Role of immature myeloid Gr-1+ cells in the development of antitumor immunity

One of the mechanisms by which tumor cells evade the immune system is the lack of proper antigen-presenting cells. Improvement in host immunity against tumor cells can be achieved by promoting the differentiation of dendritic cells (DCs) from immature myeloid cells (Gr-1(+)Ly-6C(+)F4/80(+)) that accumulate in the bone marrow and lymphoid organs of mice with large tumor burdens. The enriched immature myeloid cells inhibit T-cell proliferation and tumor-specific T-cell response, which can be reversed by the differentiation of immature myeloid cells or depletion of F4/80(+) cells. Sorted Gr-1(+)/F4/80(+) immature myeloid cells differentiated into CD11c(+) cells that express CD80 and I-A/I-E (MHC class II) in the presence of recombinant murine granulocyte macrophage colony-stimulating factor (GM-CSF). Furthermore, intratumoral gene delivery of GM-CSF not only promoted the differentiation of carboxyfluoroscein succinimidyl ester-labeled immature myeloid cells into CD11c(+) cells with the characteristics of mature DCs (CD80(+), I-A/I-E(+)) but also enhanced innate natural killer and adaptive cytolytic T-cell activities in mice treated with interleukin (IL)-12 and anti-4-1BB combination therapy. More importantly, intratumoral delivery of GM-CSF and IL-12 genes in combination with 4-1BB costimulation greatly improved the long-term survival rate of mice bearing large tumors and eradicated the untreated existing hepatic tumor. The results suggest that inducing the maturation of immature myeloid cells, thus preventing their inhibitory activity and enhancing their antigen-presenting capability, by GM-CSF gene therapy is a critically important step in the development of effective antitumor responses in hosts with advanced tumors.     

Gemcitabine selectively eliminates splenic Gr-1+/CD11b+ myeloid suppressor cells in tumor-bearing animals and enhances antitumor immune activity.

PURPOSE: Myeloid suppressor (Gr-1(+)/CD11b(+)) cells accumulate in the spleens of tumor-bearing mice where they contribute to immunosuppression by inhibiting the function of CD8(+) T cells and by promoting tumor angiogenesis. Elimination of these myeloid suppressor cells may thus significantly improve antitumor responses and enhance effects of cancer immunotherapy, although to date few practical options exist. EXPERIMENTAL DESIGN: The effect of the chemotherapy drug gemcitabine on the number of (Gr-1(+)/CD11b(+)) cells in the spleens of animals bearing large tumors derived from five cancer lines grown in both C57Bl/6 and BALB/c mice was analyzed. Suppressive activity of splenocytes from gemcitabine-treated and control animals was measured in natural killer (NK) cell lysis and Winn assays. The impact of myeloid suppressor cell activity was determined in an immunogene therapy model using an adenovirus expressing IFN-beta. RESULTS: This study shows that the chemotherapeutic drug gemcitabine, given at a dose similar to the equivalent dose used in patients, was able to dramatically and specifically reduce the number of myeloid suppressor cells found in the spleens of animals bearing large tumors with no significant reductions in CD4(+) T cells, CD8(+) T cells, NK cells, macrophages, or B cells. The loss of myeloid suppressor cells was accompanied by an increase in the antitumor activity of CD8(+) T cells and activated NK cells. Combining gemcitabine with cytokine immunogene therapy using IFN-beta markedly enhanced antitumor efficacy. CONCLUSIONS: These results suggest that gemcitabine may be a practical strategy for the reduction of myeloid suppressor cells and should be evaluated in conjunction with a variety of immunotherapy approaches.     

Expansion of myeloid immune suppressor Gr+CD11b+ cells in tumor-bearing host directly promotes tumor angiogenesis

We demonstrate a novel tumor-promoting role of myeloid immune suppressor Gr+CD11b+ cells, which are evident in cancer patients and tumor-bearing animals. These cells constitute approximately 5% of total cells in tumors. Tumors coinjected with Gr+CD11b+ cells exhibited increased vascular density, vascular maturation, and decreased necrosis. These immune cells produce high levels of MMP9. Deletion of MMP9 in these cells completely abolishes their tumor-promoting ability. Gr+CD11b+ cells were also found to directly incorporate into tumor endothelium. Consistent with this observation, Gr+CD11b+ cells acquire endothelial cell (EC) properties in tumor microenvironment and proangiogenic culture conditions. Our data provide evidence that Gr+CD11b+ cells of immune origin induced by tumors directly contribute to tumor growth and vascularization by producing MMP9 and differentiating into ECs.     

CD80 in immune suppression by mouse ovarian carcinoma-associated Gr-1+CD11b+ myeloid cells

An elevated number of Gr-1+CD11b+ myeloid cells has been described in mice bearing transplantable tumors, and has been associated with immune suppression. We examined the role of such myeloid suppressor cells in mice bearing the spontaneously transformed syngeneic mouse ovarian surface epithelial cell line, 1D8. We observed high levels of CD80 expression by Gr-1+CD11b+ cells from spleen, ascites, and tumor tissue of mice bearing 1D8 ovarian carcinoma, whereas CD40 and CD86 were absent. CD80 expression was not detected on Gr-1+CD11b+ cells from na?ve mice. However, the expression of CD80 by Gr-1+CD11b+ cells from na?ve mice was promoted by coculture with 1D8 cells. Because irradiated 1D8 cells, but not 1D8-conditioned medium, up-regulate CD80 expression by Gr-1+CD11b+ cells, this phenomenon likely requires direct interaction. Gr-1+CD11b+ cells derived from 1D8 tumor-bearing mice provided significant suppression of antigen-specific immune responses, but Gr-1+CD11b+ cells from na?ve mice did not. Both short interfering RNA-mediated knockdown and genetic knockout of CD80 expression by Gr-1+CD11b+ cells of 1D8 tumor-bearing mice alleviated the suppression of antigen-specific immune responses. Suppression via CD80 on Gr-1+CD11b+ myeloid cells was mediated by CD4+CD25+ T regulatory cells and required CD152. CD80 knockout or antibody blockade of either CD80 or CD152 retarded the growth of 1D8 tumor in mice, suggesting that expression of CD80 on Gr-1+CD11b+ myeloid cells triggered by 1D8 ovarian carcinoma suppresses antigen-specific immunity via CD152 signaling and CD4+CD25+ T regulatory cells. Thus, CD80-dependent responses to myeloid suppressor cells may contribute to tumor tolerance and the progression of ovarian carcinoma.     

CD8+ Foxp3+ regulatory T cells mediate immunosuppression in prostate cancer.

PURPOSE: Although elevated proportions of CD4(+)CD25(+) regulatory T (Treg) cells have been shown in several types of cancers, very little is known about the existence and function of CD8(+) Treg cells in prostate cancer. In this study, we investigated prostate tumor-derived CD8(+) Treg cells and their function. EXPERIMENTAL DESIGN: Tumor-infiltrating lymphocytes (TIL) from fresh tumor specimens of patients with prostate cancer were generated and subjected to phenotypic and suppressive function analyses. In particular, we investigated the role and function CD8(+) Treg cells in prostate cancer. RESULTS: We show that high percentages of CD4(+)CD25(+) T cells are probably present in the majority (70%) of prostate TILs. Remarkably, both CD4(+) and CD8(+) T-cell subpopulations possessed potent suppressive activity. T-cell cloning and fluorescence-activated cell sorting analyses showed the presence of CD8(+)CD25(+) Treg cell clones that expressed FoxP3 and suppressed na?ve T-cell proliferation, in addition to the previously known CD4(+)CD25(+) Treg cells. These CD8(+) Treg cells suppressed na?ve T-cell proliferation mainly through a cell contact-dependent mechanism. Importantly, the suppressive function of CD8(+) Treg cells could be reversed by human Toll-like receptor 8 (TLR8) signaling. CONCLUSION: Our study shows that like CD4(+)CD25(+) Treg cells, CD8(+) Foxp3(+) Treg cells present in prostate tumor-derived TILs suppress immune responses and that their suppressive function can be regulated by TLR8 ligands, raising the possibility that the manipulation of Treg cell function by TLR8 ligands could improve the efficacy of immunotherapy for prostate cancer patients.     

Abrogation of TGF beta signaling in mammary carcinomas recruits Gr-1+CD11b+ myeloid cells that promote metastasis

Aberrant TGFbeta signaling is common in human cancers and contributes to tumor metastasis. Here, we demonstrate that Gr-1+CD11b+ myeloid cells are recruited into mammary carcinomas with type II TGF beta receptor gene (Tgfbr2) deletion and directly promote tumor metastasis. Gr-1+CD11b+ cells infiltrate into the invasive front of tumor tissues and facilitate tumor cell invasion and metastasis through a process involving metalloproteinase activity. This infiltration of Gr-1+CD11b+ cells also results in increased abundance of TGF beta 1 in tumors with Tgfbr2 deletion. The recruitment of Gr-1+CD11b+ cells into tumors with Tgfbr2 deletion involves two chemokine receptor axes, the SDF-1/CXCR4 and CXCL5/CXCR2 axes. Together, these data indicate that Gr-1+CD11b+ cells contribute to TGFbeta-mediated metastasis through enhancing tumor cell invasion and metastasis.     

CD4^＋CD25^＋调节性T细胞与肿瘤的相关性研究

通过了解CD4^＋CD25^＋调节性T细胞（CD4^＋CD25^＋Treg）表面分子的特性和CD4^＋CD25^＋Treg在外周血和组织中的表达,认识CD4^＋CD25^＋Treg在肿瘤免疫调节中的作用,探索其作用的分子机制。     

CD4+ T cells are able to promote tumor growth through inhibition of tumor-specific CD8+ T-cell responses in tumor-bearing hosts

Modulation of the immune response by established tumors may contribute to the limited success of therapeutic vaccination for the treatment of cancer compared with vaccination in a preventive setting. We analyzed the contribution of the CD4+ T-cell population to the induction or suppression of tumor-specific CD8+ T cells in a tumor model in which eradication of tumors crucially depends on CD8+ T cell-mediated immunity. Vaccine-mediated induction of protective antitumor immunity in the preventive setting (i.e., before tumor challenge) was CD4+ T cell dependent because depletion of this T-cell subset prevented CD8+ T-cell induction. In contrast, depletion of CD4+ cells in mice bearing established E1A+ tumors empowered the mice to raise strong CD8+ T-cell immunity capable of tumor eradication without the need for tumor-specific vaccination. Spontaneous eradication of tumors, which had initially grown out, was similarly observed in MHC class II-deficient mice, supporting the notion that the tumor-bearing mice harbor a class II MHC-restricted CD4+ T-cell subset capable of suppressing a tumor-specific CD8+ T-cell immune response. The deleterious effects of the presence of CD4+ T cells in tumor-bearing hosts could be overcome by CD40-triggering or injection of CpG. Together these results show that CD4+ T cells with a suppressive activity are rapidly induced following tumor development and that their suppressive effect can be overcome by agents that activate professional antigen-presenting cells. These observations are important for the development of immune interventions aiming at treatment of cancer.     

Peripheral T-cell tolerance associated with prostate cancer is independent from CD4+CD25+ regulatory T cells

CD4(+)CD25(+)Foxp3(+) regulatory T cells (Treg) are thought to suppress the natural and vaccine-induced immune response against tumor-associated antigens (TAA). Here, we show that Treg accumulate in tumors and tumor-draining lymph nodes of aging transgenic adenocarcinoma of the mouse prostate (TRAMP) male mice, which spontaneously develop prostate cancer. TAA overexpression and disease progression associate also with induction of TAA-specific tolerance. TAA-specific T cells were found in the lymphoid organs of tumor-bearing mice. However, they had lost the ability to release IFN-gamma and kill relevant targets. Neither in vivo depletion of Treg by PC61 monoclonal antibody followed by repeated vaccinations with antigen-pulsed dendritic cells nor the combined treatment with 1-methyl-L-tryptophan inhibitor of the enzyme indoleamine 2,3-dyoxigenase, PC61 antibody, and dendritic cell vaccination restored the TAA-specific immune response. Treg did not seem to control the early phases of tolerance induction, as well. Indeed, depletion of Treg, starting at week 6, the age at which TRAMP mice are not yet tolerant, and prolonged up to week 12, did not avoid tolerance induction. A similar accumulation of Treg was found in the lymph nodes draining the site of dendritic cell vaccination both in TRAMP and wild-type animals. Hence, we conclude that Treg accrual is a phenomenon common to the sites of an ongoing immune response, and in TRAMP mice in particular, Treg are dispensable for induction of tumor-specific tolerance.     

CD4+ CD25+调节性T细胞及其与肿瘤的关系

CD4+CD25+调节性T细胞是体内自然存在的,能够分泌IL-4、IL-10、转化生长因子-β(TGF-β),表达IL-2Rα(CD25)、细胞毒性T淋巴细胞抗原4(CTLA-4)分子,对效应性T细胞具有抑制作用,是调节性T细胞的重要亚群,参与肿瘤的生长、自身免疫性疾病的发生及耐受移植排斥.现就该类细胞抑制作用机制的研究近况及相关免疫治疗主要是肿瘤免疫治疗的研究进展作一综述.     

CD4+CD25+调节性T细胞与抗肿瘤免疫

CD4^＋CD25^＋调节性T（CD4^＋CD25^＋Tr）细胞是一类维持机体自身耐受的T细胞亚群,分布广泛,但不同组织表型有所不同。它们可由胸腺自然产生,也可在外周血中诱导产生,其活化要依赖于特异性抗原的存在。CD4^＋CD25^＋Tr细胞发挥抑制效应是通过细胞接触依赖或分泌细胞因子这两种方式。去除CD4^＋CD25^＋Tr细胞或抑制其功能,重新募集效应性T细胞能够增强机体抗肿瘤作用,这将成为一种可行的肿瘤免疫治疗方法。     

CD4+CD25+调节性T细胞及其在肿瘤免疫治疗中的意义

CD4^＋ CD25^＋调节性T细胞是具有独特免疫调节功能的T细胞亚群。近年来研究发现各种恶性肿瘤患者外周血及肿瘤环境中该细胞比例增加，去除CD4^＋ CD25^＋调节性T细胞或封闭其抑制功能可以增强抗肿瘤免疫反应。CD4^＋ CD25^＋调节性T细胞成为肿瘤免疫治疗的新靶点。     

Hyporesponsiveness to natural killer T-cell ligand alpha-galactosylceramide in cancer-bearing state mediated by CD11b+ Gr-1+ cells producing nitric oxide

CD1d-restricted natural killer T (NKT) cells are a potential therapeutic target for cancer, for which several clinical trials have already been reported. NKT cells are specifically activated by a synthetic glycolipid, alpha-galactosylceramide (alpha-GalCer). However, it is known that, in human cancer patients, NKT cells express a degree of hyporesponsiveness to alpha-GalCer. In this study, we have examined the mechanism by which hyporesponsiveness to alpha-GalCer can be induced. In cancer-bearing mice, alpha-GalCer-induced NKT cell expansion, cytokine production, cytotoxicity, and antimetastatic effect in vivo were all significantly impaired. In fact, alpha-GalCer could eliminate metastatic disease in naive animals but failed to protect cancer-bearing mice. CD11b(+) Gr-1(+) cells were particularly increased in cancer-bearing mice and were necessary and sufficient for the suppression of the alpha-GalCer response in a nitric oxide-mediated fashion. Administration of a retinoic acid to cancer-bearing mice reduced the population of CD11b(+) Gr-1(+) cells and effectively restored alpha-GalCer-induced protection. These results show a novel feature of NKT cell function in cancer. Furthermore, our data suggest a new strategy to enhance NKT cell-mediated anticancer immune responses by suppressing CD11b(+) Gr-1(+) cell functions.     

CD4+CD25+ regulatory T cells in patients with gastrointestinal malignancies: possible involvement of regulatory T cells in disease progression

BACKGROUND: Active suppression by CD4+CD25+ regulatory T cells plays an important role in the down-regulation of the response of T cells to foreign and self antigens. Experimental tumor models in mice revealed that regulatory T cells inhibit antitumor immune responses. The purpose of the current study was to demonstrate the possible involvement of CD4+CD25+ regulatory T cells in immune system impairment in patients with gastrointestinal malignancies. METHODS: The phenotypes of lymphocytes, particularly those of CD4+CD25+ T cells, were analyzed in peripheral blood in 149 patients with gastrointestinal malignancies and in ascites in 7 patients with peritoneal dissemination. In addition, cytokine production after in vitro stimulation was examined in CD4+CD25+ and CD4+CD25- T cells isolated from patients with malignant disease. RESULTS: Compared with healthy volunteers, patients with gastrointestinal malignancies had a higher proportion of CD4+CD25+ T cells in peripheral blood, due to the presence of a drastically smaller number of CD4+CD25- T cells. Among patients with gastric carcinoma, those with higher percentages of CD4+CD25+ T cells had a poorer prognosis than did those with lower percentages. CD4+CD25+ T cells also were present in greater proportions in ascites from patients who had advanced-stage disease with peritoneal dissemination. Isolated CD4+CD25+ T cells from patients with malignant disease produced interleukin (IL)-4 and IL-10 but not IL-2 or interferon-gamma; these cells also inhibited cytokine production by CD4+CD25- T cells after in vitro stimulation. CONCLUSIONS: The relative increase in CD4+CD25+ regulatory T cells may be related to immunosuppression and tumor progression in patients with gastrointestinal malignancies. This finding suggests that the use of immunomodulatory therapy to treat patients with gastrointestinal malignancies may be an effective strategy.     

Distinct and overlapping roles of interleukin-10 and CD25+ regulatory T cells in the inhibition of antitumor CD8 T-cell responses

Lack of antitumor immunity is often related to impaired CD8 T-cell responses that could result from a poor priming capacity by tumor-infiltrating dendritic cells (TIDC) and/or further inhibition by regulatory T cells (T(reg)). Interleukin-10 (IL-10) has been implicated in the inhibition of TIDC as well as in the generation and functions of T(reg). Here, we address some of the respective and possibly overlapping roles of IL-10 and CD25+ T(reg) in CD8 antitumor immunity. Whereas tumor antigen-specific CD8 T cells proliferated in vivo in the presence of IL-10 or T(reg), optimal effector functions were observed in mice lacking both IL-10 and T(reg). Indeed, tumors grown in normal but not in IL-10-deficient or CD25-depleted mice induced tumor antigen-specific CD8 suppressor T cells. Suppression involved transforming growth factor-beta. Similarly, both IL-10 and T(reg) were responsible for impaired CD8 T cell priming by TIDCs, but IL-12 production by TIDCs was prevented only by T(reg)-independent IL-10. Subsequently, IL-10 defect and T(reg) depletion were required to achieve optimal induction of CD8 T-cell effectors by TIDC following CpG activation. Our results point out major redundant and nonredundant roles for IL-10 and T(reg) in the inhibition of TIDC-mediated generation of antitumor CD8 T-cell response.     

Multipotency of CD11bhighGr-1+ immature myeloid cells accumulating in oral squamous cell carcinoma-bearing mice

We demonstrated the accumulation of CD11b(high)Gr-1(+) cells in a murine model of squamous cell carcinoma (SCC). Inoculation of NR-S1K cells derived from oral SCC induced a rapid and clear accumulation of CD11b(high)Gr-1(+) cells in secondary lymphoid organs as well as in peripheral blood. Phenotypic and morphological analyses revealed that these CD11b(high)Gr-1(+) cells were not lymphoid lineage cells, mature dendritic cells, macrophages, or granulocytes. Although the freshly isolated CD11b(high) cells lacked antigen-presenting capacity, they acquired a potent antigen-presenting capacity that included the induction of MHC class II after culture with GM-CSF and IL-4 in vitro. These results suggest that CD11b(high) cells that accumulate in tumor-bearing hosts are immature myeloid cells, but have considerable potential to differentiate into potent antigen-presenting cells under appropriate culture conditions. The use of in vitro differentiated CD11b(high) cells may be a potential strategy for obtaining patient-matched dendritic cells for tumor immunotherapy.     

CD4+CD25+ regulatory T cells in human hematopoietic cell transplantation

Naturally occurring CD4+CD25+ regulatory T cells (T(reg)) are differentiated T lymphocytes actively involved in the control of peripheral immunity. Over the past few years, a number of animal studies have demonstrated the critical role of these cells in the outcome of allogeneic hematopoietic stem cell transplantation (HCT). In these models, T(reg) can exert a potent suppressive effect on immune effector cells reactive to host antigens and prevent graft versus host disease (GVHD) while preserving the graft-versus-leukemia effect (GVL). The present review summarizes current knowledge on the role of T(reg) populations in humans following allogeneic HCT. Recent investigations focusing on T(reg) in transplant patients have generated conflicting results mostly due to the use of different parameters to assess T(reg). Nonetheless, these studies suggested that an imbalance between T(reg) and effector cells during immune reconstitution can substantially impair regulatory mechanisms and contributes to the development of GVHD. Building on these studies, a number of therapeutic strategies are being developed to positively modulate T(reg) pools in vivo and prevent or even correct GVHD. Conversely, clinical interventions can also be envisaged to decrease T(reg) activity in vivo and enhance the GVL effect. These potential strategies are discussed herein. Coming years will undoubtedly yield additional knowledge on how to use T(reg) subsets in vivo and successfully control and modulate immune responses in patients post-HCT.     

CD4~+CD25~+调节性T细胞与抗肿瘤免疫

CD4~+CD25~+调节性T(CD4~+CD25~+Tr)细胞是一类维持机体自身耐受的T细胞亚群,分布广泛,但不同组织表型有所不同。它们可由胸腺自然产生,也可在外周血中诱导产生,其活化要依赖于特异性抗原的存在。CD4~+CD25~+Tr细胞发挥抑制效应是通过细胞接触依赖或分泌细胞因子这两种方式。去除CD4~+CD25~+Tr细胞或抑制其功能,重新募集效应性T细胞能够增强机体抗肿瘤作用,这将成为一种可行的肿瘤免疫治疗方法。     

CD4+CD25+调节性T细胞与大肠癌的关系

近来CD4 CD25 调节性T细胞对肿瘤免疫调节的抑制作用正受到越来越多的重视,许多恶性肿瘤患者外周血都存在CD4 CD25 调节性T细胞比例上调,机体抗肿瘤免疫力下降.大肠癌的发病率正逐年上升,其发病分子机制已为熟知,免疫机制研究不多,国内尚未见大肠癌与该类细胞关系的报道,现对CD4 CD25 调节性T细胞在大肠癌的发生、发展、治疗中的研究综述如下.     

CD4+CD25+CCR6+调节性T细胞在小鼠乳腺癌模型中对CD8+T细胞的抑制作用

目的：观察CD4+CD25+CCR6+调节性T细胞（简称CCR6+Tregs）体内对CD8+T细胞功能的抑制作用,并探讨其与肿瘤免疫逃逸的关系。方法：建立4T1乳腺癌细胞荷瘤裸鼠模型,FACS分选CCR6+Tregs,检测其Foxp3的表达;FACS分选4T1特异性CD8+T细胞,CFSE标记后分别与CCR6+Tregs或CCR6Tregs共同过继转输入4T1荷瘤裸鼠体内,观察荷瘤裸鼠肿瘤生长情况和小鼠存活时间;FACS检测肿瘤组织中CD8+T细胞的增殖、细胞因子IFNγ的产生和颗粒酶B的表达情况。结果：CCR6+Tregs和CCR6Tregs均高表达Foxp3;CCR6+Tregs和CD8+T细胞共转输组4T1荷瘤裸鼠肿瘤的生长明显快于CCR6Tregs共转输组和CD8+T细胞单转输组,同时该组荷瘤裸鼠生存时间也明显缩短（P<0.05）;CCR6+Tregs和CD8+T细胞共转输组CD8+T细胞的增殖、IFNγ的产生和颗粒酶B的表达均明显低于CCR6Tregs共转输组和CD8+T细胞单转输组（P<0.05）。结论：CCR6+Tregs在体内可以有效抑制CD8+T细胞的功能,其在肿瘤免疫逃逸和肿瘤发生、发展中发挥重要作用。