MRI reporter genes in the era of gene transfer

[...]PET-detectable probes, which emit radioactivity that decays over time, pose challenges in terms of safety as well as accessibility, handling and production. Realizing these drawbacks, we have demonstrated the ability to map the expression of the HSV1-tk reporter gene with an MRI-detectable reporter probe.11 To this end, a synthetic dN, the thymidine-analogue, 5-methyl-dihhydrothymidine (5-MDHT),12 was found to be a specific substrate for the HSV1-TK enzyme, thus accumulating, upon its phosphorylation, in cells engineered to express HSV1-tk. [...]5-MDHT was rationalized to be an ideal MRI reporter probe that is detected, without the need for labelling, by a technique known as chemical exchange saturation transfer (CEST).13 This makes 5-MDHT an ideal reporter probe for imaging HSV1-tk reporter gene expression with CEST-MRI. Capitalizing on this feature, we have demonstrated that another synthetic dN, pyrrolo-deoxycytidine (pdC), which generates strong CEST contrast as well, can be used as a reporter probe to image the expression of the reporter gene encoding to the enzyme Drosophila melanogaster deoxyribonucleoside kinase (Dm-dNK).15 As the CEST-MRI contrast is ‘turned on’ on demand, only upon the application of a frequency-specific, probe-selective ‘tagging’, multiple CEST reporter probes reactive to different radiofrequency pulses (Δω1, Δω2, Δω3, …) can be administered simultaneously and then mapped and presented in a pseudo-colour manner. [...]site-directed mutagenesis was applied to obtain the optimized HSV1-TK_7B and Dm-dNK_7C as ultimately orthogonal reporter genes for the 5-MDHT and pdC reporter probes, respectively (Figure 1B, bottom).