CD44v3+/CD24- cells possess cancer stem cell-like properties in human oral squamous cell carcinoma
- Authors:
- Published online on: November 20, 2015 https://doi.org/10.3892/ijo.2015.3261
- Pages: 99-109
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Copyright: © Todoroki et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
Abstract
Introduction
Recently, a number of studies identified cell surface markers, such as CD44v3 and CD24, as CSC markers (1–7). Oral squamous cell carcinoma (OSCC) is the eighth most prevalent cancer worldwide and shows high morbidity and poor survival rates (8). Despite advances in therapeutic procedures and various combinations of chemotherapeutic agents that have improved quality of life, mortality from this disease remains high because of the development of distant metastasis and the emergence of local and regional recurrences. Such treatment failure may be due to a small population of cells [cancer stem cells (CSCs)] that are responsible for tumorigenesis and contribute to resistance to conventional therapy, such as chemotherapy and radiotherapy. Therefore, identification of CSCs or cancer stem cell-like cells (CSC-LCs) may lead to the development of effective treatment. Many reports have verified the existence of CSCs in various solid neoplasms (7,9–12). With regard to OSCC, Prince et al (13) identified a population of CD44 positive tumor initiating cells in OSCC. Chen et al (14) reported that OSCC harbored potential CSC characterized by ALDH1.
CD44v3 is an alternative splicing form variant of CD44, which is a multifunctional transmembrane glycoprotein expressed in many types of cancer. With regard to OSCC, several studies have reported that CD44v3 is associated with drug resistance and unfavorable clinical outcomes (15,16).
CD24 is a 27-amino-acid single-chain protein that is O- and N-glycosylated and is bound to the extracellular matrix (17) and the extracellular membrane by a glycosylphosphatidylinositol anchor (18). Although several studies have reported that CD24 is associated with invasion, metastasis and tumor differentiation (19,20), whether CD24 expression is upregulated or downregulated with tumor invasion remains unclear. CD24 has also been studied in combination with CD44. Several studies have reported that CD44+/CD24− cells showed CSC properties in breast and prostate cancer (4–6). On the other hand, several studies have reported that CD44+/CD24+ was the CSC phenotype of pancreatic and colorectal cancer (7,21). With regard to OSCC, a few reports have shown that CD44+/CD24− may be the CSC phenotype (22,23).
In the present study, we focused on CD44v3 and CD24 and examined whether these markers have CSC properties by using two human OSCC cell lines and 50 human OSCC tissues.
Materials and methods
Cell lines and media
Two OSCC cell lines, SAS and OSC20, both derived from primary lesions of a patient with OSCC, were used in the experiment. SAS was purchased from the Health Science Research Resources Bank. OSC20 was donated by the Research Center for Innovative Cancer Therapy, Molecular Targeting Therapeutics Division, Kurume University, School of Medicine. SAS was grown in Dulbecco's modified Eagle's medium (DMEM; Nissui Seiyaku Co., Ltd., Tokyo, Japan) and Ham's F12 medium supplemented with heat-inactivated (56°C, 30 min) 5% fetal bovine serum (FBS; Bioserum, Victoria, Australia), 100 U/ml, penicillin and 100 μg streptomycin (Gibco-BRL/Life Technologies Inc., Gaitherburg, MD, USA). OSC20 was grown in Eagle's minimum essential medium (EMEM; Gibco, BRL/Life Technologies Inc.) with 5% FBS. Cells were cultured in an atmosphere of 5% CO2 in air at 37°C.
Flow cytometric analysis and separation
SAS and OSC20 cells with 80% confluence were washed once with phosphate-buffered saline (PBS), detached with accutase (Innovative Cell Technologies, Inc., San Diego, CA, USA), suspended at 1×106 cells/ml in PBS supplemented with 2% FBS, incubated with 10 μg/ml human IgG (R&D Systems, Inc., Minneapolis, MN, USA) for 15 min at room temperature, and then incubated with allophycocyanin (APC)-conjugated mouse anti-human CD44v3 (cat. no. FAB5088A; R&D Systems) combined with phycoerythrin (PE)-conjugated mouse anti-human CD24 (cat. no. 555428; BD Biosciences, San Jose, CA, USA) at 4°C for 45 min. Samples were washed, centrifuged at 500 × g for 3 min, resuspended in 2 ml cold PBS supplemented with 2% FBS, then 1 μg/ml propidium iodide (PI; BD Biosciences) was added and the cells were filtered through a 40-μm cell strainer (BD Biosciences). Analysis and separation were carried out with a FACSAria II (BD Biosciences).
Cell growth assay
A total of 2,500 cells of each of the four cell fractions, i.e., CD44v3+/CD24−, CD44v3+/CD24+, CD44v3−/CD24− and CD44v3−/CD24+ cells isolated from two cell lines were plated in 96-well plates and cultured in a CO2 incubator. The cells were harvested at 24, 48, 72 or 96 h and the proliferation was examined in colorimetric assays using 3-(4,5-dimethylthiazol-2yl-yl-)-2, 5-dimethyl tetrazolium bromide (MTT) cell growth assay kits (Chemicon, Temecula, CA, USA) as described elsewhere (24).
Sphere forming assay
Four cell fractions isolated from two cell lines (5,000 cells/dish) were cultured in serum-free medium including 10 ng/ml epidermal growth factor (EGF; Sanko Junyaku Co., Ltd., Tokyo, Japan) and 20 ng/ml basic fibroblast growth factor (bFGF) (PeproTech, Rocky Hill, NJ, USA) using ultra-low attachment 6-well plates (Corning Inc., Corning, NY, USA) for 1 week. Sphere formation was assessed by counting the number of spheres (>3 cells) under a microscope (x200).
Drug treatment assay
The four isolated cell fractions sorted from two cell lines were plated at 2,500 cells/well in 96-well plates, and the effect of CDDP (1 or 5 μM) (Nihon Kayaku, Tokyo, Japan), 5-fluorouracil (5-FU) (10 or 100 μM) (Kyowa Hakko, Tokyo, Japan), and cetuximab (100 or 1,000 nM) (Merck Serono Co., Ltd., Tokyo, Japan) was examined. Drug resistance was determined after treatment for 96 h by MTT assay.
Tumorigenicity assay
Various numbers of cells (1×103, 1×104, or 1×105) were injected subcutaneously into 4-week-old female non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice (n=5 in each group). Tumorigenic capacity was judged 8 weeks after injection. The animal procedures were approved by the Ethics Review Committee for Animal Experimentation of Kurume University School of Medicine.
cDNA preparation and quantitative real-time RT-PCR for gene expression assay
After the four fractions were isolated, total RNA was extracted using an RNAqueous Micro kit (Life Technologies, Carisbad, CA, USA), and complementary DNA (cDNA) was synthesized using the Reverse Transcription System (Promega, Madison, WI, USA) according to the manufacturer's instructions. Quantitative real-time RT-PCR (RT-qPCR) was performed to examine the expression of CSC-LC property-related genes [e.g., ABC transporter genes (ABCB1 and ABCG2), anti-apoptosis genes (BCL2 and CFLAR), self-replication genes (Oct4 and Nanog) and hypoxia-related genes [hypoxia inducible factor 1α (HIF1α)] with ABI PRISM 7500 (Applied Biosystems, Foster City, CA, USA). Gene expression assays and primer and probe mixes were used for ABCB1, ABCG2, ALDH1A1, BCL2, CFLAR, Oct4, Nanog, HIF1α, and β-actin [assay IDs (Hs 00184500_m1, Hs01053790_m1, Hs00946916_m1, Hs00608023_m1, Hs00153439_m1, Hs03666771, Hs04260366, Hs00153153_m1, and Hs99999903_m1, respectively; Applied Biosystems)], and thermal conditions were as follows: initial incubation at 95°C for 10 min, then 40 cycles alternating in turn with 95°C for 10 sec, 60°C for 20 sec, and 72°C for 15 sec, and then maintained at 72°C for 10 min. Comparative gene expression analysis was performed using the 2(−ΔΔCq) method with normalization to the level of internal control gene, β-actin.
Tissue samples
OSCC tissue samples for immunohistochemistry were obtained from 50 patients who underwent surgical resection at Kurume University Hospital between 2007 and 2008. These specimens were fixed in 10% buffered formalin followed by paraffin embedment. None of the patients had previously received any treatments, including chemotherapy or radiotherapy. This cohort was composed of 33 men and 17 women aged from 48 to 87 years (median age, 70 years). The average observation time for overall survival was 52 months for patients still alive at the time of analysis, and ranged from 1 to 95 months.
Immunohistochemical staining
All tissues were immunohistochemically examined for expression of CD44v3 and CD24 using mouse anti-human CD44v3 monoclonal antibody (VFF-327v3, 1:50 dilution; Novocastra, New Castle upon Tyne, UK) and mouse anti-human CD24 monoclonal antibody (528807, 1:100 dilution; R&D Systems). BenchMark XT (Ventana Medical Systems, Inc., Tucson, AZ, USA) was used for immunostaining.
Evaluation of staining
All slides were evaluated by two of the authors (K.T. and J.A.). CD44v3 and CD24 expressions were evaluated according to staining intensity and staining area in the non-invasive portion and invasive portion within the tumor. The staining intensity was graded as 0, negative; 1, weakly positive; 2, moderately positive; or 3, strongly positive. The staining intensity in the normal epithelium was used as an internal control. The staining area was graded as 1, 0–20%; 2, 20–40%; 3, 40–60%; 4, 60–80%; and 5, 80–100%. The scores of staining intensity and area were calculated. The expression scores of CD44v3 and CD24 were compared between the non-invasive portion and invasive portion. The expression scores of CD44v3 and CD24 were categorized as either negative or positive. The median value of the expression score was used to separate the negative and positive groups. The relationship between isolated CD44v3 and isolated CD24 immunoexpression or CD44v3/CD24 immunophenotype and each clinicopathological feature of OSCC, such as T stage, nodal status, mode of invasion (25) and histological grade, was analyzed.
Statistical analysis
JMP software version 11.0 was used for all statistical analysis. Comparisons of cell growth assay, sphere forming assay, drug resistance assay, and quantitative real-time RT-PCR assay were normally distributed and assessed by the Shapiro-Wilk test, and then, a comparison of each in vitro assay except for the sphere forming assay was performed using one-way ANOVA with Dunnet post hoc comparisons. A comparison of the sphere forming assay was performed using one-way ANOVA with the Tukey post hoc comparisons. The expression score data of the invasive portion were compared with that of the non-invasive portion using a Mann-Whitney U test. Correlation of isolated CD44v3 and CD24 immunoexpression or the CD44v3/CD24 immunophenotypes with the clinicopathological parameters was assessed by standard Chi square tests or the Fisher's exact test. The overall survival rate was defined as the interval between the diagnosis and the date of death (uncensored data) or the date of the last available clinical information (censored data). Comparison and estimation of cumulative survival rates were performed using the Kaplan-Meier curves and the log rank test. All tests were two-sided and a value of P<0.05 was considered significant.
Results
Expression of CD44v3/CD24 in SAS and OSC20
In SAS, the proportion of CD44v3+/CD24−, CD44v3+/CD24+, CD44v3−/CD24− and CD44v3−/CD24+ cells was 10.7, 48.2, 7.8 and 33.3%, respectively. Whereas, the proportion of CD44v3+/CD24−, CD44v3+/CD24+, CD44v3−/CD24− and CD44v3−/CD24+ cells was 24.1, 52.0, 11.2 and 12.7%, respectively (Fig. 1).
Biological features of sorted cell fractions in SAS and OSC20 in vitro
CD44v3+/CD24− cells in SAS showed a significantly higher proliferative ability than that of CD44v3−/CD24− and CD44v3−/CD24+ cells after culturing for 96 h (P<0.05), and, CD44v3+/CD24− cells in OSC20 showed a significantly higher proliferative ability than that of CD44v3−/CD24− cells (P<0.05) (Fig. 2A).
The sphere forming ability of CD44v3+/CD24− cells in both SAS and OSC20 was significantly higher than the other fractions (P<0.05 or P<0.001). There was also a significant difference in sphere forming ability between the two fractions, for example, between CD44v3+/CD24+ and CD44v3−/CD24−, and between CD44v3+/CD24+ and CD44v3−/CD24+ cells (P<0.001) (Fig. 2B).
After 96-h treatment with CDDP, 5-FU, or cetuximab, the sensitivity to each drug was assessed with the MTT assay in SAS and OSC20. Cell growth was significantly suppressed in cells treated with CDDP, 5-FU and cetuximab, as compared with control cells in both SAS and OSC20 (Fig. 2C).
In SAS, CD44v3+/CD24− cells had a significantly higher resistance in CDDP (1 μM) or 5-FU (10 or 100 μM) treatment than the other fractions (P<0.05 or P<0.001), and had a significantly higher resistance in CDDP (5 μM) or cetuximab (100 or 1,000 nM) treatment than CD44v3+/CD24+ and CD44v3−/CD24+ cells (P<0.001) (Fig. 2C).
In OSC20, CD44v3+/CD24− cells had a significantly higher resistance in CDDP (1 or 5 μM) or 5-FU (100 μM) or cetuximab (100 nM) treatment than CD44v3+/CD24+ cells and/or CD44v3−/CD24+ cells (P<0.05, P<0.01 or P<0.001). There was no significant difference in sensitivity in 5-FU (10 μM) or cetuximab (1,000 nM) treatment among the cell fractions (Fig. 2C).
Analysis of CSC-LC property-related gene expression in sorted cell fractions in SAS and OSC20 by RT-qPCR
We performed RT-qPCR analysis to compare CSC-LC property related gene expression in CD44v3+/CD24− cells and the other cell fractions in SAS and OSC20 cells. In SAS, CD44v3+/CD24− cells showed significantly higher mRNA expression of transporter-related genes (ABCB1 and ABCG2), ALDH1A1, anti-apoptotic gene (BCL2), and self-replication genes (Oct-4, Nanog), than the other fractions (Fig. 3).
Tumorigenicity assays in vivo in sorted cell fractions in SAS
Injection of 1×103 cells from all of the cell fractions produced no tumors in NOD/SCID mice. In contrast, two mice that received 1×104 CD44v3+/CD24− cells or CD44v3+/CD24+ cells developed tumors 8 weeks after the inoculation. In addition, injection of 1×105 cells from all of the cell fractions produced tumors in NOD/SCID mice. The ratio of tumorigenicity among CD44v3+/CD24− cell fractions and the other fractions in SAS was not significantly different (Table I).
Immunohistochemical findings and their relationship to clinicopathological features
CD44v3 and CD24 were predominantly expressed on the cell membrane and in the cytoplasm, respectively. A representative immunostaining photomicrograph of each grade is shown in Fig. 4. CD44v3 and CD24 immunoscoring of the non-invasive portion and invasive portion is shown in Fig. 5. A significant difference was observed between the expression scores of CD24 in the non-invasive portion and those in the invasive portion (P<0.01) (Fig. 5).
There was no significant correlation between isolated CD44v3 and isolated CD24 immunoexpression or CD44v3/CD24 immunophenotypes and each clinicopathological parameter in the non-invasive portion (data not shown). The association between CD44v3 and CD24 expression in the invasive portion and clinicopathological parameters is summarized in Table II. CD44v3 expression was significantly correlated with lymph node metastasis (P=0.039), but not with age, gender, T stage, mode of invasion and tumor differentiation. CD24 expression was significantly correlated with gender (P=0.01), but not with the other clinicopathological parameter. The association between CD44v3/CD24 immunophenotypes and clinicopathological parameters in 50 OSCC patients is summarized in Table III. When the 50 cases were subdivided into four groups i.e., CD44v3+/CD24−, CD44v3+/CD24+, CD44v3−/CD24− and CD44v3−/CD24+ cases, the four immunophenotypes were not significantly correlated with each clinicopathological parameter.
 | Table IIAssociation of CD44v3 and CD24 immunoexpression with clinicopathological parameters in 50 OSCC patients. |
| Table IIIAssociation of CD44v3/CD24 immunophenotype with clinicopathological parameters in 50 OSCC patients. |
Overall survival (OS) rates and CD44v3/CD24 immunophenotypes in the invasive portion
Kaplan Meier analysis established the relationship between CD44v3 expression in the invasive portion and OS (Fig. 6A). CD44v3+ cases tended to show poor OS compared with CD44v3− cases (P=0.055). The relationship between CD24 expression in the invasive portion and OS is shown in Fig. 6B. There was no significant difference in OS between CD24+ cases and CD24− cases. The Kaplan Meier curves for OS in OSCC patients that were subdivided into four groups according to CD44v3/CD24 immunophenotypes are shown in Fig. 6C. CD44v3+/CD24− cases showed significantly worse OS than CD44v3+/CD24− cases (P=0.029).
Discussion
In the present study, CD44v3+/CD24− cell fractions in SAS and OSC20 were 10.7 and 24.1%, respectively. Although the CD44v3+/CD24− cell fraction in SAS or OSC20 did not show a higher proliferative ability than each of the other fractions (CD44v3−/CD24+, CD44v3−/CD24− and CD44v3−/CD24+ cells), the CD44v3+/CD24− cell fraction showed significantly higher sphere forming ability, a higher CDDP or 5-FU resistance, suggesting that the CD44v3+/CD24− cell fraction had CSC-LC properties in SAS or OSC20.
Previous studies have reported that CSC-LCs have drug transporter genes such as ABCB1 and ABCG2 (26,27). Our real-time PCR assay found that drug transporter genes such as ABCB1 and ABCG2 were significantly more highly expressed in the CD44v3+/CD24− cell fraction than the other cell fractions in SAS. However, ABCB1 and ABCG2 are not major transporters of CDDP or 5-FU, and therefore, high expression of ABCB1 and ABCG2 is unlikely to be the explanation for the increased resistance to CDDP or 5-FU in SAS. Other studies have reported that CSC-LCs have anti-apoptotic and drug-resistant properties due to the expression of anti-apoptosis genes such as BCL2 and CFLAR (28), which contribute to the emergence of CDDP or 5-FU resistance (29–32). Our real-time PCR assays found that anti-apoptotic genes such as BCL2 and CFLAR were more highly expressed in the CD44v3+/CD24− cell fraction than the other cell fractions in SAS. These findings suggest that CD44v3+/CD24− cells have anti-apoptotic effects; therefore, they probably showed higher CDDP or 5-FU resistance. Oct-4 and Nanog are associated with self-renewal capacity (14,33). Our real-time PCR assays also found that self-replication markers such as Oct4 or Nanog were significantly more highly expressed in the CD44v3+/CD24− cell fraction than the other cell fractions in SAS.
Indeed, CD44v3+/CD24− showed the highest sphere forming abilities compared with the other fractions in both cell lines. Sphere forming assay is a representative assay used to confirm self-renewal capacity in vitro. Collectively, these findings suggest that the CD44v3+/CD24− cell fraction had a higher self-renewal capacity.
Before the presence of CSC was widely accepted in solid tumors, immunohistochemical studies on CD44, including its variant isoform such as CD44v3 and CD24 were conducted in various tumors, including OSCCs. However, the relationship between these markers and various clinicopathological findings was inconsistent in several previous studies (34–40). The present study demonstrated that CD44v3 expression in the invasive portion was slightly downregulated compared with the non-invasive portion. These findings are partially consistent with several studies on CD44 isoforms in OSCCs (36,41–43). Molecular cross-talk between tumor and host at the invasion front is probably associated with these alterations of adhesion molecules, such as CD44 isoforms (36,44,45). Our present study demonstrated CD24 expression in the invasive portion was significantly downregulated compared with expression in the non-invasive portion. In some reports, the upregulation of CD24 expression was associated with an early event of carcinogenesis in various carcinomas (35,39,46). In the present study, CD24 expression in the invasive portion was found in only 22% of cases and was not associated with prognosis. However, out of the cases without CD24 expression, the cases with unfavorable prognosis were enhanced when combined with CD44v3+ expression. As we described before, immunohistochemical analyses on CD44 and/or CD24 were inconsistent. These inconsistent results may be due to extreme variations in tissue material, varying specificity of the numerous different antibodies used, and the diverse scoring and evaluation system applied (36).
CD44v3+ cells showed a higher tumorigenicity in vivo assay compared with CD44v3− cells. These effects were almost independent of CD24 status. Also, immunohistochemical findings demonstrated that CD44v3 was more prognostic as a single marker in OSCCs. Collectively, these findings suggest that CD44v3 may be a more reliable marker for CSC-LC properties than CD24 as a single marker. The interaction of CD44 isoforms and CD24 is probably associated with each other directly or indirectly. Further studies on the association between CD44 isoforms and CD24 should be conducted to clarify the exact mechanisms.
In conclusion, the results suggest that the CD44v3+/CD24− cell population shows CSC-LC properties in a human OSCC cell line. Additionally, we presented some evidence that isolated CD44v3 immunoexpression or CD44v3+/CD24− immunophenotypes could give prognostic information associated with unfavorable clinical outcomes.
Acknowledgements
We thank Ms. Akemi Fujiyoshi for her assistance in our experiments.
References
|
Collins AT, Berry PA, Hyde C, Stower MJ and Maitland NJ: Prospective identification of tumorigenic prostate cancer stem cells. Cancer Res. 65:10946–10951. 2005. View Article : Google Scholar : PubMed/NCBI | |
|
Dalerba P, Dylla SJ, Park IK, Liu R, Wang X, Cho RW, Hoey T, Gurney A, Huang EH, Simeone DM, et al: Phenotypic characterization of human colorectal cancer stem cells. Proc Natl Acad Sci USA. 104:10158–10163. 2007. View Article : Google Scholar : PubMed/NCBI | |
|
Kalish ED, Iida N, Moffat FL and Bourguignon LY: A new CD44V3-containing isoform is involved in tumor cell growth and migration during human breast carcinoma progression. Front Biosci. 4:A1–A8. 1999. View Article : Google Scholar : PubMed/NCBI | |
|
Al-Hajj M, Wicha MS, Benito-Hernandez A, Morrison SJ and Clarke MF: Prospective identification of tumorigenic breast cancer cells. Proc Natl Acad Sci USA. 100:3983–3988. 2003. View Article : Google Scholar : PubMed/NCBI | |
|
Ma F, Li H, Wang H, Shi X, Fan Y, Ding X, Lin C, Zhan Q, Qian H and Xu B: Enriched CD44+/CD24− population drives the aggressive phenotypes presented in triple-negative breast cancer (TNBC). Cancer Lett. 353:153–159. 2014. View Article : Google Scholar : PubMed/NCBI | |
|
Hurt EM, Kawasaki BT, Klarmann GJ, Thomas SB and Farrar WL: CD44+CD24− prostate cells are early cancer progenitor/stem cells that provide a model for patients with poor prognosis. Br J Cancer. 98:756–765. 2008. View Article : Google Scholar : PubMed/NCBI | |
|
Li C, Heidt DG, Dalerba P, Burant CF, Zhang L, Adsay V, Wicha M, Clarke MF and Simeone DM: Identification of pancreatic cancer stem cells. Cancer Res. 67:1030–1037. 2007. View Article : Google Scholar : PubMed/NCBI | |
|
Petersen PE: Oral cancer prevention and control - the approach of the World Health Organization. Oral Oncol. 45:454–460. 2009. View Article : Google Scholar | |
|
Ricci-Vitiani L, Lombardi DG, Pilozzi E, Biffoni M, Todaro M, Peschle C and De Maria R: Identification and expansion of human colon-cancer-initiating cells. Nature. 445:111–115. 2007. View Article : Google Scholar | |
|
Kim CF, Jackson EL, Woolfenden AE, Lawrence S, Babar I, Vogel S, Crowley D, Bronson RT and Jacks T: Identification of bronchioalveolar stem cells in normal lung and lung cancer. Cell. 121:823–835. 2005. View Article : Google Scholar : PubMed/NCBI | |
|
Wang J, Guo LP, Chen LZ, Zeng YX and Lu SH: Identification of cancer stem cell-like side population cells in human nasopharyngeal carcinoma cell line. Cancer Res. 67:3716–3724. 2007. View Article : Google Scholar : PubMed/NCBI | |
|
Takaishi S, Okumura T, Tu S, Wang SS, Shibata W, Vigneshwaran R, Gordon SA, Shimada Y and Wang TC: Identification of gastric cancer stem cells using the cell surface marker CD44. Stem Cells. 27:1006–1020. 2009. View Article : Google Scholar : PubMed/NCBI | |
|
Prince ME, Sivanandan R, Kaczorowski A, Wolf GT, Kaplan MJ, Dalerba P, Weissman IL, Clarke MF and Ailles LE: Identification of a subpopulation of cells with cancer stem cell properties in head and neck squamous cell carcinoma. Proc Natl Acad Sci USA. 104:973–978. 2007. View Article : Google Scholar : PubMed/NCBI | |
|
Chen C, Wei Y, Hummel M, Hoffmann TK, Gross M, Kaufmann AM and Albers AE: Evidence for epithelial-mesenchymal transition in cancer stem cells of head and neck squamous cell carcinoma. PLoS One. 6:e164662011. View Article : Google Scholar : PubMed/NCBI | |
|
Wang SJ, Wong G, de Heer AM, Xia W and Bourguignon LY: CD44 variant isoforms in head and neck squamous cell carcinoma progression. Laryngoscope. 119:1518–1530. 2009. View Article : Google Scholar : PubMed/NCBI | |
|
Bourguignon LY, Wong G, Earle C and Chen L: Hyaluronan-CD44v3 interaction with Oct4-Sox2-Nanog promotes miR-302 expression leading to self-renewal, clonal formation, and cisplatin resistance in cancer stem cells from head and neck squamous cell carcinoma. J Biol Chem. 287:32800–32824. 2012. View Article : Google Scholar : PubMed/NCBI | |
|
Pirruccello SJ and LeBien TW: The human B cell-associated antigen CD24 is a single chain sialoglycoprotein. J Immunol. 136:3779–3784. 1986.PubMed/NCBI | |
|
Fischer GF, Majdic O, Gadd S and Knapp W: Signal transduction in lymphocytic and myeloid cells via CD24, a new member of phosphoinositol-anchored membrane molecules. J Immunol. 144:638–641. 1990.PubMed/NCBI | |
|
Sano A, Kato H, Sakurai S, Sakai M, Tanaka N, Inose T, Saito K, Sohda M, Nakajima M, Nakajima T, et al: CD24 expression is a novel prognostic factor in esophageal squamous cell carcinoma. Ann Surg Oncol. 16:506–514. 2009. View Article : Google Scholar | |
|
Yang XR, Xu Y, Yu B, Zhou J, Li JC, Qiu SJ, Shi YH, Wang XY, Dai Z, Shi GM, et al: CD24 is a novel predictor for poor prognosis of hepatocellular carcinoma after surgery. Clin Cancer Res. 15:5518–5527. 2009. View Article : Google Scholar : PubMed/NCBI | |
|
Yeung TM, Gandhi SC, Wilding JL, Muschel R and Bodmer WF: Cancer stem cells from colorectal cancer-derived cell lines. Proc Natl Acad Sci USA. 107:3722–3727. 2010. View Article : Google Scholar : PubMed/NCBI | |
|
Chen YC, Chen YW, Hsu HS, Tseng LM, Huang PI, Lu KH, Chen DT, Tai LK, Yung MC, Chang SC, et al: Aldehyde dehydrogenase 1 is a putative marker for cancer stem cells in head and neck squamous cancer. Biochem Biophys Res Commun. 385:307–313. 2009. View Article : Google Scholar : PubMed/NCBI | |
|
Chiu CC, Lee LY, Li YC, Chen YJ, Lu YC, Li YL, Wang HM, Chang JT and Cheng AJ: Grp78 as a therapeutic target for refractory head-neck cancer with CD24−CD44+ stemness phenotype. Cancer Gene Ther. 20:606–615. 2013. View Article : Google Scholar : PubMed/NCBI | |
|
Hisaka T, Yano H, Ogasawara S, Momosaki S, Nishida N, Takemoto Y, Kojiro S, Katafuchi Y and Kojiro M: Interferon-alphaCon1 suppresses proliferation of liver cancer cell lines in vitro and in vivo. J Hepatol. 41:782–789. 2004. View Article : Google Scholar : PubMed/NCBI | |
|
Yamamoto E, Kohama G, Sunakawa H, Iwai M and Hiratsuka H: Mode of invasion, bleomycin sensitivity, and clinical course in squamous cell carcinoma of the oral cavity. Cancer. 51:2175–2180. 1983. View Article : Google Scholar : PubMed/NCBI | |
|
Rizzo S, Hersey JM, Mellor P, Dai W, Santos-Silva A, Liber D, Luk L, Titley I, Carden CP, Box G, et al: Ovarian cancer stem cell-like side populations are enriched following chemotherapy and overexpress EZH2. Mol Cancer Ther. 10:325–335. 2011. View Article : Google Scholar : PubMed/NCBI | |
|
Hirschmann-Jax C, Foster AE, Wulf GG, Nuchtern JG, Jax TW, Gobel U, Goodell MA and Brenner MK: A distinct ‘side population’ of cells with high drug efflux capacity in human tumor cells. Proc Natl Acad Sci USA. 101:14228–14233. 2004. View Article : Google Scholar | |
|
Yajima T, Ochiai H, Uchiyama T, Takano N, Shibahara T and Azuma T: Resistance to cytotoxic chemotherapy-induced apoptosis in side population cells of human oral squamous cell carcinoma cell line Ho-1-N-1. Int J Oncol. 35:273–280. 2009.PubMed/NCBI | |
|
Yang Z, Liu Y, Liao J, Gong C, Sun C, Zhou X, Wei X, Zhang T, Gao Q, Ma D, et al: Quercetin induces endoplasmic reticulum stress to enhance cDDP cytotoxicity in ovarian cancer: Involvement of STAT3 signaling. FEBS J. 282:1111–1125. 2015. View Article : Google Scholar : PubMed/NCBI | |
|
Karaayvaz M, Zhai H and Ju J: miR-129 promotes apoptosis and enhances chemosensitivity to 5-fluorouracil in colorectal cancer. Cell Death Dis. 4:e6592013. View Article : Google Scholar : PubMed/NCBI | |
|
Longley DB, Wilson TR, McEwan M, Allen WL, McDermott U, Galligan L and Johnston PG: c-FLIP inhibits chemotherapy-induced colorectal cancer cell death. Oncogene. 25:838–848. 2006. View Article : Google Scholar | |
|
Micheau O, Solary E, Hammann A and Dimanche-Boitrel MT: Fas ligand-independent, FADD-mediated activation of the Fas death pathway by anticancer drugs. J Biol Chem. 274:7987–7992. 1999. View Article : Google Scholar : PubMed/NCBI | |
|
Murakami S, Ninomiya W, Sakamoto E, Shibata T, Akiyama H and Tashiro F: SRY and OCT4 are required for the acquisition of cancer stem cell-like properties and are potential differentiation therapy targets. Stem Cells. 33:2652–2663. 2015. View Article : Google Scholar : PubMed/NCBI | |
|
Oliveira LR, Oliveira-Costa JP, Araujo IM, Soave DF, Zanetti JS, Soares FA, Zucoloto S and Ribeiro-Silva A: Cancer stem cell immunophenotypes in oral squamous cell carcinoma. J Oral Pathol Med. 40:135–142. 2011. View Article : Google Scholar | |
|
Abdulmajeed AA, Dalley AJ and Farah CS: Putative cancer stem cell marker expression in oral epithelial dysplasia and squamous cell carcinoma. J Oral Pathol Med. 42:755–760. 2013. View Article : Google Scholar : PubMed/NCBI | |
|
Bánkfalvi A, Krassort M, Buchwalow IB, Végh A, Felszeghy E and Piffkó J: Gains and losses of adhesion molecules (CD44, E-cadherin, and beta-catenin) during oral carcinogenesis and tumour progression. J Pathol. 198:343–351. 2002. View Article : Google Scholar : PubMed/NCBI | |
|
Choi D, Lee HW, Hur KY, Kim JJ, Park GS, Jang SH, Song YS, Jang KS and Paik SS: Cancer stem cell markers CD133 and CD24 correlate with invasiveness and differentiation in colorectal adenocarcinoma. World J Gastroenterol. 15:2258–2264. 2009. View Article : Google Scholar : PubMed/NCBI | |
|
Kristiansen G, Denkert C, Schlüns K, Dahl E, Pilarsky C and Hauptmann S: CD24 is expressed in ovarian cancer and is a new independent prognostic marker of patient survival. Am J Pathol. 161:1215–1221. 2002. View Article : Google Scholar : PubMed/NCBI | |
|
Bircan S, Kapucuoglu N, Baspinar S, Inan G and Candir O: CD24 expression in ductal carcinoma in situ and invasive ductal carcinoma of breast: An immunohistochemistry-based pilot study. Pathol Res Pract. 202:569–576. 2006. View Article : Google Scholar : PubMed/NCBI | |
|
Darwish NS, Kim MA, Chang MS, Lee HS, Lee BL, Kim YI and Kim WH: Prognostic significance of CD24 expression in gastric carcinoma. Cancer Res Treat. 36:298–302. 2004. View Article : Google Scholar : PubMed/NCBI | |
|
Herold-Mende C, Seiter S, Born AI, Patzelt E, Schupp M, Zöller J, Bosch FX and Zöller M: Expression of CD44 splice variants in squamous epithelia and squamous cell carcinomas of the head and neck. J Pathol. 179:66–73. 1996. View Article : Google Scholar : PubMed/NCBI | |
|
Fonseca I, Pereira T, Rosa-Santos J and Soares J: Expression of CD44 isoforms in squamous cell carcinoma of the border of the tongue: A correlation with histological grade, pattern of stromal invasion, and cell differentiation. J Surg Oncol. 76:115–120. 2001. View Article : Google Scholar : PubMed/NCBI | |
|
Masuda M, Kuratomi Y, Shiratsuchi H, Nakashima T, Naonobu K and Komiyama S: Decreased CD44H expression in early-stage tongue carcinoma associates with late nodal metastases following interstitial brachytherapy. Head Neck. 22:662–665. 2000. View Article : Google Scholar : PubMed/NCBI | |
|
Williams HK, Sanders DS, Jankowski JA, Landini G and Brown AM: Expression of cadherins and catenins in oral epithelial dysplasia and squamous cell carcinoma. J Oral Pathol Med. 27:308–317. 1998. View Article : Google Scholar : PubMed/NCBI | |
|
Bànkfalvi A and Piffkò J: Prognostic and predictive factors in oral cancer: The role of the invasive tumour front. J Oral Pathol Med. 29:291–298. 2000. View Article : Google Scholar : PubMed/NCBI | |
|
Sagiv E, Memeo L, Karin A, Kazanov D, Jacob-Hirsch J, Mansukhani M, Rechavi G, Hibshoosh H and Arber N: CD24 is a new oncogene, early at the multistep process of colorectal cancer carcinogenesis. Gastroenterology. 131:630–639. 2006. View Article : Google Scholar : PubMed/NCBI |
