Preclinical Study of Antineoplastic Sinoporphyrin Sodium-PDT via In Vitro and In Vivo Models.

Photodynamic therapy (PDT) investigations have seen stable increases and the development of new photosensitizers is a heated topic. Sinoporphyrin sodium is a new photosensitizer isolated from Photofrin. This article evaluated its anticancer effects by clonogenic assays, MTT assays and xenograft experiments in comparison to Photofrin. The clonogenicity inhibition rates of sinoporphyrin sodium-PDT towards four human cancer cell lines ranged from 85.5% to 94.2% at 0.5 µg/mL under 630 nm irradiation of 30 mW/cm² for 180 s. For MTT assays, the IC50 ranges of Photofrin-PDT and sinoporphyrin sodium-PDT towards human cancer cells were 0.3 µg/mL to 5.5 µg/mL and 0.1 µg/mL to 0.8 µg/mL under the same irradiation conditions, respectively. The IC50 values of Photofrin-PDT and sinoporphyrin sodium-PDT towards human skin cells, HaCaT, were 10 µg/mL and 1.0 µg/mL, respectively. Esophagus carcinoma and hepatoma xenograft models were established to evaluate the in vivo antineoplastic efficacy. A control group, Photofrin-PDT group (20 mg/kg) and sinoporphyrin sodium group at three doses, 0.5 mg/kg, 1 mg/kg and 2 mg/kg, were set. Mice were injected with photosensitizers 24 h before 60 J 630 nm laser irradiation. The tumor weight inhibition ratio of 2 mg/kg sinoporphyrin sodium-PDT reached approximately 90%. Besides, the tumor growths were significantly slowed down by 2 mg/kg sinoporphyrin sodium-PDT, which was equivalent to 20 mg/kg Photofrin-PDT. In sum, sinoporphyrin sodium-PDT showed great anticancer efficacy and with a smaller dose compared with Photofrin. Further investigations are warranted.

In vivo detection of chemiluminescence to monitor photodynamic threshold dose for tumor treatment.

During photodynamic therapy (PDT) it is important to monitor the treatment progress to achieve the intended therapeutic outcome. Chemiluminescence (CL) provides a sensitive and selective means for (1)O(2) detection. In our study, by comparing CL with the corresponding in situ PDT treatment protocols and the treatment effect, the results indicate that the treatment outcome for tumors was governed by a set of dosimetry factors with a rather complicated relationship, but there was a remarkable connection between tumor treatment effect and CL. In conclusion, CL can be used in in vivo PDT to determine the dose of tumor treatment and monitor the treatment threshold. By monitoring CL it is feasible to determine PDT dose to predict tumor treatment effect.

Novel applications of diagnostic X-rays in activating a clinical photodynamic drug: Photofrin II through X-ray induced visible luminescence from "rare-earth" formulated particles.

INTRODUCTION: In this communication we report on a novel non-invasive methodology in utilizing "soft" energy diagnostic X-rays to indirectly activate a photo-agent utilized in photodynamic therapy (PDT): Photofrin II (Photo II) through X-ray induced luminescence from Gadolinium Oxysulfide (20 micron dimension) particles doped with Terbium: Gd_{2}O_{2}S:Tb. Photodynamic agents such as Photo II utilized in PDT possess a remarkable property to become preferentially retained within the tumor's micro-environment. Upon the photo-agent's activation through (visible light) photon absorption, the agents exert their cellular cytotoxicity through type I and type II pathways through extensive generation of reactive oxygen species (ROS); namely, singlet oxygen ^{1}O_{2}, superoxide anion O_{2}^{-}, and hydrogen peroxide H_{2}O_{2}, within the intra-tumoral environment. Unfortunately, due to shallow visible light penetration depth (∼ 2 mm to 5 mm) in tissues, the current PDT strategy has largely been restricted to the treatment of surface tumors, such as the melanomas. Additional invasive strategies through optical fibers are currently utilized in getting the visible light into the intended deep seated targets within the body for PDT. METHODS: X-ray induced visible luminescence from Gd_{2}O_{2}S:Tb particles were spectroscopically characterized, and the potential in-vitro cellular cytotoxicity of Gd_{2}O_{2}S:Tb particles on human glioblastoma cells (due to 48 Hrs Gd_{2}O_{2}S:Tb particle exposure) was screened through the MTS cellular metabolic assay. In-vitro human glioblastoma cellular exposures in presence of Photo II with Gd_{2}O_{2}S:Tb particles were performed in the dark in sterile 96 well tissue culture plates, and the corresponding changes in the metabolic activities of the glioblastoma due to 15 minutes of (diagnostic energy) X-ray exposure was determined 48 Hrs after treatment through the MTS assay. RESULTS: Severe suppression (> 90% relative to controls) in the cellular metabolic activity of human glioblastoma was measured due to the treatment of clinically relevant concentrations of 20 µg/ml Photo II, with Gd_{2}O_{2}S:Tb particles, and (120 kVp) diagnostic X-rays. Taken together, the in-vitro findings herein provide the basis for future studies in determining the safety and efficacy of this non-invasive X-ray induced luminescence strategy in activating photo-agent in deep seated tumors.

Antitumor effect of photodynamic therapy in mice using direct application of Photofrin dissolved in lidocaine jelly.

BACKGROUND/PURPOSE: Photodynamic therapy (PDT) is a non-invasive cancer therapy that has a strong antitumor effect with intravenous administration of Photofrin. However, Photofrin causes light hypersensitivity that impairs the quality of life (QOL) of patients, and thus an improved method of administration is needed. Here, we report the antitumor effect of local administration of Photofrin in combination with a vasodilator, lidocaine hydrochloride. METHOD: The antitumor effect was investigated in nude mice transplanted with HeLa cells. An incision was made near the tumor and Photofrin dissolved in lidocaine jelly was applied directly to the tumor. The tumor was irradiated at 100 J/cm(2) with a yttrium aluminum garnet (YAG)-dye laser (630 nm) at 2 h after the direct application and the tumor volume was measured for 30 days after PDT to investigate the antitumor effect. In some mice, the tumor was excised 24 h after PDT and the depth of necrosis was measured in the excised specimen. RESULT: The tumor was mostly necrotized by PDT following direct application of 10 mg/ml Photofrin dissolved in lidocaine jelly and the effect was greater than with direct application of Photofrin alone. The increase in tumor volume observed in control mice was significantly inhibited in mice that received PDT after direct application of Photofrin in lidocaine jelly. CONCLUSION: PDT using direct application of Photofrin in lidocaine jelly has a strong antitumor effect in mice and this approach may avoid the adverse effects of systemic Photofrin administration.

Heme oxygenase-1 protects tumor cells against photodynamic therapy-mediated cytotoxicity.

Photodynamic therapy is a promising antitumor treatment modality approved for the management of both early and advanced tumors. The mechanisms of its antitumor action include generation of singlet oxygen and reactive oxygen species that directly damage tumor cells and tumor vasculature. A number of mechanisms seem to be involved in the protective responses to PDT that include activation of transcription factors, heat shock proteins, antioxidant enzymes and antiapoptotic pathways. Elucidation of these mechanisms might result in the design of more effective combination strategies to improve the antitumor efficacy of PDT. Using DNA microarray analysis to identify stress-related genes induced by Photofrin-mediated PDT in colon adenocarcinoma C-26 cells, we observed a marked induction of heme oxygenase-1 (HO-1). Induction of HO-1 with hemin or stable transfection of C-26 with a plasmid vector encoding HO-1 increased resistance of tumor cells to PDT-mediated cytotoxicity. On the other hand, zinc (II) protoporphyrin IX, an HO-1 inhibitor, markedly augmented PDT-mediated cytotoxicity towards C-26 and human ovarian carcinoma MDAH2774 cells. Neither bilirubin, biliverdin nor carbon monoxide, direct products of HO-1 catalysed heme degradation, was responsible for cytoprotection. Importantly, desferrioxamine, a potent iron chelator significantly potentiated cytotoxic effects of PDT. Altogether our results indicate that HO-1 is involved in an important protective mechanism against PDT-mediated phototoxicity and administration of HO-1 inhibitors might be an effective way to potentiate antitumor effectiveness of PDT.

Quantitative in vitro demonstration of two-photon photodynamic therapy using photofrin and visudyne.

Photodynamic therapy (PDT), the combined action of a photosensitizer and light to produce a cytotoxic effect, is an approved therapy for a number of diseases. At present, clinical PDT treatments involve one-photon excitation of the photosensitizer. A major limitation is that damage may be caused to healthy tissues that have absorbed the drug and lie in the beam path. Two-photon excitation may minimize this collateral damage, as the probability of absorption increases with the square of the light intensity, enabling spatial confinement of the photosensitizer activation. A potential application is the treatment of the wet-form of age-related macular degeneration, the foremost cause of central vision loss in the elderly. Herein, the commercial photosensitizers Visudyne and Photofrin are used to demonstrate quantitative in vitro two-photon PDT. A uniform layer of endothelial cells (YPEN-1) was irradiated with a Ti:sapphire laser (300 fs, 865 nm, 90 MHz) using a confocal scanning microscope. Quantification of the two-photon PDT effect was achieved using the permeability stain Hoechst 33258 and a SYTOX Orange viability stain. Visudyne was found to be around seven times more effective as a two-photon photosensitizer than Photofrin under the conditions used, consistent with its higher two-photon absorption cross-section. We also demonstrate for the first time the quadratic intensity dependence of cellular two-photon PDT. This simple in vitro method for quantifying the efficacy of photosensitizers for two-photon excited PDT will be valuable to test specifically designed two-photon photosensitizers before proceeding to in vivo studies in preclinical animal models.

Sonodynamic therapy of cancer using novel sonosensitizers.

Sonodynamic therapy (SDT) of cancer is based on preferential uptake and/or retention of a sonosensitizing drug (sonosensitizer) in tumor tissues and subsequent activation of the drug by ultrasound irradiation. Ultrasound can penetrate deeply into tissues and can be focused into a small region of a tumor to activate a sonosensitizer. This is a unique advantage in the non-invasive treatment of nonsuperficial tumors when compared to laser light used for photodynamic therapy. Recently, it has been found that photochemically active porphyrins also show significant antitumor effects when activated with ultrasound. The mechanism of sonodynamic action has been suggested to involve photoexcitation of the sensitizer by sonoluminescent light, with subsequent formation of singlet oxygen. This mini-review provides a brief overview of the following four sonosensitizers useful in SDT: i) a homogeneous complex of oligomers of hematoporphyrin, Photofrin II; ii) a gallium porphyrin complex, ATX-70; iii) a hydrophilic chlorin derivative, A7X-S10, and iv) a novel porphyrin derivative devoid of photosensitivity, DCPH-P-Na (I).

[Early diagnosis of bladder cancer by special fluorescence spectrum: an in vitro experiment].

OBJECTIVE: To investigate the value of special fluorescence spectrum in the early diagnosis of bladder cancer. METHODS: Hematoporphyrin monomethyl ether (HMME) of the concentrations of 10, 100, and 200 microg/ml, as photosensitizer, was added into the sample vessels of bladder cancer tissue to be incubated for 1 - 3 h. A detecting system consisting of solid state ultraviolet laser with the wavelength of 355 nm was used to measure the fluorescence spectrum. RESULTS: The laser-induced auto-fluorescence (LIAF) of the bladder cancer tissue had characteristic double peaks at 445 - 490 nm. There was a remarkable characteristic peak, drug peak, located at 628.25 nm in the laser-induced drug-fluorescence (LIDF) of the bladder cancer tissue soaked by HMME. CONCLUSION: Photosentisizer raise the sensitivity and specificity of fluorescence spectrum in diagnosis of bladder cancer. Stable, and with high strength and visible drug peak, the LIDF is may help improve the early diagnosis of bladder cancer.

Potassium Iodide Potentiates Broad-Spectrum Antimicrobial Photodynamic Inactivation Using Photofrin.

It is known that noncationic porphyrins such as Photofrin (PF) are effective in mediating antimicrobial photodynamic inactivation (aPDI) of Gram-positive bacteria or fungi. However, the aPDI activity of PF against Gram-negative bacteria is accepted to be extremely low. Here we report that the nontoxic inorganic salt potassium iodide (KI) at a concentration of 100 mM when added to microbial cells (108/mL) + PF (10 µM hematoporphyrin equivalent) + 415 nm light (10 J/cm2) can eradicate (>6 log killing) five different Gram-negative species (Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, Proteus mirabilis, and Acinetobacter baumannii), whereas no killing was obtained without KI. The mechanism of action appears to be the generation of microbicidal molecular iodine (I2/I3-) as shown by comparable bacterial killing when cells were added to the mixture after completion of illumination and light-dependent generation of iodine as detected by the formation of the starch complex. Gram-positive methicillin-resistant Staphylococcus aureus is much more sensitive to aPDI (200-500 nM PF), and in this case potentiation by KI may be mediated mainly by short-lived iodine reactive species. The fungal yeast Candida albicans displayed intermediate sensitivity to PF-aPDI, and killing was also potentiated by KI. The reaction mechanism occurs via singlet oxygen (1O2). KI quenched 1O2 luminescence (1270 nm) at a rate constant of 9.2 × 105 M-1 s-1. Oxygen consumption was increased when PF was illuminated in the presence of KI. Hydrogen peroxide but not superoxide was generated from illuminated PF in the presence of KI. Sodium azide completely inhibited the killing of E. coli with PF/blue light + KI.

The phototoxicity of photofrin was enhanced by PEGylated liposome in vitro.

In recent years, photodynamic therapy (PDT) with a photosensitizer and laser has been given attention, especially for the treatment of superficial cancers, such as lung, gastric, bladder and cervical cancer. In this study, in order to enhance the efficacy of PDT, photofrin liposome (PF-Lip) was prepared with dimyristoylphosphatidylcholine, dimyristoylphosphatidylglycerol and cholesterol. Polyethyleneglycol modified photofrin liposome (PF-PEG-Lip) was prepared by modification of PF-Lip with monomethoxypolyethyleneglycol-2.3-dimyristoylglycerol. PF-Lip and PF-PEG-Lip entrapped with photofrin with 81.0+/-5.9 and 81.2+/-9.2%, respectively. The particle size of each liposome was 114.3+/-5.7nm (PF-Lip) and 118+/-3.5nm (PF-PEG-Lip), respectively. It was suggested that PEGylated liposomes has no effect on the trapping ratio of PF and particle size. Phototoxicity was enhanced by liposomalization, especially PEG-modification. However, PF-PEG-Lip inhibited the uptake of photofrin into tumor cells. The amount of singlet oxygen from photofrin solution (PF-sol) and each liposome was PF-PEG-Lip=PF-Lip>PF-sol. The photofrin release revel of PF-PEG-Lip was lower than that of PF-Lip. In conclusion, the phototoxicity of PF-PEG-Lip was significantly higher than that of PF-sol or PF-Lip. It is expected that formation of a fixed aqueous layer on the liposome membrane by PEGylation physically changed it into the stable state of PF-PEG-Lip.

Ascorbate reacts with singlet oxygen to produce hydrogen peroxide.

Singlet oxygen is a highly reactive electrophilic species that reacts rapidly with electron-rich moieties, such as the double bonds of lipids, thiols, and ascorbate (AscH-). The reaction of ascorbate with singlet oxygen is rapid (k = 3 x 10(8) M(-1) s(-1)). Here we have investigated the stoichiometry of this reaction. Using electrodes to make simultaneous, real-time measurements of oxygen and hydrogen peroxide concentrations, we have investigated the products of this reaction. We have demonstrated that hydrogen peroxide is a product of this reaction. The stoichiometry for the reactants of the reaction (1 1O2 + 1AscH--->1H2O2 + 1dehydroascorbic) is 1:1. The formation of H2O2 results in a very different oxidant that has a longer lifetime and much greater diffusion distance. Thus, locally produced singlet oxygen with a half-life of 1 ns to 1 micros in a biological setting is changed to an oxidant that has a much longer lifetime and thus can diffuse to distant targets to initiate biological oxidations.

Using white light during photodynamic therapy: visualization only or treatment?

BACKGROUND: Photodynamic therapy (PDT) involves selective uptake and retention of a photosensitizer in a tumor, followed by irradiation with light (usually a 630 nm diode laser), initiating tumor necrosis through formation of oxidized products or singlet oxygen. Successful PDT of early cancers of the esophagus and of Barrett's esophagus with severe dysplasia has been reported. However, side effects (edema, stricture, etc.) and treatment failure have been observed. This study aims to evaluate the possible photodynamic effect induced by illumination from the endoscope on the PDT effect, since a photon emitted to see the lesion can potentially be a photon to treat it! MATERIALS AND METHODS: Two fiber endoscopes (Olympus GIFPQ20 and Pentax FG34X) and one videoendoscope (Olympus GIFQ140) were evaluated. Output power, irradiance and emission spectrum were measured. Using the molar extinction coefficient of Photofrin and optical coefficients of the esophagus, the relative photodynamic reaction yield, determined as a function of depth, was compared with that obtained with a 630 nm diode laser. RESULTS: The irradiance at 1 and 2 cm was, respectively, 18.4 and 4.6 mW/cm (Pentax FG34X), 10.6 and 2.65 mW/cm (Olympus GIFPQ20), and 2.7 and 3.2 mW/cm (Olympus GIFQ140). The highest irradiance could lead to a relative photodynamic reaction yield at the surface of the esophagus similar to (Olympus GIFPQ20) or greater than (Pentax FG34X) that obtained using a diode laser alone. CONCLUSIONS: Our results could explain side effects sometimes observed when performing PDT. 'Endoscopic' illumination, however, could also represent an interesting alternative to the 630 nm diode laser. When using white light, superficial efficacy of PDT could be reinforced and transmural necrosis leading to perforation or stenoses reduced, since there is less red light in depth compared with a diode laser.

Time-resolved investigations of singlet oxygen luminescence in water, in phosphatidylcholine, and in aqueous suspensions of phosphatidylcholine or HT29 cells.

Singlet oxygen was generated by energy transfer from the photoexcited sensitizer, Photofrin or 9-acetoxy-2,7,12,17-tetrakis-(beta-methoxyethyl)-porphycene (ATMPn), to molecular oxygen. Singlet oxygen was detected time-resolved by its luminescence at 1270 nm in an environment of increasing complexity, water (H2O), pure phosphatidylcholine, phosphatidylcholine in water (lipid suspensions), and aqueous suspensions of living cells. In the case of the lipid suspensions, the sensitizers accumulated in the lipids, whereas the localizations in the cells are the membranes containing phosphatidylcholine. By use of Photofrin, the measured luminescence decay times of singlet oxygen were 3.5 +/- 0.5 micros in water, 14 +/- 2 micros in lipid, 9 +/- 2 micros in aqueous suspensions of lipid droplets, and 10 +/- 3 micros in aqueous suspensions of human colonic cancer cells (HT29). The decay time in cell suspensions was much longer than in water and was comparable to the value in suspensions of phosphatidylcholine. That luminescence signal might be attributed to singlet oxygen decaying in the lipid areas of cellular membranes. The measured luminescence decay times of singlet oxygen excited by ATMPn in pure lipid and lipid suspensions were the same within the experimental error as for Photofrin. In contrast to experiments with Photofrin, the decay time in aqueous suspension of HT29 cells was 6 +/- 2 micros when using ATMPn.

Feasibility of using fluoresceinyl Cypridina luciferin analog in a novel chemiluminescence method for real-time photodynamic therapy dosimetry.

Singlet oxygen ((1)O(2)) is the most important cytotoxic agent in photodynamic therapy (PDT). The feasibility of using a chemiluminescence (CL) probe, 3,7-dihydro-6-[4-(2-(N'-(5-fluoresceinyl)thioureido)ethoxy)phenyl]-2-methylimidazo{1,2-a}pyrazin-3-one sodium salt (fluoresceinyl Cypridina luciferin analog, FCLA), to monitor (1)O(2) production during PDT is evaluated in vitro. Lymphoma cells were treated with various protocols of PDT. The results show that the FCLA-CL production during PDT is linearly related to the corresponding cytotoxicity, regardless of the treatment protocol. With minimum cytotoxicity and interference to the PDT treatment outcome, the FCLA-CL system is an effective means to quantify PDT (1)O(2) production and may provide an alternative real-time dosimeter.

Modeling of a type II photofrin-mediated photodynamic therapy process in a heterogeneous tissue phantom.

We present a quantitative framework to model a Type II photodynamic therapy (PDT) process in the time domain in which a set of rate equations are solved to describe molecular reactions. Calculation of steady-state light distributions using a Monte Carlo method in a heterogeneous tissue phantom model demonstrates that the photon density differs significantly in a superficial tumor of only 3 mm thickness. The time dependences of the photosensitizer, oxygen and intracellular unoxidized receptor concentrations were obtained and monotonic decreases in the concentrations of the ground-state photosensitizer and receptor were observed. By defining respective decay times, we quantitatively studied the effects of photon density, drug dose and oxygen concentration on photobleaching and cytotoxicity of a photofrin-mediated PDT process. Comparison of the dependences of the receptor decay time on photon density and drug dose at different concentrations of oxygen clearly shows an oxygen threshold under which the receptor concentration remains constant or PDT exhibits no cytotoxicity. Furthermore, the dependence of the photosensitizer and receptor decay times on the drug dose and photon density suggests the possibility of PDT improvement by maximizing cytotoxicity in a tumor with optimized light and drug doses. We also discuss the utility of this model toward the understanding of clinical PDT treatment of chest wall recurrence of breast carcinoma.

Comparative mass spectrometric analyses of Photofrin oligomers by fast atom bombardment mass spectrometry, UV and IR matrix-assisted laser desorption/ionization mass spectrometry, electrospray ionization mass spectrometry and laser desorption/jet-cooling photoionization mass spectrometry.

Photofrin (porfimer sodium) is a porphyrin derivative used in the treatment of a variety of cancers by photodynamic therapy. This oligomer complex and a variety of porphyrin monomers, dimers and trimers were analyzed with five different mass spectral ionization techniques: fast atom bombardment, UV and IR matrix-assisted laser desorption/ionization, electrospray ionization, and laser desorption/jet-cooling photoionization. All five approaches resulted in very similar oligomer distributions with an average oligomer length of 2.7 +/- 0.1 porphyrin units. In addition to the Photofrin analysis, this study provides a side-by-side comparison of the spectra for the five different mass spectrometric techniques.

Quantum yields and kinetics of the photobleaching of hematoporphyrin, Photofrin II, tetra(4-sulfonatophenyl)-porphine and uroporphyrin.

Porphyrins used as sensitizers for the photodynamic therapy (PDT) of tumors are progressively destroyed (photobleached) during illumination. If the porphyrin bleaches too rapidly, tumor destruction will not be complete. However, with appropriate sensitizer dosages and bleaching rates, irreversible photodynamic injury to the normal tissues surrounding the tumor, which retain less sensitizer, may be significantly decreased. This paper surveys the quantum yields and kinetics of the photobleaching of four porphyrins: hematoporphyrin (HP), Photofrin II (PF II), tetra(4-sulfonatophenyl)porphine (TSPP) and uroporphyrin I (URO). The initial quantum yields of photobleaching, as measured in pH 7.4 phosphate buffer in air, were: 4.7 x 10(-5), 5.4 x 10(-5), 9.8 x 10(-6), and 2.8 x 10(-5) for HP, PF II, TSPP and URO respectively; thus, the rates of photobleaching are rather slow. Low oxygen concentration (2 microM) significantly reduced the photobleaching yields. However, D2O increased the yields only slightly, and the singlet oxygen quencher, azide, had no effect, even at 0.1 M. Photosensitizing porphyrins in body fluids, cells and tissues may be closely associated with various photooxidizable molecules and electron acceptors and donors. Therefore, selected model compounds in these categories were examined for their effects on porphyrin photobleaching. A number inhibited and/or accelerated photobleaching, depending on the compound, the porphyrin and the reaction conditions. For example, 1.0 mM furfuryl alcohol increased the photobleaching yields of HP and URO more than 5-fold, with little effect on PF II or TSPP. In contrast, the electron acceptor, methyl viologen, increased the photobleaching yield of TSPP more than 10-fold, with little accelerating effect on the other porphyrins. These results suggest that the mechanism(s) of the photobleaching of porphyrin photosensitizers in cells and tissues during PDT may be complex.

Dye-sensitized destabilization of liposomes bearing photooxidizable lipid head groups.

Liposomes were prepared from mixtures of dipalmitoyl-L-alpha-phosphatidylcholine and up to 40% mol:mol of N-stearoyl-L-histidine (NSH) in the presence of the hydrophobic sensitizer DHE. In the dark such liposomes are stable and retain entrapped salts. On photolysis with visible light, liposomes leak trapped ions at NSH concentrations greater than 10% mol:mol. Up to 15% mol:mol NSH concentration leakage is seen only during the illumination period, whereas at higher concentration the liposomes continue to leak contents after illumination and fuse to form larger structures. Photolysis of the liposomes is accompanied by oxygen uptake in proportion to the NSH concentration within the bilayer. Photocontrol of liposome permeability through oxidation of membrane additives such as NSH offers a potential means for controlled drug delivery and might be useful as an adjunct to photodynamic therapy.

Measurement of fluorophore concentrations and fluorescence quantum yield in tissue-simulating phantoms using three diffusion models of steady-state spatially resolved fluorescence.

Steady-state diffusion theory models of fluorescence in tissue have been investigated for recovering fluorophore concentrations and fluorescence quantum yield. Spatially resolved fluorescence, excitation and emission reflectance Carlo simulations, and measured using a multi-fibre probe on tissue-simulating phantoms containing either aluminium phthalocyanine tetrasulfonate (AlPcS4), Photofrin meso-tetra-(4-sulfonatophenyl)-porphine dihydrochloride The accuracy of the fluorophore concentration and fluorescence quantum yield recovered by three different models of spatially resolved fluorescence were compared. The models were based on: (a) weighted difference of the excitation and emission reflectance, (b) fluorescence due to a point excitation source or (c) fluorescence due to a pencil beam excitation source. When literature values for the fluorescence quantum yield were used for each of the fluorophores, the fluorophore absorption coefficient (and hence concentration) at the excitation wavelength (mu(a,x,f)) was recovered with a root-mean-square accuracy of 11.4% using the point source model of fluorescence and 8.0% using the more complicated pencil beam excitation model. The accuracy was calculated over a broad range of optical properties and fluorophore concentrations. The weighted difference of reflectance model performed poorly, with a root-mean-square error in concentration of about 50%. Monte Carlo simulations suggest that there are some situations where the weighted difference of reflectance is as accurate as the other two models, although this was not confirmed experimentally. Estimates of the fluorescence quantum yield in multiple scattering media were also made by determining mu(a,x,f) independently from the fitted absorption spectrum and applying the various diffusion theory models. The fluorescence quantum yields for AlPcS4 and TPPS4 were calculated to be 0.59 +/- 0.03 and 0.121 +/- 0.001 respectively using the point source model, and 0.63 +/- 0.03 and 0.129 +/- 0.002 using the pencil beam excitation model. These results are consistent with published values.

Cellular accumulation and biological activity of hematoporphyrin derivative(L) in comparison with photofrin II.

Hematoporphyrin derivative, a drug used in the photodynamic therapy of solid tumours was synthesized in the laboratory and was called Hpd(L). Physico-chemical and biological properties of this drug have been compared with Photofrin II, the commercially available drug. Both Hpd(L) and Photofrin II possess similar properties qualitatively. Quantitatively, Hpd(L) was half as active as Photofrin II in its efficacy in causing photodynamic cytotoxicity or in the optical densities at the absorption peaks. These differences could be due to the differences in the compositions. Hpd(L) is a non-purified complex mixture of a number of porphyrin derivatives whereas Photofrin II is a relatively purer compound consisting of di- and tri-hematoporphyrins linked through ether or ester bonds. In vitro cellular uptake and retention of these drugs has been found to be a passive process not involving energy expenditure. pH and temperature of the incubation media have been found to profoundly influence these processes, while a complex relation seems to exist between physiological state of a cell and accumulation of these photosensitizers.