Characterization of Biomarkers of Tumorigenic and Chemoresistant Cancer Stem Cells in Human Gastric Carcinoma

Abstract

Gastric carcinomas are heterogeneous, and the current therapy remains essentially based on surgery with conventional chemotherapy and radiotherapy. This study aimed to characterize biomarkers allowing the detection of cancer stem cells (CSC) in human gastric carcinoma of different histologic types. The primary tumors from 37 patients with intestinal- or diffuse-type noncardia gastric carcinoma were studied, and patient-derived tumor xenograft (PDX) models in immunodeficient mice were developed. The expressions of 10 putative cell surface markers of CSCs, as well as aldehyde dehydrogenase (ALDH) activity, were studied, and the tumorigenic properties of cells were evaluated by tumorsphere assays and xenografts by limiting dilution assays. We found that a subpopulation of gastric carcinoma cells expressing EPCAM, CD133, CD166, CD44, and a high ALDH activity presented the properties to generate new heterogeneous tumorspheres and tumors CD44 and CD166 were coexpressed, representing 6.1% to 37.5% of the cells; ALDH activity was detected in 1.6% to 15.4% of the cells; and the ALDH cells represented a core within the CD44/CD166 subpopulation that contained the highest frequency of tumorigenic CSCs The ALDH cells possessed drug efflux properties and were more resistant to standard chemotherapy than the ALDH cells, a process that was partially reversed by verapamil treatment. CD44 and ALDH are the most specific biomarkers to detect and isolate tumorigenic and chemoresistant gastric CSCs in noncardia gastric carcinomas independently of the histologic classification of the tumor. .

Gastric Cancer: A Stem Cell Disease?

“…Moreover, in contrast to squamous skin tumors [48] or intestinal adenomas [49], the in vivo contribution of GCSCs to tumorigenesis has not yet been fully elucidated. Nevertheless, regardless of their origin, dysplastic lesions and gastric adenocarcinomas are composed of CD44 + cells (Figure 2) [17,36] that have been recently described to possess cancer stem cell properties [50,51].…”

“…In vitro, under conditions in which cells are seeded at low-density in low-adherent plates, without serum and in the presence of some growth factors such as EGF, bFGF, and insulin, only GCSCs can survive, self-renew, and form tumorspheres [36,50,51,57]. Long self-renewal ability is evaluated after tumorsphere dissociation into single cells and several passages; indeed only GCSCs can generate tumorspheres after several passages [51,58]. In vivo, frequency of GCSCs in a given population is determined after subcutaneous xenograft in immunocompromised mice using diferent cell doses and an analysis of their ability to initiate a new heterogenous tumor after several weeks.…”

“…Immunocompromised mouse phenotypes, nonobese diabetic/severe combined immunodeiciency (NOD/SCID) and NOD/SCID/IL2Rg− (NSG), can inluence the CSC frequency as reported in the case of melanoma by Morrison's group [59]. In both methods, cells were seeded for an extreme limiting dilution assay, and a mathematical method was applied to calculate the CSC frequency in a given cell population [51,60].…”

“…We and others reported that CD44 is expressed following H. pylori infection in patients and in mouse models in the case of regenerative hyperplasia, intestinal metaplasia, dysplasia, and gastric carcinoma (Figure 2) [35, 36,50,51,65]. Histological and molecular analyses of tumor collections have shown that CD44 is positively and signiicantly associated with tumor recurrence and mortality in gastric cancer, and the expression of CD44 and CD44v has also been associated with metastasis formation [57,[65][66][67][68][69][70].…”

“…More recently, further relevant results from patient-derived xenograft models (PDXs) of gastric carcinoma have conirmed that CD44 is also a marker of GCSCs in primary gastric carcinoma. The FACS-sorted CD44 + cells, but not their CD44 − counterpart, displayed CSC properties such as growing as tumorspheres in vitro and lead to tumor formation in vivo that reconstitute the heterogeneity of the primary tumor of the patients and are more chemoresistant [51,71,72]. ESA, CD24, CD133, and CD166 are also expressed by CD44 + cells, but they do not allow a beter enrichment of GCSCs in combination with CD44 compared to CD44 alone [51,73].…”