.48(1) (dompa), and 12.53(1) (tempa). A detailed study of the dissociation kinetics of these Mn(2+) complexes indicates that the decomplexation reaction at about neutral pH occurs mainly following a spontaneous dissociation mechanism. The X-ray structure of [Mn2(nompa)2(H2O)2](ClO4)2 shows that the Mn(2+) ion is seven-coordinate in the solid state, being directly bound to five donor atoms of the ligand, the oxygen atom of a coordinated water molecule and an oxygen atom of a neighboring nompa(-) ligand acting as a bridging bidentate carboxylate group (μ-η(1)-carboxylate). Nuclear magnetic relaxation dispersion ((1)H NMRD) profiles and (17)O NMR chemical shifts and transverse relaxation rates of aqueous solutions of [Mn(nompa)](+) indicate that the Mn(2+) ion is six-coordinate in solution by the pentadentate ligand and one inner-sphere water molecule. The analysis of the (1)H NMRD and (17)O NMR data provides a very high water exchange rate of the inner-sphere water molecule (kex(298) = 2.8 × 10(9) s(-1)) and an unusually high value of the (17)O hyperfine coupling constant of the coordinated water molecule (AO/ℏ = 73.3 ± 0.6 rad s(-1)). DFT calculations performed on the [Mn(nompa)(H2O)](+)·2H2O system (TPSSh model) provide a AO/ℏ value in excellent agreement with the one obtained experimentally.

Mn-loaded apoferritin: a highly sensitive MRI imaging probe for the detection and characterization of hepatocarcinoma lesions in a transgenic mouse model

Mn-Apo is a highly sensitive MRI contrast agent consisting of ca. 1000 manganese atoms entrapped in the inner cavity of apoferritin. Part of the metallic payload is in the form of Mn(2+) ions that endow the nano-sized system with a very high relaxivity that can be exploited to detect hepatocellular carcinoma in mice. Cellular studies showed that Mn-Apo is readily taken up by normal hepatocytes via the ferritin transporting route. Conversely, hepatoma cells (HTC) displayed a markedly reduced ability to entrap Mn-Apo from the culture medium. The i.v. administration of Mn-Apo into C57BL/6 J mice resulted in a marked liver tissue hyperintensity in T(1)-weighted MR image 20 min after injection. When injected into HBV-tg transgenic mice that spontaneously develop hepatocellular carcinoma (HCC), Mn-Apo allowed a clear delineation of healthy liver tissue and tumor lesions as hyperintense and hypointense T(1)-weighted MR images, respectively. Immunohistochemistry analysis correlated Mn-Apo cellular uptake to SCARA5 receptor expression. When the MRI contrast induced by Mn-Apo was compared with that induced by Gd-BOPTA (a commercial contrast agent known to enter mouse hepatocytes through organic anion transporters) it was found that only some of the lesions were detected by both agents while others could only be visualized by one of the two. These results suggest that Mn-Apo may be useful to detect otherwise invisible lesions and that the extent of its uptake directly reports the expression/regulation of SCARA5 receptors. Mn-Apo contrast-enhanced MR images may therefore contribute to improving HCC lesion detection and characterization.

Aryl-Phosphonate Lanthanide Complexes and Their Fluorinated Derivatives: Investigation of Their Unusual Relaxometric Behavior and Potential Application as Dual Frequency 1H/19F MRI Probes

A series of low molecular weight lanthanide complexes were developed that have high (1)H longitudinal relaxivities (r1) and the potential to be used as dual frequency (1)H and (19)F MR probes. Their behavior was investigated in more detail through relaxometry, pH-potentiometry, luminescence, and multinuclear NMR spectroscopy. Fitting of the (1)H NMRD and (17)O NMR profiles demonstrated a very short water residence lifetime (<10 ns) and an appreciable second sphere effect. At lower field strengths (20 MHz), two of the complexes displayed a peak in r1 (21.7 and 16.3 mM(-1)  s(-1)) caused by an agglomeration, that can be disrupted through the addition of phosphate anions. NMR spectroscopy revealed that at least two species are present in solution interconverting through an intramolecular binding process. Two complexes provided a suitable signal in (19)F NMR spectroscopy and through the selection of optimized imaging parameters, phantom images were obtained in a MRI scanner at concentrations as low as 1 mM. The developed probes could be visualized through both (1)H and (19)F MRI, showing their capability to function as dual frequency MRI contrast agents.

Physico-chemical properties of MnII complexes formed with cis- and trans-DO2A: thermodynamic, electrochemical and kinetic studies

SYNOPSIS MnII complexes formed with cis- and trans-DO2A (DO2A=1,4,7,10-tetraazacyclododecane-1,4 (or 1,7) -diacetic acid) chelators were investigated by pH-potentiometry, 1H relaxometry, UV-vis spectrophotometry and cyclic voltammetry. The physico-chemical characteristics of MnII complexes of these structure isomers do not differ dramatically, however the cis-DO2A platform has better potential for further development. Manganese (MnII) is a promising alternative to gadolinium (GdIII) as a magnetic resonance imaging (MRI) agent. Unlike gadolinium, this biogenic metal might be better tolerated by the body, reducing the risk of toxicity associated with dissociation of the complex. Herein we report detailed equilibrium and kinetic studies performed with MnII complexes of 1,4,7,10-tetraazacyclododecane-1,4-diacetic acid (1,4-DO2A or cis-DO2A) and 1,4,7,10-tetraazacyclododecane-1,7-diacetic acid (1,7-DO2A or trans-DO2A). The protonation constants of the ligands as well as stability constants of their MnII complexes have been determined by pH-potentiometry. The stability constants of [Mn(cis-DO2A)] are slightly higher than those of [Mn(trans-DO2A)] (log KMnL=15.68 and 15.22, respectively). Cyclic voltammetric (CV) experiments performed on [Mn(cis-DO2A)] and [Mn(trans-DO2A)] revealed quasireversible systems with a half-wave potential of +636 and +705mV versus Ag/AgCl, respectively. These values indicate that the MnII ion in these complexes is more stabilized against the oxidation than in [Mn(EDTA)]2-. The kinetic inertness of the complexes has been studied in transmetallation reactions with CuII or ZnII ions. Kinetic measurements indicate that both MnII complexes primarily undergo acid catalyzed dissociation and positions of the acetate pendant arms do not influence kinetic inertness. The inertness of these complexes is comparable to that of [Mn(NOTA)]- (NOTA=1,4,7-triazacyclononane-1,4,7-triacetic acid) and about twenty times lower than that of [Mn(DOTA)]2- (DOTA=1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid). In conclusion, [Mn(cis-DO2A)] displays some very interesting features (thermodynamic and redox stability as well as kinetic inertness) which makes this complex a promising platform for the development of more efficient MnII complexes as alternatives to Gd-based MRI agents.

Lanthanide(III) complexes of some natural siderophores: A thermodynamic, kinetic and relaxometric study

Stability constants of the complexes formed between the natural trihydroxamic acids desferrioxamine B (DFB) and desferricoprogen (DFC) with Nd(III), Gd(III) and Yb(III) ions were determined using pH-potentiometry. The equilibrium in these systems can be described by models containing mononuclear protonated (Ln(HL), Ln(H2L) and Ln(H3L)), deprotonated (LnL) and ternary hydroxo Ln(H-1L) complexes, but for both ligands dinuclear complexes of low stability were also detected. The stability constants for the Ln(HDFB)(+) complexes are 11.95 (Nd(III)), 13.16 (Gd(III)) and 14.67 (Yb(III)), while these values of the Ln(DFC) complexes are considerably higher (14.42 (Nd(III)), 15.14 (Gd(III)) and 16.49 (Yb(III))). The stability constants of the complexes of DFB and DFC are much lower than those of the Ln(L)3 complexes formed with some aromatic hydroxamic acids indicating that the relatively long spacer between the hydroxamic acid moieties in DFB and DFC is unfavorable for Ln(III) complexation. The relaxometric study conducted for the Gd(HDFB)(+) species revealed an interesting pH dependence of the relaxivity associated with a large hydration number (bishydrated complex) and fast water exchange (kex=(29.9±0.4)×10(6)s(-1)), which would be favorable for CA use. However the dissociation of Gd(HDFB)(+) is fairly fast (<2ms) under all conditions employed in the present work thus the kinetically labile Gd(HDFB)(+) is not suitable for in vivo CA applications. Some low stability ternary complexes were also detected with K(Gd(HDFB)(HCO3))=17.5±1.9 and K(Gd(HDFB)(Lactate))=8.4±3.2 but in the presence of citrate and phosphate ions the Gd(HDFB)(+) complex was found to dissociate.

Complexation of Ln3+ Ions with Cyclam Dipicolinates: A Small Bridge that Makes Huge Differences in Structure, Equilibrium, and Kinetic Properties

The coordination properties toward the lanthanide ions of two macrocyclic ligands based on a cyclam platform containing picolinate pendant arms have been investigated. The synthesis of the ligands was achieved by using the well-known bis-aminal chemistry. One of the cyclam derivatives (cb-tedpa(2-)) is reinforced with a cross-bridge unit, which results in exceptionally inert [Ln(cb-tedpa)](+) complexes. The X-ray structures of the [La(cb-tedpa)Cl], [Gd(cb-tedpa)](+), and [Lu(Me2tedpa)](+) complexes indicate octadentate binding of the ligands to the metal ions. The analysis of the Yb(3+)-induced shifts in [Yb(Me2tedpa)](+) indicates that this complex presents a solution structure very similar to that observed in the solid state for the Lu(3+) analogue. The X-ray structures of [La(H2Me2tedpa)2](3+) and [Yb(H2Me2tedpa)2](3+) complexes confirm the exocyclic coordination of the metal ions, which gives rise to coordination polymers with the metal coordination environment being fulfilled by oxygen atoms of the picolinate groups and water molecules. The X-ray structure of [Gd(Hcb-tedpa)2](+) also indicates exocyclic coordination that in this case results in a discrete structure with an eight-coordinated metal ion. The nonreinforced complexes [Ln(Me2tedpa)](+) were prepared and isolated as chloride salts in nonaqueous media. However, these complexes were found to undergo dissociation in aqueous solution, except in the case of the complexes with the smallest Ln(3+) ions (Ln(3+) = Yb(3+) and Lu(3+)). A DFT investigation shows that the increased stability of the [Ln(Me2tedpa)](+) complexes in solution across the lanthanide series is the result of an increased binding energy of the ligand due to the increased charge density of the Ln(3+) ion.

Taking the next step toward inert Mn2+ complexes of open-chain ligands: the case of the rigid PhDTA ligand

In line with our research to find inert Mn(II) complexes as contrast agents for magnetic resonance imaging, we have studied the aromatic-ring rigidified EDTA-analogue o-phenylenediamine-N,N,N′,N′-tetraacetic acid (PhDTA). The protonation constants (KHi) of PhDTA and stability constants of complexes formed between this open-chain ligand and several different biogenic metal ions (Ca2+, Mg2+, Zn2+, Cu2+, Mn2+) have been determined in 0.15 M NaCl at 25 °C and compared with the values reported in the literature previously. The protonation constants are lower than those of the corresponding cis- and trans-CDTA complexes, which might be attributed to the electron withdrawing effect of the phenylene group. The lower total basicity of the ligand leads to lower stability constants for all the examined metal complexes. On the contrary, we have found that the conditional stability constants of [Mn(PhDTA)]2− and [Mn(trans-CDTA)]2− are approximately the same, as both complexes are completely formed at pH 5 and their pM values are also comparable. The relaxivity of [Mn(PhDTA)]2− is nearly identical (r1 = 3.72 mM−1 s−1) to that determined previously for the [Mn(trans-CDTA)]2− complex (r1 = 3.62 mM−1 s−1), and its pH-dependence confirms the equilibrium model used for the fitting of the titration data. The results of the kinetic studies of the metal exchange reactions reveal that the [Mn(PhDTA)]2− complex possesses a slightly better dissociation kinetics profile than that of [Mn(trans-CDTA)]2−, which has been tested in vivo recently (including human injections). The half-life of the dissociation of the complex near physiological pH at 25 °C is 19 hours. By using the rate constant calculated for the dissociation (pH = 7.4, cCu2+ = 10 μM) and the half-life of excretion (1.6 hour), the ratio of the dissociated complex is estimated to represent 8% of the injected dose. DFT studies reveal that the metal coordination environment of [Mn(PhDTA)]2− is very similar to that of [Mn(EDTA)]2−, both containing an inner-sphere water molecule. Cyclic voltammetry studies indicate that [Mn(PhDTA)]2− is slightly more resistant towards oxidation to the Mn3+ complex than the EDTA analogue.