Reporting on contrasting MRI enzyme

Researchers have developed an extracellular enzymatic gene-reporter system for magnetic resonance imaging (MRI). The system yields strong, reversible contrast changes in response to the expression of secreted alkaline phosphatase (SEAP). The products of SEAP activity can then be detected using an iron oxide based sensor.

Understanding molecular biology at the cellular level is a critical part of modern research. However, MRI is not yet up to the task of non-invasive imaging of gene expression in whole animals. Now, researchers in the US and Canada have discovered an enzymatic reporter system that is detectable by MRI and could address the issues of lack of sensitivity and spatiotemporal resolution. A reporter enzyme would combine the relatively high resolution of MRI with the ability of each genetically expressed enzyme to generate many product molecules that can be detected by MRI.

Gil Westmeyer and Alan Jasanoff of the Departments of Biological Engineering, Brain& Cognitive Sciences, and Nuclear Science & Engineering at Massachusetts Institute of Technology in Cambridge, Massachusetts, USA and Yves Durocher of the NRC Biotechnology Research Institute in Montreal, Quebec, Canada, have turned to SEAP for assistance.

SEAP is a modified form of placental alkaline phosphatase that is used as a reporter enzyme alongside optically absorbent, fluorescent, or luminescent substrates in molecular biology. In order to adapt SEAP for use with MRI, the team modified an existing sensor for adenosine, which is made by the SEAP catalysed hydrolysis of phosphorylated adenosine derivatives. The reporter enzyme thus makes product molecules that are monitored with MRI rather than acting on a contrast agent.

The adenosine sensor is controlled by a DNA aptamer that binds to adenosine. When no adenosine is present, the aptamer forms cross-links with superparamagnetic iron oxide nanoparticles added to the system. These nanoparticles are modified with reverse- complementary DNA segments. When adenosine is present, the cross-links that allow the nanoparticles to aggregate are broken and the ability of the nanoparticles to increase contrast in the MRI scan falls in-line with the rising adenosine concentration. The team modified this known adenosine sensor to weaken the cross-linking segment and its binding partner and were able to boost the sensitivity of the adenosine sensor tenfold.

Their experiments carried out with the adenosine sensor in the presence of the enzyme and its substrates showed that SEAP activity could be detected. The reversible, non-destructive use of the system is possible even in the presence of adenosine deaminase, a second enzyme that would be present in biological systems that might be studied using MRI in conjunction with the adenosine sensor.

The next step was to test their approach in a cellular context. In these experiments, SEAP could be used as a genetically encoded reporter enzyme by expressing it in HEK-293 cells. Its activity was monitored using MRI with the adenosine sensor in the presence of 2'-adenosine monophosphate (AMP) which is hydrolysed to adenosine. The combination of MRI, SEAP and modified adenosine sensor, suggests that monitoring enzyme activity, as "reported by SEAP, could soon be an entirely viable proposition in cultured cells in particular where optical assays are unreliable.

"The system generates strong T2-based contrast changes, does not involve cell delivery or the catalytic destruction of contrast agents, and is both reversible and moderately fast because of its product-dependent sensing mechanism," the team explains. It will be useful for studying opaque cells, tissue cultures and the approach might pave the way to new types of non-invasive biochemical studies on whole animals. Indeed, "MRI-based assays may be particularly beneficial for screening applications in which data from three- dimensional sample arrays may be acquired in parallel," the team adds.

The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.