Prognostic significance of NY-ESO-1 antigen and PIGR expression in esophageal tumors of CHP-NY-ESO-1-vaccinated patients as adjuvant therapy.

The aim of this study was to determine the efficacy and the biomarkers of the CHP-NY-ESO-1 vaccine complexed with full-length NY-ESO-1 protein and a cholesteryl pullulan (CHP) in patients with esophageal squamous cell carcinoma (ESCC) after surgery. We conducted a randomized phase II trial. Fifty-four patients with NY-ESO-1-expressing ESCC who underwent radical surgery following cisplatin/5-fluorouracil-based neoadjuvant chemotherapy were assigned to receive either CHP-NY-ESO-1 vaccination or observation as control. Six doses of CHP-NY-ESO-1 were administered subcutaneously once every two weeks, followed by nine more doses once every four weeks. The endpoints were disease-free survival (DFS) and safety. Exploratory analysis of tumor tissues using gene-expression profiles was also performed to seek the biomarker. As there were no serious adverse events in 27 vaccinated patients, we verified the safety of the vaccine. DFS in 2 years were 56.0% and 58.3% in the vaccine arm and in the control, respectively. Twenty-four of 25 patients showed NY-ESO-1-specific IgG responses after vaccination. Analysis of intra-cohort correlations among vaccinated patients revealed that 5% or greater expression of NY-ESO-1 was a favorable factor. Comprehensive analysis of gene expression profiles revealed that the expression of the gene encoding polymeric immunoglobulin receptor (PIGR) in tumors had a significantly favorable impact on outcomes in the vaccinated cohort. The high PIGR-expressing tumors that had higher NY-ESO-1-specific IgA response tended to have favorable prognosis. These results suggest that PIGR would play a major role in tumor immunity in an antigen-specific manner during NY-ESO-1 vaccinations. The IgA response may be relevant.

Safety and antibody immune response of CHP-NY-ESO-1 vaccine combined with poly-ICLC in advanced or recurrent esophageal cancer patients.

The nanoparticle complex of cholesteryl pullulan (CHP) and NY-ESO-1 antigen protein (CHP-NY-ESO-1) presents multiple epitope peptides to MHC class I and II pathways, leading to CD8+ and CD4+ T cell responses. Poly-ICLC is a synthetic, double-stranded RNA, an agonist of toll-like receptor (TLR)-3, and a cytoplasmic receptor of melanoma differentiation-associated gene (MDA)-5. It should be a suitable immune adjuvant of cancer vaccine to overcome the inhibitory tumor microenvironment. We conducted a phase 1 clinical trial of CHP-NY-ESO-1 with poly-ICLC in patients with advanced or recurrent esophageal cancer. CHP-NY-ESO-1/poly-ICLC (µg/mg) was administered at a dose of 200/0.5 or 200/1.0 (cohorts 1 and 2, respectively) every 2 weeks for a total of six doses. The primary endpoints were safety and immune response. The secondary endpoint was tumor response. In total, 16 patients were enrolled, and six patients in each cohort completed the trial. The most common adverse event (AE) was injection site skin reaction (86.7%). No grade 3 or higher drug-related AEs were observed. No tumor responses were observed, and three patients (30%) had stable disease. The immune response was comparable between the two cohorts, and all patients (100%) achieved antibody responses with a median of 2.5 vaccinations. Comparing CHP-NY-ESO-1 alone to the poly-ICLC combination, all patients in both groups exhibited antibody responses, but the titers were higher in the combination group. In a mouse model, adding anti-PD-1 antibody to the combination of CHP-NY-ESO-1/poly-ICLC suppressed the growth of NY-ESO-1-expressing tumors. Combining the vaccine with PD-1 blockade holds promise in human trials.

Phase I clinical trial of adoptive transfer of expanded natural killer cells in combination with IgG1 antibody in patients with gastric or colorectal cancer.

Natural killer (NK) cells exhibit strong cytotoxic activity against tumor cells without prior sensitization, and have the potential to exert antibody-dependent cellular cytotoxicity (ADCC). In this clinical trial, we examined the safety and efficacy of the use of NK cells, generated using a novel expansion system, in combination with IgG1 antibodies for the treatment of advanced gastric or colorectal cancers. Treatment consisted of trastuzumab- or cetuximab-based chemotherapy, plus adoptive NK cell therapy. For administration of expanded NK cells, dose escalation with a sequential 3 + 3 design was performed in three steps, at doses of 0.5 × 109 , 1.0 × 109 , and 2.0 × 109 cells/injection (N = 9). After 3 days of IgG1 antibody administration, patients were infused with expanded NK cells three times at triweekly intervals. NK cell populations expanded with our system were confirmed as being enriched in NK cells (median 92.9%) with high expression of NKG2D (97.6%) and CD16 (69.6%). The combination therapy was very well tolerated with no severe adverse events. Among six evaluable patients, four presented stable disease (SD) and two presented progressive disease. Of the four SD patients, three showed an overall decrease in tumor size after combination therapy. Immune monitoring suggested that combination therapy enhanced whole blood IFN-γ production and reduced peripheral regulatory T cells (Tregs). In conclusion, this phase I trial provides evidence of good tolerability, induction of Th1 immune responses, and preliminary anti-tumor activity for this combination therapy, in patients with advanced gastric and colorectal cancer that have received previous therapy.

Combination Gemcitabine and WT1 Peptide Vaccination Improves Progression-Free Survival in Advanced Pancreatic Ductal Adenocarcinoma: A Phase II Randomized Study.

We investigated the efficacy of a Wilms' tumor gene 1 (WT1) vaccine combined with gemcitabine (GEMWT1) and compared it with gemcitabine (GEM) monotherapy for advanced pancreatic ductal adenocarcinoma (PDAC) in a randomized phase II study. We randomly assigned HLA-A*02:01- or HLA-A*24:02-positive patients with advanced PDAC to receive GEMWT1 or GEM. We assessed WT1-specific immune responses via delayed-type hypersensitivity (DTH) to the WT1 peptide and a tetramer assay to detect WT1-specific cytotoxic T lymphocytes (WT1-CTL). Of 91 patients enrolled, 85 were evaluable (GEMWT1: n = 42; GEM: n = 43). GEMWT1 prolonged progression-free survival [PFS; hazard ratio (HR), 0.66; P = 0.084] and improved overall survival rate at 1 year (1-year OS%; GEMWT1: 35.7%; GEM: 20.9%). However, the difference in OS was not significant (HR: 0.82; P = 0.363). These effects were particularly evident in metastatic PDAC (PFS: HR 0.51, P = 0.0017; 1-year OS%: GEMWT1 27.3%; GEM 11.8%). The combination was well tolerated, with no unexpected serious adverse events. In patients with metastatic PDAC, PFS in the DTH-positive GEMWT1 group was significantly prolonged, with a better HR of 0.27 compared with the GEM group, whereas PFS in the DTH-negative GEMWT1 group was similar to that in the GEM group (HR 0.86; P = 0.001). DTH positivity was associated with an increase in WT1-CTLs induced by the WT1 vaccine. GEM plus the WT1 vaccine prolonged PFS and may improve 1-year OS% in advanced PDAC. These clinical effects were associated with the induction of WT1-specific immune responses. Cancer Immunol Res; 6(3); 320-31. ©2018 AACR.

The amino acid-rich elemental diet Elental® preserves lean body mass during chemo- or chemoradiotherapy for esophageal cancer.

Chemo (chemoradio) therapy can induce oral mucositis and change body composition in patients with esophageal cancer. The impact of the amino acid-rich elemental diet Elental® on oral mucositis and changes in body composition during chemo (chemoradio) therapy is unclear. Thus, the purpose of the present study was to examine the preventive effects of Elental on oral mucositis and sarcopenia progression during chemo (chemoradio) therapy for esophageal cancer. Patients were randomized to receive either azulene oral rinse (Arm 1) or Elental (Arm 2) during the treatment cycle (4 weeks). The incidence of oral mucositis and other adverse events was evaluated weekly. Body composition pre- and post-treatment cycle was measured by bioelectric impedance analysis. Thirty­three patients (17 azulene and 16 Elental) completed the study, and the groups were well matched. Elental tended to reduce the incidence of oral mucositis (Arm 1, 23.5% and Arm 2, 12.5%), but there was no statistically significant difference between the groups. The average body mass index (BMI) and body fat mass decreased significantly in both groups after the treatments. Lean body mass (LBM) was reduced in Arm 1, but was increased in Arm 2 after the treatment; the relative change of LBM after the treatment was significant between Arm 1 and Arm 2 (P=0.007). This study revealed that Elental nutrition could counteract sarcopenia development during chemoradiotherapy for esophageal cancer. These properties may lead to improvement of the quality of life and clinical outcome of esophageal cancer patients treated with chemo (chemoradio) therapy (Clinical Trial Registry ID: UMIN 000007960).

Phase I clinical trial of autologous NK cell therapy using novel expansion method in patients with advanced digestive cancer.

BACKGROUND: NK cells can destroy tumor cells without prior sensitization or immunization. Tumors often lose expression of MHC molecules and/or antigens. However, NK cells can lyse tumor cells in a non-MHC-restricted manner and independent of the expression of tumor-associated antigens. NK cells are therefore considered ideal for adoptive cancer immunotherapy; however the difficulty of obtaining large numbers of fully functional NK cells that are safe to administer deters its clinical use. This phase I clinical trial seeks to address this obstacle by first developing a novel system that expands large numbers of highly activated clinical grade NK cells, and second, determining if these cells are safe in a mono-treatment so they can be combined with other reagents in the next round of clinical trials. METHODS: Patients with unresectable, locally advanced and/or metastatic digestive cancer who did not succeed with standard therapy were enrolled. NK cells were expanded ex vivo by stimulating PBMCs with OK432, IL-2, and modified FN-CH296 induced T cells. Patients were administered autologous natural killer cell three times weekly via intravenous infusions in a dose-escalating manner (dose 0.5 × 10(9), 1.0 × 10(9), 2.0 × 10(9) cells/injection, three patients/one cohort). RESULTS: Total cell population had a median expansion of 586-fold (range 95-1102), with a significantly pure (90.96 %) NK cell population. Consequently, NK cells were expanded to approximately 4720-fold (range 1372-14,116) with cells being highly lytic in vitro and strongly expressing functional markers such as NKG2D and CD16. This NK cell therapy was very well tolerated with no severe adverse events. Although no clinical responses were observed, cytotoxicity of peripheral blood was elevated approximately twofolds up to 4 weeks post the last transfer. CONCLUSION: We successfully generated large numbers of activated NK cells from small quantities of blood without prior purification of the cells. We also determined that the expanded cells were safe to administer in a monotherapy and are suitable for the next round of clinical trials where their efficacy will be tested combined with other reagents. TRIAL REGISTRATION: UMIN UMIN000007527.

Tumor inoculation site affects the development of cancer cachexia and muscle wasting.

The phenotype and severity of cancer cachexia differ among tumor types and metastatic site in individual patients. In this study, we evaluated if differences in tumor microenvironment would affect the development of cancer cachexia in a murine model, and demonstrated that body weight, adipose tissue and gastrocnemius muscle decreased in tumor-bearing mice. Interestingly, a reduction in heart weight was observed in the intraperitoneal tumor group but not in the subcutaneous group. We evaluated 23 circulating cytokines and members of the TGF-ß family, and found that levels of IL-6, TNF-α and activin A increased in both groups of tumor-bearing mice. Eotaxin and G-CSF levels in the intraperitoneal tumor group were higher than in the subcutaneous group. Atrogin 1 and MuRF1 mRNA expressions in the gastrocnemius muscle increased significantly in both groups of tumor-bearing mice, however, in the myocardium, expression of these mRNAs increased in the intraperitoneal group but not in subcutaneous group. Based on these results, we believe that differences in microenvironment where tumor cells develop can affect the progression and phenotype of cancer cachexia through alterations in various circulating factors derived from the tumor microenvironment.

Hypersensitivity Reactions to Oxaliplatin: Identifying the Risk Factors and Judging the Efficacy of a Desensitization Protocol.

PURPOSE: We examined the clinical data of patients treated with oxaliplatin to determine the risk factors of oxaliplatin-related hypersensitivity reaction (HSR). In addition, we evaluated the efficacy of rechallenging patients with HSRs with oxaliplatin using prophylactic agents or desensitization procedures. METHODS: This study consisted of 162 patients with colorectal cancer (88 men and 74 women) who were treated consecutively at the outpatient chemotherapy department at University Hospital, Kyoto Prefectural University of Medicine. Patients underwent chemotherapy, including oxaliplatin, between March 2006 and June 2012. We analyzed the patients' clinical backgrounds (eg, age, sex, performance status, disease stage, and allergic history) to uncover any connections to the development of HSR to oxaliplatin. In addition, we rechallenged 10 patients who had oxaliplatin-related HSR using prophylactic agents or desensitization procedures. FINDINGS: Of 162 patients, 28 (17.2%) developed oxaliplatin-related HSRs (16, 2, 9 and 1 patient had grade 1, 2, 3, and 4 HSRs, respectively). The total cumulative dose of oxaliplatin at the onset of the HSR was 301 to 1126 mg/m(2) (median, 582 mg/m(2)), and the first reactions developed in these patients after 5 to 17 infusions of oxaliplatin (median, 8 infusions). Logistic regression analysis indicated that sex (male: odds ratio = 3.624; 95% CI, 1.181-11.122; P = 0.024) and eosinophil count in peripheral blood (odds ratio = 35.118; 95% CI, 1.058-1166.007; P = 0.046) were independent variables for oxaliplatin-related HSRs. Rechallenging patients with prophylactic agents was successful in 2 (28.6%) of 7 patients who successfully completed their treatment. On the other hand, all 3 patients rechallenged with oxaliplatin using a desensitization protocol successfully completed their treatment without new HSRs. IMPLICATIONS: In this retrospective study, we observed that being male and having higher counts of peripheral eosinophil could be predictors for HSR to oxaliplatin. In addition, this study confirms that oxaliplatin desensitization protocol allows patients who developed HSRs to continue with their treatment. However, the optimum desensitization protocol for oxaliplatin administration in terms of tolerability and efficacy needs to be defined.

Adoptive Transfer of MAGE-A4 T-cell Receptor Gene-Transduced Lymphocytes in Patients with Recurrent Esophageal Cancer.

PURPOSE: Preparative lymphodepletion, the temporal ablation of the immune system, has been reported to promote persistence of transferred cells along with increased rates of tumor regression in patients treated with adoptive T-cell therapy. However, it remains unclear whether lymphodepletion is indispensable for immunotherapy with T-cell receptor (TCR) gene-engineered T cells. EXPERIMENTAL DESIGN: We conducted a first-in-man clinical trial of TCR gene-transduced T-cell transfer in patients with recurrent MAGE-A4-expressing esophageal cancer. The patients were given sequential MAGE-A4 peptide vaccinations. The regimen included neither lymphocyte-depleting conditioning nor administration of IL2. Ten patients, divided into 3 dose cohorts, received T-cell transfer. RESULTS: TCR-transduced cells were detected in the peripheral blood for 1 month at levels proportional to the dose administered, and in 5 patients they persisted for more than 5 months. The persisting cells maintained ex vivo antigen-specific tumor reactivity. Despite the long persistence of the transferred T cells, 7 patients exhibited tumor progression within 2 months after the treatment. Three patients who had minimal tumor lesions at baseline survived for more than 27 months. CONCLUSIONS: These results suggest that TCR-engineered T cells created by relatively short-duration in vitro culture of polyclonal lymphocytes in peripheral blood retained the capacity to survive in a host. The discordance between T-cell survival and tumor regression suggests that multiple mechanisms underlie the benefits of preparative lymphodepletion in adoptive T-cell therapy.

Cytotoxic T lymphocyte-associated antigen 4 inhibition increases the antitumor activity of adoptive T-cell therapy when carried out with naïve rather than differentiated T cells.

Although treatment with an antibody against cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) combined with multiple therapeutic interventions has been explored, the effect of combination therapy with CTLA-4 inhibition and adoptive T-cell therapy has not been determined. In the present study, our aim was to determine whether CTLA-4 inhibition, combined with adoptive transfer of T cells at different stages of differentiation, exhibits synergistic antitumor effects in a murine colon cancer model. Mice bearing subcutaneous tumors were administered adoptive T-cell transfer of CD62Lhigh or CD62Llow cells combined with an anti-CTLA-4 antibody (α-CTLA-4) or control immunoglobulin G. Subcutaneous tumors were harvested, and the antitumor effects and helper T-cell polarization were analyzed. CTLA-4 inhibition combined with CD62Lhigh cell administration showed the strongest antitumor effect. Combination therapy increased the number of CD3+ cells within the tumor. Moreover, CTLA-4 inhibition induced polarization of T cells infiltrating the tumor toward the T helper 1 lineage, and suppressed the frequency of regulatory T cells within the tumor, particularly in combination with CD62Lhigh T-cell transfer. This is the first report demonstrating that the efficacy of α-CTLA-4 and adoptive T-cell transfer combination therapy depends on the state of differentiation of the transferred T cells. Our data support the notion that a combination of α-CTLA-4 and adoptive T-cell transfer containing an abundance of naïve phenotype cells could potentially exert antitumor effects in a clinical setting.

The inhibitory effect of heat treatment against epithelial-mesenchymal transition (EMT) in human pancreatic adenocarcinoma cell lines.

Epithelial-mesenchymal transition (EMT) plays a crucial role in cancer metastasis. In this study, we evaluated the effect of heat treatment on tumor growth factor-ß1 (TGF-ß1)-induced EMT in pancreatic cancer cells and tried to ascertain the mechanism related to any observed effects. Human pancreatic cancer cell lines (BxPC-3, PANC-1 and MIAPaCa-2) were stimulated by TGF-ß1, and evaluated for morphological changes using immunofluorescence and EMT-related factors (i.e., E-cadherin, Vimentin, Snail or ZEB-1) using RT-PCR. To examine the effect of heat on EMT, the cancer cells were heat-treated at 43°C for 1 h then stimulated with TGF-ß1. We then evaluated whether or not heat treatment changed the expression of EMT-related factors and cell migration and also whether Smad activation was inhibited in TGF-ß signaling. After being treated with TGF-ß1, pancreatic cancer cells resulted in EMT and cell migration was enhanced. Heat treatment inhibited TGF-ß1-induced changes in morphology, inhibited the expression of EMT-related factors, and attenuated TGF-ß1-induced migration in pancreatic cancer cells. Additionally, we observed that heat treatment blocked TGF-ß1-induced phosphorylation of Smad2 in PANC-1 cells. Our results suggest that heat treatment can suppress TGF-ß1-induced EMT and opens the possibility of a new therapeutic use of hyperthermia as a potential treatment for cancer metastasis.

Phase I clinical trial of fibronectin CH296-stimulated T cell therapy in patients with advanced cancer.

BACKGROUND: Previous studies have demonstrated that less-differentiated T cells are ideal for adoptive T cell transfer therapy (ACT) and that fibronectin CH296 (FN-CH296) together with anti-CD3 resulted in cultured cells that contain higher amounts of less-differentiated T cells. In this phase I clinical trial, we build on these prior results by assessing the safety and efficacy of FN-CH296 stimulated T cell therapy in patients with advanced cancer. METHODS: Patients underwent fibronectin CH296-stimulated T cell therapy up to six times every two weeks and the safety and antitumor activity of the ACT were assessed. In order to determine immune function, whole blood cytokine levels and the number of peripheral regulatory T cells were analyzed prior to ACT and during the follow up. RESULTS: Transferred cells contained numerous less-differentiated T cells greatly represented by CD27+CD45RA+ or CD28+CD45RA+ cell, which accounted for approximately 65% and 70% of the total, respectively. No ACT related severe or unexpected toxicities were observed. The response rate among patients was 22.2% and the disease control rate was 66.7%. CONCLUSIONS: The results obtained in this phase I trial, indicate that FN-CH296 stimulated T cell therapy was very well tolerated with a level of efficacy that is quite promising. We also surmise that expanding T cell using CH296 is a method that can be applied to other T- cell-based therapies. TRIAL REGISTRATION: UMIN UMIN000001835.

Ability of a novel blue laser imaging system for the diagnosis of colorectal polyps.

BACKGROUND: A new endoscope system with a laser light source, blue laser imaging (BLI), has been developed by Fujifilm that allows for narrow-band light observation. The aim of the present study was to evaluate the utility of BLI for the diagnosis of colorectal polyps. METHODS: We retrospectively analyzed 314 colorectal polyps that were examined with BLI observation at Kyoto Prefectural University of Medicine between September 2011 and January 2013. The surface and vascular patterns of polyps detected by published narrow-band imaging magnification: Hiroshima classification were used. Correlations were determined between the classifications and the histopathological diagnoses. Additionally, the ability of BLI without magnification to differentiate between neoplastic or non-neoplastic polyps was analyzed. RESULTS: A total of 41 hyperplastic polyps, 168 adenomas, 80 intramucosal cancer, 11 shallowly invaded submucosal cancer, and 14 deeply invaded submucosal cancer were analyzed.Hyperplastic polyp was observed in 100% of Type A lesions (39 lesions), adenoma was observed in 89.3% of Type B lesions (159 lesions), intramucosal cancer and shallowly invaded submucosal cancer was observed in 69.6% of Type C1 (92 lesions) and in 84.6% of Type C2 (13 lesions), and deeply invaded submucosal cancer was observed in 81.8% of Type C3 lesions (11 lesions). The overall diagnostic accuracy of BLI with magnification was 84.3%. Additionally, the diagnostic accuracy of BLI without magnification for differentiating between neoplastic and non-neoplastic polyps <10 mm in diameter was 95.2%, which was greater than that of white light (83.2%). CONCLUSION: BLI was useful for the diagnosis of colorectal polyps.

Prediction of Large Esophageal Variceal Bleeding and Subsequent Mortality.

BACKGROUND/AIMS: Ruptured esophageal varices (EV) are commonly associated with bleeding in patients with portal hypertension. As previous studies have been designed to include all viral cirrhosis patients, including those with small varices, there are no definitive reports of the risk factors for large EV (LEV) bleeding and subsequent mortality. In this study we sought to investigate these risk factors. METHODOLOGY: 16 patients with initial episodes of LEV bleeding and 56 patients who had undergone initial preventive treatment for LEV were examined. The Child-Pugh score, MELD score, and their component serological factors were examined. RESULTS: The Child-Pugh score and the MELD score were significantly lower for preventive cases. Serum albumin concentration was significantly lower in bleeding cases, as determined by univariable and multivariable analyses. In initial LEV bleeding cases, higher MELD scores and Child-Pugh scores, and lower serum albumin concentration were associated with significantly higher one-year mortality. CONCLUSIONS: These results indicate that control of the MELD score, the Child-Pugh score, and serum albumin concentration is key to improving the prognosis of patients with LEV. Especially, control of serum albumin concentration may be critical for the prevention of initial episodes of LEV bleeding and improving subsequent mortality.

Efficacy of goshajinkigan for oxaliplatin-induced peripheral neuropathy in colorectal cancer patients.

Objective. To evaluate the efficacy of Goshajinkigan for oxaliplatin-induced peripheral neuropathy in colorectal cancer patients. Patients. Colorectal cancer patients (N = 29) who received ≥4 weeks of Goshajinkigan for oxaliplatin-induced peripheral neuropathy during chemotherapy at Kyoto Prefectural University of Medicine were (Goshajinkigan group) compared to 44 patients who had not received Goshajinkigan during the same period (non-Goshajinkigan group). Main Outcome Measures. The effect of Goshajinkigan was graded as curative, effective, stabilizing, or deleterious. The relationships between the grade of peripheral neuropathy and the dose of oxaliplatin in the Goshajinkigan and non-Goshajinkigan groups were evaluated. Results. The effect of Goshajinkigan on peripheral neuropathy in the Goshajinkigan group was curative, effective, stabilizing, and deleterious in 3.4, 20.7, 69.0, and 6.9% of patients, compared to the effect in the non-Goshajinkigan group (4.5, 15.9, 45.5, and 34.1%). The ratio of deleterious effects was significantly different between these two groups (P = 0.04). A Kaplan-Meier analysis in relation to the cumulative dose of oxaliplatin showed that the incidence of grade 3 peripheral neuropathy tended to be less in the Goshajinkigan group (P = 0.05). There were no significant differences in time to treatment failure and severe adverse events between these two groups. Conclusions. Goshajinkigan prevented exacerbation of oxaliplatin-induced peripheral neuropathy. This trial is registered with UMIN000009956.

Anoxia/reoxygenation induces epithelial-mesenchymal transition in human colon cancer cell lines.

Epithelial-mesenchymal transition (EMT) is considered to be a crucial event in the development of cancer metastasis. Anoxia/reoxygenation (A/R) is known to occur in cancer tissues due to angiogenesis and changes in tissue pressure that occur during tumor growth. We investigated whether A/R induces EMT in the human colon cancer cell line HT-29. Colon cancer cells were exposed to anoxia (2 h) followed by reoxygenation (4-22 h) and evaluated for EMT changes using immunofluorescence and western blot analyses. We also investigated the expression of EMT-related transcription factors (Snail and ZEB1) using RT-PCR and evaluated the expression of NF-κB using ELISA. To determine whether NF-κB is involved in A/R-induced EMT, HT-29 cells were treated with proteasome inhibitors. Colon cancer cells exposed to A/R underwent EMT morphological changes; the cancer cells acquired a spindle-shaped phenotype. The expression of E-cadherin on the cell surface and the total amount of E-cadherin proteins were reduced after A/R. The expression of EMT-related transcription factors (Snail, ZEB1) was increased after A/R. Pretreatment with proteasome inhibitors significantly attenuated the downregulation of E-cadherin induced by A/R. These results indicate that A/R induces EMT in human colon cancer cells through an NF-κB-dependent transcriptional pathway.

BTB and CNC homolog 1 (Bach1) deficiency ameliorates TNBS colitis in mice: role of M2 macrophages and heme oxygenase-1.

BACKGROUND: BTB and CNC homolog 1 (Bach1) is a transcriptional repressor of heme oxygenase-1 (HO-1), which plays an important role in the protection of cells and tissues against acute and chronic inflammation. However, the role of Bach1 in the gastrointestinal mucosal defense system remains little understood. HO-1 supports the suppression of experimental colitis and localizes mainly in macrophages in colonic mucosa. This study was undertaken to elucidate the Bach1/HO-1 system's effects on the pathogenesis of experimental colitis. METHODS: This study used C57BL/6 (wild-type) and homozygous Bach1-deficient C57BL/6 mice in which colonic damage was induced by the administration of an enema of 2,4,6-trinitrobenzene sulfonic acid (TNBS). Subsequently, they were evaluated macroscopically, histologically, and biochemically. Peritoneal macrophages from the respective mice were isolated and analyzed. Then, wild-type mice were injected with peritoneal macrophages from the respective mice. Acute colitis was induced similarly. RESULTS: TNBS-induced colitis was inhibited in Bach1-deficient mice. TNBS administration increased the expression of HO-1 messenger RNA and protein in colonic mucosa in Bach1-deficient mice. The expression of HO-1 mainly localized in F4/80-immunopositive and CD11b-immunopositive macrophages. Isolated peritoneal macrophages from Bach1-deficient mice highly expressed HO-1 and also manifested M2 macrophage markers, such as Arginase-1, Fizz-1, Ym1, and MRC1. Furthermore, TNBS-induced colitis was inhibited by the transfer of Bach1-deficient macrophages into wild-type mice. CONCLUSIONS: Deficiency of Bach1 ameliorated TNBS-induced colitis. Bach1-deficient macrophages played a key role in protection against colitis. Targeting of this mechanism is applicable to cell therapy for human inflammatory bowel disease.

Whole blood interferon-γ levels predict the therapeutic effects of adoptive T-cell therapy in patients with advanced pancreatic cancer.

A core challenge in administering immune-based treatments for cancer is the establishment of easily accessible immunological assays that can predict patients' clinical responses to immunotherapy. In this study, our aim was to predict the therapeutic effects of adoptive T-cell therapy in patients with advanced pancreatic cancer. To do this, we evaluated whole blood cytokine levels and peripheral regulatory T cells (Tregs) in 46 patients with unresectable or recurrent pancreatic cancer who received adoptive T-cell therapy at 2-week intervals. To test immune function, venous blood was obtained from patients before the start of therapy and 2 weeks after the 4th treatment. Whole blood interferon (IFN)-α levels (after stimulation with the Sendai virus) were evaluated, as well as the levels of 9 cytokines stimulated with phytohemagglutinin [interleukin (IL)-2, IL-4, IL-5, IL-10, IL-12(p70), IL-13, tumor necrosis factor-α, IFN-γ, and granulocyte-monocyte colony-stimulating factor]. Peripheral Tregs were analyzed by flow cytometry. Using the obtained data, we then observed the relationship between these immunological parameters and clinical outcome of patients. We found that the whole blood production of IFN-γ, IL-2, IL-4, IL-5 and IL-13 significantly increased after adoptive T-cell therapy, whereas the number of peripheral Tregs did not change. Multivariate Cox proportional hazards analyses indicated that the number of peripheral Tregs before receiving adoptive T-cell therapy and the change in IFN-γ levels after adoptive T-cell therapy were independent variables predicting overall survival. The findings of this study indicate that the assay of whole blood IFN-γ production offers promise for evaluating the clinical response of patients to cancer immunotherapy.

Effects of anticancer agents on cell viability, proliferative activity and cytokine production of peripheral blood mononuclear cells.

We investigated the effects of anticancer agents on peripheral blood mononuclear cells for the purpose of providing data to support new translational chemoimmunotherapy regimens. Peripheral-blood mononuclear cells were treated with one of four anticancer agents (5-fluorouracil, irinotecan, cisplatin, and gemcitabine) for 2 h, after which cell viability was determined. For assessment of effects of each drug on proliferation and cytokine production, cells were stimulated with phytohemagglutinin for 48 h. As a result, the anticancer agents did not affect cell viability. Cell proliferation was unaffected by 5-fluorouracil and irinotecan but inhibited by cisplatin and gemcitabine. Treatment with gemcitabine enhanced the production of IFN-γ and decreased the number of regulatory T cells. gemcitabine treatment increased IFN-γ production among CD4 T cells but not among CD8 T cells. The results indicated that GEM had immunoregulatory properties that might support immune response against cancer. This finding has implications for designing chemoimmunotherapy strategies.

Endoscopic diagnosis of small intestinal diseases.

The technological development in endoscopy is directed toward improved accuracy of the diagnoses of novel diseases. The capsule endoscope and balloon-assisted endoscope are examples of such technological development. By these novel technologies, the small intestine can be examined in more detail. Therefore, an increasing number of novel diseases have been discovered, requiring the establishment of diagnosis and treatment strategies for these unknown diseases. In particular, obscure gastrointestinal bleeding, Crohn's disease, and nonsteroidal anti-inflammatory drug-induced enteropathy are of great interest to endoscopists. The capsule endoscope is the best method for screening the small intestine; however, the development of supporting methods such as the patency capsule is eagerly desired.