Fe(III) T1 MRI Probes Containing Phenolate or Hydroxypyridine-Appended Triamine Chelates and a Coordination Site for Bound Water.

Fe(III) complexes containing a triamine framework and phenolate or hydroxypyridine donors are characterized and studied as T1 MRI probes. In contrast to most Fe(III) MRI probes of linear chelates reported to date, the ligands reported here are pentadentate to give six-coordinate complexes with a coordination site for inner-sphere water. The crystal structure of the complex containing unsubstituted phenolate donors, Fe(L1)Cl, shows a six-coordinate iron center and contains a chloride ligand that is displaced in water. Two additional derivatives are sufficiently water-soluble for study as MRI probes, including a complex with a hydroxypyridine group, Fe(L2), and a hydroxybenzoic acid group, Fe(L3). The pH potentiometric titrations give protonation constants of 7.2 and 7.5 for Fe(L2) and Fe(L3), respectively, which are assigned to deprotonation of the bound water. Changes in the electronic absorbance spectra of the complexes as a function of pH are consistent with the deprotonation of phenol pendants at acidic pH values. However, the inner-sphere water ligand of Fe(L2) and Fe(L3) does not exchange rapidly on the NMR timescale at pH 6.0 or 7.4, as shown by variable-temperature 17O NMR spectroscopy. The pH-dependent proton relaxivity profiles show a maximum in relaxivity at a near-neutral pH, suggesting that exchange of the protons of the bound water is an important contribution. Competitive binding studies with ethylenediaminetetraacetic acid (EDTA) show effective stability constants for Fe(L2) and Fe(L3) at pH 7.4 with log K values of 21.1 and 20.5, respectively. These two complexes are kinetically inert in carbonate phosphate buffer at 37 °C for several hours but transfer iron to transferrin. Fe(L2) and Fe(L3) show enhanced contrast in T1-weighted imaging analyses in BALB/c mice. These studies show that Fe(L2) clears through mixed renal and hepatobiliary routes, while Fe(L3) has a similar pharmacokinetic clearance profile to a macrocyclic Gd(III) contrast agent.

Dinuclear Fe(III) Hydroxypropyl-Appended Macrocyclic Complexes as MRI Probes.

Four high-spin Fe(III) macrocyclic complexes, including three dinuclear and one mononuclear complex, were prepared toward the development of more effective iron-based magnetic resonance imaging (MRI) contrast agents. All four complexes contain a 1,4,7-triazacyclononane macrocyclic backbone with two hydroxypropyl pendant groups, an ancillary aryl or biphenyl group, and a coordination site for a water ligand. The pH potentiometric titrations support one or two deprotonations of the complexes, most likely deprotonation of hydroxypropyl groups at near-neutral pH. Variable-temperature 17O NMR studies suggest that the inner-sphere water ligand is slow to exchange with bulk water on the NMR time scale. Water proton T1 relaxation times measured for solutions of the Fe(III) complexes at pH 7.2 showed that the dinuclear complexes have a 2- to 3-fold increase in r1 relaxivity in comparison to the mononuclear complex per molecule at field strengths ranging from 1.4 T to 9.4 T. The most effective agent, a dinuclear complex with macrocycles linked through para-substitution of an aryl group (Fe2(PARA)), has an r1 of 6.7 mM-1 s-1 at 37 °C and 4.7 T or 3.3 mM-1 s-1 per iron center in the presence of serum albumin and shows enhanced blood pool and kidney contrast in mice MRI studies.

Co(II) Macrocyclic Complexes Appended with Fluorophores as paraCEST and cellCEST Agents.

Four high-spin macrocyclic Co(II) complexes with hydroxypropyl or amide pendants and appended coumarin or carbostyril fluorophores were prepared as CEST (chemical exchange saturation transfer) MRI probes. The complexes were studied in solution as paramagnetic CEST (paraCEST) agents and after loading into Saccharomyces cerevisiae yeast cells as cell-based CEST (cellCEST) agents. The fluorophores attached to the complexes through an amide linkage imparted an unusual pH dependence to the paraCEST properties of all four complexes through of ionization of a group that was attributed to the amide NH linker. The furthest shifted CEST peak for the hydroxypropyl-based complexes changed by ∼90 ppm upon increasing the pH from 5 to 7.5. At acidic pH, the Co(II) complexes exhibited three to four CEST peaks with the most highly shifted CEST peak at 200 ppm. The complexes demonstrated substantial paramagnetic water proton shifts which is a requirement for the development of cellCEST agents. The large shift in the proton resonance was attributed to an inner-sphere water at neutral pH, as shown by variable temperature 17O NMR spectroscopy studies. Labeling of yeast with one of these paraCEST agents was optimized with fluorescence microscopy and validated by using ICP mass spectrometry quantitation of cobalt. A weak asymmetry in the Z-spectra was observed in the yeast labeled with a Co(II) complex, toward a cellCEST effect, although the Co(II) complexes were toxic to the cells at the concentrations necessary for observation of cellCEST.

Modulating the Properties of Fe(III) Macrocyclic MRI Contrast Agents by Appending Sulfonate or Hydroxyl Groups.

Complexes of Fe(III) that contain a triazacyclononane (TACN) macrocycle, two pendant hydroxyl groups, and a third ancillary pendant show promise as MRI contrast agents. The ancillary group plays an important role in tuning the solution relaxivity of the Fe(III) complex and leads to large changes in MRI contrast enhancement in mice. Two new Fe(III) complexes, one with a third coordinating hydroxypropyl pendant, Fe(L2), and one with an anionic non-coordinating sulfonate group, Fe(L1)(OH2), are compared. Both complexes have a deprotonated hydroxyl group at neutral pH and electrode potentials representative of a stabilized trivalent iron center. The r1 relaxivity of the Fe(L1)(OH2) complex is double that of the saturated complex, Fe(L2), at 4.7 T, 37 °C in buffered solutions. However, variable-temperature 17O-NMR experiments show that the inner-sphere water of Fe(L1)(OH2) does not exchange rapidly with bulk water under these conditions. The pendant sulfonate group in Fe(L1)(OH2) confers high solubility to the complex in comparison to Fe(L2) or previously studied analogues with benzyl groups. Dynamic MRI studies of the two complexes showed major differences in their pharmacokinetics clearance rates compared to an analogue containing a benzyl ancillary group. Rapid blood clearance and poor binding to serum albumin identify Fe(L1)(OH2) for development as an extracellular fluid contrast agent.

A Class of FeIII Macrocyclic Complexes with Alcohol Donor Groups as Effective T1 MRI Contrast Agents.

Early studies suggested that FeIII complexes cannot compete with GdIII complexes as T1 MRI contrast agents. Now it is shown that one member of a class of high-spin macrocyclic FeIII complexes produces more intense contrast in mice kidneys and liver at 30 minutes post-injection than does a commercially used GdIII agent and also produces similar T1 relaxivity in serum phantoms at 4.7 T and 37 °C. Comparison of four different FeIII macrocyclic complexes elucidates the factors that contribute to relaxivity in vivo including solution speciation. Variable-temperature 17 O NMR studies suggest that none of the complexes has a single, integral inner-sphere water that exchanges rapidly on the NMR timescale. MRI studies in mice show large in vivo differences of three of the FeIII complexes that correspond, in part, to their r1 relaxivity in phantoms. Changes in overall charge of the complex modulate contrast enhancement, especially of the kidneys.

CoII Complexes as Liposomal CEST Agents.

Three paramagnetic CoII macrocyclic complexes containing 2-hydroxypropyl pendant groups, 1,1',1'',1'''-(1,4,8,11-tetraazacyclotetradecane-1,4,8,11-tetrayl)tetrakis- (propan-2-ol) ([Co(L1)]2+ , 1,1'-(4,11-dibenzyl-1,4,8,11-tetraazacyclotetradecane-1,8-diyl)bis(propan-2-ol) ([Co(L2)]2+ ), and 1,1'-(4,11-dibenzyl-1,4,8,11-tetraazacyclotetradecane-1,8-diyl)bis(octadecan-2-ol) ([Co(L3)]2+ ) were synthesized to prepare transition metal liposomal chemical exchange saturation transfer (lipoCEST) agents. In solution, ([Co(L1)]2+ ) forms two isomers as shown by 1 H NMR spectroscopy. X-ray crystallographic studies show one isomer with 1,8-pendants in cis-configuration and a second isomer with 1,4-pendants in trans-configuration. The [Co(L2)]2+ complex has 1,8-pendants in a cis-configuration. Remarkably, the paramagnetic-induced shift of water 1 H NMR resonances in the presence of the [Co(L1)]2+ complex is as large as that observed for one of the most effective LnIII water proton shift agents. Incorporation of [Co(L1)]2+ into the liposome aqueous core, followed by dialysis against a solution of 300 mOsm L-1 produces a CEST peak at 3.5 ppm. Incorporation of the amphiphilic [Co(L3)]2+ complex into the liposome bilayer produces a more highly shifted CEST peak at -13 ppm. Taken together, these data demonstrate the feasibility of preparing CoII lipoCEST agents.

Inner-Sphere and Outer-Sphere Water Interactions in Co(II) paraCEST Agents.

High-spin Co(II) complexes are promising for development as paraCEST agents (paraCEST = paramagnetic chemical exchange saturation transfer) for magnetic resonance imaging applications. The first examples of Co(II) paraCEST agents with bound water ligands are presented here. Four Co(II) macrocyclic complexes based on 1,4,7-triazacyclononane and containing either pendent alcohol or pendent amide groups were prepared. Two of the macrocycles encapsulate the Co(II) and contain no water ligands as shown by X-ray crystallographic studies, and two complexes have macrocycles with only five ligand donor groups to leave an open coordination site for bound water. The ionization of alcohol, water, or amide groups in the complexes was characterized by using pH potentiometry. These data show that one of the complexes has a readily deprotonated group with a pKa close to 6, which is assigned as an alcohol pendent. Amide pendents deprotonate at high pH (>8), and the water ligands of the Co(II) complexes are not deprotonated at neutral pH. All complexes produce CEST peaks through either alcohol OH or amide NH proton exchange. The water ligands exchange too rapidly to produce a CEST effect as shown by variable-temperature 17O NMR spectroscopy studies. The complexes with available coordination sites for inner-sphere water ligands produce large paramagnetic shifts and broadening of the 17O resonances of bulk water, whereas the encapsulated complexes show much less shifting and broadening of 17O resonances. All complexes produce substantial paramagnetic shifts of the 1H resonances of bulk water, which is promising for the development of supramolecular CEST agents.

A novel fluorescent probe based imprinted polymer-coated magnetite for the detection of imatinib leukemia anti-cancer drug traces in human plasma samples.

Myeloid leukemia is a chronic cancer, which associated with abnormal BCR-ABL tyrosine kinase activity. Imatinib (IMB) acts as a tyrosine kinase inhibitor and averts tumor growth in cancer cells by controlling cell division, so it is urgent to develop an effective assay to detect and monitor its IMB concentration. Therefore, an innovative fluorescent biomimetic sensor is a promising sensing material that constructed for the efficient recognition of IMB and displays excellent selectivity and sensitivity stemming from molecularly imprinted polymer@Fe3O4 (MIP@Fe3O4). The detection strategy depends on the recognition of IMB molecules at the imprinted sites in the presence of coexisting molecules, which are then transferred to the fluorescence signal. The synthesized MIP@Fe3O4 was characterized using Fourier-transform infrared spectroscopy (FT-IR), scanning electron microscopy (SEM), and atomic force microscopy (AFM). Furthermore, computational studies of the band gap (EHOMO-ELUMO) of the monomers, IMB, and their complexes were performed. These results confirmed that the copolymer is the most appropriate and has high stability (Binding energy; 0.004 x 10-19 KJ) and low reactivity. A comprehensive linear response over IMB concentrations from 5 × 10-6 mol/L to 8 × 10-4 mol/L with a low detection limit of 9.3 × 10-7 mol/L was achieved. Furthermore, the proposed technique displayed long-term stability (over 2 months), high intermediate precision (RSD<2.1 %), good reproducibility (RSD <1.9 %), and outstanding selectivity toward IMB over analogous molecules with similar chemical and spatial structure (no interference by 100 to 150-fold of the competitors). Owing to these merits, the proposed fluorescence sensor was utilized to detect IMB in drug tablets and human plasma, and satisfactory results (99.3-100.4 %) were obtained. Thus, the synthesized fluorescence sensor is a promising platform for IMB sensing in various applications.

Co(II) complexes of tetraazamacrocycles appended with amide or hydroxypropyl groups as paraCEST agents.

Co(II) complexes of 1,4,7,10-tetraazacyclododecane (CYCLEN) or 1,4,8,11-tetraazacyclotetradecane (CYCLAM) with 2-hydroxypropyl or carbamoylmethyl (amide) pendants are studied with the goal of developing paramagnetic chemical exchange saturation transfer (paraCEST) agents. Single-crystal X-ray diffraction studies show that two of the coordination cations with hexadentate ligands, [Co(DHP)]2+ and [Co(BABC)]2+, form six-coordinate complexes; whereas two CYCLEN-based complexes with potentially octadentate ligands, [Co(THP)]2+ and [Co(HPAC)]2+, are seven-coordinate with only three of the four pendant groups bound to the metal center. 1H NMR spectra of these complexes suggest that the six-coordinate complexes are present as a single isomer in aqueous solution. For the complexes which are seven-coordinate in the solid state, one is highly fluxional in aqueous solution on the NMR time scale ([Co(HPAC)]2+), whereas the NMR spectrum of [Co(THP)]2+ is consistent with an eight-coordinate complex with all pendants bound. Co(II) complexes of CYCLEN derivatives show CEST effects of low intensity that are assigned to NH or OH groups of the pendants. One complex, [Co(DHP)]2+, shows a highly-shifted CEST peak at 113 ppm versus bulk water, attributed to OH protons. However, the CEST effect is largest for two Co(II) CYCLAM-based complexes with coordinated amide groups that undergo NH proton exchange. All five complexes are inert towards dissociation in buffered solutions containing carbonate and phosphate and towards trans-metalation by excess Zn(II). These data give insight into the production of an intense CEST effect for tetraazamacrocyclic complexes with pendant groups containing NH or OH exchangeable protons. The intense and highly shifted CEST peak(s) of the CYCLAM-based complexes suggest that they are promising for further development as paraCEST agents.

Liposomal Fe(III) Macrocyclic Complexes with Hydroxypropyl Pendants as MRI Probes.

Paramagnetic liposomes containing Fe(III) complexes were prepared by incorporation of mononuclear (Fe(L1) or Fe(L3)) or dinuclear (Fe2(L2)) coordination complexes of 1,4,7-triazacyclononane macrocycles containing 2-hydroxypropyl pendant groups. Two different types of paramagnetic liposomes were prepared. The first type, LipoA, has the mononuclear Fe(L1) complex loaded into the internal aqueous core. The second type, LipoB, has the amphiphilic Fe(L3) complex inserted into the liposomal bilayer and the internal aqueous core loaded with either Fe(L1) (LipoB1) or Fe2(L2) (LipoB2). LipoA enhances both T1 and T2 water proton relaxation rates. Treatment of LipoA with osmotic gradients to produce a nonspherical liposome produces a liposome with a chemical exchange saturation transfer effect as shown by an asymmetry analysis but only at high osmolarity. LipoB1, which contains an amphiphilic complex in the liposomal bilayer, produced a broadened Z-spectrum upon treatment of the liposome with osmotic gradients. The r1 relaxivity of LipoB1 and LipoB2 were higher than the r1 relaxivity of LipoA on a per Fe basis, suggesting an important contribution from the amphiphilic Fe(III) center. The r1 relaxivities of paramagnetic liposomes are relatively constant over a range of magnetic field strengths (1.4-9.4 T), with the ratio of r2/r1 substantially increasing at high field strengths. MRI studies of LipoB1 in mice showed prolonged contrast enhancement in blood compared to the clinically employed Gd(DOTA), which was injected at a 2-fold higher dose per metal than the Fe(III)-loaded liposomes.

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.