New CETP inhibitor K-312 reduces PCSK9 expression: a potential effect on LDL cholesterol metabolism.

Despite significant reduction of cardiovascular events by statin treatment, substantial residual risk persists, driving emerging needs for the development of new therapies. We identified a novel cholesteryl ester transfer protein (CETP) inhibitor, K-312, that raises HDL and lowers LDL cholesterol levels in animals. K-312 also suppresses hepatocyte expression of proprotein convertase subtilisin/kexin 9 (PCSK9), a molecule that increases LDL cholesterol. We explored the underlying mechanism for the reduction of PCSK9 expression by K-312. K-312 inhibited in vitro human plasma CETP activity (IC50; 0.06 µM). Administration of K-312 to cholesterol-fed New Zealand White rabbits for 18 wk raised HDL cholesterol, decreased LDL cholesterol, and attenuated aortic atherosclerosis. Our search for additional beneficial characteristics of this compound revealed that K-312 decreases PCSK9 expression in human primary hepatocytes and in the human hepatoma cell line HepG2. siRNA silencing of CETP in HepG2 did not compromise the suppression of PCSK9 by K-312, suggesting a mechanism independent of CETP. In HepG2 cells, K-312 treatment decreased the active forms of sterol regulatory element-binding proteins (SREBP-1 and -2) that regulate promoter activity of PCSK9. Chromatin immunoprecipitation assays demonstrated that K-312 decreased the occupancy of SREBP-1 and SREBP-2 on the sterol regulatory element of the PCSK9 promoter. PCSK9 protein levels decreased by K-312 treatment in the circulating blood of cholesterol-fed rabbits, as determined by two independent mass spectrometry approaches, including the recently developed, highly sensitive parallel reaction monitoring method. New CETP inhibitor K-312 decreases LDL cholesterol and PCSK9 levels, serving as a new therapy for dyslipidemia and cardiovascular disease.

Design and discovery of 2-oxochromene derivatives as liver X receptor ß-selective agonists.

In an attempt to molecularly design liver X receptor (LXR) ß-selective agonists, we discovered that the combination of the 2-oxochromene moiety (head) and the imidazoline-2,4-dione moiety (tail) plays an important role in the expression potency and selectivity toward LXRß. We synthesized a series of 2-oxochromene derivatives and identified 43 as a LXRß-selective agonist that increased the HDL-C level without significantly elevating the TG level and resulted in a decreased lipid-accumulation area in the aortic arch in a high-fat-and-cholesterol-fed Bio F1B hamster. In this manuscript, we report the design, synthesis and pharmacology of these 2-oxochromene derivatives.

Virtual bronchoscopic navigation combined with endobronchial ultrasound to diagnose small peripheral pulmonary lesions: a randomised trial.

BACKGROUND: Bronchoscopy using endobronchial ultrasound (EBUS) can help to diagnose small peripheral pulmonary lesions. However, although biopsy sites can be confirmed, a bronchoscope cannot be guided in EBUS. Virtual bronchoscopic navigation (VBN) can guide a bronchoscope with virtual images, but its value has not been confirmed. METHODS: This prospective multicentre study examines the value of VBN-assisted EBUS for diagnosing small peripheral pulmonary lesions. 199 patients with small peripheral pulmonary lesions (diameter ≤30 mm) were randomly assigned to VBN-assisted (VBNA) or non-VBN-assisted (NVBNA) groups. A bronchoscope was introduced into the target bronchus of the VBNA group using the VBN system. Sites of specimen sampling were verified using EBUS with a guide sheath under fluoroscopy. RESULTS: The diagnostic yield was higher for the VBNA than for the NVBNA group (80.4% vs 67.0%; p = 0.032). The duration of the examination and time elapsed until the start of sample collection were reduced in the VBNA compared with the NVBNA group (median (range), 24.0 (8.7-47.0) vs 26.2 (11.6-58.6) min, p = 0.016) and 8.1 (2.8-39.2) vs 9.8 (2.3-42.3) min, p = 0.045, respectively). The only adverse event was mild pneumothorax in a patient from the NVBNA group. CONCLUSIONS: The diagnostic yield for small peripheral pulmonary lesions is increased when VBN is combined with EBUS. Clinical trial number UMIN000000569.

Clinical investigation of malignant mesothelioma in Japan.

PURPOSE: The asbestos-related problems caused much social concern; however, no large-scale study was conducted about clinical features of MM in Japan. Patients with MM who have a history of occupational asbestos exposure (AE) are provided worker's compensation in Japan. However, only about 10% of MM cases were actually claimed and compensated. So there is still controversy over the association between MM and AE. The aim of this study is to investigate the clinical features of MM. We also aimed to clarify the association between MM and occupational AE in Japan. METHODS: We examined the clinical features of MM cases. Clinical information was obtained including gender, age, site of origin, pathological subtype, radiological findings, and treatment outcome. To investigate the association between MM and AE, investigators interviewed all patients regarding work and residential history. RESULTS: Between January 2005 and December 2007, 105 cases (median age: 63 years, range 35-80, male/female: 88/17) were diagnosed with MM in the Rosai Hospital group and related facilities. Among them, 94(89.5%) cases originated in the pleura, 7(6.7%) in the peritoneum, 2(1.9%) in the pericardium, and 1(0.9%) in the tunica vaginalis testis. There were 69(65.7%) epithelioid, 19(18.1%) biphasic, 16(15.2%) sarcomatoid, and 1 unclassified pathological subtypes of MM. A favorable survival rate was indicated in the patient group of MPM that underwent surgery compared to others, though it was not statistically significant (P = 0.1743). The occupational AE was indicated in 89 cases (84.8%). Three patients had no history of occupational AE, but lived with someone who was in an occupation that handled asbestos. There were two patients in which AE was indicated in their life environment. Altogether, AE was indicated in 93(88.6%) patients. CONCLUSIONS: This study stresses the urgent need for physicians to acknowledge the association between MM and AE, and to inquire thoroughly regarding AE to the patients with MM.

Combined survival analysis of prospective clinical trials of gefitinib for non-small cell lung cancer with EGFR mutations.

PURPOSE: Somatic mutations of the epidermal growth factor receptor (EGFR) gene are associated with an increased response to gefitinib in patients with non-small cell lung cancer. We have examined the impact of gefitinib on progression-free survival and overall survival in patients with EGFR mutation-positive non-small cell lung cancer. EXPERIMENTAL DESIGN: We searched for all clinical trials that prospectively evaluated the efficacy of gefitinib for advanced non-small cell lung cancer with EGFR mutations in Japan. We did a combined analysis based on individual patient data from the identified trials. RESULTS: Seven eligible trials were identified for a total of 148 non-small cell lung cancer patients with EGFR mutations. The overall response rate to gefitinib was 76.4% [95% confidence interval (95% CI), 69.5-83.2]. The median progression-free survival and overall survival were 9.7 months (95% CI, 8.2-11.1) and 24.3 months (95% CI, 19.8-28.2), respectively. Good performance status and chemotherapy-naïve status were significantly associated with a longer progression-free survival or overall survival. Of the 148 patients, 87 received gefitinib as a first-line therapy, whereas 61 received systemic chemotherapy before gefitinib treatment. The median progression-free survival after the start of first-line therapy was significantly longer in the gefitinib-first group than in the chemotherapy-first group (10.7 versus 6.0 months; P < 0.001), whereas no significant difference in median overall survival was apparent between the two groups (27.7 versus 25.7 months; P = 0.782). CONCLUSIONS: Gefitinib monotherapy confers substantial clinical benefit in terms of progression-free survival and overall survival in non-small cell lung cancer patients with EGFR mutations. Randomized trials comparing chemotherapy with gefitinib as a first-line treatment are warranted in such patients.

Phase I study of amrubicin and vinorelbine in non-small cell lung cancer previously treated with platinum-based chemotherapy.

BACKGROUND: Combination chemotherapy comprising amrubicin and vinorelbine as a second-line therapy for advanced non-small cell lung cancer (NSCLC) has not been fully evaluated. To determine the maximum tolerated dose (MTD) and recommended dose (RD), the present phase I study examined patients with advanced NSCLC. METHODS: The subjects were nine patients with histologically confirmed advanced NSCLC, Eastern Cooperative Oncology Group performance status 0-1, prior platinum-based first-line chemotherapy, and measurable or evaluable lesions. Treatment consisted of five dose levels, with amrubicin 35-45 mg/m2 administered as a 5-min intravenous infusion on days 1-3 and vinorelbine 15-25 mg/m2 given as a 1-h intravenous infusion on days 1 and 8, every 3 weeks. RESULTS: All patients had received carboplatin and paclitaxel as first-line therapy. Dose-limiting toxicity (DLT) was seen in two of six patients (febrile neutropenia and deep vein thrombosis ) at level 1, allowing us to conduct level 2. At level 2, all three patients experienced DLT (leucopenia > or =4 days in one patient; febrile neutropenia in three patients; and infection in two patients), and this level was determined as the MTD. Subsequently, level 1 (amrubicin 35 mg/m2 and vinorelbine 15 mg/m2) was defined as the RD. Responses in the nine patients included a partial response in one patient and stable disease in four patients. CONCLUSION: As second-line therapy, the RD of the combination of amrubicin and vinorelbine is 35 mg/m2 and 15 mg/m2, respectively. Further study should proceed to clarify the efficacy of this regimen.

First-line gefitinib for patients with advanced non-small-cell lung cancer harboring epidermal growth factor receptor mutations without indication for chemotherapy.

PURPOSE: This multicenter phase II study was undertaken to investigate the efficacy and feasibility of gefitinib for patients with advanced non-small-cell lung cancer (NSCLC) harboring epidermal growth factor receptor (EGFR) mutations without indication for chemotherapy as a result of poor performance status (PS). PATIENTS AND METHODS: Chemotherapy-naïve patients with poor PS (patients 20 to 74 years of age with Eastern Cooperative Oncology Group PS 3 to 4, 75 to 79 years of age with PS 2 to 4, and >or= 80 years of age with PS 1 to 4) who had EGFR mutations examined by the peptide nucleic acid-locked nucleic acid polymerase chain reaction clamp method were enrolled and received gefitinib (250 mg/d) alone. RESULTS: Between February 2006 and May 2007, 30 patients with NSCLC and poor PS, including 22 patients with PS 3 to 4, were enrolled. The overall response rate was 66% (90% CI, 51% to 80%), and the disease control rate was 90%. PS improvement rate was 79% (P < .00005); in particular, 68% of the 22 patients improved from >or= PS 3 at baseline to

Concentrations of clarithromycin and active metabolite in the epithelial lining fluid of patients with Mycobacterium avium complex pulmonary disease.

OBJECTIVES: Clarithromycin (CAM) is widely accepted for the treatment of Mycobacterium avium complex (MAC) pulmonary diseases. This study measured (a) the concentrations of CAM and its active metabolite (14OH-CAM) in bronchial epithelial lining fluid (ELF) obtained by bronchoscopic microsampling (BMS), and (b) the minimal inhibitory concentrations (MIC) of CAM for each MAC isolate. METHODS: We studied eight patients with MAC pulmonary disease treated with oral CAM, 400 (n=4) or 800 (n=4) mg/day. BMS was performed 3h after the last CAM dose, and the concentrations of CAM and 14OH-CAM were measured in ELF collected from the diseased and normal contralateral pulmonary segments, and in serum. RESULTS: The mean+/-SEM ELF concentrations of CAM (23.85+/-7.64 microg/ml) and CAM+14OH-CAM (28.71+/-8.37 microg/ml) in the 800 mg/day treatment group were significantly higher than in the 400mg/day group (7.48+/-2.58 microg/ml and 9.63+/-2.99 microg/ml, respectively; both p<0.05), while the serum concentrations were similar in both groups. In both treatment groups, the mean ELF concentrations sampled from diseased and normal segments were higher than the MIC against MAC isolates. The mean ELF concentration of CAM in the 400mg/day treatment group was <8 mircog/ml (the breakpoint concentration of CAM against M. avium recommended by the Clinical Laboratories Standards Institute), while the mean concentration in the 800 mg/day treatment group was >8 microg/ml. CONCLUSION: These observations suggest that CAM, 800 mg/day, is an appropriate dose to treat MAC pulmonary disease, and prevent its spread from diseased to non-diseased lung segments.

Randomized phase II trial comparing amrubicin with topotecan in patients with previously treated small-cell lung cancer: North Japan Lung Cancer Study Group Trial 0402.

PURPOSE: Amrubicin, a new anthracycline agent, and topotecan are both active for previously treated small-cell lung cancer (SCLC). No comparative study of these agents has been reported. This randomized phase II study was conducted to select amrubicin or topotecan for future evaluation. PATIENTS AND METHODS: Patients with SCLC previously treated with platinum-containing chemotherapy were randomly assigned to receive amrubicin (40 mg/m(2) on days 1 through 3) or topotecan (1.0 mg/m(2) on days 1 through 5). Patients were stratified by Eastern Cooperative Oncology Group performance status (0, 1, or 2) and type of relapse (chemotherapy sensitive or refractory). The primary end point was overall response rate (ORR), and secondary end points were progression-free survival (PFS), overall survival, and toxicity profile. RESULTS: From February 2004 to July 2007, 60 patients were enrolled, and 59 patients (36 patients with sensitive and 23 patients with refractory relapse) were assessable for efficacy and safety evaluation. Neutropenia was severe, and one treatment-related death owing to infection was observed in the amrubicin arm. ORRs were 38% (95% CI, 20% to 56%) for the amrubicin arm and 13% (95% CI, 1% to 25%) for the topotecan arm. In sensitive relapse, ORRs were 53% for the amrubicin arm and 21% for the topotecan arm. In refractory relapse, ORRs were 17% for the amrubicin arm and 0% for the topotecan arm. Median PFS was 3.5 months for patients in the amrubicin arm and 2.2 months for patients in the topotecan arm. Multivariate analysis revealed that amrubicin has more influence than topotecan on overall survival. CONCLUSION: Amrubicin may be superior to topotecan with acceptable toxicity for previously treated patients with SCLC. Further evaluation of amrubicin for relapsed SCLC is warranted.

Prolonged survival of patients with lung adenocarcinoma expressing XAGE-1b and HLA class I antigens.

XAGE-1b is a cancer/testis antigen that has been shown to be expressed at a significant frequency and to be immunogenic in non-small cell lung cancer (NSCLC). In the present study, we investigated correlations between XAGE-1b expression and NSCLC patient survival. XAGE-1b expression was examined immunohistochemically using USO9-13, an anti-XAGE-1b monoclonal antibody, in 121 NSCLCs (83 adenocarcinomas and 38 other histological types). XAGE-1b expression was observed in 27 (32.5%) adenocarcinoma specimens. In the other histological types, positive staining was observed in only 1 specimen. HLA class I expression in these samples was assessed previously. XAGE-1b expression had no correlation with overall survival. However, both XAGE-1b and HLA class I expression correlated with prolonged survival (P = 0.019). Moreover, expression of XAGE-1b combined with down-regulated HLA class I expression correlated with poor survival (P = 0.01). The density of cancer nest-infiltrating CD8+ T-cells in tumors expressing both XAGE-1b and HLA class I was higher than that in other groups. The findings suggest that XAGE-1b and HLA class I expression elicited a CD8+ T-cell response against minimal residual disease after surgery and resulted in prolonged survival of NSCLC patients.

Efficacy and safety of pemetrexed in combination with cisplatin for malignant pleural mesothelioma: a phase I/II study in Japanese patients.

BACKGROUND: Pemetrexed in combination with cisplatin (Pem/Cis) is used globally for the treatment of malignant pleural mesothelioma (MPM). This Phase I/II study was conducted to determine the recommended dose (RD) (Phase I) of Pem/Cis, and evaluate the efficacy and safety (Phase II) in Japanese MPM patients. METHODS: Key eligibility criteria were histologic diagnosis of MPM incurable by surgery, no prior chemotherapy, and a performance status 0-1. Under full vitamin supplementation, pemetrexed was intravenously administered on Day 1 of a 21-day cycle, followed by cisplatin. A cohort of six patients, starting from pemetrexed 500 mg/m(2) and cisplatin 75 mg/m(2) (Level 1), were studied in the dose-escalation Phase I (Step 1). The RD determined in Step 1 was carried forward into Phase II (Step 2). Planned number of patients treated with Pem/Cis was 18-38. RESULTS: In Step 1, 13 patients were enrolled: seven in Level 1 and six in Level -1 (pemetrexed 500 mg/m(2), cisplatin 60 mg/m(2)). Two of six evaluable patients had dose-limiting toxicities (pneumonitis and neutropenia) in Level 1, establishing Level 1 as the RD. In Step 2, 12 patients were enrolled, for a total of 19 patients treated at the RD. Seven patients achieved a partial response among these patients, for a response rate of 36.8% (95% confidence interval: 16.3-61.6); overall survival was 7.3 months. One drug-related death occurred due to worsening of a pre-existing pneumonia. Common grade 3/4 toxicities were neutropenia and decreased-hemoglobin. CONCLUSION: The Pem/Cis combination provides promising activity and an acceptable safety profile for chemonaive Japanese MPM patients with the same recommend dosage and schedule used in rest of the world.

Surmounting tumor-induced immune suppression by frequent vaccination or immunization in the absence of B cells.

Tumor-induced immune suppression is one of the most difficult obstacles to the success of tumor immunotherapy. Here, we show that established tumors suppress CD8 T cell clonal expansion in vivo, which is normally observed in tumor-free mice upon antigen-specific glycoprotein (gp) 96-chaperone vaccination. Suppression of CD8 T-cell expansion by established tumors is independent of tumor-associated expression of the antigen that is recognized by the CD8-T-cell receptor. Vaccination of tumor-bearing mice is associated with increased cellular recruitment to the vaccine site compared with tumor-free mice. However, rejection of established, suppressive tumors required frequent (daily) gp96 vaccination. B cells are known to attenuate T helper cell-1 responses. We found that in B-cell deficient mice, tumor rejection of established tumors can be achieved by a single vaccination. Accordingly, in tumor-free B-cell deficient mice, cognate CD8 cytotoxic T lymphocyte clonal expansion is enhanced in response to gp96-chaperone vaccination. The data have implications for the study of tumor-induced immune suppression and for translation of tumor immunotherapy into the clinical setting. Frequent vaccination with cellular vaccines and concurrent B-cell depletion may greatly enhance the activity of anticancer vaccine therapy in patients.

Recurrent gefitinib-induced interstitial lung disease.

Gefitinib, the epidermal growth factor receptor tyrosine kinase inhibitor, is effective for patients with non-small cell lung cancer. However, a serious adverse effect, interstitial lung disease (ILD), has been reported. The re-administration of gefitinib might be considered when there is no other choice of treatment and a therapeutic effect can be expected; however, there is no published data on the safety of restarting gefitinib after its discontinuation in cases suspected of having gefitinib-induced ILD. We report a case with recurrent gefitinib-induced ILD, which suggests that re-administration of gefitinib should be considered cautiously in patients who have previously developed gefitinib-induced ILD.

Pharmacokinetics of clarithromycin in bronchial epithelial lining fluid.

BACKGROUND AND OBJECTIVE: BAL is an established technique for measuring antibiotic concentrations in the epithelial lining fluid (ELF) of the bronchiolar-alveolar regions. However, the results may not reflect concentrations in bronchial regions. Bronchoscopic microsampling (BMS) is a technique for repeated sampling of bronchial ELF. The objective of the present study was to determine the time versus concentration profile of clarithromycin and its active metabolite, 14-hydroxy-clarithromycin, in bronchial ELF, as determined by BMS. METHODS: BMS was performed at 1, 2, 3, 5 and 10 h after a single oral administration of 200 mg clarithromycin in five healthy volunteers. BAL was performed 3 h after administration to determine clarithromycin concentrations in alveolar ELF and alveolar macrophages (AM). RESULTS: The maximum concentration (C(max)) of clarithromycin was 0.36 +/- 0.07 mg/L in serum and 1.44 +/- 0.49 mg/L in bronchial ELF (P < 0.01). C(max) for 14-hydroxy-clarithromycin was 0.34 +/- 0.13 mg/L in serum and 0.68 +/- 0.34 mg/L in bronchial ELF. The area under the concentration-time curve from 0 to 10 h (AUC(0-10)) for clarithromycin was 2.10 +/- 0.49 mg.h/L for serum and 7.37 +/- 2.07 mg.h/L for bronchial ELF (P < 0.01). The concentrations of clarithromycin in alveolar ELF and AM, 3 h after oral administration, were 4.84 +/- 3.39 mg/L and 10.7 +/- 8.7 mg/L, respectively. CONCLUSIONS: A single oral dose of clarithromycin produces a significantly higher C(max) and AUC(0-10) for clarithromycin in bronchial ELF than in serum, and higher concentrations in alveolar ELF and AM than in serum. BMS might be useful for measuring the pharmacokinetic profile of clarithromycin in bronchial ELF.

Anti-OX40 monoclonal antibody therapy in combination with radiotherapy results in therapeutic antitumor immunity to murine lung cancer.

The therapeutic effect of agonistic anti-OX40 (CD134) monoclonal antibody (mAb) in combination with radiotherapy was evaluated in a murine lung cancer model. After intradermal transplantation of ovalbumin (OVA)-transfected Lewis lung carcinoma, C57BL/6 mice were irradiated locally with a single dose of 20 Gy in combination with an intratumoral injection of anti-OX40 mAb at 50 microg on day 4 after transplantation, which is when the major axis of the inoculated tumor reached a diameter of 7-9 mm. On days 8, 11, and 14, the tumor-bearing mice were further treated with the same dose of anti-OX40 mAb. Anti-OX40 mAb in combination with radiotherapy prolonged survival and provided greater efficacy than either single treatment against well-established tumors. An in vivo depletion study suggested that therapeutic immunity was mainly CD8(+) T-cell dependent. OX40(+)CD8(+) T cells were augmented in draining lymph nodes obtained from irradiated mice compared with those from non-irradiated mice. OVA-major histocompatibility complex tetramer(+) CD8(+) T cells had been strongly recruited to the draining lymph nodes obtained from mice treated with anti-OX40 mAb in combination with radiotherapy, and strong antigen-specific cytotoxicity was confirmed by a (51)Cr-release assay. Moreover, a tumor-rechallenge model indicated that this combination therapy induced durable tumor immunity. Thus, anti-OX40 mAb in combination with radiotherapy may potentially help the management of patients with lung cancer.

Steep dose-response relationship for stage I non-small-cell lung cancer using hypofractionated high-dose irradiation by real-time tumor-tracking radiotherapy.

PURPOSE: To investigate the clinical outcomes of patients with pathologically proven, peripherally located, Stage I non-small-cell lung cancer who had undergone stereotactic body radiotherapy using real-time tumor tracking radiotherapy during the developmental period. METHODS AND MATERIALS: A total of 41 patients (25 with Stage T1 and 16 with Stage T2) were admitted to the study between February 2000 and June 2005. A 5-mm planning target volume margin was added to the clinical target volume determined with computed tomography at the end of the expiratory phase. The gating window ranged from +/-2 to 3 mm. The dose fractionation schedule was 40 or 48 Gy in four fractions within 1 week. The dose was prescribed at the center of the planning target volume, giving more than an 80% dose at the planning target volume periphery. RESULTS: For 28 patients treated with 48 Gy in four fractions, the overall actuarial survival rate at 3 years was 82% for those with Stage IA and 32% for those with Stage IB. For patients treated with 40 Gy in four fractions within 1 week, the overall actuarial survival rate at 3 years was 50% for those with Stage IA and 0% for those with Stage IB. A significant difference was found in local control between those with Stage IB who received 40 Gy vs. 48 Gy (p = 0.0015) but not in those with Stage IA (p = 0.5811). No serious radiation morbidity was observed with either dose schedule. CONCLUSION: The results of our study have shown that 48 Gy in four fractions within 1 week is a safe and effective treatment for peripherally located, Stage IA non-small-cell lung cancer. A steep dose-response curve between 40 and 48 Gy using a daily dose of 12 Gy delivered within 1 week was identified for Stage IB non-small-cell lung cancer in stereotactic body radiotherapy using real-time tumor tracking radiotherapy.

Diagnosis of peripheral pulmonary lesions using a bronchoscope insertion guidance system combined with endobronchial ultrasonography with a guide sheath.

We developed a bronchoscope insertion guidance system that produces virtual images by extracting the bronchi by automatic threshold adjustment, and searching for the bronchial route to the determined target. We used this system in combination with a thin bronchoscope and endobronchial ultrasonography with a guide sheath (EBUS-GS), and evaluated its practicability, usefulness and safety. The subjects were 31 patients with 32 peripheral pulmonary lesions. Computed tomography (CT) data were transferred into this system, and virtual bronchial images were automatically produced by setting the lesion as the target. While virtual images with the target were displayed for comparison with real images by the system, a thin bronchoscope was advanced to the target bronchus. Transbronchial biopsy (TBB) was then performed by EBUS-GS. The system automatically produced virtual images to a median of fifth- (third- to seventh-) order bronchi. In all patients, the thin bronchoscope could be guided along the planned route, and observation to a median of fifth- (third- to seventh-) order bronchi was possible. Thirty lesions (93.8%) were successfully visualized by EBUS, and 27 (84.4%) could be pathologically diagnosed. In lesions < or =30mm in size, the EBUS visualization yield was 91.7% (22/24), and the diagnostic yield was 79.2% (19/24). The median total examination time was 22.3 (9.8-41.5) min. In summary, using the bronchoscope insertion guidance system, virtual images can be readily produced, and the bronchoscope can be successfully guided to the target. This method is promising as a routine examination method in the biopsy of peripheral pulmonary lesions.

Immunotherapy with dendritic cells pulsed with tumor-derived gp96 against murine lung cancer is effective through immune response of CD8+ cytotoxic T lymphocytes and natural killer cells.

BACKGROUND AND PURPOSE: Immunization with heat shock proteins, gp96, elicits specific protective immunity against parent tumors. However, it is marginally effective as a therapeutic tool against established tumors. In the present study, we evaluated the efficacy and mechanism of immunotherapy with bone marrow-derived dendritic cells (DCs) pulsed with tumor-derived gp96 against murine lung cancer. METHODS: Mice were transplanted subcutaneously with ovalbumin (OVA)-transfected Lewis Lung Cancer (LLC-OVA) cells and immunized with gp96 derived from LLC-OVA, DCs, or DCs pulsed with gp96 derived from LLC-OVA. RESULTS: The antitumor effect was significantly enhanced in the mice immunized with DCs pulsed with gp96 derived from LLC-OVA, compared to mice immunized with gp96 or DCs (P<0.05). The antitumor effect was significantly dependent on natural killer (NK) cells and CD8(+) cells and partially dependent on CD4(+) cells. Analysis by laser confocal microscopy demonstrated that gp96 was shown on the cell surface at 15 min, and after 30 min internalized in the endosomes and not in the endoplasmic reticulum or lysosomes. OVA-specific(+) CD8(+) cells were more readily recruited into the draining lymph nodes and higher CD8(+) cytotoxic T cell activity against LLC-OVA was observed in splenocytes from mice immunized with DCs pulsed with gp96 derived from LLC-OVA. Re-challenge of the surviving mice with LLC-OVA tumors after the initial tumor inoculation showed dramatic retardation in tumor growth. CONCLUSION: In conclusion, immunotherapy of DCs pulsed with tumor-derived gp96 against murine lung cancer is effective through immune response of CD8(+) cytotoxic T lymphocytes and NK cells.

Histopathologic consideration of fiducial gold markers inserted for real-time tumor-tracking radiotherapy against lung cancer.

PURPOSE: Internal fiducial gold markers, safely inserted with bronchoscopy, have been used in real-time tumor-tracking radiotherapy for lung cancer. We investigated the histopathologic findings at several points after the insertion of the gold markers. METHODS AND MATERIALS: Sixteen gold markers were inserted for preoperative marking in 7 patients who subsequently underwent partial resection of tumors by video-assisted thoracoscopic surgery within 7 days. RESULTS: Fibrotic changes and hyperplasia of type 2 pneumocytes around the markers were seen 5 or 7 days after insertion, and fibrin exudation without fibrosis was detected 1 or 2 days after insertion. CONCLUSIONS: Because fibroblastic changes start approximately 5 days after gold marker insertion, real-time tumor-tracking radiotherapy should be started >5 days after gold marker insertion.

Combination tumor immunotherapy with radiotherapy and Th1 cell therapy against murine lung carcinoma.

Mice bearing established Lewis lung carcinoma (LLC) expressing model tumor antigen, ovalbumin (OVA) (LLC-OVA) marginally responded to local radiotherapy, but none of the mice was cured. In contrast, treatment of the tumor-bearing mice with intratumoral injection of tumor-specific T helper type 1 (Th1) cells and tumor antigen (OVA) after radiotherapy dramatically prolonged the survival days and induced complete cure of the mice at high frequency (80%). Radiation therapy combined with Th1 cells or OVA alone showed no significant therapeutic activity against LLC-OVA. Such a strong therapeutic activity was not induced by intratumoral injection of Th1 cells plus OVA. Compared with other treatment, radiation therapy combined with Th1 cells and OVA was superior to induce the generation of OVA/H-2(b) tetramer(+) tumor-specific cytotoxic T lymphocyte (CTL) with a strong cytotoxicity against LLC-OVA in draining lymph node (DLN). Moreover, the combined therapy is demonstrated to inhibit the growth of tumor mass, which grew at contralateral side. These results indicated that radiotherapy combined with Th1 cell/vaccine therapy induced a systemic antitumor immunity. These findings suggested that combination therapy with radiotherapy and Th1 cell/vaccine therapy may become a practical strategy for cancer treatment.