Application of lower fluence rate for less microvasculature damage and greater cell-killing during photodynamic therapy

During the process of photodynamic therapy (PDT), problems arise such as stasis or occlusion of microvasculature, tumor oxygen depletion, and photosensitizer bleaching. This study shows that the first problem could be reduced by using a lower fluence rate light source in PDT. Microvasculature damage was studied experimentally in hematoporphyrin derivative–mediated PDT against light fluence rate, and, to some extent, less microvasculature damage was induced under 75 mW/cm² illumination than under 150 mW/cm². Histology of vessels at the end of PDT showed that vessel damage could be observed in both groups, indicating that the microvasculature would eventually be damaged as long as the administration of light fluence was sufficient and regardless of the illuminating fluence rates. Thus microvasculature damage induced by low fluence rate illumination could also be effective in tumor control after PDT. The cell-killing experiment was performed in vitro and designed so that cell-killing rate was influenced only by light characteristics. The higher cell-killing rate caused by 75 mW/cm² illumination indicated that lower fluence rate light could enhance the light absorbency or decrease the bleaching of photosensitizer.An erratum to this article can be found at     

Applicator for light delivery and in situ light dosimetry during endobronchial photodynamic therapy: First measurements in humans

This paper presents a design of an applicator for light delivery and light dosimetry during endobronchial photodynamic therapy (EB-PDT). The design incorporates a linear diffuser that is fixed in the centre of the lumen by a steel spring basket that does not block air flow. An isotropic light detector is included in this design, to measure the light fluence actually delivered to the bronchial mucosa surface. The applicator is designed for use with common bronchoscopy equipment, and can be used with bronchoscopes with a large biopsy channel (≈3 mm). The first clinical measurements were performed and caused no additional discomfort to the (nonphotosensitized) patients. The data showed considerable inter-patient variability of the light fluence rate measured as a result of fixed output power of the diffuser. This fact and the expected strong dependence of the fluence rate on the lumen diameter stress the importance of in situ fluence rate measurement for a proper evaluation of the relationship between light fluence and the biological response of EB-PDT.     

The Calibration of fibre optic probes used for light dosimetry measurements during photodynamic therapy

We describe the calibration of fibre optic probes used to perform in vivo light dosimetry studies during the treatment of skin lesions by photodynamic therapy. Results from six individual detectors show that the linearity of the calibration and the calculated radiant energy fluence rate within a liquid phantom are independent of the type and sensitivity of the probe. The method of calibration is also shown to yield the optical interaction coefficients of the phantom.     

The optical properties of skin tumours measured during superficial photodynamic therapy

Accurate measurement of light distribution in tissue can improve the knowledge of in vivo tissue optical properties, which is essential for precise dosimetry calculations during photodynamic therapy (PDT). In our application of PDT, superficial skin lesions are treated by topical administration of 5-aminolaevulinic acid followed by surface illumination using 630 nm laser light. Several small detector probes inserted under the skin surface prior to illumination record the light intensity at different depths throughout the treatment. Results from 11 patients are presented. These results indicate light distributions described by the diffusion theory for light transport in tissue. The accumulated data do not imply one specific set of optical parameters for the skin, but indicate that although the depth of light penetration is constant for all patients, the variation in skin colour is significant when performing dosimetry calculations. The average value of effective attenuation coefficient was found to be 0.359 mm⁻¹ (±9.5%) and the coefficientk that is used to quantify the build-up of subsurface fluence rate varies between 0.12 and 8.23.     

Photofrin-mediated photodynamic therapy of rat palatal mucosa: Normal tissue effects and light dosimetry

Photodynamic therapy (PDT) is a treatment modality with potential application for premalignant lesions and squamous cell carcinoma of the oral mucosa. PDT in principle has dual selectivity. This may result from a preferential retention of the photosensitizer in target tissue. In addition, the photodynamic activity will be limited to the irradiated area because PDT will not affect tissues in the absence of excitation light. The specificity of PDT is limited by the fact that normal tissues also retain the photosensitizer to some degree, which makes these tissues susceptible to PDT damage. To optimize PDT for oral malignancies, a study was undertaken on normal tissue to investigate the responses in rat palatal mucosa and surrounding anatomical structures. Eighty male Wistar rats were used in the study. Photofrin was administered i.v. at four doses (0, 2.5, 5 or 10 mg kg^–1 body weight). Irradiation for PDT was performed 24 h later. An argon pumped dye laser system was used to produce light of two different treatment wavelengths (514.5 and 625 nm), and various energy density levels (0, 25, 50, 100 or 200 J cm^–2). Early effects of PDT were studied at 2 days and late effects at 2 months after treatment. Twenty-four hours after i.v. administration of Photofrin, it was found that PDT affects normal tissues of the oral cavity both macroscopically and microscopically. Combinations of photosensitizer doses 5 mg kg^–1 and light doses100 J cm^–2 caused severe and permanent damage to the palatal mucosa and adjacent normal structures such as palatal bone and dentition.Light scattering and internal reflection usually raise the fluence rate in tissue above the irradiance of the incident beam. In an additional study using six male Wistar rats, the energy fluence rate at two treatment wavelengths (514.5 and 625 nm) was measured ex vivo in the palatal mucosa and adjacent anatomical structures. As expected, the energy fluence rates were wavelength, tissue and depth dependent. At the air-mucosa boundary, light of 625 nm was found to have a three-times higher fluence rate than the primary incident beam. Under similar conditions, the fluence rate of 514.5 nm was found to be less, but still twice as high as the primary incident beam. At deeper levels of the rat maxilla, fluence rates were still elevated compared with the incident beam. For 625 nm light, this phenomenon was observed up to the level of the nasal cavity. These increased fluence rates could largely explain the pattern of damage to normal mucosa and surrounding anatomical structures.     

Light delivery and light dosimetry for photodynamic therapy

Photodynamic therapy (PDT) has attracted attention because it was considered to be a selective form of cancer treatment causing minimal damage to normal tissues. This is not exactly true, because the ratio between the photosensitizer concentrations in tumour and surrounding normal tissues is not always much more than one. Nevertheless, tumour destruction by PDT with relatively little damage to normal tissue is possible in many cases. This requires sophisticated light delivery and/or light dosimetry techniques. In this respect the limited penetration of light into biological tissues can sometimes be useful. In this paper a qualitative and sometimes quantitative discussion is given of the physical phenomena determining the energy fluence in a biological tissue. Most important is light scattering, the contribution of which depends on the geometrical conditions. Finite beam surface irradiation, irradiation of hollow organs (bladder) and interstitial irradiation are discussed separately. The emphasis is on light dose and light dose distribution. It is emphasized that PDT dosimetry in general is complicated by photosensitizer distribution (which is usually not known), by photobleaching of the sensitizer, by possible effects of hyperthermia, and by changes in optical properties during and as a result of PDT.     

Standardization of intralipid for light scattering in clinical photodynamic therapy

Intralipid is a light scattering medium used in photodynamic therapy (PDT) in both phantom studies and to obtain optimum light distributions from clinical light delivery systems. Light delivery to a target is one variable in PDT over which we have direct control, and should be as accurate as possible. Variations in the scattering ability of Intralipid from batch to batch will lead to an unpredictable light distribution and undesirable PDT effects. Furthermore, 10% and 20% stock solutions, when diluted to similar final concentrations, have different scattering abilities which will lead to inaccuracies in studies measuring scattering coefficients and imprecision in light distributions from light delivery systems. Batch to batch variations of similar stock solutions should lead to minimal differences in outputs from light delivery systems. A simple and quick assay system is described to define the concentration of Intralipid based on its light scattering ability, which enables testing of solutions prior to use. In the clinical setting where accurate light distribution is required, we suggest that only one stock solution (e.g. 20%) is used and that the final dilution scattering ability is checked using this assay system.     

Laser light application and light monitoring for photodynamic therapy in hollow organs

Photodynamic therapy (PDT) of tumours in hollow organs requires light application devices which enable homogeneous illumination of the tissue surface in hollow organs. This paper presents two laser light application systems generating a homogeneous light distribution and one monitoring unit which detects variations in the applied laser light intensity during treatment. A cylindrical light diffuser has been developed for PDT of cylindrical organs (e.g. bronchus, oesophagus). This system guarantees a defined homogeneity of the laser light intensity. An exact positioning to a defined treatment area is possible. Adjacent tissue is prevented from laser light irradiation. A special lens system has been developed for the irradiation of flat surfaces (e.g. skin, carina, organ wall of the large bowel and the stomach). The illumination area is defined by the distance to the tissue and the large aperture. The size of the system allows for it to pass through the biopsy channel of conventional endoscopes. A monitoring device recently developed helps to detects of the fibre, changes in the medium around the fibre tip, and variations of the laser output power during laser treatment. All devices serve for evaluating an accurate light dosimetry in clinical PDT.     

Coagulation of blood at the tip of optical fibres used for light delivery in photodynamic therapy

Optimal delivery of light to the tumour is of considerable importance in photodynamic therapy. The most effective way of delivering laser light to the tumour tissue is through an implanted optical fibre. In order to investigate the possible effects taking place at the tips of fibres implanted in tissue, fibres were used to deliver light to human blood and the transmission of light by the blood was measured at different power levels. The maximum power level which could be delivered without charring or coagulation at the fibre tip was measured for five different fibres. Three plane cut fibres and two with 1.5 cm long diffusing tips were studied. Charring and coagulation, which resulted in practically no light being delivered more than 0.5 mm from the fibre tip, were observed at relatively low-output powers (70-130 mW) for all the plane-cut fibres. This is less than the level required to deliver a clinically useful dose in a reasonable time. In contrast, neither charring nor coagulation was observed at the diffusing tips for output powers up to 1.1 W and consequently these should be the fibres of choice for interstitial photodynamic therapy. Observed changes in light transmission through the blood with increasing output power indicate that, for accurate light dosimetry, a means of monitoring delivered light in vivo during photodynamic therapy is essential.     

Factors controlling the selectivity and efficiency of tumour damage in photodynamic therapy

The scope and potential of the photodynamic therapy of tumors can be enhanced through an adequate control of the factors which improve the selectivity of tumour targeting by the systemically injected photosensitizer and increase the efficiency of photosensitized tumour damage. Promising results are obtained by using hydrophobic photosensitizers which can be specifically transported and released to the tumour by serum lipoproteins, especially low-density lipoproteins. The photosensitizer molecule should possess those structural features which induce a high probability of photoactivation by 700–800 nm light, as well as a high yield of long-lived triplet state. The use of liposome-delivered Zn-phthalocyanine as a second generation phototherapeutic agent for tumours is proposed.     

A comparison of novel light sources for photodynamic therapy

A diode laser, light-emitting diode (LED) array bandwidth 25 nm, full width half maximum (FWHM) and filtered arc lamp (bandwidth 40 nm, FWHM), all with peak emission at about 650 nm, suitable for the photosensitizer tetra(meta-hydroxyphenyl)chlorin (mTHPC), were compared with a copper vapour laser pumped dye laser, using depth of necrosis in normal rat liver as a measure of photodynamic effect. A three-way comparison between a DL10K dye laser, the LED array and the filtered arc lamp resulted in mean depths of necrosis of 4.64, 4.29 and 4.04 mm, respectively, at 20 J cm^-2, the values for the laser and arc lamp being significantly different at the 5% level. A further comparison of a narrower linewidth DL20K dye laser with the LED array, using a light dose of 20 J cm^-2, showed a significant difference between the mean depths of necrosis of 4.97 and 4.05 mm, respectively (p=0.01). A final study, comparing the DL20K dye laser with the diode laser and a light dose of 10 J cm^-2, demonstrated no significant difference in depths of necrosis (3.23 and 3.25 mm, respectively). The results obtained in the three studies are attributed to the relative bandwidths of light emission for the various sources. A simple mathematical model is presented explaining the results in terms of the relative activation of the photosensitizer and the consequent threshold fluence required for the induction of necrosis. It is concluded that, in order to achieve the same depth of effect as a laser when using the broad band sources, the incident fluence would have to be approximately doubled. However, when the low cost and ease of use of the non-laser sources are taken into consideration, these devices are likely to find widespread applications in clinical photodynamic therapy.     

A comparison of gold-vapour and dye lasers for photodynamic therapy

This paper compares the relative merits of the continuous wave argon-pumped dye laser and the pulsed gold-vapour laser as used clinically for photodynamic therapy. At comparable power and energy outputs, the biological effect of the two appears to be the same. However, for 1 W output (a suitable level for clinical use), the gold laser is simpler and easier to install and run, although it requires a larger diameter fibre for light delivery (0.6 mm v. 0.2 mm). The wavelength of the dye laser is tunable, whereas that of the gold is not, although the gold laser can be easily converted to a copper-vapour laser which can pump a tunable dye laser.     

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.     

光动力学治疗对瘢痕成纤维细胞增殖及其细胞周期的影响

目的 探讨血卟啉单甲醚-光动力学疗法对体外培养瘢痕成纤维细胞的作用.方法 原代培养瘢痕组织来源的成纤维细胞后,建立细胞系,实验采用第4～6代细胞.银染细胞后镜下观察,并计算核仁组成区的相对面积(I.S%);采用流式细胞技术,分析光动力学治疗后各细胞周期中细胞分布比例,并计算增殖指数.结果 ①光动力学治疗后瘢痕成纤维细胞内I.S%显著降低;②流式细胞仪检测结果表明光动力学治疗组S期峰低于对照组,S期细胞比例显著降低,为(11.2±2.3)%;③计算对照组和光动力学治疗组的增殖指数,分别为(35.0±3.4)%和(27.2±3.1)%,两者比较差异有统计学意义(P<0.05).结论 血卟啉单甲醚-光动力学疗法能够抑制瘢痕成纤维细胞的过度增殖,改变其细胞周期分布.     

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.     

Tetra(m-hydroxyphenyl)chlorin clinical photodynamic therapy of early bronchial and oesophageal cancers

There have been few studies to date of clinical photodynamic therapy (PDT) with tetra(m-hydroxyphenyl)chlorin (mTHPC). This paper describes the results of the authors' experience with this second-generation photosensitizer, used in the treatment of 40 early cancers of the oesophagus and the bronchi. Surface illumination of the tumour was performed, in most cases, 4 days after intravenous injection of 0.15 mg kg^–1 of mTHPC, using 652 nm or 514 nm continuous wave laser light. Endoscopic follow-up with biopsies and brushings was possible for 35 tumours: 27 (77%) showed no recurrence after disease-free follow-ups that ranged from 3 to 38 months. Major complications, all of which were after red light illumination, included one bronchial stenosis, one oesophagotracheal fistula and two probable occult perforations of the oesophagus. Photodynamic therapy with green light renders such perforations of the oesophageal wall essentially impossible at the applied conditions, and appears not to reduce the efficacy of the treatment. Skin photosensitization, which was never observed later than the first week after injection, occurred in 12 patients. Hence, PDT with mTHPC is a safe and effective treatment for early carcinomas of the oesophagus and the tracheobronchial tree.     

Phase III randomized trial of surgery with or without intraoperative photodynamic therapy and postoperative immunochemotherapy for malignant pleural mesothelioma

Background: Patients with malignant pleural mesothelioma (MPM) usually die of progressive local disease. This report describes the results of a Phase III trial comparing maximum debulking surgery and postoperative cisplatin, interferon α-2b, and tamoxifen (CIT) immunochemotherapy with and without intraoperative photodynamic therapy (PDT) to determine (1) whether such a multimodal approach can be performed with minimum morbidity and mortality in malignant pleural mesothelioma (MPM), and (2) whether first-generation (i.e., 630-nm laser light, Photofrin II) intrapleural PDT impacts on local recurrence or survival. Methods: From July 1993 to June 1996, 63 patients with localized MPM were randomized to either PDT or no PDT. The tumors of 15 patients could not be debulked to 5 mm. Patients assigned to PDT (n=25) and no PDT (n=23) were similar with respect to age, sex, tumor volume, and histology. Results: The type of resection (11 pleurectomies and 14 pneumonectomies vs. 12 pleurectomies and 11 pneumonectomies), length postoperative stay, and ICU time were comparable (PDT vs. no PDT). There was one operative death (hemorrhage), and each group had two bronchopleural fistulas. Postoperative staging divided patients into the following categories: stage I: PDT, 2, no PDT, 2; stage II: PDT, 2, no PDT, 2; stage III, PDT, 21; no PDT, 17; stage IV, PDT, 0; 0; no PDT, 2. Comparable numbers of CIT cycles were delivered. Median survival for the 15 non-debulked patients was 7.2 months, compared to 14 months for the 48 patients on protocol. There were no differences in median survival (14.4 vs. 14.1 months) or median progression-free time (8.5 vs. 7.7 months), and sites of first recurrence were similar. Conclusions: Aggressive multimodal therapy can be delivered for patients with higher stage MPM. First-generation PDT does not prolong survival or increase local control for MPM. Presented at the 50th Annual Cancer Symposium of The Society of Surgical Oncology, Chicago, Illinois, March 20–23, 1997.     

Performance of a dedicated light delivery and dosimetry device for photodynamic therapy of nasopharyngeal carcinoma: phantom and volunteer experiments

The objective of this study was to develop a light delivery and measurement device for photodynamic therapy (PDT) in the nasopharyngeal cavity, which achieves a homogeneous and reproducible fluence rate distribution to a target area and provides proper shielding of predefined risk areas.MATERIALS AND METHODS: A flexible silicone applicator was developed, incorporating light delivery and dosimetry fibers. The applicator can be inserted through the mouth and fixed in the nasopharyngeal cavity. Tissue optical phantoms were prepared on the basis of optical properties measured in vivo using diffuse reflectance spectroscopy (DRS). The fluence rate over the length of the applicator surface was measured in air, in tissue optical phantoms and in five healthy volunteers. RESULTS: The fluence rate distribution over the applicator surface in air and tissue optical phantom was found to be more homogeneous (SD/mean 3.8% and 18.3%, respectively) than the fluence rate distribution in five volunteers (SD/mean ranging from 19% up to 52%). The maximum observed fluence rate build-up in the nasopharynx varied between subjects and ranged from a factor of 4.1-6.9. Shielding of the risk area such as the soft palate and tongue was effective. CONCLUSIONS: In air and in tissue optical phantoms the fluence rate distribution of the device was highly homogeneous. The observed inter-subject and intra-subject variations in fluence rate in healthy volunteers originated from differences in optical properties and nasopharyngeal geometry. Light delivery based on a single tissue surface measurement will not be adequate. In situ dosimetric measurements are required to determine the light fluence delivered to a geometrically complex site such as the nasopharynx. These observations should be taken in consideration when developing light applicators for PDT of the nasopharynx and other non-uniform surfaces.     

晚期胆管癌姑息性切除、光动力治疗及支架引流疗效的Meta分析

目的评价晚期胆管癌患者姑息性R1、R2切除与支架引流治疗的疗效及光动力治疗与支架引流的疗效。 方法计算机检索PUBMED、中国生物医学文献数据库(CBMdisc)、万方数据库、中国学术期刊全文数据库(CNKI),查找1980年1月至2013年12月发表的有关对比分析胆管癌姑息性(R1、R2)切除与支架引流治疗及光动力治疗与支架引流治疗效果的随机对照试验文献。按照纳入与排除标准选择文献、提取资料、评价质量后,采用RevMan 5.2软件进行Meta分析。 结果共有9篇研究585例患者纳入姑息性R1、R2切除和支架引流分析,其中姑息性切除组208例,支架引流组377例。共有4篇研究纳入光动力治疗和支架引流分析,包括252例患者,其中光动力治疗组132例,引流治疗组120例。分析显示:总体并发症发生率、死亡率、1年生存率指标中,姑息性R1、R2切除组与引流治疗组相比1年生存人数较多,差异有统计学意义[(OR 0.79,95%CI 0.42~1.50,P=0.48)、(RD-0.00,95%CI-0.04~0.04,P=0.94)、(OR 2.87,95%CI 1.82~4.54,P<0.05)],胆瘘发生率、胆管炎或胆道梗阻发生率差异无统计学意义[(OR 1.74,95%CI 0.73~4.17,P=0.21)、(OR 0.50, 95%CI 0.22~1.12,P=0.09)]。光动力治疗组和支架引流的疗效对比中光动力治疗术后生存时间较支架引流明显延长。 结论晚期胆管癌患者R1、R2切除及光动力疗效均好于单纯支架引流,但光动力治疗与手术相比是一种安全、恢复快、创伤小的治疗方式。