Shui Cao

Myeloid-Derived Suppressor Cells Suppress Antitumor Immune Responses through IDO Expression and Correlate with Lymph Node Metastasis in Patients with Breast Cancer

Myeloid-derived suppressor cells (MDSCs) represent heterogeneous immunosuppressive cells in multiple cancer types and display potent immunosuppressive activity on T cells. We have shown the increased expression of IDO in breast cancer. Because IDO plays a pivotal role in immune tolerance via suppressing T cell function, the aim of this study was to investigate the expression of IDO in MDSCs in breast cancer and its role in MDSC-mediated inhibition of immune surveillance. The proportion of MDSCs with the phenotype of CD45+CD13+CD33+CD14−CD15− significantly increased in primary cancer tissues and patients’ peripheral blood. IDO expression was significantly upregulated in MDSCs isolated from fresh breast cancer tissues (fresh MDSCs [fMDSCs]), which correlated with increased infiltration of Foxp3+ regulatory T cells in tumors and lymph node metastasis in patients. fMDSCs inhibited IL-2 and anti-CD3/CD28 mAb-induced T cell amplification and Th1 polarization but stimulated apoptosis in T cells in an IDO-dependent manner. CD33+ progenitors isolated from healthy donors’ umbilical cord blood were cocultured with breast cancer cell line MDA-MB-231 cells to induce MDSCs. IDO expression was upregulated in induced MDSCs, which required phosphorylation of STAT3, but not STAT1. IDO was required for induced MDSCs’ immunosuppressive activity on T cells, which was blocked by IDO inhibitor 1-methyl-L-tryptophan or STAT3 antagonist JSI-124. Consistently, increased STAT3 phosphorylation level was found in fMDSCs. Together, our findings suggest that STAT3-dependent IDO expression mediates immunosuppressive effects of MDSCs in breast cancer. Thus, inhibition of MDSC-induced T cell suppression by blocking IDO may represent a previously unrecognized mechanism underlying immunotherapy for breast cancer.

Dendritic cell-activated cytokine-induced killer cells enhance the anti-tumor effect of chemotherapy on non-small cell lung cancer in patients after surgery

BACKGROUND AIMS Cytokine-induced killer (CIK) cells have shown cytolytic activity against several tumor cells in vitro and in animal tumor models. Furthermore, CIK cells activated by dendritic cell (DC) stimulation show increased anti-tumor activity. This study aimed to evaluate the clinical efficacy of DC-activated CIK cell treatment following regular chemotherapy and the effects of this therapy on immune responses in patients with non-small cell lung cancer (NSCLC) after surgery. METHODS A paired study, with 42 patients in each group with stage I-IIIa NSCLC after surgery, was performed. Patients received chemotherapy alone (CT) or chemotherapy and DC-activated CIK cell treatment (immuno-CT). Disease-free survival (DFS) and overall survival were evaluated. CIK cell cytotoxicity against tumor cells was detected using a lactate dehydrogenase-based method. Serum cytokine levels in the immuno-CT group were detected using cytokine antibody arrays. RESULTS The cytotoxicity of CIK cells was significantly enhanced by DC activation. The 2-year overall survival rate in the immuno-CT group was significantly improved compared with the CT group (94.7 +/- 3.6% versus 78.8 +/- 7.0%, P < 0.05). The 2-year DFS of these two groups showed no significant difference. DC-activated CIK cell treatment increased production of cytokines that have known anti-tumor effects, including IFN-gamma, MIG, TNF-alpha and TNF-beta, in patients who had no progression, but they were not found in patients who developed recurrence/metastasis. CONCLUSIONS This study suggests that the role of DC-activated CIK cells in improvement of chemotherapy for malignant tumor treatment is associated with up-regulation of the production of cytokines involved in the anti-tumor effect.

MiR-205 in cancer: An angel or a devil?

MicroRNAs (MiRNAs) are small non-coding RNAs that regulate their target genes expression at the post-transcriptional level. As accumulating properties of miR-205 have been identified, complex roles of miR-205 in tumor initiation and progression are emerging. MiR-205 acts either as a tumor suppressor through inhibiting proliferation and invasion, or as an oncogene through facilitating tumor initiation and proliferation, depending on the specific tumor context and target genes. In this review, we focus on the properties of miR-205 in cancers to shed light on better management of various fatal malignancies. Moreover, we discuss epigenetics that may account for the fluctuation of miR-205 expression. In addition, we sketch a network of miR-205 and its targets to further elucidate the mechanisms through which miR-205 exerts its multiple functions.

Upregulated Expression of Indoleamine 2, 3-Dioxygenase in Primary Breast Cancer Correlates with Increase of Infiltrated Regulatory T Cells In Situ and Lymph Node Metastasis

IDO has been reported to induce immunotolerance and promote metastasis in solid malignancy, but the mechanisms involved were not fully understood. In this study, the expression of IDO in primary breast cancer was examined and the correlation between the expression levels of IDO and the densities of Foxp3+ Tregs in situ was studied. The IDO stably-expressing CHO cells(IDO/CHO) were generated to evaluate the induction of Foxp3+ Tregs after coculturing with CD3+ T cells in vitro. The IDO expression in cancer was higher than that in benign diseases both at RNA and protein levels. The IDO expression was significantly upregulated in tumors of more advanced stages and with more extensive lymph node metastasis, and displayed positive linear correlation with the density of Foxp3+ Tregs. We further demonstrated that CD4+CD25+CD127− Tregs could be amplified by coculturing CD3+ T cells with IDO/CHO cells in vitro which displayed increasing Foxp3 expression both at mRNA and protein levels. Our results implied that up-regulation of IDO in primary breast cancer may inhibit local immune surveillance and promote metastasis by favoring development and infiltration of Foxp3+ Tregs in the tumor microenvironment.

Maintenance therapy with autologous cytokine-induced killer cells in patients with advanced epithelial ovarian cancer after first-line treatment.

Cytokine-induced killer (CIK) cells have shown cytolytic ability against ovarian cancer cells in vitro and in vivo. This study was aimed to evaluate the clinical efficacy of maintenance therapy of CIK cells in patients with advanced epithelial ovarian cancer after first-line treatment. A paired study was performed in patients with stages IIB–IV epithelial ovarian cancer after cytoreductive surgery followed by 6–8 courses of carboplatin/paclitaxel chemotherapy. A total of 92 patients who achieved complete remission after first-line treatment were enrolled in this study. Forty-six patients in the treatment group received CIK cells transfusion monthly, whereas the other 46 patients in the control group received observation with follow-up. Progression-free survival (PFS), overall survival (OS), and toxicity were evaluated. Our results showed that median PFS was 37.7 months in the treatment group and 22.2 months in the control group (P=0.004). However, although median OS in the treatment group (61.5 mo) was longer than that in the control group (55.9 mo), there was no significant difference (P=0.289). The subgroup analysis revealed that the survival advantage of PFS from immunotherapy was independent of the extent of debulking surgery and pathologic stage. After 2 courses of CIK cells transfusion, the proportion of CD4+CD25+CD127− regular T cells in the peripheral blood significantly decreased (P=0.006). No grades III and IV adverse reaction were found during CIK cells infusion. Maintenance therapy with CIK cells improved the PFS in patients with advanced ovarian cancer after first-line treatment with slight side effects. However, the benefits with respect to OS are still pending.

Can the dual-functional capability of CIK cells be used to improve antitumor effects?

Cytokine-induced killer (CIK) cells, which display both potent anti-tumor ability of T lymphocytes and non-major histocompatibility complex (MHC) restricted killing tumor cells capacity of natural killer (NK) cells are capable of recognizing and lysing a broad array of tumor targets. They have begun to be used in clinical care with good prospects for treatment success. CIK cells are a heterogeneous cell population that contain CD3(+)CD56(+) cells, CD3(-)CD56(+) natural killer (NK) cells and CD3(+)CD56(-) T cells on which much attention has been focused. This review will summarize the connections and differences among CD3(+)CD56(+)CIK cells, CD3(-)CD56(+) NK cells and CD3(+)CD56(-) T cells in the following aspects: the main cell surface molecule, killing mechanism, and clinical applications so that treatment with CIK cells can be optimized and further to enhance the antitumor effect.

Immunotherapy with cytokine-induced killer cells as an adjuvant treatment for advanced gastric carcinoma: A retrospective study of 165 patients

BACKGROUND Cytokine-induced killer (CIK) cells have demonstrated antitumor effects in vitro and in vivo. The purpose of this study was to evaluate the effect of CIK cell treatment as an adjuvant immunotherapy on the prognosis of gastric carcinoma in patients after surgery. METHODS The patients with stage II-III gastric carcinoma after gastrectomy, including 53 patients receiving autologous CIK cell treatment combined with chemotherapy (CIK group) and 112 patients in the corresponding period receiving chemotherapy alone (control group), were retrospectively studied. The patients in the CIK group were matched to those in the control group regarding the sex and age of patients, tumor site, histological type, pathological grade, tumor size, clinical stage, and chemotherapy plan. Progression-free survival (PFS) and overall survival (OS) were evaluated. RESULTS The 5-year OS rate in the CIK group was significantly improved compared to that in the control group (56.6% vs. 26.8%, p=0.014). The 5-year PFS rate in the CIK group was also significantly improved compared to that in the control group (49.1% vs. 24.1%, p=0.026). The median PFS (36.0 months) and OS (96.0 months) in the CIK group were significantly prolonged than those in the control group (23.0 months for median PFS and 32.0 months for median OS, p=0.028 and p=0.003). No serious side effect was observed in the CIK group. CONCLUSIONS This study suggests that immunotherapy with CIK cells may serve as an adjuvant treatment to prolong the survival of patients with stage II-III gastric carcinoma.

Increased prevalence of regulatory T cells in the lung cancer microenvironment: a role of thymic stromal lymphopoietin

Expansion of CD4+CD25+ regulatory T cells (Tregs) in tumor microenvironment was one of the mechanisms by which cancer cells escaped host defense. Thymic stromal lymphopoietin (TSLP) contributes to the generation of natural Tregs in thymus. Therefore, the purpose of this report was to investigate the role of TSLP in the increasing prevalence of Tregs in lung cancer microenvironment. The expression ratio of TSLP protein in tumor tissues was significantly increased compared with that in benign lesion and non-cancer lung tissue. The prevalence of Tregs in tumor microenvironment was correlated with the expression of TSLP in lung cancer. Dendritic cells (DCs) were induced from peripheral blood mononuclear cells (PBMCs) collected from lung cancer patients and left unstimulated (imDCs) or exposed to hTSLP (TSLP-DCs) or LPS (LPS-DCs). TSLP-DCs expressed intermediate levels of CD83 and high levels of CD86, CD11C, and HLA-DR, which showed a characteristic of less mature DCs. TSLP-DCs secreted low levels of IL-6, IL-12, IL-10, TNF-α and IFN-γ, and high levels of TGF-β and MDC. The percentage of Tregs in CD4+CD25− T cells cocultured with TSLP-DCs group was statistically higher than that of LPS-DCs and imDCs. Transwell assays showed that TSLP-DCs exhibited increased ability to attract the migration of CD4+CD25− Tregs, when compared with imDCs. These results indicated that TSLP proteins were expressed in lung tumor tissue and correlated with the prevalence of Tregs. TSLP-DCs could induce CD4+CD25− T cells to differentiate into CD4+CD25+foxp3+ T cells and the migration of CD4+CD25+ T cells.

CD4+T Cells in CIKs (CD4+ CIKs) Reversed Resistance to Fas-Mediated Apoptosis Through CD40/CD40L Ligation Rather Than IFN-γ Stimulation

BACKGROUND Cytokine-induced killer cells (CIKs) are nonspecific antitumor effectors with superior advantages. CD4+ CIKs can induce Fas-dependent apoptosis in sensitive Raji cells. Here, a Fas-dependent apoptosis was detected in resistant breast cancer MDA-MB-231 cells, and underlying mechanisms were discriminated. METHODS Amplification of CIKs and purification of CD4+ CIKs were performed in 15 patients with malignant solid tumors. The expression of CD40L and soluble cytokines in CD4+ CIKs were analyzed. The apoptotic rates of tumor cells and the expression of Fas on membranes were detected using flow cytometry assay. The specific blocking antibodies against FasL, CD40L, and interferon-gamma (IFN-gamma) were added to abolish their effects. The changes of 4 apoptosis-related genes (Bcl-2, Bax, Fas-associating protein with death domain [FADD], and FLICE inhibitory protein [c-FLIP]) in MDA-MB-231 cells cocultured with CD4+ CIKs were measured by real-time quantitative reverse-transcriptase polymerase chain reaction after 6 hours and 24 hours with or without blocking antibodies. RESULTS Upregulated expression of membrane-attached CD40L and dramatically increased secretion of soluble CD40L and IFN-gamma were identified in CD4+ CIK. The susceptibility to Fas-mediated apoptosis of insensitive MDA-MB-231 cells was elevated after being pretreated with supernatants from CD4+ CIK. After coculture with CD4+ CIK, apoptosis in MDA-MB-231 cells paralleled with enhanced expression of Fas was blocked fully by either anti-FasL or anti-CD40L, but only partly by anti-IFN-gamma antibodies. The anti-CD40L monoclonal antibody (McAb) rather than anti-IFN-gamma McAb induced significant increase of c-FLIP, which negatively correlated with the apoptosis observed in MDA-MB-231 cells. CONCLUSIONS Apoptosis in MDA-MB-231 cells induced by CD4+ CIK is Fas-dependent. The reversion of Fas resistance is mediated through CD40/CD40L ligation rather than IFN-gamma stimulation by inhibiting synthesis of c-FLIP.

Alloreactive natural killer cells promote haploidentical hematopoietic stem cell transplantation by expansion of recipient-derived CD4+CD25+ regulatory T cells

Alloreactive NK cells (Allo‐NKs) have been shown to exert advantageous effects on the outcomes of haploidentical hematopoietic stem cell transplantation (Haplo‐HSCT) for cancer treatment. However, the mechanisms of action of Allo‐NKs remain unclear. We established a novel Haplo‐HSCT conditioning regimen composed of Allo‐NKs and a low dose of immunosuppressive drugs (Allo‐NKs + Chemo) to investigate alternative mechanisms besides direct cytotoxicity. The inhibitory effects of different cell subsets on the donor–recipient mixed lymphocyte reactions (MLRs) were evaluated after Haplo‐HSCT. The quantities and functions of CD4+CD25+ regulatory T cells (Tregs) and dendritic cells (DCs) in the spleen and the thymus were examined. Our results showed that the Allo‐NKs + Chemo regimen induced systemic tolerance, and that CD4+CD25+ Tregs played a significant role in inducing and maintaining systemic tolerance after Haplo‐HSCT. Alloreactive NK cells promoted the expansion of recipient‐derived CD4+CD25+CD127− Tregs in the thymus and the spleen which could be amplified in vitro by the immature donor‐derived DC subset isolated from the thymus of Allo‐NKs + Chemo‐treated mice. Our findings suggested that Allo‐NKs are capable of inducing systemic tolerance after Haplo‐HSCT by assembling donor‐derived immature DCs to expand recipient‐derived Treg cells in the thymus.