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要旨:
The development of bioresponsive magnetic resonance imaging (MRI) contrast agents (CAs) specific to monitoring Ca2+ fluctuations are of increasing interest for fMRI studies of neural activity. Such probes can provide key information regarding their microenvironment through changes in MR signal which in turn can lead to vital information concerning the functioning of tissue being extracted. Thus far, a number of CAs sensitive to Ca2+ have been developed ranging from ‘small’ molecular systems to larger nano-sized derivatives. Here, an extension to this ever growing field with the development of a range monomeric, multimeric and nano-sized Ca2+-responsive smart contrast agents (SCAs) is described. A range of bioresponsive dendrimeric CAs with different structures and charge distributions are described in the pursuit of probes for T1-weighted imaging and r2/r1 ratiometric imaging probes. The use of nano-sized platforms enabled higher Gd3+ loading and slower diffusion rates, which are favoured characteristics for in vivo applications. The impact of structural and charge changes resulted in significant consequences for the performance of the probes as Ca2+-responsive MRI CAs. The most active probe displayed common changes in r1 while also exhibiting a remarkable increase in the r2/r1 ratio, greater than that previously achieved. Further investigation revealed that only through a synergistic combination of an increase in q with a change in size and rigidity of the conjugate could such relaxometric changes be realised. This ultimately provided significant insights into the behaviour of such dendrimeric systems and provided a model in which future preparations should be based in the development of T1-weighted and r2/r1 ratiometric probes to visualise Ca2+ fluctuations dynamically. Deeper structural studies were performed on two monomeric systems in which the linker length between the MR reporting moiety and the bioresponsive unit were extended. Various studies revealed significant differences in relaxometric behaviour between the probes. Characterisation with a range of techniques revealed structural changes in the complex coordination environment between the ‘off’ and ‘on’ states which is expected for such systems. Furthermore, the diffusive behaviour of each complex described systems which do not significantly change upon Ca2+ coordination. The results of this study revealed how subtle structural changes can significantly impact the performance of a SCA, thus helping to identify the requirements for future probes. The final parts of this work focused on employing solid phase synthetic techniques as an alternative to standard solution phase chemistry in the preparation of more ‘complex’ SCA derivatives. In one approach, a functionalised bismacrocyclic derivative was assembled on solid phase through the use of multiple building blocks in a straightforward manner. The potency of this probe was confirmed by relaxometric titrations. In a second study, a targeted multimeric probe consisting of three SCA monomers and the RGD peptide sequence was developed. This multimer showed significant increases in relaxivity upon Ca2+ addition. The use of solid phase protocols in both of these cases allowed for more complex SCAs to be developed, which would otherwise be extremely difficult following solution phase protocols. Furthermore, the use of peptide scaffolds allows for simple customisation in which multimeric or multifunctional probes can be developed, providing an additional synthetic tool for chemists attempting to develop bioresponsive MRI CAs.
