Hemoporfin-mediated photodynamic therapy for the treatment of port-wine stain birthmarks in pediatric patients

Abstract: Port-wine stain is a type of common congenital superficial telangiectasia in the dermal layer mostly occurring on the forehead, face, and neck. The affected skin shows abnormal red or purple lesions, which darken and thicken. Nodular changes may develop with advancing age. If untreated, port-wine stains scarcely regress, which can have serious lifelong psychological impact on patients and affect their quality of life. In this report, we focused on two cases of port-wine stain in pediatric patients; the youngest patient was only 1.5 years old. During and after treatment, Hemoporfin-mediated photodynamic therapy features fewer adverse reactions, short light protection period, easy nursing, and good efficacy.

Hematoporphyrin monomethyl ether combined with He-Ne laser irradiation-induced apoptosis in canine breast cancer cells through the mitochondrial pathway

Hematoporphyrin monomethyl ether (HMME) combined with He-Ne laser irradiation is a novel and promising photodynamic therapy (PDT)-induced apoptosis that can be applied in vitro on canine breast cancer cells. However, the exact pathway responsible for HMME-PDT in canine breast cancer cells remains unknown. CHMm cells morphology and apoptosis were analyzed using optical microscope, terminal deoxynucleotidyl transferase dUTP nick end labeling fluorescein staining and DNA ladder assays. Apoptotic pathway was further confirmed by Real-time-polymerase chain reaction and Western blotting assays. Our results showed that HMME-PDT induced significant changes in cell morphology, such as formation of cytoplasmic vacuoles and the gradual rounding of cells coupled with decreased size and detachment. DNA fragmentation and cell death was shown to occur in a time-dependent manner. Furthermore, HMME-PDT increased the activities of caspase-9 and caspase-3, and released cytochrome c from mitochondria into the cytoplasm. HMME-PDT also significantly increased both mRNA and protein levels of Bax and decreased P53 gene expression in a time-dependent manner, while the mRNA and protein expression of Bcl-2 were repressed. These alterations suggest that HMME-PDT induced CHMm cell apoptosis via the mitochondrial apoptosis pathway and had anti-canine breast cancer effects in vitro.

Deciphering the binding modes of hematoporphyrin to bovine serum albumin.

Interaction of proteins with small molecules is important in understanding delivery and transport of different therapeutic agents, including drugs. In the present study, we investigated the interaction between hematoporphyrin (HP), the principal component of photosensitizing drug with bovine serum albumin (BSA) in aqueous buffer solution using UV-Vis absorption spectroscopy and fluorescence measurements. The results were further substantiated by molecular docking and molecular dynamics (MD) simulation. Our results revealed that fluorescence of BSA was dominantly quenched by the ground-state complex formation with HP accompanied by the electronic energy transfer (EET) to the later. We experimentally determined the thermodynamic parameters such as G0, H0, and S0 for the HP-BSA system which were -35.5 kJ mole-1, -56.4 kJ mole-1 and -0.06 kJ mole-1 K-1, respectively. These parameters suggested hydrogen-bonding and Van der Waals forces playing major role in the complexation. This was also supported by the binding energy parameters calculated by molecular docking. Moreover, the experimentally determined G0 nicely correlated with those determined by molecular docking and MD-simulation. Further, computational results clearly showed that the binding of HP with BSA in the subdomains IB and IIA.

Photodynamic inactivation of four Candida species induced by photogem®

This study evaluated the in vitro susceptibility of C. albicans, C. dubliniensis, C. tropicalis and C. krusei to photodynamic therapy (PDT) induced by Photogem® and light emitting diode (LED). Suspensions of each Candida strain were treated with three photosensitizer (PS) concentrations (10, 25 and 50 mg/L) and exposed to 18.0, 25.5 and 37.5 J/cm² LED light fluences (λ ~ 455 nm). Control suspensions were treated only with PS concentrations, only exposed to the LED light fluences or not exposed to LED light or PS. Sixteen experimental conditions were obtained and each condition was repeated three times. From each sample, serial dilutions were obtained and aliquots were plated on Sabouraud Dextrose Agar. After incubation of plates (37 ºC for 48 hours), colonies were counted (cfu/mL) and the data were statistically analyzed by ANOVA and the Tukey test (α=0.05). Complete killing of C. albicans was observed after 18.0 J/cm² in association with 50 mg/L of PS. C. dubliniensis were inactivated after 18.0 J/cm² using 25 mg/L of PS. The inactivation of C. tropicalis was observed after photosensitization with 25 mg/L and subsequent illumination at 25.5 J/cm². For C. krusei, none of the associations between PS and light resulted in complete killing of this species. PDT proved to be effective for the inactivation of C. albicans, C. dubliniensis and C. tropicalis. In addition, reduction in the viability of C. krusei was achieved with some of the PS and light associations.

Avaliação do efeito citotóxico da terapia fotodinâmica associada ao LED e ao Photogem® sobre a mucosa bucal de rato / Toxicity of the photodynamics therapy with LED associated to Photogem®: an in vivo study

A utilização da PDT para tratamento de diferentes tipos de infecções, tal como a candidose bucal, tem sido estudada. Entretanto, poucos são os dados científicos que relatam os possíveis efeitos tóxicos dessa terapia. Dessa forma, o objetivo deste estudo foi avaliar os efeitos da irradiação na mucosa bucal de ratos com LED azul (de 460 nm e potência de 200 mW/cm2) em presença do fotossensibilizador (FS) Photogem®, em duas diferentes concentrações (500 mg/L e 1000 mg/L). Para isso, foram utilizados 101 ratos (Rattus Norvegicus Albinus Holtzman) distribuídos em 6 grupos, de acordo com os seguintes tratamentos: Grupo 1 ­ controle; Grupo 2 ­ aplicação do FS (500 mg/L); Grupo 3 ­ aplicação do FS (500 mg/L) e irradiação com LED; Grupo 4 - aplicação do FS (1000 mg/L); Grupo 5 ­ aplicação do FS (1000 mg/L) e irradiação com LED; e Grupo 6 ­ irradiação com LED. O FS foi aplicado por 30 minutos (tempo de pré-incubação) e o tempo de irradiação da mucosa foi de 20 minutos (dose de 144 J/cm2 ). Decorridos os 4 períodos de avaliação propostos (0 dia, 1dia, 3 dias e 7 dias), os animais tiveram a mucosa palatina fotografada para análise macroscópica, sendo então imediatamente sacrificados para remoção cirúrgica do palato e posterior análise em microscopia de luz e de fluorescência. Um mapeamento térmico foi realizado a fim de avaliar a variação de temperatura ocorrida no tecido durante a irradiação com LED. Macroscopicamente, em todos os grupos experimentais e para todos os períodos de avaliação propostos na presente pesquisa, observou-se que a mucosa apresentava-se intacta, com aspecto de normalidade semelhante ao do Grupo 1 (controle). Microscopicamente, alterações teciduais, caracterizadas especialmente por discreta inflamação, puderam ser observadas na mucosa palatina de apenas 4 de um total de 80 animais submetidos a PDT. A penetração do fotossensibilizador na mucosa tratada pôde ser observada por meio da emissão de fluorescência do Photogem® , tendo este FS se mantido presente apenas no tecido epitelial. O mapeamento térmico revelou que a temperatura aumentou de 35ºC para 41ºC durante 20 minutos de irradiação. Dentro das condições experimentais avaliadas, foi possível concluir que a PDT, utilizando Photogem® nas concentrações de 500 mg/L e 1000 mg/L associado ou não à irradiação com LED (dose de 144 J/cm2), não foi tóxica para a mucosa palatina de ratos

The use of PDT has been investigated for the treatment of different types of infection, like oral candidosis. There are, however, few research-based data that report the possible toxic effects of this therapy. Therefore, this study evaluated the effects of irradiating the palatal mucosa of rats with blue LED (460 nm; 200 mW/cm²) in the presence of the photosensitizer Photogem® at two concentrations (500 and 1000 mg/L). Then, 101 rats (Rattus norvegicus albinus Holtzman) were randomly distributed in six groups, according to the treatment performed on the palatal mucosa: Group 1: control; Group 2: Photogem® (500 mg/L); Group 3: Photogem® (500 mg/L) + blue LED; Group 4 - Photogem® (1000 mg/L); Group 5: (1000 mg/L) + blue LED; and Group 6: blue LED. The exposure times to the photosensitizing agent and to the light source were 30 min (pre-incubation time) and 20 min (144 J/cm2 energy density), respectively. At 0, 1, 3 and 7 days posttreatment, the animals had their palatal mucosa photographed for macroscopic analysis and were immediately sacrificed. The palate was removed for further analysis by light and fluorescence microscopy. Thermal mapping was made to evaluate the temperature change occurred in the tissue during LED irradiation. In all experimental groups and periods, the macroscopic analysis revealed intact mucosa with normal aspect similar to that of Group 1 (control). Tissue alterations, characterized primarily by a mild inflammation, were observed microscopically on the mucosa of only 4 out of 80 animals subjected to PDT. Photosensitizer penetration into the treated mucosa was identified by the fluorescence emitted by Photogem® and was limited to the epithelial layer. The thermal mapping revealed a temperature increase from 35 to 41ºC during the 20-min irradiation. In conclusion, under the tested conditions, PDT using Photogem® at 500 and 1000 mg/L concentrations associated or not to LED irradiation (144 J/cm2) was not toxic to the rat palatal mucosa

Effect of ultrasound activating hematoporphyrin on the activities of antioxidative enzymes in mouse hepatoma 22 / 生物医学工程学杂志

This investigation was made with regard to the influences of ultrasound combined with hematoporphyrin on the activities of antioxidative enzyme in ascites hepatoma 22 (H-22) tumor cells, and to a better understanding of the potential biological mechanism of sonodynamic therapy which involved the damage to cells. Combined with 100 microg/ml hematoporphyrin, high intensity focused ultrasound sonication at a frequency of 1.43 MHz and an intensity level of 2.0 W/cm2 was delivered to H-22 tumor cells for 1 min. The viability of cells was evaluated by typan-blue blue exclusion test. The intracellular reactive oxygen species (ROS) levels were determined by 2',7'-dichlorofluorescein diacetata (DCFH-DA). Enzymatic chemical methods were used to measure the activities of key antioxidative enzymes. The results indicated that the cell damage rate of ultrasound combined with hematoporphyrin was significantly higher than that of the treatment with ultrasound alone, and hematoporphyrin alone had no killing effect on H-22 cells. The level of ROS in cell suspension was significantly increased, and the key antioxidative enzyme activities were obviously decreased after treatment with the combined use of ultrasound and hematoporphyrin. We speculated that the decreased activities of key antioxidative enzymes in cells might be involved in mediating the killing effect on H22 cells in sonodynamic therapy.

Efetividade da terapia fotodinâmica na inativação de Candida spp / Effectiveness of photodynamic therapy on the inactivation for Candida spp

O surgimento da resistência antifúngica aos tratamentos convencionais tem proporcionado o desenvolvimento de novas modalidades terapêuticas para o tratamento da candidose bucal. Nesse contexto, a utilização da PDT vem sendo sugerida como método alternativo para a inativação de microrganismos patogênicos. Este estudo avaliou a efetividade da PDT na inativação de C. albicans e C. glabrata, ATCC e resistente a fluconazol, por meio da utilização do agente fotossensibilizador Photogem® e da iluminação com LEDs de comprimento de onda azul. Inicialmente, os microrganismos avaliados foram inoculados em tubos de ensaio contendo meio de cultura líquido e incubados overnight a 37ºC. Em seguida, foram obtidas suspensões celulares das espécies de Candida avaliadas. Essas suspensões foram transferidas para placas de 96 orifícios, tratadas com cinco diferentes concentrações de Photogem® (2,5; 5; 10; 25 e 50mg/L) e expostas a quatro doses de luz (10,5; 18; 25,5 e 37,5J/cm2). Suspensões adicionais foram tratadas somente com as cinco concentrações do fotossensibilizador ou apenas com as quatro doses de luz. Cada condição experimental foi realizada três vezes. Após a realização desses experimentos, foram obtidas diluições seriadas de cada amostra (10-1 a 10-3), e alíquotas de 25 µL dessas diluições foram plaqueadas, em triplicatas, em Sabouraud Dextrose Agar. Adicionalmente, alíquotas de 25 µL foram removidas das cavidades das placas de orifícios e transferidas diretamente para um quadrante da placa de Petri, sem a realização de diluição. As placas foram incubadas a 37ºC por 48 horas. Após a incubação, foi realizada a contagem das colônias viáveis (ufc/mL), e os valores obtidos foram analisados com o teste t de Student (p < 0,05). Os 16 resultados demonstraram que a inativação de Candida spp. ocorreu de forma concentração/dose dependente, que resultou na completa inativação desses microrganismos em determinadas condições experimentais. A dose de luz mínima que promoveu a completa inativação das duas origens de C. albicans foi 18 J/cm2, em associação com 50mg/L de Photogem® . Após a aplicação de 25,5 e 37,5 J/cm2, baixas concentrações de fotossensibilizador foram requeridas para a inativação total dessa espécie, sendo que diferenças estatisticamente significantes foram apontadas entre os valores obtidos para as duas origens de C. albicans. Também foram observadas diferenças estatisticamente significantes na obtenção da inativação total das duas origens de C. glabrata. Para a ATCC, não houve crescimento de colônias viáveis após o tratamento com 10, 25 e 50 mg/L de Photogem® seguido de iluminação a 37,5 J/cm2 . No entanto, somente as concentrações de 25 e 50 mg/L foram capazes de eliminar a C. glabrata resistente a fluconazol, nas doses de 25,5 e 37,5 J/cm2. Assim, a fotoativação do Photogem® pela luz do LED demonstrou efetividade na inativação das duas espécies de Candida avaliadas, ATCC e resistente a fluconazol

Oral candidosis is an opportunistic infection that affects a significant percentage of the population. The oral infection caused by Candida spp. is usually treated with topical and systemic antifungal drugs. However, the widespread use of these agents has resulted in an alarming increase in the rate of antifungal resistance. Recently, the photodynamic therapy (PDT) has been studied as an alternative modality of killing microorganisms, including viruses, fungi and bacteria. The aim of this study was to determine whether Candida albicans and C. glabrata, ATCC and fluconazole-resistant strains, could be photosensitized by Photogem® in combination with blue Light Emitted Diode (LED). Suspensions of each Candida strain, containing viable cells per milliliter, were treated with five concentrations of Photogem® (2.5, 5, 10, 25 and 50 mg/l), followed by LED irradiation in four light doses (10.5, 18, 25.5 and 37.5 J/cm2). Each experimental condition was carried out in triplicate and repeated tree times. From each sample, serial dilutions were obtained and aliquots of 25 µl of each dilution were plated on Sabouraud Dextrose Agar. All plates were incubated at 37°C for 48 hours. After incubation, colonies were counted (CFU/ml) and the data were statistically analyzed by the Student's t test (p < 0.05). The results demonstrated a concentration/dose-dependent pattern of inactivation, that resulted in complete elimination of all Candida evaluated. The minimal light dose for the complete inactivation of both C. albicans source was 18 J/cm2 , in conjunction with 50 mg/l of Photogem®. After 25.5 and 37.5 J/cm2 , a lower concentration of Photogem® was required to totally inactivate C. albicans ATCC (5 and 2.5 mg/l) in comparison with C. albicans fluconazole-resistant (10 and 5 mg/l). There were statistically significant differences in the log (CFU/ml) for the minimal light dose to complete inactivation of both C. glabrata sources. For C. glabrata ATCC, no viable cells were detected after treatment with 10, 25.5 or 50 mg/l of Photogem® followed by 37.5 J/cm2 , while the association of 25 mg/l with 25.5 J/cm2 was sufficient to totally inactivate the C. glabrata fluconazole-resistant. The photoactivation of Photogem® by blue LED light proved to be effective for the inactivation of fluconazole-resistant and ATCC strains of C. albicans and C. glabrata

Effect of HMME-PDT on hyperplastic scar in rabbit ear model / 中华整形外科杂志

<p><b>OBJECTIVE</b>To investigate the effect of HMME-PDT (Hematoporphyrin Monomethyl Ether-Photodynamic therapy) on Hyperplastic scar in the rabbit ear.</p><p><b>METHODS</b>The acute model of dermal Hyperplastic scar in the rabbit ear was established. 24 scars were randomly divided into 2 groups: the experimental group (n = 12) received HMME-PDT treatment, and the controlled group (n = 12) received no special treatment. Specimens were harvested from scars on postoperative 28 day. Scar hyper plasty and collagen fibers were observed by haematoxylin-eosin staining and Van-Gieson staining respectively. The microvessel density was calculated under microscope.</p><p><b>RESULTS</b>Compared with the controlled group, HMME-PDT treatment in the experimental group reduced scar formation, decreased the microvessel density and prevented excess collagen deposition at the wound site.</p><p><b>CONCLUSIONS</b>HMME-PDT may play a role in inhibiting hyperplastic scar in rabbit ear.</p>

Hematoporphyrin derivative-mediated photodynamic therapy for human colon carcinoma: a comparative study with LoVo and CoLo205 cells in vitro / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the killing effect of photodynamic therapy (PDT) mediated by hematoporphyrin derivative (HpD) on human colon carcinoma LoVo and CoLo205 cells in vitro.</p><p><b>METHODS</b>LoVo and CoLo205 cells cultured in vitro were incubated in the presence of 0.5, 1.0, 1.5, 2.0, 2.5, 3.0, 3.5, and 4.0 microg/ml HpD for 4 h and exposed to different light doses delivered using a semiconductor laser at 630 nm with the energy density of 2, 5, 10, and 20 J/cm(2). After further culture for 24 h, the survival rate of LoVo and CoLo205 cells were analyzed by MTT assay, and the cellular fluorescence intensities of HpD were measured with a luminescence spectrometer.</p><p><b>RESULTS</b>HpD-PDT resulted in effective cell killing to a comparable magnitude in LoVo and CoLo205 cells cultured in vitro (P>0.05). The killing effects were positively correlated with the concentration of HpD and the dosage of laser irradiation. Exposure to 20 J/cm(2) resulted in an IC(50) of LoVo and CoLo205 cells of 0.4 and 0.6 microg/ml respectively, which were not significantly different (P>0.05). The cellular HpD fluorescence intensities were also similar between the two cells.</p><p><b>CONCLUSION</b>HpD-PDT may effectively kill LoVo and CoLo205 cells cultured in vitro.</p>

Outcome of Photodynamic Therapy for Early Esophageal Cancer

BACKGROUND/AIMS: Endoscopic treatment as an alternative to surgery has become increasingly popular for improving the quality of life. Although photodynamic therapy (PDT) has been used for the endoscopic treatment of digestive cancer, its curative efficacy remains unclear. We evaluated the curative efficacy of PDT in superficial esophageal cancer in inoperable patients. METHODS: Ten male patients with histologically proven early esophageal cancer (surgery was contraindicated for age > 80 years, surgery was contraindicated, Karnofsky performance status of at least 30%, or refusal of surgery) were intravenously injected with a hematoporphyrin derivative (2 mg/kg), and PDT was performed 48 h later. The response to treatment was assessed by gastroscopy with biopsies. RESULTS: The mean follow-up period was 27.6 months (range, 9.6-58.7 months). Endoscopic ultrasonography revealed that all ten cases were at tumor stage T1. Complete remission (CR) to initial and subsequent PDT was observed in all patients. For the CR cases, the recurrence rate was 10% (1/10) and the time from initial PDT to recurrence was 9.6 months. CONCLUSIONS: For patients in whom surgery is risky or refused, PDT may represent an acceptable alternative treatment modality, especially for superficial esophageal cancer without lymph node metastasis. However, a study involving long-term follow-up in a large population is needed for confirmation.

Preparation of hematoporphyrin-herceptin photoimmunoconjugate for photoimmunotherapy / 南方医科大学学报

<p><b>OBJECTIVE</b>To prepare photoimmunoconjugate of hematoporphyrin (HP) and herceptin, and study its killing and apoptosis-inducing effect on tumor cells BT-474.</p><p><b>METHODS</b>HP-herceptin photoimmunoconjugate was synthesized with EDCI as the condensator. After exposure of the cells to 630 nm laser, the killing effect of the conjugate and cell apoptosis were evaluated by MTT assay and flow cytometry.</p><p><b>RESULTS</b>Compared with free HP at equivalent dose, the immune reactivity, killing effect and the apoptosis-inducing effect of HP-herceptin immunoconjugate on BT-474 cells was enhanced (P<0.05).</p><p><b>CONCLUSION</b>The killing effect of HP-herceptin immunoconjugate is stronger than free HP on BT-474 cells.</p>

Gadolinium-hematoporphyrin: new potential MRI contrast agent for detection of breast cancer cell line

Gadolinium-porphyrins have been synthesized and are currently being investigated as magnetic resonance imaging [MRI] contrast agents. This study aimed to synthesize Gd-hematoporphyrin and applicate it for in vitro detection of breast cancer cell line [MCF-7]. The naturally occurring porphyrin [hematoporphyrin] was inserted with gadolinium [III] nitrate hexahydrate to yield Gd-H. T1 relaxation times and signal enhancement of the contrast agents were presented, and the results were compared. UV spectrophotometer measured the attachment of Gd to the cell membrane of MCF-7. Most of gadolinium chloride [GdCl3] was found in the washing solution, indicate that it didn't fixed to the breast cell membranes during incubation. Gd-DTPA showed some uptake into the MCF-7 cell membranes with incubation, however, its uptake was significantly lower than Gd-H. Good cell memberan uptake of Gd-porphyrin is comparable to controls, indicating selective delivery it to the breast cell line and considerable potency in diagnostic MR imaging for detection of breast cancer

Photogem Induces Necrosis in Various Uterine Cervical Cancer Cell Lines by PDT / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: In order to elucidate the antitumor effect of photodynamic therapy (PDT), using a derivative of the photosensitizing agent hematoporphyrin (Photogem) and a diode laser, the cell death of uterine cancer cell lines (CaSki, HT3, HeLa, and SKOV-3), and mice transplanted with TC-1 lung cancer cells, were evaluated. MATERIALS AND METHODS: The morphological changes, MTT assay, flow cytometry, cytotoxicity and tumor growth inhibition study were evaluated at various time intervals after the PDT. RESULTS: The results showed that the survival rates of each cell line decreased with time and dose response after performing the PDT. Also, the PDT-induced damage of cancer cells was almost entirely confined to necrosis of the tumor cells in the early time courses. The irradiation of CaSki cells in the presence of Photogem induced plasma membrane disruption and cell shrinkage, indicating the plasma membrane as the main target for Photogem. In the in vivo experiment, significantly longer survival and a significantly smaller tumor size were seen over the time courses of the Photogem with irradiation compared to the untreated control groups; resorption of the tumor was also observed after the PDT treatment. CONCLUSION: Collectively, our results indicated that Photogem possesses anti-tumor effects, and necrosis-like death, with plasma membrane damage, was postulated to be the principal mechanism of the antitumor effect of the PDT using Photogem.

Terapia fotodinâmica em hiperoxigenaçäo hiperbárica / Photodynamic therapy in hyperbaric hyperoxia

A Terapia Fotodinâmica (PDT) tem-se mostrado uma técnica seletiva promissora no tratamento do câncer. Após injeção intravenosa de um sensibilizador, que é retido por várias horas em células neoplásias, o tecido é irradiado por um laser, e devido a um mecanismo de transferência de energia não radiativa, agentes citotóxicos são produzidos, induzindo as células tumorais à morte. A técnica de hiperoxigenação hiperbárica (HBO) permite elevar o suprimento de oxigênio molecular, nos tecidos tumorais permitindo potencializar a PDT. Para avaliar o efeito combinado (PDT+HBO), desenvolvemos um modelo experimental que consiste na irradiação de tumores sólidos subcutâneos em dorso de ratos.

Photodynamic Therapy (PDT) have been demonstrated a very promising selectivity potential in cancer treatment. After an intravenous infusion of sensitizes, which is retained for severa! hours inside the neoplasic cells, the tissue is irradiated with a laser beam, and due a non-radiactive transfer mechanism, cytotoxic agents are produced, inducing death of the tumor cells. Hyperbaric hyperoxia (HBO) allows the tissues to achieve high supply of molecular oxygen. ln order to evaluate this combined effect of both therapies an experimental model was developed, creating a tumor mass in the dorsal subcutaneous tissue of rats.

A simple in vitro method to detect singlet oxygen and to compare photodynamic activity using alkaline phosphatase.

A simple, sensitive and reliable in vitro method based on photodynamic inactivation of alkaline phosphatase to detect singlet oxygen and for evaluating relative photosensitizing efficiencies of photosensitizers such as hematoporphyrin (Hp) and phthalocyanines has been developed and compared with photobleaching of p-nitroso dimethyl aniline (RNO) and photooxidation of L-tryptophan. Inactivation of alkaline phosphatase is dependent both on light fluence and sensitizer concentration. Scavengers like mannitol and azide anion indicated the involvement of singlet oxygen in the deactivation of alkaline phosphatase, since azide anion provided concentration dependent protection whereas mannitol had no effect and that compared to ordinary water, photoinactivation of alkaline phosphatase was three times higher in 65% D2O. Alkaline phosphatase appears to be resistant to free radical attack (particularly to OH radicals) since hydrogen peroxide alone or in presence of ferrous ions did not reduce the enzyme activity and mannitol or azide anion gave no significant protection when alkaline phosphatase was irradiated with Co-60 gamma rays up to 2 K Gy. With the present method using red light, the chloroaluminium phthalocyanine sulphonates prepared by sulphonation showed higher and the corresponding condensation product lower photodynamic activity; Hp being intermediate and Mn- and Gd-phthalocyanines had no photodynamic activity.

Effects of 2-deoxy-D-glucose on the photosensitisation-induced bioenergetic changes in Saccharomyces cerevisiae as observed by in vivo NMR spectroscopy.

Haematoporphyrin (HP), a photosensitiser used for photodynamic therapy (PDT) of tumours, has been observed to affect the cellular energy metabolism both in the absence and presence of light. The effects of HP alone and in combination with 2-deoxy-D-glucose (2-DG) on photosensitisation-induced bioenergetic changes in yeast (Saccharomyces cerevisiae) were monitored and compared using in vivo NMR spectroscopy. Presence of HP was seen to reduce polyphosphates and inorganic phosphate in the dark. Upon photo-irradiation, polyphosphates and sugar phosphates decrease drastically with concomitant rise in inorganic phosphates. Presence of 2-DG in the medium also induced a decrease in polyphosphates and nucleotide triphosphates and the build-up of 2-deoxy-D-glucose-6-phosphate (2-DG-6-P) was clearly detectable. The combination of 2-DG and HP followed by photo-irradiation, however, induced a significant reduction in intracellular pH and beta phosphate of ATP/inorganic phosphate (beta-ATP/Pi) ratio decreased to a larger extent as compared to similar treatment without 2-DG. These observations confirm that polyphosphates are utilised as phosphogen, as phosphate store to be used during phosphate deprivation and as alternative energy source under conditions of energy deficiency induced by HP-PDT and/or 2-DG. The present results further suggest that photodynamic therapy could be made more effective in conjunction with 2-DG administration.

Interdependence of drug dose, incubation time & light dose on hematoporphyrin derivative induced photoinactivation of brain tumour cells.

The relationship between photosensitizer concentration, light dose, incubation time and cellular damage in human cerebral glioma cells in culture, was studied. Cells were incubated with hematoporphyrin derivative (Hpd) for different durations at 37 degrees C. Immediately after specified period of incubation, cells were irradiated with white light. Cellular damage was assessed by colony forming ability of cells. A progressive reduction in the surviving fraction was observed as a function of drug and light dose. The survival curves were of exponential nature with an initial shoulder. The cell survival was found to be dependent on the time of incubation with Hpd. These results suggest that photodynamic cellular damage can be enhanced at low drug and light dose by increasing the incubation time.

Effects of some divalent metal ions on the aging phenomenon of hematoporphyrin and photofrin II.

Dilute aqueous solutions of hematoporphyrin (Hp) and its derivative (Hpd or PF II) have been found to undergo a transformation (aging) on keeping at room temperature leading to (i) shift of the Soret band from 395 nm to 405 nm, (ii) disappearance of visible bands I (610 nm) and IV (503 nm) and (iii) shift of the first emission band from 615 nm to 580 nm. The transformation was concentration dependent. The effects of concentration and temperature on the absorption spectra were much more pronounced in Hp than in Photofrin II (PF II). Variation of pH resulted in changes in the relative intensities of the absorption bands, possibly due to formation of different ionic species at different pH. The rate of transformation was accelerated in the presence of Zn ions (0.01 microM) and considerably increased at higher (50 microM) concentration. The effect of Cu ions was different from the effect of aging. It formed the metal-chelate even when present in very small amounts. The results (absorption and fluorescence analysis) suggest that in dilute solutions (conc. less than or equal to 2 microM) of Hp and PF II, Zn ions present in glass and water as impurity, deform the porphyrin nucleus leading to changes in the conjugated ring symmetry and hence changes in the absorption and fluorescence spectra, while in higher concentrations (greater than 2 microM) it forms the metal chelate as evidenced by their absorption and fluorescence spectra.

Effect of a newly developed hematoporphyrin derivative preparation on oral flora during orthodontic treatment

This study was designed to assess the effect of topical porphyrin application on some bacterial and fungal species of oral flora after using orthodontic treatment [ODT] which indicated to correct abnormalities of the teeth of patient. A novel porphyrin derivative was used. It was concluded that, all Gram-negative strains appeared to be resistant, except neisseria. Gram-positive strains were significantly inhibited due to porphyrin application. Oral fungus as C and ida albicans were significantly inhibited after porphyrin application

Inducing effect of hematoporphyrin derivative (HpD) on cell sister chromatid exchanges (SCE) in vitro / 中华肿瘤杂志

The mutagenic effect of HpD on cell SCE and the reactions of cell SCE to different sources of light combined with HpD were studied using V79 cells. There were 6 doses of HpD: 1 microgram/ml, 3 micrograms/ml, 5 micrograms/ml, 10 micrograms/ml, 50 micrograms/ml and 100 micrograms/ml. The dose of 5 micrograms/ml is equal to the maximum dose of HpD used in the clinic (HpD per milliliter of patient's blood). Our experiments demonstrated that when the cells were cultured in the dark and HpD was added to the medium no more than 5 micrograms/ml, the SCE frequencies were not increased. The cells were irradiated with different sources of light without HpD, both the fluorescence and ultraviolet light could promote SCE but the light of daylight lamp and red light did not increase it. But when HpD was added into culture medium at the dose of less than 5 micrograms/ml, every light could increase the cell SCE intensively except the daylight lamp light. The red light was more notable than the others by relation analysis.