Name: utilizing high resolution ultrasound to monitor tumor onset and growth in genetically engineered pancreatic cancer models
Uploaded: 2018-04-07T13:00:00
Description: Watch this Scientific Journal Video about Utilizing High Resolution Ultrasound to Monitor Tumor Onset and Growth in Genetically Engineered Pancreatic Cancer Models at JoVE.com

This research was supported by the Deutsche Krebshilfe (Max Eder Group to AN: 110972), a DGVS doctoral thesis scholarship (to SMB), and an Else-Kr&#246;ner-Fresenius-Foundation scholarship (to RGG) at the University Medical Center Goettingen. We thank Jutta Blumberg and Ulrike Wegner for expert technical assistance. We also thank all animal technicians at the animal facility of the University Medical Center Goettingen for mouse keeping. All experiments were performed according to German animal welfare regulations.

This protocol is in accordance with the animal care guidelines at the University Medical Centre Goettingen, Germany (33.9-42502-04-15/2056). Depending on specific requirements of individual animal review boards, some of the protocol steps could be modified accordingly.
1. Abdominal Palpation of KPC Mice
<ol>
	<li>To avoid unnecessary ultrasound scans, palpate the mouse abdomen to identify mice which might possibly bear an intraabdominal lesion and should subsequently undergo abdominal ultras...

<ol>
	<li>Kersten, K., de Visser, K. E., van Miltenburg, M. H., Jonkers, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetically+engineered+mouse+models+in+oncology+research+and+cancer+medicine.">Genetically engineered mouse models in oncology research and cancer medicine.</a> EMBO Molecular Medicine. 9 (2), 137-153 (2017).</li><li>Olive, K. P., Politi, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Translational therapeutics in genetically engineered mouse models of cancer. (2), 131-143 (2014).</li><li>Westphalen, C. B., Olive, K. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetically+engineered+mouse+models+of+pancreatic+cancer.">Genetically engineered mouse models of pancreatic cancer.</a> The Cancer Journal. 18 (6), 502-510 (2012).</li><li>Hingorani, S. R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trp53R172H+and+KrasG12D+cooperate+to+promote+chromosomal+instability+and+widely+metastatic+pancreatic+ductal+adenocarcinoma+in+mice.">Trp53R172H and KrasG12D cooperate to promote chromosomal instability and widely metastatic pancreatic ductal adenocarcinoma in mice.</a> Cancer Cell. 7 (5), 469-483 (2005).</li><li>Frese, K. K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=nab-Paclitaxel+potentiates+gemcitabine+activity+by+reducing+cytidine+deaminase+levels+in+a+mouse+model+of+pancreatic+cancer.">nab-Paclitaxel potentiates gemcitabine activity by reducing cytidine deaminase levels in a mouse model of pancreatic cancer.</a> Cancer Discovery. 2 (3), 260-269 (2012).</li><li>Paredes, J. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+non-invasive+method+of+quantifying+pancreatic+volume+in+mice+using+micro-MRI.">A non-invasive method of quantifying pancreatic volume in mice using micro-MRI.</a> PLoS One. 9 (3), e92263 (2014).</li><li>Boj, S. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Organoid+models+of+human+and+mouse+ductal+pancreatic+cancer.">Organoid models of human and mouse ductal pancreatic cancer.</a> Cell. 160 (1-2), 324-338 (2015).</li><li>Aung, W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Immunotargeting+of+Integrin+alpha6beta4+for+Single-Photon+Emission+Computed+Tomography+and+Near-Infrared+Fluorescence+Imaging+in+a+Pancreatic+Cancer+Model.">Immunotargeting of Integrin alpha6beta4 for Single-Photon Emission Computed Tomography and Near-Infrared Fluorescence Imaging in a Pancreatic Cancer Model.</a> Molecular Imaging. 15, (2016).</li><li>Akladios, C. Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contribution+of+microCT+structural+imaging+to+preclinical+evaluation+of+hepatocellular+carcinoma+chemotherapeutics+on+orthotopic+graft+in+ACI+rats.">Contribution of microCT structural imaging to preclinical evaluation of hepatocellular carcinoma chemotherapeutics on orthotopic graft in ACI rats.</a> Bulletin du Cancer. 98 (2), 120-132 (2011).</li><li>Neesse, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=CTGF+antagonism+with+mAb+FG-3019+enhances+chemotherapy+response+without+increasing+drug+delivery+in+murine+ductal+pancreas+cancer.">CTGF antagonism with mAb FG-3019 enhances chemotherapy response without increasing drug delivery in murine ductal pancreas cancer.</a> Proceedings of the National Academy of Science USA. 110 (30), 12325-12330 (2013).</li><li>Sastra, S. A., Olive, K. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantification+of+murine+pancreatic+tumors+by+high-resolution+ultrasound.">Quantification of murine pancreatic tumors by high-resolution ultrasound.</a> Methods in Molecular Biology. 980, 249-266 (2013).</li></ol>

With this protocol, a detailed description for quantifying pancreatic tumors using high-resolution abdominal ultrasound imaging in genetically engineered mouse models is provided. Recently, Sastra et al. published a detailed description how to quantify pancreatic tumors in mouse models, but no visualized instructions about the preparation and handling as prerequisite for all further steps were shown11. The overall goal of this manuscript is to provide a comprehensive visual guide for high...

Genetically engineered mouse models have gained an increasing importance in cancer research due to their ability to closely recapitulate the complex nature of human carcinogenesis1,2,3. One of the most frequently used models to study pancreatic cancer development, progression and therapeutic response is characterized by an activating mutation in the Kras oncogene combined with an inactivation of the tumor suppressor p534. This LSL-KrasG12D/+; LSL-Trp53R172H/+; Pdx-1-Cre (KPC) mouse model mimics the step-wise progression from pre-invasive pancreatic intraepithelial neoplasia (PanIN) lesions to invasive carcinoma. Phenotypically, nearly all mice develop PDAC within the first six months after birth. However, compared to transplanted models, the KPC model reveals a highly variable tumor onset from 8 weeks to several months4. Once pancreatic tumors reach a certain size (5-9 mm in diameter), tumor growth accelerates rapidly and mice will have to be enrolled in preclinical trials5. Therefore, the exact detection of tumor onset and tumor size is an essential prerequisite for preclinical study logistics and therapy monitoring. In general, several approaches like magnetic resonance imaging (MRI)6, computed tomography scanning7,8,9 or high resolution ultrasound can be employed to conduct tumor screening and therapy10. Each technique has its advantages and drawbacks. Although MRI-or computed tomography (CT) -imaging allows high resolution data acquisition as well as accurate volume calculation, prolonged examination time under general sedation, and very expensive equipment is required, and does not permit frequent scanning over a long period of time. In contrast, small animal ultrasonography is an established method that can be employed to screen for abdominal pathologies in mice11. Advantages of this imaging method are short scanning times, high resolution, and the possibility to use doppler ultrasound or contrast enhanced ultrasound (CEUS) to visualize perfusion of organs in parallel. However, anatomical knowledge, 3D imagination and thorough practical training are required for correct image interpretation.
In the following article, a detailed protocol for utilizing high resolution ultrasound in the KPC model is provided. Furthermore, standard ultrasound images are depicted and labelled with organ structures to facilitate orientation for the investigator.

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			<td>Visual Sonics Vevo2100 High Resolution Ultrasound System, including imaging stage and anesthesia line</td>
			<td>FUJIFILM VisualSonics Inc, Canada</td>
			<td>VS-11945</td>
			<td></td>
		</tr>
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			<td>Vevo 2100 MicroScan Transducer MS-550-D (22-55MHz)</td>
			<td>FUJIFILM VisualSonics Inc, Canada</td>
			<td>VS-11874</td>
			<td></td>
		</tr>
		<tr>
			<td>Vevo Anesthesia System (anesthesia induction chamber with fresh and waste gas inlet)</td>
			<td>FUJIFILM VisualSonics Inc, Canada</td>
			<td>SA-12055</td>
			<td></td>
		</tr>
		<tr>
			<td>Vevo Imaging Station (working stage with nose cone for anesthesia supply)</td>
			<td>FUJIFILM VisualSonics Inc, Canada</td>
			<td>SA-11982</td>
			<td></td>
		</tr>
		<tr>
			<td>&#160;electronic pet clippers</td>
			<td>Panasonic Marketing Europe, Germany</td>
			<td>5025232484324</td>
			<td>Panasonic ER-PA10-s</td>
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			<td>Labotect Hot plate</td>
			<td>Labor tech G&#246;ttingen, Germany</td>
			<td>13854</td>
			<td></td>
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			<td>eye cream (ophthalmic ointment)</td>
			<td>Sch&#252;lke&#38;Mayr, Germany</td>
			<td>9080249</td>
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			<td>veterinary isoflurane</td>
			<td>Abbvie, Germany</td>
			<td>4831867</td>
			<td></td>
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			<td>depilatory cream</td>
			<td>RB healthcare UK, United Kingdom</td>
			<td>8218535</td>
			<td></td>
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			<td>70% ethanol (v/v) in distilled water</td>
			<td>TH. Geyer, Germany</td>
			<td>22941000</td>
			<td></td>
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			<td>ultrasound gel</td>
			<td>Asmuth, Germany</td>
			<td>13477</td>
			<td></td>
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			<td>tissue wipes</td>
			<td>Kimberly-Clark Germany, Germany</td>
			<td>7558</td>
			<td></td>
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			<td>cotton tips</td>
			<td>Meditrade, Germany</td>
			<td>75481116</td>
			<td></td>
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		<tr>
			<td>glass bowl for ultrasound gel</td>
			<td>ARC France, France</td>
			<td>H1149</td>
			<td></td>
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		<tr>
			<td>water bowl</td>
			<td>W &#38; P Trading Co., USA</td>
			<td>B00K2P6PLQ</td>
			<td></td>
		</tr>
		<tr>
			<td>gauze sponges</td>
			<td>Fuhrmann, Germany</td>
			<td>960504</td>
			<td></td>
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utilizing high resolution ultrasound to monitor tumor onset and growth in genetically engineered pancreatic cancer models

The LSL-KrasG12D/+; LSL-Trp53R172H/+; Pdx-1-Cre (KPC) mouse model represents an established and frequently used transgenic model to evaluate novel therapies in pancreatic cancer. Tumor onset is variable in the KPC model between 8 weeks and several months. Therefore, non-invasive imaging tools are required to screen for tumor onset and monitor for response to treatment. To address this issue, different approaches have emerged over the last years. High resolution ultrasound has major advantages such as non-invasiveness, fast session times and a high image resolution without radiation exposure. However, ultrasound in mice is not trivial and sufficient anatomical knowledge and practical skills are required to successfully perform high resolution ultrasound in preclinical pancreatic cancer models. With the following article, a detailed hands-on guide for abdominal ultrasound in murine models with a particular focus on endogenous pancreatic cancer models is presented. Furthermore, a summary of common mistakes and how to avoid them is provided.

Ultrasound imaging is a versatile and non-invasive technique that is used to address several issues in murine models of human diseases. Compared to all other imaging approaches major advantages are high-throughput, cost efficiency, short acquisition time and real-time imaging. However, this tool needs expertise to generate accurate, high quality images. Particularly in the case of unwanted artefacts at least some experience with ultrasound imaging in general is very helpful. In relation t...

Watch this Scientific Journal Video about Utilizing High Resolution Ultrasound to Monitor Tumor Onset and Growth in Genetically Engineered Pancreatic Cancer Models at JoVE.com

Utilizing High Resolution Ultrasound to Monitor Tumor Onset and Growth in Genetically Engineered Pancreatic Cancer Models

This article describes the utilization of high-resolution ultrasound in genetically engineered pancreatic cancer mice. The primary aim is to provide a detailed instruction for detection and evaluation of endogenous pancreatic tumors.

The LSL-KrasG12D/+; LSL-Trp53R172H/+; Pdx-1-Cre (KPC) mouse model represents an established and frequently used transgenic model to evaluate novel ...

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