Redesigned reporter gene for improved proton exchange-based molecular MRI contrast

• Published in: Scientific reports Vol. 10; no. 1; p. 20664
• Main Authors: Perlman, Or, Ito, Hirotaka, Gilad, Assaf A., McMahon, Michael T., Chiocca, E. Antonio, Nakashima, Hiroshi, Farrar, Christian T.
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 26.11.2020; Nature Publishing Group
• Subjects: 631/1647/1511; 631/1647/245; 631/67/2321; 639/166/985; Amino Acid Sequence; Animals; Brain cancer; Brain Neoplasms - genetics; Brain Neoplasms - pathology; Brain tumors; Contrast Media - metabolism; Deoxyribonucleic acid; DNA; Female; Gene transfer; Genes, Reporter - genetics; HEK293 Cells; Humanities and Social Sciences; Humans; LRP protein; Lysates; Lysine; Magnetic resonance imaging; Magnetic Resonance Imaging - methods; Mice; Mice, Inbred C57BL; multidisciplinary; Neuroimaging; Nucleotide sequence; Oncolysis; Oncolytic Virotherapy - methods; Polymerase chain reaction; Protons; Recombination; Reporter gene; Science; Science (multidisciplinary); Tumor cells
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-020-77576-z

Reporter gene imaging allows for non-invasive monitoring of molecular processes in living cells, providing insights on the mechanisms underlying pathology and therapy. A lysine-rich protein (LRP) chemical exchange saturation transfer (CEST) MRI reporter gene has previously been developed and used to image tumor cells, cardiac viral gene transfer, and oncolytic virotherapy. However, the highly repetitive nature of the LRP reporter gene sequence leads to DNA recombination events and the expression of a range of truncated LRP protein fragments, thereby greatly limiting the CEST sensitivity. Here we report the use of a redesigned LRP reporter (rdLRP), aimed to provide excellent stability and CEST sensitivity. The rdLRP contains no DNA repeats or GC rich regions and 30% less positively charged amino-acids. RT-PCR of cell lysates transfected with rdLRP demonstrated a stable reporter gene with a single distinct band corresponding to full-length DNA. A distinct increase in CEST-MRI contrast was obtained in cell lysates of rdLRP transfected cells and in in vivo LRP expressing mouse brain tumors ( p = 0.0275 , n = 10).