Rapid Isolation of Gastric Adenocarcinoma Cancer Stem Cells as a Target for Autologous Dendritic Cell-Based Immunotherapy

Document Type : Research Paper


1 Cellular and Molecular Research Center, Birjand University of Medical Sciences, Birjand, Iran

2 Immunology Department, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran.

3 Immunology Department, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran

4 Department of Laboratory Sciences, School of Paramedical Sciences, Mashhad University of Medical Sciences, Mashhad, Iran

5 Medical Genetics Research Center, Mashhad University of Medical Sciences, Mashhad, Iran


Background: Gastric cancer (GC) is a malignancy cause associated with a high death rate in the world. Cancer stem cells 
(CSCs) are a rare immortal subpopulation of cells within tumors with characteristics of the ability to self-renew, initiate 
tumor, and differentiate into defined progenies as well as and high resistance to conventional therapies. 
Objectives: Despite the use of surgery and chemotherapy for GC therapy, there are no efficient therapeutic protocols for 
it to date. Therefore, rapid isolation of CSCs in order to therapeutic targets, especially immunotherapy is very important.
Materials and Methods: Cancerous cell suspension isolated from patients with GC was cultured in the serum-free 
medium containing EGF, bFGF, LIF, and heparin under non-adherent culture conditions to generate spheres. Expression 
of mRNA level stemness transcription factors (OCT4, SOX2, SALL4, and Cripto-1), CD44 variable isoforms (CD44s, 
CD44v3, CD44v6, CD44V8-10) of spheroid-forming single cells compared with gastric normal tissue cells using real 
time PCR and molecules of CD44, CD54, and EpCAM as gastric CSC markers, and stemness factor Oct4 using flow 
cytometry, as well as tumorgenicity using subcutaneous injection of sphere-forming cells to nude mice were investigated.
Results: Few cancerous cells isolated from patients with GC were able to generate three-dimensional spheroid colonies in 
the serum-free medium containing EGF, bFGF, LIF, and heparin under non-adherent culture conditions, and form xenograft 
tumors in immunodeficient nude mice after subcutaneous injection. Spheroid-forming single cells upregulated stemness 
transcription factors OCT4, SOX2, SALL4, and Cripto-1 that are associated with pluripotency and self-renewal and CD44 
isoforms (CD44s, CD44v3, CD44v6, CD44V8-10) compared with gastric normal tissue cells. Finally, molecules of CD44, 
CD54, and EpCAM as gastric CSC markers and stemness factor Oct4 were expressed in sphere-forming cells.
Conclusion: We suggested that the sphere formation and tumorigenicity assays are two procedures, leading to the rapid 
isolation of cancer cells with certain stem-like properties in order to target CSCs using autologous dendritic cell therapy, 
especially in patients with advanced disease.


Main Subjects

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