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PARACEST redox sensors containing the NAD+/NADH mimic N-methylated quinolinium moiety as redox active functional group have been designed and synthesized. The Eu3+-complex with two quinolinium moieties was nearly completely silent in CEST in the oxidized form, but “turns on” upon reduction with β-NADH. The CEST effect of the Eu3+- complex containing only one quinolinium group was much less redox responsive but showed an unexpected sensitivity to pH in the physiologically relevant pH range.

The chemical exchange saturation transfer (CEST) efficiency for a series Eu3+-based tetraamide complexes bearing p-substituents on a single coordinating pendant arm is highly sensitive to water exchange rates. The CEST effect increases in the order Me < MeO < F approximately CO2tBu < CN < H. These results show that CEST contrast can be modulated by changes in electron density at a single ligating atom, and this forms the basis of creating imaging agents that respond to chemical oxidation and reduction.

Ln(S-THP) 3+ complexes are paramagnetic chemical exchange saturation transfer agents (PARACEST) agents for magnetic resonance imaging (MRI) (S-THP = (1S,4S,7S,10S)-1,4,7,10-tetrakis(2-hydroxypropyl)-1,4,7,10-tetraazacyclododecane, Ln(III) = Ce(III), Eu(III), Yb(III)). CEST spectra at 11.7 T show that the PARACEST effect of these complexes is enhanced at neutral pH in buffered solutions containing 100 mM NaCl upon addition of 1-2 equivalents of diethylphosphate (DEP). CEST images of phantoms at 4.7 T confirm that DEP enhances the properties of Yb(S-THP) 3+ as a PARACEST MRI agent in buffered solutions at neutral pH and 100 mM NaCl. Studies using 1 H NMR, direct excitation Eu(III) luminescence spectroscopy, and UV-visible spectroscopy show that DEP is an outersphere ligand. Dissociation constants for [Ln(S-THP)(OH 2 )](DEP) are 1.9 mM and 2.8 mM for Ln(III) = Yb(III) at pH 7.0 and Eu(III) at pH 7.4, respectively. Related ligands including phosphorothioic acid, O,O-diethylester, ethyl methylphosphonate, O-(4-nitrophenylphosphoryl)choline and cyclic 3,5-adenosine monophosphate do not activate PARACEST. BNPP (bis(4-nitrophenyl phosphate) activates PARACEST of Ln(S-THP) 3+ (Ln(III) = Eu(III), Yb(III)), albeit less effectively than does DEP. This data shows that binding through second coordination sphere interactions is selective for phosphate diesters with two terminal oxygens and two identical ester groups. A crystal structure of [Eu(S-THP)(OH 2 )]((O 2 NPhO) 2 PO 2 ) 2 (CF 3 SO 3 ) · 2H 2 O · iPrOH has two outersphere BNPP anions that form hydrogen bonds to the alcohol groups of the macrocycle and the bound water ligand. This structure supports 1 H NMR spectroscopy studies showing that outersphere interactions of the phosphate diester with the alcohol protons modulate the rate of alcohol proton exchange to influence the PARACEST properties of the complex. Further, DEP interacts only with the non-ionized form of the complex, Ln(S-THP)(OH 2 ) 3+ contributing to the pH dependence of the PARACEST effect.

Chemical exchange-dependent saturation transfer and paramagnetic chemical exchange-dependent saturation transfer are agent-mediated contrast mechanisms that depend on saturating spins at the resonant frequency of the exchangeable protons on the agent, thereby indirectly saturating the bulk water. In general, longer saturating pulses produce stronger chemical and paramagnetic exchange-dependent saturation transfer effects, with returns diminishing for pulses longer than T 1 . This could make imaging slow, so one approach to chemical exchange-dependent saturation transfer imaging has been to follow a long, frequency-selective saturation period by a fast imaging method. A new approach is to insert a short frequency-selective saturation pulse before each spatially selective observation pulse in a standard, two-dimensional, gradient-echo pulse sequence. Being much less than T 1 apart, the saturation pulses have a cumulative effect. Interleaved, multislice imaging is straightforward. Observation pulses directed at one slice did not produce observable, unin- (2) demonstrated that low-molecular-weight compounds with slowly exchanging ÀNH or ÀOH protons can alter tissue contrast via chemical exchange dependent saturation transfer (CEST) of chemically shifted, presaturated spins to bulk water. Images using CEST are typically generated by subtracting images obtained by applying a radiofrequency saturation pulse at the resonant frequency of the exchanging protons from images obtained by applying the saturation pulse at a control frequency. More recently, this CEST approach has been extended to exogenous paramagnetic lanthanide chelates with slow water exchange to produce MR contrast based on the paramagnetic CEST (PARAC-EST) effect (3,4). These PARACEST agents have advantages over more conventional T 1 -shortening agents because they are activated selectively by an excitation pulse applied at a specific frequency. Targeted CEST/ PARACEST agents with different targets and different resonant frequencies for the exchanging protons may be injected together, yet imaged separately in a single examination (5). The CEST/PARACEST effect has been applied with responsive agents (6) to produce images sensitive to a variety of tissue parameters, including temperature (7), presence of enzymes (8), pH (2,9), and concentrations of glucose (10,11), lactate (12), and zinc (13). Also, since the relaxivities of these PARACEST agents are low, endogenous MR contrast is preserved until external radiofrequency saturation is applied and there is no interference with subsequent imaging with T 1 -shortening agents.In practice, the external radiofrequency saturation pulse is applied for a time longer than the T 1 of the bulk water protons. Woessner et al. (14) have published numerical solutions to the Bloch equations that show that both CEST and PARACEST systems approach steady state under these conditions. As was recently pointed out by Liu et al. (15) and Sun et al. (16), the most straightforward acquisition schemes for PARACEST and CEST MRI invol...

A novel approach for the design of responsive paramagnetic chemical exchange saturation transfer (PARACEST) magnetic resonance imaging (MRI) agents has been developed where the signal is “turned on” by altering the longitudinal relaxation time (T1) of bulk water protons. To demonstrate this approach, a model Eu(DOTA-tetraamide) complex (DOTA = 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) containing two nitroxide free radical units was synthesized. The nitroxide groups substantially shortened the T1 of the bulk water protons which, in turn, resulted in quenching of the CEST signal. Reduction of paramagnetic nitroxide moieties to a diamagnetic species resulted in the appearance of CEST. The modulation of CEST by T1 relaxation provides a new platform for designing biologically responsive MRI agents.

Purpose The water molecule exchange rates in a series of DyDOTA-(amide)X chelates were fine-tuned to maximize the effects of T2-exchange line broadening and improve T2 contrast. Methods Four DyDOTA-(amide)X chelates having a variable number of glycinate side-arms were prepared and characterized as T2-exchange agents. The non-exchanging DyTETA chelate was also used to measure the bulk water T2 reduction due solely to T2*. The total transverse relaxivity (r2tot) at 22, 37, and 52 °C for each chelate was measured in vitro at 9.4 T (400 MHz) by fitting plots of total T2−1 versus concentration. The water molecule exchange rates for each complex were measured by fitting 17O line-width versus temperature data taken at 9.4 T (54.3 MHz). Results The measured transverse relaxivities due to water molecule exchange (r2ex) and bound water lifetimes (τM) were in excellent agreement with Swift-Connick theory, with DyDOTA-(gly)3 giving the largest r2ex = 11.8 s−1 mM−1 at 37 °C. Conclusion By fine-tuning the water molecule exchange rate at 37 °C, the transverse relaxivity has been increased by 2 to 30 times compared to previously studied Dy3+-based chelates. Polymerization or dendrimerization of the optimal chelate could yield a highly sensitive, molecule-sized T2 contrast agent for improved molecular imaging applications.