A combination therapy of photodynamic therapy (PDT) and airway stent placement using a transparent silicone stent.

The aim of this study was to evaluate the feasibility of a combination therapy of photodynamic therapy (PDT) and airway stent placement using a transparent silicone stent (gold studded stent [GSS]). Laser irradiation (664 nm, continuous wave) was performed through the GSS using a straight and cylindrical fiber 1.0 cm away from a power meter. There are two types of GSS: the TD type for the trachea and the BD type for the bronchus. Laser outputs were set to 150 mW, 180 mW, 210 mW, 240 mW, 270 mW, and 300 mW. The laser powers passing through the both types of GSS were measured three times for each outputs and the averages were calculated. Based on the results, animal experiment was performed using two female pigs. Under general anesthesia, a GSS (BD type) was inserted into trachea of pigs, and PDT using NPe6 as a photosensitizer was performed by 100 J/cm2 laser irradiation on parts of the trachea with and without a GSS. Immediately after and 1 week after PDT, pig tracheas were harvested and histological analysis was performed. Histological analysis of areas with or without the stent showed edematous changes between the cartilage and submucosal layer immediately after PDT, and necrotic changes 1 week later. The effectiveness of NPe6-PDT for pigs' trachea covered by the stent was same as trachea without the stent. The use of a GSS may enable PDT to be effective even in the area covered by the stent.

Tumor volume determines the feasibility of cell-free DNA sequencing for mutation detection in non-small cell lung cancer.

Next-generation sequencing (NGS) and digital PCR technologies allow analysis of the mutational profile of circulating cell-free DNA (cfDNA) in individuals with advanced lung cancer. We have now evaluated the feasibility of cfDNA sequencing for mutation detection in patients with non-small cell lung cancer at earlier stages. A total of 150 matched tumor and serum samples were collected from non-small cell lung cancer patients at stages IA-IIIA. Amplicon sequencing with DNA extracted from tumor tissue detected frequent mutations in EGFR (37% of patients), TP53 (39%), and KRAS (10%), consistent with previous findings. In contrast, NGS of cfDNA identified only EGFR, TP53, and PIK3CA mutations in three, five, and one patient, respectively, even though adequate amounts of cfDNA were extracted (median of 4936 copies/mL serum). Next-generation sequencing showed a high accuracy (98.8%) compared with droplet digital PCR for cfDNA mutation detection, suggesting that the low frequency of mutations in cfDNA was not due to a low assay sensitivity. Whereas the yield of cfDNA did not differ among tumor stages, the cfDNA mutations were detected in seven patients at stages IIA-IIIA and at T2b or T3. Tumor volume was significantly higher in the cfDNA mutation-positive patients than in the negative patients at stages T2b-T4 (159.1 ± 58.0 vs. 52.5 ± 9.9 cm3 , P = 0.014). Our results thus suggest that tumor volume is a determinant of the feasibility of mutation detection with cfDNA as the analyte.

[Photodynamic Therapy for Lung Cancer].

In Japan, Photodynamic therapy (PDT) is recommended as a treatment option for centrally located early-stage lung cancers (CLELCs). It is a minimally invasive treatment with excellent anti-tumor effects. The 2nd generation photosensitizer, talaporfin sodium has strong anti-tumor effects with much less photosensitivity than porfimer sodium. Moreover, the laser equipment is compact and portable, and talaporfin sodium is now the current mainstay of PDT for lung cancer. For successful PDT, accurate evaluation of tumor extent and bronchial invasion is crucial. Detailed examination of the tumor using autofluorescence bronchoscopy and endobronchial ultrasonography or optical coherence tomography is extremely useful before PDT. At present, PDT has become the 1st choice of treatment for CLELC in institutions with the necessary equipment. It can also be effective for advanced lung cancer causing tracheobronchial obstruction. With such advances in PDT for CLELC, we are expanding the indications of PDT for not only CLELC, but also peripheral type lung cancer.

DNA methylation is globally disrupted and associated with expression changes in chronic obstructive pulmonary disease small airways.

DNA methylation is an epigenetic modification that is highly disrupted in response to cigarette smoke and involved in a wide spectrum of malignant and nonmalignant diseases, but surprisingly not previously assessed in small airways of patients with chronic obstructive pulmonary disease (COPD). Small airways are the primary sites of airflow obstruction in COPD. We sought to determine whether DNA methylation patterns are disrupted in small airway epithelia of patients with COPD, and evaluate whether changes in gene expression are associated with these disruptions. Genome-wide methylation and gene expression analysis were performed on small airway epithelial DNA and RNA obtained from the same patient during bronchoscopy, using Illumina's Infinium HM27 and Affymetrix's Genechip Human Gene 1.0 ST arrays. To control for known effects of cigarette smoking on DNA methylation, methylation and gene expression profiles were compared between former smokers with and without COPD matched for age, pack-years, and years of smoking cessation. Our results indicate that aberrant DNA methylation is (1) a genome-wide phenomenon in small airways of patients with COPD, and (2) associated with altered expression of genes and pathways important to COPD, such as the NF-E2-related factor 2 oxidative response pathway. DNA methylation is likely an important mechanism contributing to modulation of genes important to COPD pathology. Because these methylation events may underlie disease-specific gene expression changes, their characterization is a critical first step toward the development of epigenetic markers and an opportunity for developing novel epigenetic therapeutic interventions for COPD.

[Present status of interventional pulmonology].

Endobronchial intervention has been well established in therapeutic and palliative methods for patients with malignant and benign airway obstruction. Central airway tumor obstruction could lead to significant dyspnea or hemoptysis and dramatically lower the quality of life. Such symptoms can be relieved immediately via bronchoscopic intervention. Endobronchial therapies such as neodymium yttrium aluminum garnet( Nd:YAG) laser therapy, argon plasma coagulation and microwave coagulation or stent placement are well-established treatment options. Each of these modalities has its specific advantages and disadvantages. Laser therapy is direct thermal ablation applied to endobronchial tumors. It has sufficient power to vaporize tissues and produces an excellent coagulation effect. However, the risk of perforation is high. On the other hand, photodynamic therapy has the curative potential for patients with early superficial lung cancer using low power laser. Moreover, it has recently been approved for the treatment of other cancers including advanced lung cancer. Stent placement is an effective treatment to re-establish airway patency and immediately relieve the patient from life-threatening dyspnea. Stent restenosis, migration, mucus plug and hemoptysis are common short-term and longterm problems. It is important to select the appropriate treatment method for each patient to avoid complications and achieve successful results.

Frontiers in bronchoscopic imaging.

Bronchoscopy is a minimally invasive method for diagnosis of diseases of the airways and the lung parenchyma. Standard bronchoscopy uses the reflectance/scattering properties of white light from tissue to examine the macroscopic appearance of airways. It does not exploit the full spectrum of the optical properties of bronchial tissues. Advances in optical imaging such as optical coherence tomography (OCT), confocal endomicroscopy, autofluorescence imaging and laser Raman spectroscopy are at the forefront to allow in vivo high-resolution probing of the microscopic structure, biochemical compositions and even molecular alterations in disease states. OCT can visualize cellular and extracellular structures at and below the tissue surface with near histological resolution, as well as to provide three-dimensional imaging of the airways. Cellular and subcellular imaging can be achieved using confocal endomicroscopy or endocytoscopy. Contrast associated with light absorption by haemoglobin can be used to highlight changes in microvascular structures in the subepithelium using narrow-band imaging. Blood vessels in the peribronchial space can be displayed using Doppler OCT. Biochemical compositions can be analysed with laser Raman spectroscopy, autofluorescence or multispectral imaging. Clinically, autofluorescence and narrow-band imaging have been found to be useful for localization of preneoplastic and neoplastic bronchial lesions. OCT can differentiate carcinoma in situ versus microinvasive cancer. Endoscopic optical imaging is a promising technology that can expand the horizon for studying the pathogenesis and progression of airway diseases such as COPD and asthma, as well as to evaluate the effect of novel therapy.

Effect of photodynamic therapy (PDT) on a rat model of bleomycin-induced interstitial pneumonia.

BACKGROUND: Even if lung cancer is detected at an early stage, surgery may be difficult in patients with severe comorbidities, like interstitial pneumonia (IP). Radiation therapy cannot be performed due to the high risk of acute IP exacerbation. Therefore, an effective alternative, such as photodynamic therapy (PDT), is required. To prove that acute exacerbation is not induced after PDT in peripheral lung cancer, we investigated the effects of PDT on IP rat models. METHODS: Bleomycin (BLM) was administered intratracheally. Seven days after administration, left thoracotomy was performed. Talaporfin sodium was injected, and diode laser irradiation (664 nm, 150mW, 100J/cm2) was performed. Seven days after PDT, the whole blood and left lungs were collected. A total of 23 rats, comprising BLM + PDT (n = 4), BLM + non-PDT (n = 10), non-BLM + PDT (n = 2), non-BLM + non-PDT (n = 5), and two rats that died immediately after PDT were observed. Serum levels of Krebs von den Lungen-6, surfactant protein-D, lactate dehydrogenase, and serum C-reactive protein were measured. Fibrosis and macrophage scorings, and the ​​collagen fibers percentage were examined by staining with hematoxylin and eosin, Elastica van Gieson, anti-α smooth muscle antibody, and anti-CD68 antibodies. RESULTS: There was no remarkable difference in the values of each marker in fibrosis and macrophage scores with or without PDT. In case of death, fibrosis was mild, and PDT was not affected. CONCLUSIONS: In IP rat models, PDT did not induce lung fibrosis or acute exacerbation.

Molecular determinants of photodynamic therapy for lung cancers.

BACKGROUND AND OBJECTIVES: PDT induces apoptosis, inflammatory reactions, immune reactions, and damage to the microvasculature around the tumors. The mechanisms responsible for the anticancer effects of Photofrin-PDT and NPe6-PDT differ somewhat. To select a photosensitizer for lung cancer treatment and to improve the efficacy of PDT, the mechanisms of action for PDT using Photofrin or NPe6 must be elucidated and the phenomena validated by analyzing molecular determinants from clinical samples. STUDY DESIGN/MATERIALS AND METHODS: We examined the role of immunological reactions in the anti-tumor effects of PDT using cytokine-overexpressing cells and investigated whether the anti-apoptotic protein Bcl-2 may be a molecular target. Moreover, we investigated the association between ATP-binding cassette transporter proteins such as breast cancer-resistant protein (BCRP), which can pump out some types of photosensitizer, and the efficacy of PDT using clinical samples from 81 early lung cancer lesions treated with PDT between 1998 and 2006 at the Tokyo Medical University Hospital. RESULTS: Photofrin-PDT damaged Bcl-2 and rapidly induced apoptosis, but NPe6-PDT did not damage Bc-2 nor did it induce morphologically typical apoptosis. However, NPe6-PDT exerted a strong anti-tumor effect, regardless of the overexpression of Bcl-2. By analyzing the BCRP-overexpressing cells, Photofrin, but not NPe6, was found to be a substrate of BCRP. All 81 lung cancer lesions were BCRP-positive; as Photofrin was found to be a substrate of BCRP, the expression of BCRP significantly affected the efficacy of Photofrin-PDT. However, NPe6-PDT exerted a strong antitumor effect regardless of BCRP expression, and the complete response rate after NPe6-PDT was much higher than that after Photofrin-PDT. CONCLUSIONS: Our translational research suggests that NPe6-PDT may be superior to Photofrin-PDT for the treatment of lung caner, and individualized approaches to PDT based on the expression status of Bcl-2 and/or BCRP may improve the efficacy of PDT in patients with lung cancers.

Clinical trial of photodynamic therapy for peripheral-type lung cancers using a new laser device in a pilot study.

INTRODUCTION/AIM: Photodynamic therapy (PDT) involves the use of a tumor-specific photosensitizer and laser irradiation, and is one of the treatment options recommended for early centrally located lung cancers, but not yet for peripheral-type lung cancers. We developed a new laser probe, the composite-type optical fiberscope (COF), which allows accurate laser irradiation of a cancer lesion with simultaneous visualization of the lesion. In this study, we attempted a new endobronchial PDT technique using the new laser probe, and evaluated the effectiveness and feasibility of this novel PDT technique for peripheral lung cancers. METHODS: This phase I study was conducted in 7 patients with peripheral lung cancers (primary tumor ≤20 mm in diameter). We performed endobronchial PDT for these patients using the new laser probe and talaporfin sodium as the photosensitizer. RESULTS: We performed PDT for 3 patients with peripheral lung cancer using a laser dose of 50 J/cm2 at 120 mW, and confirmed the feasibility of using this dose. Then, we escalated the laser dose to 100 J/cm2 in 4 additional patients. A total of 7 patients met our inclusion criteria. Evaluation at 2 weeks and 3 months after the PDT revealed no complication such as pneumonia or pneumothorax. At the evaluation conducted 6 months later, we found CR in 3 cases and SD in the remaining 4 cases. CONCLUSION: PDT was found to be a feasible and non-invasive treatment modality for early peripheral-type lung cancer. In the future, PDT could become a standard treatment option for peripheral-type lung cancer.

[Laser therapy for endobronchial malignancies].

Photodynamic therapy (PDT), neodymium yttrium aluminum garnet (Nd-YAG) laser therapy, electrocautery and microwave coagulation therapy are therapeutic options available for management of endobronchial malignancies. All of these treatment modalities have been used for both palliation of late obstructing cancers, and more recently have been used as primary treatment of early stage lung cancers. Only PDT has the curative potential for patients with early superficial squamous cell carcinoma. Nd-YAG laser therapy is used for direct thermal ablation of tissue in endobronchial malignancy. This equipment is the most widely used type of laser for bronchoscopic interventions because it has sufficient power to vaporize tissues and produces an excellent coagulation effect. But the risks of perforation and bleeding are high. Endobronchial electrocautery is the use of high-frequency electrical current that generates heat due to tissue resistance, resulting in destruction of tissue. Argon plasma coagulation (APC) is a form of noncontact electrocoagulation. The risks of perforation and igniting are much lower than with the Nd-YAG laser therapy. Microwave coagulation therapy refers to the use of all electromagnetic methods for inducing tumor destruction by using devices with frequencies of 2450 MHz. It is important to select these treatment methods appropriately according to each case.

Tailor-made approach to photodynamic therapy in the treatment of cancer based on Bcl-2 photodamage.

It is very important to elucidate the mechanism of action and identify the molecular determinant of photodynamic medicine, in order to increase the number of clinical applications of photodynamic therapy (PDT) and perform personalized medicine. We have previously reported that PDT using some photosensitizers, such as phthalocyanine 4 (Pc 4) damages the anti-apoptotic protein Bcl-2, and that Bcl-2 is a molecular PDT target using a mitochondrion-targeting photosensitizer. In this study, we examined the molecular targets of Photofrin-PDT and NPe6-PDT, which are approved for early stage lung cancers by the Japanese Ministry of Health Labor and Welfare, by evaluating the photodamage to Bcl-2 using Western blot analysis. Our results showed that Photofrin-PDT damaged Bcl-2, induced morphologically typical apoptosis, and demonstrated equal sensitivity between MCF-7c3 cells (human breast cancer cells expressing stably transfected procaspase-3) and Bcl-2 overexpressing cells, MCF-7c3-GFP-Bcl-2 cells, with a clonogenic assay. However, NPe6-PDT did not damage Bcl-2 and took longer to induce typical apoptosis compared with Photofrin-PDT. MCF-7c3-GFP-Bcl-2 cells were considerably more resistant to the lethal effects of NPe6-PDT than parental MCF-7c3 cells. In conclusion, Photofrin-PDT damages different molecular targets, and our data indicate that the extent of Bcl-2 photodamage can determine the sensitivity of cancer cells to apoptosis and to overall cell killing caused by PDT using Photofrin, but not the lysosomal targeting NPe6. The application of these findings to clinical PDT may depend on the levels of the Bcl-2 proteins in the tumor being treated, and the tailor-made medicine based on the Bcl-2 photodamage may overcome any resistance afforded by elevated amounts of Bcl-2.

Photodynamic therapy for lung cancers based on novel photodynamic diagnosis using talaporfin sodium (NPe6) and autofluorescence bronchoscopy.

BACKGROUND: We had previously developed the possibility of use of a photodynamic diagnosis (PDD) system using a tumor-selective photosensitizer and laser irradiation for the early detection and photodynamic therapy (PDT) for centrally located early lung cancers. Recently, we established the autofluorescence diagnosis system integrated into a videoendoscope (SAFE-3000) as a very useful technique for the early diagnosis of lung cancer. PATIENTS AND METHODS: Twenty-nine patients (38 lesions) with centrally located early lung cancer received PDD and PDT using the second-generation photosensitizer, talaporfin sodium (NPe6). Just before the PDT, we defined the tumor margin accurately using the novel PDD system SAFE-3000 with NPe6 and a diode laser (408nm). RESULTS: Red fluorescence emitted from the tumor by excitation of the photosensitizer by the diode laser (408nm) from SAFE-3000 allowed accurate determination of the tumor margin just before the PDT. The complete remission (CR) rate following NPe6-PDT in the cases with early lung cancer was 92.1% (35/38 lesions). We also confirmed the loss of red fluorescence from the tumors immediately after the PDT using SAFE-3000. We confirmed that all the NPe6 in the tumor had been excited and photobleached by the laser irradiation (664nm) and that no additional laser irradiation was needed for curative treatment. CONCLUSIONS: This novel PDD system using SAFE-3000 and NPe6 improved the quality and efficacy of PDT and avoided misjudgement of the dose of the photosensitizer or laser irradiation in PDT. PDT using NPe6 will become a standard option of treatments for centrally located early lung cancer.

Skin fluorescence following photodynamic therapy with NPe6 photosensitizer.

BACKGROUND: The second-generation photosensitizer NPe6 has strong anti-tumor effects with a much shorter photosensitive period than the first-generation photosensitizer Photofrin. Although photosensitive period has been reduced, skin photosensitivity is still a major side effect of photodynamic therapy (PDT). Therefore, we conducted a prospective study to investigate whether the NPe6 fluorescence intensity in skin after PDT could be measured effectively in human patients to improve the management of a patient's photosensitive period. METHODS: The NPe6 fluorescence measurements using a constructed fluorescence sensing system at the inside of the arm were acquired prior to and 5 and 10min after NPe6 administration as well as at the time of PDT (4-5h after administration), at discharge (2 or 3days after PDT), and at 1 or 2 weeks after PDT. Participants were interviewed as to whether they had any complications at 2 weeks after PDT. RESULTS: Nine male patients and one female patient entered this study. Nine patients were inpatients and one patient was an outpatient. All of the measurements of NPe6 fluorescence in the skin could be obtained without any complications. The spectral peak was detected at the time of discharge (2-3days after administration) in most cases and it decreased at 1 or 2 weeks after PDT. CONCLUSIONS: The fluorescence of NPe6 in the skin could be detected feasibly using the fluorescence sensing system in human patients. Measuring the relative concentration of NPe6 in the skin indirectly by measuring fluorescence intensity might be useful to predict the period of skin photosensitivity after PDT.

Lysosomal cathepsin initiates apoptosis, which is regulated by photodamage to Bcl-2 at mitochondria in photodynamic therapy using a novel photosensitizer, ATX-s10 (Na).

ATX-s10 is a novel and second-generation photosensitizer for photodynamic therapy (PDT). In order to conduct clinical trials of ATX-s10-PDT and/or extend its clinical applications, it is very important to elucidate the mechanisms of the action of ATX-s10-PDT. We examined the apoptic response against ATX-s10-PDT using a Bcl-2 or Bcl-2 mutant overexpressing cells. Using fluorescent microscopy, ATX-s10 localized not only to mitochondria but also to lysosomes and possibly other intracellular organelles, but not to the plasma membrane or the nucleus. These results suggest that ATX-s10-PDT can damage mitochondria and lysosomes. By Western blot analysis, ATX-s10-PDT damaged Bcl-2, which localized preferentially at mitochondrial membranes, and caused Bcl-2 to cross-link immediately after laser irradiation. However, ATX-s10-PDT was not able to rapidly induce morphologically typical apoptosis (i.e. chromatin condensation and fragmentation) as PDT using mitochondria targeted photosensitizers, such as phthalocyanine 4 (Pc 4). Pharmacological inhibitions of lysosomal cytokine protease cathepsins, such as cathepsin B and D, protected MCF-7c3 cells (human breast cancer cells expressing stably transfected procaspase-3) from apoptosis caused by ATX-s10-PDT. Overexpression of wild-type Bcl-2 or Bcl-2Delta33-54 resulted in relative resistance of cells to ATX-s10-PDT, as assessed by the degree of morphological apoptosis or loss of clonogenicity. We conclude that lysosomal damage by ATX-s10-PDT can initiate apoptotic response and this apoptotic pathway can be regulated by photodamage to Bcl-2 via mitochondrial damage.

Combination effect of photodynamic therapy using NPe6 with pemetrexed for human malignant pleural mesothelioma cells.

To identify a possible new treatment modality for malignant pleural mesothelioma (MPM), we examined whether combination treatment consisting of pemetrexed chemotherapy and photodynamic therapy (PDT) using the photosensitizer NPe6, enhanced the antitumor effect in both in vitro and in vivo models. We also investigated preclinical treatment schedules. Four human malignant mesothelioma cell lines (MSTO­211H, H2052, H2452 and H28) were assayed using the WST assay after treatment with pemetrexed and NPe6­PDT. The treatment schedule for the combination treatment was examined using nude mice. Pemetrexed pre­treatment enhanced the lethal effect of NPe6­PDT in the four malignant mesothelioma cell lines, but NPe6­PDT followed by pemetrexed treatment did not enhance cell lethality in the in vitro assay. Pemetrexed pre­treatment did not enhance the intracellular accumulation of NPe6, which is one of the determinants of the antitumor effect of PDT. In nude mice injected with MSTO­211H cells and then treated using a combination of pemetrexed and NPe6­PDT (10 mg/kg NPe6, 10 J/cm(2) laser irradiation), the tumor volume decreased by 50% but subsequently increased, reaching the pre­treatment value after 14 days. Pemetrexed treatment followed by NPe6­PDT resulted in an 80% reduction in the tumor size and inhibited re­growth. NPe6­PDT followed by pemetrexed treatment resulted in a 60% reduction in tumor size but did not inhibit re­growth. NPe6­PDT induced the expression of thymidylate synthase (TS), which confers resistance to pemetrexed, and NPe6­PDT followed by pemetrexed treatment did not enhance the treatment outcome in vivo. In conclusion, combination treatment, consisting of pemetrexed followed by NPe6­PDT, should be further investigated as a new treatment modality for MPM. In the future, this combination treatment may contribute to a reduction in local recurrence and a prolonged survival period in patients with MPM.

Reproducibility of optical coherence tomography airway imaging.

Optical coherence tomography (OCT) is a promising imaging technique to evaluate small airway remodeling. However, the short-term insertion-reinsertion reproducibility of OCT for evaluating the same bronchial pathway has yet to be established. We evaluated 74 OCT data sets from 38 current or former smokers twice within a single imaging session. Although the overall insertion-reinsertion airway wall thickness (WT) measurement coefficient of variation (CV) was moderate at 12%, much of the variability between repeat imaging was attributed to the observer; CV for repeated measurements of the same airway (intra-observer CV) was 9%. Therefore, reproducibility may be improved by introduction of automated analysis approaches suggesting that OCT has potential to be an in-vivo method for evaluating airway remodeling in future longitudinal and intervention studies.

Validation of airway wall measurements by optical coherence tomography in porcine airways.

Examining and quantifying changes in airway morphology is critical for studying longitudinal pathogenesis and interventions in diseases such as chronic obstructive pulmonary disease and asthma. Here we present fiber-optic optical coherence tomography (OCT) as a nondestructive technique to precisely and accurately measure the 2-dimensional cross-sectional areas of airway wall substructure divided into the mucosa (WAmuc), submucosa (WAsub), cartilage (WAcart), and the airway total wall area (WAt). Porcine lung airway specimens were dissected from freshly resected lung lobes (N = 10). Three-dimensional OCT imaging using a fiber-optic rotary-pullback probe was performed immediately on airways greater than 0.9 mm in diameter on the fresh airway specimens and subsequently on the same specimens post-formalin-fixation. The fixed specimens were serially sectioned and stained with H&E. OCT images carefully matched to selected sections stained with Movat's pentachrome demonstrated that OCT effectively identifies airway epithelium, lamina propria, and cartilage. Selected H&E sections were digitally scanned and airway total wall areas were measured. Traced measurements of WAmuc, WAsub, WAcart, and WAt from OCT images of fresh specimens by two independent observers found there were no significant differences (p>0.05) between the observer's measurements. The same wall area measurements from OCT images of formalin-fixed specimens found no significant differences for WAsub, WAcart and WAt, and a small but significant difference for WAmuc. Bland-Altman analysis indicated there were negligible biases between the observers for OCT wall area measurements in both fresh and formalin-fixed specimens. Bland-Altman analysis also indicated there was negligible bias between histology and OCT wall area measurements for both fresh and formalin-fixed specimens. We believe this study sets the groundwork for quantitatively monitoring pathogenesis and interventions in the airways using OCT.

In vivo lung microvasculature visualized in three dimensions using fiber-optic color Doppler optical coherence tomography.

For the first time, the use of fiber-optic color Doppler optical coherence tomography (CDOCT) to map in vivo the three-dimensional (3-D) vascular network of airway segments in human lungs is demonstrated. Visualizing the 3-D vascular network in the lungs may provide new opportunities for detecting and monitoring lung diseases such as asthma, chronic obstructive pulmonary disease, and lung cancer. Our CDOCT instrument employs a rotary fiber-optic probe that provides simultaneous two-dimensional (2-D) real-time structural optical coherence tomography (OCT) and CDOCT imaging at frame rates up to 12.5 frames per second. Controlled pullback of the probe allows 3-D vascular mapping in airway segments up to 50 mm in length in a single acquisition. We demonstrate the ability of CDOCT to map both small and large vessels. In one example, CDOCT imaging allows assignment of a feature in the structural OCT image as a large (∼1 mm diameter) blood vessel. In a second example, a smaller vessel (∼80 µm diameter) that is indistinguishable in the structural OCT image is fully visualized in 3-D using CDOCT.

Klotho is a novel biomarker for good survival in resected large cell neuroendocrine carcinoma of the lung.

BACKGROUND: In terms of prognosis, large cell neuroendocrine carcinoma (LCNEC) differs distinctively from other non-small cell lung cancers, with the prognosis of LCNEC being poor, even for early-stage disease. Improvements in survival require a biomarker capable of defining a subset of patients destined to do poorly so that these patients can be targeted for additional therapies, including chemotherapy. In this study, we focused on the Klotho gene, which is an anti-aging gene known to be a potential tumor suppressor. We investigated whether the immunohistochemical expression of Klotho can predict survival patients with resected LCNEC. METHODS: The histological characteristics of patients receiving an initial diagnosis of LCNEC (n=30) at Tokyo Medical University Hospital were retrospectively reviewed, and multiple variables including stage, lymphangioinvasion, lymph node status and the expression of Klotho as identified using an immunohistochemical analysis, were assessed. RESULTS: Immunostaining for Klotho was mostly cytoplasmic, and Klotho expression was seen in 10 patients (33.3%) but not in 20 patients (66.7%). The expression of Klotho was significantly associated with a good outcome of resected patients with LCNEC and Klotho(-) was associated with increased LCNEC risk by multivariate analysis (hazard ratio 4.92, 95% confidence interval 1.04-23.24, p=0.044). Neither lymph node status nor lymphangioinvasion were significantly associated with a poor survival. However, among patients without lymph node metastasis or angioinvasion, the survival benefit of Klotho expression in the primary tumor was significantly higher, compared with that of patients without Klotho expression. CONCLUSION: Klotho staining provides a new biomarker for a good outcome in patients with LCNEC, especially among patients without lymph node metastasis or lymphangioinvasion.