Regulatory T cells and cytokines in malignant pleural effusions secondary to mesothelioma and carcinoma

Immunotherapy against a variety of malignancies, including pleural-based malignancies, has shown promise in animal models and early human clinical trials, but successful efforts will need to address immunosuppressive factors of the tumor and host, particularly certain cytokines and CD4(+) CD25(+) regulatory T cells (Treg). Here, we evaluated the cellular and cytokine components of malignant pleural effusions from 44 patients with previously diagnosed mesothelioma, non-small cell lung cancer (NSCLC), or breast cancer and found significant differences in the immune profile of pleural effusions secondary to mesothelioma vs. carcinoma. Although a high prevalence of functionally suppressive CD4(+) CD25(+) T cells was found in carcinomatous pleural effusions, mesothelioma pleural effusions contained significantly fewer CD4(+) CD25(+) T cells. Activated CD8(+) T cells in pleural fluid were significantly more prevalent in mesothelioma than carcinoma. However, there is clear patient-to-patient variability and occasional mesothelioma patients with high percentages of CD4(+) CD25(+) pleural effusion T cells and low percentages of CD8(+) CD25(+) pleural effusion T cells can be identified. Mesothelioma pleural effusions contained the highest concentrations of the immunosuppressive cytokine transforming growth factor (TGF)-beta. Thus, the contribution of cellular and cytokine components of immunosuppression associated with malignant pleural effusions varies by tumor histology and by the individual patient. These results have implications for the development of immunotherapy directed to the malignant pleural space, and suggest the need to tailor immunotherapy to overcome immunosuppressive mechanisms in tumor environments.     

恶性胸腔积液来源树突状细胞对自体肿瘤 浸润性淋巴细胞的作用

 目的探讨恶性胸腔积液来源DCs（dentritic cells,DCs）对自体肿瘤浸润性淋巴细胞（tumor infiltration lymphocytes, TILs）增殖及杀伤肿瘤细胞能力的影响。方法用体外培养方法从16例肺癌患 者恶性胸腔积液来源分离单个核细胞,再用密度梯度离心辅以免疫磁珠分选细胞,用白细胞介素4（IL-4 ）、肿瘤坏死因子-α（TNF-α）、粒-单核细胞刺激因子（GM-CSF）等诱导出DCs,用流式细胞仪和电子 显微镜鉴定分离DCs;用IL-2、植物血凝素诱导同一患者胸腔积液单个核细胞中的TILs,用HEA-125磁珠分 选纯化肿瘤细胞,3H-TdR渗入法检测DCs对TILs增殖能力;MTT法检测TILs对肿瘤细胞杀伤活性。结果从肺 癌患者恶性胸腔积液单个核细胞中可诱导成熟DCs,电镜和光镜观察结果显示,这类DCs具有典型的DC形态 ,高表达HLA-ABC、HLA-DR、CD86,较高表达CD80、CD54,也表达CD83、CD1a。DCs可明显促进TILs增殖能 力（增加约1.7倍）。以TILs本身为对照,DCs激活的TILs对肿瘤细胞杀伤活性明显增加［从（31.80± 14.05）%提高到（51.89±13.27）%,P<0.05］。结论肺癌患者恶性胸腔积液单个核细胞中可诱导成熟DCs ,这种DCs可刺激自体TILs扩增,增加TILs杀伤肿瘤细胞的活性。     

Silencing IDO in dendritic cells: A novel approach to enhance cancer immunotherapy in a murine breast cancer model

Cancer immunotherapeutic agents (vaccines) in the form of antigen‐loaded dendritic cells (DCs) reached an important milestone with the recent approval of Provenge, the first DC vaccine for treatment of prostate cancer. Although this heralds a new era of tumor immunotherapy, it also highlights the compelling need to optimize such DC‐based therapies as they are increasingly tested and used to treat human patients. In this study we sought to augment and enhance the antitumor activity of a DC‐based vaccine using siRNA to silence expression of immunosuppressive enzyme indoleamine 2,3‐dioxygenase (IDO) in DCs. We report here that DCs loaded with tumor antigens, but with siRNA‐silenced IDO expression, were introduced into 4T1 breast tumor‐bearing mice, the treatment: (i) lengthened the time required for tumor onset, (ii) decreased tumor size compared to tumors grown for equal lengths of time in mice treated with antigen‐loaded DCs without IDO silencing and (iii) reduced CD4⁺ and CD8⁺ T cell apoptosis. Furthermore, immunization with IDO‐silenced DCs enhanced tumor antigen‐specific T cell proliferation and CTL activity, and decreased numbers of CD4⁺CD25⁺Foxp3⁺ T_reg. This study provides evidence to support silencing of immunosuppressive genes (IDO) as an effective strategy to enhance the efficacy of DC‐based cancer immunotherapeutic.     

Altered CD94/NKG2A and perforin expression reduce the cytotoxic activity in malignant pleural effusions

CD94/NKG2A is an inhibitory receptor expressed by NK cells and cytotoxic lymphocytes and, upon activation by HLA-E, downregulates the cytolytic activities of these cells thus representing a tumour immune escape mechanism.This study was aimed at assessing whether cytotoxic lymphocytes (CD8+) and NK cells from malignant pleural effusions have a deregulated expression of CD94/NKG2A.The expression of membrane CD94/NKG2A and perforin was evaluated by flow-cytometry in CD8+ and NK cells from pleural effusions and autologous peripheral blood of cancer (n = 19) and congestive heart failure (CHF) (n = 11) patients. Intracellular CD94/NKG2A expression was evaluated by flow-cytometry in pleural effusion CD8+ and NK cells from cancer patients (n = 10). Cytotoxic activity against cancer cells exerted by pleural and autologous peripheral blood T lymphocytes from cancer patients was assessed by flow-cytometry assay.Pleural CD8+ from cancer patients showed a reduced expression of membrane CD94/NKG2A and perforin when compared to autologous peripheral blood and CHF pleural effusions. Reduced numbers of NK cells were present in pleural effusions from both cancer and CHF patients. Pleural NK from cancer patients showed a reduced expression of membrane CD94/NKG2A and perforin when compared to autologous peripheral blood. Pleural T lymphocytes from cancer patients exhibited a reduced cytotoxic activity against cancer cells when compared to autologous peripheral blood T lymphocytes. The intracellular expression of CD94/NKG2A in CD8+ and NK cells from cancer patients was higher than membrane expression.In conclusion, this study provides compelling evidences of new mechanisms underlying the reduced host defence against cancer within the pleural space.     

从恶性胸腔积液体外诱导成熟树突细胞

背景与目的:树突细胞(dendriticcells,DCs)具有激活初始T细胞、引发抗原特异性免疫反应的特性。本研究拟用体外培养的方法,从肺癌患者恶性胸腔积液中诱导出功能健全的DCs。方法:取16例肺癌患者胸腔积液500～1000ml,用密度梯度离心辅以免疫磁珠分选的方法,分离出DCs前体细胞,加入IL鄄4、GM鄄CSF、TNF鄄α诱导出DCs。用电镜和光镜观察培养的DCs,用流式细胞仪检测DCs的表面分子。将不同培养时间的DCs和肿瘤浸润性淋巴细胞(tumorinfiltrativelymphocytes,TILs)作混合淋巴细胞反应,了解DCs对TILs的激活作用。结果:在体外能诱导出恶性胸腔积液来源的功能健全的DCs,电镜和光镜分析表明,这类DCs具有成熟DCs的典型形态;当DCs体外培养到第48h时,表面分子的表达率与培养0h、24h、92h、192h的DCs比较相对较高;DCs可使TILs细胞扩增。结论:可从肺癌患者恶性胸腔积液中诱导具有成熟功能的DCs。     

Demonstration of anti-tumor activity of oncolytic measles virus strains in a malignant pleural effusion breast cancer model

Breast cancer is the second leading cause of malignant effusions in cancer patients. Pleural effusion indicates incurable disease with limited palliative treatment options and poor outcome. Here, we demonstrate the therapeutic efficacy of measles virus (MV) vaccine strain derivative against malignant pleural effusion in an MDA-MB-231 xenograft model of advanced breast cancer. Both systemic intravenous (i.v.) and intrapleural (t.t.) administered virus caused massive infection and syncytia formation in the pleural tumor deposits. Intrapleural administration of 1.5 × 10⁶ plaque-forming units (PFU) total dose of MV significantly improved median survival by approximately 80% compared to the control animal group. Furthermore, we tested human dendritic cells as carriers for delivery of oncolytic MV infection to breast cancer pleural metastases. Carrier-delivered MV infection prevented accumulation of the pleural exudate and also significantly improved the survival of the treated mice. This is the first demonstration of the therapeutic potential of oncolytic virotherapy against malignant pleural effusions in a pre-clinical model of advanced breast cancer.     

New approaches in the diagnostic procedure of malignant pleural effusions

The content of carcinoembryonic antigen (CEA), carbohydrate antigen 19-9 (Ca 19-9), carbohydrate antigen 15-3 (Ca 15-3) and the expression of LewisY related carbohydrate antigens in benign and malignant pleural effusion were determined. These included 35 malignant pleural effusions: 13 breast cancers, 12 lung cancers (6 squamous cell carcinomas, 5 adenocarcinomas and 1 microcytoma), 2 mesotheliomas, 1 epithelioma, 1 kidney cancer, 1 hepatocarcinoma, 1 colon carcinoma, 3 lymphomas, 1 osteosarcoma and 9 benign pleural effusions. We showed that pleural fluid content of CEA, Ca 19-9 and Ca 15-3 were higher in malignant than in benign effusions. However CEA levels in squamous lung cancers were very high in both serum and pleural fluids whereas its levels were only slightly above the cut-off in breast cancers and in lung adenocarcinomas. Serum and pleural fluid Ca 15-3 values were higher in breast and in lung cancers with the highest values in the patients with breast cancer. Furthermore, the LewisY related carbohydrate antigens, evaluated by the reactivity of the cell extracts to MAb B3, were expressed only in breast cancers. These data suggest that pleural fluid content of CEA, and Ca 15-3 associated with the immunoblotting of cell extracts with MAb B3 appear to be very useful to improve the diagnosis of malignant pleural effusions.     

Presence of innate lymphoid cells in pleural effusions of primary and metastatic tumors: Functional analysis and expression of PD-1 receptor

The tumor microenvironment (TM) contains a wide variety of cell types and soluble factors capable of suppressing immune responses. While the presence of NK cells in pleural effusions (PE) has been documented, no information exists on the presence of other innate lymphoid cell (ILC) subsets and on the expression of programmed cell death-1 (PD-1) in NK and ILC. The presence of ILC was assessed in PE of 54 patients (n = 33 with mesothelioma, n = 15 with adenocarcinoma and n = 6 with inflammatory pleural diseases) by cell staining with suitable antibody combinations and cytofluorimetric analysis. The cytokine production of ILC isolated from both PE and autologous peripheral blood was analyzed upon cell stimulation and intracytoplasmic staining. We show that, in addition to NK cells, also ILC1, ILC2 and ILC3 are present in malignant PE and that the prevalent subset is ILC3. PE-ILC subsets produced their typical sets of cytokines upon activation. In addition, we analyzed the PD-1 expression on NK/ILC by multiparametric flow-cytometric analysis, while the expression of PD-1 ligand (PD-L1) was evaluated by immunohistochemical analysis. Both NK cells and ILC3 expressed functional PD-1, moreover, both tumor samples and malignant PE-derived tumor cell lines were PD-L1⁺ suggesting that the interaction between PD-1⁺ILC and PD-L1⁺tumor cells may hamper antitumor immune responses mediated by NK and ILC.     

Blood monocytes sample MelanA/MART1 antigen for long‐lasting cross‐presentation to CD8+ T cells after differentiation into dendritic cells

Human blood monocytes are very potent to take up antigens. Like macrophages in tissue, they efficiently degrade exogenous protein and are less efficient than dendritic cells (DCs) at cross‐presenting antigens to CD8⁺ T cells. Although it is generally accepted that DCs take up tissue antigens and then migrate to lymph nodes to prime T cells, the mechanisms of presentation of antigens taken up by monocytes are poorly documented so far. In the present work, we show that monocytes loaded in vitro with MelanA long peptides retain the capacity to stimulate antigen‐specific CD8⁺ T cell clones after 5 days of differentiation into monocytes‐derived dendritic cells (MoDCs). Tagged‐long peptides can be visualized in electron‐dense endocytic compartments distinct from lysosomes, suggesting that antigens can be protected from degradation for extended periods of time. To address the pathophysiological relevance of these findings, we screened blood monocytes from 18 metastatic melanoma patients and found that CD14⁺ monocytes from two patients effectively activate a MelanA‐specific CD8 T cell clone after in vitro differentiation into MoDCs. This in vivo sampling of tumor antigen by circulating monocytes might alter the tumor‐specific immune response and should be taken into account for cancer immunotherapy.     

Diagnostic value of metabolic phenotypes in malignant pleural effusions: expression of GLUT1 and CAIX by immunocytochemistry

BACKGROUND:

The sensitivity of conventional cytology for the detection of malignant cells in pleural effusion is insufficient. GLUT1 and CAIX are the hallmarks of metabolic change in cancer cells. The aim of this study was to evaluate the usefulness of GLUT1 and CAIX expression to the detection of cancer cells in pleural effusions.

METHODS:

A total of 150 pleural effusions were subjected to immunocytochemical staining for GLUT1 and CAIX expression. According to their cytological diagnosis and etiology, these included 58 benign effusions, 38 probable malignant effusions, 7 atypical effusions, and 47 malignant effusions,.

RESULTS:

None of the benign effusions showed GLUT1 or CAIX expression, but probable malignant effusions and malignant effusions significantly expressed GLUT1 and CAIX with a positive result in 74.5% and 63.8% of the malignant effusions, respectively, with 100% specificity. When the combination of both markers was evaluated, GLUT1 and CAIX displayed a higher diagnostic performance, ie, a sensitivity of 76.6%, specificity of 100%, and accuracy of 89.5%. A statistically significant positive correlation between GLUT1 and CAIX expression was observed. In addition, 18% of cases with cells resembling mesothelial cell hyperplasia in probable malignant effusions and 71.4% of cases with atypical cells of uncertain significance in atypical effusions were highlighted on GLUT1 or/and CAIX immunocytochemical stains.

CONCLUSIONS:

Immunocytochemical staining for GLUT1 and CAIX may be a complementary tool for the detection of malignant pleural effusions and is helpful in distinguishing cancer cells from reactive mesothelial cells. A combination of GLUT1 and CAIX immunocytochemical staining can give a higher diagnostic performance. Cancer (Cancer Cytopathol) 2011;. © 2011 American Cancer Society.     

Increased frequency of nonconventional double positive CD4CD8 αβ T cells in human breast pleural effusions

Breast cancer remains a leading cause of cancer‐related death within the female population. Immunotherapy is expected to provide additional therapeutic benefits but has met so far limited success. This may be due in part to the poor understanding of immune responses to breast cancer. Although CD4⁺ and CD8⁺ T lymphocytes infiltrate these tumors, the phenotype and functions of these cells remain ill defined. This study was designed to investigate further about these questions, taking advantage of multiparameter flow cytometry on lymphocytes derived from peripheral blood, solid tumors, metastatic lymph nodes and pleural effusions samples of patients with breast cancer. Results showed that, in addition to conventional CD4⁺ and CD8⁺ αβ T cells, individual tumors and most pleural effusions contained significant fractions of unconventional double positive (DP) CD4⁺CD8⁺ αβ T cells. These DP T cells displayed the phenotype and cytotoxic potential of effector/memory activated CD8⁺ T cells but differed essentially from these cells by a high production of IL‐5 and IL‐13. The increased frequency of DP T cells in advanced breast cancer and their high lytic potential and original cytokine profile suggest that this T‐cell subset may play a specific role in the regulation of immune responses to human breast cancer. © 2009 UICC     

Lysis of cancer cells by autologous T cells in breast cancer pleural effusates treated with anti-EpCAM BiTE antibody MT110

In the present study, the efficacy of a new drug, i.e. the bispecific single-chain antibody MT110 targeting the epithelial antigen EpCAM and the T-cell antigen CD3 was tested ex vivo in malignant pleural effusions (MPEs). EpCAM⁺ epithelial cells were found in 78% of the MPEs (n = 18). Ex vivo treatment of seven MPEs resulted in a dose-dependent specific lysis of 37 ± 27% (±SD) EpCAM⁺ cells with 10 ng/ml (P = 0.03) and 57 ± 29.5% EpCAM⁺ cells with 1,000 ng/ml MT110 (P = 0.016) after 72 h. As a prerequisite for redirected lysis, stimulation of the autologous CD4⁺ and CD8⁺ cells in MPE by 1,000 ng/ml MT110 resulted in 21 ± 17% CD4⁺/CD25⁺ and 29.4 ± 22% CD8⁺/CD25⁺ cells (P = 0.016, respectively) after 72 h. This was confirmed by a 22-fold release of TNF-α and 230-fold release of IFN-γ (1,000 ng/ml, 48 h, P = 0.03, respectively). Thus, relapsed breast cancer patients resistant to standard treatment might benefit from targeted therapy using MT110.     

Antigen spreading-induced CD8+T cells confer protection against the lethal challenge of wild-type malignant mesothelioma by eliminating myeloid-derived suppressor cells

A key focus in cancer immunotherapy is to investigate the mechanism of efficacious vaccine responses. Using HIV-1 GAG-p24 in a model PD1-based DNA vaccine, we recently reported that vaccine-elicited CD8⁺ T cells conferred complete prevention and therapeutic cure of AB1-GAG malignant mesothelioma in immunocompetent BALB/c mice. Here, we further investigated the efficacy and correlation of protection on the model vaccine-mediated antigen spreading against wild-type AB1 (WT-AB1) mesothelioma. We found that this vaccine was able to protect mice completely from three consecutive lethal challenges of AB1-GAG mesothelioma. Through antigen spreading these animals also developed tumor-specific cytotoxic CD8⁺ T cells, but neither CD4⁺ T cells nor antibodies, rejecting WT-AB1 mesothelioma. A majority of these protected mice (90%) were also completely protected against the lethal WT-AB1 challenge. Adoptive cell transfer experiments further demonstrated that antigen spreading-induced CD8⁺ T cells conferred efficacious therapeutic effects against established WT-AB1 mesothelioma and prevented the increase of exhausted PD-1⁺ and Tim-3⁺ CD8⁺ T cells. A significant inverse correlation was found between the frequency of functional PD1⁻Tim3⁻ CD8⁺ T cells and that of MDSCs or tumor mass in vivo. Mechanistically, we found that WT-AB1 mesothelioma induced predominantly polymorphonuclear (PMN) MDSCs in vivo. In co-cultures with efficacious CD8⁺ T cells, a significant number of PMN-MDSCs underwent apoptosis in a dose-dependent way. Our findings indicate that efficacious CD8⁺ T cells capable of eliminating both tumor cells and MDSCs are likely necessary for fighting wild-type malignant mesothelioma.     

Clinical application of adoptive immunotherapy by cytotoxic T lymphocytes induced from tumor-infiltrating lymphocytes

The tumor-infiltrating lymphocytes (TIL) were cultured with interleukin 2 (IL-2) to induce the cytotoxic T lymphocytes possessing autologous tumor-killing activity from 21 cancer patients (11 with solid tumor and 10 with malignant peritoneal or pleural effusions), and transferred into 7 patients as IL-2-activated TIL adoptively. The clinical application of activated TIL by adoptive transfer could result the complete regression of malignant pleural effusions in a patient with pancreatic cancer, and the nearly complete regression of malignant ascites in a patient with gastric cancer. The autologous tumor cells were isolated at the purity of more than 90% by Ficoll-Hypaque and Percoll discontinuous gradients, and then the TIL were cultured with IL-2 until 4 weeks. The optimal concentration of IL-2 was 1,500 IU/ml to obtain maximum proliferation and autologous tumor killing activity. The cytotoxic activities of activated TIL at 3 weeks-incubation was 72 +/- 15, 42 +/- 26, 27 +/- 21 and 22 +/- 15% against K562, Daudi, KATO-III and autologous tumor, respectively. By negative selection method, it was clarified that the killer cells recognizing autologous tumor consisted of CD4 or CD8 positive T lymphocyte in 43% of patients. The CD8 positive cells and CD56 positive cells increased, the CD4 positive cells and CD16 positive cells decreased by flow cytometry. The activated TIL could lyse not only cultured tumor cell lines, also other autologous tumor cells. The CD56+ cells were isolated by the Panning method, these cells could not lyse autologous tumor cells. Thus, it was indicated that the cytotoxic T lymphocytes recognizing autologous tumor could be generated from TIL and the adoptive immunotherapy of activated TIL was effective in cancer therapy.     

Group II phospholipase A2 is increased in peritoneal and pleural effusions in patients with various types of cancer

Serum levels of group II phospholipase A2 (PLA2) have been reported to be associated with stage of disease in cancer patients. These levels are also related to the malignant potential in tissues, and are an important prognostic factor. We radioimmunoassayed group II PLA2 levels in pleural and peritoneal effusions from patients with various cancers. We also investigated the production of group II PLA2 in cells in effusions from cancer patients by Northern blotting, immunocytochemistry and in situ hybridization. Immunoreactive group II PLA2 levels were significantly higher in effusions from 47 patients with various cancers, compared with those in sera and cirrhotic ascites. There was no significant correlation between group II PLA2 levels in effusions and those in sera. Group II PLA2 mRNA was expressed at a high level in cells from effusions, by Northern blot analysis, but not in those cells from blood. The localization of group II PLA2 protein and mRNA was intense in carcinoma cells and CD68-positive macrophages, determined by immunocytochemistry and in situ hybridization. In addition, IL-6 and IL-8 levels were significantly higher in effusions, in comparison with those in sera from patients, suggesting that cancer cells and macrophages produce group II PLA2 by IL-6. These group II PLA2 levels are apparently significantly increased in effusions, and the carcinoma cells and macrophages produce group II PLA2, as noted in effusions from patients with various cancers. Int. J. Cancer 74:245-250, 1997. © 1997 Wiley-Liss, Inc.     

胸腔积液中肿瘤坏死因子、癌胚抗原和神经元特异性烯醇化酶的检测及诊断价值研究

目的探讨检测胸水中肿瘤坏死因子（TNF-α）、癌胚抗原（CEA）和神经元特异性烯醇化酶（NSE）对胸腔积液的诊断价值。方法采用电化学发光酶免疫分析法检测59例结核性胸水和48例肺癌性胸水患者胸水中TNF-α、CEA和NSE水平。结果结核性胸水中TNF-α水平显著高于肺癌性胸水（P〈0.05）。肺癌性胸水中CEA和NSE明显高于结核性胸水（P〈0.01）。肺腺癌胸水中CEA升高最明显,非小细胞肺癌胸水中NSE升高最显著。联合检测CEA及NSE,诊断敏感度92.0%,准确度86.3%。结论检测TNF-α、CEA和NSE对结核性胸水和肺癌性胸水的诊断及鉴别诊断有较高的临床价值,联合检测胸水CEA和NSE可提高肺癌诊断敏感度。     

Diagnostic utility of CYFRA 21-1, carcinoembryonic antigen, CA 125, neuron specific enolase, and squamous cell antigen level determinations in the serum and pleural fluid of patients with pleural effusions.

BACKGROUND: To the authors' knowledge the role of tumor marker determination in the differential diagnosis of pleural effusions has not been established definitively. The current article reports the results of a study of CYFRA 21-1, carcinoembryonic antigen (CEA), cancer antigen 125 (CA 125), squamous cell antigen (SCC), and neuron specific enolase (NSE) in the serum and pleural fluid of patients with pleural effusions of diverse etiologies. METHODS: One hundred forty-six patients with pleural effusions (43 malignant, 47 tuberculous, 32 miscellaneous benign, and 24 paramalignant) were studied prospectively. Levels of CYFRA 21-1, CA 125, CEA, NSE, and SCC were measured by radioimmunoassay in the pleural fluid in all patients and in the serum in 118 patients. RESULTS: There were no significant differences between the serum and pleural fluid levels of tumor markers with the exception of CA 125, which was higher in the pleural fluid. With maximum specificity, the highest sensitivity in the diagnosis of pleural malignancy was obtained with a combination of CYFRA 21-1 (with a cutoff value of 150 U/L), CEA (with a cutoff value of 40 ng/mL), and CA 125 (with a cutoff value of 1000 ng/mL) in pleural fluid. NSE and SCC added no diagnostic value. The simultaneous use of tumor markers and cytology in pleural fluid increased the sensitivity from 55.8% to 81%. CONCLUSIONS: These findings suggest that a combination of CYFRA 21-1, CEA, and CA 125 in the pleural fluid can be a useful addition to pleural cytology in the diagnosis of malignant pleural effusion.     

Identification and characterization of cancer stem cells in ovarian yolk sac tumors

Recent evidence supports the cancer stem cell theory, that is, that malignant tumors arise from cells termed cancer stem cells or tumor‐initiating cells that have the ability to self‐renew and are responsible for maintaining the tumor. Cells with marked tumor‐initiating capacity have recently been identified in a number of solid tumors. CD133 (PROM1, human prominin‐1) has been used as a marker to detect stem cells (progenitor cells) and cancer stem cells (tumor‐initiating cells) in various tissues. Ovarian yolk sac tumors (YSTs) are rare and highly malignant. The present study was designed to evaluate the tumor‐forming ability of CD133⁺ cells in ovarian YST cell lines and to examine the characteristics of CD133⁺ cells, such as cell growth and invasiveness. Our data suggest ovarian YST to be maintained by a rare fraction of cancer stem‐like cells that express the cell surface marker CD133. (Cancer Sci 2010)     

A universal anti-cancer vaccine: Chimeric invariant chain potentiates the inhibition of melanoma progression and the improvement of survival

For many years, clinicians and scientists attempt to develop methods to stimulate the immune system to target malignant cells. Recent data suggest that effective cancer vaccination requires combination immunotherapies to overcome tumor immune evasion. Through presentation of both MHC-I and II molecules, DCs-based vaccine platforms are effective in generating detectable CD4 and CD8 T cell responses against tumor-associated antigens. Several platforms include DC transfection with mRNA of the desired tumor antigen. These DCs are then delivered to the host and elicit an immune response against the antigen of interest. We have recently established an mRNA genetic platform which induced specific CD8⁺ cytotoxic T cell response by DC vaccination against melanoma. In our study, an MHC-II mRNA DCs vaccine platform was developed to activate CD4⁺ T cells and to enhance the anti-tumor response. The invariant chain (Ii) was modified and the semi-peptide CLIP was replaced with an MHC-II binding peptide sequences of melanoma antigens. These chimeric MHC-II constructs are presented by DCs and induce proliferation of tumor specific CD4⁺ T cells. When administered in combination with the MHC-I platform into tumor bearing mice, these constructs were able to inhibit tumor growth, and improve mouse survival. Deciphering the immunological mechanism of action, we observed an efficient CTLs killing in addition to higher levels of Th1 and Th2 subsets in the groups immunized with a combination of the MHC-I and MHC-II constructs. These universal constructs can be applied in multiple combinations and offer an attractive opportunity to improve cancer treatment.