本橋 新一郎

BACKGROUND: We have previously identified the cancer-type organic anion transporting polypeptide 1B3 (Ct-OATP1B3) mRNA in several human colon and lung cancer tissues. Ct-OATP1B3 is a variant of the liver-type OATP1B3 (Lt-OATP1B3) mRNA, which is a hepatocyte plasma membrane transporter with broad substrate specificity. However, in cancer tissues, both the detailed characteristics of Ct-OATP1B3 mRNA expression and its biological functions remain unclear. With this point in mind, we sought to characterize Ct-OATP1B3 mRNA expression in colon and lung cancer tissues. In addition, we attempted to obtain functional implication of Ct-OATP1B3 in cancer cells. METHODS: Matched pairs of cancer and normal tissues were collected from 39 colon cancer and 28 lung cancer patients. The OATP1B3 mRNA expression levels in each of these tissues were separately determined by quantitative real-time polymerase chain reaction. Mann-Whitney U test and Fisher's exact test were used in statistical analysis. The Ct-OATP1B3 functional expression in colon cancer cells was then examined by Western blotting and transport analyses. RESULTS: Ct-OATP1B3 mRNA, but not Lt-OATP1B3 mRNA, was abundantly expressed in colon cancer tissues at a higher detection frequency (87.2%) than that of the adjacent normal tissues (2.6%). Furthermore, it was found that Ct-OATP1B3 mRNA expression was often detected in early colon cancer stages (88.9%, n = 18), and that its expression was associated with well-differentiated colon cancer statuses. On the other hand, Ct-OATP1B3 mRNA also showed a predominant and cancer-associated expression profile in lung tissues, although at frequencies and expression levels that were lower than those obtained from colon cancer. As for attempts to clarify the Ct-OATP1B3 functions, neither protein expression nor transport activity could be observed in any of the cell lines examined. CONCLUSIONS: Based on the unique characteristics of the Ct-OATP1B3 mRNA expression profile identified in this study, Ct-OATP1B3 mRNA can be expected to become a biomarker candidate for use in colon (and lung) cancer diagnosis. Simultaneously, our results advance the possibility that Ct-OATP1B3 might play yet unidentified roles, in addition to transporter function, in cancer cell biology.

Human natural killer T (NKT) cells are characterized by their expression of an invariant T cell antigen receptor alpha chain variable region encoded by a V alpha 24J alpha 18 rearrangement. These NKT cells recognize a-galactosylceramide (a-GalCer) in conjunction with the MHC class l-like CD1d molecule and bridge the innate and acquired immune systems to mediate efficient and augmented immune responses. A prime example of one such function is adjuvant activity: NKT cells augment anti-tumor responses because they can rapidly produce large amounts of IFN-gamma, which acts on NK cells to eliminate MHC negative tumors and also on CD8 cytotoxic T cells to kill MHC positive tumors. Thus, upon administration of alpha-GalCer-pulsed DCs, both MHC negative and positive tumor cells can be effectively eliminated, resulting in complete tumor eradication without tumor recurrence. Clinical trials have been completed in a cohort of 17 patients with advanced non-small cell lung cancers and 10 cases of head and neck tumors. Sixty percent of advanced lung cancer patients with high IFN-gamma production had significantly prolonged median survival times of 29.3 months with only the primary treatment. In the case of head and neck tumors, 10 patients who completed the trial all had stable disease or partial responses 5 weeks after the combination therapy of alpha-GalCer-DCs and activated NKT cells. We now focus on two potential powerful treatment options for the future. One is to establish artificial adjuvant vector cells containing tumor mRNA and alpha-GalCer/CD1d. This stimulates host NKT cells followed by DC maturation and NK cell activation but also induces tumor-specific long-term memory CD8 killer T cell responses, suppressing tumor metastasis even 1 year after the initial single injection. The other approach is to establish induced pluripotent stem (iPS) cells that can generate unlimited numbers of NKT cells with adjuvant activity. Such iPS-derived NKT cells produce IFN-gamma in vitro and in vivo upon stimulation with alpha-GalCer/DCs, and mediated adjuvant effects, suppressing tumor growth in vivo.

To develop more effective vaccines and strategies to regulate chronic inflammatory diseases, it is important to understand the mechanisms of immunological memory. Factors regulating memory CD4(+) T helper (Th)-cell pool size and function remain unclear, however. We show that activation of type I invariant natural killer T (iNKT) cells with glycolipid ligands and activation of type II natural killer T (iNKT) cells with the endogenous ligand sulfatide induced dramatic proliferation and expansion of memory, but not naive, CD4 T cells. NKT cell-induced proliferation of memory Th1 and Th2 cells was dependent largely on the production of IL-2, with Th2-cell proliferation also affected by loss of IL-4. Type II NKT cells were also required for efficient maintenance of memory CD4 T cells in vivo. Activation of iNKT cells resulted in up-regulation of IFN-gamma expression by memory Th2 cells. These IFN-gamma-producing memory Th2 cells showed a decreased capability to induce Th2 cytokines and eosinophilic airway inflammation. Thus, activated NKT cells directly regulate memory CD4 T-cell pool size and function via the production of cytokines in vivo.

Organic anion transporting polypeptide 1B3 (OATP1B3) is a hepatocyte plasma membrane protein that transports various endogenous and xenobiotic compounds. Although it is exclusively expressed in the human liver under normal conditions, OATP1B3 can be also expressed in various human cancer tissues that have been associated with prognosis and clinical outcomes. However, despite the potential significance of the latter finding, no experimental evidence addressing the molecular entity of cancer-associated OATP1B3 has been provided to date. In this paper, we report the identification of a new OATP1B3 mRNA isoform expressed in human colon and lung cancer tissues, which we named cancer-type OATP1B3 (Ct-OATP1B3). Our results also make known a previously unidentified transcription start site and an alternative promoter region, localized at intron 2, from which Ct-OATP183 mRNA is generated. Isoform specific mRNA quantification showed that the Ct-OATP1B3 mRNA level was strikingly higher than that of Lt-OATP1B3 mRNA in human cancer tissues. In addition, the results showed that the translation occurred at three out of four open reading frames. To summarize, our results clearly demonstrate that the newly-identified Ct-OATP1B3 (but not Lt-OATP1B3) is the primary mRNA isoform, at least in the human cancerous samples we have examined. In line with the possibility that its translation products play important biological roles in cancer cells, we strongly believe that the existence of Ct-OATP1B3 should be taken into account during future studies of OATP1B3 associated with cancer prognosis and clinical outcomes. (C) 2012 Elsevier Inc. All rights reserved.

Invariant NKT (iNKT) cells constitute a distinct lymphocyte subset, and upon activation, iNKT cells modulate the function of a wide variety of other immune cells including anti-tumor effector cells in both a direct and indirect manner. Decreased numbers and a reduced function of iNKT cells have been observed in patients with various malignant diseases, thus correlating with a poor clinical outcome. Therefore, therapeutic intervention strategies aimed at the recovery of functional iNKT cells would be an appropriate rationale for the treatment of cancer. Early clinical trials of iNKT cell-based immunotherapy demonstrated that the infusion of ligand-pulsed antigen presenting cells and/or in vitro activated iNKT cells was safe and well tolerated. This review summarizes the results of a series of clinical trials for lung cancer and head and neck cancer patients in Chiba University Hospital, Japan, and discusses iNKT cell-induced immune responses particularly those in the tumor microenvironment. (C) 2011 Elsevier Inc. All rights reserved.

Functionally polarized helper T cells (Th cells) play crucial roles in the induction of tumor immunity. There is considerable knowledge about the contributions of IFN-producing Th1 cells that supports the role of cytotoxic cluster of differentiation (CD8) T cells and natural killer (NK) cells, but much less is known about how IL-4-producing Th2 cells contribute to tumor immunity. In this study, we investigated the cellular and molecular mechanisms employed by memory Th2 cells in sustaining tumor immunity by using a mouse model system wherein ovalbumin (OVA) is used as a specific tumor antigen. In this model, we found that OVA-specific memory Th2 cells exerted potent and long-lasting antitumor effects against NK-sensitive OVA-expressing tumor cells, wherein antitumor effects were mediated by NK cells. Specifically, NK cell cytotoxic activity and expression of perforin and granzyme B were dramatically enhanced by the activation of memory Th2 cells. Interleukin 4 (IL-4) produced by memory Th2 cells in vivo was critical for the antitumor effects of the NK cells, which IL-4 directly stimulated to induce their perforin-and granzyme-B-dependent cytotoxic activity. Our findings show that memory Th2 cells can induce potent antitumor immunity through IL-4-induced activation of NK cells, suggesting potential applications in cellular therapy for cancer patients. Cancer Res; 71(14); 4790-8. (C) 2011 AACR.

V alpha 24 natural killer T (NKT) cells have potent anti-tumor activity. We performed a phase 11 clinical study in patients with head and neck squamous cell carcinoma (HNSCC) using ex vivo expanded V alpha 24 NKT cells and alpha-galactosylceramide (alpha GalCer; KRN7000)-pulsed antigen-presenting cells (APCs) to investigate the efficacy and induction of NKT cell-specific immune responses. The subjects were 10 patients with locally recurrent and operable HNSCC. One course of nasal submucosal administration of alpha GalCer-pulsed APCs and intra-arterial infusion of activated NKT cells via tumor-feeding arteries was given before salvage surgery. Anti-tumor effects, MKT cell-specific immune responses in extirpated cancer tissue and peripheral blood, safety, and pathological effects were evaluated. Five cases achieved objective tumor regression. The number of NKT cells increased in cancer tissues in 7 cases and was associated with tumor regression. The combination therapy induced NKT cell-specific immune responses in cancer tissues that were associated with beneficial clinical effects.. (C) 2010 Elsevier Inc. All rights reserved.

Invariant natural killer T (iNKT) cells are a conserved T cell sublineage and an important component of the innate immune system. The invariant T cell antigen receptor (TCR) chain on iNKT cells interacts with glycolipid presented via CD1d on antigen-presenting cells (APCs), resulting in the production of a variety of cytokines, and thus bridging the innate and adaptive immune systems. In this review, we discuss two strategies of immune modulation that target iNKT cells using either liposomal α-galactosylceramide (α-GalCer) or α-GalCer-loaded APCs. Liposomal α- GalCer generates regulatory iNKT cells, which serve to induce regulatory T cells (Treg) and can be used to diminish immune responses as is seen in autoimmunity and allergic diseases. In contrast, α-GalCer-pulsed APCs generate stimulatory iNKT cells capable of releasing pro-inflammatory cytokines and leading to adaptive immune responses that can be used for treating malignancies. Here, we summarize the modalities used to manipulate the dual nature of iNKT cell function and their tremendous potential in treating both allergic and malignant disease. © 2010 Bentham Science Publishers Ltd.

Invariant natural killer T(iNKT) cells have adjuvant activity due to their ability to produce large amounts of IFN-gamma, which activates other cells in innate and acquired systems, and orchestrates protective immunity. Based on these adjuvant mechanisms, we developed iNKT cell-targeted adjuvant therapy and carried out a phase I/IIa trial on advanced lung cancer patients. The patient group with increased numbers of IFN gamma-producing cells showed prolonged survival with a median survival time of 31.9 months. Sixty percent of the patients in this group survived for more than 2 years with only a primary treatment and without tumor progression and metastasis. (C) 2009 Elsevier Ltd. All rights reserved.

The aim of this study was to evaluate the molecular influence of chronic obstructive pulmonary diseases (COPD) on the pathogenesis of non-small cell lung cancer (NSCLC). The methylation profiles of 12 genes, and the epidermal growth factor receptor (EGFR) and KRAS mutations were determined for samples from 229 NSCLC patients. In addition, protein expression of EGFR and HER2 in 116 NSCLCs was analyzed based on the presence or absence of COPD. IL-12R beta 2 and Wif-1 methylation and HER2 overexpression were more frequent events in the COPD group. Eighty nonmalignant lung tissues had no correlation with any molecular changes between the COPD and the non-COPD group. EGFR mutation was significantly higher in the non-COPD group, while EGFR expression was inversely correlated with %FEV1.0. In the COPD group, unmethylated SPARC and sFRP-2 genes or a negative CpG island methylator phenotype (CIMP) was a negative prognostic factor, while methylation of p16(INK4A) and WNT antagonist genes was a negative prognostic factor in the non-COPD group. Novel characteristics of COPD-related NSCLC were identified by examination of methylation profiles and alterations of EGFR signaling. In consideration of the high sensitivity to smoking in patients with COPD, NSCLC with COPD might be a distinct population of smoke-related NSCLC, the genetic profile of which is quite different from non-COPD NSCLC.

The aim of this clinical trial was to investigate the feasibility of intra-arterial infusion of in vitro-expanded V alpha 24 natural killer T (NKT) cells combined with submucosal injection of alpha-galactosylceramide (KRN7000; alpha GalCer)-pulsed antigen-presenting cells (APC). A phase I clinical study was carried out in patients with head and neck squamous cell carcinoma (HNSCC). Patients with locally recurrent HNSCC refractory to standard therapy were eligible. Eight patients received super-selective transcatheter intra-arterial infusion of activated V alpha 24 NKT cells into tumor-feeding arteries and nasal submucosal injections of alpha GalCer-pulsed APC twice with a 1-week interval. V alpha 24 NKT cell-specific immune responses, safety, and antitumor effects were evaluated. The number of V alpha 24 NKT cells and interferon-gamma-producing cells in peripheral blood mononuclear cells increased in seven out of eight patients enrolled. Grade 3 toxicity with a pharyngocutaneous fistula related to local tumor reduction was observed in one patient and mild adverse events with grade 1-2 symptoms occurred in seven patients. Regarding the clinical responses, three cases exhibited a partial but significant response, four were classified as stable disease, and one patient continued to develop progressive disease. The use of the intra-arterial infusion of activated V alpha 24 NKT cells and the submucosal injection of alpha GalCer-pulsed APC has been shown to induce significant antitumor immunity and had beneficial clinical effects in the management of advanced HNSCC. The use of such therapeutic modalities may be helpful in the management of tumors and therefore needs to be explored in further detail. The clinical trial registration number was UMIN000000722. (Cancer Sci 2009; 100: 1092-1098).

Several studies have described p16(INK4A) and prostaglandin E2 (PGE2) co-alterations in various solid tumors, including non-small cell lung cancer (NSCLC). In this study, we examined the correlation between PGE2 receptor 2 (EP2) expression and p16(INK4A) methylation in NSCLC, and the association with clinicopathological features and prognostic significance. We retrospectively reviewed 88 NSCLC patients who underwent resection from July 1993 to May 1997. The tumors included 43 adenocarcinomas, 39 squamous cell carcinomas, and 6 large cell carcinomas. EP2 expression was determined by immunostaining, and p16(INK4A) methylation was analyzed by methylation specific PCR. EP2 was overexpressed in 44% of NSCLC patients, 61% of adenocarcinoma cases, 28% of squamous cell carcinoma cases, and 33% of large cell carcinoma cases. EP2 expression positively correlated with lymph node metastasis (P=0.034), especially in patients with squamous cell carcinoma (P < 0.009). Methylation of p16(INK4A) was detected in 34% of NSCLC patients, 23% of adenocarcinoma cases, 44% of squamous cell carcinoma cases, and 50%, of large cell carcinoma cases. In patients with squamous cell carcinoma, EP2 overexpression correlated with poor prognosis with a relative risk of 2.4 (confidence interval 2.1-51.8, P < 0.003), and positively correlated with p16(INK4A) methylation (P < 0.024). Adenocarcinoma patients with p16(INK4A) methylation had poor prognosis with a relative risk of 2.4 (confidence interval 1.8-69.7, P < 0.009), but this was not. correlated with EP2 expression. In conclusion, EP2 overexpression was common in NSCLCs, especially in adenocarcinomas. Synchronous alteration of p16(INK4A) and EP2 may accelerate progression of squamous cell carcinomas. These two alterations may differentially affect pathogenesis among subtypes of NSCLC.

To evaluate the safety, immune responses, and antitumor responses after the administration of a-galactosylceramide (alpha GalCer) KRN7000-pulsed PBMC cultured with IL-2 and GM-CSF (IL-2/GM-CSF-cultured PBMCs), a phase I-II study in patients with non-small cell lung cancer was conducted. Patients with advanced non-small cell lung cancer or recurrent lung cancer refractory to the standard therapy were eligible. alpha GalCer-pulsed IL-2/GM-CSF-cultured PBMCs (I X 10(9)/m(2)) were i.v. administered four times. Immune responses were monitored weekly. Twenty-three patients were enrolled in this study and 17 cases (73.9%) completed. No severe adverse event related to the treatment was observed. After the injection of aGalCer-pulsed IL-2/GM-CSF-cultured PBMCs, an increased number of IFN-gamma-producing cells in the peripheral blood were detected in 10 patients (58.8%). Five cases remained as stable disease, and the remaining 12 cases were evaluated as progressive disease. The estimated median survival time (MST) of the 17 cases was 18.6 mo (range, 3.8 to 36.3 mo). Ten patients who displayed increased IFN-gamma-producing cells (>= 2-fold) showed prolonged MST (31.9 mo; range, 14.5 to 36.3 mo) as compared with poor-responder patients (n = 7) MST (9.7 mo; range, 3.8 to 25.0 mo) (log-rank test,p = 0.0015). The administration of aGalCer-pulsed IL-2/GM-CSF-cultured PBMCs was well tolerated and was accompanied by the successful induction of NKT cell-dependent immune responses. The increased IFN-gamma-producing cells that result from aGalCer stimulation in PBMCs were significantly associated with prolonged MST. These results are encouraging and warrant further evaluation for survival benefit of this immunotherapy. The Journal of Immunology, 2009, 182: 2492-2501.

Human invariant Valpha24 Natural Killer T (NKT) cells are unique lymphocyte subsets, characterized by an invariant T-cell receptor Valpha24 chain paired with Vbeta11. Recent findings have highlighted the role of NKT cells in tumor immunity. Valpha24 NKT cells are activated by a specific glycolipid ligand, alpha-Galactosylceramide and rapidly produce high levels of cytokines upon stimulation, thereby modulating other immune cells such as NK cells antigen-specific CD4+ and CD8+ T cells and dendritic cells. Abnormalities in the numbers and functions of Valpha24 NKT cells have been observed in patients with various malignant diseases. Therefore, therapeutic strategies have recently focused on the reconstitution of an adequate number of functionally sufficient Valpha24 NKT cells which is thought to be logical and reasonable for cancer treatment. The quantitative alteration and functional impairment of circulating Valpha24 NKT cells are herein reviewed in various cancer-bearing patients and the progress to date in the clinical applications of NKT cell-based tumor immunotherapy is summarized.

Human V alpha 24 invariant natural killer T (iNKT) cells are a distinct lymphocyte population, characterized by an invariant T-cell receptor V alpha 24 chain paired mainly with V alpha 11. V alpha 24 iNKT cells are activated by a glycolipid ligand - alpha-galactosylceramide - and produce a large amount of Th1 and Th2 cytokines, thereby modulating the function of other cells. iNKT cells have the capability to control a wide variety of immune responses, including antitumor immunity. Abnormalities in the number and function of V alpha 24 NKT cells have been observed in patients with malignant diseases accompanied with a poor clinical outcome. Therefore, therapeutic strategies that focused on the restoration of V alpha 24 iNKT cell population and function would be a reasonable rationale for the treatment of cancer. In this article, the progress to date in the clinical studies of iNKT cell-based immunotherapy is briefly reviewed and the role of V alpha 24 iNKT cells in cancer immunotherapy is highlighted.

Chemokines play an important role in the pathogenesis of non-small cell lung cancer (NSCLC). However, aberrant methylation of CXCL12 has not been examined in NSCLC. CXCL12 mRNA expression and methylation were examined in 17 NSCLC cell lines by RT-PCR and methylation-specific PCR (MSP). MSP was performed on 236 tumor specimens from NSCLC patients who received curative intent surgery. CXCL12 and CXCR4 protein expression was examined in 90 of the 236 NSCLC specimens by immunohistochernistry. Down-regulation of CXCL12 expression was found in 10 of 17 (59%) NSCLC cell lines compared with normal bronchial cells. Treatment of 8 expression-negative cell lines with a demethylating agent restored expression in all cases. Twelve cell lines (71%) showed aberrant methylation, and good concordance between methylation and expression was present. Aberrant methylation occurred in 85 out of 236 (36%) primary NSCLCs in a tumor-specific manner. In multivariate analysis, CXCL12 methylation correlated strongly and independently with prognosis both in all patients with NSCLCs and in those with stage I NSCLCs (hazard ratio=1.68, P=0.015 and hazard ratio=3.58, P=0.017). Secreted protein CXCL12 and its receptor CXCR4 were abundant in NSCLC cells (72 out of 90, 80%; 57 out of 90, 63%) and correlated with the progression of NSCLCs. In conclusion, epigenetic silencing of CXCL12 is a frequent event in NSCLCs, and could be an independent and powerful prognostic marker in patients with NSCLCs and those with stage I disease. Analysis for CXCL12 may provide novel opportunities for prognosis and therapy of resected NSCLCs.

Human invariant V alpha 24 natural killer T (NKT) cells are a novel, distinct lymphocyte population, characterized by an invariant T-cell receptor V alpha 24 chain paired with V beta 11. V alpha 24 NKT cells are activated by a specific glicolipid ligand, alpha-GalCer, and rapidly produce a large amount of Th1 and Th2 cytokines, thereby modulating other immune cells such as antigen-specific CD4 and CD8 T cells, NK cells, and dendritic cells. Recent studies have shown that NKT cells play pivotal regulatory roles in many immune responses, including antitumor immunity. We herein review the quantitative alteration and functional deterioration of circulating V alpha 24 NKT cells in various cancer-bearing patients. We also summarize the recent progress in the clinical studies of NKT cell-based tumor immunotherapy. Novel immunological results including the increased peripheral blood V alpha 24 NKT cells and IFN-producing cells after the immunotherapy were revealed. The details of the safety profile and the antitumor responses were also disclosed. Although the objective clinical responses still remain unclear, some encouraging results have emerged. Therefore, NKT cell-based immunotherapy may potentially be an effective strategy for the treatment of cancer patients.

Background Human V alpha 24 natural killer T (NKT) cells are activated by the specific ligand, alpha-galactosylceramide (alpha-GalCer), in a CD1d-dependent manner. Potent anti-tumor activity of activated NKT cells has been previously demonstrated. Methods We conducted a phase I study with alpha-GalCer-pulsed antigen presenting cells (APCs) administered in the nasal submucosa of patients with head and neck cancer, and evaluated the safety and feasibility of such a treatment. Nine patients with unresectable or recurrent head and neck cancer received two treatments 1 week apart, of 1 x(8) of alpha-GalCer-pulsed autologous APCs into the nasal submucosa. Results During the clinical study period, no serious adverse events (Common Terminology Criteria for Adverse Events version 3.0 greater than grade 3) were observed. After the first and the second administration of alpha-GalCer-pulsed APCs, an increased number of NKT cells was observed in four patients and enhanced natural killer activity was detected in the peripheral blood of eight patients. Conclusion The administration of alpha-GalCer-pulsed APCs into the nasal submucosa was found to be safe and induce anti-tumor activity in some patients.

Purpose: The Wnt and epidermal growth factor receptor (EGFR) signaling pathways play crucial roles in the pathogenesis of a variety of malignant tumors. Although the details of each cascade are understood, very little is known about their collective effects in non -small cell lung cancer (NSCLC). Experimental Design: A total of 238 NSCLC samples were examined for methylation of Wnt antagonists [secreted frizzled-related protein (sFRP)-1, sFRP-2, sFRP-5,Wnt inhibitory factor-1, and Dickkopf-3] and for EGFR and KRAS mutations. Protein expression levels of P-catenin were assayed in 91 of the 238 NSCLCs. Results: We found that (a) aberrant methylation of Wnt antagonists is common in NSCLCs; (b) methylation of sFRP-2 is more prevalent in females, nonsmokers, and adenocarcinoma cases; (c) Dickkopf-3 methylation is significantly associated with a poor prognosis in adenocarcinomas; (d) there is a positive correlation between activated EGFR mutation and nuclear accumulation of beta-catenin; (e) KRAS mutation and aberrant methylation of Wnt antagonists are positively correlated; and (f) EGFR mutation is significantly associated with a good prognosis in tumors lacking methylated Wnt antagonist genes. Conclusions: These results contribute to a better understanding of the cross-talk between the Wnt and EGFR signaling pathways and help foster development of chemotherapeutic treatments in NSCLCs.

Purpose: Human V alpha 24 natural killer T (V alpha 24 NKT) cells bearing an invariant V alpha 24J alpha Q antigen receptor are activated by a glicolipid ligand a-galactosylceramide (alpha GalCer; KRN7000) in a CD1d-dependent manner. The human V alpha 24 NKTcells activated with alpha GalCer and interleukin-2 have been shown to produce large amounts of cytokines, such as IFN-gamma, and also exerting a potent killing activity against various tumor cell lines. We did a phase I study with autologous activated V alpha 24 NKT cell therapy. Experimental Design: Patients with advanced or recurrent non-small cell lung cancer received i.v. injections of activated V alpha 24 NKTcells (level 1: 1 x 10(7)/m(2) and level 2: 5 x 107/m(2)) to test the safety, feasibility, and clinical response of this therapeutic strategy. Immunomonitoring was also done in all cases. Results: Six patients were enrolled in this study. No severe adverse events were observed during this study in any patients. After the first and second injection of activated V alpha 24 NKTcells, an increased number of peripheral blood V alpha 24 NKTcells was observed in two of three cases receiving a level 2 dose of activated V alpha 24 NKTcells. The number of IFN-gamma-producing cells in peripheral blood mononuclear cells increased after the administration of activated V alpha 24 NKT cells in all three cases receiving the level 2 dose. No patient was found to meet the criteria for either a partial or a complete response. Conclusions: The clinical trial with activated V alpha 24 NKT cell administration was well tolerated and carried out safely with minor adverse events even in patients with advanced diseases.

A unique lymphocyte population, CD1d-restricted NKT cells, has been revealed to be a key player in both the innate and acquired immune responses, including antitumor effects. Recent studies revealed that at least two subsets of CD1d-restricted NKT cells exist: type I, having invariant V alpha 14 receptor; and type II, having heterogeneous non-V alpha 14 receptor. The specific glycolipid ligand, alpha-GalCer, effectively stimulates mouse and human type I NKT cells. The activation of type I NKT cells substantially influences function of other various cell types, particularly DC, NK cells, CD4 Th1 cells, and CD8 cytotoxic T cells, all contributing to the antitumor immune responses. Recent studies also indicated that, unlike type I NKT cells, type II NKT cells have a potential to repress antitumor immune responses. In this review, we summarize the characteristics of the antitumor immune responses mediated by both mouse and human CD1d-restricted NKT cells and discuss their potential in clinical applications against cancer.

We report on a case of a complete resection for bronchogenic carcinoma necessitating right upper sleeve lobectomy with prosthetic replacement of right pulmonary artery (PA) and superior vena cava (SVC). A 74-year-old male with squamous cell carcinoma had a tumor which extended to the right main bronchus, right PA and SVC. After reconstruction of the SVC with a ringed polytetrafluoroethylene (PTFE) graft between the left brachiocephalic vein (BCV) and right atrial auricle, the tumor was completely resected en bloc. Bronchial anastomosis followed by the prosthetic reconstructions of PA and SVC between the right BCV and the origin of SVC were performed. Pathological staging was t4n2m0 (stage IIIB). The postoperative course was uneventful and the patient discharged from the hospital on the 29th postoperative day. He has been doing well without recurrence and keeping a good graft patency for more than 3 years.

Background. Treatment of pulmonary metastases from colorectal cancer by excision has increased rapidly, but reports on indications and prognostic factors are inconsistent. We sought to identify poor prognostic factors preoperatively and to retrospectively evaluate preoperative clinical indications for surgery. Methods. A total of 75 patients with colorectal cancer had pulmonary metastases excised from 1986 to 2003. Tumor size, number, laterality, hilar or mediastinal lymphadenopathy, and carcinoembryonic antigen level were possible risk factors for metastatic tumors, with primary site of colorectal tumor, disease-free interval, and hepatectomy for liver metastasis possible risk factors for primary tumors. Prognostic factors in univariate and multivariate analyses also included age and sex. Results. Five-year survival rates were 41.3% after pulmonary excision and 73.1% after primary colorectal resection. Three factors identified as significant by univariate log-rank test for overall survival after pulmonary resection were carcinoembryonic antigen (p < 0.0001), tumor laterality (p = 0.0205), and number of pulmonary metastases (p = 0.0028). Multivariate analysis found that carcinoembryonic antigen, tumor number, tumor size, and patient's age were also independent prognostic factors. In contrast, carcinoembryonic antigen, number of metastases, and disease-free interval predicted prognosis after primary colorectal resection. Prior hepatectomy for metastases did not influence prognosis after pulmonary metastasectomy. Conclusions. Elevated carcinoembryonic antigen level and multiple metastases are preoperative predictors of poor prognosis after resection of pulmonary metastases from colorectal cancer. Survival rate is sufficient to justify pulmonary metastasectomy if there is no local or distant metastatic lesion other than in the liver; if needed, sequential pulmonary and hepatic metastasectomy can be performed.

Human invariant V alpha 24(+) natural killer T (NKT) cells display potent antitumor activity upon stimulation. Activation of endogenous V alpha 24(+) NKT cells would be one strategy for the treatment of cancer patients. For example, dendritic cells (DCs) loaded with a glycolipid NKT cell ligand, alpha-galactosylceramide (alpha GalCer, KRN7000), are a possible tool for the activation and expansion of functional V alpha 24(+) NKT cells in vivo. In this report, we demonstrate that the levels of expansion and the ability to produce IFN-gamma of V alpha 24(+) NKT cells induced by alpha GalCer-loaded whole PBMCs cultured with IL-2 and GM-CSF (IL-2/GM-CSF-cultured PBMCs) were superior to those of cells induced by monocyte-derived CD11c(+) DCs (moDCs) developed with IL-4 and GM-CSF. Interestingly, CD11c(+) cells in the IL-2/GM-CSF-cultured PBMCs showed a mature phenotype without further stimulation and exerted potent stimulatory activity on V alpha 24(+) NKT cells to enable them to produce IFN-gamma preferentially at an extent equivalent to mature moDCs induced by stimulation with LPS or a cytokine cocktail. Cocultivation with CD11c(-) cells in the IL-2/GM-CSF-cultured PBMCs induced maturation of moDCs. In particular, CD11c(-)CD3(+) T cells appeared to play important roles in DC maturation. In addition, TNF-alpha was preferentially produced by CD11c(-)CD3(+) T cells in IL-2/GM-CSF-cultured PBMCs and was involved in the maturation of moDCs. Thus, the maturation of DCs induced by CD11c(-) T cells through TNF-alpha production appears to result in the efficient expansion and activation of V alpha 24(+) NKT cells to produce IFN-gamma preferentially. (c) 2005 Wiley-Liss, Inc.

In 1999, the World Health Organization introduced a new classification of a rare thymic cancer called papillary adenocarcinoma of the thymus gland. In this study we report on a case of papillary thymic carcinoma in a 29-year-old woman. Histologically, the tumor consisted of a papillary component admixed with a solid component, a component that is commonly found in type A thymoma.

A unique lymphocyte population, V alpha 14 NKT cells, has recently been revealed to be a key player in the immune responses against tumors. Activation of V alpha 14 NKT cells affects various cell types, particularly dendritic cells (DCs), NK cells, CD4 Th1 cells, and CD8 cytotoxic T cells in the innate and acquired immune systems, eventually resulting in the enhanced activation of NKT cell-mediated cellular cascade in the anti-tumor responses. The specific ligand, alpha-galactosylceramide (alpha-GalCer), effectively stimulates mouse and human NKT cells, making NKT cells an ideal target for the development of cancer immunotherapy. Clinical trials using alpha-GalCer have actually started in several centers in the world. In this review, we summarize the Va14 NKT cell-mediated cellular cascade in the anti-tumor response in mice and discuss potential clinical applications of alpha-GalCer-pulsed DC therapy.

Neurogenic benign tumors arising from the trachea and bronchus are relatively rare. We experienced three cases of neurofibroma of the bronchus which were successfully treated by transbronchial electrical snaring and Nd-YAG laser abrasion. The first was a 67-year-old man with right lung cancer, who was pointed out to have a neurofibroma in the left main bronchus. The second was a 34-year-old man with an obstruction in the right main bronchus due to neurofibroma. The third was a 66-year-old woman with a complete obstruction in the left main bronchus due to schwannoma. All patients were successfully treated to remove the tumors and obtain a patency of the bronchus by transbronchial electrical snaring and Nd-YAG laser abrasion. We also review 23 reported cases of endobronchial neurogenic tumors and discuss the efficacy of endoscopic treatments for endobronchial neurogenic tumors.

Papillary thyroid carcinoma with massive invasion into the great veins of the neck and mediastinum has rarely been reported and is thought to have a poor prognosis. Here we report successful management of a case of papillary thyroid carcinoma with extensive invasion into the left internal jugular vein, left brachiocephalic vein, and superior vena cava, followed by reconstruction of the superior vena cava using an artificial graft. The operation was conducted to prevent sudden death due to complete obstruction of venous flow, improve the patient's quality of life, and prolong survival. The patient has survived for more than two years after surgery, with good general condition.

Interleukin 6 (IL-6) facilitates the differentiation of B cells to immunoglobulin-secreting cells and is reported to be a proliferative factor in some tumors. In this study, we examined IL-6 production in non-small cell lung carcinoma (NSCLC) and the proliferation of tumor cells following IL-6 treatment in vitro and in vivo. We analyzed the expression of IL-6 mRNA and protein in a series of 15 human lung cancer cell lines (four adenocarcinomas, five squamous cell carcinomas, two large cell carcinomas, and four small cell carcinomas) by reverse transcriptase polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA). We established an IL-6-producing cell line (ABC-1#IL-6) by transfecting a human IL-6 cDNA into a human non-IL-6-producing NSCLC cell line (ABC-1). These two cell lines were used to determine tumor cell proliferation both in vivo and in vitro in order to clarify the effect of IL-6 on tumor growth and metastasis. Athymic nude mice, SCID mice, and BALB/c mice were subcutaneously inoculated with these two cell lines, and body weight, tumor growth, and tumor doubling time were measured. The presence of IL-6 and tumor-infiltrating lymphocytes (TILs) within tumor tissues was examined by immunohistochemical staining. Results: Eight of 15 (53%) lung cancer cell lines expressed both IL-6 mRNA and protein. Tumor lesions of both cell lines developed in nude and SCID mice, although no such lesions of either cell lines developed in BALB/c mice. The tumor doubling time in nude and SCID mice was 2.97+/-1.22 days and 2.45+/-1.32 days, respectively, in mice inoculated with the cell line ABC-1#IL-6. These doubling times were statistically significantly shorter than those evident in mice inoculated with the control original ABC-1 cell line (nude, p=0.0337; SCID, p=0.0119; unpaired t-test). The rates of cell proliferation in vitro of the ABC-1#IL-6 and original ABC-1 cells lines were comparable (p=0.1441, unpaired t-test). Immunohistochemical staining revealed strong IL-6 expression in tumors derived from the IL-6-producing cell line but not in tumors derived from the original ABC-1 cell line (both in nude and SCID mice). Conclusion: 53% of lung cancer cell lines produce IL-6 mRNA and protein. Although IL-6 itself does not influence tumor cell proliferation in vitro, an association between IL-6 expression and tumor proliferation was found in vivo in nude and SCID mice. An anti-IL-6 reagent could provide a novel therapeutic strategy in patients with IL-6-producing lung tumors.

Purpose: The aim of this study was to determine whether collagen XVIII expression is correlated with circulating serum endostatin and whether this has any prognostic value in patients with non-small cell lung cancer (NSCLC). Experimental Design: Serum endostatin levels were measured quantitatively by a competitive enzyme immunoassay, and collagen XVIII expression in tumor tissue was investigated with an immunohistochemical method in a series of 94 patients who underwent surgery for NSCLC. Results: Sixty cases (63.8%) had positive immunohistochemical staining with anticollagen XVIII polyclonal antibodies, including strongly positive staining in 11 (11.7%) cases. The mean (+/- SD) serum endostatin level was 41.6 +/- 34.4 ng/ml in the patient group and 16.3 +/- 10.3 ng/ml in the control group (P < 0.0001). The It cases who were strongly collagen XVIII-positive had significantly higher serum endostatin levels than the cases who were negative or weakly positive (P = 0.0297). The 5-year survival rates of negative, weakly positive, and strongly positive patients were 77.8%, 56.9%, and 43.8%, respectively. The cases with strongly positive collagen XVIII expression had a significantly poorer outcome than cases with negative expression (P = 0.0027). A multivariate analysis with Cox proportional hazards model for disease-specific survival revealed that expression of collagen XVIII (strongly positive versus negative; weakly positive versus negative), tumor classification, and regional lymph node classification were independent prognostic factors. Conclusions: Our results suggest that expression of collagen XVIII in tumor tissue is strongly associated with a poorer outcome in NSCLC and correlates with elevated levels of circulating serum endostatin.

Sarcoidosis is a systemic disorder associated with granuloma characterized by an abnormal T(h)1-type cytokine production and accumulation of T(h)1 CD4 T cells in the granuloma lesions, suggesting an importance of T(h)1 responses in sarcoidosis. However, the pathogenesis of sarcoidosis remains to be solved. Here, we investigated the nature of V(alpha)24 NKT cells with immunoregulatory functions in sarcoidosis. Patients with non-remitting sarcoidosis displayed a decrease in the number of V(alpha)24 NKT cells in peripheral blood, but an accumulation of these cells in granulomatous lesions. When stimulated with the specific glycolipid ligand, alpha-galactosylceramide, peripheral blood V(alpha)24 NKT cells from patients with non-remitting disease produced significantly less IFN-gamma than those from healthy volunteers, but normal levels of IL-4. The reduced IFN-gamma production was observed only in V(alpha)24 NKT cells and not conventional CD4 T cells, but was normal in patients with remitting disease, suggesting that non-remitting sarcoidosis involves an insufficient IFN-gamma production of V(alpha)24 NKT cells which is well correlated with disease activity. Thus, these results suggest that V(alpha)24 NKT cells play a crucial role in the disease status of sarcoidosis.

Background. The aim of this study was to evaluate the influence of chronic obstructive pulmonary diseases (COPD) on postoperative pulmonary function and to elucidate the factors for decreasing the reduction of pulmonary function after lobectomy. Methods. We conducted a retrospective chart review of 521 patients who had undergone lobectomy for lung cancer at Chiba University Hospital between 1990 and 2000. Forty-eight patients were categorized as COPD, defined as percentage of predicted forced expiratory volume at 1 second (FEV1) less than or equal to 70% and percentage of FEV1 to forced vital capacity less than or equal to 70%. The remaining 473 patients were categorized as non-COPD. Results. Although all preoperative pulmonary function test data and arterial oxygen tension were significantly lower in the COPD group, postoperative arterial oxygen tension and FEV1 were equivalent between the two groups, and the ratio of actual postoperative to predicted postoperative FEV1 was significantly better in the COPD group (p < 0.001). With multivariable analysis, COPD and pulmonary resection of the lower portion of the lung (lower or middle-lower lobectomies) were identified as independent factors for the minimal deterioration of FEV1. Actual postoperative FEV1 was 15% lower and higher than predicted, respectively, in the non-COPD patients with upper portion lobectomy and the COPD patients with lower portion lobectomy. Finally, we created a new equation for predicting postoperative FEV1, and it produced a higher coefficient of determination (R-2) than the conventional one. Conclusions. The postoperative ventilatory function in patients with COPD who had lower or middle-lower lobectomies was better preserved than predicted. (C) 2003 by The Society of Thoracic Surgeons.

Background: Certain immune functions are known to be impaired in human beings exposed to Antarctic winter; in particular, decreased amounts of serum proinflammatory cytokines, such as TNF-alpha and IL-1, were noted. It is not known, however, whether this exposure has any effect on T-cell-mediated acquired immune functions. Objectives: This study aims to investigate whether exposure to Antarctic winter has any effect on T cell-dependent immune functions. Methods: We assessed changes in various immunologic indicators. including serum levels of various cytokines, peripheral blood Valpha24Vbeta11 natural killer T cell numbers, and T(H)1/T(H)2 ratios of 40 Japanese personnel exposed to an Antarctic winter. Also, a 2-month inland traverse was executed during the isolation, and the effect on the above indicators was assessed. Results: All subjects were healthy during the Antarctic isolation. The levels of serum TNF-alpha, IL-1Ra, IL-6, and IL-1beta were dramatically reduced and remained at low levels throughout the isolation. The decrease in the levels of TNF-alpha and IL-1Ra was more pronounced during the inland traverse than during the rest of the isolation. The percentage of Valpha24Vbeta11 natural killer T cells was significantly increased at the midpoint of the isolation. Most interestingly, T(H)1/T(H)2 ratio was increased significantly, and this T(H)1 bias was most prominent at the late point of the isolation. Conclusions: Exposure to an Antarctic winter appeared to induce T(H)1-skewed immunity in human beings. (J Allergy Clin Immunol 2003;111:1353-60.).

We report the case of a 60-year-old woman referred to us after chest X-ray and mobile computed tomography screening detected an 8-mm nodule in right S2. Transbronchial aspiration cytology suggested a pulmonary metastasis from colorectal cancer. Therefore, we performed a colonoscopy and found a polypoid lesion, 2 cm in diameter, in the sigmoid colon. An analysis of a biopsy specimen from this polypoid lesion confirmed adenocarcinoma. Surgical resection of the primary sigmoid colon cancer was subsequently performed, followed 4 weeks later by a right S2 segmentectomy to remove the lung metastasis. The patient is currently well without any clinical signs of recurrence, 44 months after her operation.

Human Valpha24 NKT cells bearing an invariant Valpha24JalphaQ antigen receptor, the counterpart of murine Valpha14 NKT cells, are activated by a specific ligand, alpha-GalCer, in a CD1d-dependent manner. Here, we demonstrate decreased numbers of circulating Valpha24 NKT cells in patients with primary lung cancer compared to healthy volunteers. However, Valpha24 NKT cells and DCs from lung cancer patients were functionally normal, even in the presence of tumor. Furthermore, levels of Valpha24 NKT cells in surgically resected lung tissue appeared to be equivalent to those of Valpha14 NKT cells in the mouse lung. Levels of Valpha24 NKT cells in the tumor tissue itself were increased about 2.5 times. Administration of alpha-GalCer-pulsed DCs expanded Valpha14 NKT cells in the lung more than 10 times, and the increased levels were sustained for 1 week. This may explain the previous finding that alpha-GalCer-pulsed DCs exerted strong antitumor activity in mouse lung tumor metastatic models. The potential use of alpha-GalCer-pulsed DCs for immunotherapy aimed at activating endogenous Valpha24 NKT cells in the lung of cancer patients is discussed. (C) 2002 Wiley-Liss, Inc.

NKT cells, a novel murine lymphoid lineage bearing an invariant T cell receptor encoded by Valpha14 and Jalpha281 gene segments, recognize a specific ligand glycolipid, alpha-galactosylceramide (alpha-GalCer) in a CD1d-dependent manner. Recent research has revealed that activated Valpha14 NKT cells have dramatic antitumor effects against a wide variety of tumor cell lines in vivo and in vitro. Here, we demonstrate strong in vivo antitumor effects brought about by treatment with alpha-GalCer-pulsed dendritic cells in comparison with in vitro-activated Valpha14 NKT cells. Furthermore, we show a significant expansion of endogenous Valpha14 NKT cells in the lung following the administration of alpha-GalCer-pulsed dendritic cells. The feasibility of immunotherapy with alpha-GalCer-pulsed dendritic cells is discussed.