The interaction between gut microbiome and anti-tumor drug therapy

A large number of symbiotic gut microbiome exists in the human gastrointestinal micro-ecosystem. The daily diet, lifestyle, and body constitution influence the type and quantity of gut microbiome in the body. Increasing evidence demonstrates that the gut microbiome can affect tumor development and progress. We discuss in this paper how the gut microbiome impacts tumor pathology through DNA damage, production of dietary and microbial metabolites, altered cellular signaling pathways, immune system suppression, and involvement in pro-inflammatory pathways changing gut microbiome composition. The gut microbiome acts on different types of the anti-tumor drug through bacterial translocation, immuno-modulation, metabolic modulation, enzymatic degradation, and reduction of microbial diversity. This article summarized the aforementioned by reviewing recent studies on the interaction among the gut microbiome, tumor development, and antitumor drugs.     

Inhibition of vascular adhesion protein-1 enhances the anti-tumor effects of immune checkpoint inhibitors

Modulation of the immunosuppressive tumor microenvironment (TME) is essential for enhancing the anti-tumor effects of immune checkpoint inhibitors (ICIs). Adhesion molecules and enzymes such as vascular adhesion protein-1 (VAP-1), which are expressed in some cancers and tumor vascular endothelial cells, may be involved in the generation of an immunosuppressive TME. In this study, the role of VAP-1 in TME was investigated in 2 murine colon cancer models and human cancer cells. Intraperitoneal administration of the VAP-1-specific inhibitor U-V296 inhibited murine tumor growth by enhancing IFN-γ-producing tumor antigen-specific CD8⁺ T cells. U-V296 exhibited significant synergistic anti-tumor effects with ICIs. In the TME of mice treated with U-V296, the expression of genes associated with M2-like macrophages, Th2 cells (Il4, Retnla, and Irf4), angiogenesis (Pecam1), and fibrosis (Acta2, Loxl2) were significantly decreased, and the Th1/Th2 balance was increased. H₂O₂, an enzymatic product of VAP-1, which promoted the production of IL-4 by mouse Th2 and inhibited IFN-γ by mouse Th1 and human tumor-infiltrating lymphocytes, was decreased in tumors and CD31⁺ tumor vascular endothelial cells in the TMEs of mice treated with VAP-1 inhibitor. TCGA database analysis showed that VAP-1 expression was a negative prognostic factor in human cancers, exhibiting a significant positive correlation with IL-4, IL4R, and IL-13 expression and a negative correlation with IFN-γ expression. These results indicated that VAP-1 is involved in the immunosuppressive TMEs through H₂O₂-associated Th2/M2 conditions and may be an attractive target for the development of combination cancer immunotherapy with ICIs.     

The inhibitory effects of probiotics on colon cancer cells: in vitro and in vivo studies

BackgroundColorectal cancer (CRC) is the most common gastrointestinal malignancy. And probiotics may have the function of preventing colon cancer. The aim of this study was to investigate the inhibitory effects of a probiotic mixture on colorectal cancer and its potential mechanisms.MethodsThe effects of the probiotic mixture on proliferation and metastasis of mouse colon cancer CT26 cells were assessed by probiotics and cells co-culture assay, Cell Counting Kit-8 assay, colony formation assay, wound-healing assay, as well as migration and invasion assays. And CT26 cells were also transplanted into BALB/c mice to construct transplanted tumor animal model. The mice were randomly divided into two groups, control group and probiotic mixture intragastric administration group, after injection 21 days the tumor size and infiltration of immune cells in the tumor or spleen tissues were analyzed by hematoxylin and eosin (HE) and immunohistochemistry (IHC) staining.ResultsThe probiotic mixture significantly inhibited the proliferation, invasion, and migration ability of CT26 cells compare to the control cells (P<0.05). In the animal experiments, the tumor volume of mice that had been fed the probiotic mixture was significantly smaller than that of the control group (P<0.05). Compared with control mice, more apoptotic cells and infiltration of immune cells were showed in the tumor tissues of the mice treated with the probiotic mixture, and an increased number of CD8+ cells in the tumor and spleen tissues but no significant change in tissues.ConclusionsThese results suggested that the probiotic mixture could inhibit the growth of CT26 tumors and induce an immune response in vivo. The probiotic mixture also inhibited the invasion, migration, and proliferation ability of CT26 cells in vitro.     

Temperature as a modulator of the gut microbiome: what are the implications and opportunities for thermal medicine?

Abstract

There is substantial research being conducted on the relationships between the gut microbiome, the immune response and health and disease. Environmental temperature and heat stress are known to modify the gut microbiome. Changes in core temperature have been linked, in multiple phyla, to altered microbiome composition and function. This raises the question of whether local/regional or whole body thermal therapies which target tumors in the abdomen, peritoneal cavity, or pelvis influence the gut microbiome. To date, there is little information on whether thermal therapy exerts positive or negative effects on the microbiome. This is an intriguing question since there is growing interest in the immunological impact of various thermal therapies. The goal of this brief review is to highlight research on how environmental conditions, particularly temperature (internal as well as external temperatures) influences the gut microbiome. Given the potential for temperature shifts to modulate gut microbe function and composition, it is likely that various forms of thermal therapy, including hyperthermic intraperitoneal chemotherapy (HIPEC), deep regional, and whole body hyperthermia influence the microbiome in ways that are currently not appreciated. More research is needed to determine whether thermal therapy induced changes in the microbiome occur, and whether they are beneficial or detrimental to the host. Currently, although approaches to microbiome modification such as dietary intervention, fecal transfer, probiotics and prebiotics are being developed, the potential of temperature manipulation has, as yet, not been explored. Therefore, new research could reveal whether perturbations of the microbiome composition that have negative health consequences (dysbiosis) could be an important target for treatment by thermal medicine.     

Role of intestinal microbiome in American ginseng-mediated colon cancer protection in high fat diet-fed AOM/DSS mice

Objective

Chronic intestinal inflammation is a risk factor for colorectal cancer (CRC) initiation and development. Diets that are rich in Western style fats have been shown to promote CRC. This study was conducted to investigate the role of intestinal microbiome in American ginseng-mediated CRC chemoprevention in a mouse model. The population and diversity of enteric microbiome were evaluated after the ginseng treatment.

Methods

Using an azoxymethane (AOM)/dextran sulfate sodium (DSS)-induced gut inflammation and tumorigenesis mouse model, the effects of oral American ginseng on high fat diet-associated enteric pathology were determined. After establishment of a 16S rRNA illumina library from fecal samples, MiSeq sequencing was carried out to reveal the microbial population. The alpha and beta diversities of microbiome were analyzed.

Results

American ginseng significantly attenuated AOM/DSS-induced colon inflammation and tumorigenesis by reducing the colitis score and colon tumor multiplicity. The MiSeq results showed that the majority of sequences fell into three phyla: Firmicutes, Bacteroidetes and Verrucomicrobia. Further, two significant abundance shifts at the family level, Bacteroidaceae and Porphyromonadaceae, were identified to support ginseng’s anti-colitis and anti-tumor effects. In addition, alpha and beta diversity data demonstrated that ginseng led to a profound recovery from the AOM/DSS-induced dysbiosis in the microbial community.

Conclusion

Our results suggest that the CRC chemopreventive effects of American ginseng are mediated through enteric microbiome population-shift recovery and dysbiosis restoration. Ginseng’s regulation of the microbiome balance contributes to the maintenance of enteric homeostasis.

     

MHC-dependent cytolysis of autologous tumor cells by lymphocytes infiltrating urothelial carcinomas

Tumor-infiltrating lymphocytes (TIL) were grown from 23 urothelial carcinomas. Phenotyping analysis showed that the TIL cultures were mainly CD3⁺. Although CD4⁺ and CD8⁺ T-cell sub-sets were grown in culture, CD4⁺ T-cell sub-sets predominated over CD8⁺ T cells. Immunohistochemical studies performed on 5 tumor specimens confirmed this observation, and indicated that CD4⁺ T cells surrounded the tumor islets, whereas CD8⁺ T lymphocytes were localized among the tumor cells. Five short-term carcinoma cell lines established from these urothelial tumors were used as target cells in cytolysis assays in order to investigate the functional anti-tumor activity of autologous TIL. TIL from 4/5 tumors were lytic and 3 TIL lines displayed MHC-class-I-dependent cytotoxicity directed against autologous tumor cells. CD4⁺ T-cell-depletion experiments performed on TIL line 07 confirmed that CD8⁺ MHC-class-I-dependent CTL were the predominant effectors. Finally, experiments performed on 6 allogeneic urothelial-cancer cell lines matched for HLA-class-I molecules showed that TIL07 exhibited selective lytic activity toward tumor 07. These data indicate that CD8⁺ MHC-class-I-dependent CTL present in urothelial carcinomas are functional and may participate in the anti-tumor immune response. Int. J. Cancer 71:585-594, 1997. © 1997 Wiley-Liss, Inc.     

Gut microbiome in non-alcoholic fatty liver disease associated hepatocellular carcinoma: Current knowledge and potential for therapeutics

Metabolic diseases such as nonalcoholic fatty liver disease (NAFLD) are rising in incidence and are an increasingly common cause of cirrhosis and hepatocellular carcinoma (HCC). The gut microbiome is closely connected to the liver via the portal vein, and has recently been identified as a predictor of liver disease state. Studies in NAFLD, cirrhosis and HCC have identified certain microbial signatures associated with these diseases, with the disease-associated microbiome changes collectively referred to as dysbiosis. The pathophysiologic underpinnings of these observations are an area of ongoing investigation, with current evidence demonstrating that the gut microbiome can influence liver disease and carcinogenesis via effects on intestinal permeability (leaky gut) and activation of the innate immune system. In the innate immune system, pathogen recognition receptors (Toll like receptors) on resident liver cells and macrophages cause liver inflammation, fibrosis, hepatocyte proliferation and reduced antitumor immunity, leading to chronic liver disease and carcinogenesis. Dysbiosis-associated changes include increase in secondary bile acids and reduced expression of FXR (nuclear receptor), which have also been associated with deleterious effects on lipid and carbohydrate metabolism associated with progressive liver disease. Longitudinal experimental and clinical studies are needed in different populations to examine these questions further. The role of therapeutics that modulate the microbiome is an emerging field with experimental studies showing the potential of diet, probiotics, fecal microbiota transplantation and prebiotics in improving liver disease in experimental models. Clinical studies are ongoing with preliminary evidence showing improvement in liver enzymes and steatosis. The microbial profile is different in responders to cancer immunotherapy including liver cancer, but whether or not manipulation of the microbiome can be utilized to affect response is being investigated.     

The Gut Microbiome and Obesity

The gut microbiome consists of trillions of bacteria which play an important role in human metabolism. Animal and human studies have implicated distortion of the normal microbial balance in obesity and metabolic syndrome. Bacteria causing weight gain are thought to induce the expression of genes related to lipid and carbohydrate metabolism thereby leading to greater energy harvest from the diet. There is a large body of evidence demonstrating that alteration in the proportion of Bacteroidetes and Firmicutes leads to the development of obesity, but this has been recently challenged. It is likely that the influence of gut microbiome on obesity is much more complex than simply an imbalance in the proportion of these phyla of bacteria. Modulation of the gut microbiome through diet, pre- and probiotics, antibiotics, surgery, and fecal transplantation has the potential to majorly impact the obesity epidemic.     

Efficacy and safety of probiotics in the treatment of Candida‐associated stomatitis

This study aimed at evaluating the short‐term efficacy and safety of probiotics as an aid in the treatment of Candida‐associated stomatitis in a randomised controlled trial. A total of 65 patients were randomly assigned to receive oral local antifungal agents alone (gargle 2% sodium bicarbonate solution for 30 s, wait 10 min and then apply 2% nystatin paste) or these agents plus local probiotics (the mixture of Bifidobacterium longum, Lactobacillus bulgaricus and Streptococcus thermophilus) three times per day for 4 weeks. Parameters related to hyperaemia, visual analogue scale scores, culture of resting saliva and a lingual dorsum swab and adverse reactions were assessed or recorded in the beginning, middle and end of treatment. Although the baseline characteristics of the participants were similar, both groups showed a significant reduction in pain level and hyperaemia on the tongue mucosa (P = 0.000) after 4‐week application. However, despite the reduction in hyperaemia in the probiotic group, these improvements did not display statistically significant differences. The detection rate of Candida spp. was 100% before treatment and 8.21% in the experimental group and 34.6% in the control group after treatment. The detection rate of Candida spp. decreased (P = 0.000) in both groups and was significantly lower in the probiotic group than the control group (P = 0.038). Other analysed micro‐organisms, including the decreased detection rate for Lactobacillus spp. (P = 0.049) and the increased detection rate for Staphylococcus epidermidis (P = 0.019), did not display consistent change trends in the probiotics group. Compared with conventional antifungal therapies for oral candidiasis, the inclusion of locally administered probiotics helped improve certain clinical conditions and reduced the prevalence of Candida spp., although the impact of probiotics on oral bacterial species remains to be further studied.     

VEGFR2 blockade augments the effects of tyrosine kinase inhibitors by inhibiting angiogenesis and oncogenic signaling in oncogene-driven non-small-cell lung cancers

Molecular agents targeting the epidermal growth factor receptor (EGFR)-, anaplastic lymphoma kinase (ALK)- or c-ros oncogene 1 (ROS1) alterations have revolutionized the treatment of oncogene-driven non-small-cell lung cancer (NSCLC). However, the emergence of acquired resistance remains a significant challenge, limiting the wider clinical success of these molecular targeted therapies. In this study, we investigated the efficacy of various molecular targeted agents, including erlotinib, alectinib, and crizotinib, combined with anti-vascular endothelial growth factor receptor (VEGFR) 2 therapy. The combination of VEGFR2 blockade with molecular targeted agents enhanced the anti-tumor effects of these agents in xenograft mouse models of EGFR-, ALK-, or ROS1-altered NSCLC. The numbers of CD31-positive blood vessels were significantly lower in the tumors of mice treated with an anti-VEGFR2 antibody combined with molecular targeted agents compared with in those of mice treated with molecular targeted agents alone, implying the antiangiogenic effects of VEGFR2 blockade. Additionally, the combination therapies exerted more potent antiproliferative effects in vitro in EGFR-, ALK-, or ROS1-altered NSCLC cells, implying that VEGFR2 inhibition also has direct anti-tumor effects on cancer cells. Furthermore, VEGFR2 expression was induced following exposure to molecular targeted agents, implying the importance of VEGFR2 signaling in NSCLC patients undergoing molecular targeted therapy. In conclusion, VEGFR2 inhibition enhanced the anti-tumor effects of molecular targeted agents in various oncogene-driven NSCLC models, not only by inhibiting tumor angiogenesis but also by exerting direct antiproliferative effects on cancer cells. Hence, combination therapy with anti-VEGFR2 antibodies and molecular targeted agents could serve as a promising treatment strategy for oncogene-driven NSCLC.     

Anti-human lung giant cell cancer (PG) effect of human LAK cellsin vitro and in nude mice

Human LAK cells were prepared by culturing normal human peripheral blood mononuclear cells (PBMC) with or without rIL-2 and assayed for T cell surface markers as well as anti-tumor activity against PGin vitro and in nude mice. Although the percentages of T₃, T₄ and T₈ positive cells in rIL-2-activated cells did not differ significantly from those of control cellsin vitro, the former showed stronger cytotoxicity than control cells to PG tumor cellsin vitro. In vivo, LAK cells completely inhibited the growth of PG tumor in nude mice, whereas PBMC control cells were to be of no effect. The anti-tumor effect of human LAK cells in nude mice may offer a useful model to study the role of human LAK cells against human tumorin vivo.     

Prospective study of oral microbiome and colorectal cancer risk in low-income and African American populations

Oral microbiome may play an important role in cancer pathogenesis. However, no study has prospectively investigated the association of the oral microbiome with subsequent risk of developing colorectal cancer (CRC). We conducted a nested case–control study including 231 incident CRC cases and 462 controls within the Southern Community Cohort Study with 75% of the subjects being African-Americans. The controls were individually matched to cases based on age, ethnic group, smoking, season-of-study enrollment and recruitment method. Oral microbiota were assessed using 16S rRNA gene sequencing in pre-diagnostic mouth rinse samples. Multiple bacterial taxa showed an association with CRC risk at p <0.05. Oral pathogens Treponema denticola and Prevotella intermedia were associated with an increased risk of CRC, with odds ratios (ORs) and 95% confidence intervals (CIs) of 1.76(1.19–2.60) and 1.55(1.08–2.22), respectively, for the individuals carrying these bacteria compared to non-carriers. In the phylum Actinobacteria, Bifidobacteriaceae was more abundant among CRC patients than among controls. In the phylum Bacteroidetes, Prevotella denticola and Prevotella sp. oral taxon 300 were associated with an increased CRC risk, while Prevotella melaninogenica was associated with a decreased risk of CRC. In the phylum Firmicutes, Carnobacteriaceae, Streptococcaceae, Erysipelotrichaceae, Streptococcus, Solobacterium, Streptococcus sp. oral taxon 058 and Solobacterium moorei showed associations with a decreased risk of CRC. Most of these associations were observed among both African- and European-Americans. Most of the associations were not significant after Bonferroni correction for multiple testing, which may be conservative. Our study suggests that the oral microbiome may play a significant role in CRC etiology.     

Immuno-oncology-microbiome axis of gastrointestinal malignancy

Research on the relationship between the microbiome and cancer has been controversial for centuries. Recent works have discovered that the intratumor microbiome is an important component of the tumor microenvironment (TME). Intratumor bacteria, the most studied intratumor microbiome, are mainly localized in tumor cells and immune cells. As the largest bacterial reservoir in human body, the gut microbiome may be one of the sources of the intratumor microbiome in gastrointestinal malignancies. An increasing number of studies have shown that the gut and intratumor microbiome play an important role in regulating the immune tone of tumors. Moreover, it has been recently proposed that the gut and intratumor microbiome can influence tumor progression by modulating host metabolism and the immune and immune tone of the TME, which is defined as the immuno-oncology-microbiome (IOM) axis. The proposal of the IOM axis provides a new target for the tumor microbiome and tumor immunity. This review aims to reveal the mechanism and progress of the gut and intratumor microbiome in gastrointestinal malignancies such as esophageal cancer, gastric cancer, liver cancer, colorectal cancer and pancreatic cancer by exploring the IOM axis. Providing new insights into the research related to gastrointestinal malignancies.     

The biological characteristics of adenovirus-mediated il-18 gene-modified murine colorectal adenocarcinoma cellin vivo andin vitro

Objective: Interleukin 18 (IL-18) is a strong activator of NK cells and promotes the generation of IL-2, IFN-γ and GM-CSF. In the present study, we constructed adenovirus encoding IL-18 gene (AdIL-18), and observed the biological characteristics of IL-18 gene-modified murine colorectal adenocarcinoma cell (CT26)in vivo andin vitro. Methods: Gene modification was mediated by adenovirus. The proliferation of the cells was determined by MTT and IL-18 was assayed by ELISA. The cytotoxicity of NK and CTL was detected by four-hour⁵¹Cr release assay. Results: IL-18 gene modification had no effect on the proliferation and morphology of CT-26 cellsin vitro, but the growth of IL-18-modified CT26 cells was obviously inhibitedin vivo. In addition, although IL-18-modified CT26 cells could form tumor nodulesin vivo as well as LacZ-modified CT26 cells or wild-type CT26 cells, the mean survival time of the mice inoculated with IL-18-modified CT26 cells was significantly prolonged as compared with that of control groups. Thus, the anti-tumor immune responses were induced in the group of mice inoculated with IL-18-modified CT26 cells, which might be related to the activation of NK cells and CTL. However, all the three groups ultimately died of tumor. Conclusion: IL-18-modified CT26 cells could induce the anti-tumor immune responses incompletely, which required other factors for effective anti-tumor responses. Foundation item: This work was supported by National Natural Science Foundation of China (No. 39970689) and Natural Science Foundation of Shanghai (No. 99QB14047). Biography: SONG Wen-gang (1964-), associate professor, majors in cancer immunology, present address: Taishan Medical College.     

Anti-tumor effect and its mechanisms of ursolic acid on human esophageal carcinoma cell Eca-109 <Emphasis Type="Italic">in vivo</Emphasis>

Objective： To investigate the anti-tumor effect and possible mechanisms of ursolic acid on human esophageal carcinoma in vivo. Methods： Atransplanted tumor model by injecting Eca-109 cells into subcutaneous tissue of BALB/c nude mice was established. 40 nude mice bearing tumors were randomly divided into 4 groups and 0.2 ml saline or 0.2 ml ursolic acid （25-100 mg·kg^-1·d^-1） was injected into abdominal cavity respectively once everyday and lasted for fourteen days. The changes of tumor volume were measured continuously and tumor inhibition rate was calculated. The morphological changes of apoptosis were observed by electron microscope. The expressions of COX-2, bcl-2 and Bax protein in transplanted tumors were detected by immunohistochemistry. At last the PGE2 level of transplanted tumors was detected byradioimmunoassay. Results： Treatment of nude mice with 25, 50, or 100 mg·kg^-1·d^-1 of ursolic acid significantly inhibited the growth of the human esophageal carcinoma tumor in nude mice and induced Eca-109 cells apoptosis as demonstrated by electron microscopy analyses. The expressions of COX-2 and bcl-2 in the transplanted tumors were decreased in ursolic acid groups, while the Bax increased. The PGE2 level of transplanted tumors was decreased in ursolic acid groups with adose-relatedmanner. Conclusion： Ursolic acid has anti-tumor effects against human esophageal carcinoma cells in vivo, which are likely mediated via induction of tumor cell apoptosis and inhibition of COX-2 and PGE2.     

In vivo anti-tumor effects of local adoptive transfer of mouse and human cultured lymphoid cells

Mouse and human lymphoid cells were cultivated in the presence of T-cell growth factor (TCGF) and evaluated for their in vivo anti-tumor effect in mice. Cultured spleen cells of normal BALB/c mice or of mice bearing the M109 tumor had a high level of cytotoxic acitivity in vitro against a variety of tumor target cells and had characteristics of natural killer cells. These cultured cells were evaluated for their in vivo cytotoxic activity by admixture with [¹²⁵l]dUrd-labelled M109 tumor cells (2 × 10⁵) at a 30:1 ratio and inoculation of the mixture into the footpads of BALB/c mice. The level of radioactivity remaining in the footpad was determined at various periods following inoculation of radiolabelled tumor cells. The presence of cultured cells in the inocula caused a marked decrease in the footpad radioactivity by 24 h. However, the slope of clearance of the radiolabel then became similar to that in the control mice. The cultured cells delayed, but did not prevent, tumor appearance, and did not influence the subsequent rate of growth of the tumors. The transient effects of the cultured cells contrasted with the more prolonged in vivo effects of alloimmune lymphocytes, and this may be due to their short survival period. Ninety-nine percent of [¹²⁵l]dUrd-labelled cultured mouse lymphoid cells were eliminated within 48 h of i.f.p. inoculation. Cultured human lymphoid cells, initiated from the blood of normal donors, also had a high level of cytotoxic activity in vitro and were evaluated for their in vivo effects by intra-footpad inoculation into nude mice, together with radiolabelled human tumor cell lines, G-ll or HT-29. In vivo cytotoxic activity of the human cultured lymphoid cells correlated with their cytotoxic effect in vitro. These results indicate that cultured mouse and human effector cells have appreciable in vivo cytotoxic effects against tumor cell lines. However, the transient duration of these effects may limit their immunotherapeutic potential.     

Suppression of Anti-tumor CD4+ T Cell Responsiveness in the Tumor-bearing State and Its Recovery in in vitro Culture Free of Tumor Burden

We investigated whether the responsiveness of anti-tumor CD4⁺ T cells suppressed in the tumor-bearing state is reversed in conditions free of tumor burden. Spleen cells from BALB/c mice bearing a syngeneic tumor (CSA1M) 1–3 wk after inoculation with CSA1M cells produced interleukin-2 (IL-2) and IL-4 upon in vitro culture without addition of exogenous tumor antigens. This lymphokine production was achieved through collaboration between anti-CSA1M CD4⁺ T cells and antigen-presenting cells (APC) that had been pulsed with CSA1M tumor antigens in vivo in the tumor-bearing state. However, spleen cells from late (8–10 wk) tumor-bearing stages produced reduced levels of lymphokine production despite the presence of comparable proportions of CD4⁺ T cells. Because APC in these cell populations exhibited enhanced capacities to present tumor antigens, reduced responsiveness was ascribed to the dysfunction of CD4⁺ T cells themselves. When spleen cells from early tumor-bearing mice were preincubated for 1–2 days and recultured in fresh medium, the magnitude of lymphokine production by these cells was not changed. In contrast, the same protocol of preincubation and reculture for cells from late tumor-bearing mice resulted in the recovery of anti-tumor lymphokine-producing capacity. The recovered capacity was comparable to or slightly higher than that expressed by cells from early tumor-bearing stages. Since the CD4⁺ T cell content did not significantly differ before and after preincubation, enhanced lymphokine production was due to the recovered responsiveness of anti-tumor CD4⁺ helper T cells. The recovery of anti-tumor responsiveness was also induced in vivo by tumor removal at the late tumor-bearing stage: spleen cells from mice 2–4 wk after tumor resection efficiently produced IL-2 and IL-4. These results indicate that the immunodysfunction of anti-tumor CD4⁺ T cells induced in the tumor-bearing state is reversible because release from tumor burden either by preincubation in vitro or by tumor removal in vivo results in almost complete recovery of the potent anti-tumor responsiveness initially expressed.     

Interleukin-6 functions as an autocrine growth factor in human bladder carcinoma cell lines in vitro

Interleukin (IL)-6 is reported to function as a growth factor for renal and prostatic carcinomas. We conducted the present study to define the role of IL-6 in the growth of normal and neoplastic urothelial cells. Human bladder carcinoma cell lines (253J, RT4 and T24) and primary cultured human urothelial cells derived from normal ureters were used. Recombinant human IL-6 stimulated the growth of bladder carcinoma cell lines far better than that of normal urothelial cells (p < 0.001). All carcinoma cell lines tested produced and released IL-6, whereas normal urothelial cells did so only at marginal levels. Furthermore, treatment with lipopolysaccharide derived from Escherichia coli, tumor necrosis factor-α or IL-1 increased IL-6 secretion by bladder carcinoma cell lines but not by normal urothelial cells. Growth of bladder carcinoma cells was significantly inhibited by anti-IL-6 neutralizing antibody or the anti-sense oligonucleotide for IL-6 cDNA. We conclude that IL-6 functions as an autocrine growth factor for bladder carcinoma cells but not for normal urothelial cells and that it may be a factor accounting for the marked enhancement of inflammation-associated bladder carcinogenesis and tumor growth. Int. J. Cancer 72:149–154, 1997. © 1997 Wiley-Liss Inc.     

High‐mobility group box‐1 contributes tumor angiogenesis under interleukin‐8 mediation during gastric cancer progression

Many soluble factors are involved in tumor angiogenesis. Thus, it is valuable to identify novel soluble factors for effective control of tumor angiogenesis in gastric cancer (GC). We investigated the role of extracellular high‐mobility group box‐1 (HMGB1) and its associated soluble factors in the tumor angiogenesis of GC. Clinically, we measured serum levels of HMGB1 and GC‐associated cytokines/chemokines using GC serum samples (n = 120), and calculated microvessel density (MVD) by CD34 immunostaining using human GC tissues (n = 27). Then we analyzed the correlation of serum HMGB1 levels with MVD or that with cytokine/chemokine levels by linear regression. As in vitro angiogenesis assay for HMGB1, HUVEC migration and capillary tube formation assay were carried out using different histological types of human GC cells (N87 and KATOIII). CD34‐positive microvessels were detected from early GC, but MVD increased according to GC stages, and were closely correlated with serum HMGB1 levels (R = 0.608, P = 0.01). The HUVECs cultured in conditioned media derived from rhHMGB1‐treated or HMGB1‐TF GC cells showed remarkably enhanced migration and tube formation activities. These effects were abrogated by anti‐HMGB1 antibody or HMGB1 siRNA in both N87 and KATOIII cells (all P < 0.05). Among tested cytokines/chemokines, interleukin‐8 (IL‐8) was the most remarkable cytokine correlated with serum HMGB1 (P < 0.001), and enhanced HUVEC migration and tube formation activities by rhHMGB1 or HMGB1‐TF were significantly reversed by IL‐8 inhibition. These results indicate overexpressed HMGB1 contributes to tumor angiogenesis through IL‐8 mediation, and combined targeting of HMGB1 and IL‐8 can control tumor angiogenesis in GC.     

Immunosuppressive activity is attenuated by Astragalus polysaccharides through remodeling the gut microenvironment in melanoma mice

Astragalus polysaccharides (APS), the main effective component of Astragalus membranaceus, can inhibit tumor growth, but the underlying mechanisms remain unclear. Previous studies have suggested that APS can regulate the gut microenvironment, including the gut microbiota and fecal metabolites. In this work, our results showed that APS could control tumor growth in melanoma-bearing mice. It could reduce the number of myeloid-derived suppressor cells (MDSC), as well as the expression of MDSC-related molecule Arg-1 and cytokines IL-10 and TGF-β, so that CD8⁺ T cells could kill tumor cells more effectively. However, while APS were administered with an antibiotic cocktail (ABX), MDSC could not be reduced, and the growth rate of tumors was accelerated. Consistent with the changes in MDSC, the serum levels of IL-6 and IL-1β were lowest in the APS group. Meanwhile, we found that fecal suspension from mice in the APS group could also reduce the number of MDSC in tumor tissues. These results revealed that APS regulated the immune function in tumor-bearing mice through remodeling the gut microbiota. Next, we focused on the results of 16S rRNA, which showed that APS significantly regulated most microorganisms, such as Bifidobacterium pseudolongum, Lactobacillus johnsonii and Lactobacillus. According to the Spearman analysis, the changes in abundance of these microorganisms were related to the increase of metabolites like glutamate and creatine, which could control tumor growth. The present study demonstrates that APS attenuate the immunosuppressive activity of MDSC in melanoma-bearing mice by remodeling the gut microbiota and fecal metabolites. Our findings reveal the therapeutic potential of APS to control tumor growth.