Physicochemical and MRI characterization of Gd3+-loaded polyamidoamine and hyperbranched dendrimers.

Generation 4 polyamidoamine (PAMAM) and, for the first time, hyperbranched poly(ethylene imine) or polyglycerol dendrimers have been loaded with Gd3+ chelates, and the macromolecular adducts have been studied in vitro and in vivo with regard to MRI contrast agent applications. The Gd3+ chelator was either a tetraazatetracarboxylate DOTA-pBn4- or a tetraazatricarboxylate monoamide DO3A-MA3- unit. The water exchange rate was determined from a 17O NMR and 1H Nuclear Magnetic Relaxation Dispersion study for the corresponding monomer analogues [Gd(DO3A-AEM)(H2O)] and [Gd(DOTA-pBn-NH2)(H2O)]- (kex298=3.4 and 6.6x10(6) s-1, respectively), where H3DO3A-AEM is {4-[(2-acetylaminoethylcarbamoyl)methyl]-7,10-bis(carboxymethyl-1,4,7,10-tetraazacyclododec-1-yl)}-acetic acid and H4DOTA-pBn-NH2 is 2-(4-aminobenzyl)-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid. For the macromolecular complexes, variable-field proton relaxivities have been measured and analyzed in terms of local and global motional dynamics by using the Lipari-Szabo approach. At frequencies below 100 MHz, the proton relaxivities are twice as high for the dendrimers loaded with the negatively charged Gd(DOTA-pBn)- in comparison with the analogous molecule bearing the neutral Gd(DO3A-MA). We explained this difference by the different rotational dynamics: the much slower motion of Gd(DOTA-pBn)--loaded dendrimers is likely related to the negative charge of the chelate which creates more rigidity and increases the overall size of the macromolecule compared with dendrimers loaded with the neutral Gd(DO3A-MA). Attachment of poly(ethylene glycol) chains to the dendrimers does not influence relaxivity. Both hyperbranched structures were found to be as good scaffolds as regular PAMAM dendrimers in terms of the proton relaxivity of the Gd3+ complexes. The in vivo MRI studies on tumor-bearing mice at 4.7 T proved that all dendrimeric complexes are suitable for angiography and for the study of vasculature parameters like blood volume and permeability of tumor vessels.

Positively charged GdIII cryptates: slow, associative water exchange.

A combined variable-temperature and multiple field 17O NMR, EPR and NMRD study has been performed for the first time on gadolinium(III) complexes of cryptand ligands, L1 and L2, where L1 contains three 2,2'-bipyridine units ([bpy.bpy.bpy]) and L2 is the disubstituted methyl ester derivative of L1. The experimental data have been analysed in a simultaneous fit in order to determine parameters for water exchange, rotational dynamics and electronic relaxation for both complexes. The cryptates have three water molecules in the inner sphere which exchange with a rate of k(ex)298 = 1.8 x 10(6) s(-1) and 0.97 x 10(6) s(-1) for [GdL1(H2O)3]3+ and [GdL2(H2O)3)]3+, respectively. The k(ex)298 values obtained for these positively charged cryptates are smaller than those of the negatively charged Gd-poly(amino carboxylate) complexes. The water exchange mechanism was assessed for [GdL2(H2O)3]3+ by variable-pressure 17O NMR relaxation measurements. Based on the activation volume, DeltaV++ = -2.5 cm3 mol(-1), the water exchange is an associative interchange process. The proton relaxivities, r1, of the cryptate complexes are 9.79 mM(-1) s(-1) for [GdL1(H2O)3]3+ and 11.18 mM(-1) s(-1) for [GdL2(H2O)3]3+ (298 K, 20 MHz), which, due to the presence of three inner sphere water molecules, represent much higher values than those obtained for Gd3+ poly(amino carboxylate) complexes of similar molecular weight.

Equilibrium and NMR studies on GdIII, YIII, CuII and ZnII complexes of various DTPA-N,N''-bis(amide) ligands. Kinetic stabilities of the gadolinium(III) complexes.

Three DTPA-derivative ligands, the non-substituted DTPA-bis(amide) (L(0)), the mono-substituted DTPA-bis(n-butylamide) (L(1)) and the di-substituted DTPA-bis[bis(n-butylamide)] (L(2)) were synthesized. The stability constants of their Gd3+ complexes (GdL) have been determined by pH-potentiometry with the use of EDTA or DTPA as competing ligands. The endogenous Cu2+ and Zn2+ ions form ML, MHL and M(2)L species. For the complexes CuL(0) and CuL(1) the dissociation of the amide hydrogens (CuLH(-1)) has also been detected. The stability constants of complexes formed with Gd3+, Cu2+ and Zn2+ increase with an increase in the number of butyl substituents in the order ML(0) < ML(1) < ML(2). NMR studies of the diamagnetic YL(0) show the presence of four diastereomers formed by changing the chirality of the terminal nitrogens of their enantiomers. At 323 K, the enantiomerization process, involving the racemization of central nitrogen, falls into the fast exchange range. By the assignment and interpretation of 1H and 13C NMR spectra, the fractions of the diastereomers were found to be equal at pH = 5.8 for YL(0). The kinetic stabilities of GdL(0), GdL(1) and GdL(2) have been characterized by the rates of the exchange reactions occurring between the complexes and Eu3+, Cu2+ or Zn2+. The rates of reaction with Eu3+ are independent of the [Eu3+] and increase with increasing [H+], indicating the rate determining role of the proton assisted dissociation of complexes. The rates of reaction with Cu2+ and Zn2+ increase with rising metal ion concentration, which shows that the exchange can take place with direct attack of Cu2+ or Zn2+ on the complex, via the formation of a dinuclear intermediate. The rates of the proton, Cu2+ and Zn2+ assisted dissociation of Gd3+ complexes decrease with increasing number of the n-butyl substituents, which is presumably the result of steric hindrance hampering the formation or dissociation of the intermediates. The kinetic stabilities of GdL(0) and GdL(1) at pH = 7.4, [Cu2+] = 1 x 10(-6) M and [Zn(2+)] = 1 x 10(-5) M are similar to that of Gd(DTPA)2-, while the complex GdL2 possesses a much higher kinetic stability.

Fine-tuning water exchange on Gd(III) poly(amino carboxylates) by modulation of steric crowding.

In the objective of optimizing water exchange rate on stable, nine-coordinate, monohydrated Gd(III) poly(amino carboxylate) complexes, we have prepared monopropionate derivatives of DOTA4- (DO3A-Nprop4-) and DTPA5- (DTTA-Nprop5-). A novel ligand, EPTPA-BAA(3-), the bisamylamide derivative of ethylenepropylenetriamine-pentaacetate (EPTPA5-) was also synthesized. A variable temperature 17O NMR study has been performed on their Gd(III) complexes, which, for [Gd(DTTA-Nprop)(H2O)]2- and [Gd(EPTPA-BAA)(H2O)] has been combined with multiple field EPR and NMRD measurements. The water exchange rates, k(ex)(298), are 8.0 x 10(7) s(-1), 6.1 x 10(7) s(-1) and 5.7 x 10(7) s(-1) for [Gd(DTTA-Nprop)(H2O)]2-, [Gd(DO3A-Nprop)(H2O)]- and [Gd(EPTPA-BAA)(H2O)], respectively, all in the narrow optimal range to attain maximum proton relaxivities, provided the other parameters (electronic relaxation and rotation) are also optimized. The substitution of an acetate with a propionate arm in DTPA5- or DOTA4- induces increased steric compression around the water binding site and thus leads to an accelerated water exchange on the Gd(III) complex. The k(ex) values on the propionate complexes are, however, lower than those obtained for [Gd(EPTPA)(H2O)]2- and [Gd(TRITA)(H2O)]- which contain one additional CH(2) unit in the amine backbone as compared to the parent [Gd(DTPA)(H2O)]2- and [Gd(DOTA)(H2O)]-. In addition to their optimal water exchange rate, [Gd(DTTA-Nprop)(H2O)]2- has, and [Gd(DO3A-Nprop)(H2O)]- is expected to have sufficient thermodynamic stability. These properties together make them prime candidates for the development of high relaxivity, macromolecular MRI contrast agents.