An ultrastructural study of human bladder cancer treated by photodynamic therapy

An electron microscopic study has been carried out biopsy material of transitional cell carcinoma of human bladder, taken before and after photodynamic therapy (PDT), from 14 patients. It has been demonstrated that bladder cancer is highly sensitive to PDT using haematoporphyrin derivative (HPD, made in China) and laser irradiation. It was found that vascular endothelium within the tumour tissue was very sensitive to PDT, showing distinct changes as early as immediately after completion of a 20-min irradiation. Twenty-four hours after PDT, almost all the capillaries examined were necrotic and broken down into small fragments. The cancer cells were less sensitive to PDT, being damaged later and less seriously than the blood vessels. It has been concluded that in PDT-treated tumours the vascular endothelium is damaged primarily while the cancer cells are destroyed, to a considerable extent, secondarily as the consequence of structural damage to capillaries and functional disturbance in the microcirculation. We would speculate that the main factor influencing the final response is the actual concentration of HPD in various types of cell in the tumours.     

Ultrastructural changes in normal rat colon induced by photodynamic therapy

Despite the fact that photodynamic therapy (PDT) is a promising new technique for the treatment of gastrointestinal tumours, its effect on normal mucosa has so far received little attention. With a view to understanding the changes that occur in normal colon mucosa, we have examined the ultrastructure of normal rat colon mucosa after PDT. Female Wistar rats were treated with dihaematoporphyrin derivative (Photofrin^®) administered i.v. at a concentration of 5 mg kg^?1 body weight. Twenty-four hours after injection lesions were produced using a special device and irradiation was carried out with an argon-ion pumped-dye laser system (λ=630 nm, energy density 150 J cm^?2 and 30 J cm^?2; power density 100 mW cm^?2). At different points of time animals were sacrificed, tissues specimens were removed and a light and ultrastructural examination was carried out. At 90 min after treatment with 30 J cm^?2, superficial epithelial cells showed vacuoles in the basal part and were separating from the basement membrane. Capillaries were engored with erythrocytes and 24h after treatment there was destruction of capillaries and massive haemorrhage into the lamina propria. Immediately after treatment with 150 J cm^?2 there was sloughing of superficial epithelial cells from the basement membrane and crypt cells contained numerous vacuoles. After 24h there was complete necrosis of the mucosa. The action of PDT applied to normal rat colon mucosa is time- and energy-density-dependent and manifests as primary cytotoxic effects on superficial epithelial and crypt cells. Secondarily, there is destruction of the capillary system in the lamina propria, which leads to haemorrhagic necrosis of the mucosa.     

Development of an alternative light source to lasers for photodynamic therapy: 2. Comparative in vivo tumour response characteristics

The performance of a low cost, table-top/portable light source was tested against an argon ion pumped dye laser for in vivo photodynamic therapy (PDT). The prototype delivers up to 1 W via a 4 mm flexible lightguide within a 30 nm bandwidth centred at any wavelength from 300 nm to 1200 nm at fluence rates of up to 8 W cm^–2. An in situ bioassay using regrowth delay of tumour T50/80 was used to quantify the relative efficacy of the prototype with a laser. The tumours were sensitized with haematoporphyrin derivative (HpD) and externally irradiated. There was no significant difference in the response of the tumour to treatment between the two light sources (p = 0.69). Mean growth delays ranged from 2 days (light dose 10 J cm^–2) to 20 days (light dose 100 J cm^–2). The estimate for the difference in means (laser minus prototype growth delay) was only 0.66 days and was not statistically significant. This in vivo study demonstrates that the prototype is equivalent to a laser in PDT effect. The device has low capital/running cost, is simple to use and is one of the most powerful, spectrally efficient non-laser PDT sources available.     

Experimental photodynamic therapy with tetrapropyl-porphycene: Ultrastructural studies on the mechanism of tumour photodamage

Red light irradiation of a transplanted MS-2 fibrosarcoma in mice at 24 h after injection of liposome-zbound tetra-n-propyl-porphycene (TPP, 2mg kg^–1 b.w.) caused an efficient tumour necrosis. Electron microscopy analysis of tumour specimens taken at different times after the phototherapeutic treatment showed the development of direct damage of malignant cells between 3 and 6 h; the earliest detectable alterations occurred at the level of mitochondria. The endocellular damage gradually progressed with extensive vacuolization of the cytoplasm and, at later stages, formation of pyknotic nuclei. On the other hand, the vascular system of the tumour appeared to be well preserved up to about 9 h, when several endothelial alterations were detected. The damage of the tumour tissue was essentially complete 24 h after the phototreatment. The pattern of tumour modification is consistent with a preferential transport and tumour release of the liposome-bound TPP by low-density lipoproteins.     

Optimizing light dosimetry in photodynamic therapy of the bronchi by fluorescence spectroscopy

Under identical conditions (drug and light dose, timing), the results of photodynamic therapy (PDT) of carcinomas of the bronchi with tetra(meta-hydroxyphenyl)chlorin (mTHPC) show large variations between patients. Before patients underwent PDT treatment, the mTHPC level was measured in the lesion, the normal surrounding tissue and the oral cavity, with an apparatus based on fluorescence spectroscopy. The fluctuations in degree of tissue reaction and tumour destruction between patients could be explained by individual variations in the mTHPC level in the mucosa of the bronchi. The patients who showed the highest mTHPC fluorescence signal also had the strongest response to PDT. In addition, a correlation between the mTHPC level in the oral cavity and bronchial mucosa was found. It is concluded that PDT can be improved by measuring the mTHPC level in the bronchi or the oral cavity before treatment by fluorescence spectroscopy, and then by adjusting the light dose to be applied to the observed mTHPC level.     

Sensitization and photodynamic therapy of normal pancreas,duodenum and bile ducts in the hamster using 5-aminolaevulinic acid

Photodynamic therapy (PDT) using 5-aminolaevulinic-acid-(ALA)-induced protoporphyrin IX (PPIX) increases survival in hamsters with pancreatic cancer. However, experiments with other photosensitizers on this model show a high risk of duodenal perforation. In this paper, the pharmacokinetics and PDT effects of ALA on normal tissues in the pancreatobiliary region are presented. Using quantitative fluorescence microscopy, maximum PPIX fluorescence was seen in the bile ducts, less in the duodenal mucosa and least in the muscularis propria and pancreas. For PDT, light was delivered either using a bare fibre touching the tissue (single-point illumination), or irradiating a 1.5 cm diameter circular area. Single-point PDT (50 J) produced only localized reversible damage without perforation. Surface irradiation of the whole periampullary region (50 J cm^–2) caused extensive damage, sometimes with perforation. Before PDT can be used safely to treat tumours of the pancreas and bile duct, further studies are necessary to understand its effect on larger areas of normal tissue.     

Gross and microscopic changes in the viscera induced by photodynamic therapy applied to the lower abdomen of intact rats.

Photodynamic therapy (PDT) is a promising approach to the treatment of cancer. Preferential retention of the photosensitizer by malignant tissue has been considered a hallmark of this treatment modality. However, photosensitivity can be observed in normal, non-neoplastic tissues, and the present study investigated the effects of PDT treatment on the abdomen of intact rats. A circular region (1 cm diameter) on the shaved abdomen of Fischer rats, pretreated 24 h prior with Photofrin II, was irradiated for 30 min at 632 nm. Control animals received either photoradiation or Photofrin II administration. Subsequent lesions were observed in the irradiated skin, its associated abdominal wall, and the underlying gut in rats receiving Photofrin II and laser irradiation. All tissues were not equally sensitive to PDT treatment. Gut lesions were consistently more severe than were skin and abdominal wall injuries. By 24 hr after treatment, the gut manifested a transmural hemorrhagic necrosis, while the irradiated skin and abdominal wall were edematous, with an inflammatory infiltrate in the dermis and around occasional swollen myocytes. These results indicate that superficial lesions induced by PDT may not be reliable indicators of the extent of deeper PDT tissue damage. Further, it may be possible to take advantage of this discrepancy in tissue sensitivity and treat deep tissues through less sensitive superficial tissues.     

Photodynamic therapy in the treatment of malignant tumours of the upper aerodigestive tract

This is the report of a pilot study in which the technique of photodynamic therapy (PDT)—in which malignant tissues are destroyed by light after being previously photosensitized by haematoporphyrin or its derivative—was performed using a high power argon-dye laser system which operated at 630±5 nm and optical fibres with either a microlens or a diffusing tip.A total of thirty-seven patients were utilized for this study, including twenty-five patients affected by esophageal carcinoma, five by recurrences of head and neck cancer, four by gastric cancer and three by lung cancer. Among the sixteen cases with superficial or localized cancer of the esophagus, complete and partial responses were observed in seven and six patients, respectively. In advanced and recurring cancers less satisfactory results were obtained. Thus, PDT seems to be useful in cases of small superficial cancers, either primary or recurrent after previous treatments.