Reduction Of Antitumor Immunity Research Articles

Barriers to antibody therapy in solid tumors, and their solutions.

Antibody drugs have become the mainstream of cancer treatment due to advances in cancer biology and Ab engineering. However, several barriers to Ab therapy have also been identified. These include various mechanisms for Ab drug resistance, such as heterogeneity of antigen expression in tumor cells and reduction in antitumor immunity due to expression diversity, polymorphism of Fc receptors (FcR) in effector cells, and reduced function of effector cells. Countermeasures to each resistance mechanism are being investigated. This review focuses on barriers that impede the delivery of Ab drugs due to features of the solid tumor microenvironment. Unlike hematological malignancies, in which the target tumor cells are in blood vessels, clinical solid tumors contain cancer stroma, which interferes with the delivery of Ab drugs. In addition, the cancer mass itself interferes with the penetration of Ab drugs. In this article, I will consider the etiology of cancer stroma and propose a new Ab drug development strategy for solid cancer treatment centering on cancer stromal targeting (CAST) therapy using anti‐insoluble fibrin Ab‐drug conjugate (ADC), which can overcome the cancer stroma barrier. The recent success of ADCs, chimeric antigen receptor T cells (CAR‐Ts), and Bi‐specific Abs is changing the category of Ab drugs from molecular‐targeted drugs based on growth signal inhibition to cancer‐specific targeted therapies. Therefore, at the end of this review, I argue that it is time to reorient the concept of Ab drug development.

Effect of IL-15 addition on asbestos-induced suppression of human cytotoxic T lymphocyte induction

BackgroundAsbestos fibers possess tumorigenicity and are thought to cause mesothelioma. We have previously reported that exposure to asbestos fibers causes a reduction in antitumor immunity. Asbestos exposure in the mixed lymphocyte reaction (MLR) showed suppressed induction of cytotoxic T lymphocytes (CTLs), accompanied by a decrease in proliferation of CD8+ T cells. Recently, we reported that asbestos-induced suppression of CTL induction is not due to insufficient levels of interleukin-2 (IL-2). In this study, we continue to investigate the mechanism responsible for the effect of asbestos fibers on the differentiation of CTLs and focus on interleukin-15 (IL-15) which is known to be a regulator of T lymphocyte proliferation.MethodsFor MLR, human peripheral blood mononuclear cells (PBMCs) were cultured with irradiated allogenic PBMCs upon exposure to chrysotile B asbestos at 5 μg/ml for 7 days. After 2 days of culture, IL-15 was added at 1 ng/ml. After 7 days of MLR, PBMCs were collected and analyzed for phenotypic and functional markers of CD8+ T cells with fluorescence-labeled anti-CD3, anti-CD8, anti-CD45RA, anti-CD45RO, anti-CD25, and anti-granzyme B antibodies using flow cytometry. To examine the effect of IL-15 on the expression level of intracellular granzyme B in proliferating and non-proliferating CD8+ lymphocytes, PBMCs were stained using carboxyfluorescein diacetate succinimidyl ester (CFSE) and then washed and used for the MLR.ResultsIL-15 addition partially reversed the decrease in CD3+CD8+ cell numbers and facilitated complete recovery of granzyme B+ cell percentages. IL-15 completely reversed the asbestos-induced decrease in percentage of granzyme B+ cells in both non-proliferating CFSE-positive and proliferating CFSE-negative CD8+ cells. The asbestos-induced decrease in the percentage of CD25+ and CD45RO+ cells in CD8+ lymphocytes was not reversed by IL-15.ConclusionThese findings indicate that CTLs induced upon exposure to asbestos possess dysfunctional machinery that can be partly compensated by IL-15 supplementation, and that IL-15 is more effective in the recovery of proliferation and granzyme B levels from asbestos-induced suppression of CTL induction compared with IL-2.

Ingredients such as trehalose and hesperidin taken as supplements or foods reverse alterations in human Tcells, reducing asbestos exposure-induced antitumor immunity.

Exposure of human immune cells to asbestos causes a reduction in antitumor immunity. The present study aimed to investigate the recovery of reduced antitumor immunity by several ingredients taken as supplements or foods, including trehalose (Treh) and glycosylated hesperidin (gHesp). Peripheral blood CD4+ cells were stimulated with IL-2, anti-CD3 and anti-CD28 antibodies for 3 days, followed by further stimulation with IL-2 for 7 days. Subsequently, cells were stimulated with IL-2 for an additional 28 days. During the 28 days, cells were cultured in the absence or presence of 50 μg/ml chrysotile asbestos fibers. In addition, cells were treated with 10 mM Treh or 10 μM gHesp. Following culture for 28 days, reverse transcription-quantitative PCR was performed to assess the expression levels of transcription factors, cytokines and specific genes, including matrix metalloproteinase-7 (MMP-7), nicotinamide nucleotide transhydrogenase (NNT) and C-X-C motif chemokine receptor 3, in unstimulated cells (fresh) and cells stimulated with PMA and ionomycin (stimuli). The results demonstrated that compared with the control group, chrysotile-exposure induced alterations in MMP-7, NNT and IL-17A expression levels were not observed in the 'Treh' and 'gHesp' groups in stimulated cells. The results suggested that Treh and gHesp may reverse asbestos exposure-induced reduced antitumor immunity in T helper cells. However, further investigation is required to confirm the efficacy of future trials involving the use of these compounds with high-risk human populations exposed to asbestos, such as workers involved in asbestos-handling activities.

Enhanced expression of nicotinamide nucleotide transhydrogenase (NNT) and its role in a human T cell line continuously exposed to asbestos

The effects of asbestos fibers on human immune cells have not been well documented. We have developed a continuously exposed cell line model using the human T-lymphotropic virus 1 (HTLV-1)-immortalized human T cell line MT-2. Sublines continuously exposed to chrysotile (CH) or crocidolite (CR) showed acquired resistance to asbestos-induced apoptosis following transient and high-dose re-exposure with fibers. These sublines in addition to other immune cells such as natural killer cells or cytotoxic T lymphocytes exposed to asbestos showed a reduction in anti-tumor immunity. In this study, the expression of genes and molecules related to antioxidative stress was examined. Furthermore, complexes related to oxidative phosphorylation were investigated since the production of reactive oxygen species (ROS) is important when considering the effects of asbestos in carcinogenesis and the mechanisms involved in resistance to asbestos-induced apoptosis. In sublines continuously exposed to CH or CR, the expression of thioredoxin decreased. Interestingly, nicotinamide nucleotide transhydrogenase (NNT) expression was markedly enhanced. Thus, knockdown of NNT was then performed. Although the knockdown clones did not show any changes in proliferation or occurrence of apoptosis, these clones showed recovery of ROS production with returning NADPH/NADP+ ratio that increased with decreased production of ROS in continuously exposed sublines. These results indicated that NNT is a key factor in preventing ROS-induced cytotoxicity in T cells continuously exposed to asbestos. Considering that these sublines showed a reduction in anti-tumor immunity, modification of NNT may contribute to recovery of the anti-tumor effects in asbestos-exposed T cells.

CD147‑mediated reprogrammed glycolytic metabolism potentially induces immune escape in the tumor microenvironment (Review).

Impaired antitumor immunity or induced immunosuppression in the tumor microenvironment contributes significantly to tumor progression and resistance to immunotherapy. It is becoming increasingly recognized that dynamic metabolic programming orchestrates appropriate immune responses, whereas incorrect metabolic reprogramming may underlie aberrant immune remodeling. Furthermore, pathways that control cellular metabolism and immune cell function by transcriptional and post‑transcriptional mechanisms are intimately interlinked, including hypo-xia‑inducible factor 1α, c‑Myc and phosphatidylinositol 3‑kinase/protein kinase B/mammalian target of rapamycin signaling. Immunometabolism is an emerging research field involving investigation of the interaction between immunological and metabolic processes. It is likely that high levels of nutrient competition and metabolic interplay exist between tumor cells and infiltrating immune cells in the local tumor milieu, which consequently leads to a reduction in antitumor immunity or immune cell dysfunction. Recently, a metabolic molecular mechanism responsible for the tumorigenic capacity of cluster of differentiation (CD)147, which exhibits high expression on the surface of various malignant tumor cells and is associated with tumor progression via multiple non‑metabolic molecular mechanisms, was identified. The aim of the present review was to focus on the glycolytic mechanism mediated by the upregulation of CD147 in tumors and tumor‑imposed metabolic restrictions on tumor‑infiltrating immune cells, and the consequent immunological hyporesponsiveness. Cellular metabolism is becoming increasingly acknowledged as a key regulator of T‑cell function, specification and fate, and the manipulation of metabolic programming may elucidate therapeutic options for immunological disorders and tumor immunotherapy.

Aberrant expression of FoxP3 in a human Tcell line possessing regulatory T cell‑like function and exposed continuously to asbestos fibers.

Prompted by the known carcinogenic activity of asbestos, our investigations revealed that asbestos causes a reduction in antitumor immunity. One mechanism involves the enhancement of regulatory T (Treg) cell function and volume assayed using MT‑2 original cells (Org), an HTLV‑1 immortalized human Tcell line which possesses Treg‑like function. Continuous and relatively low‑dose exposure of MT‑2 to asbestos fibers yielded sublines resistant to asbestos‑induced apoptosis and enhanced Treg function via cell‑cell contact mechanisms and increased the production of soluble factors such as interleukin (IL)‑10 and transforming growth factor (TGF)‑β. Additionally, cell cycle progression was accelerated in these sublines. Subsequently, the status of the Treg‑specific transcription factor FoxP3 was examined. Unexpectedly, FoxP3 mRNA levels decreased in the sublines, although significant changes in protein expression were absent. Methylation analysis of CpG sites located in the promoter region of FoxP3 in original MT‑2 cells and sublines showed almost complete methylation in Org and slight hypomethylation in the sublines. Although treatment with the demethylating agent 5‑aza‑deoxycytidine tended to upregulate FoxP3 expression, the methylation status did not match the mRNA expression and enhanced function. Additionally, the expression of other transcription factors related to Treg did not differ between Org and subline CB1. Collectively, aberrant expression and methylation patterns of FoxP3 were detected in human Tcells continuously exposed to asbestos, although cell function was enhanced by asbestos exposure. Future analyses to identify factors responsible for Treg functional enhancements induced by asbestos, such as the investigation of surface molecules, are needed for the development of strategies to prevent the occurrence of asbestos‑induced cancers.

Inflammatory Alteration of Human T Cells Exposed Continuously to Asbestos.

Asbestos is a known carcinogen and exposure can lead to lung cancer and malignant mesothelioma. To examine the effects of asbestos fibers on human immune cells, the human T cell leukemia/lymphoma virus (HTLV)-1 immortalized human T cell line MT-2 was employed. Following continuous exposure to asbestos fibers for more than eight months, MT-2 sublines showed acquisition of resistance to asbestos-induced apoptosis with decreased death signals and increased surviving signals. These sublines showed various characteristics that suggested a reduction in anti-tumor immunity. On the other hand, inflammatory changes such as expression of MMP7, CXCR5, CXCL13 and CD44 was found to be markedly higher in sublines continuously exposed to asbestos compared with original MT-2 cells. All of these molecules contribute to lung inflammation, T and B cell interactions and connections between mesothelial cells and T cells. Thus, further investigation focusing on these molecules may shed light on the role of chronic inflammation caused by asbestos exposure and the occurrence of malignant mesothelioma. Finally, regarding peripheral T cells from healthy donors (HD) and asbestos-exposed patients with pleural plaque (PP) or malignant pleural mesothelioma (MPM), following stimulation of CD4+ T cells, T cells from MPM patients showed reduced potential of interferon (IFN)-γ expression. Moreover, levels of interleukin (IL)-6, one of the most important cytokines in chronic inflammation, in cultured supernatants were higher in PP and MPM patients compared with HD. Overall, asbestos-induced chronic inflammation in the lung as well as the pleural cavity may facilitate the onset of asbestos-induced cancers due to alterations in the interactions among fibers, immune cells such as T and B cells and macrophages, and mesothelial and lung epithelial cells. Further investigations regarding chronic inflammation caused by asbestos fibers may assist in identifying molecular targets for preventive and therapeutic strategies related to the effects of asbestos exposure.

Induction of IL-17 production from human peripheral blood CD4+ cells by asbestos exposure.

We have previously reported that chronic, recurrent and low-dose exposure to asbestos fibers causes a reduction in antitumor immunity. Investigation of natural killer (NK) cells using an invitro cell line model and comprising invitro activation using freshly isolated NK cells co-cultured with chrysotile fibers, as well as NK cells derived from asbestos-exposed patients with pleural plaque (PP) or malignant mesothelioma (MM), revealed decreased expression of NK cell activating receptors such as NKG2D, 2B4 and NKp46. An invitro differentiation and clonal expansion model for CD8+ cytotoxic T lymphocytes (CTLs) showed reduced cytotoxicity with decreased levels of cytotoxic molecules such as granzymeB and perforin, as well as suppressed proliferation of CTLs. Additionally, analysis of T helper cells showed that surface CXCR3, chemokine receptor, and the productive potential of interferon (IFN)γ were reduced following asbestos exposure in an invitro cell line model and in peripheral CD4+ cells of asbestos-exposed patients. Moreover, experiments revealed that asbestos exposure enhanced regulatory Tcell (Treg) function. This study also focused on CXCR3 expression and the Th-17 cell fraction. Following activation with T-cell receptor and co-culture with various concentrations of chrysotile fibers using freshly isolated CD4+ surface CXCR3 positive and negative fractions, the intracellular expression of CXCR3, IFNγ and IL-17 remained unchanged when co-cultured with chrysotile. However, subsequent re-stimulation with phorbol 12-myristate 13-acetate (PMA) and ionomycin resulted in enhanced IL-17 production and expression, particularly in CD4+ surface CXCR3 positive cells. These results indicated that the balance and polarization between Treg and Th-17 fractions play an important role with respect to the immunological effects of asbestos and the associated reduction in antitumor immunity.

Accelerated cell cycle progression of human regulatory Tcell-like cell line caused by continuous exposure to asbestos fibers.

Asbestos exposure causes malignant tumors such as lung cancer and malignant mesothelioma. Based on our hypothesis in which continuous exposure to asbestos of immune cells cause reduction of antitumor immunity, the decrease of natural killer cell killing activity with reduction of NKp46 activating receptor expression, inhibition of cytotoxic T cell clonal expansion, reduced CXCR3 chemokine receptor expression and production of interferon-γ production in CD4+ T cells were reported using cell line models, freshly isolated peripheral blood immune cells from health donors as well as asbestos exposed patients such as pleural plaque and mesothelioma. In addition to these findings, regulatory T cells (Treg) showed enhanced function through cell-cell contact and increased secretion of typical soluble factors, interleukin (IL)-10 and transforming growth factor (TGF)-β, in a cell line model using the MT-2 human polyclonal T cells and its sublines exposed continuously to asbestos fibers. Since these sublines showed a remarkable reduction of FoxO1 transcription factor, which regulates various cell cycle regulators in asbestos-exposed sublines, the cell cycle progression in these sublines was examined and compared with that of the original MT-2 cells. Results showed that cyclin D1 expression was markedly enhanced, and various cyclin-dependent kinase-inhibitors were reduced with increased S phases in the sublines. Furthermore, the increase of cyclin D1 expression was regulated by FoxO1. The overall findings indicate that antitumor immunity in asbestos-exposed individuals may be reduced in Treg through changes in the function and volume of Treg.

Antitumor immunologic reactivity in the relatively early period after cryosurgery: Experimental studies in the rat

An experimental investigation was performed on antitumor immunity in the relatively early postoperative period after cryosurgery, using a metastasizing rat's mammary tumor, MRMT-1. Two weeks after its inoculation, surgical excision of the tumor, cryosurgery, surgical excision plus inoculation with freezing-thawing produced vaccine, or surgical excision plus fasting for 72 hr was performed, and postoperative follow-up was done on incidences of metastases, those of metastatic death, etc. Specific immunologic reactivity was examined in the surgical excision (SE) and cryosurgery (CR) groups. The FTV and fasting groups showed more metastatic deaths as compared with the SE group. The CR and SE groups did not differ significantly from each other in incidences of lung and lymph node metastases. Specific footpad reactivity at 2 and 3 weeks after treatment was lower in the CR group than in the SE group. Winn's neutralization assay showed an inhibition of tumor growth at 1 and 3 week(s) after treatment both in the SE and in the CR groups, the inhibitory effect tending to be lower in the latter. Inactivated serum obtained at 1 week after treatment showed a facilitation of tumor growth in the SE group and a tendency of tumor suppression in the CR group, showing a significant difference between them. A mild reduction in antitumor immunity seen in the relatively early postoperative period following cryosurgery probably was not due to a blocking effect by superfluous antigens. Rather it was considered to be due to activation of suppressor cells, consequent on cryosurgical stress, and/or slow and steady absorption of antigens.