CD133 expression is an independent prognostic marker for low survival in colorectal cancer

Colon cancer cells have previously been demonstrated to contain a subpopulation of CD133+ tumour cells that have the ability to initiate tumour growth and are thus referred to as colon cancer-initiating cells or colon cancer stem cells (CSCs). As CD133 is currently one of the best markers to characterise colon CSCs, we analysed CD133+ tumour cells in colorectal cancer specimens using immunohistochemistry. We show that CD133 detection is specific and that the CD133 antigen is localised on the glandular-luminal surface of colon cancer cells, whereas undifferentiated tumour cells at the front of invasion are CD133-. In addition, CD133+ cells are characterised in situ by lack of CK20 expression, whereas they are positive for EpCAM. Moreover, we show that CD133 expression in colorectal cancer is an independent prognostic marker that correlates with low survival in a stratified patient collective. Our results indicate that in colorectal cancer, the CD133+ tumour cells can be detected by immunohistochemistry, which facilitates their further characterisation in situ.     

Clinical outcomes of mild hyperthermia for locally advanced rectal cancer treated with preoperative radiochemotherapy

Purpose: The aim of this report was to determine the impact of hyperthermia (HT) on preoperative radiochemotherapy for locally advanced rectal cancer.

Materials and methods: Between 1996 and 2007, 235 patients with locally advanced rectal cancer were treated with concurrent preoperative radiochemotherapy with or without HT. The total dose of radiotherapy was 39.6?Gy for 109 patients (group A) and 45?Gy for 126 patients (group B). Two or three cycles of chemotherapy were administered. Hyperthermia was given immediately after radiotherapy.

Results: In the HT subgroup of group A, more patients achieved down-staging of T stage when compared to the non-HT subgroup (57.9% versus 38%, p?=?0.047). For the cN+ subgroup of all patients, the number of patients with ypN+ were significantly less in the HT subgroup (25% versus 50%, p?=?0.022). In group A, HT appeared to reduce distant metastasis, increase disease-free survival, and improve overall survival.

Conclusions: HT seemed to increase the response of both primary tumour and lymph nodes to preoperative radiochemotherapy in patients with locally advanced rectal cancer. The relationship between increased response by HT and survival should be confirmed by a large prospective randomised trial.     

Long-term local control and survival after preoperative radiochemotherapy in combination with deep regional hyperthermia in locally advanced rectal cancer

Purpose: The aim of this study was to evaluate the impact of deep regional hyperthermia on long-term local control and survival in locally advanced non-metastatic rectal cancer. Methods: In total 103 patients with locally advanced non-metastatic rectal cancer were treated preoperatively with either neoadjuvant radiochemotherapy alone (n?=?43) or the same treatment with additional deep regional hyperthermia (n?=?60). The two groups were compared with respect to local control, overall survival (OS), disease-free survival (DFS), and distant metastases-free survival (DMFS). Results: Patients receiving additional hyperthermia had excellent long-term local control with a 5-year Kaplan-Meier estimate of 98% compared with 87% in the radiochemotherapy only group (p?=?0.09). Five-year rates for OS (88% versus 76%, p?=?0.08), DFS (77% versus 73%, p?=?n.s.) and DMFS (75% versus 77%, p?=?n.s.) were not statistically different between the two groups. Conclusion: Radiochemotherapy combined with hyperthermia results in excellent long-term local control.     

The correlation between the levels of tissue inhibitor of metalloproteinases 1 in plasma and tumour response and survival after preoperative radiochemotherapy in patients with rectal cancer

Background

The aim of this study was to analyse whether the level of tissue inhibitor of metalloproteinases (TIMP) 1 is associated with the tumour response and survival to preoperative radiochemotherapy in rectal cancer patients.

Patients and methods.

Ninety-two patients with histologically confirmed non-metastatic rectal cancer of clinical stage I– III were treated with preoperative radiochemotherapy, surgery and postoperative chemotherapy. Plasma TIMP-1 concentrations were measured prior to the start of the treatment with an enzyme-linked immunosorbent assay (ELISA).

Results

Median follow-up time was 68 months (range: 3–93 months) while in survivors it was 80 months (range: 68–93 months). The 5-year locoregional control (LRC), disease-free survival (DFS), disease-specific survival (DSS) and overall survival (OS) rates for all patients were 80.2%, 56.4%, 63.7% and 52.2%, respectively. The median TIMP-1 level was 185 ng/mL (range: 22–523 ng/mL) and the mean level (±standard deviation) was 192 (±87) ng/mL. Serum TIMP-1 levels were found to be significantly increased in patients with preoperative CRP>12 mg/L and in those who died from rectal cancer or had cT4 tumours. No correlation was established for age, gender, carcinoembriogenic antigene (CEA) level, platelets count, histopathological grade, response to preoperative therapy, resectability and disease reappearance. On univariate analysis, various parameters favourably influenced one or more survival endpoints: TIMP-1 <170 ng/mL, CRP <12 mg/L, platelets count <290 10E9/L, CEA <3.4mg/L, age <69 years, male gender, early stage disease (cN0 and/or cT2–3), radical surgery (R0) and response to preoperative radiochemotherapy. In multivariate model, LRC was favourably influenced by N-downstage, DFS by lower CRP and N-downstage, DSS by lower CRP and N-downstage and OS by lower TIMP-1 level, lower CRP and N-downstage.

Conclusions

Although we did not find any association between pretreatment serum TIMP-1 levels and primary tumour response to preoperative radiochemotherapy in our cohort of patients with rectal cancer, TIMP-1 levels were recognized as an independent prognostic factor for OS in these patients.     

CD133-positive tumor cell content is a predictor of early recurrence in colorectal cancer

Background

The aims of this study were to demonstrate the tumorigenicity of CD133+ colon cancer cells in vitro, analyze the correlations between spheroid formation and clinicopathologic variables, and screen for overexpressed genes in CD133+ colon cancer stem cells. Moreover, the aim of this study was to establish a living tumor tissue bank using surgically resected specimens.

Methods

Using LoVo cell line, we isolated CD133+ cells and performed clonogenic assay and animal experiment to test tumorigenicity of CD133+ cells. Twenty-nine surgical samples were freshly collected from 27 patients who received curative or palliative surgery, and the samples were mechanically and enzymatically dissociated into single cells.

Results

We confirmed the enhanced tumorigenicity of CD133+ cells isolated from LoVo cell line both in vitro and in vivo. Of these 29 samples, 8 (28%) contained >3% CD133+ cells. Sphere formation was significantly higher in samples from patients with lymphatic invasion than in those without lymphatic invasion [54.5% (6/11) vs. 12.5% (2/16); P=0.033] and in samples containing >3% of CD133+ cells than in those containing ≤3% of CD133+ cells [36.4% (4/11) vs. 0% (0/16); P=0.019].

Conclusions

These findings indicate that CD133 is a valid marker for identifying cancer stem cells from fresh surgically resected colorectal cancer tissues. Furthermore, we successfully established a living tumor tissue bank using surgically resected colorectal tissues with a viability of >70%.     

CD133 is indicative for a resistance phenotype but does not represent a prognostic marker for survival of non‐small cell lung cancer patients

Despite advances in anticancer treatment, lung cancer still has poor prognosis. Recently, a cancer stem cell (CSC) hypothesis has emerged describing a small subset of tumor cells with stem cell properties. CSCs found in many solid tumors express CD133 antigen on the cell surface. The presence of CSC is correlated with poor survival of patients with glioblastomas, colon or prostate cancers. In this study, we evaluated whether CD133 expression in non‐small cell lung cancer (NSCLC) has a prognostic value in patients' survival. We also analyzed whether CD133 positivity of NSCLC correlates with the expression of resistance‐related proteins, angiogenic factors, oncogenes, proliferative activity or apoptosis. CD133 expression was retrospectively examined in a total of 88 cases of previously untreated NSCLC by immunohistochemistry. We found no correlation between CD133 positivity or the amount of CD133⁺ cells with NSCLC patients' survival, expression of oncogenes c‐myc, c‐N‐ras, c‐jun, c‐fos, c‐erbB1, c‐erbB2 or p53, angiogenic factors VEGF, VEGFR‐1, FGF, FGFR‐1, tissue factor and with proliferative activity or apoptosis in NSCLC tissues. However, there was a significant association between the expression of resistance‐related proteins glutathione S‐transferase, thymidylate synthase, catalase, O⁶‐methylguanine‐DNA methyltransferase and p170 and CD133. Because CD133 expression is linked to a resistant phenotype, detection of CD133⁺ cells may be useful to predict efficacy of cytotoxic therapy but CD133 is not a strong prognostic parameter for survival of patients with NSCLC.     

Impact of CD133 positive stem cell proportion on survival in patients with glioblastoma multiforme

Background

The aim of the study was to assess the impact of CD133-positive (CD133+) cancer stem cell proportions on treatment results of glioblastoma multiforme (GBM) patients.

Patients and methods

Patients with GBM (n = 42) received postoperative radiotherapy (± chemotherapy). Surgically excised GBM tissue sections were immunohistochemically examined for CD133 expression. The proportions of CD133+ GBM cells were determined (%). The proportion of CD133+ GBM stem cells was established by 2 independent researchers whose results were in good accordance (R = 0.8, p < 0.01). Additionally, CD133 expression levels were correlated with patients overall survival.

Results

The proportion of CD133+ cells varied between patients, being from 0.5% to 82%. Mean and median proportions of CD133+ cells of the entire study group were 33% ± 24% (mean ± SD) and 28%, respectively. Clinical data do not support the association between higher proportion of stem cells and the aggressiveness of GBM. Median survival time of the study group was 10.0 months (95% CI 9.0–11.0). The survival time clearly depended on the proportion of CD133+ cells (log rank test, p = 0.02). Median survival times for patients with low (< median) and high (≥ median) proportion of CD133+ cells were 9.0 months (95% CI 7.6–10.5) and 12.0 months (95% CI 9.3–14.7), respectively. In multivariate analysis, the proportion of CD133+ cells emerged as a significant independent predictor for longer overall survival (HR 2.0, 95% CI 1.0–3.8, p = 0.04).

Conclusions

In patients with higher stem cell proportion, significantly longer survival times after postoperative radiotherapy were achieved. Underlying reasons and possible higher sensitivity of GBM stem cells to fractionated radio-therapy should be clarified in further studies.     

Expression of the stem cell marker CD133 in recurrent glioblastoma and its value for prognosis

BACKGROUND:

Experimental data suggest that glioblastoma cells expressing the stem cell marker CD133 play a major role in radiochemoresistance and tumor aggressiveness. To date, however, there is no clinical evidence that the fraction of CD133‐positive cells in glioblastoma that recurs after radiochemotherapy may be relevant for prognosis.

METHODS:

The authors used immunohistochemistry to assess CD133 expression in 37 paired glioblastoma samples, including 1 primary tumor sample and 1 recurrent tumor sample, after patients received adjuvant radiochemotherapy. To assess the actual composition of the CD133‐positive glioblastoma cell population, fluorescence‐associated cell sorting (FACS) analysis was used to sort CD133‐positive/CD45‐negative cells that were assayed for tumor‐specific chromosomal aberrations using interphase fluorescence in situ hybridization. To rule out endothelial precursor cells, CD133‐positive fractions also were assayed with anti‐CD34 by FACS.

RESULTS:

In recurrent glioblastomas, the percentage of CD133‐positive cells was increased by 4.6‐fold compared with the percentage in primary glioblastomas, although, in some tumors, it increased up to 10‐fold and 20‐fold. Unexpectedly, the increase in CD133 expression was associated significantly with longer survival after tumor recurrence. An analysis of tumor‐specific chromosomal aberrations and in vivo studies revealed that the CD133‐positive cell compartment of recurrent glioblastoma was composed of both cancer stem cells and nontumor neural stem cells. The latter cells represented from 20% to 60% of the CD133‐positive cell population, and their relative percentage favorably affected the survival of patients with recurrent glioblastoma. Endothelial CD133‐positive/CD34‐positive precursors did not contribute to the CD133‐positive cell population.

CONCLUSIONS:

The authors hypothesized that, similar to the phenomenon described in glioblastoma models, neural stem/progenitor cells that are recruited by the tumor from surrounding brain may exert an antitumorigenic effect. Cancer 2011. © 2010 American Cancer Society.     

CD133与肿瘤干细胞研究进展

越来越多的证据表明肿瘤中存在肿瘤干细胞(cancer stem cells),并且其与肿瘤的增殖、转移、复发和对放化疗不敏感关系密切.因此,肿瘤治疗应当针对肿瘤干细胞,通过特异表面标记分选肿瘤干细胞是研究其生长特点的前提.近年来,CD133为研究最多的在于细胞(stem cell)和多种组织肿瘤干细胞表面独立表达的特异标记分子.通过CD133可以分选干细胞、前体细胞和肿瘤干细胞.众多研究表明,CD133~+肿瘤细胞与肿瘤的自我更新、分化潜能、信号传导调控、药物耐受、复发和预后等均有相关性.CD133~+细胞有望在干细胞相关疾病的治疗和肿瘤靶向治疗中发挥巨大作用.     

Characterization of the conversion between CD133+ and CD133- cells in colon cancer SW620 cell line

The state of cancer stem cells (CSC) under reversible fluctuations, which has been revealed in breast cancer cells most recently, suggests that subpopulations with distinct phenotypes and functions within cancer cells can undergo inter-conversion. To investigate the possibility in colon cancer cells, we employed CD133 as the CSC marker, and characterized CD133 expression pattern and the biological features of the CD133⁺ and CD133^- subsets. Flow cytometry revealed that CD133 was bimodally expressed in SW620 cells among eight colon cancer cell lines. The CD133⁺ clonal SW620 cells displayed a differential gene expression profile, higher cellular reactive oxygen species (ROS), enhanced tumorigenesis and resistance to 5-fluorouracil. The conversion in term of the CD133 phenotype of the sorted cells was observed in vitro and in vivo. The fraction of the CD133⁺ cells decreased from 99% to 80% in the sorted CD133⁺ population while rising from 5 to 10% in the sorted CD133^- population during the first 20-day cultivation and then stayed almost unchanged. A fraction (about 20%) of the CD133⁺ clonal cells lost their CD133 marker while about 10% of the CD133^- clonal cells acquired the CD133 marker. 5-Azacytidine enhanced the fraction of the CD133⁺ cells in both of the CD133⁺ and CD133^- clonal cells. Our data demonstrate that CD133 expression is dynamic and reversible, and reveal the inter-conversion between the CD133⁺ and the CD133^- SW620 cells, suggesting that the CD133 phenotype of SW620 cell population is retained by the conversion between the two cell subsets.     

Cisplatin selects for CD133+ cells in lung cancer cells

Objective Platinum-based chemotherapy is the first-line treatment for non-small cell lung cancer,but the chemoresistance of tumor cells continues to be a considerable challenge in the management of NSCLCs,leading to recurrence of most patients.CD133(prominin-1)is a five-transmembrane glycoprotein,and recent evidence suggests that CD133+cells are the cause of drug resistance and tumor recurrence.In this study,the correlation between cisplatin and CD133+cells was investigated systematically.Methods Four lung cancer cell lines,including A549,H460,801D and H1299,were treated with different concentrations of cisplatin.Cell viability was determined by MTT assay.Sphere-forming assay was performed to detect the capability of sphere-forming.CD133+cells was detected by BD FACScaliber flow cytometer.Results The results showed that cisplatin could increase the number of CD133+cells in both time-and dose-dependent manner.The enrichment would weaken but the proportion of CD133+cells was still higher than the basic level as incubation time extended after cisplatin was withdrawn.Compared with adherent culture,the proportion of CD133+cells was higher when the cells were maintained suspension culture.The proportion of CD133+cells significantly increased when cisplatin was provided in suspension culture.Conclusion These results revealed that cisplatin induces the enrichment of CD133+cells and CD133 is a new therapeutic target.Our data partially explained drug resistance to second-line chemotherapy in cisplatin-treated patients with NSCLCs.     

Modulation of invasive properties of CD133(+) glioblastoma stem cells: A role for MT1‐MMP in bioactive lysophospholipid signaling

Future breakthroughs in cancer therapy must accompany targeted agents that will neutralize cancer stem cells response to circulating growth factors. Since the brain tissue microenvironmental niche is a prerequisite for expression of the stem cell marker CD133 antigen in brain tumors, we investigated the invasion mechanisms specific to CD133(+) U87 glioblastoma cells in response to lysophosphatidic acid (LPA) and sphingosine 1‐phosphate (S1P), two circulating bioactive lysophospholipids and potent inducers of cancer. A CD133(+) U87 glioma cell population was isolated from parental U87 glioblastoma cells using magnetic cell sorting technology. The CD133(+)‐enriched cell population grew as neurospheres and showed enhanced maximal response to both LPA (～5.0‐fold) and S1P (～2.5‐fold) at 1 µM when compared to parental U87 cells. The increased response to LPA in CD133(+) cells, reflected by increased levels of phosphorylated ERK, was found independent of the cooperative functions of the membrane‐type‐1 matrix metalloproteinase (MT1‐MMP), while this cooperativity was essential to the S1P response. Quantitative RT‐PCR was performed and we found higher gene expression levels of the S1P receptors S1P1 and S1P2, and of the LPA receptor LPA1 in CD133(+) cells than in their parental U87 cells. These increased levels reflected those observed from in vivo experimental U87 tumor implants. Our data suggest that the CD133(+) cell subpopulation evokes most of the lysophospholipid response within brain tumors through a combined regulation of S1P/LPA cell surface receptors signaling and by MT1‐MMP. The emergence of lead compounds targeting the stem cell niche and S1P/LPA signaling in CD133(+) cancer cells is warranted. © 2009 Wiley‐Liss, Inc.     

CD133 negative glioma cells form tumors in nude rats and give rise to CD133 positive cells

CD133 is a cell surface marker expressed on progenitors of haematopoietic and endothelial cell lineages. Moreover, several studies have identified CD133 as a marker of brain tumor-initiating cells. In this study, human glioblastoma multiforme biopsies were engrafted intracerebrally into nude rats. The resulting tumors were serially passaged in vivo, and monitored by magnetic resonance imaging. CD133 expression was analyzed at various passages. Tumors initiated directly from the biopsies expressed little or no CD133, and showed no contrast enhancement suggesting an intact blood-brain barrier. During passaging, the tumors gradually displayed more contrast enhancement, increased angiogenesis and a shorter survival. Real-time qPCR and immunoblots showed that this was accompanied by increased CD133 expression. Primary biopsy spheroids and xenograft tumors were subsequently dissociated and flow sorted into CD133 negative and CD133 positive cell populations. Both populations incorporated BrdU in cell culture, and expressed the neural precursor marker nestin. Notably, CD133 negative cells derived from 6 different patients were tumorgenic when implanted into the rat brains. For 3 of these patients, analysis showed that the resulting tumors contained CD133 positive cells. In conclusion, we show that CD133 negative glioma cells are tumorgenic in nude rats, and that CD133 positive cells can be obtained from these tumors. Upon passaging of the tumors in vivo, CD133 expression is upregulated, coinciding with the onset of angiogenesis and a shorter survival. Thus, our findings do not suggest that CD133 expression is required for brain tumor initiation, but that it may be involved during brain tumor progression.     

肝细胞性肝癌组织CD133+细胞肿瘤干细胞特性的研究

目的 越来越多的研究表明,肝癌细胞系中能检测到肿瘤干细胞(cancer stem cells,CSCs)的存在.本研究旨在探讨从肝细胞性肝癌(hepatocellular carcinoma,HCC)组织样本中分离获得的CD133+细胞是否具有CSCs特性.方法 将2014-02-01-2015-06-30广西医科大学附属肿瘤医院肝胆外科25例手术获得的新鲜HCC组织和对应癌旁组织,采用酶消化法分别消化成单个肝癌细胞和单个肝细胞,利用流式细胞术检测部分单个肝癌细胞和单个肝细胞CD133的表达率.用剩余的单个肝癌细胞进行原代培养,流式细胞术将培养获得的肝癌细胞分选为CD133+和CD133-细胞,通过平板克隆形成实验、肿瘤球形成实验和裸鼠移植瘤形成实验对比分析这两组细胞的CSCs特性.结果 25例HCC组织中CD133的表达率为3.8％～8.3％,平均值为(5.8±1.6)％,而癌旁组织CD133的表达率为0.1％～0.4％,平均值为(0.2±0.1)％,两者比较差异有统计学意义,t=17.12,P＜0.001.CD133+和CD133-细胞的平均克隆率分别为(25.2±0.8)％和(7.6±0.8)％,两者比较差异有统计学意义,t=81.95,P＜0.001.CD133+和CD133-细胞的平均成球率分别为(20.3±0.6)％和(12.5±1.4)％,两者比较差异有统计学意义,t=68.17,P＜0.001.CD133+细胞的裸鼠移植瘤形成能力明显高于CD133-细胞.结论 从HCC组织样本中分离获得的CD133+细胞具有明显的CSCs特性.     

结直肠癌组织中CD133和Bmi-1的表达及临床意义

目的:探讨肿瘤干细胞标志物CD133和Bmi-1在结直肠癌组织中的表达,及其与临床病理因素的关系。方法:应用免疫组织化学方法检测60例手术切除的结直肠癌、癌旁和正常组织中CD133、Bmi-1的表达,对各指标之间的相关关系及其与性别、年龄、肿瘤的部位和大小、病理分级、侵袭深度、淋巴结转移、pTNM分期之间的关系进行分析。结果:结直肠癌组织中CD133、Bmi-1阳性表达率分别为50％（30/60）和73.33％（44/60）,显著高于癌旁及正常组织（P<0.01）;CD133的表达与淋巴结转移、pTNM分期和病理分级相关（P<0.05）,与其他临床病理参数均无关;Bmi-1的表达与淋巴结转移、pTNM分期相关（P<0.01）,与其他临床病理参数均无关;CD133和Bmi-1的表达呈正相关（P<0.05）。结论:结直肠癌组织中存在CD133+肿瘤细胞,即肿瘤干细胞;CD133+肿瘤细胞和Bmi-1表达在结直肠癌的发生、发展中起重要作用。肿瘤干细胞标志物CD133和Bmi-1表达可能成为判断肿瘤的恶性程度和预后的重要指标。