Tetsuya Okunaka

Phase II clinical study of photodynamic therapy using mono-l-aspartyl chlorin e6 and diode laser for early superficial squamous cell carcinoma of the lung

Photofrin is the most commonly used photosensitizer for photodynamic therapy (PDT). The major side effect of Photofrin is cutaneous photosensitivity. A second generation photosensitizer, mono-L-aspartyl chlorin e6 (NPe6) has shown anti-tumor efficacy and rapid clearance from skin. Therefore, we conducted a phase II clinical study to investigate the anti-tumor effects and safety of NPe6 in patients with early superficial squamous cell carcinoma of the lung. Enrollment criteria consisted of endoscopically evaluated early stage lung cancer with normal chest X-ray and CT images, no lymph node or distant metastasis. Tumors were located no more peripherally than subsegmental bronchi, the peripheral margin had to visible, and the tumor size had to not more than 2 cm in diameter. The histologic type of the tumor had to squamous cell carcinoma. Laser irradiation (100 J/cm2) using a diode laser was performed at 4 h after administration of NPe6 (40 mg/m2). Among 41 patients with 46 lesions, 40 with 45 lesions were eligible for safety evaluation, and 35 patients with 39 lesions were judged as eligible for efficacy evaluation. No serious adverse drug reactions were observed. Disappearance of skin photosensitivity was recognized within 2 weeks in 28 of 33 patients (84.8%) and in all the other seven patients first tested at 15-18 days. Complete response (CR) was seen in 84.6% of lesions (82.9% of patients). This study demonstrated excellent anti-tumor effects and safety, especially low skin photosensitivity in patients with early stage lung cancer. PDT using the second generation photosensitizer NPe6 and a diode laser will likely become a standard modality of PDT for central type early superficial squamous cell carcinoma of the lung.

Photodynamic Therapy (PDT) for Lung Cancers

Photodynamic therapy (PDT), a treatment for cancer, uses a photosensitizer and laser irradiation to produce reactive oxygen in cells. In Japan, the United States, and many other countries, PDT is a treatment option for stage 0 (TisN0M0) and stage I (T1N0M0) centrally located early stage lung cancer. PDT can preserve lung function, can be repeated, and can be combined with other therapeutic modalities such as chemotherapy. Recently, mono-l-aspartyl chlorine e6 (NPe6, Laserphyrin), a second-generation photosensitizer with lower photosensitivity than Photofrin (porfimer sodium), was approved by the Japanese government and a phase II clinical study using NPe6 with a new diode laser demonstrated an excellent antitumor effect and low skin photosensitivity. We expect PDT to be widely employed in many fields and the applications of PDT to be extended because of the decreasing cost of laser equipment and lower systemic photosensitivity induced by the photosensitizer. The purpose of this review is to introduce not only recent clinical trials of PDT for centrally located early lung cancer, but also new applications of PDT for cases of peripheral-type, early-stage lung cancers. We also discuss the applications of PDT for advanced lung cancer and combined therapy using PDT and other treatments for lung cancer.

LOCALIZATION OF MONO‐L‐ASPARTYL CHLORIN e6 (NPe6) IN MOUSE TISSUES

Abstract It is known that HpD is retained longer by malignant tissue than normal tissue and is therefore a useful material for photodynamic therapy (PDT). Currently, vigorous research is being conducted throughout the world to discover a new material which can have greater cancer cell affinity than hematoporphyrin derivative (HpD) and will be used effectively for PDT. Investigation has been conducted to determine the spectral characteristics and cancer cell affinity of NPe6, a recently developed material.

A Comparison between Argon‐dye and Excimer‐dye Laser for Photodynamic Effect in Transplanted Mouse Tumor

Photodynamic therapy (PDT) utilizing a hematoporphyrin derivative (HpD) as a sensitizer has become a viable option for the local treatment of neoplastic disease. The argon‐dye laser system is commonly used as a light source in this treatment modality. The excimer‐dye laser, on the other hand, delivers high‐energy red light in a pulsatile fashion. In this investigation, we treated BALB/c mice bearing mouse kidney sarcoma cell tumors with PDT using HpD at the dose of 5 mg/kg body weight as a photosensitizer and either a standard argon‐dye laser or the pulsatile excimer‐dye laser as the light source. At equal light energy doses (50 J/cm2), necrotic changes at depths averaging 4 mm from the tumor surface were obtained with the argon‐dye laser (200 mW power output) while tumor necrosis at depths exceeding 15 mm from the tumor surface was obtained using the excimer‐dye laser (6 mJ/pulse, 5 Hz). To determine the best conditions for photoirradiation with the excimer‐dye laser, tumor‐bearing mice were treated with different total light doses (10, 30 and 50 J/cm2), dose rates (1, 3 and 6 mJ/cm2), and frequencies (5,15 and 50 Hz) of light exposure. Our results indicate that the optimal effects obtained with the excimer‐dye laser are related to the total light dose used and the dose rate, but not to the frequency of light exposure.

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‐α, 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‐α 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‐α 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‐α 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.

Photodynamic therapy for multiple primary bronchogenic carcinoma

In recent years, multiple primary lung cancers have been reported with greater frequency, partly as a result of technologic advances in the detection of lung cancer and therapeutic achievements in its management. Photodynamic therapy (PDT) is a relatively new therapy used with increasing frequency in the treatment of a wide variety of malignancies, including central lung cancers. In PDT, the differential retention of an injected photosensitizer by malignant tissue is exploited by treatment with a low‐power laser beam delivered endoscopically. Since 1980, 145 patients with central lung cancers, including 35 cases of endoscopically evaluated early‐stage lesions were treated with PDT at Tokyo Medical College. Thirteen of these 145 patients had multiple primary bronchogenic carcinomas, five cases of which were synchronous with the rest, metachronous. Three of 13 patients with multiple tumors had early‐stage lesions and were treated with endoscopic PDT alone. In the other ten cases, PDT was used to treat accessible early‐stage foci although operative excision was required for advanced lesions. Mean survival after PDT, alone or in combination with surgery, was 38 months (range, 14 to 87 months), and seven patients remain alive to date. It was concluded that PDT is useful in extending the therapeutic options for, and improving the prognosis of patients with, multiple primary bronchogenic carcinomas.

Photodynamic therapy of esophageal carcinoma.

SummaryPhotodynamic therapy (PDT) utilizing either hematoporphyrin derivative or Photofrin II is proving to be an effective modality in the treatment of early superficial (ES) or advanced invasive (AI) carcinoma of the esophagus. An argon-pumped dye laser was used to deliver 630 nm light via quartz fibers passed through the biopsy channel of a gastroscope after intravenous injection of photosensitizer. Between 1982 and 1989, 20 patients (ES=6; AI=14) were treated in this manner. Complete remission was obtained in 4 of 6 ES cases, and the mean survival after PDT alone or in combination with other therapy was 27 months. Five patients remain alive to date. In the AI group, significant remissions were obtained in 6 cases while partial remissions were observed in another 8. The mean dysphagia grade improved from 4.0 to 2.8. We conclude that PDT is efficacious in the treatment of ES esophageal cancer, where complete remission may be achieved, and as palliative therapy in advanced cases to alleviate dysphagia.

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.

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.

Histopathological evaluation of fluorescence bronchoscopy using resected lungs in cases of lung cancer.

Objective evaluation of the performance of autofluorescence bronchoscopy based on analysis of thin sections of the bronchus of resected lungs was performed and compared with the results of preoperative autofluorescence bronchoscopy. Conventional bronchoscopy and autofluorescence bronchoscopy were performed prior to surgery for lung cancer. Thin sections of the bronchus were obtained from the resected specimens. The thin sections were pathologically analyzed and the diagnostic accuracy of endoscopy was calculated. The subjects were 30 consecutive operable lung cancer cases who received white light and autofluorescence bronchoscopy before operation. A total of 163 thin sections of the bronchi in the resected lungs were made. The sensitivity of white light bronchoscopy for cancer was 90 and 31% for dysplasia. The respective figures for autofluorescence bronchoscopy were 97 and 50% for cancer and dysplasia. The specificity of white light and autofluorescence was 88 and 84%, respectively. The diagnostic accuracy of autofluorescence bronchoscopy was objectively confirmed. Autofluorescence examination showed better sensitivity for cancerous/precancerous lesions and the evaluation of the extent of cancer invasion was accurate.