Name: monitoring breast cancer growth and metastatic colony formation in mice using bioluminescence
Uploaded: 2021-11-05T13:00:00
Description: Watch this Scientific Journal Video about Monitoring Breast Cancer Growth and Metastatic Colony Formation in Mice using Bioluminescence at JoVE.com

We thank the members of the Y.D.S. laboratory. We would like to thank The Wohl Institute for Translational Medicine at the Hadassah Medical Center, Jerusalem, for providing the small animal imaging facility. This study was supported by Research Career Development Award from the Israel Cancer Research Fund.

All mouse experiments were carried out under the Hebrew University Institutional Animal Care and Use Committee-approved protocol MD-21-16429-5. In addition, the Hebrew University is certified by the Association for Assessment and Accreditation of Laboratory Animal Care (AAALAC).
1. Cell line maintenance
NOTE: The human breast cancer cell lines (MCF-7, MDA-MB-468, and MDA-MB-231) were used in this protocol.
<ol>
	<li>Culture all the breast cancer cell lines in Dulb...

<ol>
	<li>Waks, A. G., Winer, E. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Breast+cancer+treatment:+A+review.">Breast cancer treatment: A review.</a> JAMA. 321 (3), 288-300 (2019).</li><li>Jin, X., Mu, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Targeting+breast+cancer+metastasis.">Targeting breast cancer metastasis.</a> Breast Cancer: Basic and Clinical Research. 9, 23-34 (2015).</li><li>Saha, D., 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=In+vivo+bioluminescence+imaging+of+tumor+hypoxia+dynamics+of+breast+cancer+brain+metastasis+in+a+mouse+model.">In vivo bioluminescence imaging of tumor hypoxia dynamics of breast cancer brain metastasis in a mouse model.</a> Journal of Visualized Experiments: JoVE. (56), e3175 (2011).</li><li>Rashid, O. M., 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=Is+tail+vein+injection+a+relevant+breast+cancer+lung+metastasis+model.">Is tail vein injection a relevant breast cancer lung metastasis model.</a> Journal of Thoracic Disease. 5 (4), 385-392 (2013).</li><li>Fantozzi, A., Christofori, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mouse+models+of+breast+cancer+metastasis.">Mouse models of breast cancer metastasis.</a> Breast Cancer Research. 8 (4), 212 (2006).</li><li>Kocat&#252;rk, B., Versteeg, H. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Orthotopic+injection+of+breast+cancer+cells+into+the+mammary+fat+pad+of+mice+to+study+tumor+growth.">Orthotopic injection of breast cancer cells into the mammary fat pad of mice to study tumor growth.</a> Journal of Visualized Experiments: JoVE. (96), e51967 (2015).</li><li>Baker, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+whole+picture.">The whole picture.</a> Nature. 463 (7283), 977-979 (2010).</li><li>Wang, Y., Tseng, J. -. C., Sun, Y., Beck, A. H., Kung, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Noninvasive+imaging+of+tumor+burden+and+molecular+pathways+in+mouse+models+of+cancer.">Noninvasive imaging of tumor burden and molecular pathways in mouse models of cancer.</a> Cold Spring Harbor Protocols. 2015 (2), 135-144 (2015).</li><li>Kim, J. E., Kalimuthu, S., Ahn, B. -. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+cell+tracking+with+bioluminescence+imaging.">In vivo cell tracking with bioluminescence imaging.</a> Nuclear Medicine and Molecular Imaging. 49 (1), 3-10 (2015).</li><li>Paschall, A. V., Liu, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+orthotopic+mouse+model+of+spontaneous+breast+cancer+metastasis.">An orthotopic mouse model of spontaneous breast cancer metastasis.</a> Journal of Visualized Experiments: JoVE. (114), (2016).</li><li>Morten, B. C., Scott, R. J., Avery-Kiejda, K. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+Three+Different+Methods+for+Determining+Cell+Proliferation+in+Breast+Cancer+Cell+Lines.">Comparison of Three Different Methods for Determining Cell Proliferation in Breast Cancer Cell Lines.</a> Journal of Visualized Experiments. (115), e54040 (2016).</li><li>Zimmerman, M., Hu, X., Liu, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+metastasis+and+CTL+adoptive+transfer+immunotherapy+mouse+model.">Experimental metastasis and CTL adoptive transfer immunotherapy mouse model.</a> Journal of Visualized Experiments: JoVE. (45), e2077 (2010).</li><li>Lv, X., 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=Orthotopic+transplantation+of+breast+tumors+as+preclinical+models+for+breast+cancer.">Orthotopic transplantation of breast tumors as preclinical models for breast cancer.</a> Journal of Visualized Experiments: JoVE. (159), e61173 (2020).</li><li>Cheng, R. Y. S., 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=Studying+triple+negative+breast+cancer+using+orthotopic+breast+cancer+model.">Studying triple negative breast cancer using orthotopic breast cancer model.</a> Journal of Visualized Experiments: JoVE. (157), e60316 (2020).</li><li>Bajikar, S. S., 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=Tumor-suppressor+inactivation+of+GDF11+occurs+by+precursor+sequestration+in+triple-negative+breast+cancer.">Tumor-suppressor inactivation of GDF11 occurs by precursor sequestration in triple-negative breast cancer.</a> Developmental Cell. 43 (4), 418-435 (2017).</li><li>Khatib, 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=The+glutathione+peroxidase+8+(GPX8)/IL-6/STAT3+axis+is+essential+in+maintaining+an+aggressive+breast+cancer+phenotype.">The glutathione peroxidase 8 (GPX8)/IL-6/STAT3 axis is essential in maintaining an aggressive breast cancer phenotype.</a> Proceedings of the National Academy of Sciences of the United States of America. 117 (35), 21420-21431 (2020).</li></ol>

Animal-based experiments are obligatory for cancer research7,8,9, and indeed many protocols have been developed3,6,10,11,12,13,14. However, most of these studies determined the biological effect onl...

Breast cancers are frequent types of cancer worldwide, with approximately 250,000 new cases diagnosed each year in the United States1. Despite its high incidence, a new set of anticancer drugs has significantly improved breast cancer patient outcomes2. However, these treatments are still inadequate, as many patients experience disease relapse and metastatic spread to vital organs2, which is the primary cause of patient morbidity and mortality. Therefore, one of the main challenges in breast cancer research is identifying the molecular mechanisms regulating the formation of distal metastases to develop new means to inhibit their development.
Cancer metastasis is a dynamic process in which cells detach from the primary tumor and invade neighboring tissues through the blood circulation. Thus, animal models in which the cells undergo a similar metastatic cascade can facilitate the identification of the mechanisms that govern this process3,4. Additionally, these in vivo models are essential for developing breast cancer therapeutic agents5,6. However, these orthotopic models cannot indicate the actual tumor growth kinetics as the effect is only determined upon termination. Therefore, we established a luciferase-based tool to detect tumor development and metastatic colonization in real time. Additionally, these cells express GFP to detect the metastatic colonies. This approach is relatively simple and does not involve any invasive procedures3. Thus, combining luciferase and fluorescence detection is a helpful strategy to advance the preclinical studies of breast cancer therapeutics and disease management.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>1.7 mL eppendorf tubes</td>
			<td>Lifegene</td>
			<td>LMCT1.7B-500</td>
			<td></td>
		</tr>
		<tr>
			<td>10 &#181;L tips</td>
			<td>Lifegene</td>
			<td>LRT10</td>
			<td></td>
		</tr>
		<tr>
			<td>1000 &#181;L tips</td>
			<td>Lifegene</td>
			<td>LRT1000</td>
			<td></td>
		</tr>
		<tr>
			<td>15 mL tubes</td>
			<td>Lifegene</td>
			<td>LTB15-500</td>
			<td></td>
		</tr>
		<tr>
			<td>200 &#181;L tips</td>
			<td>Lifegene</td>
			<td>LRT200</td>
			<td></td>
		</tr>
		<tr>
			<td>6 well cell culture plate</td>
			<td>COSTAR</td>
			<td>3516</td>
			<td></td>
		</tr>
		<tr>
			<td>96 well Plates BLACK flat bottom</td>
			<td>Bar Naor</td>
			<td>BN30496</td>
			<td></td>
		</tr>
		<tr>
			<td>Automated Cell Counters</td>
			<td>Thermofisher</td>
			<td>A50298</td>
			<td></td>
		</tr>
		<tr>
			<td>BD FACSAria III sorter</td>
			<td>BD</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>BD Microlance 3 Needles 27 G (3/4&#39;&#39;)</td>
			<td>BD</td>
			<td>302200</td>
			<td></td>
		</tr>
		<tr>
			<td>BD Plastipak Syringes 1 mL x 120</td>
			<td>BD</td>
			<td>303172</td>
			<td></td>
		</tr>
		<tr>
			<td>Corning 100 mm x 20 mm Style Dish</td>
			<td>CORNING</td>
			<td>430167</td>
			<td></td>
		</tr>
		<tr>
			<td>Corning 150 mm x 20 mm Style Dish</td>
			<td>CORNING</td>
			<td>430599</td>
			<td></td>
		</tr>
		<tr>
			<td>Countess cell counting chamber slides</td>
			<td>Thermofisher</td>
			<td>C10228</td>
			<td></td>
		</tr>
		<tr>
			<td>Dulbecco&#39;s modified Eagle&#39;s medium (DMEM), high glucose, no glutamine</td>
			<td>Biological Industries</td>
			<td>01-055-1A</td>
			<td></td>
		</tr>
		<tr>
			<td>Eclipse 80i microscope</td>
			<td>Nikon</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>eppendorf Centrifuge 5810 R</td>
			<td>Sigma Aldrich</td>
			<td>EP5820740000</td>
			<td></td>
		</tr>
		<tr>
			<td>Fetal Bovine Serum (FBS)</td>
			<td>Biological Industries</td>
			<td>04-127-1A</td>
			<td></td>
		</tr>
		<tr>
			<td>FUW GFP</td>
			<td></td>
			<td></td>
			<td>Gifted from Dr. Yossi Buganim&#39;s lab (Hebrew University of Jerusalem)</td>
		</tr>
		<tr>
			<td>HEK293T</td>
			<td></td>
			<td></td>
			<td>Gifted from Dr. Lior Nissim&#39;s lab (Hebrew University of Jerusalem)</td>
		</tr>
		<tr>
			<td>Isoflurane, USP Terrell</td>
			<td>Piramal</td>
			<td>NDC 66794-01-25</td>
			<td></td>
		</tr>
		<tr>
			<td>IVIS Spectrum In Vivo Imaging System</td>
			<td>Perkin Elmer</td>
			<td>124262</td>
			<td></td>
		</tr>
		<tr>
			<td>L-Glutamine Solution</td>
			<td>Biological industries</td>
			<td>03-020-1A</td>
			<td></td>
		</tr>
		<tr>
			<td>Living Image Software</td>
			<td>PerkinElmer</td>
			<td></td>
			<td>bioluminescence measurement</td>
		</tr>
		<tr>
			<td>MCF-7</td>
			<td>ATCC</td>
			<td>ATCC HTB-22</td>
			<td></td>
		</tr>
		<tr>
			<td>MDA-MB-231</td>
			<td>ATCC</td>
			<td>ATCC HTB-26</td>
			<td></td>
		</tr>
		<tr>
			<td>MDA-MB-468</td>
			<td>ATCC</td>
			<td>ATCC HTB-132</td>
			<td></td>
		</tr>
		<tr>
			<td>Pasteur pipettes</td>
			<td>NORMAX</td>
			<td>2430-475</td>
			<td></td>
		</tr>
		<tr>
			<td>PBS</td>
			<td>Hylabs</td>
			<td>BP655/500D</td>
			<td></td>
		</tr>
		<tr>
			<td>pCMV-dR8.2-dvpr</td>
			<td>Addgene</td>
			<td>#8455</td>
			<td>Provided by David M. Sabatini&#8217;s lab (Whitehead institute, Boston, USA)</td>
		</tr>
		<tr>
			<td>pCMV-VSV-G</td>
			<td>Addgene</td>
			<td>#8454</td>
			<td>Provided by David M. Sabatini&#8217;s lab (Whitehead institute, Boston, USA)</td>
		</tr>
		<tr>
			<td>Penicillin-Streptomycin Solution</td>
			<td>Biological Industries</td>
			<td>03-031-1B</td>
			<td></td>
		</tr>
		<tr>
			<td>Petri dish 90 mm (90x15)</td>
			<td>MINI PLAST</td>
			<td>820-090-01-017</td>
			<td></td>
		</tr>
		<tr>
			<td>Pipettes 10ml</td>
			<td>Lifegene</td>
			<td>LG-GSP010010S</td>
			<td></td>
		</tr>
		<tr>
			<td>Pipettes 25ml</td>
			<td>Lifegene</td>
			<td>LG-GSP010050S</td>
			<td></td>
		</tr>
		<tr>
			<td>Pipettes 5ml</td>
			<td>Lifegene</td>
			<td>LG-GSP010005S</td>
			<td></td>
		</tr>
		<tr>
			<td>pLX304 Luciferase-V5 blast plasmid</td>
			<td>Addgene</td>
			<td>#98580</td>
			<td></td>
		</tr>
		<tr>
			<td>Polybrene</td>
			<td>Sigma Aldrich</td>
			<td>#107689</td>
			<td></td>
		</tr>
		<tr>
			<td>Prism 9</td>
			<td>GraphPad</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Reagent Reservoirs</td>
			<td>Bar Naor</td>
			<td>BN20621STR200TC</td>
			<td></td>
		</tr>
		<tr>
			<td>SMZ18 Stereo microscopes</td>
			<td>Nikon</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium Chloride</td>
			<td>Bio-Lab</td>
			<td>190359400</td>
			<td></td>
		</tr>
		<tr>
			<td>Syringe filters</td>
			<td>Lifegene</td>
			<td>LG-FPV403030S</td>
			<td></td>
		</tr>
		<tr>
			<td>Trypan Blue 0.5% solution</td>
			<td>Biological industries</td>
			<td>03-102-1B</td>
			<td></td>
		</tr>
		<tr>
			<td>Trypsin EDTA Solution B (0.25%), EDTA (0.05%)</td>
			<td>Biological Industries</td>
			<td>03-052-1a</td>
			<td></td>
		</tr>
		<tr>
			<td>Vacuum driven Filters</td>
			<td>SOFRA LIFE SCIENCE</td>
			<td>SPE-22-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Virusolve</td>
			<td></td>
			<td>disinfectant</td>
			<td></td>
		</tr>
		<tr>
			<td>VivoGlo Luciferin, In Vivo Grade</td>
			<td>Promega</td>
			<td>P1043</td>
			<td></td>
		</tr>
		<tr>
			<td>X-tremeGENE HP DNA Transfection Reagent</td>
			<td>Sigma Aldrich</td>
			<td>#6366236001</td>
			<td></td>
		</tr>
	</tbody>
</table>

monitoring breast cancer growth and metastatic colony formation in mice using bioluminescence

Breast cancer is a frequent heterogeneous malignancy and the second leading cause of mortality in women, mainly due to distant organ metastasis. Several animal models have been generated, including the widely used orthotopic mouse models, where cancer cells are injected into the mammary fat pad. However, these models cannot help monitor tumor growth kinetics and metastatic colonization. Cutting-edge tools to monitor cancer cells in real time in mice will significantly advance the understanding of tumor biology. 
Here, breast cancer cell lines stably expressing luciferase and green fluorescent protein (GFP) were established. Specifically, this technique contains two sequential steps initiated by measuring the luciferase activity in vitro and followed by the implantation of the cancer cells into mammary fat pads of nonobese diabetic-severe combined immunodeficiency (NOD-SCID) mice. After the injection, both the tumor growth and metastatic colonization are monitored in real time by the noninvasive bioluminescence imaging system. Then, the quantification of GFP-expressing metastases in the lungs will be examined by fluorescence microscopy to validate the observed bioluminescence results. This sophisticated system combining luciferase and fluorescence-based detection tools evaluates cancer metastasis in vivo, which has great potential for use in breast cancer therapeutics and disease management.

We generated breast cancer cell lines (MDA-MB-231, MCF-7, and MDA-MB-468) expressing GFP and luciferase vectors. Specifically, this was achieved by a sequential infection. First, the breast cancer cell lines were infected with a lentivirus vector expressing fluorescent GFP. The GFP-positive cells (GFP+) were sorted 2 days post-infection (Figure 1A,B) and infected with the pLX304 Luciferase-V5 vector. Then, blasticidin was used to select for luciferase to generate ...

Watch this Scientific Journal Video about Monitoring Breast Cancer Growth and Metastatic Colony Formation in Mice using Bioluminescence at JoVE.com

Monitoring Breast Cancer Growth and Metastatic Colony Formation in Mice using Bioluminescence

Here, we describe a noninvasive monitoring method involving luciferase and green fluorescent protein expression in various breast cancer cell lines. This protocol provides a technique to monitor tumor formation and metastatic colonization in real time in mice.

Breast cancer is a frequent heterogeneous malignancy and the second leading cause of mortality in women, mainly due to distant organ metastasis. Several ...

Using this sophisticated, non-invasive tool, we can real-time monitor tumor growth kinetics and metastatic colonization in mice, which has a great potential in breast cancer therapeutic and disease management. Combining luciferase and fluorescence detection is a helpful strategy to advance the preclinical studies of breast cancer progression and disease management. This can be applied for anticancer drug studies.This protocol is not restricted to breast cancer and could be applied to other carcinomas, such as lung and pancreatic. Start by growing the MCF-7, MDA-MB-468, and MDA-MB-231 GFP and luciferase-positive cells separately in a 16-centimeter plate to 80%confluency. After harvesting the cells by trypsinization, seed an increasing number of cells in each well into a black 96-well plate.Then, fill all the wells with 100 microliters of DMEM, and incubate for 16 to 24 hours in 37 degrees Celsius, 5%CO2 incubator. Prepare luciferin solution in PBS at a 1.5-milligrams-per-milliliter concentration, and store the solution at minus 20 degrees Celsius. Post-incubation, wash the cells once with PBS gently before adding 100 microliters of luciferin solution into each well.Wait for two minutes, and then measure the luciferase activity in all the breast cancer cells using bioluminescence. Before injecting the mouse, transfer five million MCF-7 and MDA-MB-468 cells into 200 microliters of PBS and two million MDA-MB-231 GFP and luciferase-positive cells into 100 microliters PBS. Prepare the anesthetized mouse for injection by placing a cone over the head in a supine position.Next, wipe the abdominal area of the mouse above the mammary gland with ethanol using a cotton swab, and lift the fourth mammary gland with forceps before inserting a 27-gauge needle under the fat pad to slowly inject 100 microliters of the prepared cell suspension. After the injection, take the mouse out of the hood and transfer it to a new cage under observation until it returns to consciousness. Before the bioluminescence detection, restrain the conscious mouse by holding its neck with the left hand and tilting the hand to the left, resulting in the face of the mouse upward with the lower body in a supine position.Inject 100 microliters of 30-milligrams-per-milliliter luciferin intraperitoneally into the lower left abdominal quadrant of the mouse using a one-milliliter syringe. Keep the mouse for seven minutes without anesthesia, followed by three minutes within the anesthesia chamber before measuring the tumor kinetics. While the mouse is under incubation, open the software and initialize the imaging system by clicking the Initialize button.Keep the setup in auto exposure with exposure time in the auto at 60 seconds, binning medium at an aperture of f/stop one, excitation filter blocked, and emission filter open. When initialization ends, select Imaging Wizard and Bioluminescence and then click Next, Open Filter to select mouse in the image subject. In the field view, select stage C at 10 centimeters and subject height at 1.50 centimeters.Then, click Image Setup Stage to C.Ensure that the mouse is placed on the stage in the proper supine position before closing the door and clicking the Acquire button. Wait for an image to appear on the screen. Repeat the procedure for the other mouse.To detect lung metastasis, cover the strong signal of the primary tumor using a thick cardboard paper and expose only the ventral side of the lungs towards the camera. Capture the image using the same parameters as described earlier. The breast cancer cell lines were infected with a lentivirus vector expressing fluorescent GFP and subject to FAC sorting.The GFP-positive cells were sorted two days post-infection, plated and visualized by fluorescence microscope. The in vitro luciferase activity was confirmed with a cell number-dependent increase in the luciferase activity levels. In addition, a linear correlation was found between the luciferase activity and the cell number.After injecting the breast cancer cell lines, the mice were subjected to bioluminescence reading every two weeks to determine the tumor growth kinetics. The tumor growth kinetics was faster in the MDA-MB-231 cell lines than in the less aggressive cell lines, MCF-7 and MDA-MB-468. The luminescence activity was quantified for three cell lines.Six weeks post-injection, the harvested tumors were found to maintain the GFP expression. Also, positive bioluminescence readings were obtained from the lung of the whole mouse in real time. The lung was harvested to verify positive metastatic colonies, and the metastatic colonies were observed for GFP and bioluminescence.It is more important to be careful while performing the injection procedure. An improper injection may lead to deviation in the tumor growth rate or absence of tumor. We can examine the lung metastasis in vivo and ex vivo by detecting GFP expression.In addition, we can also study the histopathology of lungs by immunohistochemistry-based staining. This technique can be useful to explore the new questions, such as the effect of drug efficacy studies.

Research

JoVE Journal

Cancer Research

Utilizing Functional Genomics Screening to Identify Potentially Novel Drug Targets in Cancer Cell Spheroid Cultures

The identification of functional driver events in cancer is central to furthering our understanding of cancer biology and indispensable for the discovery ...

Labeling of Breast Cancer Patient-derived Xenografts with Traceable Reporters for Tumor Growth and Metastasis Studies

The use of preclinical models to study tumor biology and response to treatment is central to cancer research. Long-established human cell lines, and many ...

Practical Considerations in Studying Metastatic Lung Colonization in Osteosarcoma Using the Pulmonary Metastasis Assay

The pulmonary metastasis assay (PuMA) is an ex vivo lung explant and closed cell culture system that permits researchers to study the biology of lung ...

Preclinical Assessment of the Bioactivity of the Anticancer Coumarin OT48 by Spheroids, Colony Formation Assays, and Zebrafish Xenografts

In vitro and in vivo pre-clinical screening of novel therapeutic agents are an essential tool in cancer drug discovery. Although human cancer cell lines ...

Orthotopic Injection of Breast Cancer Cells into the Mice Mammary Fat Pad

A proper animal model is crucial for a better understanding of diseases. Animal models established by different methods (subcutaneous injections, ...

Performing Data Mining And Integrative Analysis Of Biomarker in Breast Cancer Using Multiple Publicly Accessible Databases

In recent years, emerging databases were designed to lower the barriers for approaching the intricate cancer genomic datasets, thereby, facilitating ...

In Vivo Inhibition of MicroRNA to Decrease Tumor Growth in Mice

MicroRNAs (miRNAs) are important regulators of gene expression through their ability to destabilize mRNA and inhibit translation of target mRNAs. An ...

Studying Triple Negative Breast Cancer Using Orthotopic Breast Cancer Model

Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype with limited therapeutic options. When compared to patients with less ...

Combined Use of Tail Vein Metastasis Assays and Real-Time In Vivo Imaging to Quantify Breast Cancer Metastatic Colonization and Burden in the Lungs

Metastasis is the main cause of cancer-related deaths and there are limited therapeutic options for patients with metastatic disease. The identification ...

Orthotopic Transplantation of Breast Tumors as Preclinical Models for Breast Cancer

Preclinical models that faithfully recapitulate tumor heterogeneity and therapeutic response are critical for translational breast cancer research. ...