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We examined effects of fluence rate on the photobleaching of the photosensitizer Pc 4 during photodynamic therapy (PDT) and the relationship between photobleaching and tumor response to PDT. BALB/c mice with intradermal EMT6 tumors were given 0.03 mg kg−1 Pc 4 by intratumor injection and irradiated at 667 nm with an irradiance of 50 or 150 mW cm−2 to a fluence of 100 J cm−2. While no cures were attained, significant tumor growth delay was demonstrated at both irradiances compared with drug-only controls. There was no significant difference in tumor responses to these two irradiances ( P = 0.857). Fluorescence spectroscopy was used to monitor the bleaching of Pc 4 during irradiation, with more rapid bleaching with respect to fluence shown at the higher irradiance. No significant correlation was found between fluorescence photobleaching and tumor regrowth for the data interpreted as a whole. Within each treatment group, weak associations between photobleaching and outcome were observed. In the 50 mW cm−2 group, enhanced photobleaching was associated with prolonged growth delay ( P = 0.188), while at 150 mW cm−2 this trend was reversed ( P = 0.308). Thus, it appears that Pc 4 photobleaching is not a strong predictor of individual tumor response to Pc 4-PDT under these treatment conditions. [ABSTRACT FROM AUTHOR]

Photodynamic therapy (PDT) is a clinically approved, minimally invasive therapeutic procedure that can exert a selective cytotoxic activity toward malignant cells. The procedure involves administration of a photosensitizing agent followed by irradiation at a wavelength corresponding to an absorbance band of the sensitizer. In the presence of oxygen, a series of events lead to direct tumor cell death, damage to the microvasculature, and induction of a local inflammatory reaction. Clinical studies revealed that PDT can be curative, particularly in early stage tumors. It can prolong survival in patients with inoperable cancers and significantly improve quality of life. Minimal normal tissue toxicity, negligible systemic effects, greatly reduced long-term morbidity, lack of intrinsic or acquired resistance mechanisms, and excellent cosmetic as well as organ function-sparing effects of this treatment make it a valuable therapeutic option for combination treatments. With a number of recent technological improvements, PDT has the potential to become integrated into the mainstream of cancer treatment. CA Cancer J Clin 2011. © 2011 American Cancer Society, Inc. [ABSTRACT FROM AUTHOR]

We present an in vivo fluorescence microscopic evaluation of intratumor distribution of the photosensitizer mono-L-aspartylchlorin-e6 (NPe6) in an intradermal mouse EMT6 tumor model. Although the identification of favorable photophysical and pharmacological properties has led to the development of new photosensitizers in photodynamic therapy, their intratumor distribution kinetics have remained relatively understudied. In this study, we used confocal fluorescence microscopy to follow the transport of NPe6 in vivo after systemic administration through the tail vein. Labeling of vasculature using fluorophore-conjugated anti-CD31 antibodies allows visualization of the uptake of NPe6 in tumor and normal vessels and its partitioning kinetics into the adjacent parenchyma for 3 hours after injection. During the initial 60 minutes after injection, the drug is predominantly confined to the vasculature. Subsequently, it significantly redistributes throughout the extravascular regions with no discernable difference in its extravasation rate between tumor and normal tissues. Further, we investigate the sensitizer's altered intratumor distribution in response to photodynamic therapy irradiation and observe that treatment-induced changes in vessel permeability caused enhanced accumulation of NPe6 in the extravascular space. Our findings are of immediate clinical relevance and demonstrate the importance of an in vivo imaging approach to examine the dynamic process of intratumor drug distribution. [ABSTRACT FROM AUTHOR]

We have developed a comprehensive theoretical model for rigorously describing the spatial and temporal dynamics of oxygen (3O2) consumption and transport and microscopic photodynamic dose deposition during photodynamic therapy (PDT) in vivo. Previously published models have been improved by considering perfused vessels as a time-dependent 3O2 source and linking the 3O2 concentration in the vessel to that within the tissue through the Hill equation. The time-dependent photochemical 3O2 consumption rate incorporates sensitizer photobleaching effects and an experimentally determined initially nonuniform photosensitizer distribution. The axial transport of 3O2 is provided for in the capillaries and in the surrounding tissue. A self-sensitized singlet oxygen (1O2)-mediated bleaching mechanism and the measured, initially nonuniform distribution of meso-tetrahydroxyphenyl chlorin at 3 h after intravascular administration were used to demonstrate the capabilities of the model. Time-evolved distributions of 3O2 concentration were obtained by numerically solving two-dimensional diffusion-with-reaction equations both in the capillary and the adjacent tissue. Using experimentally established physiological and photophysical parameters, the mathematical model allows computation of the dynamic variation of hemoglobin-3O2 saturation (SO2) within the vessels, irreversible sensitizer degradation due to photobleaching, and the microscopic distributions of 3O2, sensitizer concentration, and 1O2 dose deposition under various irradiation conditions. The simulations reveal severe axial gradients in 3O2 and in photodynamic dose deposition in response to a wide range of clinically relevant treatment parameters. Thus, unlike former Krogh cylinder-based models, which assume a constant 3O2 concentration at the vessel, this new model identifies conditions in which 3O2 depletion and minimal deposition of reacting 1O2 exist near the end of axial segments of vessels and shows that treatment-limiting 3O2 depletion is induced at fluence rates as low as 10 mW cm-2. These calculations also demonstrate that intercapillary heterogeneity of photosensitizer contributes significantly to the distribution of photodynamic dose. This more rigorous mathematical model enables comparison with experimentally observable, volume-averaged quantities such as SO2 and the loss of sensitizer fluorescence through bleaching that have not been included in previous analyses. Further, it establishes some of the intrinsic limitations of such measurements. Specifically, our simulations demonstrate that tissue measurements of SO2 and of photobleaching are necessarily insensitive to microscopic heterogeneity of photodynamic dose deposition and are sensitive to intercapillary spacing. Because prior knowledge of intercapillary distances in tumors is generally unavailable, these measurements must be interpreted with caution. We anticipate that this model will make useful dosimetry predictions that should inform optimal treatment conditions and improve current clinical protocols. [ABSTRACT FROM AUTHOR]

Genetically-encoded photosensitizers produce reactive oxygen species (ROS) in response to light. Transgenic expression of fusion proteins can target the photosensitizers to specific cell regions and permit the spatial and temporal control of ROS production. These ROS-generating proteins (RGPs) are widely used for cell ablation, mutagenesis and chromophore-assisted light inactivation of target proteins. However, the species produced by RGPs are unclear due to indirect measures with confounding interpretations. Recently, the RGP mini “Singlet Oxygen Generator” (miniSOG) was engineered from Arabidopsis thaliana phototropin 2. While miniSOG produces singlet oxygen ( 1 O 2 ), the contribution of superoxide (O 2 • - ) to miniSOG-generated ROS remains unclear. We measured the light-dependent O 2 • - production of purified miniSOG using HPLC separation of dihydroethidium (DHE) oxidation products. We demonstrate that DHE is insensitive to 1 O 2 and establish that DHE is a suitable indicator to measure O 2 • - production in a system that produces both 1 O 2 and O 2 • - . We report that miniSOG produces both 1 O 2 and O 2 • - , as can its free chromophore, flavin mononucleotide. miniSOG produced O 2 • - at a rate of ~4.0 µmol O 2 • - /min/µmol photosensitizer for an excitation fluence rate of 5.9 mW/mm 2 at 470 ± 20 nm, and the rate remained consistent across fluences (light doses). Overall, the contribution of O 2 • - to miniSOG phenotypes should be considered. [ABSTRACT FROM AUTHOR]

BACKGROUND Topical photodynamic therapy (PDT) for selected nonmelanoma skin cancer using 5-aminolevulinic acid (ALA) or methyl aminolevulinate (MAL) has yielded high long-term complete response rates with very good cosmesis. Pain during light activation of the photosensitizer can be a serious adverse event. A 2-step irradiance protocol has previously been shown to minimize ALA-PDT pain. OBJECTIVE To determine the irradiance-dependent pain threshold for MAL-PDT, to adapt the 2-step protocol to a light-emitting diode (LED) light source, and assess clinical response. METHODS In this prospective study, 25 superficial basal cell carcinoma (sBCC) received an initial irradiance by laser at 40 or 50 mW/cm², or LED at 35 mW/cm² followed by an irradiance at 70 mW/cm² for a total of 75 J/cm². Pain levels were recorded for both irradiance steps. Efficacy was assessed at 6, 12, or 24 months. RESULTS Pain was mild in the 40/70 mW/cm² laser cohort. Three instances of irradiance-limiting pain occurred at 50/70 mW/cm². Pain was minimal in the 35/70 mW/cm² LED cohort. Clinical response rates were 80% in the 50/70 mW/cm² laser cohort and 90% in the 35/70 mW/cm² LED cohort. CONCLUSION Topical PDT can be effectively delivered to sBCC with minimal treatment-related pain by a 2-step irradiance protocol. [ABSTRACT FROM AUTHOR]