Influence of angiogenetic factors and matrix metalloproteinases upon tumour progression in non-small-cell lung cancer.

We attempted to investigate immunohistochemical expression of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), platelet-derived growth factor (PD-ECGF), c-erbB-2, matrix metalloproteinase-2 (MMP-2), and MMP-9 using surgical specimens of 119 non-small-cell lung carcinoma (NSCLC) cases and to evaluate the relationship between the expression levels of each molecule and clinicopathological factors or prognosis. VEGF expression levels were significantly associated with the local invasion (P = 0.0001), lymph node involvement (pN-factor) (P = 0.0019), pathological stage (p-stage) (P = 0.0027) and lymphatic permeation (P = 0.0389). PD-ECGF expression levels were associated with pN-factor (P = 0.0347). MMP-2 expression levels were associated with pN-factor (P = 0.004) and lymphatic permeation (P = 0.0056). Also, MMP-9 expression levels showed a significant correlation to local invasion (P = 0.0012), pN-factor (P = 0.0093) and p-stage (P = 0.0142). Multivariate analysis showed VEGF to be the most related to local invasion (P = 0.0084), and MMP-2 was the only factor with significant independent impact on lymphatic permeation (P = 0.0228). Furthermore, log-rank analysis showed significant association with poor survival by VEGF, bFGF, MMP-2 and MMP-9. Especially, combined overexpression of VEGF and MMP-2 revealed poor prognosis, our study might provide a basis for the better evaluation of biological characteristics and a new therapeutic strategy based on chemotherapy.

[Preoperative PDT for early stage lung cancer accompanied with infiltration to the central airway].

For the treatment of central type lung cancers, it is often necessary to perform bronchoplasty or bi-lobectomy even in early stage cases in which tumor invasion is found at the bronchial bifurcation. To solve this contradictory situation, we applied PDT for central type early stage lung cancer to reduce the extent of superficial infiltration, enabling a simple surgical technique or decreasing the amount of resected lung parenchyma. Among 7 patients, the simplification of surgical technique and reduction of the range of resection were possible in 2 cases each respectively. However, these objectives were not satisfied in the remaining 3 cases and the operations performed were those that had been originally scheduled before PDT. The current problems are the establishment of appropriate laser irradiation technique and accurate assessment for extension of invasion.

Increased cytotoxic effects of photodynamic therapy in IL-6 gene transfected cells via enhanced apoptosis.

PDT has been reported to induce cancer cell expression of cytokines, such as IL-6 and TNF-alpha, but it has been unclear whether cytokine expression by cancer cells is directly related to the antitumor effect of PDT. We treated Lewis lung carcinoma (LLC) cells with a new photosensitizer, mono-L-aspartyl chlorin e6 (NPe6) and light from a diode laser and found that expression of the mRNA of IL-2, IL-6, and TNF-alpha was increased by NPe6-mediated-PDT 6 hr later. To elucidate the mechanism of the direct anti-tumor effect of cytokine expression, we examined the photosensitivity of cytokine-gene-transfected cells, namely LLC-IL-2, LLC-IL-6, and LLC-TNF-alpha cells, by MTT assay. The IL-6 gene transfected, LLC-IL-6 cells were significantly more sensitive to cytotoxic effects than the parent LLC cells and other cytokine gene-transfected cells. This finding indicates that IL-6 expression modulates cellular sensitivity to PDT and that IL-2 and TNF-alpha expressions does not. In addition, the apoptosis of LLC-IL-6 cells induced by NPe6-PDT was greater than in the other cells as determined by DNA fragmentation and staining of apoptotic nuclei. Because IL-6 has been reported to induce apoptosis by downregulating expression of Bcl-2, we analyzed the expression of apoptosis-related Bcl-2, Bax, and cytochrome C by Western blot analysis. Decreased expression of Bcl-2 and cytochrome C was observed in both LLC cells and LLC-IL-6 cells. Bax protein increased in a time-dependent manner, and the ratio of Bax to Bcl-2 rose markedly after PDT in LLC-IL-6 cells. These results suggest that the increased sensitivity of LLC-IL-6 cells to PDT-induced cytotoxicity results from the high ratio of Bax to Bcl-2 in the IL-6-dependent apoptotic pathway. In conclusion, IL-6 expression plays a role in cellular sensitivity to PDT, and combination of IL-6 and PDT may provide a new strategy for cancer treatment.

Early hilar lung cancer--risk for multiple lung cancers and clinical outcome.

In early hilar lung cancer patients, multiple lung cancers frequently develop. The clinical outcome of such patients were studied. A total of 91 patients, 88 men and three women, who were endoscopically diagnosed with early hilar lung cancer were studied retrospectively. Surgery was performed in 46 patients, while organ-sparing treatment, including photodynamic therapy (PDT), Nd-YAG (neodymium-yttrium, argon, garnet) laser vaporization, and radiotherapy, were done for 45 patients. During follow-up, newly developed lung cancers and/or malignancies in other organs were recorded. The average smoking index (cigarettes per day x years) was 1040. Synchronous and/or metachronous multiple lung cancers developed in 26/91 patients (28.6%). Malignancies in other organs were found in 12/91 (13.2%). The smoking index of patients with multiple lung cancers was significantly higher than for other patients. The overall 5 year survival rate was 70.7% in all patients, 76.0% in the surgery group, and 64.4% in the nonsurgery group. The lung cancer-specific 5 year survival rate was 89.8% in all patients, 89.3% in the surgery group, and 90.5% in the nonsurgery group. Early hilar lung cancer frequently accompanies other lung cancers or malignancies in other organs. A favorable prognosis can be obtained with organ-sparing treatment.

K-ras mutation in sputum of primary lung cancer patients does not always reflect that of cancerous cells.

K-ras mutation in sputum was examined using mutant-allele-specific amplification method among 100 primary lung cancer and 15 non-oncological patients. K-ras mutation was detected in 11 out of 59 adenocarcinoma cases (18.6%), 5 out of 32 squamous cell carcinoma cases (15.6%), 2 out of 4 large cell carcinoma cases (50.0%) and 3 out of 15 non-oncological disease cases (20.0%). In the 18 cases of primary lung cancer K-ras mutation was examined in both sputum and the resected specimen of the primary lesion. In 5 cases K-ras mutation in sputum was detected without K-ras mutation in primary lesion. Therefore, these findings suggested that K-ras mutation in sputum may not be directly related to that of the primary lesion.

[Preoperative photodynamic therapy for lung cancer].

In Japan, photodynamic therapy (PDT) for locally superficial hilar lung cancer is common and its indications have already been determined. In order to improve the survival rate for lung cancer patients, it is necessary to increase the number of operable cases and preserve pulmonary function, and it thus would be beneficial to reduce the extent of resection. In our institution, preoperative PDT has been employed as one option to reduce the extent of resection. Nineteen patients who were initially planned to undergo pneumonectomy underwent successful lobectomy. For multiple primary lung cancer lesions, combined use of PDT was carried out in 29 cases. For early-stage lung cancer, PDT was performed and for advanced lesions surgery was selected. These new options may contribute to the management of advanced lung cancer.

[Bronchofiberscopy].

New diagnostic modalities have been used in conjunction with endoscopy for early detection of lung cancer. Videoendoscope is routinely used instead of fiberoptic bronchoscope. Fluorescence diagnosis has been proved to be useful in detecting subtle lesions which might be invisible by conventional endoscopy in central airway. Also, a number of small peripheral lesions has increased by the helical CT. CT guided transbronchial lung as well as needle cytology are indicated for definitive diagnosis of such lesions. Endobronchial Ultrasonography is employed to evaluate the depth of cancer invasion of the bronchus and lymph node swelling around the bronchus. It should be helpful in staging of lung cancer and selecting therapy.

[Molecular biological diagnosis].

Biological behavior of lung cancer was evaluated by basic study. Malignancy Associated Change is the concept that the nuclear features of normal cells in the vicinity of cancer show subtle morphological difference from those of healthy individuals. The difference was recognized by high resolution cytometry and the expression of MAC cells was correlated with the degree of abnormality of chest diseases. Comparative genomic hybridization and fluorescence in situ hybridization were performed to investigate genetic abnormality of. Multiple genetic abnormalities and chromosomal instability showed poor prognosis. Two dimensional electrophoresis was employed to detect the expression of the specific protein of lung cancer. TAO2 was proved to be specific to well differentiated adenocarcinoma. Also, metabolic analysis will be employed for cell analysis.

Cellular immunologic parameters related to age, gender, and stage in lung cancer patients.

Several immunologic parameters have been reported to correlate with the clinicopathologic status of lung cancer patients. However, these studies were based on relatively small numbers of patients and often yielded conflicting results. We prospectively studied cellular immunologic parameters related to age, gender, and stage in lung cancer patients. We obtained pretreatment peripheral blood samples from 287 lung cancer patients. Lymphocyte subsets (percentage of lymphocytes positive for CD3, CD4, CD8, HLA-DR, or representing FcgammaR IIIa-positive T cells), natural killer (NK) cell activity, and lymphoblastogenesis (LB) after stimulation by phytohemagglutinin (PHA) were evaluated. Significant decline was seen in older patients in percentages of cells positive for CD3 or CD4, in the CD4/CD8 ratio and in LB. The percentage of FcgR IIIa-positive T cells increased with age. LB as well as CD4 positivity were significantly greater in women than in men. NK cell activity showed the greatest cytotoxic responses in stage IIIA, with significantly less response in stage IV than in IIIA. Node-negative patients showed higher reactivities for LB and lower positivity for HLA-DR than node-positive patients. Patients with no distant metastases had a higher level of NK cell activity than patients with distant metastases. Immune parameters are variously related to age, gender, and the stage in lung cancer patients, some may prove to be useful predictors of survival.

Localization of experimental submucosal esophageal tumor in rabbits by using mono-L-aspartyl chlorin e6 and long-wavelength photodynamic excitation.

BACKGROUND AND OBJECTIVE: To increase the applicability of photodynamic diagnosis with regard to deep-seated tumor, we illuminated tumors with a long-wavelength laser beam after photosensitization with mono-L-aspartyl chlorin e6 (NPe6). STUDY DESIGN/MATERIALS AND METHODS: Rabbits with VX2 esophageal tumors were divided into four groups. The control group was not treated, and the other three groups were injected with 1, 2.5, and 5 mg/kg mono-L-aspartyl chlorin e6 (NPe6), respectively. After excitation with a 664-nm laser beam (10 mW, 10 seconds), the fluorescence image and the relative fluorescence intensity (tumor/normal tissue) were recorded every 2 hours up to 8 hours by a newly developed diode laser endoscopic fluorescence imaging system. The tissue concentration of NPe6 was examined by high performance liquid chromatography at 2, 4, and 6 hours after injection with 1 and 5 mg/kg NPe6. RESULTS: The diode laser endoscopic fluorescence imaging system was able to selectively detect fluorescence from submucosal tumor by comparison with the surrounding normal mucosa after NPe6 injection. The fluorescence intensity correlated with NPe6 dose, selectively accumulated in the tumor tissue and relative intensity peaked at 6 hours after injection. No fluorescent images were detected in controls. CONCLUSION: Given intravenously, NPe6 at a dose of 5 mg/kg and excited with a 664-nm wavelength laser beam 6 hours later can define experimentally induced deep-seated esophageal carcinoma in rabbits, by using an endoscopic fluorescence imaging system.

An early stage diffuse B-cell lymphoma within a visible site of bronchofiberscope accompanied by peripheral lung cancer.

We report a case of non-Hodgkin's lymphoma found at the orifice of right B2 accompanied by peripheral lung cancer in a 66-year-old asymptomatic man. Chest X-ray films showed a mass shadow in the left lower lung field. Transbronchial lung biopsy of left S9 demonstrated squamous cell carcinoma. Simultaneously wall thickening at the orifice of the right B2 was found coincidentally. The biopsy specimen from that site showed non-Hodgkin's lymphoma (diffuse B-cell type). After left lower lobectomy, systemic chemotherapy was performed. It is rare for malignant lymphoma to be recognized bronchofiberscopically.

Comparison of endoscopic features of early-stage squamous cell lung cancer and histological findings.

Seventy cases with early-stage central-type squamous cell carcinoma were treated surgically between 1984 and 1993 in seven participating institutes. We classified endoscopic features of early-stage central-type squamous cell carcinoma into three types (hypertrophic type, nodular type and polypoid type). After surgery we investigated the relationship between endoscopic features and both the area of superficial extent and depth of carcinoma invasion based on histopathological investigations of the surgical specimens. In 66.7% of the hypertrophic type lesions cancer cells did not invade into the cartilaginous layer, and only 4.8% of this type showed tumour invasion beyond the bronchial cartilage. On the other hand, a few nodular and polypoid type cases showed in-situ carcinoma or carcinoma with invasion from the subepithelial layer to the muscle layer, and in approximately 20% the these types we observed carcinoma invasion beyond the cartilaginous layer, which was not suitable for photodynamic therapy. Also, concerning the greatest dimension 24 out of 35 lesions (68.6%) less than 10 mm in the greatest dimension were evaluated as either in-situ carcinoma or micro-invasive tumour within the muscle layer. The endoscopic features can provide a basis for the determination of therapeutic strategy in early-stage central-type lung cancer.

The relationship between p21/waf1 expression patterns and cell proliferation during the tumorigenesis of the bronchus.

Each developmental stage in the process towards bronchial squamous cell carcinoma (normal epithelium, squamous metaplasia and early stage squamous cell carcinoma including in situ carcinoma) was examined for p21/waf1 protein expression and cell proliferation using MIB1. P21/waf1 immunoreactivity was classified into four patterns: predominantly cytoplasmic staining, exclusively nuclear staining, both nuclear and cytoplasmic staining and negative. The cases with predominantly cytoplasmic staining showed suppression of cell proliferation. Most cases with either negative or exclusively nuclear staining revealed high cell proliferation. The simultaneous evaluation of p21/waf1 and cell proliferation is valuable for clinical determination of the high risk for malignant transformation.

Cytologic features based on the expression of E-cadherin and catenins in lung adenocarcinoma.

OBJECTIVE: To investigate the relationship between E-cadherin-associated cell-to-cell adhesion and cytologic features in preoperative cytologic lung adenocarcinoma specimens. STUDY DESIGN: Evaluation of the relationship between cell-to-cell adhesion, formation of cellular clusters and frequency of single cells in 31 cases of primary lung adenocarcinoma, collected by brush and needle cytology preoperatively. RESULTS: Most cases with remarkable overlapping of cells in compact cellular clusters and a few solitary cells maintained cell-to-cell adhesion. Cellular clusters that had a slight tendency to overlap, a small cell-to-cell adhesion area and a high frequency of solitary cells tended to lack E-cadherin-associated cell-to-cell adhesion. CONCLUSION: Formation of cellular clusters and the appearance of solitary cancer cells are closely related to E-cadherin-associated cell-to-cell adhesion. Therefore, it is highly likely that cytologic features may indicate malignant behavior, such as local invasion and lymph node metastasis, in primary lung adenocarcinoma.

Early detection of bronchial lesions using lung imaging fluorescence endoscope.

The performance of the Lung Imaging Fluorescence Endoscope (LIFE) system was compared with conventional bronchoscopy in 158 patients: 68 patients with invasive cancer, 42 patients with abnormal sputum cytology findings (12 early cancer and 26 dysplasia), 17 cases with resected lung cancer and 31 smokers with symptoms. The respective results of conventional bronchoscopy and LIFE for detection of dysplasia were; sensitivity 52% and 90% (biopsy basis), 62% and 92% (patient basis). Fluorescence bronchoscopy may be an important adjunct to conventional bronchoscopy to improve the localization of subtle lesions of bronchus.

Early Detection of Bronchial Lesions Using System of Autofluorescence Endoscopy (SAFE) 1000.

Recently several endoscopic fluorescence detection systems have been developed. In some of them, laser light was used for the excitation of autofluorescence, and sophisticated techniques were also necessary to amplify the fluorescence signal as well.The result of fluorescence diagnosis using a simple system with a conventional Xenon lamp excitation and an image intensifier is reported. The respective results of sensitivity and positive predictive values of cancer plus dysplasia were 66%, and 62% by standard bronchoscopy and 92% and 88% by the newly developed autofluorescence system. In this paper, developed endoscope for detection of tissue/mucosal autofluorescence without the application of any photosensitizing agents or use of any lasers is evaluated.