Name: establishment of gastric cancer patient-derived xenograft models and primary cell lines
Uploaded: 2019-07-19T14:00:00
Description: Watch this Scientific Journal Video about Establishment of Gastric Cancer Patient-derived Xenograft Models and Primary Cell Lines Video at JoVE.com

This work was supported by the National Natural Science Foundation of China (81572392); the National Key Research and Development Program of China (2016YFC1201704); Tip-top Scientific and Technical Innovative Youth Talents of Guangdong Special Support Program (2016TQ03R614).
We specifically thank Guangzhou Sagene Biotech Co., Ltd. for aid in the preparation of the figures.

This human study was approved by the Institutional Ethics Review Board of Sun Yat-sen University Cancer Center (SYSUCC, Guangzhou, China). The animal study was approved by the Institutional Animal Care and Use Committee of Sun Yat-sen University.&#160;Note: all experiments were performed in compliance with the relevant laws and institutional guidelines, including the Guideline for occupational exposure protection against blood-borne pathogens.
1. Sample preparation
<ol>
	<li>Obtain gastric c...

<ol>
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Gastric cancer is an aggressive disease with limited therapeutic options; thus, models of gastric cancer have become a critical resource to enable functional research studies with direct translation to the clinic4,8,17. Here, we have described the methods and protocol of establishing gastric cancer PDX models and primary cell lines. Importantly, both morphological and biological characteristics of gastric cancer specimens were m...

Gastric cancer is the fifth-most common cancer worldwide and the third leading cause of cancer death. In 2018, over 1,000,000 new cases of gastric cancer were diagnosed globally, and an estimated 783,000 people were killed by this disease1. The incidence and mortality of gastric cancer remain very high in northeastern Asian countries2,3. Despite significant progress in the field of cancer therapeutics, the prognosis of patients with advanced gastric cancer remains poor, with a five-year survival rate of approximately 25%4,5,6,7,. Thus, there is an urgent need for the development of new therapeutic strategies for gastric cancer
The treatment of gastric cancer is challenging because of its high heterogeneity8,9.&#160;Thus, the question of how to address the challenges of tumor heterogeneity to realize precision medicine is central to cancer research. In vitro and in vivo models play crucial roles in elucidating the heterogeneous mechanisms and biology of gastric cancer. However, although there are numerous gastric cancer cell lines and many conventional transgenic mouse models for preclinical research, the disadvantages of these models limit their applications10. Because the characteristics of these models have changed in culture, they no longer model tumor heterogeneity, and their responses have not been able to predict responses in humans11. These issues severely limit the possibility of identifying subgroups of cancer patients that will respond to targeted drugs. The short-term culture of primary tumors provides a relatively rapid and personalized way to investigate anticancer pharmacological properties, which will likely be the hallmark of personalized cancer treatment.
Patient-derived xenografts (PDXs) are preferred as an alternative preclinical model for drug response profiling12. In addition, PDX models offer a powerful tool for studying the initiation and progression of cancer13,14. PDX models preserve the histologic appearance of cancer cells, retain intratumoral heterogeneity, and better reflect the relevant human components of the tumor microenvironment15,16. However, the limitation of the widely used PDX models is the low success rate for establishing and serially propagating human solid tumors. In this study, decently successful methods for establishing PDX models and primary cell lines are described.

<table>
	<tbody>
		<tr>
			<td>40 &#956;m Cell Strainer</td>
			<td>Biologix, Shandong, China</td>
			<td>15-1040</td>
			<td></td>
		</tr>
		<tr>
			<td>Biological Microscope</td>
			<td>OLYMPUS, Tokyo, Japan</td>
			<td>OLYMPUS CKX41</td>
			<td></td>
		</tr>
		<tr>
			<td>Centrifuge</td>
			<td>Eppendorf, Mittelsachsen, Germany.</td>
			<td>5427R</td>
			<td></td>
		</tr>
		<tr>
			<td>CO2 Incubator</td>
			<td>Thermo Fisher Scientific, Carlsbad, California, USA</td>
			<td>HERACELL 150i</td>
			<td></td>
		</tr>
		<tr>
			<td>DPBS</td>
			<td>Basalmedia Technology, Shanghai, China</td>
			<td>L40601</td>
			<td></td>
		</tr>
		<tr>
			<td>Electro-Thermostatic Water Cabinet</td>
			<td>Yiheng, Shanghai, China</td>
			<td>DK-8AXX</td>
			<td></td>
		</tr>
		<tr>
			<td>Fetal bovine serum</td>
			<td>Wisent Biotechnology, Vancouver, Canada</td>
			<td>86150040</td>
			<td></td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Baxter, China</td>
			<td>CN2L9100</td>
			<td></td>
		</tr>
		<tr>
			<td>Live Tissue Kit Cryo Kit</td>
			<td>Celliver Biotechnology, Shanghai, China</td>
			<td>LT2601</td>
			<td></td>
		</tr>
		<tr>
			<td>Live Tissue Thaw Kit</td>
			<td>Celliver Biotechnology, Shanghai, China</td>
			<td>LT2602</td>
			<td></td>
		</tr>
		<tr>
			<td>NSG</td>
			<td>Biocytogen, Beijing, China</td>
			<td>B-CM-002-4-5W</td>
			<td></td>
		</tr>
		<tr>
			<td>Penicilin&#38;streptomycin</td>
			<td>Thermo Fisher Scientific, Carlsbad, California, USA</td>
			<td>15140122</td>
			<td></td>
		</tr>
		<tr>
			<td>Red blood cell lysis buffer</td>
			<td>Solarbio, Beijing, China</td>
			<td>R1010</td>
			<td></td>
		</tr>
		<tr>
			<td>RPMI-1640 medium</td>
			<td>Thermo Fisher Scientific, Carlsbad, California, USA</td>
			<td>8118367</td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical Suture Needles with Thread</td>
			<td>LingQiao, Ningbo, China</td>
			<td>3/8 arc 4&#215;10</td>
			<td></td>
		</tr>
		<tr>
			<td>Tissue-processed molds and auxiliary blades</td>
			<td>Celliver Biotechnology, Shanghai, China</td>
			<td>LT2603</td>
			<td></td>
		</tr>
		<tr>
			<td>Trypsin-EDTA</td>
			<td>Thermo Fisher Scientific, Carlsbad, California, USA</td>
			<td>2003779</td>
			<td></td>
		</tr>
		<tr>
			<td>Type 1 collagenase</td>
			<td>Thermo Fisher Scientific, Carlsbad, California, USA</td>
			<td>17100017</td>
			<td></td>
		</tr>
	</tbody>
</table>

establishment of gastric cancer patient-derived xenograft models and primary cell lines

The use of preclinical models to advance our understanding of tumor biology and investigate the efficacy of therapeutic agents is key to cancer research. Although there are many established gastric cancer cell lines and many conventional transgenic mouse models for preclinical research, the disadvantages of these in vitro and in vivo models limit their applications. Because the characteristics of these models have changed in culture, they no longer model tumor heterogeneity, and their responses have not been able to predict responses in humans. Thus, alternative models that better represent tumor heterogeneity are being developed. Patient-derived xenograft (PDX) models preserve the histologic appearance of cancer cells, retain intratumoral heterogeneity, and better reflect the relevant human components of the tumor microenvironment. However, it usually takes 4-8 months to develop a PDX model, which is longer than the expected survival of many gastric patients. For this reason, establishing primary cancer cell lines may be an effective complementary method for drug response studies. The current protocol describes methods to establish PDX models and primary cancer cell lines from surgical gastric cancer samples. These methods provide a useful tool for drug development and cancer biology research.

Here, tumor tissues from an operation were preserved in stock solution until the next step. Within 4 hours, tumor tissues were cut into small pieces and implanted into the dorsal flanks of NSG mice that had been anesthetized using isoflurane-soaked cotton. Tumors larger than 1 cm3 could be resected for implantation into new mice (Figure 1) or sliced carefully and preserved in liquid nitrogen following the protocol. In this study, the first-generation tumors grew more slowly than t...

Watch this Scientific Journal Video about Establishment of Gastric Cancer Patient-derived Xenograft Models and Primary Cell Lines Video at JoVE.com

Establishment of Gastric Cancer Patient-derived Xenograft Models and Primary Cell Lines Video (Video) | JoVE

The current protocol describes methods to establish patient-derived xenograft (PDX) models and primary cancer cell lines from surgical gastric cancer samples. The methods provide a useful tool for drug development and cancer biology research.

Establishment of Gastric Cancer Patient-derived Xenograft Models and Primary Cell Lines

The use of preclinical models to advance our understanding of tumor biology and investigate the efficacy of therapeutic agents is key to cancer research. ...

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