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<p>Ferritins are iron storage proteins found in different species. Through ferroxidase, they convert iron from its toxic, ferrous state (Fe<sup>2+</sup>) into a non-toxic, ferric state (Fe<sup>3+</sup>). When present in its ferric state, iron acts as a paramagnetic agent and causes a shortening of the transversal relaxation (T2) in MRI. Depending on the concentration of iron, this can lead to a visible change in contrast. <a href="#bib5" class="forward-ref">[5]</a></p> | <p>Ferritins are iron storage proteins found in different species. Through ferroxidase, they convert iron from its toxic, ferrous state (Fe<sup>2+</sup>) into a non-toxic, ferric state (Fe<sup>3+</sup>). When present in its ferric state, iron acts as a paramagnetic agent and causes a shortening of the transversal relaxation (T2) in MRI. Depending on the concentration of iron, this can lead to a visible change in contrast. <a href="#bib5" class="forward-ref">[5]</a></p> | ||
− | <p>Bacterioferritin is one of the three forms of ferritin-like proteins found in bacteria. It has been shown that overexpression of bacterioferritin in <span class="bacterium">E. coli</span> Nissle can lead to a visible contrast change in MRI, which allows for visualization of the bacteria (Figure 2).<a href="#bib6" class="forward-ref">[6]</a></p> | + | <p>Bacterioferritin is one of the three forms of ferritin-like proteins found in bacteria. It has been shown that overexpression of bacterioferritin in <span class="bacterium">E. coli</span> Nissle 1917 can lead to a visible contrast change in MRI, which allows for visualization of the bacteria (Figure 2).<a href="#bib6" class="forward-ref">[6]</a></p> |
</section> | </section> | ||
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<img alt="FIXME" | <img alt="FIXME" | ||
src="https://static.igem.org/mediawiki/2017/c/c9/T--ETH_Zurich--Circuit_MRI_figure3.png"/> | src="https://static.igem.org/mediawiki/2017/c/c9/T--ETH_Zurich--Circuit_MRI_figure3.png"/> | ||
− | <figcaption>Figure 3. | + | <figcaption>Figure 3. Bacterioferritin in our system. Once the <a href="https://2017.igem.org/Team:ETH_Zurich/Circuit/Fa_Tumor_Sensor">Tumor Sensing</a> has been activated, both the <a href="https://2017.igem.org/Team:ETH_Zurich/Circuit/Fc_Anti_Cancer_Toxin">Anti-Cancer Toxin</a> azurin and the MRI Contrast Agent bacterioferritin are produced. Azurin will accumulate inside of the bacteria until it is ready for release. Bacterioferritin will take up iron and produce a change in the T2 signal in MRI.</figcaption> |
</figure> | </figure> | ||
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<ul> | <ul> | ||
<li>the bacteria have colonized the correct location,</li> | <li>the bacteria have colonized the correct location,</li> | ||
− | <li>there | + | <li>there are enough bacteria to produce a full dose of the <a href="https://2017.igem.org/Team:ETH_Zurich/Circuit/Fc_Anti_Cancer_Toxin">Anti-Cancer Toxin</a>, so the risk of sub-dosing and eventually producing a drug-resistant tumor is minimized, and </li> |
<li>the <a href="https://2017.igem.org/Team:ETH_Zurich/Circuit/Fc_Anti_Cancer_Toxin">Anti-Cancer Toxin</a>, located on the same plasmid and under the control of the same promoter as the MRI Contrast Agent, has started to accumulate. </li> | <li>the <a href="https://2017.igem.org/Team:ETH_Zurich/Circuit/Fc_Anti_Cancer_Toxin">Anti-Cancer Toxin</a>, located on the same plasmid and under the control of the same promoter as the MRI Contrast Agent, has started to accumulate. </li> | ||
</ul> | </ul> |
Revision as of 17:51, 1 November 2017
![CATE logo](https://static.igem.org/mediawiki/2017/d/d6/T--ETH_Zurich--LogoMinimal.png)
Function B: MRI Contrast Agent
This is a detailed description of an individual function of our circuit. To access other functions and get an overview of the whole circuit, visit the Circuit page.
Introduction
Since the beginnings of modern medicine, doctors have been eager to see what's happening on the inside of the body rather than to just treat blindly.
Therefore, it comes as no surprise that one of the characteristics that the ideal bacterial cancer therapeutic should posses is external detectability. The ability to see from the outside what's happening on the inside would allow the doctor to get critical information about the state of the tumor, the success of localization and the efficacy of treatment. [1]
![](https://static.igem.org/mediawiki/2017/9/9e/T--ETH_Zurich--fb_figure1.png)
The Imaging Modality
To achieve this in CATE, the first step was to choose the imaging modality. Several methods have already been used to visualize contrast-producing bacteria, such as Positron Emission Tomography (PET), Bioluminescence Imaging (BLI), Fluorescence Imaging, Computed Tomography (CT) and Magnetic Resonance Imaging (MRI). [2][3][4] To pick one, we carefully evaluated advantages and disadvantages of these different techniques (Table 1).
Our final decision: MRI. We chose MRI because it is a well-established imaging method in humans, readily available in the clinics. It offers good spatial resolution and does not use ionizing radiation, which means it is in no way harmful. Finally, focused ultrasound, an integral part of our Heat Sensor, uses exclusively MRI image guidance. Therefore, for both theoretical and practical reasons, we decided to incorporate an MRI reporter gene into CATE.
BLI | FLUORESCENCE | PET | MRI | CT | |
---|---|---|---|---|---|
ADVANTAGES | sensitive non-toxic |
sensitive non-toxic |
suitable for humans very sensitive |
suitable for humans spatial resolution non-toxic |
suitable for humans spatial resolution temporal resolution |
DISADVANTAGES | only small animals | only small animals external light source limited depth |
radioactive source expensive spatial resolution |
slow (ca. 30 min/scan) expensive less sensitive than PET |
ionizing radiation |
The MRI Reporter
![FIXME](https://static.igem.org/mediawiki/2017/9/96/T--ETH_Zurich--fb_figure2.png)
From Hill, Philip J., et al. PLoS One 6.10 (2011): e25409 under the terms of the Creative Commons Attribution License.
We have looked into several different MRI reporters and consulted professor Markus Rudin, an expert in molecular imaging from the Institute of Biomedical Engineering at ETH Zurich. After careful consideration, we choose to work with a ferritin-like bacterial protein, bacterioferritin.
Ferritins are iron storage proteins found in different species. Through ferroxidase, they convert iron from its toxic, ferrous state (Fe2+) into a non-toxic, ferric state (Fe3+). When present in its ferric state, iron acts as a paramagnetic agent and causes a shortening of the transversal relaxation (T2) in MRI. Depending on the concentration of iron, this can lead to a visible change in contrast. [5]
Bacterioferritin is one of the three forms of ferritin-like proteins found in bacteria. It has been shown that overexpression of bacterioferritin in E. coli Nissle 1917 can lead to a visible contrast change in MRI, which allows for visualization of the bacteria (Figure 2).[6]
Bacterioferritin and CATE
In our design, we make bacterioferritin a reporter of passing through Checkpoint 1. Once enough bacteria have colonized the tumor and overproduction of lactate has been sensed, the AND-gate of the Tumor Sensor is activated. This leads to expression of the MRI contrast agent and the Anti-Cancer Toxin (Figure 3).
![FIXME](https://static.igem.org/mediawiki/2017/c/c9/T--ETH_Zurich--Circuit_MRI_figure3.png)
While the Anti-Cancer Toxin will accumulate inside of CATE until it is ready to be released during Cell Lysis, the MRI Contrast Agent will take up iron and create a change in the MRI signal. The change in the signal will alert the doctor of the following:
- the bacteria have colonized the correct location,
- there are enough bacteria to produce a full dose of the Anti-Cancer Toxin, so the risk of sub-dosing and eventually producing a drug-resistant tumor is minimized, and
- the Anti-Cancer Toxin, located on the same plasmid and under the control of the same promoter as the MRI Contrast Agent, has started to accumulate.
References
- Forbes, Neil S. "Engineering the perfect (bacterial) cancer therapy." Nature reviews. Cancer 10.11 (2010): 785. doi: 10.1038/nrc2934
- Cronin, M., et al. "Bacterial vectors for imaging and cancer gene therapy: a review." Cancer gene therapy 19.11 (2012): 731. doi: 10.1038/cgt.2012.59
- Gilad, Assaf A., and Mikhail G. Shapiro. "Molecular Imaging in Synthetic Biology, and Synthetic Biology in Molecular Imaging." Molecular Imaging and Biology 19.3 (2017): 373-378. doi: 10.1007/s11307-017-1062-1
- Lyons, Scott K., P. Stephen Patrick, and Kevin M. Brindle. "Imaging mouse cancer models in vivo using reporter transgenes." Cold Spring Harbor Protocols 2013.8 (2013): pdb-top069864. doi: 10.1101/pdb.top069864
- Cohen, Batya et al. “Ferritin as an Endogenous MRI Reporter for Noninvasive Imaging of Gene Expression in C6 Glioma Tumors.” Neoplasia (New York, N.Y.) 7.2 (2005): 109–117. Print. doi: 10.1593/neo.04436
- Hill, Philip J., et al. "Magnetic resonance imaging of tumors colonized with bacterial ferritin-expressing Escherichia coli." PLoS One 6.10 (2011): e25409. doi: 10.1371/journal.pone.0025409