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It is well known that bacteria are able to sense their environment, differentiate between cell types, and deliver proteins to eukaryotic cells. {{:Team:TU-Eindhoven/Template:Ref | id=VoigtControlledInvasionCancerCellsI | author=J.C. Anderson, E.J. Clarke, A.P. Arkin, and C.A. Voigt | title=Environmentally Controlled Invasion of Cancer Cells by Engineered Bacteria | journal=Molecular Biology | edition=355 | pages=619-627 | year=2006 }} Furthermore, they are relatively easy to manipulate genetically and possess motility abilities, allowing them to reach specific targets e.g. tumors. These characteristics make them an intriguing alternative to traditional approaches used for treating or studying a certain disease. <!--Literature shows that in recent years, the use of bacteria for cancer therapies has gain popularity and it is now a very attractive and promising research area. {{:Team:TU-Eindhoven/Template:Ref | id=GaoCureCancerI | author=K. Gao | title=E. coli: The Cure for Cancer? | journal=The Triple Helix | edition= | pages=12-13 | year=2010 }}{{:Team:TU-Eindhoven/Template:Ref | id=BugajNextGenTher | author=L.J. Bugaj, D.V. Schaffer | title=Bringing next-generation therapeutics to the clinic through synthetic biology | journal=Current Opinion in Chemical Biology | edition=16 | pages=355-361 | year=2012 }}{{:Team:TU-Eindhoven/Template:Ref | id=KhanBacCan | author=A.A. Khan | title=Bacteria and cancer | journal=Springer | edition= | pages= | year=2012 }}{{:Team:TU-Eindhoven/Template:Ref | id=LeeBacTumTar | author=C.H. Lee | title=Engineering bacteria toward tumor targeting for cancer treatment: current state and perspectives | journal=Appl Microbiol Biotechnol | edition=93 | pages=517-523 | year=2012 }}{{:Team:TU-Eindhoven/Template:Ref | id=MengeshaBactVect | author=A. Mengesha | title=Use of Non-Pathogenic Bacteria as Vectors for Targeted Gene Expression in Cancer Gene Therapy | journal=Procopia | edition= | pages= | year=2009 }}--> | It is well known that bacteria are able to sense their environment, differentiate between cell types, and deliver proteins to eukaryotic cells. {{:Team:TU-Eindhoven/Template:Ref | id=VoigtControlledInvasionCancerCellsI | author=J.C. Anderson, E.J. Clarke, A.P. Arkin, and C.A. Voigt | title=Environmentally Controlled Invasion of Cancer Cells by Engineered Bacteria | journal=Molecular Biology | edition=355 | pages=619-627 | year=2006 }} Furthermore, they are relatively easy to manipulate genetically and possess motility abilities, allowing them to reach specific targets e.g. tumors. These characteristics make them an intriguing alternative to traditional approaches used for treating or studying a certain disease. <!--Literature shows that in recent years, the use of bacteria for cancer therapies has gain popularity and it is now a very attractive and promising research area. {{:Team:TU-Eindhoven/Template:Ref | id=GaoCureCancerI | author=K. Gao | title=E. coli: The Cure for Cancer? | journal=The Triple Helix | edition= | pages=12-13 | year=2010 }}{{:Team:TU-Eindhoven/Template:Ref | id=BugajNextGenTher | author=L.J. Bugaj, D.V. Schaffer | title=Bringing next-generation therapeutics to the clinic through synthetic biology | journal=Current Opinion in Chemical Biology | edition=16 | pages=355-361 | year=2012 }}{{:Team:TU-Eindhoven/Template:Ref | id=KhanBacCan | author=A.A. Khan | title=Bacteria and cancer | journal=Springer | edition= | pages= | year=2012 }}{{:Team:TU-Eindhoven/Template:Ref | id=LeeBacTumTar | author=C.H. Lee | title=Engineering bacteria toward tumor targeting for cancer treatment: current state and perspectives | journal=Appl Microbiol Biotechnol | edition=93 | pages=517-523 | year=2012 }}{{:Team:TU-Eindhoven/Template:Ref | id=MengeshaBactVect | author=A. Mengesha | title=Use of Non-Pathogenic Bacteria as Vectors for Targeted Gene Expression in Cancer Gene Therapy | journal=Procopia | edition= | pages= | year=2009 }}--> | ||
- | According to different researches, genetically engineered bacteria can be delivered as therapy given that several bacteria, e.g. as Escherichia, Clostridium, and Salmonella, naturally target and accumulate in tumor regions when injected into the bloodstream. {{:Team:TU-Eindhoven/Template:RefAgain | id=LeeBacTumTar }} Bacteria are able to invade tumors due to the hypoxic microenvironment, poor immune surveillance and increased availability of nutrients. The most current research being done with bacteria and tumors focuses on inducing cell death within the tumor by means of a cytotoxic agent that is delivered by the bacteria. {{:Team:TU-Eindhoven/Template:RefAgain | id=BugajNextGenTher }}{{:Team:TU-Eindhoven/Template:RefAgain | id=LeeBacTumTar }} However there is another potential application for bacteria in what concerns tumors which is the imaging of the latter | + | According to different researches, genetically engineered bacteria can be delivered as therapy given that several bacteria, e.g. as Escherichia, Clostridium, and Salmonella, naturally target and accumulate in tumor regions when injected into the bloodstream. {{:Team:TU-Eindhoven/Template:RefAgain | id=LeeBacTumTar }} Bacteria are able to invade tumors due to the hypoxic microenvironment, poor immune surveillance and increased availability of nutrients. The most current research being done with bacteria and tumors focuses on inducing cell death within the tumor by means of a cytotoxic agent that is delivered by the bacteria. {{:Team:TU-Eindhoven/Template:RefAgain | id=BugajNextGenTher }}{{:Team:TU-Eindhoven/Template:RefAgain | id=LeeBacTumTar }} However there is another potential application for bacteria in what concerns tumors which is the imaging of the latter. Bacterial-aided delivery of imaging agents has allowed tumor visualization though PET {{:Team:TU-Eindhoven/Template:Ref | id=BraderAlEColiTumorDect | author=P. Brader, J. Stritzker, C.C. Riedl, P. Zanzonico, S. Cai, E.M. Burnazi, E.R. Ghani, H. Hricak, A.A. Szalay, Y. Fong et al | title=Escherichia coli Nissle 1917 facilitates tumor detection by positron emission tomography and optical imaging | journal=Clin Cancer Res | edition=14 | pages=2295-2302 | year=2008 }}{{:Team:TU-Eindhoven/Template:Ref | id=SogAlPETBact | author=S.A. Soghomonyan, M. Doubrovin, J. Pike, X. Luo, M. Ittensohn, J.D. Runyan, j. Balatoni, R. Finn, J.G. Tjuvajev, R. Blasberg et al | title=Positron emission tomography (PET) imaging of tumorlocalized Salmonella expressing HSV1-TK | journal=Cancer Gene Ther | edition=12 | pages=101-108 | year=2004 }}, MRI {{:Team:TU-Eindhoven/Template:Ref | id=BenoitAlMRI | author=M.R. Benoit, D. Mayer, Y. Barak, I.Y. Chen, W. Hu, Z. Cheng, S.X. Wang, D.M. Spielman, S.S. Gambhir, A. Matin | title=Visualizing implanted tumors in mice with magnetic resonance imaging using magnetotactic bacteria | journal=Clin Cancer Res | edition=15 | pages=5170-5177 | year=2009 }}{{:Team:TU-Eindhoven/Template:Ref | id=DresselaersAlMRI | author=T. Dresselaers, J. Theys, S. Nuyts, B. Wouters, E. de Bruijn, J. Anne, P. Lambin, P. Van Hecke, W. Landuyt | title=Non-invasive 19F MR spectroscopy of 5-fluorocytosine to 5-fluorouracil conversion by recombinant Salmonella in tumours | journal=Br J Cancer | edition=89 | pages=1796-1801 | year=2003 }}{{:Team:TU-Eindhoven/Template:Ref | id=HillAlMRI | author=P.J. Hill, J. Stritzker, M. Scadeng, U. Geissinger, D. Haddad, T.C. Basse- Lu¨ sebrink, U. Gbureck, P. Jakob, A.A. Szalay | title=Magnetic resonance imaging of tumors colonized with bacterial ferritinexpressing Escherichia coli | journal=PLoS One | edition=6 | pages= | year=2011 }}, fluorescence {{:Team:TU-Eindhoven/Template:Ref | id=YuAlFluor | author=Y.A. Yu, S. Shabahang, T.M. Timiryasova, Q. Zhang, R. Beltz, I. Gentschev, W. Goebel, A.A. Szalay | title=Visualization of tumors and metastases in live animals with bacteria and vaccinia virus encoding light emitting proteins | journal=Nat Biotechnol | edition=22 | pages=313-320 | year=2004 }}{{:Team:TU-Eindhoven/Template:Ref | id=ZhaoAlFluor | author=M. Zhao, M. Yang, X.-M. Li, P. Jiang, E. Baranov, S. Li, M. Xu, S. Penman, R.M. Hoffman | title=Tumor-targeting bacterial therapy with amino acid auxotrophs of GFP-expressing Salmonella typhimurium | journal=Proc Natl Acad Sci USA | edition=102 | pages=755-760 | year=2005 }} and bioluminescence {{:Team:TU-Eindhoven/Template:Ref | id=ChengAlBiolum | author=C.M. Cheng, Y.L. Lu, K.H. Chuang, W.C. Hung, J. Shiea, Y.C. Su, C.H. Kao, B.M. Chen, S. Roffler, T.L. Cheng | title=Tumor-targeting prodrug-activating bacteria for cancer therapy | journal=Cancer Gene Ther | edition=15 | pages=393-401 | year=2008 }}{{:Team:TU-Eindhoven/Template:Ref | id=CroninAlImBiolum | author=M. Cronin, A.R. Akin, S.A. Collins, J.Meganck, J.-B. Kim, C.K. Baban, S.A. Joyce, G.M. van Dam, N. Zhang, D. van Sinderen et al | title=High resolution in vivo bioluminescent imaging for the study of bacterial tumour targeting | journal=PLoS One | edition=7 | pages= | year=2011 }}. The potential of these capabilities lies in the possibility of simultaneously treating and visualizing a specific tumor progression and metastasis in a non-invasive manner. Additionally, targeted fluorescent protein delivery to tumors has been demonstrated to aid surgeons in tumor removal procedures. {{:Team:TU-Eindhoven/Template:Ref | id=NguyenAlFluor | author=Q.T. Nguyen, E.S. Olson, T.A. Aguilera, T. Jiang, M. Scadeng, L.G. Ellies, R.Y Tsien | title=Surgery with molecular fluorescence imaging using activatable cell-penetrating peptides decreases residual cancer and improves survival | journal=Proc Natl Acad Sci USA | edition= | pages= | year=2010 }} Tumor-specific bacterial delivery of imaging proteins may increase the opportunities of visualizing and treating different types of tumors, with a higher signal-to-noise ratio between tumor and non-tumor tissue. Given that the CEST contrast production from our device is triggered by hypoxic environment, it could also be possible to link it to bacterial infections considering that hypoxia is also a feature of tissues experiencing a bacterial infection. {{:Team:TU-Eindhoven/Template:Ref | id=SawyerOxInf | author=R. G. Sawyer, M. D. Spengler, R. B. Adams, T. L. Pruett | title=The peritoneal environment during infection: the effect of monomicrobial and polymicrobial bacteria on pO2 and pH | journal=Ann Surg | edition=213 | pages=253-260 | year=1991 }} <!--The hypoxic state is due to increased oxygen consumption by the proliferation of bacteria and vasoconstriction of vessels in the area. --> |
- | Inspired by this type of research | + | Inspired by this type of research, we propose two main applications which follow the same idea of targeting a specific region and delivering a compound to it. The first application is intended to be used in humans for imaging tumors when sensing a hypoxic environment. The second application is meant for bacterial tracking research in animals by systemic administration of a bacterial concentrated solution. |
==First Application: Tumor Imaging== | ==First Application: Tumor Imaging== |
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Contents |
Where and How to Use Our Device
It is well known that bacteria are able to sense their environment, differentiate between cell types, and deliver proteins to eukaryotic cells. VoigtControlledInvasionCancerCellsIJ.C. Anderson, E.J. Clarke, A.P. Arkin, and C.A. Voigt, Environmentally Controlled Invasion of Cancer Cells by Engineered Bacteria. Molecular Biology 355, 619-627 (2006) Furthermore, they are relatively easy to manipulate genetically and possess motility abilities, allowing them to reach specific targets e.g. tumors. These characteristics make them an intriguing alternative to traditional approaches used for treating or studying a certain disease.
According to different researches, genetically engineered bacteria can be delivered as therapy given that several bacteria, e.g. as Escherichia, Clostridium, and Salmonella, naturally target and accumulate in tumor regions when injected into the bloodstream. LeeBacTumTar Bacteria are able to invade tumors due to the hypoxic microenvironment, poor immune surveillance and increased availability of nutrients. The most current research being done with bacteria and tumors focuses on inducing cell death within the tumor by means of a cytotoxic agent that is delivered by the bacteria. BugajNextGenTherLeeBacTumTar However there is another potential application for bacteria in what concerns tumors which is the imaging of the latter. Bacterial-aided delivery of imaging agents has allowed tumor visualization though PET BraderAlEColiTumorDectP. Brader, J. Stritzker, C.C. Riedl, P. Zanzonico, S. Cai, E.M. Burnazi, E.R. Ghani, H. Hricak, A.A. Szalay, Y. Fong et al, Escherichia coli Nissle 1917 facilitates tumor detection by positron emission tomography and optical imaging. Clin Cancer Res 14, 2295-2302 (2008)SogAlPETBactS.A. Soghomonyan, M. Doubrovin, J. Pike, X. Luo, M. Ittensohn, J.D. Runyan, j. Balatoni, R. Finn, J.G. Tjuvajev, R. Blasberg et al, Positron emission tomography (PET) imaging of tumorlocalized Salmonella expressing HSV1-TK. Cancer Gene Ther 12, 101-108 (2004), MRI BenoitAlMRIM.R. Benoit, D. Mayer, Y. Barak, I.Y. Chen, W. Hu, Z. Cheng, S.X. Wang, D.M. Spielman, S.S. Gambhir, A. Matin, Visualizing implanted tumors in mice with magnetic resonance imaging using magnetotactic bacteria. Clin Cancer Res 15, 5170-5177 (2009)DresselaersAlMRIT. Dresselaers, J. Theys, S. Nuyts, B. Wouters, E. de Bruijn, J. Anne, P. Lambin, P. Van Hecke, W. Landuyt, Non-invasive 19F MR spectroscopy of 5-fluorocytosine to 5-fluorouracil conversion by recombinant Salmonella in tumours. Br J Cancer 89, 1796-1801 (2003)HillAlMRIP.J. Hill, J. Stritzker, M. Scadeng, U. Geissinger, D. Haddad, T.C. Basse- Lu¨ sebrink, U. Gbureck, P. Jakob, A.A. Szalay, Magnetic resonance imaging of tumors colonized with bacterial ferritinexpressing Escherichia coli. PLoS One 6, (2011), fluorescence YuAlFluorY.A. Yu, S. Shabahang, T.M. Timiryasova, Q. Zhang, R. Beltz, I. Gentschev, W. Goebel, A.A. Szalay, Visualization of tumors and metastases in live animals with bacteria and vaccinia virus encoding light emitting proteins. Nat Biotechnol 22, 313-320 (2004)ZhaoAlFluorM. Zhao, M. Yang, X.-M. Li, P. Jiang, E. Baranov, S. Li, M. Xu, S. Penman, R.M. Hoffman, Tumor-targeting bacterial therapy with amino acid auxotrophs of GFP-expressing Salmonella typhimurium. Proc Natl Acad Sci USA 102, 755-760 (2005) and bioluminescence ChengAlBiolumC.M. Cheng, Y.L. Lu, K.H. Chuang, W.C. Hung, J. Shiea, Y.C. Su, C.H. Kao, B.M. Chen, S. Roffler, T.L. Cheng, Tumor-targeting prodrug-activating bacteria for cancer therapy. Cancer Gene Ther 15, 393-401 (2008)CroninAlImBiolumM. Cronin, A.R. Akin, S.A. Collins, J.Meganck, J.-B. Kim, C.K. Baban, S.A. Joyce, G.M. van Dam, N. Zhang, D. van Sinderen et al, High resolution in vivo bioluminescent imaging for the study of bacterial tumour targeting. PLoS One 7, (2011). The potential of these capabilities lies in the possibility of simultaneously treating and visualizing a specific tumor progression and metastasis in a non-invasive manner. Additionally, targeted fluorescent protein delivery to tumors has been demonstrated to aid surgeons in tumor removal procedures. NguyenAlFluorQ.T. Nguyen, E.S. Olson, T.A. Aguilera, T. Jiang, M. Scadeng, L.G. Ellies, R.Y Tsien, Surgery with molecular fluorescence imaging using activatable cell-penetrating peptides decreases residual cancer and improves survival. Proc Natl Acad Sci USA , (2010) Tumor-specific bacterial delivery of imaging proteins may increase the opportunities of visualizing and treating different types of tumors, with a higher signal-to-noise ratio between tumor and non-tumor tissue. Given that the CEST contrast production from our device is triggered by hypoxic environment, it could also be possible to link it to bacterial infections considering that hypoxia is also a feature of tissues experiencing a bacterial infection. SawyerOxInfR. G. Sawyer, M. D. Spengler, R. B. Adams, T. L. Pruett, The peritoneal environment during infection: the effect of monomicrobial and polymicrobial bacteria on pO2 and pH. Ann Surg 213, 253-260 (1991)
Inspired by this type of research, we propose two main applications which follow the same idea of targeting a specific region and delivering a compound to it. The first application is intended to be used in humans for imaging tumors when sensing a hypoxic environment. The second application is meant for bacterial tracking research in animals by systemic administration of a bacterial concentrated solution.
First Application: Tumor Imaging
![TU-Eindhoven Images ApplicationIDiagram.png](/wiki/images/2/2a/TU-Eindhoven_Images_ApplicationIDiagram.png)
The idea behind the first application is to generate the CEST MRI contrast agent on site within the tumor region, facilitating in this manner tumor localization. Considering that the contrast agent will be generated only if the bacteria sense a hypoxic environment, it can be said that using our contrast agent would allow physicians to identify tumors in an accurate way. presents a graphic description of the process that would be followed by using the CEST MRI contrast agent to image tumors in humans. First, a bacterial concentrated solution is injected into the patient. Then, the bacteria in the solution travel through the bloodstream without alerting the immune system. Third, they accumulate in the tumor region due to their ability of sensing hypoxic environments. The anaerobic conditions activate the CEST proteins production. Once the CEST proteins have been produced, the CEST MRI can be taken and analyzed. Finally, the bacteria can be disabled and killed by means of activating the kill switch. The switch is enabled by the systemic administration of the prodrug Ganciclovir which induces apoptosis only in the cells that contain the HSV-thymidine kinase, in this case the CEST proteins producing bacteria.
In order to use our device for this application it is necessary to modify the chassis to include two crucial components that enable the bacteria to travel through the bloodstream without raising an alert in the immune system and die when the kill switch is activated. The first change that needs to be done to the chassis is to modify its surface by exchanging the carbohydrate chains commonly found on it for a carbohydrate layer instead. This alteration will prevent innate and adaptive immune responses, allowing the bacteria to freely travel to the tumor region without being engulfed by phagocytic cells. This solution to the immune system’s response was based on the solutions implemented by the Berkeley iGEM team in 2007 and the research of Dr. J. Christopher Anderson, an assistant professor of bioengineering at the University of California, Berkeley. The Berkeley iGEM team in 2007 proposed to display K1 carbohydrates. K1 carbohydrates are long linear polymers of sialic acid that extend about half the diameter of the bacteria’s surface. Given that polysialic acid is a typical coating on mammalian cells, the human immune system does not recognize K1 as foreign and hence the bacteria are disguised and granted access through the bloodstream. To display K1 carbohydrate in our bacteria it is necessary to also express O antigens, they suggest expressing antigen O16. The second modification to the chassis is to include HSV-thymidine kinase for the kill mechanism to function adequately.
Results:Pharmacokinetic modeling of the distribution of the CEST protein producing bacteria in the human body and their elimination after CEST MRI
A pharmacokinetic model of the administration and elimination of the CEST protein producing bacteria concentrated solution was performed based on parameters from the available literature. The model is described in detail in the Drylab section.
Second Application: Tracking of Bacteria in Bacterial Infections Research
![TU-Eindhoven Images ApplicationIIDiagram.png](/wiki/images/4/44/TU-Eindhoven_Images_ApplicationIIDiagram.png)
Similarly to the first application, the second application also requires the systemic administration of the CEST MRI protein producing bacteria in a concentrated solution. The idea behind this application is to use our device as a tool for bacterial infections research, by allowing the researchers to trace the bacteria inside the animal specimen subject to research. explains the general process that should be followed within this application. First the bacterial concentrated solution is injected intravenously. Once in the bloodstream the CEST contrast producing bacteria travel through it until they find their target, which varies according to the infection being studied. Then a CEST MR image is acquired and the data is analyzed. Although according to the literature hypoxia is a suitable trigger for this application given that it is a characteristic of tissues experiencing bacterial infections, it could be possible to tune the activation mechanism based on a certain group of bacterial infections.
Considering that both applications are similar in principle, no specific model was developed for the second application, but it is possible to modify the parameters of the pharmacokinetic modeled used in the tumor imaging application to fit the requirements of the second application.
Strengths and Weaknesses of our device in both applications
The ideas we suggest with the two applications previously described are novel in the sense that they take the principles of CEST MRI and delivery systems and apply it to in vivo production of a CEST contrast agent to enhance the acquired MR images. As every other application, what we propose also has advantages and disadvantages. The following table presents the most important strengths and weaknesses that the proposed applications comprise.
Application | Strengths | Weaknesses |
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Tumor Imaging |
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Bacterial Tracking |
The strongest point of what we propose is the possibility of producing in situ only the required amount of CEST contrast for the image acquisition, summed to the fact that it enables the physician or researcher to easily localize the damaged region. As for the greatest weakness they both have, it is not the potential mutation risks that the device could develop with bacterial replication, the chances of the device presenting a mutation that inactivates the safety locks and jeopardizes the functionality of the device and health of the patient are one per million. GaoCureCancerI The greatest risk to the success of these applications is the difficulties society might have with coping with the idea of injecting live bacteria into a patient’s bloodstream as a tool for studying, detecting, and treating diseases like cancer or bacterial infections. This main obstacle can only be surpassed by providing people with the proper unbiased knowledge sources related to synthetic biology and the use of bacteria for clinical and research applications. Therefore, as our human practice project, we have developed an online source of knowledge with links to a variety of papers and other sources related to a specific topic or area of synthetic biology. In this webpage anybody can read about a variety of myths and facts that have been revised by different experts, and are now available as a trustful unbiased source. To know more about it please visit our human practice section in the Wiki.
References