Name: pre-clinical orthotopic murine model of human prostate cancer
Uploaded: 2016-08-29T14:00:00
Description: Watch this Scientific Journal Video about Pre-clinical Orthotopic Murine Model of Human Prostate Cancer at JoVE.com

We thank Dr. Roger Erickson for his support and assistance with the preparation of the manuscript. This work was supported by the National Cancer Institute at the National Institutes of Health through grant numbers RO1CA160079, RO1CA138642, UO1CA184966 and VA funded program project number 1P1 BX001604.

सभी जानवरों को शामिल प्रक्रियाओं के लिए प्रोटोकॉल की समीक्षा की और एक संस्थागत पशु की देखभाल और उपयोग समिति (IACUC) द्वारा अनुमोदित किया जाना चाहिए। देखभाल और प्रयोगशाला पशुओं के उपयोग के लिए आधिकारिक तौ...

<ol>
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J., 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=Targeting+platelet-derived+growth+factor+receptor+on+endothelial+cells+of+multidrug-resistant+prostate+cancer.">Targeting platelet-derived growth factor receptor on endothelial cells of multidrug-resistant prostate cancer.</a> J Natl Cancer Inst. 98 (11), 783-793 (2006).</li><li>Park, S. I., 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=Targeting+SRC+family+kinases+inhibits+growth+and+lymph+node+metastases+of+prostate+cancer+in+an+orthotopic+nude+mouse+model.">Targeting SRC family kinases inhibits growth and lymph node metastases of prostate cancer in an orthotopic nude mouse model.</a> Cancer Res. 68 (9), 3323-3333 (2008).</li><li>Zhang, J., 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=AFAP-110+is+overexpressed+in+prostate+cancer+and+contributes+to+tumorigenic+growth+by+regulating+focal+contacts.">AFAP-110 is overexpressed in prostate cancer and contributes to tumorigenic growth by regulating focal contacts.</a> J Clin Invest. 117 (10), 2962-2973 (2007).</li><li>Park, S. I., Kim, S. J., McCauley, L. K., Gallick, G. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pre-clinical+mouse+models+of+human+prostate+cancer+and+their+utility+in+drug+discovery.">Pre-clinical mouse models of human prostate cancer and their utility in drug discovery.</a> Curr Protoc Pharmacol. Chapter 14, Unit 14.15 (2010).</li><li>Johnson, L. C., 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=Longitudinal+live+animal+micro-CT+allows+for+quantitative+analysis+of+tumor-induced+bone+destruction.">Longitudinal live animal micro-CT allows for quantitative analysis of tumor-induced bone destruction.</a> Bone. 48 (1), 141-151 (2011).</li><li>Steinbauer, 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=GFP-transfected+tumor+cells+are+useful+in+examining+early+metastasis+in+vivo,+but+immune+reaction+precludes+long-term+tumor+development+studies+in+immunocompetent+mice.">GFP-transfected tumor cells are useful in examining early metastasis in vivo, but immune reaction precludes long-term tumor development studies in immunocompetent mice.</a> Clin Exp Metastasis. 20 (2), 135-141 (2003).</li><li>Yang, 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=A+fluorescent+orthotopic+bone+metastasis+model+of+human+prostate+cancer.">A fluorescent orthotopic bone metastasis model of human prostate cancer.</a> Cancer Res. 59 (4), 781-786 (1999).</li><li>Stephenson, R. 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=Metastatic+model+for+human+prostate+cancer+using+orthotopic+implantation+in+nude+mice.">Metastatic model for human prostate cancer using orthotopic implantation in nude mice.</a> J Natl Cancer Inst. 84 (12), 951-957 (1992).</li><li>Hoffman, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Orthotopic+metastatic+mouse+models+for+anticancer+drug+discovery+and+evaluation:+a+bridge+to+the+clinic.">Orthotopic metastatic mouse models for anticancer drug discovery and evaluation: a bridge to the clinic.</a> Invest New Drugs. 17 (4), 343-359 (1999).</li><li>Wang, X., An, Z., Geller, J., Hoffman, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-malignancy+orthotopic+nude+mouse+model+of+human+prostate+cancer+LNCaP.">High-malignancy orthotopic nude mouse model of human prostate cancer LNCaP.</a> Prostate. 39 (3), 182-186 (1999).</li><li>An, Z., Wang, X., Geller, J., Moossa, A. R., Hoffman, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Surgical+orthotopic+implantation+allows+high+lung+and+lymph+node+metastatic+expression+of+human+prostate+carcinoma+cell+line+PC-3+in+nude+mice.">Surgical orthotopic implantation allows high lung and lymph node metastatic expression of human prostate carcinoma cell line PC-3 in nude mice.</a> Prostate. 34 (3), 169-174 (1998).</li><li>Kim, S. J., 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=Reduced+c-Met+expression+by+an+adenovirus+expressing+a+c-Met+ribozyme+inhibits+tumorigenic+growth+and+lymph+node+metastases+of+PC3-LN4+prostate+tumor+cells+in+an+orthotopic+nude+mouse+model.">Reduced c-Met expression by an adenovirus expressing a c-Met ribozyme inhibits tumorigenic growth and lymph node metastases of PC3-LN4 prostate tumor cells in an orthotopic nude mouse model.</a> Clin Cancer Res. 9 (14), 5161-5170 (2003).</li></ol>

यह पांडुलिपि एक मानव ओर्थोटोपिक प्रोस्टेट कैंसर xenograft माउस मॉडल की स्थापना के लिए एक विस्तृत प्रक्रिया का वर्णन करता है। यह मॉडल immunocompromised चूहों के पृष्ठीय प्रोस्टेटिक खण्डों में मानव प्रोस्टेट कैंसर को?...

प्रोस्टेट कैंसर संयुक्त राज्य अमेरिका में पुरुषों के बीच कैंसर मौतों (9%) के दूसरे सबसे प्रचलित कारण, अगले फेफड़े और श्वसनी (28%) 1 के कैंसर के लिए है। हाल के आंकड़ों के अनुसार, यह अनुमान है कि 220, 800 नव निदान प्रोस्टेट कैंसर के मामलों और 27, 540 लोगों की मृत्यु 2015 1 में घटित होगा। पंचवर्षीय प्रारंभिक चरण में प्रोस्टेट कैंसर के रिश्तेदार जीवित रहने की दर&gt; 99% है, जबकि उन्नत metastatic रोग की है कि केवल 28% 1। उन्नत metastatic रोग के उपचार के लिए एक बड़ी चुनौती इस रोग की प्रवृत्ति अंतर्निहित, अन्य अंगों को metastasize करने के लिए विशेष रूप से हड्डी है, जो प्रोस्टेट कैंसर के लिए एक लगातार साइट है आणविक तंत्र की समझ की कमी है। इसलिए, वहाँ आदेश उन्नत metastatic रोग 2,3 के लिए प्रगति के खिलाफ प्रभावी उपचार परहेजों का विकास करने के लिए इन प्रोस्टेट ट्यूमर की आणविक मेकअप अध्ययन करने के लिए एक स्पष्ट की जरूरत है। प्रोस्टेट ट्यूमर प्रदर्शनी higप्रगति के लिए एक अच्छी तरह से परिभाषित मार्ग के बिना ज जैविक विविधता। मेटास्टेसिस अक्सर ट्यूमर invasiveness 4 का कोई पूर्व संकेत के साथ होते हैं। यह नैदानिक ​​विविधता प्रोस्टेट कैंसर की आणविक विविधता के लिए जिम्मेदार ठहराया है। इन घातक ट्यूमर के आणविक मेकअप को समझना इस रोग के लिए बेहतर नैदानिक ​​और चिकित्सीय रणनीतियों डिजाइन करने के लिए महत्वपूर्ण है। नतीजतन, प्रोस्टेट कैंसर अनुसंधान वर्तमान समझ और मेटास्टेसिस को रोकने पर ध्यान केंद्रित किया है। पूर्व नैदानिक ​​इन विवो माउस मॉडल विकल्प उन्नत मेटास्टेटिक रोग से प्रोस्टेट कैंसर की प्रगति के आणविक तंत्र को समझने के लिए की एक किस्म की पेशकश करते हैं। इसके अलावा, इन मॉडलों को इस बीमारी के खिलाफ नए चिकित्सकीय रणनीति के preclinical मूल्यांकन के लिए महत्वपूर्ण हैं। सबसे अधिक इस्तेमाल पशु मॉडल ट्रांसजेनिक माउस मॉडल, पूंछ-नस में इंजेक्शन, इंट्रा-हृदय आरोपण और मानव ओर्थोटोपिक माउस मॉडल शामिल हैं। ट्रांसजेनिक पढ़ाई के समय consumi हैंएनजी और मनुष्यों के साथ कि चूहों में प्रोस्टेट कैंसर के विकास के संबंध परिवर्तनशीलता 11 से पता चला है। सहज मेटास्टेटिक माउस मॉडल में, कोशिकाओं के संचलन में सीधे इंजेक्ट कर रहे हैं और हालांकि, वे तेजी से बदलाव का समय है, वे प्राथमिक ट्यूमर या metastatic झरना 5 में प्रारंभिक कदम का अध्ययन करने के लिए इस्तेमाल नहीं किया जा सकता। Orthotopic जेनोग्राफ्ट मॉडल हड्डी मेटास्टेटिक घावों, प्रोस्टेट कैंसर मेटास्टेसिस के आम साइट के विकास की सीमा है। बहरहाल, मानव ओर्थोटोपिक प्रोस्टेट कैंसर xenograft माउस मॉडल अच्छी तरह से विशेषता है और व्यापक रूप से प्राथमिक ट्यूमर के विकास, ट्यूमर और अंग microenvironment, मेटास्टेटिक रोग और चिकित्सकीय हस्तक्षेप 6 के लिए प्रयोगात्मक दवाओं के उपयोग के प्रारंभिक चरण के बीच परस्पर बात की आणविक घटनाओं का अध्ययन करने के लिए प्रयोग किया जाता है , 7,8-11।

<table border="0" cellpadding="0" cellspacing="0" width="732">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:229px;">PC3 prostate cancer cell line&#160;</td>
			<td style="width:188px;">ATCC</td>
			<td style="width:111px;">CRL-1435</td>
			<td style="width:204px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Minimum Essential Medium (MEM)&#160;</td>
			<td>GIBCO,Life Technology</td>
			<td>11095-080</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">PBS</td>
			<td>GIBCO,Life Technology</td>
			<td>10010-023</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">FBS</td>
			<td>GIBCO,Life Technology</td>
			<td>10437-028</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Zeocin</td>
			<td>Invitrogen,Life Technology</td>
			<td>R250-01</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Trypsin&#160;</td>
			<td>GIBCO,Life Technology</td>
			<td>25300-54</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">IVIS&#160;</td>
			<td>Xenogen-Caliper</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Insulin Syringes (300ul, 28.5g)</td>
			<td>Becton Dickinson</td>
			<td>309300</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Mice</td>
			<td>Charles River Laboratories, Inc</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Alcohol Swabs</td>
			<td>MEDEquip Depot</td>
			<td>326895 BD</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">PVP Iodine Prep Pad</td>
			<td>MEDEquip Depot</td>
			<td>C12400PDI</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Surgical CatGut Chromic Suture</td>
			<td>Demetech</td>
			<td>CC224017F0P</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Matrigel</td>
			<td>Corning</td>
			<td>354248</td>
			<td></td>
		</tr>
	</tbody>
</table>

pre-clinical orthotopic murine model of human prostate cancer

To study the multifaceted biology of prostate cancer, pre-clinical in vivo models offer a range of options to uncover critical biological information about this disease. The human orthotopic prostate cancer xenograft mouse model provides a useful alternative approach for understanding the specific interactions between genetically and molecularly altered tumor cells, their organ microenvironment, and for evaluation of efficacy of therapeutic regimens. This is a well characterized model designed to study the molecular events of primary tumor development and it recapitulates the early events in the metastatic cascade prior to embolism and entry of tumor cells into the circulation. Thus it allows elucidation of molecular mechanisms underlying the initial phase of metastatic disease. In addition, this model can annotate drug targets of clinical relevance and is a valuable tool to study prostate cancer progression. In this manuscript we describe a detailed procedure to establish a human orthotopic prostate cancer xenograft mouse model.

पीछे प्रोस्टेटिक पालि में PC3M ल्यूक-सी 6 कोशिकाओं के ओर्थोटोपिक आरोपण के बाद, चूहों साप्ताहिक प्रयोग के पाठ्यक्रम (- बी चित्रा 5 ए) से अधिक कोशिकाओं के उपनिवेशवाद और ट्यूमर के विक?...

Watch this Scientific Journal Video about Pre-clinical Orthotopic Murine Model of Human Prostate Cancer at JoVE.com

Pre-clinical Orthotopic Murine Model of Human Prostate Cancer

Prostate cancer is the second most common cause of cancer-related deaths in the United States. An orthotopic cancer model provides a useful approach to understand the biology of prostate cancer and to evaluate the efficacy of therapeutic regimens. This protocol describes detailed steps necessary to establish an orthotopic prostate cancer mouse model.

पूर्व नैदानिक ​​Orthotopic Murine मानव प्रोस्टेट कैंसर के मॉडल

To study the multifaceted biology of prostate cancer, pre-clinical in vivo models offer a range of options to uncover critical biological information ...

The overall goal of this procedure is to establish a mouse orthotopic prostate cancer xenograft model that provides a useful approach to studying the molecular events of primary tumor development in the early events of the metastatic cascade. This model can help answer key questions regarding the spastic interactions between genetically and molecularly altered tumor cells in the micro-environment of the growing tumor. It can also help in the pre-clinical evaluation of new therapeutic approaches against the human prostate cancer.The main advantage of this technique over the spontaneous metastatic models are that it promotes the study of the molecular events of primary tumor development and also the recapitulation of the initial phase of metastasis prior to entry and embolism of tumor cells into the circulation. Orthotopic model of prostate cancer is one of the best pre-clinical models to understand how prostate cancer cells metastasize to different organs. Most prostate cancer patients die of metastasis, and therefore this model is very important to investigate the molecular mechanisms of metastasis.Using this model we can design better strategies for the treatment of metastasis by using neural compounds, those are specific to metastatic pathways. Demonstrating this procedure will be Melissa Colden, a grad student and Dr.Varahram Shahryari, a research associate from our laboratory. Before beginning the procedure, wash the tumor cell culture of interest with PBS and incubate the cells with 2 milliliters of 05 percent trypsin for 3 to 5 minutes in the cell culture incubator.When the cells have detached, stop the reaction with 5 milliliters of complete medium and count the cells by trypan blue exclusion. Dilute the cells to a 2.5 times 10 to the 5 cells per 10 microliters of medium concentration, and mix the cell suspension with 10 microliters of basement membrane-like extracellular matrix extract. Hold the cells on ice, then place an anesthetized six to eight week old immunocompromised male experimental mouse in the supine position on a sterile heating pad and confirm a lack of response to toe pinch.Apply ointment to the animals eyes and remove the hair from the lower abdomen. Disinfect the surgical area with 10 percent povidone iodine scrub, followed by 70 percent ethanol about 1 to 2 centimeters above the penis sheath, and about 2 to 3 centimeters below the bottom of the rib cage. Use a pair of fine forceps to lift 2 millimeters of skin above the preputial gland.Using a scalpel, make a 1 centimeter mid-line incision through the skin, followed by a 1 centimeter incision through the muscle layer. Locate the yellow, light brown spherical bladder directly below the incision. Then use the fine forceps to grip the bladder and to lift the organ up and out of the body cavity toward the penis sheath to expose the seminal vesicles.With a wet cotton swab in each hand, carefully externalize the vesicles one at a time, and lay them top down on the outer surface of the abdomen with the bladder in the middle. Next, use the cotton swabs to gently tilt back the seminal vesicles at the point of insertion near the bladder neck towards the penis sheath so that the two dorsal prostate lobes are clearly visible. Now use a micro-pipette to re-suspend the cells in a single-cell suspension and label them with luciferin.Aspirate 25 microliters of the cells into a 0.5 cc syringe equipped with a 28.5 gauge needle and transfer the mouse under a dissecting microscope. Then, insert the syringe needle into a dorsal prostatic lobe and slowly inject 20 microliters of the cell suspension until a bulla formation is observed. When all of the cells have been administered, retract the needle, pressing lightly on the injection site with a cotton swab for a few seconds to prevent leakage.Then use the swabs to carefully return the vesicles to the body cavity one at a time, followed by the bladder. After placing the organs back into the body cavity, suture the muscle layer first in an interrupted pattern with absorbable 4-0 chromic cat gut sutures, followed by the skin closure with non-absorbable 4-0 nylon surgical sutures. The skin can also be pulled together and closed with surgical clamps to close the incision completely.Then, immediately image the animal, and return the mouse to a clean cage under a warming lamp with monitoring until it is fully recovered. Following the orthotopic implantation of luciferin-labeled prostate cancer cells into the posterior prostatic lobe, as just demonstrated, weekly imaging using live animal bioluminescence imaging reveals a successful colonization of the prostate lobes by the tumor cells. Indeed, an increase in the bioluminescence over time is indicative of the enlargement of the primary tumor over the course of the experiment, confirming the effectiveness of this technique in non-invasive, radiographic, fluorescent or luminescent imaging for tumor growth and metastatic lesion monitoring.While attempting this procedure, it is important to remember that intra-prostatic injection requires open abdominal surgery. So, it should be performed in a pathogen-free environment using proper aseptic surgical techniques. Working with human cancer cells can be extremely hazardous and precautions, such as wearing personal protective equipment, and working within biosafety level two containment should always be taken while performing this procedure.This model paved the way for the transplant of histologically intact tumor fragments to examine the effects of potential drug therapies on tumor growth or original lymph node metastasis. After watching this video, you should have a good understanding of how to implant cancer cells into the dorsal prostatic lobes for the establishment of an orthotopic human prostate cancer xenograft mouse model.

Research

JoVE Journal

Medicine

An Orthotopic Model of Murine Bladder Cancer

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In this straightforward procedure, bladder tumors are established in female C57 mice through the use of catheterization, local cauterization, and ...

Combination Radiotherapy in an Orthotopic Mouse Brain Tumor Model

एक Orthotopic माउस ब्रेन ट्यूमर मॉडल में संयोजन रेडियोथेरेपी

Glioblastoma multiforme (GBM) are the most common and aggressive adult primary brain tumors1. In recent years there has been substantial progress in the ...

Development of Obliterative Bronchiolitis in a Murine Model of Orthotopic Lung Transplantation

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Orthotopic lung transplantation in rats was first reported by Asimacopoulos and colleagues in 1971 1. Currently, this method is well accepted and ...

An Orthotopic Bladder Cancer Model for Gene Delivery Studies

जीन डिलीवरी के अध्ययन के लिए एक orthotopic मूत्राशय कैंसर के मॉडल

Bladder cancer is the second most common cancer of the urogenital tract and novel therapeutic approaches that can reduce recurrence and progression are ...

Formation of Human Prostate Epithelium Using Tissue Recombination of Rodent Urogenital Sinus Mesenchyme and Human Stem Cells

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Progress in prostate cancer research is severely  limited by the availability of human-derived and hormone-na&iuml;ve model systems,  which limit our ...

Bioluminescent Orthotopic Model of Pancreatic Cancer Progression

अग्नाशय के कैंसर प्रगति की bioluminescent Orthotopic के लिए मॉडल

Pancreatic cancer has an extremely poor five-year survival rate of 4-6%.  New therapeutic options are critically needed and depend on improved ...

An Orthotopic Murine Model of Human Prostate Cancer Metastasis

मानव प्रोस्टेट कैंसर मेटास्टेसिस के एक orthotopic murine मॉडल

Our laboratory has developed a novel orthotopic implantation model of  human prostate cancer (PCa). As PCa death is not due to the primary tumor, but  ...

Isolation of Cancer Stem Cells From Human Prostate Cancer Samples

मानव प्रोस्टेट कैंसर के नमूनों से कैंसर स्टेम कोशिकाओं के अलगाव

The cancer stem cell (CSC) model has been considerably revisited over the last two decades. During this time CSCs have been identified and directly ...

पूर्व नैदानिक ​​Orthotopic Murine मानव प्रोस्टेट कैंसर के मॉडल

पूर्व नैदानिक Orthotopic Murine मानव प्रोस्टेट कैंसर के मॉडल