PROJECT SUMMARY
Magnetic resonance (MR) reporter genes have the potential to monitor transgene expression non-invasively in
real time at high resolution. These genes can be applied to interrogate the efficacy of gene therapy, to monitor
viral therapeutics and viral gene delivery, to assess cellular differentiation, cell trafficking, and specific
metabolic activity, and also assess changes in the microenvironment. Efforts toward the development of MR
reporter genes have been made for over a decade, but, despite these efforts, the field is still in its early
developmental stage. This reflects the fact that there are numerous complications, caused by the low
sensitivity of detection, the need for substrates with their associated undesirable pharmacokinetics, and/or the
difficult and, in some cases, delayed interpretation of signal changes.
We have previously demonstrated that many of these challenges can be overcome with the use of a lysine rich
protein (LRP) reporter gene, that is detectable by chemical exchange saturation transfer (CEST) MRI.
However, to mature the CEST reporter gene technology and bring it towards clinical translation, its sensitivity
and specificity need to be improved. In particular, the LRP reporter gene specificity is limited by the fact that
the lysine amide exchangeable protons of LRP have the same chemical shift as amide protons from
endogenous proteins. It is therefore difficult to distinguish the reporter CEST contrast from the background
CEST contrast, both of which may be changing with time. The specificity is further limited by the sensitivity of
the CEST contrast to intracellular pH where the qualitative CEST contrast cannot distinguish between
exchange rate and concentration effects. Finally, a decrease in cytosolic pH, observed in many disease
pathologies, reduces the amide proton exchange rate and hence the CEST reporter sensitivity.
We therefore propose to develop improved MRI reporter genes and quantitative MRI detection methods that
will facilitate the clinical translation of these methods for imaging biological therapeutics, such as oncolytic
virotherapy. We hypothesize that CEST reporter genes with improved sensitivity and specificity along with
improved quantitative CEST methods will enable viral infection and replication to be monitored longitudinally
throughout OV tumor therapy. To test this hypothesis and establish the clinical potential of MRI reporter genes
we will capitalize on two transformative technologies developed in our labs; (Aim 1) an artificial intelligence
based genetic programming algorithm will be used for optimizing the sensitivity and specificity of the CEST
reporter gene and (Aim 2) a CEST magnetic resonance fingerprinting (MRF) method will be used for the rapid
quantification of both the reporter protein concentration and chemical exchange rate. (Aim 3) These methods
will be validated for imaging oncolytic viral infection and replication in mouse glioblastoma tumor models.